Wonnacott S


Full name : Wonnacott Susan

First name : Susan

Mail : Department of Biology & Biochemistry, University of Bath, Bath BA2 7AY

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Country : United Kingdom

Email : bsssw@bath.ac.uk

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Website : \/\/www.bath.ac.uk\/bio-sci\/contacts\/academics\/sue_wonnacott\/

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References (139)

Title : Integration of inhibitory and excitatory effects of alpha7 nicotinic acetylcholine receptor activation in the prelimbic cortex regulates network activity and plasticity - Udakis_2016_Neuropharmacol_105_618
Author(s) : Udakis M , Wright VL , Wonnacott S , Bailey CP
Ref : Neuropharmacology , 105 :618 , 2016
Abstract : Cognitive and attentional processes governed by the prefrontal cortex (PFC) are influenced by cholinergic innervation. Here we have explored the role of alpha7 nicotinic acetylcholine receptors (nAChRs) as mediators of cholinergic signalling in the dorsomedial (prelimbic) PFC, using mouse brain slice electrophysiology. Activation of alpha7 nAChRs located on glutamatergic terminals and cell soma of GABAergic interneurons increased excitation and inhibition, respectively, in layer V of the prelimbic cortex. These actions were distinguished by their differential dependence on local acetylcholine (ACh): potentiation of endogenous cholinergic signalling with the positive allosteric modulator, PNU-120596, enhanced spontaneous excitatory events, an effect that was further increased by inhibition of acetylcholinesterase. In contrast, alpha7 nicotinic modulation of inhibitory signalling required addition of exogenous agonist (PNU-282987) as well as PNU-120596, and was unaffected by acetylcholinesterase inhibition. Thus alpha7 nAChRs can bi-directionally regulate network activity in the prelimbic cortex, depending on the magnitude and localisation of cholinergic signalling. This bidirectional influence is manifest in dual effects of alpha7 nAChRs on theta-burst-induced long-term potentiation (LTP) in layer V of the prelimbic cortex. Antagonism of alpha7 nAChRs significantly decreased LTP implicating a contribution from endogenous ACh, consistent with the ability of local ACh to enhance glutamatergic signalling. Exogenous agonist plus potentiator also decreased LTP, indicative of the influence of this drug combination on inhibitory signalling. Thus alpha7 nAChRs make a complex contribution to network activity and synaptic plasticity in the prelimbic cortex.
ESTHER : Udakis_2016_Neuropharmacol_105_618
PubMedSearch : Udakis_2016_Neuropharmacol_105_618
PubMedID: 26921769

Title : Pharmacological differences between rat frontal cortex and hippocampus in the nicotinic modulation of noradrenaline release implicate distinct receptor subtypes - Kennett_2012_Nicotine.Tob.Res_14_1339
Author(s) : Kennett A , Heal DJ , Wonnacott S
Ref : Nicotine Tob Res , 14 :1339 , 2012
Abstract : INTRODUCTION: Noradrenergic mechanisms in frontal cortex and hippocampus are relevant to attentional and stress-related aspects of addiction, respectively. Nicotinic receptors (nAChRs) modulate the release of noradrenaline (NA) in these tissues. This study determined if similar subtypes of nAChR regulate NA release in rat frontal cortex and hippocampus.
METHODS: The release of [(3)H]-NA from rat tissue prisms was characterized in a 96-well plate assay. In vivo microdialysis was used to monitor NA overflow from rat frontal cortex and hippocampus in conscious freely moving rats.
RESULTS: [(3)H]-NA release from frontal cortex prisms was more sensitive to nicotinic agonists than release from hippocampal prisms. The beta2-selective agonist 5-iodo-A-85380 was 1000-fold more potent in frontal cortex compared with hippocampus. Agonist-evoked [(3)H]-NA release was inhibited by the beta2-selective antagonist dihydro-beta-erythroidine (DHbetaE) in frontal cortex, whereas in hippocampal tissue, DHbetaE had no effect. In vivo, 5-iodo-A-85380 (1, 100 muM) applied locally via the dialysis probe, significantly increased NA overflow, compared with basal release, in frontal cortex but not in hippocampus.
CONCLUSIONS: These data support the modulation of NA release by different nAChR subtypes in frontal cortex and hippocampus. The pharmacological profile for rat hippocampus is consistent with previous studies, implicating alpha3beta4* nAChRs in the modulation of NA release in this tissue. nAChRs having this function in frontal cortex are pharmacologically distinct and correspond to beta2-containing nAChRs.
ESTHER : Kennett_2012_Nicotine.Tob.Res_14_1339
PubMedSearch : Kennett_2012_Nicotine.Tob.Res_14_1339
PubMedID: 22614547

Title : Xenopus laevis RIC-3 enhances the functional expression of the C. elegans homomeric nicotinic receptor, ACR-16, in Xenopus oocytes - Bennett_2012_J.Neurochem_123_911
Author(s) : Bennett HM , Lees K , Harper KM , Jones AK , Sattelle DB , Wonnacott S , Wolstenholme AJ
Ref : Journal of Neurochemistry , 123 :911 , 2012
Abstract : RIC-3 enhances the functional expression of certain nicotinic acetylcholine receptors (nAChRs) in vertebrates and invertebrates and increases the availability of functional receptors in cultured cells and Xenopus laevis oocytes. Maximal activity of RIC-3 may be cell-type dependent, so neither mammalian nor invertebrate proteins is optimal in amphibian oocytes. We cloned the X. laevis ric-3 cDNA and tested the frog protein in oocyte expression studies. X. laevis RIC-3 shares 52% amino acid identity with human RIC-3 and only 17% with that of Caenorhabditis elegans. We used the C. elegans nicotinic receptor, ACR-16, to compare the ability of RIC-3 from three species to enhance receptor expression. In the absence of RIC-3, the proportion of oocytes expressing detectable nAChRs was greatly reduced. Varying the ratio of acr-16 to X. laevis ric-3 cRNAs injected into oocytes had little impact on the total cell current. When X. laevis, human or C. elegans ric-3 cRNAs were co-injected with acr-16 cRNA (1 : 1 ratio), 100 muM acetylcholine induced larger currents in oocytes expressing X. laevis RIC-3 compared with its orthologues. This provides further evidence for a species-specific component of RIC-3 activity, and suggests that X. laevis RIC-3 is useful for enhancing the expression of invertebrate nAChRs in X. laevis oocytes.
ESTHER : Bennett_2012_J.Neurochem_123_911
PubMedSearch : Bennett_2012_J.Neurochem_123_911
PubMedID: 22970690

Title : Neurotoxicity induced by okadaic acid in the human neuroblastoma SH-SY5Y line can be differentially prevented by alpha7 and beta2* nicotinic stimulation - Del Barrio_2011_Toxicol.Sci_123_193
Author(s) : Del Barrio L , Martin-de-Saavedra MD , Romero A , Parada E , Egea J , Avila J , McIntosh JM , Wonnacott S , Lopez MG
Ref : Toxicol Sci , 123 :193 , 2011
Abstract : A good model of neuronal death that reproduces the characteristic tau (tau) hyperphosphorylation of Alzheimers disease is the use of okadaic acid (OA). The aim of this study was to determine the contribution of alpha7 and beta2* nicotinic acetylcholine receptor (nAChR) subtypes to neuroprotection against OA in the SH-SY5Y cell line by using the selective alpha7 and beta2* nAChR agonists PNU 282987 and 5-Iodo-A85380, respectively. The results of this study show that both alpha7 and beta2* nAChR can afford neuroprotection against OA-induced neurotoxicity. Protection mediated by alpha7 nAChRs was independent of Ca(2+) and involved the intracellular signaling pathway Janus Kinase-2/Phosphatidylinositol-3-kinase/Akt. When Ca(2+) entry was promoted through the alpha7 nAChR by using the alpha7-selective positive allosteric modulator PNU 120596, protection was lost. By contrast, protection mediated by beta2* nAChRs was Ca(2+) dependent and implicated the signaling pathways PI3K/Akt and extracellular regulated kinase 1/2. Both alpha7 and beta2* nAChR activation converged on downregulation of GSK-3beta and reduction of tau phosphorylation in cells undergoing cell death induced by OA. Therefore, targeting nAChR could offer a strategy for reducing neurodegeneration secondary to hyperphosphorylation of protein tau.
ESTHER : Del Barrio_2011_Toxicol.Sci_123_193
PubMedSearch : Del Barrio_2011_Toxicol.Sci_123_193
PubMedID: 21715663

Title : alpha6beta2* and alpha4beta2* nicotinic acetylcholine receptors as drug targets for Parkinson's disease - Quik_2011_Pharmacol.Rev_63_938
Author(s) : Quik M , Wonnacott S
Ref : Pharmacol Rev , 63 :938 , 2011
Abstract : Parkinson's disease is a debilitating movement disorder characterized by a generalized dysfunction of the nervous system, with a particularly prominent decline in the nigrostriatal dopaminergic pathway. Although there is currently no cure, drugs targeting the dopaminergic system provide major symptomatic relief. As well, agents directed to other neurotransmitter systems are of therapeutic benefit. Such drugs may act by directly improving functional deficits in these other systems, or they may restore aberrant motor activity that arises as a result of a dopaminergic imbalance. Recent research attention has focused on a role for drugs targeting the nicotinic cholinergic systems. The rationale for such work stems from basic research findings that there is an extensive overlap in the organization and function of the nicotinic cholinergic and dopaminergic systems in the basal ganglia. In addition, nicotinic acetylcholine receptor (nAChR) drugs could have clinical potential for Parkinson's disease. Evidence for this proposition stems from studies with experimental animal models showing that nicotine protects against neurotoxin-induced nigrostriatal damage and improves motor complications associated with l-DOPA, the "gold standard" for Parkinson's disease treatment. Nicotine interacts with multiple central nervous system receptors to generate therapeutic responses but also produces side effects. It is important therefore to identify the nAChR subtypes most beneficial for treating Parkinson's disease. Here we review nAChRs with particular emphasis on the subtypes that contribute to basal ganglia function. Accumulating evidence suggests that drugs targeting alpha6beta2* and alpha4beta2* nAChR may prove useful in the management of Parkinson's disease.
ESTHER : Quik_2011_Pharmacol.Rev_63_938
PubMedSearch : Quik_2011_Pharmacol.Rev_63_938
PubMedID: 21969327

Title : Glutamate-dopamine crosstalk in the rat prefrontal cortex is modulated by Alpha7 nicotinic receptors and potentiated by PNU-120596 - Livingstone_2010_J.Mol.Neurosci_40_172
Author(s) : Livingstone PD , Dickinson JA , Srinivasan J , Kew JN , Wonnacott S
Ref : Journal of Molecular Neuroscience , 40 :172 , 2010
Abstract : The aim of this study was to explore the modulation by alpha7 nicotinic receptors (nAChRs) of dopamine and glutamate release in the rat prefrontal cortex where these receptors are implicated in attentional processes and are therapeutic targets for cognitive deficits. The presence of presynaptic alpha7 nAChRs on glutamate terminals is supported by the ability of the subtype-selective agonist Compound A to evoke [(3)H]D-aspartate release from synaptosomes: This response was potentiated by the selective allosteric potentiator PNU-120596 and blocked by alphabungarotoxin. Compound A also evoked dopamine overflow in the prefrontal cortex in vivo, and this was potentiated by PNU-120596. alpha7 nAChR-evoked [(3)H]dopamine release from tissue prisms in vitro was blocked by antagonists of NMDA and AMPA receptors. These data are consistent with a model in which alpha7 nAChRs present on glutamate terminals increase glutamate release that (1) contributes to presynaptic facilitation and synaptic plasticity and (2) co-ordinately enhances dopamine release from neighbouring boutons.
ESTHER : Livingstone_2010_J.Mol.Neurosci_40_172
PubMedSearch : Livingstone_2010_J.Mol.Neurosci_40_172
PubMedID: 19688191

Title : Oligomerisation differentially affects the acute and chronic actions of amyloid-beta in vitro - Innocent_2010_Neuropharmacol_59_343
Author(s) : Innocent N , Evans N , Hille C , Wonnacott S
Ref : Neuropharmacology , 59 :343 , 2010
Abstract : Key neuropathological hallmarks of Alzheimer's disease include the accumulation of amyloid-beta (Abeta), disruption of Ca(2+) homeostasis and neurodegeneration. However, the physical nature of the toxic Abeta species is controversial. Here, we examined the effect of aging on acute and chronic actions of Abeta peptides: changes in intracellular Ca(2+) and toxic responses, respectively. Acute application of Abeta(1-42) to PC12 cells potentiated KCl-evoked increases in Ca(2+), while chronic application decreased mitochondrial function with concomitant perturbation of membrane integrity and activation of apoptosis in PC12 cells, and reduced neurite length and synaptogenesis in rat cortical neurons. Both the acute and chronic effects of Abeta(1-42) were prevented by the anti-oligomerisation peptide D-KLVFFA, implicating oligomeric structures. The generation of a range of oligomeric species by aging Abeta(1-42) at 37 degrees C for different times was supported by thioflavin T fluorescence and atomic force microscopy. Abeta(1-42) aged for 24 h maximally potentiated KCl-evoked increases in Ca(2+), and this correlated with oligomers composed of 3-6 monomers, as judged by size exclusion filtration. Aging for 72 or 96 h, which generated fibrillar structures, was less efficacious. The Abeta(25-35) fragment that lacks the self-recognition element targeted by D-KLVFFA failed to potentiate KCl-evoked increases in Ca(2+). However, Abeta(25-35) was more efficacious than Abeta(1-42) at decreasing cellular functions when applied chronically. The acute and chronic effects of Abeta(1-42) also showed differential sensitivity to blockade of voltage operated Ca(2+) channels. These results suggest that the acute effects of Abeta(1-42) on Ca(2+) signals do not underpin the toxic responses measured, although both acute and chronic effects are promoted by small oligomeric species.
ESTHER : Innocent_2010_Neuropharmacol_59_343
PubMedSearch : Innocent_2010_Neuropharmacol_59_343
PubMedID: 20388522

Title : Molecular determinants for competitive inhibition of alpha4beta2 nicotinic acetylcholine receptors - Iturriaga-Vasquez_2010_Mol.Pharmacol_78_366
Author(s) : Iturriaga-Vasquez P , Carbone A , Garcia-Beltran O , Livingstone PD , Biggin PC , Cassels BK , Wonnacott S , Zapata-Torres G , Bermudez I
Ref : Molecular Pharmacology , 78 :366 , 2010
Abstract : The Erythrina alkaloids erysodine and dihydro-beta-erythroidine (DHbetaE) are potent and selective competitive inhibitors of alpha4beta2 nicotinic acetylcholine receptors (nAChRs), but little is known about the molecular determinants of the sensitivity of this receptor subtype to inhibition by this class of antagonists. We addressed this issue by examining the effects of DHbetaE and a range of aromatic Erythrina alkaloids on [(3)H]cytisine binding and receptor function in conjunction with homology models of the alpha4beta2 nAChR, mutagenesis, and functional assays. The lactone group of DHbetaE and a hydroxyl group at position C-16 in aromatic Erythrina alkaloids were identified as major determinants of potency, which was decreased when the conserved residue Tyr126 in loop A of the alpha4 subunit was substituted by alanine. Sensitivity to inhibition was also decreased by substituting the conserved aromatic residues alpha4Trp182 (loop B), alpha4Tyr230 (loop C), and beta2Trp82 (loop D) and the nonconserved beta2Thr84; however, only alpha4Trp182 was predicted to contact bound antagonist, suggesting alpha4Tyr230, beta2Trp82, and beta2Thr84 contribute allosterically to the closed state elicited by bound antagonist. In addition, homology modeling predicted strong ionic interactions between the ammonium center of the Erythrina alkaloids and beta2Asp196, leading to the uncapping of loop C. Consistent with this, beta2D196A abolished sensitivity to inhibition by DHbetaE or erysodine but not by epierythratidine, which is not predicted to form ionic bonds with beta2Asp196. This residue is not conserved in subunits that comprise nAChRs with low sensitivity to inhibition by DHbetaE or erysodine, which highlights beta2Asp196 as a major determinant of the receptor selectivity of Erythrina alkaloids.
ESTHER : Iturriaga-Vasquez_2010_Mol.Pharmacol_78_366
PubMedSearch : Iturriaga-Vasquez_2010_Mol.Pharmacol_78_366
PubMedID: 20547737

Title : Increase in locomotor activity after acute administration of the nicotinic receptor agonist 3-bromocytisine in rats - Abin-Carriquiry_2010_Eur.J.Pharmacol_634_89
Author(s) : Abin-Carriquiry JA , Urbanavicius J , Scorza C , Rebolledo-Fuentes M , Wonnacott S , Cassels BK , Dajas F
Ref : European Journal of Pharmacology , 634 :89 , 2010
Abstract : Nicotinic acetylcholine receptors influence striatal dopaminergic activity and its outcome on motor behavior. For these reasons, nicotinic receptors have been considered as therapeutically relevant targets for Parkinson's disease, in which a dramatic loss of dopamine affects motor functions. The aim of the present work was to compare the effects on locomotor activity induced by the nicotinic agonist cytisine and two brominated derivatives, 5- and 3-bromocytisine (5-BrCy and 3-BrCy) using nicotine for comparison. After acute systemic administration of the agonists only 3-BrCy induced an increase in locomotor activity. To study the mechanism of action involved in this increase we co-administered 3-BrCy with the nicotinic antagonist mecamylamine and also examined 3-BrCy's effects in rats pre-treated with the long acting nicotinic antagonist chlorisondamine, administered directly in the dorsal and ventral striatum. We studied the role of the dopaminergic system by co-administration of the D2 dopamine receptor antagonist, haloperidol. The results indicate that the increase in motor activity elicited by 3-BrCy was mediated by nicotinic receptors in the dorsal and ventral striatum and depends on the interaction of nicotinic receptors with the dopaminergic system. We conclude that 3-BrCy might be a new tool to study the modulation of the dopaminergic system by nicotinic receptors and their behavioral implications.
ESTHER : Abin-Carriquiry_2010_Eur.J.Pharmacol_634_89
PubMedSearch : Abin-Carriquiry_2010_Eur.J.Pharmacol_634_89
PubMedID: 20184877

Title : In silico characterization of cytisinoids docked into an acetylcholine binding protein - Abin-Carriquiry_2010_Bioorg.Med.Chem.Lett_20_3683
Author(s) : Abin-Carriquiry JA , Zunini MP , Cassels BK , Wonnacott S , Dajas F
Ref : Bioorganic & Medicinal Chemistry Lett , 20 :3683 , 2010
Abstract : Homology models of nicotinic acetylcholine receptors (nAChRs) suggest that subtype specificity is due to non-conserved residues in the complementary subunit of the ligand-binding pocket. Cytisine and its derivatives generally show a strong preference for heteromeric alpha4beta2* nAChRs over the homomeric alpha7 subtype, and the structural modifications studied do not cause large changes in their nAChR subtype selectivity. In the present work we docked cytisine, N-methylcytisine, and several pyridone ring-substituted cytisinoids into the crystallographic structure of the Lymnaea stagnalis acetylcholine binding protein (AChBP) co-crystallized with nicotine (1UW6). The graphical analysis of the best poses showed that cytisinoids have weak interactions with the side chains of the non-conserved amino acids in the complementary subunit justifying the use of PDB 1UWB as a surrogate for nAChR. Furthermore, we found a high correlation (R(2)=0.96) between the experimental pIC(50) values at alpha4beta2* nAChR and docking energy (S) of the best cytisinoid poses within the AChBP. Due to the quality of the correlation we suggest that this equation might be used as a predictive model to propose new cytisine-derived nAChRs ligands. Our docking results also suggest that further structural modifications of these cytisinoids will not greatly alter their alpha4beta2*/alpha7 selectivity.
ESTHER : Abin-Carriquiry_2010_Bioorg.Med.Chem.Lett_20_3683
PubMedSearch : Abin-Carriquiry_2010_Bioorg.Med.Chem.Lett_20_3683
PubMedID: 20493692

Title : Subtype-selective nicotinic agonists enhance olfactory working memory in normal rats: a novel use of the odour span task - Rushforth_2010_Neurosci.Lett_471_114
Author(s) : Rushforth SL , Allison C , Wonnacott S , Shoaib M
Ref : Neuroscience Letters , 471 :114 , 2010
Abstract : Nicotinic agonists have been shown to enhance performance in cognitive tasks based on attention and memory. The aim of this study was to use a test of olfactory working memory; the odour span task (OST) in rodents, to investigate the effects of subtype-specific nicotinic agonists on working memory in normal rats. Rats were trained in a non-matching to sample (NMTS) rule and then the full OST, which involved identifying a novel odour from an increasing number of presented odours. Male hooded Lister rats were treated with nicotine, selective nicotinic agonists or vehicle (saline). In order to validate the task, muscarinic and nicotinic receptor antagonists were also examined. Nicotine at both 0.05 and 0.1mg/kg significantly increased mean span length in the OST. The selective alpha 4 beta 2 nicotinic receptor agonist metanicotine (0.1mg/kg s.c.) and the selective alpha 7 nicotinic receptor agonist (R)-N-(1-azabicyclo[2.2.2]oct-3-yl)(5-(2-pyridyl)thiophene-2-carboxamide) (compound A, 10mg/kg i.p.) also improved performance. In contrast, mecamylamine and scopolamine significantly decreased mean span length. These findings suggest a role for the activation of both alpha 4 beta 2 and alpha 7 subtypes of neuronal nicotinic receptor in mediating enhancements of olfactory working memory capacity in normal, non-compromised rats. These nicotinic receptor subtypes may therefore prove to be useful targets for the development of novel treatments for neuropsychiatric disorders that involve cognitive dysfunction.
ESTHER : Rushforth_2010_Neurosci.Lett_471_114
PubMedSearch : Rushforth_2010_Neurosci.Lett_471_114
PubMedID: 20083163

Title : Associated proteins: The universal toolbox controlling ligand gated ion channel function - Araud_2010_Biochem.Pharmacol_80_160
Author(s) : Araud T , Wonnacott S , Bertrand D
Ref : Biochemical Pharmacology , 80 :160 , 2010
Abstract : Ligand gated ion channels are integral multimeric membrane proteins that can detect with high sensitivity the presence of a specific transmitter in the extracellular space and transduce this signal into an ion flux. While these receptors are widely expressed in the nervous system, their expression is not limited to neurons or their postsynaptic targets but extends to non-neuronal cells where they participate in many physiological responses. Cells have developed complex regulatory mechanisms allowing for the precise control and modulation of ligand gated ion channels. In this overview the roles of accessory subunits and associated proteins in these regulatory mechanisms are reviewed and their relevance illustrated by examples at different ligand gated ion channel types, with emphasis on nicotinic acetylcholine receptors. Dysfunction of ligand gated ion channels can result in neuromuscular, neurological or psychiatric disorders. A better understanding of the precise function of associated proteins and how they impact on ligand gated ion channels will provide new therapeutic opportunities for clinical intervention.
ESTHER : Araud_2010_Biochem.Pharmacol_80_160
PubMedSearch : Araud_2010_Biochem.Pharmacol_80_160
PubMedID: 20346921

Title : alpha7 and non-alpha7 nicotinic acetylcholine receptors modulate dopamine release in vitro and in vivo in the rat prefrontal cortex - Livingstone_2009_Eur.J.Neurosci_29_539
Author(s) : Livingstone PD , Srinivasan J , Kew JN , Dawson LA , Gotti C , Moretti M , Shoaib M , Wonnacott S
Ref : European Journal of Neuroscience , 29 :539 , 2009
Abstract : Nicotine enhances attentional and working memory aspects of executive function in the prefrontal cortex (PFC) where dopamine plays a major role. Here, we have determined the nicotinic acetylcholine receptor (nAChR) subtypes that can modulate dopamine release in rat PFC using subtype-selective drugs. Nicotine and 5-Iodo-A-85380 (beta2* selective) elicited [(3)H]dopamine release from both PFC and striatal prisms in vitro and dopamine overflow from medial PFC in vivo. Blockade by dihydro-beta-erythroidine supports the participation of beta2* nAChRs. However, insensitivity of nicotine-evoked [(3)H]dopamine release to alpha-conotoxin-MII in PFC prisms suggests no involvement of alpha6beta2* nAChRs, in contrast to the striatum, and this distinction is supported by immunoprecipitation of nAChR subunits from these tissues. The alpha7 nAChR-selective agonists choline and Compound A also promoted dopamine release from PFC in vitro and in vivo, and their effects were enhanced by the alpha7 nAChR-selective allosteric potentiator PNU-120596 and blocked by specific antagonists. DNQX and MK801 inhibited [(3)H]dopamine release evoked by choline and PNU-120596, suggesting crosstalk between alpha7 nAChRs, glutamate and dopamine in the PFC. In vivo, systemic (but not local) administration of PNU-120596, in the absence of agonist, facilitated dopamine overflow in the medial PFC, consistent with the activation of extracortical alpha7 nAChRs by endogenous acetylcholine or choline. These data establish that both beta2* and alpha7 nAChRs can modulate dopamine release in the PFC in vitro and in vivo. Through their distinct actions on dopamine release, these nAChR subtypes could contribute to executive function, making them specific therapeutic targets for conditions such as schizophrenia and attention deficit hyperactivity disorder.
ESTHER : Livingstone_2009_Eur.J.Neurosci_29_539
PubMedSearch : Livingstone_2009_Eur.J.Neurosci_29_539
PubMedID: 19187266

Title : Drug discrimination and neurochemical studies in alpha7 null mutant mice: tests for the role of nicotinic alpha7 receptors in dopamine release - Quarta_2009_Psychopharmacology.(Berl)_203_399
Author(s) : Quarta D , Naylor CG , Barik J , Fernandes C , Wonnacott S , Stolerman IP
Ref : Psychopharmacology (Berl) , 203 :399 , 2009
Abstract : RATIONALE: The nicotine discriminative stimulus has been linked to beta2-containing (beta2*) nicotinic receptors, with little evidence of a role for alpha7 nicotinic receptors, because nicotine discrimination was very weak in beta2 null mutant mice but normal in alpha7 mutants. OBJECTIVES: As both alpha7 and beta2* nicotinic receptors have been implicated in nicotine-stimulated dopamine overflow, this study focused on the dopamine-mediated element in the nicotine stimulus by examining cross-generalisation between amphetamine and nicotine. MATERIALS AND
METHODS: Male alpha7 nicotinic receptor null mutant mice and wild-type controls were bred in-house and trained to discriminate nicotine (0.8 mg/kg) or (+)-amphetamine (0.6 mg/kg) from saline in a two-lever procedure with a tandem VI-30 FR-10 schedule of food reinforcement. Dopamine release from striatal slices was determined in parallel experiments.
RESULTS: An alpha7 nicotinic receptor-mediated component of dopamine release was demonstrated in tissue from wild-type mice using choline as a selective agonist. This response was absent in tissue from null mutant animals. The mutation did not influence acquisition of drug discriminations but subtly affected the results of cross-generalisation tests. In mice trained to discriminate nicotine or amphetamine, there was partial cross-generalisation in wild-type mice and, at certain doses, these effects were attenuated in mutants. Further support for an alpha7 nicotinic receptor-mediated component was provided by the ability of the alpha7 nicotinic receptor antagonist methyllycaconitine to attenuate responses to nicotine and amphetamine in wild-type mice.
CONCLUSIONS: These findings support the concept of an alpha7 nicotinic receptor-mediated dopaminergic element in nicotine discrimination, warranting further tests with selective dopamine agonists.
ESTHER : Quarta_2009_Psychopharmacology.(Berl)_203_399
PubMedSearch : Quarta_2009_Psychopharmacology.(Berl)_203_399
PubMedID: 18758759

Title : Nicotinic acetylcholine receptors and the ascending dopamine pathways - Livingstone_2009_Biochem.Pharmacol_78(7)_744
Author(s) : Livingstone PD , Wonnacott S
Ref : Biochemical Pharmacology , 78 :744 , 2009
Abstract : Nicotinic acetylcholine receptors (nAChRs) are widely expressed in midbrain dopamine neurons that project to dorsal striatum, nucleus accumbens and prefrontal cortex. Thus nAChRs can influence the functions of these three pathways, notably motor control, 'reward' and executive function, respectively. Diverse subtypes of nAChRs have been identified on dopamine cell bodies and terminals as well as on neighbouring afferents and interneurons. Here we review the molecular and cellular mechanisms through which nAChRs exert their influence on these pathways in rodents.
ESTHER : Livingstone_2009_Biochem.Pharmacol_78(7)_744
PubMedSearch : Livingstone_2009_Biochem.Pharmacol_78(7)_744
PubMedID: 19523928

Title : Poster: Distinct pharmacological profiles for nicotinic AChR-evoked noradrenaline release in rat frontal cortex and hippocampus -
Author(s) : Kennett A , Heal D , Wonnacott S
Ref : Biochemical Pharmacology , 78 :915 , 2009

Title : Poster: Nicotinic acetylcholine receptors differentially regulate phosphorylation of dopamine target cell proteins in the rat prefrontal cortex -
Author(s) : Livingstone PD , Hockley J , Wonnacott S
Ref : Biochemical Pharmacology , 78 :909 , 2009

Title : Molecular and cellular mechanisms of action of nicotine in the CNS - Barik_2009_Handb.Exp.Pharmacol__173
Author(s) : Barik J , Wonnacott S
Ref : Handbook of Experimental Pharmacology , :173 , 2009
Abstract : Nicotine achieves its psychopharmacological effects by interacting with nicotinic acetylcholine receptors (nAChRs) in the brain. There are numerous subtypes of nAChR that differ in their properties, including their sensitivity to nicotine, permeability to calcium and propensity to desensitise. The nAChRs are differentially localised to different brain regions and are found on presynaptic terminals as well as in somatodendritic regions of neurones. Through their permeability to cations, these ion channel proteins can influence both neuronal excitability and cell signalling mechanisms, and these various responses can contribute to the development or maintenance of dependence. However, many questions and uncertainties remain in our understanding of these events and their relevance to tobacco addiction. In this chapter, we briefly overview the fundamental characteristics of nAChRs that are germane to nicotine's effects and then consider the cellular responses to acute and chronic nicotine, with particular emphasis on dopamine systems because they have been the most widely studied in the context of nicotine dependence. Where appropriate, methodological aspects are critically reviewed.
ESTHER : Barik_2009_Handb.Exp.Pharmacol__173
PubMedSearch : Barik_2009_Handb.Exp.Pharmacol__173
PubMedID: 19184650

Title : Gates and filters: unveiling the physiological roles of nicotinic acetylcholine receptors in dopaminergic transmission - Wonnacott_2008_Br.J.Pharmacol_153 Suppl 1_S2
Author(s) : Wonnacott S
Ref : British Journal of Pharmacology , 153 Suppl 1 :S2 , 2008
Abstract : Neuronal nicotinic acetylcholine receptors (nAChRs) in the brain have been enigmatic players on the cerebral stage. As ligand-gated ion channels they were expected to mediate fast cholinergic transmission, yet their influence appears to be modulatory. Two reviews in this issue of the BJP consider the relationship between nAChRs and endogenous ACh, with respect to the modulation of dopaminergic signalling. In his review, Maskos posits that in midbrain dopamine neurons, somatodendritic nAChRs activated by cholinergic inputs from the pedunculopontine tegmental nucleus (PPTg) and laterodorsal tegmental nucleus (LDTg) serve as a 'gate' that facilitates the switch to burst firing. In the terminal field, Exley and Cragg argue that nAChRs function as a 'presynaptic filter' to enhance the contrast between single and repetitive spike firing. Thus somatodendritic and presynaptic nAChRs exert subtle and complementary influences in responding to cholinergic inputs.
ESTHER : Wonnacott_2008_Br.J.Pharmacol_153 Suppl 1_S2
PubMedSearch : Wonnacott_2008_Br.J.Pharmacol_153 Suppl 1_S2
PubMedID: 18246098

Title : Smoking cessation 2008 -
Author(s) : Wonnacott S
Ref : IDrugs , 11 :256 , 2008
PubMedID: 18379957

Title : Presynaptic alpha 7- and beta 2-containing nicotinic acetylcholine receptors modulate excitatory amino acid release from rat prefrontal cortex nerve terminals via distinct cellular mechanisms - Dickinson_2008_Mol.Pharmacol_74_348
Author(s) : Dickinson JA , Kew JN , Wonnacott S
Ref : Molecular Pharmacology , 74 :348 , 2008
Abstract : Nicotine can enhance working memory and attention. Activation of both alpha7 and beta2(*) nicotinic acetylcholine receptors (nAChRs) in the prefrontal cortex (PFC) has been implicated in these processes. The ability of presynaptic nAChRs to modulate neurotransmitter release, notably glutamate release, is postulated to contribute to nicotine's effects. We have examined the cellular mechanisms underlying alpha7 and beta2(*) nAChR-mediated [(3)H]d-aspartate release from the PFC in vitro. Using the alpha7 and beta2(*) nAChR-selective agonists (R)-N-(1-azabicyclo[2.2.2]-oct-3-yl)(5-(2-pyridyl)thiophene-2-carboxamide) (compound A) and 5-iodo-3-(2(S)-azetidinylmethoxy)pyridine (5-iodo-A-85380), respectively, in conjunction with inhibitors of voltage-operated Ca(2+) channels (VOCCs) and intracellular Ca(2+) stores, we show that [(3)H]d-aspartate release evoked by activation of beta2(*) nAChRs occurs via VOCCs. In contrast, alpha7 nAChR-evoked release was unaffected by VOCC blockers but was abolished by modulators of Ca(2+) stores, including ryanodine. The alpha7 nAChR ligand alpha-bungarotoxin and ryanodine receptors were colocalized to a subpopulation of PFC synaptosomes. Compound A-evoked [(3)H]d-aspartate release was also blocked by mitogen-activated protein kinase kinase 1 inhibitors, implicating extracellular signal-regulated kinase (ERK)1/2 in alpha7 nAChR-evoked exocytosis. Western blotting confirmed that compound A, but not 5-iodo-A-85380, application increased ERK2 phosphorylation in PFC synaptosomes, and this was dependent on ryanodine-sensitive stores. Compound A also promoted synapsin-1 phosphorylation at ERK1/2-dependent sites, in a ryanodine-sensitive manner. Thus, beta2(*) and alpha7 nAChR subtypes in the PFC mediate [(3)H]d-aspartate release via distinct mechanisms as a result of their differential coupling to VOCCs and Ca(2+)-induced Ca(2+) release (CICR), respectively. The ability of alpha7 nAChRs to promote the phosphorylation of presynaptic ERK2 and synapsin-1, downstream of CICR, provides a potential mechanism for presynaptic facilitation in the PFC.
ESTHER : Dickinson_2008_Mol.Pharmacol_74_348
PubMedSearch : Dickinson_2008_Mol.Pharmacol_74_348
PubMedID: 18445710

Title : Alpha-conotoxin Arenatus IB[V11L,V16D] [corrected] is a potent and selective antagonist at rat and human native alpha7 nicotinic acetylcholine receptors - Innocent_2008_J.Pharmacol.Exp.Ther_327_529
Author(s) : Innocent N , Livingstone PD , Hone A , Kimura A , Young T , Whiteaker P , McIntosh JM , Wonnacott S
Ref : Journal of Pharmacology & Experimental Therapeutics , 327 :529 , 2008
Abstract : A recently developed alpha-conotoxin, alpha-conotoxin Arenatus IB-[V11L,V16D] (alpha-CtxArIB[V11L,V16D]) [corrected], is a potent and selective competitive antagonist at rat recombinant alpha7 nicotinic acetylcholine receptors (nAChRs), making it an attractive probe for this receptor subtype. alpha7 nAChRs are potential therapeutic targets that are widely expressed in both neuronal and non-neuronal tissues, where they are implicated in a variety of functions. In this study, we evaluate this toxin at rat and human native nAChRs. Functional alpha7 nAChR responses were evoked by choline plus the allosteric potentiator PNU-120596 [1-(5-chloro-2,4-dimethoxy-phenyl)-3-(5-methyl-isoxazol-3-yl)-urea] in rat PC12 cells and human SH-SY5Y cells loaded with calcium indicators. alpha-CtxArIB[V11L,V16D] specifically inhibited alpha7 nAChR-mediated increases in Ca2+ in PC12 cells. Responses to other stimuli, 5-I-A-85380 [5-iodo-3-(2(S)-azetidinylmethoxy)pyridine dihydrochloride], nicotine, or KCl, that did not activate alpha7 nAChRs were unaffected. Human alpha7 nAChRs were also sensitive to alpha-CtxArIB[V11L, V16D]; acetylcholine-evoked currents in Xenopus laevis oocytes expressing human alpha7 nAChRs were inhibited by alpha-CtxArIB[V11L,V16D] (IC(50), 3.4 nM) in a slowly reversible manner, with full recovery taking 15 min. This is consistent with the time course of recovery from blockade of rat alpha7 nAChRs in PC12 cells. alpha-CtxArIB[V11L,V16D] inhibited human native alpha7 nAChRs in SHSY5Y cells, activated by either choline or AR-R17779 [(2)-spiro[1-azabicyclo[2.2.2]octane-3,59-oxazolidin]-29-one] plus PNU-120596. Rat brain alpha7 nAChRs contribute to dopamine release from striatal minces; alpha-CtxArIB[V11L,V16D] (300 nM) selectively inhibited choline-evoked dopamine release without affecting responses evoked by nicotine that activates heteromeric nAChRs. This study establishes that alpha-CtxArIB[V11L,V16D] selectively inhibits human and rat native alpha7 nAChRs with comparable potency, making this a potentially useful antagonist for investigating alpha7 nAChR functions.
ESTHER : Innocent_2008_J.Pharmacol.Exp.Ther_327_529
PubMedSearch : Innocent_2008_J.Pharmacol.Exp.Ther_327_529
PubMedID: 18664588

Title : In vivo modulation of dopaminergic nigrostriatal pathways by cytisine derivatives: implications for Parkinson's Disease - Abin-Carriquiry_2008_Eur.J.Pharmacol_589_80
Author(s) : Abin-Carriquiry JA , Costa G , Urbanavicius J , Cassels BK , Rebolledo-Fuentes M , Wonnacott S , Dajas F
Ref : European Journal of Pharmacology , 589 :80 , 2008
Abstract : Nicotinic acetylcholine receptor agonists are considered potential pharmacological agents for Parkinson's disease treatment, due to their ability to improve experimental Parkinson symptomatology, reduce 3,4-dihydroxy-L-phenylalanine-induced dyskinesias and stop the neurodegenerative process at an experimental level. In the present work, the ability of the nicotinic agonist cytisine and two halogenated derivatives (3-bromocytisine and 5-bromocytisine) to induce striatal dopamine release was characterized in vivo by microdialysis. Cytisine, 5-bromocytisine and nicotine were much more efficacious than 3-bromocytisine in eliciting dopamine release in response to their local application through the microdialysis probe. Moreover, the agonists were intermittently administered before and after an intranigral injection of 6-hydroxydopamine (6-OHDA), and striatal dopamine tissue levels were assessed 8 days after the lesion. Both cytisine and its 5-bromo derivative (but not the 3-bromo derivative) significantly prevented the decrease of striatal dopamine tissue levels induced by 6-OHDA. These results suggest that the efficacy of nicotinic agonists to stimulate dopamine release in vivo through presynaptic nicotinic receptors could be related to their potential to induce striatal protection.
ESTHER : Abin-Carriquiry_2008_Eur.J.Pharmacol_589_80
PubMedSearch : Abin-Carriquiry_2008_Eur.J.Pharmacol_589_80
PubMedID: 18589414

Title : Differential coupling of alpha7 and non-alpha7 nicotinic acetylcholine receptors to calcium-induced calcium release and voltage-operated calcium channels in PC12 cells - Dickinson_2007_J.Neurochem_100_1089
Author(s) : Dickinson JA , Hanrott KE , Mok MH , Kew JN , Wonnacott S
Ref : Journal of Neurochemistry , 100 :1089 , 2007
Abstract : Neuronal nicotinic acetylcholine receptors (nAChRs) are ligand-gated cation channels that can modulate various neuronal processes by altering intracellular Ca(2+) levels. Following nAChR stimulation Ca(2+) can enter cells either directly, through the intrinsic ion channel, or indirectly following voltage-operated Ca(2+) channel (VOCC) activation; Ca(2+) levels can subsequently be amplified via Ca(2+)-induced Ca(2+) release from intracellular stores. We have used subtype-selective nAChR agonists to investigate the Ca(2+) sources contributing to alpha7 and non-alpha7 nAChR-mediated increases in intracellular Ca(2+) in PC12 cells. Application of the alpha7 nAChR positive allosteric modulator PNU 120596 (10 mum), in conjunction with the alpha7 nAChR agonist, compound A [(R)-N-(1-azabicyclo[2.2.2]oct-3-yl)(5-(2-pyridyl)thiophene-2-carboxamide), 10 nm], produces a rapid increase in fluo-3 fluorescence that is prevented by the selective alpha7 nAChR antagonist alpha-bungarotoxin. The non-alpha7 nAChR agonist 5-Iodo-A-85380 produces alpha-bungarotoxin-insensitive increases in intracellular Ca(2+) (EC(50) = 11.2 mum). Using these selective agonists or KCl in conjunction with general and selective VOCC inhibitors, we demonstrate that the primary route of Ca(2+) entry following either non-alpha7 nAChR activation or KCl stimulation is via L-type VOCCs. In contrast, the alpha7 nAChR-mediated response is unaffected by VOCC blockers but is inhibited by modulators of intracellular Ca(2+) stores. These results indicate that alpha7 and non-alpha7 nAChRs are differentially coupled to Ca(2+)-induced Ca(2+) release and VOCCs, respectively.
ESTHER : Dickinson_2007_J.Neurochem_100_1089
PubMedSearch : Dickinson_2007_J.Neurochem_100_1089
PubMedID: 17181555

Title : Nicotine induces tyrosine hydroxylase plasticity in the neurodegenerating striatum - Urbanavicius_2007_J.Neurochem_102_723
Author(s) : Urbanavicius J , Ferreira M , Costa G , Abin-Carriquiry JA , Wonnacott S , Dajas F
Ref : Journal of Neurochemistry , 102 :723 , 2007
Abstract : It has been shown that nicotine prevents the loss of dopamine (DA) in the corpus striatum (CS) after 6-hydroxydopamine injection in the substantia nigra. To study the role of the enzyme tyrosine hydroxylase (TH; EC in this experimental paradigm, we have examined its activity by assessing the accumulation of l-3,4-dihydroxyphenylalanine after inhibiting the subsequent enzyme in the DA synthetic pathway, aromatic l-amino acid decarboxylase, with 3-hydroxybenzylhydrazine. In addition the amount of TH protein was assessed by western blotting and its distribution in the CS was examined using immunohistochemical methods. 6-hydroxydopamine injection produced a significant decrease in DA levels and l-3,4-dihydroxyphenylalanine accumulation, as well as decreases in TH protein and TH immunoreactive fibres in the CS. After nicotine treatment, the decrease in TH protein in the CS was significantly reduced, with a concomitant preservation of TH activity, but nicotine did not alter the number of TH immunoreactive fibres. The activity and amount of TH did not change in the contralateral (intact) CS. Thus, nicotine induces long lasting TH plasticity in the degenerating CS. A synergistic action of nicotine-activated and lesion-originated signals appears necessary for the expression of this neuronal molecular plasticity.
ESTHER : Urbanavicius_2007_J.Neurochem_102_723
PubMedSearch : Urbanavicius_2007_J.Neurochem_102_723
PubMedID: 17437548

Title : Differential effects of acute and chronic nicotine on Elk-1 in rat hippocampus - Nuutinen_2007_Neuroreport_18_121
Author(s) : Nuutinen S , Barik J , Jones IW , Wonnacott S
Ref : Neuroreport , 18 :121 , 2007
Abstract : Transcriptional regulation is central to the long-term effects of drugs of abuse. Activation of the extracellular signal-regulated kinase (ERK1/2) pathway underlies plasticity changes that accompany drug use. One target of ERK1/2 activation is the Ets-like transcription factor Elk-1. We show here that nicotine modulates Elk-1 in the rat hippocampus in a spatially and temporally specific manner. In-vitro nicotine (1 muM) activated Elk-1 in hippocampal slices. In-vivo acute nicotine (0.4 mg/kg) activated Elk-1 in the CA1 area but not in the dentate gyrus. Chronic nicotine for 14 days changed the level of total Elk-1 but not its phosphorylation state. Thus, Elk-1 regulation of transcriptional events may contribute to nicotine-induced changes in the hippocampus.
ESTHER : Nuutinen_2007_Neuroreport_18_121
PubMedSearch : Nuutinen_2007_Neuroreport_18_121
PubMedID: 17301675

Title : Development of an anti-cotinine vaccine to potentiate nicotine-based smoking cessation strategies - Oliver_2007_Vaccine_25_7354
Author(s) : Oliver JL , Pashmi G , Barnett P , Mettens P , Biemans R , Monteyne P , Palmantier R , Gallagher T , Ramaya S , Wonnacott S
Ref : Vaccine , 25 :7354 , 2007
Abstract : Nicotine replacement therapies (NRT) have limited success in smoking cessation. The efficacy of nicotine may be compromised by its main metabolite, cotinine. An anti-cotinine vaccine to remove this antagonism could enhance the efficacy of NRT. We show that cotinine is a weak nicotinic agonist and decreases responses to nicotine, consistent with antagonism through receptor desensitisation. trans-4-Thiol cotinine was coupled to tetanus toxoid, and rats immunised repeatedly. Vaccination raised antibodies specific for cotinine that do not recognise other metabolites or nicotine. Increased serum cotinine concentrations following nicotine administration indicate sequestration of cotinine by antibodies, encouraging further evaluation of this vaccine in behavioural models of nicotine addiction and relapse.
ESTHER : Oliver_2007_Vaccine_25_7354
PubMedSearch : Oliver_2007_Vaccine_25_7354
PubMedID: 17870213

Title : Comparison of the effects of bupropion and nicotine on locomotor activation and dopamine release in vivo - Sidhpura_2007_Biochem.Pharmacol_74(8)_1292
Author(s) : Sidhpura N , Redfern P , Rowley H , Heal D , Wonnacott S
Ref : Biochemical Pharmacology , 74 :1292 , 2007
Abstract : Bupropion is an atypical anti-depressant that is approved for smoking cessation. In addition to inhibiting dopamine reuptake, bupropion has been reported to block nicotinic acetylcholine receptors in vitro, and this action might contribute to its efficacy for smoking cessation. In this study we investigated if nicotinic receptor-mediated responses in vivo are decreased in the presence of a behaviorally effective dose of bupropion. In separate experiments we measured locomotor activation and dopamine overflow in the nucleus accumbens core, using in vivo microdialysis in freely moving rats. Bupropion (30 mg/kg i.p.) increased locomotor activity, which remained elevated for up to 2 h. Nicotine (0.4 mg/kg s.c.) also increased locomotor activity but for a shorter duration. When given 20 min after bupropion, hyperlocomotion was significantly enhanced, compared to the response to either nicotine or bupropion alone, consistent with the effects of the two drugs being additive. Systemic administration of bupropion (30 mg/kg i.p.) also elicited a significant increase in dopamine overflow (113+/-16% above basal levels). Nicotine (3 mM; delivered into the nucleus accumbens core via the microdialysis probe) increased dopamine overflow by 126 +/- 35%. Nicotine delivered during the response to bupropion resulted in enhanced dopamine overflow of 294 +/- 50%, also consistent with the actions of the two drugs being additive. This study suggests that behaviorally effective concentrations of bupropion in the rat do not diminish the effects of nicotine by blocking nicotinic receptors.
ESTHER : Sidhpura_2007_Biochem.Pharmacol_74(8)_1292
PubMedSearch : Sidhpura_2007_Biochem.Pharmacol_74(8)_1292
PubMedID: 17678630

Title : Comparison of the effects of bupropion on nicotinic receptor-evoked [(3)H]dopamine release from rat striatal synaptosomes and slices - Sidhpura_2007_Eur.J.Pharmacol_567_102
Author(s) : Sidhpura N , Redfern P , Wonnacott S
Ref : European Journal of Pharmacology , 567 :102 , 2007
Abstract : Tobacco smoking is a nicotine addiction, mediated in part by the ability of nicotine to elicit dopamine release, as a result of the stimulation of nicotinic acetylcholine receptors associated with dopaminergic pathways. The smoking cessation agent bupropion is an inhibitor of the dopamine transporter, but has also been shown to inhibit nicotinic acetylcholine receptors. To assess the relative impact of its actions at these two targets, we have examined the effects of bupropion on nicotine-evoked [(3)H]dopamine release from rat striatal synaptosomes and slices, in the absence of any other transporter inhibitor. Bupropion (10 microM) significantly decreased nicotine-evoked [(3)H]dopamine release by approximately 50% in both preparations, consistent with the blockade of nicotinic receptors. In support of this interpretation, bupropion also selectively inhibited nicotine-evoked Ca(2+) increases in SH-SY5Y cells. In striatal slices (but not in synaptosomes) the concentration-response profile for bupropion has an inverted 'u' shape, with a decrease in nicotine-evoked [(3)H]dopamine release also observed in the presence of 0.1 microM bupropion. This effect of 0.1 microM bupropion (but not 10 microM bupropion) was reversed by the dopamine D(2) receptor antagonist raclopride. We propose that modest blockade of the dopamine transporter by low concentrations of bupropion results in feedback inhibition via dopamine D(2) autoreceptors. This is overcome at higher concentrations of bupropion, before inhibition of nicotinic receptors occurs. Therefore bupropion's inhibition of the dopamine transporter and nicotinic receptors appears to be separated with respect to concentration.
ESTHER : Sidhpura_2007_Eur.J.Pharmacol_567_102
PubMedSearch : Sidhpura_2007_Eur.J.Pharmacol_567_102
PubMedID: 17477919

Title : 6-hydroxydopamine-induced apoptosis is mediated via extracellular auto-oxidation and caspase 3-dependent activation of protein kinase Cdelta - Hanrott_2006_J.Biol.Chem_281_5373
Author(s) : Hanrott K , Gudmunsen L , O'Neill MJ , Wonnacott S
Ref : Journal of Biological Chemistry , 281 :5373 , 2006
Abstract : 6-Hydroxydopamine is a neurotoxin commonly used to lesion dopaminergic pathways and generate experimental models for Parkinson disease, however, the cellular mechanism of 6-hydroxydopamine-induced neurodegeneration is not well defined. In this study we have explored how 6-hydroxydopamine neurotoxicity is initiated. We have also investigated downstream signaling pathways activated in response to 6-hydroxydopamine, using a neuronal-like, catecholaminergic cell line (PC12 cells) as an in vitro model system. We have shown that 6-hydroxydopamine neurotoxicity is initiated via extracellular auto-oxidation and the induction of oxidative stress from the oxidative products generated. Neurotoxicity is completely attenuated by preincubation with catalase, suggesting that hydrogen peroxide, at least in part, evokes neuronal cell death in this model. 6-Hydroxydopamine does not initiate toxicity by dopamine transporter-mediated uptake into PC12 cells, because both GBR-12909 and nisoxetine (inhibitors of dopamine and noradrenaline transporters, respectively) failed to reduce toxicity. 6-Hydroxydopamine has previously been shown to induce both apoptotic and necrotic cell-death mechanisms. In this study oxidative stress initiated by 6-hydroxydopamine caused mitochondrial dysfunction, activation of caspases 3/7, nuclear fragmentation, and apoptosis. We have shown that, in this model, proteolytic activation of the proapoptotic protein kinase Cdelta (PKCdelta) is a key mediator of 6-hydroxydopamine-induced cell death. 6-Hydroxydopamine induces caspase 3-dependent cleavage of full-length PKCdelta (79 kDa) to yield a catalytic fragment (41 kDa). Inhibition of PKCdelta (with rottlerin or via RNA interference-mediated gene suppression) ameliorates the neurotoxicity evoked by 6-hydroxydopamine, implicating this kinase in 6-hydroxydopamine-induced neurotoxicity and Parkinsonian neurodegeneration.
ESTHER : Hanrott_2006_J.Biol.Chem_281_5373
PubMedSearch : Hanrott_2006_J.Biol.Chem_281_5373
PubMedID: 16361258

Title : Nicotinic receptors modulate transmitter cross talk in the CNS: nicotinic modulation of transmitters - Wonnacott_2006_J.Mol.Neurosci_30_137
Author(s) : Wonnacott S , Barik J , Dickinson J , Jones IW
Ref : Journal of Molecular Neuroscience , 30 :137 , 2006
Abstract : Neuronal nicotinic acetylcholine receptors (nAChRs) in the CNS appear to exert a predominantly modulatory influence on brain mechanisms, despite being fast-acting ligand-gated ion channels. Many nAChRs have an extrasynaptic location on somatodendritic regions or presynaptic terminals. They influence local excitability by depolarization and can initiate short- and long-term changes by interfacing with Ca2+ signaling pathways (Dajas- Bailador and Wonnacott, 2004). The modulation of neurotransmitter release by presynaptic nAChRs is well-documented (Wonnacott, 1997): Both Na+ and Ca2+ fluxes associated with nAChR activation can influence transmitter release. It is also emerging that nAChRs, especially the alpha7 subtype, can exert an indirect effect on transmitter release, through modulation of amino acid transmitters. This complex scenario facilitates transmitter cross talk, which is the subject of this short review.
ESTHER : Wonnacott_2006_J.Mol.Neurosci_30_137
PubMedSearch : Wonnacott_2006_J.Mol.Neurosci_30_137
PubMedID: 17192660

Title : The synthesis and nicotinic binding activity of (+\/-)-epiquinamide and (+\/-)-C(1)-epiepiquinamide - Kanakubo_2006_Bioorg.Med.Chem.Lett_16_4648
Author(s) : Kanakubo A , Gray D , Innocent N , Wonnacott S , Gallagher T
Ref : Bioorganic & Medicinal Chemistry Lett , 16 :4648 , 2006
Abstract : The synthesis of (+/-)-epiquinamide 1 and (+/-)-C(1)-epiepiquinamide 2 based on the use of a Curtius rearrangement to introduce the C(1) amino residue is reported. In a competition binding assay for [(3)H]epibatidine binding to rat brain membranes neither (+/-)-1 nor (+/-)-2 showed any significant level of nicotinic activity.
ESTHER : Kanakubo_2006_Bioorg.Med.Chem.Lett_16_4648
PubMedSearch : Kanakubo_2006_Bioorg.Med.Chem.Lett_16_4648
PubMedID: 16784846

Title : Alpha7 nicotinic acetylcholine receptor expression in Alzheimer's disease: receptor densities in brain regions of the APP(SWE) mouse model and in human peripheral blood lymphocytes - Jones_2006_J.Mol.Neurosci_30_83
Author(s) : Jones IW , Westmacott A , Chan E , Jones RW , Dineley K , O'Neill MJ , Wonnacott S
Ref : Journal of Molecular Neuroscience , 30 :83 , 2006
Abstract : The brains of people with Alzheimer's disease (AD) display several characteristic pathological features, including deposits (plaques) of beta-amyloid 1-42 (Abeta1-42), intraneuronal accumulations (tangles) of hyperphosphorylated tau, degeneration of the basal forebrain cholinergic pathway, and gliosis. Abeta1-42 plaques develop in specific brain regions, including hippocampus and cortex, as well as in the vasculature. Abeta1-42 might promote neurodegeneration through the induction of free radicals and disruption of Ca2+ homeostasis, giving rise to the symptoms of AD. Abeta1-42 interacts with the alpha7 subtype of the nicotinic acetylcholine receptor (alpha7 nAChR), which is widely expressed throughout the central and peripheral nervous systems, as well as in several nonneuronal loci, such as epithelial cells, lymphoid tissues, and peripheral blood lymphocytes. Western blot and autoradiographic analyses indicate that the alpha7 nAChR subunit protein is up-regulated in human brain samples from Alzheimer patients, as well as in animal models of AD (Dineley et al., 2001; Bednar et al., 2002), and might be involved in nicotine-mediated reduction of Abeta1-42 deposition (Hellstrom et al., 2004), although the nature of this relationship remains ill-defined. We have undertaken a semiquantitative histological evaluation of alpha7 nAChR expression in a mouse model of AD pathology, as well as a comparison of alpha7 nAChR levels in lymphocytes from AD patients and control subjects.
ESTHER : Jones_2006_J.Mol.Neurosci_30_83
PubMedSearch : Jones_2006_J.Mol.Neurosci_30_83
PubMedID: 17192639

Title : Indirect modulation by alpha7 nicotinic acetylcholine receptors of noradrenaline release in rat hippocampal slices: interaction with glutamate and GABA systems and effect of nicotine withdrawal - Barik_2006_Mol.Pharmacol_69_618
Author(s) : Barik J , Wonnacott S
Ref : Molecular Pharmacology , 69 :618 , 2006
Abstract : Nicotinic acetylcholine receptors (nAChRs) can modulate transmitter release. Striatal [(3)H]dopamine ([(3)H]DA) release is regulated by presynaptic nAChR on dopaminergic terminals and alpha7 nAChR on neighboring glutamatergic afferents. Here, we explored the role of alpha7 nAChR in the modulation of [(3)H]noradrenaline ([(3)H]NA) release from rat hippocampal slices. The nicotinic agonist anatoxin-a (AnTx) evoked monophasic [(3)H]NA release (EC(50) = 1.2 microM) that was unaffected by alpha-conotoxin-MII or dihydro-beta-erythroidine, antagonists of alpha3/alpha6beta2* and beta2* nAChR, respectively. In contrast AnTx-evoked striatal [(3)H]DA release was biphasic (EC(50) = 138.9 nM; 7.1 microM) and blocked by these antagonists. At a high AnTx concentration (25 microM), alpha7 nAChR antagonists (methyllycaconitine, alpha-conotoxin-ImI) and glutamate receptor (GluR) antagonists [kynurenic acid, 6,7-dinitroquinoxaline-2,3-dione (DNQX)] partially inhibited [(3)H]NA release. The alpha7 nAChR-selective agonist choline evoked [(3)H]NA release (E(max) = 33% of that of AnTx) that was blocked by GluR antagonists, supporting a model in which alpha7 nAChRs trigger glutamate release that subsequently stimulates [(3)H]NA release. A GABAergic component was also revealed: choline-evoked [(3)H]NA release was partially blocked by the GABA(A) receptor antagonist bicuculline, and coapplication of bicuculline and DNQX fully abolished this response. These findings support alpha7 nAChR on GABAergic neurons that can promote GABA release which, in turn, leads to [(3)H]NA release, probably by disinhibition. To investigate the impact of long-term nicotine exposure on this model, rats were exposed for 14 days to nicotine (4 mg/kg/day) with or without 3 or 7 days of withdrawal. alpha7 nAChR responses were selectively and transiently up-regulated after 3 days of withdrawal. This functional up-regulation could contribute to the withdrawal effects of nicotine.
ESTHER : Barik_2006_Mol.Pharmacol_69_618
PubMedSearch : Barik_2006_Mol.Pharmacol_69_618
PubMedID: 16269536

Title : C3-halogenation of cytisine generates potent and efficacious nicotinic receptor agonists - Abin-Carriquiry_2006_Eur.J.Pharmacol_536_1
Author(s) : Abin-Carriquiry JA , Voutilainen MH , Barik J , Cassels BK , Iturriaga-Vasquez P , Bermudez I , Durand C , Dajas F , Wonnacott S
Ref : European Journal of Pharmacology , 536 :1 , 2006
Abstract : Neuronal nicotinic acetylcholine receptors subserve predominantly modulatory roles in the brain, making them attractive therapeutic targets. Natural products provide key leads in the quest for nicotinic receptor subtype-selective compounds. Cytisine, found in Leguminosae spp., binds with high affinity to alpha4beta2* nicotinic receptors. We have compared the effect of C3 and C5 halogenation of cytisine and methylcytisine (MCy) on their interaction with native rat nicotinic receptors. 3-Bromocytisine (3-BrCy) and 3-iodocytisine (3-ICy) exhibited increased binding affinity (especially at alpha7 nicotinic receptors; Ki approximately 0.1 microM) and functional potency, whereas C5-halogenation was detrimental. 3-BrCy and 3-ICy were more potent than cytisine at evoking [3H]dopamine release from striatal slices (EC50 approximately 11 nM), [3H]noradrenaline release from hippocampal slices (EC50 approximately 250 nM), increases in intracellular Ca2+ in PC12 cells and inward currents in Xenopus oocytes expressing human alpha3beta4 nicotinic receptor (EC50 approximately 2 microM). These compounds were also more efficacious than cytisine. C3-halogenation of cytisine is proposed to stabilize the open conformation of the nicotinic receptor but does not enhance subtype selectivity.
ESTHER : Abin-Carriquiry_2006_Eur.J.Pharmacol_536_1
PubMedSearch : Abin-Carriquiry_2006_Eur.J.Pharmacol_536_1
PubMedID: 16563372

Title : Cellular responses to nicotinic receptor activation are decreased after prolonged exposure to galantamine in human neuroblastoma cells - Barik_2005_Br.J.Pharmacol_145_1084
Author(s) : Barik J , Dajas-Bailador F , Wonnacott S
Ref : British Journal of Pharmacology , 145 :1084 , 2005
Abstract : In this study, we have examined cellular responses of neuroblastoma SH-SY5Y cells after chronic treatment with galantamine, a drug used to treat Alzheimer's disease that has a dual mechanism of action: inhibition of acetylcholinesterase and allosteric potentiation of nicotinic acetylcholine receptors (nAChR). Acute experiments confirmed that maximum potentiation of nicotinic responses occurs at 1 microM galantamine; hence this concentration was chosen for chronic treatment. Exposure to 1 microM galantamine for 4 days decreased Ca(2+) responses (by 19.8+/-3.6%) or [(3)H]noradrenaline ([(3)H]NA) release (by 23.9+/-3.3%) elicited by acute application of nicotine. KCl-evoked increases in intracellular Ca(2+) were also inhibited by 10.0+/-1.9% after 4 days' treatment with galantamine. These diminished responses are consistent with the downregulation of downstream cellular processes. Ca(2+) responses evoked by activation of muscarinic acetylcholine receptors were unaffected by chronic galantamine treatment. Exposure to the more potent acetylcholinesterase inhibitor rivastigmine (1 microM) for 4 days failed to alter nicotine-, KCl-, or muscarinic receptor-evoked increases in intracellular Ca(2+). These observations support the hypothesis that chronic galantamine exerts its effects through interaction with nAChR in this cell line. Exposure to 10 microM nicotine for 4 days produced decreases in acute nicotine- (18.0+/-3.5%) and KCl-evoked Ca(2+) responses (10.6+/-2.5%) and nicotine-evoked [(3)H]NA release (26.0+/-3.3%) that are comparable to the effects of a corresponding exposure to galantamine. Treatment with 1 microM galantamine did not alter numbers of [(3)H]epibatidine-binding sites in SH-SY5Y cells, in contrast to 62% upregulation of these sites in response to 10 microM nicotine. Thus, chronic galantamine acts at nAChR to decrease subsequent functional responses to acute stimulation with nicotine or KCl. This effect appears to be independent of the upregulation of nAChR-binding sites.
ESTHER : Barik_2005_Br.J.Pharmacol_145_1084
PubMedSearch : Barik_2005_Br.J.Pharmacol_145_1084
PubMedID: 15937519

Title : Nicotine: from molecular mechanisms to behaviour - Wonnacott_2005_Curr.Opin.Pharmacol_5_53
Author(s) : Wonnacott S , Sidhpura N , Balfour DJ
Ref : Curr Opin Pharmacol , 5 :53 , 2005
Abstract : The addictive potential of nicotine is clearly recognized by the tenacity of tobacco smoking for most users, and has prompted extensive psychopharmacological studies in animals. In parallel, the interaction of nicotine with the many subtypes of its eponymous receptor has been the focus of molecular and cellular investigations. More recently, a convergence of these approaches has been stimulated by the generation of transgenic animals, which facilitates analysis of the impact of molecular changes on behaviour. Nicotine, like other addictive drugs including psychomotor stimulants, promotes dopamine release in the nucleus accumbens. This transmitter system has been a major focus of both neurochemical and behavioural investigations, although recently the pre-eminence of this system in nicotine dependence has been challenged. Complexities in the brain circuitry (including the subdivisions of the nucleus accumbens) and differences between behavioural models help to rationalise the current controversy.
ESTHER : Wonnacott_2005_Curr.Opin.Pharmacol_5_53
PubMedSearch : Wonnacott_2005_Curr.Opin.Pharmacol_5_53
PubMedID: 15661626

Title : From ligand design to therapeutic efficacy: the challenge for nicotinic receptor research - Cassels_2005_Drug.Discov.Today_10_1657
Author(s) : Cassels BK , Bermudez I , Dajas F , Abin-Carriquiry JA , Wonnacott S
Ref : Drug Discov Today , 10 :1657 , 2005
Abstract : S-Nicotine, the principal psychoactive constituent of Nicotiana tabacum, underpins addiction to tobacco smoking. Although tobacco consumption is a leading cause of death worldwide, nicotine itself is also proposed to have potential therapeutic benefits for a diverse range of conditions. Nicotine interacts with its cognate receptors in the central nervous system to exert a predominantly modulatory influence, making neuronal nicotinic receptors attractive therapeutic targets. Here, we focus on three natural products as lead compounds for drug discovery programs, nicotine, epibatidine and cytisine, and consider the aims and limitations that shape these drug discovery endeavors.
ESTHER : Cassels_2005_Drug.Discov.Today_10_1657
PubMedSearch : Cassels_2005_Drug.Discov.Today_10_1657
PubMedID: 16376826

Title : Why doesn't nicotinic ACh receptor immunoreactivity knock out? - Jones_2005_Trends.Neurosci_28_343
Author(s) : Jones IW , Wonnacott S
Ref : Trends in Neurosciences , 28 :343 , 2005
Abstract : Immunochemical analyses of protein expression and localization rely on the specificity of primary immunoreagents. A recent report, using transgenic mice, casts doubt on the specificity of three antibodies commonly used to immunolocalize alpha7 nicotinic ACh receptors. These data highlight the conundrum facing histologists--how 'real' is the labelling they see?
ESTHER : Jones_2005_Trends.Neurosci_28_343
PubMedSearch : Jones_2005_Trends.Neurosci_28_343
PubMedID: 15979499

Title : Presynaptic alpha7 and non-alpha7 nicotinic acetylcholine receptors modulate [3H]d-aspartate release from rat frontal cortex in vitro - Rousseau_2005_Neuropharmacol_49_59
Author(s) : Rousseau SJ , Jones IW , Pullar IA , Wonnacott S
Ref : Neuropharmacology , 49 :59 , 2005
Abstract : The presynaptic nicotinic modulation of glutamatergic transmission in the CNS has been associated with activation of the alpha7 subtype of nicotinic acetylcholine receptor (nAChR) in sub-cortical regions, whereas in the frontal cortex, non-alpha7 nAChRs have been implicated. The aim of this investigation was to directly characterise nAChR-evoked release of excitatory amino acids from rat frontal cortex, by monitoring the release of [3H]D-aspartate from superfused synaptosomes or minces. Co-administration of a nAChR agonist with a depolarising stimulus enhanced [3H]D-aspartate release above the effect of depolarising agent alone. This enhancement was blocked by the nicotinic antagonist mecamylamine. Other experiments revealed that in the absence of a depolarising stimulus, the nAChR agonists nicotine, epibatidine and anatoxin-a could evoke the release of [3H]D-aspartate in a Ca2+- and concentration-dependant manner. Differential sensitivity to the alpha7- and beta2*-selective nAChR antagonists alpha-bungarotoxin (alpha-Bgt) and dihydro-beta-erythroidine (DHbetaE) implicated two nAChR subtypes (alpha7 and beta2*), and this was supported by using the subtype-selective agonists choline (10 mM; alpha7 selective, blocked by alpha-Bgt but not by DHbetaE) and 5-Iodo-A-85380 (10 nM; beta2*-selective, blocked by DHbetaE but not by alpha-Bgt). Immunocytochemistry showed that alpha-Bgt labelling was associated with structures immunopositive for vesicular glutamate transporters, in both frontal cortex sections and synaptosome preparations, supporting the presence of alpha7 nAChR on glutamatergic terminals in rat frontal cortex.
ESTHER : Rousseau_2005_Neuropharmacol_49_59
PubMedSearch : Rousseau_2005_Neuropharmacol_49_59
PubMedID: 15992581

Title : Nicotinic acetylcholine receptors and the regulation of neuronal signalling - Dajas-Bailador_2004_Trends.Pharmacol.Sci_25_317
Author(s) : Dajas-Bailador F , Wonnacott S
Ref : Trends in Pharmacological Sciences , 25 :317 , 2004
Abstract : Neuronal nicotinic acetylcholine (nACh) receptors in the brain are more commonly associated with modulatory events than mediation of synaptic transmission. nACh receptors have a high permeability for Ca(2+), and Ca(2+) signals are pivotal in shaping nACh receptor-mediated neuromodulatory effects. In this review, we consider the mechanisms through which nACh receptors convert rapid ionic signals into sustained, wide-ranging phenomena. The complex Ca(2+) responses that are generated after activation of nACh receptors can transmit information beyond the initial domain and facilitate the interface with many intracellular processes. These mechanisms underlie the diverse repertoire of neuronal activities of nicotine in the brain, from the enhancement of learning and memory, to addiction and neuroprotection.
ESTHER : Dajas-Bailador_2004_Trends.Pharmacol.Sci_25_317
PubMedSearch : Dajas-Bailador_2004_Trends.Pharmacol.Sci_25_317
PubMedID: 15165747

Title : Alpha bungarotoxin-1.4 nm gold: a novel conjugate for visualising the precise subcellular distribution of alpha 7* nicotinic acetylcholine receptors - Jones_2004_J.Neurosci.Methods_134_65
Author(s) : Jones IW , Barik J , O'Neill MJ , Wonnacott S
Ref : Journal of Neuroscience Methods , 134 :65 , 2004
Abstract : Alpha 7 subunit-containing nicotinic acetylcholine receptors (alpha7* nAChR) are involved in a variety of functions in the mammalian brain, including modulating neurotransmitter release and synaptic plasticity. Identifying the precise cellular distribution of alpha7* nAChRs with respect to the local neurochemical environment is crucial to understanding these biological roles. Current strategies for localising alpha7* nAChRs at the subcellular level have limitations. Anti-alpha7 subunit antibodies detect both assembled and unassembled subunits whereas biotinylated alphabungarotoxin (alphaBgt) only binds to assembled alpha7* nAChRs, but interpretation of labelling is marred by co-detection of endogenous tissue biotin. To overcome these problems, we have characterised a novel 1.4 nm gold alphaBgt conjugate used to directly localise alpha7* nAChR. Gold conjugation does not significantly decrease binding affinity, and gold alphaBgt specifically labels alpha7* nAChR in both unfixed and aldehyde-fixed tissue at the light and electron microscope levels, labelling being abolished in the presence of excess competing toxin. At the ultrastructural level, gold alphaBgt is associated with neuronal membranes and located at axon-dendritic synapses in the rat hippocampus CA1 stratum radiatum. These results reveal gold alphaBgt to be a valuable new tool in elucidating the functional neuroanatomy of alpha7* nAChR in the central nervous system.
ESTHER : Jones_2004_J.Neurosci.Methods_134_65
PubMedSearch : Jones_2004_J.Neurosci.Methods_134_65
PubMedID: 15102504

Title : Precise localization of alpha7 nicotinic acetylcholine receptors on glutamatergic axon terminals in the rat ventral tegmental area - Jones_2004_J.Neurosci_24_11244
Author(s) : Jones IW , Wonnacott S
Ref : Journal of Neuroscience , 24 :11244 , 2004
Abstract : Alpha7 neuronal nicotinic acetylcholine receptors (nAChRs) constitute one of the predominant nAChR subtypes in the mammalian brain. Within the ventral tegmental area (VTA), nicotine application, paired with postsynaptic stimulation, contributes to a form of long-term potentiation, an effect attributed to presynaptic alpha7 nAChRs on glutamatergic afferents (Mansvelder and McGehee, 2000). The aim of this study was to examine the precise subcellular distribution of alpha7 nAChRs in the adult rat VTA to establish whether these receptors are indeed present on glutamatergic axon terminals and to determine their relationship with cholinergic afferents. The spatial relationship between alpha7 nAChRs, labeled using the alpha7 nAChR-specific antagonist alpha-bungarotoxin, and the local neurochemical environment was investigated by the application of multiple labeling strategies with antibodies against tyrosine hydroxylase, vesicular glutamate transporters (VGluTs), vesicular acetylcholine transporter, and glial fibrillary acidic protein. alpha7 nAChRs were localized at both somatodendritic and presynaptic loci within the VTA: on subpopulations of dopaminergic and nondopaminergic neurons and glutamatergic and nonglutamatergic terminals. There was no detectable alpha7 nAChR expression within astrocytes in the VTA. Most alpha7 nAChRs were cytoplasmic (82%), and the remainder were associated with the plasma membrane. Most presynaptic receptors (75%) were on glutamatergic axon terminals, with similar levels of alpha-bungarotoxin binding present on both VGluT1- and VGluT2-immunoreactive boutons. Both preembedding and postembedding electron microscopy revealed that presynaptic alpha7 nAChRs are often located at extrasynaptic (27%) and perisynaptic (61%) loci. alpha7 nAChRs were not associated with cholinergic synapses, consistent with their activation by a paracrine mode of acetylcholine or choline delivery.
ESTHER : Jones_2004_J.Neurosci_24_11244
PubMedSearch : Jones_2004_J.Neurosci_24_11244
PubMedID: 15601930

Title : Functional responses and subunit composition of presynaptic nicotinic receptor subtypes explored using the novel agonist 5-iodo-A-85380 - Mogg_2004_Neuropharmacol_47_848
Author(s) : Mogg AJ , Jones FA , Pullar IA , Sharples CG , Wonnacott S
Ref : Neuropharmacology , 47 :848 , 2004
Abstract : The novel compound 5-iodo-A-85380 binds with higher affinity to alpha4beta2* nicotinic acetylcholine receptors (nAChR), compared with other nAChR subtypes (Mukhin et al., 2000). In the present study, we have confirmed that in competition binding assays for three major nAChR subtypes, 5-iodo-A-85380 is 850 and 27,000-fold more potent at rat brain alpha4beta2* binding sites than at alpha3beta4 and alpha7 subtypes, respectively. In functional assays, 5-iodo-A-85380 potently activated (EC50 12-35 nM) both alpha-CTx-MII-sensitive and -insensitive components of [3H]dopamine release from rat striatal synaptosomes, corresponding to alpha6beta2* and alpha4beta2* nAChR, respectively. 5-Iodo-A-85380 was markedly less potent at eliciting [3H]ACh release from rat interpeduncular nucleus synaptosomes, [3H]noradrenaline release from rat hippocampal slices, and Ca2+ increases in a cell line expressing rat alpha3beta4 nAChR (EC50 = 5, 3.2, 1.6 microM, respectively). As predicted by ligand binding studies, 5-iodo-A-85380 is a more discriminating agonist than the parent compound epibatidine. However, it is not specific for alpha4beta2* nAChR as it also potently activates alpha6beta2* nAChR.
ESTHER : Mogg_2004_Neuropharmacol_47_848
PubMedSearch : Mogg_2004_Neuropharmacol_47_848
PubMedID: 15527819

Title : Alpha-conotoxins as tools for the elucidation of structure and function of neuronal nicotinic acetylcholine receptor subtypes - Nicke_2004_Eur.J.Biochem_271_2305
Author(s) : Nicke A , Wonnacott S , Lewis RJ
Ref : European Journal of Biochemistry , 271 :2305 , 2004
Abstract : Cone snails comprise approximately 500 species of venomous molluscs, which have evolved the ability to generate multiple toxins with varied and often exquisite selectivity. One class, the alpha-conotoxins, is proving to be a powerful tool for the differentiation of nicotinic acetylcholine receptors (nAChRs). These comprise a large family of complex subtypes, whose significance in physiological functions and pathological conditions is increasingly becoming apparent. After a short introduction into the structure and diversity of nAChRs, this overview summarizes the identification and characterization of alpha-conotoxins with selectivity for neuronal nAChR subtypes and provides examples of their use in defining the compositions and function of neuronal nAChR subtypes in native vertebrate tissues.
ESTHER : Nicke_2004_Eur.J.Biochem_271_2305
PubMedSearch : Nicke_2004_Eur.J.Biochem_271_2305
PubMedID: 15182346

Title : Attentional effects of nicotinic agonists in rats - Hahn_2003_Neuropharmacol_44_1054
Author(s) : Hahn B , Sharples CG , Wonnacott S , Shoaib M , Stolerman IP
Ref : Neuropharmacology , 44 :1054 , 2003
Abstract : Nicotine can increase stimulus detection, response rate and speed in the five-choice serial reaction time task, a rodent test of attention. In the present experiments, four other nicotinic agonists with different pharmacological profiles were compared in the same procedure. The response profile of epibatidine resembled that previously obtained with nicotine in that response accuracy was enhanced and omission errors and correct response latency decreased. ABT-418 transiently increased accuracy in the first 10 min of test sessions and reduced response latency. Isoarecolone caused a dose-related increase in accuracy, but had no effect on omissions or response latency. This absence of effects on response rate- or speed-related measures may be related to its previously reported reduced ability to release dopamine as compared with nicotine. The alpha7-agonist AR-R17779 was without effect on any measure, indicating that this receptor subtype may not mediate nicotinic effects on attention. Affinity constants of compounds, determined in competition binding assays targeting the alpha4beta2, alpha7, alpha3beta4 and alpha3beta2* nAChR subtypes, could not explain the differential behavioural effects observed. Differences in their functional efficacy at nAChR subtypes may instead be responsible. The finding that attentional performance and response rate and speed can be selectively modulated by nicotinic agonists is encouraging for the development of drugs with therapeutic properties similar to those of nicotine but with reduced unwanted effects.
ESTHER : Hahn_2003_Neuropharmacol_44_1054
PubMedSearch : Hahn_2003_Neuropharmacol_44_1054
PubMedID: 12763099

Title : Synthesis of two fluoro analogues of the nicotinic acetylcholine receptor agonist UB-165 - Sutherland_2003_J.Org.Chem_68_2475
Author(s) : Sutherland A , Gallagher T , Sharples CG , Wonnacott S
Ref : J Org Chem , 68 :2475 , 2003
Abstract : Two racemic fluoropyridine analogues 4 and 5 of the potent nicotinic agonist UB-165 have been synthesized. Halogenated pyridines 7 and 12 provided the organometallic reagents needed for the Negishi and Suzuki coupling reactions used for the preparation of 4 and 5, and the N-vinyloxycarbonyl protecting group of 8 and 15 was cleaved using a novel trifluoroacetic acid-mediated deprotection protocol. Analogue 4 retained high binding affinity at rat brain alpha4beta2 and alpha7 nicotinic receptors.
ESTHER : Sutherland_2003_J.Org.Chem_68_2475
PubMedSearch : Sutherland_2003_J.Org.Chem_68_2475
PubMedID: 12636420

Title : Neuroprotection by nicotine against hypoxia-induced apoptosis in cortical cultures involves activation of multiple nicotinic acetylcholine receptor subtypes - Hejmadi_2003_Mol.Cell.Neurosci_24_779
Author(s) : Hejmadi MV , Dajas-Bailador F , Barns SM , Jones B , Wonnacott S
Ref : Molecular & Cellular Neurosciences , 24 :779 , 2003
Abstract : Activation of neuronal nicotinic acetylcholine receptors (nAChR) by nicotine has been suggested to protect neurons against a hypoxic insult. The objective of this study was to examine the nature of cell death induced by acute hypoxia in rat primary cortical cultures and the neuroprotective potential of nicotine in ameliorating these processes. Neuronal cell death induced by a 4-h exposure to hypoxia (0.1% O(2)) was apoptotic, as shown by TUNEL staining and assays monitoring DNA strand breaks and caspase-3/7 activity. The presence of nicotine (10 microM) during the hypoxic insult protected a subpopulation of susceptible neurones against DNA damage and apoptosis induced by oxygen deprivation. This protective effect of nicotine was prevented by a 30-min pre-incubation with either 100 nM alpha-bungarotoxin or 1 microM dihydro-beta-erythroidine, but not 1 microM atropine, suggesting that activation of at least two subtypes of nAChR, alpha7 and beta2* nAChR, is involved in mediating nicotine neuroprotection.
ESTHER : Hejmadi_2003_Mol.Cell.Neurosci_24_779
PubMedSearch : Hejmadi_2003_Mol.Cell.Neurosci_24_779
PubMedID: 14664825

Title : The allosteric potentiation of nicotinic acetylcholine receptors by galantamine is transduced into cellular responses in neurons: Ca2+ signals and neurotransmitter release - Dajas-Bailador_2003_Mol.Pharmacol_64_1217
Author(s) : Dajas-Bailador FA , Heimala K , Wonnacott S
Ref : Molecular Pharmacology , 64 :1217 , 2003
Abstract : Neuronal nicotinic acetylcholine receptors (nAChR) modulate a variety of cellular responses, including Ca2+ signals and neurotransmitter release, which can influence neuronal processes such as synaptic efficacy and neuroprotection. In addition to receptor activation through the agonist binding site, an allosteric modulation of nAChR has also been described for a novel class of allosteric ligands. Of these, the acetylcholinesterase inhibitor and Alzheimer drug galantamine represents the prototypical allosteric ligand, based on its potentiation of nAChR-evoked single-channel and whole-cell currents. The aim of this study was to establish whether the allosteric potentiation of nAChR currents is transduced in downstream cellular responses to nAChR activation, namely increases in intracellular Ca2+ and [3H]noradrenaline release. In SH-SY5Y cells, galantamine potentiated nicotine-evoked increases in intracellular Ca2+ and [3H]noradrenaline release with a bell-shaped concentration-response profile; maximum enhancement of nicotine-evoked responses occurred at 1 muM galantamine. This potentiation was blocked by mecamylamine, whereas galantamine had no effect on these measures in the absence of nicotine. Galantamine did not compete for radioligand binding to the agonist binding sites of several nAChR subtypes, consistent with an allosteric mode of action. Unlike galantamine, the acetylcholinesterase inhibitors rivastigmine and donepezil did not potentiate nAChR-mediated responses, whereas donepezil was a reasonably potent inhibitor of nicotine- and KCl-evoked increases in Ca2+. nAChR-mediated [3H]noradrenaline release from hippocampal slices was also potentiated by galantamine, with an additional component attributable to acetylcholinesterase inhibition and subsequent increase in acetylcholine. These results indicate that the allosteric regulation of nAChR results in the potentiation of receptor-dependent cellular processes relevant to many of the physiological consequences of nAChR activation.
ESTHER : Dajas-Bailador_2003_Mol.Pharmacol_64_1217
PubMedSearch : Dajas-Bailador_2003_Mol.Pharmacol_64_1217
PubMedID: 14573772

Title : Effect of smoking and transdermal nicotine on colonic nicotinic acetylcholine receptors in ulcerative colitis - Richardson_2003_QJM_96_57
Author(s) : Richardson CE , Morgan JM , Jasani B , Green JT , Rhodes J , Williams GT , Lindstrom JM , Wonnacott S , Peel S , Thomas GA
Ref : Qjm , 96 :57 , 2003
Abstract : BACKGROUND: Ulcerative colitis (UC) is a disease largely of non-smokers, in which nicotine is of therapeutic value. The mode of action is unknown, but may involve nicotinic acetylcholine receptors (nAChRs) in the bowel wall. AIM: To investigate the presence of nAChRs in rectal mucosa, and the effect of smoking and nicotine on their expression. DESIGN: Prospective case-control study.
METHODS: In situ hybridization (ISH) and immunocytochemistry (ICC) were used to show alpha3 nAChRs in colonic mucosa. Rectal mucosa was examined from controls (n=55) and patients with inactive UC (n=62), both smokers and non-smokers, by ICC, using two antibodies to show the density and distribution of receptors in the mucosa. Non-smokers with UC (n=43) were given transdermal nicotine or placebo patches for 6 months, and rectal biopsies, taken before and after treatment, were examined by ICC to show nAChRs.
RESULTS: In normal colon, ISH and ICC showed alpha3 subunit in a wide variety of cells, including mucosal epithelium. In rectal biopsies, neither smoking nor nicotine influenced the expression of alpha3 immunoreactivity in epithelium, either in controls or UC. However, controls had a significantly greater density of immunodetectable mucosal epithelium alpha3 subunit, compared with UC patients. DISCUSSION: The presence of nAChRs in colonic epithelium may be pertinent to the beneficial effect of nicotine in UC, but since neither smoking nor nicotine treatment is associated with any change in the expression of epithelial alpha3 nAChRs, the effect may be due to functional changes in the receptor. The decreased number of alpha3 nAChRs in UC compared with controls may be related to an increased cell turnover in UC.
ESTHER : Richardson_2003_QJM_96_57
PubMedSearch : Richardson_2003_QJM_96_57
PubMedID: 12509650

Title : Synthesis and nicotinic binding of novel phenyl derivatives of UB-165. Identifying factors associated with alpha7 selectivity - Karig_2003_Bioorg.Med.Chem.Lett_13_2825
Author(s) : Karig G , Large JM , Sharples CG , Sutherland A , Gallagher T , Wonnacott S
Ref : Bioorganic & Medicinal Chemistry Lett , 13 :2825 , 2003
Abstract : Four racemic phenyl-substituted analogues 3-6 of the potent nicotinic agonist UB-165 1 have been synthesised and evaluated against the alpha(4)beta(2), alpha(3)beta(4), and alpha(7) neuronal nicotinic receptors. The 2'-phenyl derivative 3 shows no activity at these major receptor subtypes, while the 4'-phenyl analogue 4 shows an enhanced level of alpha(7) selectivity as compared to UB-165 and deschloro UB-165 2. These results are discussed within the context of recent pharmacophore models.
ESTHER : Karig_2003_Bioorg.Med.Chem.Lett_13_2825
PubMedSearch : Karig_2003_Bioorg.Med.Chem.Lett_13_2825
PubMedID: 14611837

Title : Effects of chronic drug treatments on increases in intracellular calcium mediated by nicotinic acetylcholine receptors in SH-SY5Y cells - Ridley_2002_Br.J.Pharmacol_135_1051
Author(s) : Ridley DL , Pakkanen J , Wonnacott S
Ref : British Journal of Pharmacology , 135 :1051 , 2002
Abstract : 1. SH-SY5Y cells express alpha7 and alpha3* subtypes of nicotinic acetylcholine receptors (AChR). Numbers of these receptors are upregulated by chronic treatment with nicotinic agonists or KCl. In this study we have examined the functional consequences of these drug treatments on nicotine- or KCl-evoked increases in [Ca(2+)](i), in SH-SY5Y cells. 2. In untreated cells, nicotine increased [Ca(2+)](i) (EC(50) 7.5 microM). Responses to 10 microM nicotine were abolished by the non-selective nicotinic antagonist mecamylamine and were partially blocked by alpha7-selective antagonists, the alpha3beta2*-selective antagonist alpha-conotoxin-MII, and by cadmium and verapamil. 3. After treatment for 4 days with nicotinic agonists, nicotine-evoked increases in [Ca(2+)](i) were significantly decreased by about 25%. Nicotine-evoked responses were paradoxically increased in the presence of acute methyllycaconitine (MLA; an alpha7-selective antagonist) although other alpha7-selective antagonists were without effect, while alpha-conotoxin-MII gave a partial inhibition. The increase observed with MLA was abolished by mecamylamine but not by alpha-conotoxin-MII and was still observed 24 h after chronic nicotine treatment. 4. After treatment for 4 days with KCl, nicotine-evoked increases in [Ca(2+)](i) were also decreased by 25%, but acute MLA was without effect. Responses to 20 mM KCl were unchanged by prior treatment with nicotine or KCl. Treatment for 4 days with 5 microM verapamil reduced responses to both nicotine and KCl by about 50%. 5. Multiple nicotinic AChR subtypes contribute to nicotine-evoked increases in [Ca(2+)](i) in SH-SY5Y cells. Responses to acute nicotine are reduced after chronic nicotine or KCl treatment, with loss of the component attributed to the alpha7 subtype. However, in nicotine-treated cells this effect is reversed when nicotine stimulation is applied in the presence of acute MLA. The antagonist may assist in converting a non-functional alpha7 nicotinic AChR to a conducting state.
ESTHER : Ridley_2002_Br.J.Pharmacol_135_1051
PubMedSearch : Ridley_2002_Br.J.Pharmacol_135_1051
PubMedID: 11861334

Title : Methyllycaconitine is a potent antagonist of alpha-conotoxin-MII-sensitive presynaptic nicotinic acetylcholine receptors in rat striatum - Mogg_2002_J.Pharmacol.Exp.Ther_302_197
Author(s) : Mogg AJ , Whiteaker P , McIntosh JM , Marks M , Collins AC , Wonnacott S
Ref : Journal of Pharmacology & Experimental Therapeutics , 302 :197 , 2002
Abstract : The plant alkaloid methyllycaconitine (MLA) is considered to be a selective antagonist of the alpha7 subtype of neuronal nicotinic acetylcholine receptor (nAChR). However, 50 nM MLA partially inhibited (by 16%) [(3)H]dopamine release from rat striatal synaptosomes stimulated with 10 microM nicotine. Other alpha7-selective antagonists had no effect. Similarly, MLA (50 nM) inhibited [(3)H]dopamine release evoked by the partial agonist (2-chloro-5-pyridyl)-9-azabicyclo[4.2.1]non-2-ene (UB-165) (0.2 microM) by 37%. In both cases, inhibition by MLA was surmountable with higher agonist concentrations, indicative of a competitive interaction. At least two subtypes of presynaptic nAChR can modulate dopamine release in the striatum, and these nAChR are distinguished by their differential sensitivity to alpha-conotoxin-MII (alpha-CTx-MII). MLA was not additive with a maximally effective concentration of alpha-CTx-MII (100 nM) in inhibiting [(3)H]dopamine release elicited by 10 microM nicotine or 0.2 microM UB-165, suggesting that both toxins act at the same site. This was confirmed in quantitative binding assays with (125)I-alpha-CTx-MII, which displayed saturable specific binding to rat striatum and nucleus accumbens with B(max) values of 9.8 and 16.5 fmol/mg of protein, and K(d) values of 0.63 and 0.83 nM, respectively. MLA fully inhibited (125)I-alpha-CTx-MII binding to striatum and nucleus accumbens with a K(i) value of 33 nM, consistent with the potency observed in the functional assays. We speculate that MLA and alpha-CTx-MII interact with a presynaptic nAChR of subunit composition alpha3/alpha6beta2beta3* on dopamine neurons. The use of MLA as an alpha7-selective antagonist should be exercised with caution, especially in studies of nAChR in basal ganglia.
ESTHER : Mogg_2002_J.Pharmacol.Exp.Ther_302_197
PubMedSearch : Mogg_2002_J.Pharmacol.Exp.Ther_302_197
PubMedID: 12065717

Title : Synthesis and pharmacological characterization of novel analogues of the nicotinic acetylcholine receptor agonist (+\/-)-UB-165 - Sharples_2002_J.Med.Chem_45_3235
Author(s) : Sharples CG , Karig G , Simpson GL , Spencer JA , Wright E , Millar NS , Wonnacott S , Gallagher T
Ref : Journal of Medicinal Chemistry , 45 :3235 , 2002
Abstract : (+/-)-UB-165 (1) is a potent neuronal nicotinic acetylcholine receptor (nAChR) ligand, which displays functional selectivity between nAChR subtypes. Using UB-165 as a lead structure, two classes of racemic ligands were synthesized and assessed in binding assays for three major nAChR subtypes (alpha4beta2, alpha3beta4, and alpha7). The first class of compounds comprises the three pyridine isomers 4-6, corresponding to the 3-, 2-, and 4-substituted pyridine isomers, respectively. Deschloro UB-165 (4) displayed a 2-3-fold decrease in affinity at alpha4beta2 and alpha3beta4 nAChR subtypes, as compared with (+/-)-UB-165, while at the alpha7 subtype a 31-fold increase in affinity was observed. At each of the nAChR subtypes, high affinity binding was dependent on the presence of a 3-substituted pyridine, and the other isomers, 5 and 6, resulted in marked decreases in binding affinities. The second class of compounds is based on replacing the pyridyl unit of 1 with a diazine moiety, giving pyridazine (7), pyrimidine (8), and pyrazine (9), which retain the "3-pyridyl" substructure. Modest reductions in binding affinity were observed for all of the diazine ligands at all nAChR subtypes, with the exception of 7, which retained potency comparable to that of 4 in binding to alpha7 nAChR. In functional assays at the alpha3beta4 nAChR, all analogues 4-9 were less potent, as compared with 1, and the rank order of functional potencies correlated with that of binding potencies. Computational studies indicate that the 3-substituted pyridine 4 and 2-substituted pyridine 5, as well as the diazine analogues 7-9, all conform to a distance-based pharmacophore model recently proposed for the alpha4beta2 receptor. However, the nicotinic potencies of these ligands vary considerably and because 5 lacks appreciable nicotinic activity, it is clear that further refinements of this model are necessary in order to describe adequately the structural and electronic demands associated with this nAChR subtype. This rational series of compounds based on UB-165 presents a systematic approach to defining subtype specific pharmacophores.
ESTHER : Sharples_2002_J.Med.Chem_45_3235
PubMedSearch : Sharples_2002_J.Med.Chem_45_3235
PubMedID: 12109907

Title : Intracellular Ca2+ signals evoked by stimulation of nicotinic acetylcholine receptors in SH-SY5Y cells: contribution of voltage-operated Ca2+ channels and Ca2+ stores - Dajas-Bailador_2002_J.Neurochem_81_606
Author(s) : Dajas-Bailador FA , Mogg AJ , Wonnacott S
Ref : Journal of Neurochemistry , 81 :606 , 2002
Abstract : Neuronal nicotinic acetylcholine receptors (nAChR) can regulate several neuronal processes through Ca2+-dependent mechanisms. The versatility of nAChR-mediated responses presumably reflects the spatial and temporal characteristics of local changes in intracellular Ca2+ arising from a variety of sources. The aim of this study was to analyse the components of nicotine-evoked Ca2+ signals in SH-SY5Y cells, by monitoring fluorescence changes in cells loaded with fluo-3 AM. Nicotine (30 microm) generated a rapid elevation in cytoplasmic Ca2+ that was partially and additively inhibited (40%) by alpha7 and alpha3beta2* nAChR subtype selective antagonists; alpha3beta4* nAChR probably account for the remaining response (60%). A substantial blockade (80%) by CdCl2 (100 microm) indicates that voltage-operated Ca2+ channels (VOCC) mediate most of the nicotine-evoked response, although the alpha7 selective antagonist alpha-bungarotoxin (40 nm) further decreased the CdCl2- resistant component. The elevation of intracellular Ca2+ levels provoked by nicotine was sustained for at least 10 min and required the persistent activation of nAChR throughout the response. Intracellular Ca2+ stores were implicated in both the initial and sustained nicotine-evoked Ca2+ responses, by the blockade observed after ryanodine (30 microm) and the inositoltriphosphate (IP3)-receptor antagonist, xestospongin-c (10 microm). Thus, nAChR subtypes are differentially coupled to specific sources of Ca2+: activation of nAChR induces a sustained elevation of intracellular Ca2+ levels which is highly dependent on the activation of VOCC, and also involves Ca2+ release from ryanodine and IP3-dependent intracellular stores. Moreover, the alpha7, but not alpha3beta2* nAChR, are responsible for a fraction of the VOCC-independent nicotine-evoked Ca2+ increase that appears to be functionally coupled to ryanodine sensitive Ca2+ stores.
ESTHER : Dajas-Bailador_2002_J.Neurochem_81_606
PubMedSearch : Dajas-Bailador_2002_J.Neurochem_81_606
PubMedID: 12065669

Title : Nicotine activates the extracellular signal-regulated kinase 1\/2 via the alpha7 nicotinic acetylcholine receptor and protein kinase A, in SH-SY5Y cells and hippocampal neurones - Dajas-Bailador_2002_J.Neurochem_80_520
Author(s) : Dajas-Bailador FA , Soliakov L , Wonnacott S
Ref : Journal of Neurochemistry , 80 :520 , 2002
Abstract : Neuronal nicotinic acetylcholine receptors (nAChR) can modulate many cellular mechanisms, such as cell survival and memory processing, which are also influenced by the serine/threonine protein kinases ERK1/2. In SH-SY5Y cells and hippocampal neurones, nicotine (100 microM) increased the activity of ERK1/2. This effect was Ca2+ dependent, and prevented by the alpha7 nAChR antagonist alpha-bungarotoxin (alpha-Bgt) and an inhibitor (PD98059) of the upstream kinase MEK. To determine the intervening steps linking Ca2+ entry to MEK-ERK1/2 activation, inhibitors of Ca2+-dependent kinases were deployed. In SH-SY5Y cells, selective blockers for PKC (Ro 31-8220), CaM kinase II (KN-62) or PI3 kinase (LY 294002) failed to inhibit the nicotine-evoked increase in ERK1/2 activity. In contrast, two structurally different inhibitors of PKA (KT 5720 and H-89) completely prevented the nicotine-dependent increase in ERK1/2 activity. Inhibition of the nicotine-evoked increase in ERK1/2 activity by H-89 was also observed in hippocampal cultures. Down stream of PKA, the activity of B-Raf was significantly decreased by nicotine in SH-SY5Y cells, as determined by direct measurement of MEK1 phosphorylation or in vitro kinase assays, whereas the modulation of MEK1 phosphorylation by Raf-1 tended to increase. Thus, this study provides evidence for a novel signalling route coupling the stimulation of alpha7 nAChR to the activation of ERK1/2, in a Ca2+ and PKA dependent manner.
ESTHER : Dajas-Bailador_2002_J.Neurochem_80_520
PubMedSearch : Dajas-Bailador_2002_J.Neurochem_80_520
PubMedID: 11905997

Title : Megacystis-microcolon-intestinal hypoperistalsis syndrome and the absence of the alpha3 nicotinic acetylcholine receptor subunit - Richardson_2001_Gastroenterology_121_350
Author(s) : Richardson CE , Morgan JM , Jasani B , Green JT , Rhodes J , Williams GT , Lindstrom JM , Wonnacott S , Thomas GA , Smith V
Ref : Gastroenterology , 121 :350 , 2001
Abstract : BACKGROUND & AIMS: The megacystis-microcolon-intestinal hypoperistalsis syndrome (MMIHS) is a rare disease of childhood that presents early with intestinal hypoperistalsis, hydronephrosis, and hydroureters. Transgenic mice that lack the alpha3 subunit containing nicotinic acetylcholine (nAChR) have a phenotype similar to that of MMIHS.
METHODS: We examined the expression of this subunit in control and MMIHS tissue derived from patients using in situ hybridization (ISH) and immunocytochemistry (ICC).
RESULTS: In controls, both techniques showed a wide distribution of alpha3 nAChRs present in ganglion cells, muscle, and epithelium. By contrast, most MMIHS tissue gave negative staining with ISH and variable results with ICC.
CONCLUSIONS: These observations are consistent with a lack of alpha3 nAChRs contributing to the pathogenesis of MMIHS.
ESTHER : Richardson_2001_Gastroenterology_121_350
PubMedSearch : Richardson_2001_Gastroenterology_121_350
PubMedID: 11487544

Title : Differential effects of chronic drug treatment on alpha3* and alpha7 nicotinic receptor binding sites, in hippocampal neurones and SH-SY5Y cells - Ridley_2001_Br.J.Pharmacol_133_1286
Author(s) : Ridley DL , Rogers A , Wonnacott S
Ref : British Journal of Pharmacology , 133 :1286 , 2001
Abstract : 1. The aim of this study was to compare the effects of chronic treatment (for 4 or 7 days) with nicotinic drugs and 20 mM KCl on numbers of surface alpha7 nicotinic AChR, identified by [(125)I]-alpha bungarotoxin (alpha-Bgt) binding, in primary hippocampal cultures and SH-SY5Y cells. Numbers of alpha3* nicotinic AChR were also examined in SH-SY5Y cells, using [(3)H]-epibatidine, which is predicted to label the total cellular population of predominantly alpha3beta2* nicotinic AChR under the conditions used. 2. All the nicotinic agonists examined, the antagonists d-tubocurarine and methyllycaconitine, and KCl, upregulated [(125)I]-alpha Bgt binding sites by 20 - 60% in hippocampal neurones and, where examined, SH-SY5Y cells. 3. Upregulation of [(125)I]-alpha-Bgt binding sites by KCl was prevented by co-incubation with the L-type Ca2+ channel blocker verapamil or the Ca2+-calmodulin dependent kinase II (CaM-kinase II) inhibitor KN-62. Upregulation of [(125)I]-alpha-Bgt binding sites by nicotine or 3,[(4-dimethylamino) cinnamylidene] anabaseine maleate (DMAC) was insensitive to these agents. 4. [(3)H]-Epibatidine binding sites in SH-SY5Y cells were not affected by KCl but were upregulated in a verapamil-insensitive manner by nicotine and DMAC. KN-62 itself provoked a 2 fold increase in [(3)H]-epibatidine binding. The inactive analogue KN-04 had no effect, suggesting that CaM-kinase II plays a role in regulating numbers of alpha3* nicotinic AChR. 5. These data indicate that numbers of alpha3* and alpha7 nicotinic AChR are modulated differently. Nicotinic agonists and KCl upregulate alpha7 nicotinic AChR through distinct cellular mechanisms, the latter involving L-type Ca2+ channels and CaM-kinase II. In contrast, alpha3* nicotinic AChR are not upregulated by KCl. This difference may reflect the distinct physiological roles proposed for alpha7 nicotinic AChR.
ESTHER : Ridley_2001_Br.J.Pharmacol_133_1286
PubMedSearch : Ridley_2001_Br.J.Pharmacol_133_1286
PubMedID: 11498514

Title : Presynaptic localisation of the nicotinic acetylcholine receptor beta2 subunit immunoreactivity in rat nigrostriatal dopaminergic neurones - Jones_2001_J.Comp.Neurol_439_235
Author(s) : Jones IW , Bolam JP , Wonnacott S
Ref : Journal of Comparative Neurology , 439 :235 , 2001
Abstract : Nicotinic acetylcholine receptors (nAChR) are widely distributed in the central nervous system, where they exert a modulatory influence on synaptic transmission. For the striatum, pharmacological evidence supports the presence of presynaptic alpha3beta2* and alpha4beta2* nAChR that modulate dopamine release from nigrostriatal terminals. The objective of this study was to examine the precise subcellular distribution of the nAChR beta2 subunit in these neurones and its localisation at presynaptic sites. Double immunolabelling with tyrosine hydroxylase (TH) at the confocal level revealed that the cell bodies and axon terminals (synaptosomes) of nigrostriatal neurones were also immunoreactive for the nAChR beta2 subunit. Double-preembedding electron microscopy confirmed that beta2-immunogold labelling was enriched in TH-positive terminals in the dorsal striatum. Quantitative analysis of doubly immunogold-labelled sections in postembedding electron microscopy showed that 86% of TH-positive axonal boutons are also labelled for the nAChR beta2 subunit, whereas 45% of beta2 subunit-immunolabeled boutons do not contain TH. Thus the beta2 subunit is localised within at least two populations of axon terminals in the dorsal striatum. In these structures, 15% of beta2 subunit immunoreactivity was at the plasma membrane but was rarely associated with synapses. These findings are compatible with functional presynaptic beta2-containing nAChR that may be stimulated physiologically by acetylcholine that diffuses from synaptic or nonsynaptic sites of acetylcholine release. These results demonstrate the presynaptic localisation of an nAChR subunit in nigrostriatal dopaminergic neurones, providing morphological evidence for the presynaptic nicotinic modulation of dopamine release.
ESTHER : Jones_2001_J.Comp.Neurol_439_235
PubMedSearch : Jones_2001_J.Comp.Neurol_439_235
PubMedID: 11596051

Title : [3H]-Methyllycaconitine: a high affinity radioligand that labels invertebrate nicotinic acetylcholine receptors - Lind_2001_Insect.Biochem.Mol.Biol_31_533
Author(s) : Lind RJ , Hardick DJ , Blagbrough IS , Potter BV , Wolstenholme AJ , Davies AR , Clough MS , Earley FG , Reynolds SE , Wonnacott S
Ref : Insect Biochemistry & Molecular Biology , 31 :533 , 2001
Abstract : Nicotinic acetylcholine receptors (nAChR) of insect and other invertebrates are heterogeneous and new tools are needed to dissect their multiplicity. [(3)H]-Methyllycaconitine ([(3)H]-MLA) is a novel radioligand which is a potent antagonist at vertebrate alpha7-type nAChR. Putative invertebrate nAChR of the aphid Myzus persicae, the moths Heliothis virescens and Manduca sexta, the fly Lucilia sericata, and the squid Loligo vulgaris were investigated in radioligand binding studies with [(3)H]-MLA. Saturable binding was consistent with a single class of high affinity binding sites for each of these invertebrates, characterised by a dissociation constant, K(d), of approximately 1 nM and maximal binding capacities, B(max), between 749 and 1689 fmol/mg protein for the insects and 14,111 fmol/mg protein for squid. [(3)H]-MLA binding to M. persicae membranes was characterised in more detail. Kinetic analysis demonstrated rapid association in a biphasic manner and slow, monophasic dissociation. Displacement studies demonstrate the nicotinic character of [(3)H]-MLA binding sites. Data for all nicotinic ligands, except MLA itself, are consistent with displacement from a high and a low affinity site, indicating that displacement is occurring from two or more classes of nicotinic binding site that are not distinguished by MLA itself. Autoradiographic analysis of the distribution of [(3)H]-MLA binding sites in Manduca sexta shows discrete labelling of neuropil areas of the optic and antennal lobes.
ESTHER : Lind_2001_Insect.Biochem.Mol.Biol_31_533
PubMedSearch : Lind_2001_Insect.Biochem.Mol.Biol_31_533
PubMedID: 11267892

Title : Involvement of protein kinase C in the presynaptic nicotinic modulation of [(3)H]-dopamine release from rat striatal synaptosomes - Soliakov_2001_Br.J.Pharmacol_132_785
Author(s) : Soliakov L , Wonnacott S
Ref : British Journal of Pharmacology , 132 :785 , 2001
Abstract : 1. Presynaptic nicotinic ACh receptors modulate transmitter release in the brain. Here we report their interactions with protein kinase C (PKC) with respect to [(3)H]-dopamine release from rat striatal synaptosomes, monitored by superfusion. 2. Two specific PKC inhibitors, Ro 31-8220 (1 microM) and D-erythro-sphingosine (10 microM) significantly reduced (by 51 and 26% respectively) [(3)H]-dopamine release stimulated by anatoxin-a (AnTx), a potent and selective agonist of nicotinic ACh receptors. The inactive structural analogue of Ro 31-8220, bisindolylmaleimide V (1 microM) had no effect. 3. Two phorbol esters, PDBu (1 microM) and PMA (1 microM) potentiated AnTx-evoked [(3)H]-dopamine release by 50 - 80%. This was Ca(2+)-dependent and prevented by PKC inhibitors. In the absence of nicotinic agonist, phorbol esters enhanced basal release through a PKC-independent mechanism. 4. A (86)Rb(+) efflux assay of nicotinic ACh receptor function confirmed that Ro 31-8220 has no nonspecific effect on presynaptic nicotinic ACh receptors. 5. These results suggest that PKC is activated by nicotinic ACh receptor stimulation and mediates a component of AnTx-evoked [(3)H]-dopamine release. In addition, independent activation of PKC can further amplify the response, offering a potential mechanism for receptor crosstalk.
ESTHER : Soliakov_2001_Br.J.Pharmacol_132_785
PubMedSearch : Soliakov_2001_Br.J.Pharmacol_132_785
PubMedID: 11159732

Title : Presynaptic Neuronal Nicotinic Receptors: Pharmacology, Heterogeneity, and Cellular Mechanisms -
Author(s) : Kaiser S , Soliakov L , Wonnacott S
Ref : Handbook of Experimental Pharmacology , 144 :193 , 2000

Title : Presynaptic nicotinic receptors modulating dopamine release in the rat striatum - Wonnacott_2000_Eur.J.Pharmacol_393_51
Author(s) : Wonnacott S , Kaiser S , Mogg A , Soliakov L , Jones IW
Ref : European Journal of Pharmacology , 393 :51 , 2000
Abstract : The modulation of striatal dopamine release by presynaptic nicotinic acetylcholine receptors is well documented for both synaptosomes and slices. Because the latter retain local anatomical integrity, we have compared [3H]dopamine release evoked by the nicotinic receptor agonists (-)-nicotine and (+/-)-anatoxin-a from striatal synaptosome and slice preparations in parallel. At higher agonist concentrations, mecamylamine-sensitive [3H]dopamine release was greater from slices, indicative of an additional component, and this increase was abolished by glutamate receptor antagonists. To begin to examine the localisation of specific nicotinic acetylcholine receptor subtypes in the striatum, immunogold electron microscopy was undertaken with the beta2-specific monoclonal antibody 270. In striatal sections, gold particles were associated with symmetric synapses (dopaminergic) but were absent from asymmetric synapses (glutamatergic). Surface labelling of striatal synaptosomes with gold particles was also demonstrated. Taken together, these results are consistent with dopamine release mediated by beta2-containing nicotinic acetylcholine receptors on dopamine terminals, while non-beta2-containing nicotinic acetylcholine receptors may enhance dopamine release indirectly by releasing glutamate from neighbouring terminals.
ESTHER : Wonnacott_2000_Eur.J.Pharmacol_393_51
PubMedSearch : Wonnacott_2000_Eur.J.Pharmacol_393_51
PubMedID: 10770997

Title : alpha-bungarotoxin-sensitive nicotinic receptors indirectly modulate [(3)H]dopamine release in rat striatal slices via glutamate release - Kaiser_2000_Mol.Pharmacol_58_312
Author(s) : Kaiser S , Wonnacott S
Ref : Molecular Pharmacology , 58 :312 , 2000
Abstract : Nicotinic agonists elicit the release of dopamine from striatal synaptosomes by acting on presynaptic nicotinic acetylcholine receptors (nAChRs) on dopamine nerve terminals. Both alpha3beta2* and alpha4beta2 nAChR subtypes (but not alpha7* nAChRs) have been implicated. Here, we compared nAChR-evoked [(3)H]dopamine release from rat striatal synaptosome and slice preparations by using the nicotinic agonist anatoxin-a. In the more integral slice preparation, the concentration-response curve for anatoxin-a-evoked [(3)H]dopamine release was best fitted to a two-site model, giving EC(50) values of 241 nM and 5.1 microM, whereas only the higher-affinity component was observed in synaptosome preparations (EC(50) = 134 nM). Responses to a high concentration of anatoxin-a (25 microM) in slices (but not in synaptosomes) were partially blocked by ionotropic glutamate receptor antagonists (kynurenic acid, 6,7-dinitroquinoxaline-2,3-dione) and by alpha7*-selective nAChR antagonists (alpha-bungarotoxin, alpha-conotoxin-ImI, methyllycaconitine) in a nonadditive manner. In contrast, the alpha3beta2-selective nAChR antagonist alpha-conotoxin-MII partially inhibited [(3)H]dopamine release from both slice and synaptosome preparations, stimulated with both low (1 microM) and high (25 microM) concentrations of anatoxin-a. Antagonism by alpha-conotoxin-MII was additive with that of alpha7*-selective antagonists. These data support a model in which alpha7* nAChRs on striatal glutamate terminals elicit glutamate release, which in turn acts at ionotropic glutamate receptors on dopamine terminals to stimulate dopamine release. In addition, non-alpha7* nAChRs on dopamine terminals also stimulate dopamine release. These observations have implications for the complex cholinergic modulation of inputs onto the major efferent neurons of the striatum.
ESTHER : Kaiser_2000_Mol.Pharmacol_58_312
PubMedSearch : Kaiser_2000_Mol.Pharmacol_58_312
PubMedID: 10908298

Title : The alpha7 nicotinic acetylcholine receptor subtype mediates nicotine protection against NMDA excitotoxicity in primary hippocampal cultures through a Ca(2+) dependent mechanism - Dajas-Bailador_2000_Neuropharmacol_39_2799
Author(s) : Dajas-Bailador FA , Lima PA , Wonnacott S
Ref : Neuropharmacology , 39 :2799 , 2000
Abstract : Neuronal nicotinic acetylcholine receptors (nAChR) have been suggested to play a role in a variety of modulatory and regulatory processes, including neuroprotection. Here we have characterized the neuroprotective effects of nicotine against an excitotoxic insult in primary hippocampal cultures. Exposure of hippocampal neurons to 200 microM NMDA for 1 h decreased cell viability by 25+/-5%, an effect blocked by NMDA receptor antagonists. Nicotine (10 microM) counteracted the NMDA-induced cell death when co-incubated with NMDA or when present subsequent to the NMDA treatment. Nicotine protection was prevented by 1 microM MLA, confirming that it was mediated by nAChR, and by 1 microM alpha-bungarotoxin, demonstrating that the alpha7 nAChR subtype was responsible. Both the NMDA evoked neurotoxicity and nicotine neuroprotection were Ca(2+)-dependent. In Fura-2-loaded hippocampal neurons, nicotine (10 microM) and NMDA (200 microM) acutely increased intracellular resting Ca(2+) from 70 nM to 200 and 500 nM, respectively. Responses to NMDA were unaffected by the presence of nicotine. (45)Ca(2+) uptake after a 1 h exposure to nicotine or NMDA also demonstrated quantitative differences between the two drugs. This study demonstrates that the alpha7 subtype of nAChR can support neuronal survival after an excitotoxic stimulus, through a Ca(2+) dependent mechanism that operates downstream of NMDA receptor activation.
ESTHER : Dajas-Bailador_2000_Neuropharmacol_39_2799
PubMedSearch : Dajas-Bailador_2000_Neuropharmacol_39_2799
PubMedID: 11044750

Title : UB-165: a novel nicotinic agonist with subtype selectivity implicates the alpha4beta2* subtype in the modulation of dopamine release from rat striatal synaptosomes - Sharples_2000_J.Neurosci_20_2783
Author(s) : Sharples CG , Kaiser S , Soliakov L , Marks MJ , Collins AC , Washburn M , Wright E , Spencer JA , Gallagher T , Whiteaker P , Wonnacott S
Ref : Journal of Neuroscience , 20 :2783 , 2000
Abstract : Presynaptic nicotinic acetylcholine receptors (nAChRs) on striatal synaptosomes stimulate dopamine release. Partial inhibition by the alpha3beta2-selective alpha-conotoxin-MII indicates heterogeneity of presynaptic nAChRs on dopamine terminals. We have used this alpha-conotoxin and UB-165, a novel hybrid of epibatidine and anatoxin-a, to address the hypothesis that the alpha-conotoxin-MII-insensitive subtype is composed of alpha4 and beta2 subunits. UB-165 shows intermediate potency, compared with the parent molecules, at alpha4beta2* and alpha3-containing binding sites, and resembles epibatidine in its high discrimination of these sites over alpha7-type and muscle binding sites. (+/-)-Epibatidine, (+/-)-anatoxin-a, and (+/-)-UB-165 stimulated [(3)H]-dopamine release from striatal synaptosomes with EC(50) values of 2.4, 134, and 88 nM, and relative efficacies of 1:0.4:0.2, respectively. alpha-Conotoxin-MII inhibited release evoked by these agonists by 48, 56, and 88%, respectively, suggesting that (+/-)-UB-165 is a very poor agonist at the alpha-conotoxin-MII-insensitive nAChR subtype. In assays of (86)Rb(+) efflux from thalamic synaptosomes, a model of an alpha4beta2* nAChR response, (+/-)-UB-165 was a very weak partial agonist; the low efficacy of (+/-)-UB-165 at alpha4beta2 nAChR was confirmed in Xenopus oocytes expressing various combinations of human nAChR subunits. In contrast, (+/-)-UB-165 and (+/-)-anatoxin-a were similarly efficacious and similarly sensitive to alpha-conotoxin-MII in increasing intracellular Ca(2+) in SH-SY5Y cells, a functional assay for native alpha3-containing nAChR. These data support the involvement of alpha4beta2* nAChR in the presynaptic modulation of striatal dopamine release and illustrate the utility of exploiting a novel partial agonist, together with a selective antagonist, to dissect the functional roles of nAChR subtypes in the brain.
ESTHER : Sharples_2000_J.Neurosci_20_2783
PubMedSearch : Sharples_2000_J.Neurosci_20_2783
PubMedID: 10751429

Title : Characterisation of the binding of [3H]methyllycaconitine: a new radioligand for labelling alpha 7-type neuronal nicotinic acetylcholine receptors - Davies_1999_Neuropharmacol_38_679
Author(s) : Davies AR , Hardick DJ , Blagbrough IS , Potter BV , Wolstenholme AJ , Wonnacott S
Ref : Neuropharmacology , 38 :679 , 1999
Abstract : Methyllycaconitine (MLA), a norditerpenoid alkaloid isolated from Delphinium seeds, is one of the most potent non-proteinacious ligands that is selective for alpha bungarotoxin-sensitive neuronal nicotinic acetylcholine receptors (nAChR). [3H]MLA bound to rat brain membranes with high affinity (Kd = 1.86 +/- 0.31 nM) with a good ratio of specific to non-specific binding. The binding of [3H]MLA was characterised by rapid association (t 1/2 = 2.3 min) and dissociation (t 1/2 = 12.6 min) kinetics. The radioligand binding displayed nicotinic pharmacology, consistent with an interaction with alpha bungarotoxin-sensitive nAChR. The snake alpha-toxins, alpha bungarotoxin and alpha cobratoxin, displaced [3H]MLA with high affinity (Ki = 1.8 +/- 0.5 and 5.5 +/- 0.9 nM, respectively), whereas nicotine was less potent (Ki = 6.1 +/- 1.1 microM). The distribution of [3H]MLA binding sites in crudely dissected rat brain regions was identical to that of [125I] alpha bungarotoxin binding sites, with a high binding site density in hippocampus and hypothalamus, but low density in striatum and cerebellum. [3H]MLA also labelled a sub-population of binding sites which are not sensitive to the snake alpha toxins, but which did not differ significantly from the major population with respect to their other pharmacological properties or regional distribution. [3H]MLA, therefore, is a novel radiolabel for characterising alpha 7-type nAChR. A good signal to noise ratio and rapid binding kinetics provide advantages over the use of radiolabelled alpha bungarotoxin for rapid and accurate equilibrium binding assays.
ESTHER : Davies_1999_Neuropharmacol_38_679
PubMedSearch : Davies_1999_Neuropharmacol_38_679
PubMedID: 10340305

Title : International Union of Pharmacology. XX. Current status of the nomenclature for nicotinic acetylcholine receptors and their subunits -
Author(s) : Lukas RJ , Changeux JP , Le Novere N , Albuquerque EX , Balfour DJ , Berg DK , Bertrand D , Chiappinelli VA , Clarke PB , Collins AC , Dani JA , Grady SR , Kellar KJ , Lindstrom JM , Marks MJ , Quik M , Taylor P , Wonnacott S
Ref : Pharmacol Rev , 51 :397 , 1999
PubMedID: 10353988

Title : Nicotine: not just for cigarettes anymore -
Author(s) : Wonnacott S , Marks MJ
Ref : Drug Discov Today , 4 :490 , 1999
PubMedID: 10529765

Title : An autoradiographic study of the distribution of binding sites for the novel alpha7-selective nicotinic radioligand [3H]-methyllycaconitine in the mouse brain - Whiteaker_1999_Eur.J.Neurosci_11_2689
Author(s) : Whiteaker P , Davies AR , Marks MJ , Blagbrough IS , Potter BV , Wolstenholme AJ , Collins AC , Wonnacott S
Ref : European Journal of Neuroscience , 11 :2689 , 1999
Abstract : [3H]-Methyllycaconitine ([3H]-MLA) is a new radioligand with selectivity for alpha7-type neuronal nicotinic acetylcholine receptors (nAChRs). In our previous study [Davies, A.R.L., Hardick, D.J., Blagbrough, I.S., Potter, B.V.L., Wolstenholme, A.J. & Wonnacott, S. (1999) Neuropharmacology, 38, 679-690], this radioligand labelled a single class of site in rat brain membranes; its pharmacology and distribution in crudely dissected brain regions closely paralleled that of the well-established alpha7-ligand [125I]-alpha-bungarotoxin. However, a small population of [3H]-MLA binding sites was apparently insensitive to alpha-bungarotoxin. Here we have extended the study to mouse brain, using autoradiography to examine the distribution of [3H]-MLA and [125I]-alpha-bungarotoxin binding sites. [3H]-MLA labelled a single class of site in mouse brain membranes with a KD of 2.2 nM and a Bmax of 45.6 fmol/mg protein. Specific binding, defined by unlabelled MLA (Ki = 0.69 nM), was completely inhibited by (-)-nicotine (Ki = 1.62 microM), whereas alpha-bungarotoxin inhibited only 85% of specific binding (Ki = 3.5 nM). The distributions of [125I]-alpha-bungarotoxin and [3H]-MLA binding sites were compared by autoradiography, and binding was quantitated in 72 brain regions. Binding of both radioligands was highly correlated, with highest densities in the dorsal tegmental nucleus of the pons, colliculi and hippocampus. Serial sections labelled with [3H]-MLA in the absence or presence of unlabelled MLA or alpha-bungarotoxin provided no evidence for any alpha-bungarotoxin-resistant binding. The results are discussed in terms of binding sites that are inaccessible to alpha-bungarotoxin in membrane preparations. This study demonstrates the utility of [3H]-MLA for characterization of alpha7-type nicotinic receptors in mammalian brain, and suggests that it labels a population identical to that defined by [125I]-alpha-bungarotoxin.
ESTHER : Whiteaker_1999_Eur.J.Neurosci_11_2689
PubMedSearch : Whiteaker_1999_Eur.J.Neurosci_11_2689
PubMedID: 10457165

Title : Characterization of a nicotinic acetylcholine receptor from the insect Manduca sexta - Eastham_1998_Eur.J.Neurosci_10_879
Author(s) : Eastham HM , Lind RJ , Eastlake JL , Clarke BS , Towner P , Reynolds SE , Wolstenholme AJ , Wonnacott S
Ref : European Journal of Neuroscience , 10 :879 , 1998
Abstract : Manduca sexta is a nicotine-insensitive insect, the larval form of which feeds on tobacco. It has been postulated that its nicotine insensitivity may reflect the presence of a modified nicotinic acetylcholine receptor whose alpha subunits lack the amino acid residues necessary for binding nicotine: we have performed ligand binding assays and molecular cloning to examine this hypothesis. [125I]alpha-bungarotoxin bound specifically to both larval and adult membranes, with Kd values of 7.6 and 6.5 nM and Bmax values of 119 and 815 fmol/mg protein, respectively. The pharmacological profile of [1251]alpha-bungarotoxin binding was similar in both tissues. In particular, nicotine (Ki values: 1.6 microM and 2 microM for larvae and adults, respectively) competed with an affinity similar to that found for nicotine-sensitive insects. No alpha-bungarotoxin-insensitive binding sites labelled by [3H]epibatidine could be detected. Using the alpha-like subunit from the locust Schistocerca gregaria to probe two cDNA libraries, and by inverse PCR on circularized genomic DNA from Manduca sexta, we have obtained overlapping cDNA clones that contain the complete coding sequence of a putative nicotinic subunit from Manduca sexta (MARA1). No other alpha-subunit cDNAs were isolated using this probe, although it hybridized to multiple bands on Southern blots. The sequence of MARA1 is consistent with an alpha-like subunit capable of binding alpha-bungarotoxin, and it retains all those amino acids implicated in nicotine binding to vertebrate nicotinic receptors. Taken together, these findings provide no support for the hypothesis that the nicotine insensitivity of Manduca sexta is the result of a nicotinic receptor with diminished nicotine binding.
ESTHER : Eastham_1998_Eur.J.Neurosci_10_879
PubMedSearch : Eastham_1998_Eur.J.Neurosci_10_879
PubMedID: 9753155

Title : Differential inhibition by alpha-conotoxin-MII of the nicotinic stimulation of [3H]dopamine release from rat striatal synaptosomes and slices - Kaiser_1998_J.Neurochem_70_1069
Author(s) : Kaiser SA , Soliakov L , Harvey SC , Luetje CW , Wonnacott S
Ref : Journal of Neurochemistry , 70 :1069 , 1998
Abstract : The presynaptic nicotinic modulation of dopamine release from striatal nerve terminals is well established, but the subtype(s) of neuronal nicotinic acetylcholine receptor (nAChR) underlying this response has not been identified. Recently, alpha-conotoxin-MII has been reported to inhibit potently and selectively the rat alpha3beta2 combination of nAChR subunits. Here we have synthesised the peptide, confirmed its specificity, and examined its effect on the (+/-)-anatoxin-a-evoked release of [3H]dopamine from rat striatal synaptosomes and slices. Alpha-conotoxin-MII (112 nM) completely blocked acetylcholine-evoked currents of alpha3beta2 nAChRs expressed in Xenopus oocytes (IC50 = 8.0 +/- 1.1 nM). Pairwise combinations of other nicotinic subunits were not blocked by 112 nM alpha-conotoxin-MII. On perfused striatal synaptosomes and slices, alpha-conotoxin-MII dose-dependently inhibited [3H]dopamine release evoked by 1 microM (+/-)-anatoxin-a with IC50 values of 24.3 +/- 2.9 and 17.3 +/- 0.1 nM, respectively. The dose-response curve was shifted to the right with increasing agonist concentrations. However, the maximal inhibition of responses achieved by alpha-conotoxin-MII (112 nM) was 44.9 +/- 5.4% for synaptosomes and 25.0 +/- 4.1% for slices, compared with an inhibition by 10 microM mecamylamine of 77.9 +/- 3.7 and 88.0 +/- 2.1%, respectively. These results suggest the presence of presynaptic alpha3beta2-like nAChRs on striatal dopaminergic terminals, but the incomplete block of (+/-)-anatoxin-a-evoked [3H]dopamine release by alpha-conotoxin-MII also supports the participation of nAChRs composed of other subunits. The lower inhibition found in slices is consistent with an additional indirect nicotinic stimulation of dopamine release via an alpha-conotoxin-MII-insensitive nAChR.
ESTHER : Kaiser_1998_J.Neurochem_70_1069
PubMedSearch : Kaiser_1998_J.Neurochem_70_1069
PubMedID: 9489727

Title : Agonist-induced up-regulation of alpha4beta2 nicotinic acetylcholine receptors in M10 cells: pharmacological and spatial definition - Whiteaker_1998_Mol.Pharmacol_53_950
Author(s) : Whiteaker P , Sharples CG , Wonnacott S
Ref : Molecular Pharmacology , 53 :950 , 1998
Abstract : Chronic nicotine up-regulates the number of high affinity nicotinic acetylcholine receptors (nAChRs) in mammalian brain. Here, we studied up-regulation of the nAChR composed of alpha4 and beta2 subunits in the M10 cell line by using [3H]epibatidine to measure nAChR in cells in situ and in membrane preparations. Cultures were exposed to drugs for 2 days before assay. All agonists up-regulated [3H]epibatidine binding sites with EC50 values typically 10-100-fold higher than their respective Ki values from competition binding assays. Maximum up-regulation ranged from 40% to 250% above control values. Maximally effective concentrations of the less efficacious agonists methylcarbamylcholine or (+/-)-epibatidine together with nicotine resulted in less up-regulation than that produced by nicotine alone, showing that they are partial up-regulatory agonists. The antagonists dihydro-beta-erythroidine, methyllycaconitine, d-tubocurarine, hexamethonium, decamethonium, and mecamylamine either failed to up-regulate [3H]epibatidine binding sites or up-regulated mildly at high concentrations. When tested at non-up-regulating concentrations, only d-tubocurarine significantly inhibited agonist-induced up-regulation; this inhibition seemed to be noncompetitive. Comparison of [3H]epibatidine displacement in intact M10 cells and membrane preparations by membrane-impermeant ligands indicated that 85% of [3H]epibatidine binding sites are intracellular. On chronic treatment with agonist, the proportion of surface receptors did not change significantly, indicating that most up-regulated [3H]epibatidine binding sites are internal. However, up-regulation is mediated at the cell surface because the impermeant ligand tetramethylammonium was as efficacious as nicotine in eliciting up-regulation, and methylcarbamylcholine (i.e., impermeant but with low efficacy) blocked nicotine induced up-regulation. Thus, agonists elicit up-regulation (mainly of intracellular receptors) by interacting with cell surface nAChRs that are not compatible with either an active or high affinity desensitized conformation.
ESTHER : Whiteaker_1998_Mol.Pharmacol_53_950
PubMedSearch : Whiteaker_1998_Mol.Pharmacol_53_950
PubMedID: 9584223

Title : Structure-activity studies of bicyclic and tricyclic analogues of methyllycaconitine -
Author(s) : Davies AR , Hardick DJ , Blagbrough IS , Potter BV , Wolstenholme AJ , Wonnacott S
Ref : Biochemical Society Transactions , 25 :545S , 1997
PubMedID: 9388759

Title : Presynaptic nicotinic modulation of dopamine release in the three ascending pathways studied by in vivo microdialysis: comparison of naive and chronic nicotine-treated rats - Marshall_1997_J.Neurochem_68_1511
Author(s) : Marshall DL , Redfern PH , Wonnacott S
Ref : Journal of Neurochemistry , 68 :1511 , 1997
Abstract : The modulation of dopamine release by presynaptic nicotinic receptors in vitro is well established, but the significance of this effect in vivo is unclear. We have characterised the effect of nicotine, locally applied via a microdialysis probe, on dopamine release from the terminal regions of three ascending dopaminergic pathways in conscious, freely moving rats. Nicotine caused a dose-dependent increase in dopamine release in the striatum, the nucleus accumbens, and, to a lesser extent, the frontal cortex. Metabolite levels were unaltered by any concentration of nicotine. Prior administration of mecamylamine via the probe abolished the nicotine-evoked increase in dopamine release, confirming the mediation of nicotinic receptors. The dose dependence of mecamylamine-sensitive, nicotine-evoked dopamine release was similar in all three brain regions. However, 10(-5) M tetrodotoxin totally blocked nicotine-stimulated dopamine release in the striatum and the accumbens but not the cortex. Daily subcutaneous injections of nicotine (0.4 mg kg-1 for 7 days) increased the response to a subsequent local application of nicotine in the striatum, and a similar trend was found in the other brain areas. The same daily dose of nicotine given as a continuous infusion had no effect, whereas infusion of 4 mg kg-1 day-1 increased the response to a subsequent nicotine challenge. The localisation and regulation of nicotinic receptors in the terminal fields of dopaminergic pathways are discussed.
ESTHER : Marshall_1997_J.Neurochem_68_1511
PubMedSearch : Marshall_1997_J.Neurochem_68_1511
PubMedID: 9084421

Title : Differential upregulation of alpha 7 and alpha 3 subunit-containing nicotinic acetylcholine receptors in rat hippocampal and PC12 cell cultures -
Author(s) : Rogers AT , Wonnacott S
Ref : Biochemical Society Transactions , 25 :544S , 1997
PubMedID: 9388758

Title : Parallel increases in [alpha-125I]bungarotoxin binding and alpha 7 nicotinic subunit immunoreactivity during the development of rat hippocampal neurons in culture - Samuel_1997_Neurosci.Lett_222_179
Author(s) : Samuel N , Wonnacott S , Lindstrom JM , Futerman AH
Ref : Neuroscience Letters , 222 :179 , 1997
Abstract : Previous studies have shown that hippocampal neurons cultured at high density express alpha-bungarotoxin binding sites and have alpha 7 nicotinic acetylcholine receptor subunit immunoreactivity [Barrantes, G.E., Rogers, A.T., Lindstrom, J. and Wonnacott, S., Brain Res., 672 (1995) 228-236]. We now examine both of these parameters in well-characterized hippocampal neurons cultured at sufficiently low densities to resolve individual neurons and their processes. The specific binding of [alpha-125I]bungarotoxin is first detectable after 3 days in culture and increases during the next 12 days in culture, reaching a maximum of approximately 30,000 binding sites per cell. This is accompanied, over the same timecourse, by an increase in immunoreactivity for two antibodies that specifically bind to the alpha 7 subunit. Both cell bodies and processes were labelled by 9 days in culture. The timecourse of alpha 7-type nicotinic receptor expression resembles that previously described for synapse formation in hippocampal cultures.
ESTHER : Samuel_1997_Neurosci.Lett_222_179
PubMedSearch : Samuel_1997_Neurosci.Lett_222_179
PubMedID: 9148244

Title : Pharmacology of nicotinic acetylcholine receptor (nAChR) upregulation in the transfected cell line, M10 -
Author(s) : Whiteaker P , Sharples CG , Wonnacott S
Ref : Biochemical Society Transactions , 25 :550S , 1997
PubMedID: 9388764

Title : ABT-089 [2-methyl-3-(2-(S)-pyrrolidinylmethoxy)pyridine]: I. A potent and selective cholinergic channel modulator with neuroprotective properties - Sullivan_1997_J.Pharmacol.Exp.Ther_283_235
Author(s) : Sullivan JP , Donnelly-Roberts D , Briggs CA , Anderson DJ , Gopalakrishnan M , Xue IC , Piattoni-Kaplan M , Molinari E , Campbell JE , McKenna DG , Gunn DE , Lin NH , Ryther KB , He Y , Holladay MW , Wonnacott S , Williams M , Arneric SP
Ref : Journal of Pharmacology & Experimental Therapeutics , 283 :235 , 1997
Abstract : Accumulating preclinical and clinical evidence data suggests that compounds that selectively activate neuronal nicotinic acetylcholine receptor (nAChR) subtypes may have therapeutic utility for the treatment of several neurological disorders. In the present study, the in vitro pharmacological properties of the novel cholinergic channel modulator ABT-089 [2-methyl-3-(2-(S)-pyrrolidinylmethoxy)pyridine], are described. In radioligand binding studies, ABT-089 was shown to display selectivity toward the high-affinity (-)-cytisine binding site present on the alpha4beta2 nAChR subtype (Ki = 16 nM) relative to the [125I]alpha-bungarotoxin binding site present on the alpha7 (Ki > or = 10,000 nM) and alpha1beta1deltagamma (Ki > 1000 nM) nAChR subtypes. In cation flux and channel current studies, ABT-089 displayed a more complex profile than (-)-nicotine having agonist, partial agonist and inhibitory activities depending on the nAChR subtype with which it interacts. ABT-089 differentially stimulated neurotransmitter release. The compound displayed a similar potency and efficacy to (-)-nicotine to facilitate ACh release (ABT-089, EC50 = 3 microM; (-)-nicotine, EC50 = 1 microM), but was markedly less potent and less efficacious than (-)-nicotine to stimulate dopamine release (ABT-089, EC50 = 1.1 microM; (-)-nicotine, EC50 = 0.04 microM). Additionally, ABT-089 was neuroprotective against the excitotoxic insults elicited by exposure to glutamate in both rat cortical cell cultures (EC50 = 10 +/- 3 microM) and differentiated human IMR32 cells (EC50 = 3 +/- 2 microM). The differential full agonist/partial agonist profile of ABT-089, as compared with (-)-nicotine and ABT-418, illustrates the complexity of nAChR activation and the potential to target responses at subclasses of the neuronal and peripheral receptors.
ESTHER : Sullivan_1997_J.Pharmacol.Exp.Ther_283_235
PubMedSearch : Sullivan_1997_J.Pharmacol.Exp.Ther_283_235
PubMedID: 9336329

Title : Presynaptic nicotinic ACh receptors - Wonnacott_1997_Trends.Neurosci_20_92
Author(s) : Wonnacott S
Ref : Trends in Neurosciences , 20 :92 , 1997
Abstract : Nicotinic ACh (nACh) receptors in the CNS are composed of a diverse array of subunits and have a range of pharmacological properties. However, despite the fact that they are ligand-gated cation channels, their physiological functions have not been determined. This has led to increased interest in presynaptic nACh receptors that act to modulate the release of transmitter from presynaptic terminals.
ESTHER : Wonnacott_1997_Trends.Neurosci_20_92
PubMedSearch : Wonnacott_1997_Trends.Neurosci_20_92
PubMedID: 9023878

Title : Nudicauline and elatine as potent norditerpenoid ligands at rat neuronal alpha-bungarotoxin binding sites: importance of the 2-(methylsuccinimido)benzoyl moiety for neuronal nicotinic acetylcholine receptor binding - Hardick_1996_J.Med.Chem_39_4860
Author(s) : Hardick DJ , Blagbrough IS , Cooper G , Potter BV , Critchley T , Wonnacott S
Ref : Journal of Medicinal Chemistry , 39 :4860 , 1996
Abstract : Methyllycaconitine (MLA, 1) is a novel, potent probe for mammalian and insect nicotinic acetylcholine receptors (nAChR) and displays remarkable selectivity toward neuronal [125I]-alpha-bungarotoxin (alpha BgTX) binding sites that correspond to alpha 7-type nAChR in mammalian brain. We have shown that, among a number of selected norditerpenoid alkaloids, elatine (2) and nudicauline (3) are equipotent with, or better than, MLA (1) in binding to brain [125I]-alpha BgTX binding sites, with IC50 values of 6.1, 1.7, and 7.6 nM, respectively. The 2-((S)-methylsuccinimido)benzoyl moiety of these ligands is crucial for high-affinity binding, whereas structural modifications to the norditerpenoid core of the ligand can be tolerated without loss of activity or selectivity. In addition to MLA (1), elatine (2), and nudicauline (3), we have examined lycoctonine (4), inuline (6), lappaconitine (7), N-desacetyllappaconitine (8), delsoline (10), delcorine (11), deltaline (12), condelphine (13), and karacoline (14). This study therefore extends the range of norditerpenoids, other than MLA, which can be used to probe this important class of nAChR. All 12 alkaloids were assessed for activity at [3H]nicotine binding sites which are considered to represent alpha 4 beta 2 nAChR. Furthermore, the 1H and 13C NMR spectroscopic data of MLA and elatine have been critically compared.
ESTHER : Hardick_1996_J.Med.Chem_39_4860
PubMedSearch : Hardick_1996_J.Med.Chem_39_4860
PubMedID: 8941400

Title : Tetrodotoxin-sensitivity of nicotine-evoked dopamine release from rat striatum - Marshall_1996_Neuropharmacol_35_1531
Author(s) : Marshall D , Soliakov L , Redfern P , Wonnacott S
Ref : Neuropharmacology , 35 :1531 , 1996
Abstract : Recent observations from synaptosome preparations have questioned the tetrodotoxin (TTX) insensitivity of nicotine-evoked release in the striatum, a characteristic previously considered diagnostic of presynaptically located nicotinic acetylcholine receptors (nAChRs). Therefore, we have undertaken a comparison of nicotine-evoked dopamine release in the presence of TTX from the rat striatum in vitro, using synaptosomes and brain slices, and in vivo, using microdialysis. In P2 and Percoll-purified synaptosome preparations, 1.5 microM TTX partially inhibited nicotine-evoked [3H]dopamine release by 54% and 37%, respectively, whereas in more intact preparations (brain slices and microdialysis) TTX completely inhibited mecamylamine-sensitive nicotine-stimulated dopamine release. These results suggest that caution should be exercised in the interpretation of TTX sensitivity of nicotine-evoked responses with regard to the location of nAChRs.
ESTHER : Marshall_1996_Neuropharmacol_35_1531
PubMedSearch : Marshall_1996_Neuropharmacol_35_1531
PubMedID: 9025100

Title : Relationship between up-regulation of nicotine binding sites in rat brain and delayed cognitive enhancement observed after chronic or acute nicotinic receptor stimulation - Abdulla_1996_Psychopharmacology.(Berl)_124_323
Author(s) : Abdulla FA , Bradbury E , Calaminici MR , Lippiello PM , Wonnacott S , Gray JA , Sinden JD
Ref : Psychopharmacology (Berl) , 124 :323 , 1996
Abstract : (-)-Nicotine tartrate (2 mg/kg), and a nicotinic agonist, RJR 2403 (1.4 mg/kg), and antagonist, mecamylamine (1 mg/kg), were administered to separate groups of rats SC twice daily for 10 days. Two other groups received the same doses of nicotine or RJR 2403 for 1 day followed by saline for 9 days. Twenty-four hours after the final injection, the rats were compared to a 10-day saline-injected group on acquisition of a hidden platform position in the Morris water maze (20 trials, 30-min inter-trial interval). The rats were killed 48 h after the last drug injection and frontal, entorhinal and posterior cingulate cortex and dorsal and ventral hippocampus assayed for [3H]-nicotine binding density. Chronic nicotine significantly increased the number of frontal and entorhinal cortical and dorsal hippocampal, but not posterior cingulate cortical or ventral hippocampal, nicotinic receptors, and improved rate of learning. Chronic mecamylamine and RJR 2403 also significantly increased the number of nicotinic receptors in frontal cortex, though not other regions, but retarded rate of learning. Nicotine given for 1 day 11 days earlier marginally increased nicotinic receptors in entorhinal cortex (but not other regions) and significantly increased rate of learning, though significantly less than 10-day nicotine. Entorhinal cortical and dorsal hippocampal nicotinic receptor numbers were positively associated with rate of learning but not performance at asymptote. Thus cognitive enhancement after chronic nicotine is in part a delayed consequence of nicotine administration 11 days earlier, and may reflect regional changes in nicotinic receptor up-regulation.
ESTHER : Abdulla_1996_Psychopharmacology.(Berl)_124_323
PubMedSearch : Abdulla_1996_Psychopharmacology.(Berl)_124_323
PubMedID: 8739547

Title : Pharmacological characterization of a nicotinic autoreceptor in rat hippocampal synaptosomes - Wilkie_1996_Neurochem.Res_21_1141
Author(s) : Wilkie GI , Hutson P , Sullivan JP , Wonnacott S
Ref : Neurochem Res , 21 :1141 , 1996
Abstract : The modulation of [3H]ACh release by nicotinic compounds was studied in superfused rat hippocampal synaptosomes loaded with [3H]choline, (-)-Nicotine (0.1-10 microM) evoked a dose-dependent increase in [3H]ACh release; higher concentrations were less effective. Nicotine-evoked release was Ca(2+)-dependent, and blocked by the nicotinic antagonists dihydro-beta-erythroidine, mecamylamine, and pempidine. The alpha 7-selective antagonist methyllycaconitine did not inhibit nicotine-evoked release when tested at 1 microM, although at 10 microM some attenuation of the response was observed. Six agonists tested were equally efficacious in stimulating [3H]ACh release, as judged by the maximum responses, and gave the following EC50 values: (+/-)-epibatidine 0.12 microM; (+)-anatoxin-a 0.14 microM; (-)-nicotine 0.99 microM; (-)-cytisine 1.06 microM; ABT-418 2.6 microM; isoarecolone 43 microM. Each agonist generated a "bell-shaped" dose response curve, suggesting desensitisation at higher concentrations. This is supported by analysis of repetitive stimulation with (-)-nicotine and (-)-cytisine: S2/S1 ratios declined sharply with increasing concentration, whereas subsequent KC1-evoked release remained constant. These results are discussed in terms of possible nicotinic receptor subtypes that might be present on hippocampal nerve terminals.
ESTHER : Wilkie_1996_Neurochem.Res_21_1141
PubMedSearch : Wilkie_1996_Neurochem.Res_21_1141
PubMedID: 8897478

Title : Voltage-sensitive Ca2+ channels involved in nicotinic receptor-mediated [3H]dopamine release from rat striatal synaptosomes - Soliakov_1996_J.Neurochem_67_163
Author(s) : Soliakov L , Wonnacott S
Ref : Journal of Neurochemistry , 67 :163 , 1996
Abstract : The potent nicotinic agonist anatoxin-a elicits mecamylamine-sensitive [3H]dopamine release from striatal synaptosomes, and this action is both Na+ and Ca2+ dependent and is blocked by Cd2+. This suggests that stimulation of presynaptic nicotinic receptors results in Na+ influx and local depolarisation that activates voltage-sensitive Ca2+ channels, which in turn provide the Ca2+ for exocytosis. Here we have investigated the subtypes of Ca2+ channels implicated in this mechanism. [3H]-Dopamine release evoked by anatoxin-a (1 microM) was partially blocked by 20 microM nifedipine, whereas KCl-evoked release was insensitive to the dihydropyridine. However, a 86Rb+ efflux assay of nicotinic receptor function suggested that nifedipine has a direct effect on the receptor, discrediting the involvement of L-type channels. The N-type Ca2+ channel blocker omega-conotoxin GVIA (1 microM) blocked anatoxin-a-evoked [3H]dopamine release by 60% but had no significant effect on 86Rb+ efflux; release evoked by both 15 and 25 mM KCl was inhibited by only 30%. The P-type channel blocker omega-agatoxin IVA (90 nM) also inhibited KCl-evoked release by approximately 30%, whereas anatoxin-a-evoked release was insensitive. The Q-type channel blocker omega-conotoxin MVIIC (1 microM) had no effect on either stimulus. These results suggest that presynaptic nicotinic receptors on striatal nerve terminals promote [3H]dopamine release by activation of N-type Ca2+ channels. In contrast, KCl-evoked [3H]dopamine release appears to involve both N-type and P-type channels.
ESTHER : Soliakov_1996_J.Neurochem_67_163
PubMedSearch : Soliakov_1996_J.Neurochem_67_163
PubMedID: 8666987

Title : Conversion of the sodium channel activator aconitine into a potent alpha 7-selective nicotinic ligand - Hardick_1995_FEBS.Lett_365_79
Author(s) : Hardick DJ , Cooper G , Scott-Ward T , Blagbrough IS , Potter BV , Wonnacott S
Ref : FEBS Letters , 365 :79 , 1995
Abstract : Methyllycaconitine (MLA) is a competitive antagonist of nicotinic acetylcholine receptors, with a remarkable preference for neuronal [125I]alpha Bgt binding sites. We have begun to investigate the structural basis of its potency and subtype selectivity. MLA is a substituted norditerpenoid alkaloid linked to a 2-(methylsuccinimido)benzoyl moiety. Hydrolysis of the ester bond in MLA to produce lycoctonine diminished affinity for rat brain [125I]alpha Bgt binding sites 2500-fold and abolished affinity for [3H]nicotine and muscle [125I]alpha Bgt binding sites. The voltage-gated Na+ channel activator aconitine, also a norditerpenoid alkaloid, but with significant structural differences from lycoctonine, displayed comparable weak or absent nicotinic activity. Addition of a 2-(methylsuccinimido)benzoyl sidechain to O-demethylated aconitine, to mimic MLA, abolished Na+ channel activation and conferred nanomolar affinity for brain [125I]alpha Bgt binding sites, comparable to that of MLA. We propose that the ester-linked 2-(methylsuccinimido)benzoyl group is necessary for nicotinic potency, but alpha 7 selectivity resides in the norditerpenoid core of the molecule.
ESTHER : Hardick_1995_FEBS.Lett_365_79
PubMedSearch : Hardick_1995_FEBS.Lett_365_79
PubMedID: 7774720

Title : Nicotine-induced upregulation of alpha bungarotoxin (alpha Bgt) binding sites in cultured rat hippocampal neurons -
Author(s) : Rogers AT , Wonnacott S
Ref : Biochemical Society Transactions , 23 :48S , 1995
PubMedID: 7758761

Title : Sensitivity of rat frontal cortical neurones to nicotine is increased by chronic administration of nicotine and by lesions of the nucleus basalis magnocellularis: comparison with numbers of [3H]nicotine binding sites - Abdulla_1995_Synapse_21_281
Author(s) : Abdulla FA , Calaminici M , Wonnacott S , Gray JA , Sinden JD , Stephenson JD
Ref : Synapse , 21 :281 , 1995
Abstract : The effects of chronic nicotine treatment and of unilateral AMPA lesion of the nucleus basalis magnocellularis (nbm) on the sensitivity of frontal cortical neurones to iontophoretically applied nicotine were studied. Chronic nicotine treatment increased the number of [3H]nicotine binding sites from 2.9 to 3.9 pmol g-1 wet weight, and increased the proportion of cortical neurones responding to nicotine from 32.3% to 60.0%. After unilateral nbm lesions, the densities of AChE-positive fibers and [3H]nicotine binding sites were reduced by approximately 97% and 55%, respectively, and the proportion of neurones responding to nicotine increased from 32.3% to 53.8%. The two treatments, chronic nicotine administration and nbm lesion, also increased the size of individual neuronal responses, prolonged their duration, and shortened the response latency. Responses to glutamate were unaffected by either procedures. The results show that the increase in [3H]nicotine binding produced by chronic nicotine administration is associated with an increased response to iontophoretically applied nicotine, suggesting that the receptor upregulation induced by the chronic treatment were functional. Less easily explained is the association between increased sensitivity of frontal cortical neurons to nicotine after nbm lesion with a decreased receptor density. It is suggested that a substantial proportion of nicotinic receptors are located presynaptically, and that their loss after lesion concealed an upregulation at postsynaptic sites.
ESTHER : Abdulla_1995_Synapse_21_281
PubMedSearch : Abdulla_1995_Synapse_21_281
PubMedID: 8869158

Title : Anatoxin-a is a potent agonist of the nicotinic acetylcholine receptor of bovine adrenal chromaffin cells - Molloy_1995_Eur.J.Pharmacol_289_447
Author(s) : Molloy L , Wonnacott S , Gallagher T , Brough PA , Livett BG
Ref : European Journal of Pharmacology , 289 :447 , 1995
Abstract : (+)-Anatoxin-a is a neurotoxic alkaloid produced by the cyanobacterium Anabaena flos-aquae. In this study synthetic (+/-)-anatoxin-a was tested on isolated bovine adrenal chromaffin cells to determine its ability to evoke secretion of endogenous catecholamines through neuronal-type nicotinic receptor activation. Anatoxin-a was found to act as a potent agonist of the secretory response of chromaffin cells with an EC50 of 1-2 microM, compared with an EC50 of 4-5 microM for nicotine. The cells responded to anatoxin-a and nicotine with bell-shaped concentration-response curves consistent with desensitisation at concentrations of anatoxin-a greater than 5 microM and of nicotine greater than 20 microM. The secretion of catecholamines stimulated by anatoxin-a was completely inhibited in a non-competitive manner by the nicotinic antagonist mecamylamine with an IC50 of 0.4-0.5 microM. In the presence of depolarising concentrations of K+ (15 or 50 mM), anatoxin-a increased the secretion of catecholamines in a concentration-dependent manner up to the same maximum as that achieved by anatoxin-a alone. It is concluded that anatoxin-a acts as a potent and selective nicotinic agonist, capable of evoking secretion of endogenous catecholamines from chromaffin cells via their neuronal-type nicotinic receptor.
ESTHER : Molloy_1995_Eur.J.Pharmacol_289_447
PubMedSearch : Molloy_1995_Eur.J.Pharmacol_289_447
PubMedID: 7556413

Title : A nicotinic acetylcholine receptor subunit from insect brain forms a non-desensitising homo-oligomeric nicotinic acetylcholine receptor when expressed in Xenopus oocytes - Amar_1995_Neurosci.Lett_199_107
Author(s) : Amar M , Thomas P , Wonnacott S , Lunt GG
Ref : Neuroscience Letters , 199 :107 , 1995
Abstract : The locust alpha-like nicotinic receptor subunit alpha L1 was expressed in Xenopus oocytes. Small but reproducible currents were elicited by application of high concentrations of nicotine, demonstrating that alpha L1 is capable of forming homo-oligomeric channels. Nicotine-evoked currents were blocked by alpha-bungarotoxin and methyllycaconitine. Comparison with chick alpha 7 receptors showed that the two receptors differ with respect to nicotine sensitivity and time course of evoked currents. Nicotine dose-response curves gave EC50 values of 24 and 830 microM for alpha 7 and alpha L1 respectively. Whereas alpha 7 responses showed characteristic fast onset and rapid desensitization within 3 s, alpha L1 currents displayed a slow onset and showed no tendency to desensitize during 45 s of agonist application. Thus alpha L1 is a novel nicotine subunit for the further exploration of structure-function relationships of ligand-gated ion channels. The question of the subunit composition of native insect receptors remains open.
ESTHER : Amar_1995_Neurosci.Lett_199_107
PubMedSearch : Amar_1995_Neurosci.Lett_199_107
PubMedID: 8584235

Title : Locomotor activation and dopamine release produced by nicotine and isoarecolone in rats - Whiteaker_1995_Br.J.Pharmacol_116_2097
Author(s) : Whiteaker P , Garcha HS , Wonnacott S , Stolerman IP
Ref : British Journal of Pharmacology , 116 :2097 , 1995
Abstract : 1. Isoarecolone was approximately 250 times less potent than nicotine as an inhibitor of [3H]-nicotine binding to rat brain membranes. Isoarecolone failed to inhibit the binding of the nicotinic ligand [125I]-alpha-bungarotoxin or of the muscarinic ligand [3H]-QNB. 2. Nicotine (0.01-30 microM) evoked the release of [3H]-dopamine from striatal and frontal cortex synaptosomes, with EC50 values of approximately 0.5 microM in each case. This release was largely mecamylamine-sensitive. 3. Isoarecolone (1-200 microM) evoked predominantly mecamylamine-sensitive dopamine release from both striatal and cortical synaptosomes, with a potency at least 20 times less than that of nicotine. The maximum effect of isoarecolone was less than that of nicotine, particularly in the frontal cortex preparation. 4. In control rats treated chronically with saline, neither nicotine nor isoarecolone had clear effects on locomotor activity at the doses tested. Chronic treatment with nicotine clearly sensitized rats to the locomotor activating effect of isoarecolone was seen at a dose about 40 times larger than that of nicotine. 5. The low potency and efficacy of isoarecolone in facilitating sensitized locomotor activity resembled its lower potency and efficacy, compared with nicotine, in evoking dopamine release in vitro. The agonist profile of the nicotinic receptor population mediating dopamine release may determine the pharmacological characteristics of consequent locomotor behaviour.
ESTHER : Whiteaker_1995_Br.J.Pharmacol_116_2097
PubMedSearch : Whiteaker_1995_Br.J.Pharmacol_116_2097
PubMedID: 8640351

Title : Nicotine increases intracellular calcium in rat hippocampal neurons via voltage-gated calcium channels - Barrantes_1995_Neurosci.Lett_196_101
Author(s) : Barrantes GE , Murphy CT , Westwick J , Wonnacott S
Ref : Neuroscience Letters , 196 :101 , 1995
Abstract : The effect of nicotinic receptor activation on intracellular calcium concentrations ([Ca2+]i) was quantitated in populations of cultured hippocampal neurons loaded with Fura-2. Nicotine (50 microM) and cytisine (50 microM) increased [Ca2+]i by 100%. This response was abolished in the presence of the nicotinic antagonist methyllycaconitine (MLA) whereas KCl-evoked increases in [Ca2+]i were insensitive to MLA. Glial cultures were unaffected by nicotine, although they did respond to glutamate with increased [Ca2+]i. In hippocampal neurons, responses to nicotinic agonists and KCl were dependent on the presence of extracellular Ca2+ and were similarly sensitive (85% inhibition) to CdCl2. These results are consistent with the presence of functional nicotinic receptors on hippocampal neurons. The receptors appear to elevate [Ca2+]i by promoting the influx of extracellular Ca2+ through voltage-gated calcium channels.
ESTHER : Barrantes_1995_Neurosci.Lett_196_101
PubMedSearch : Barrantes_1995_Neurosci.Lett_196_101
PubMedID: 7501232

Title : alpha-Bungarotoxin binding sites in rat hippocampal and cortical cultures: initial characterisation, colocalisation with alpha 7 subunits and up-regulation by chronic nicotine treatment - Barrantes_1995_Brain.Res_672_228
Author(s) : Barrantes GE , Rogers AT , Lindstrom JM , Wonnacott S
Ref : Brain Research , 672 :228 , 1995
Abstract : High density neuronal cultures from rat E18 hippocampus and cortex have been characterised with respect to cholinergic binding sites. No specific binding of [3H]nicotine or [3H]cytisine to live cells in situ was detected although the limit for detection was estimated to be 30 fmol/mg protein. Muscarinic binding sites labelled with [3H]QNB were present at a density of 0.75 pmol/mg protein. [125I]alpha-Bungarotoxin (alpha Bgt) bound to hippocampal cultures with a Bmax of 128 fmol/mg protein and a Kd of 0.6 nM; cortical cultures expressed five times fewer [125I]alpha-Bgt binding sites. Fluorescence cytochemistry with rhodamine-alpha-Bgt indicated that 95% of hippocampal neurons were labelled, compared with only 36% of cortical neurons. Average densities of 4 x 10(4) and 2 x 10(4) binding sites/cell were calculated for hippocampal and cortical cultures, respectively. Double labelling experiments with mAb307 (which recognises the rat alpha 7 nicotinic receptor subunit) and rhodamine-alpha-Bgt gave coincident labelling patterns, supporting the correlation between the alpha 7 subunit and Bgt-sensitive neuronal nicotinic receptor. Treatment of hippocampal cultures with 10 microM nicotine for 14 days elicited a 40% increase in the numbers of [125I]alpha-Bgt binding sites, mimicking the up-regulation observed in in vivo studies. Primary cultures offer a useful in vitro system for investigating the expression and regulation of brain alpha-Bgt-sensitive receptors.
ESTHER : Barrantes_1995_Brain.Res_672_228
PubMedSearch : Barrantes_1995_Brain.Res_672_228
PubMedID: 7749744

Title : Anatoxin-a-evoked [3H]dopamine release from rat striatal synaptosomes - Soliakov_1995_Neuropharmacol_34_1535
Author(s) : Soliakov L , Gallagher T , Wonnacott S
Ref : Neuropharmacology , 34 :1535 , 1995
Abstract : Presynaptic nicotinic acetylcholine receptors on striatal nerve terminals modulate the release of dopamine. Using rat striatal synaptosomes loaded with [3H]dopamine, we have characterized the action of the selective nicotinic agonist, (+/-)anatoxin-a, with respect to [3H]dopamine release, in order to explore the mechanisms coupling nicotinic receptor activation to exocytosis. Anatoxin-a evoked [3H]dopamine release in a concentration-dependent and mecamylamine-sensitive manner, EC50 = 0.11 microM. The maximum [3H]dopamine release elicited by anatoxin-a was only 20% of the maximum elicited by KCl depolarization; there was no additivity between anatoxin-a and sub-maximal concentrations of KCl. Both agents stimulated Ca(2+)-dependent release that was equally sensitive to inhibition by 200 microM Cd2+. This result suggests that anatoxin-a-stimulated exocytosis is mediated by Ca2+ influx via voltage-sensitive Ca2+ channels, with little contribution from Ca2+ entering directly through the nicotinic receptor channel. This view is supported by the abolition of anatoxin-a-evoked [3H]dopamine release in Na(+)-depleted medium. A partial (40%) inhibition by tetrodotoxin was observed. These data suggest that activation of presynaptic nicotinic acetylcholine receptors by anatoxin-a results in an influx of Na+, producing sufficient local depolarization to open voltage-sensitive Ca2+ and Na+ channels. The latter may then amplify the response, activating further Ca2+ channels. The particular voltage-sensitive Ca2+ channels involved remain to be determined.
ESTHER : Soliakov_1995_Neuropharmacol_34_1535
PubMedSearch : Soliakov_1995_Neuropharmacol_34_1535
PubMedID: 8606800

Title : Neurotoxins: nature's untapped bounty -
Author(s) : Wonnacott S , Dajas F
Ref : Trends in Pharmacological Sciences , 15 :1 , 1994
PubMedID: 8140650

Title : Nicotinic acetylcholine receptors in primary cultures of hippocampal neurons: pharmacology and Ca++ permeability -
Author(s) : Barrantes GE , Westwick J , Wonnacott S
Ref : Biochemical Society Transactions , 22 :294S , 1994
PubMedID: 7821553

Title : Behavioural and ligand-binding studies in rats with 1-acetyl-4-methylpiperazine, a novel nicotinic agonist - Garcha_1993_Psychopharmacology.(Berl)_110_347
Author(s) : Garcha HS , Thomas P , Spivak CE , Wonnacott S , Stolerman IP
Ref : Psychopharmacology (Berl) , 110 :347 , 1993
Abstract : The novel nicotinic agonist 1-acetyl-4-methylpiperazine (AMP) has been studied in ligand-binding and behavioural studies. AMP methiodide potently inhibited [3H]-(-)-nicotine and [125I]-alpha-bungarotoxin binding to P2 membranes from rat brain and [125I]-alpha-bungarotoxin binding to rat skeletal muscles. AMP HCl also inhibited nicotinic binding, but it was 100 times less potent than AMP methiodide. In behavioural studies, AMP HCl reduced locomotor activity of experimentally naive rats and mecamylamine blocked this effect. In rats receiving (-)-nicotine chronically, AMP HCl did not increase locomotor activity consistently or to the same extent as (-)-nicotine. In rats trained to discriminate (-)-nicotine from saline in a two-bar operant conditioning procedure with food reinforcement, there was generalization to AMP HCl, but only at doses that reduced the overall rate of responding. The potency and effectiveness of AMP relative to (-)-nicotine varied across the different behavioural procedures. The results suggest that the pharmacodynamic action of AMP differs from that of (-)-nicotine and that it usefully extends the range of agonists that can be used as probes for central nicotinic mechanisms.
ESTHER : Garcha_1993_Psychopharmacology.(Berl)_110_347
PubMedSearch : Garcha_1993_Psychopharmacology.(Berl)_110_347
PubMedID: 7831430

Title : Upregulation of nicotinic receptors following continuous infusion of nicotine is brain-region-specific - Sanderson_1993_Brain.Res_617_349
Author(s) : Sanderson EM , Drasdo AL , McCrea K , Wonnacott S
Ref : Brain Research , 617 :349 , 1993
Abstract : Rats receiving 4 mg nicotine/kg/day via implanted minipumps sustained plasma nicotine concentrations of 40 ng/ml throughout two weeks of nicotine infusion. Numbers of brain [3H]nicotine binding sites were increased by about 50% in cortex and hippocampus whereas numbers of [3H]nicotine binding sites in striatum were unaffected by nicotine treatment at either of the timepoints examined (7, 14 days). Cortical [125I] alpha-bungarotoxin and [3H]QNB binding sites were also unchanged. The regional selectivity of nicotinic receptor modulation may reflect the low dose of nicotine used and the mode of administration. The changes observed may be pertinent to the continuous administration of nicotine in man, via transdermal nicotine patches.
ESTHER : Sanderson_1993_Brain.Res_617_349
PubMedSearch : Sanderson_1993_Brain.Res_617_349
PubMedID: 8402163

Title : Hippocampal nicotinic autoreceptors modulate acetylcholine release -
Author(s) : Wilkie GI , Hutson PH , Stephens MW , Whiting P , Wonnacott S
Ref : Biochemical Society Transactions , 21 :429 , 1993
PubMedID: 8359505

Title : Methyl lycaconitine: A novel nicotinic antagonist - Drasdo_1992_Mol.Cell.Neurosci_3_237
Author(s) : Drasdo A , Caulfield M , Bertrand D , Bertrand S , Wonnacott S
Ref : Molecular & Cellular Neurosciences , 3 :237 , 1992
Abstract : Methyllycaconitine (MLA), a natural toxin from Delphinium seeds, was investigated for its ability to antagonize nicotinic responses in several preparations representing different subtypes of neuronal nicotinic acetylcholine receptor. A presynaptic nicotinic receptor mediating dopamine release from rat striatal synaptosomes was blocked by 10 muM MLA, in good agreement with its Ki of 4 muM for inhibition of [(3)H]nicotine binding to striatal membranes. Nicotinic responses in rat superior cervical ganglia were similarly blocked by MLA, and this inhibition was readily reversible. Functional expression of the chick alpha3nalpha1 and alpha4nalpha1 receptor subtypes in Xenopus oocytes confirmed that MLA is a nicotinic antagonist at both receptor subtypes, with IC(50) values of 0.08 and 0.65 muM, respectively. Inhibition by MLA was voltage independent and competitive with agonist concentration. Thus MLA is a useful addition to the nicotinic pharmacopoeia.
ESTHER : Drasdo_1992_Mol.Cell.Neurosci_3_237
PubMedSearch : Drasdo_1992_Mol.Cell.Neurosci_3_237
PubMedID: 19912865

Title : Homoanatoxin: a potent analogue of anatoxin-A - Wonnacott_1992_Biochem.Pharmacol_43_419
Author(s) : Wonnacott S , Swanson KL , Albuquerque EX , Huby NJ , Thompson P , Gallagher T
Ref : Biochemical Pharmacology , 43 :419 , 1992
Abstract : The natural toxin anatoxin-a (AnTx) is a potent nicotinic agonist that is valuable for the study of nicotinic receptors. We have synthesized 2-(propan-1-oxo-1-yl)-9-azabicyclo[4.2.1]non-2-ene, the homologue of AnTx in which the side-chain is extended by one methylene unit from a methyl to an ethyl ketone. This chemistry would allow the generation of a tritiated product and the homologue, designated homoanatoxin (HomoAnTx), has been characterized here with that aim in mind. In competition binding assays at neuronal nicotinic ligand binding sites characterized by [3H]nicotine and [125I]-alpha bungarotoxin, HomoAnTx retained the same potency as the parent molecule, with Ki values of 7.5 nM and 1.1 microM, respectively. In contrast, it showed little inhibition of muscarinic binding defined by [3H]-quinuclidinyl benzilate. HomoAnTx is a potent nicotinic agonist in frog muscle contracture assays, having four times the potency of carbamylcholine and one tenth of the activity of AnTx itself. The N-methylated version of HomoAnTx was more than two orders of magnitude weaker in both functional and binding assays. The successful synthesis of HomoAnTx with retention of high nicotinic potency offers a route for the generation of novel, potent radiolabelled nicotinic ligands.
ESTHER : Wonnacott_1992_Biochem.Pharmacol_43_419
PubMedSearch : Wonnacott_1992_Biochem.Pharmacol_43_419
PubMedID: 1540199

Title : Blockade of nicotinic currents in hippocampal neurons defines methyllycaconitine as a potent and specific receptor antagonist - Alkondon_1992_Mol.Pharmacol_41_802
Author(s) : Alkondon M , Pereira EF , Wonnacott S , Albuquerque EX
Ref : Molecular Pharmacology , 41 :802 , 1992
Abstract : Methyllycaconitine, a toxin isolated from the seeds of Delphinium brownii, inhibited acetylcholine- and anatoxin-induced whole-cell currents in cultured fetal rat hippocampal neurons, at picomolar concentrations. This antagonism was specific, concentration dependent, reversible, and voltage independent. Furthermore, methyllycaconitine inhibited 125I-alpha-bungarotoxin binding to adult rat hippocampal membranes, protected against the alpha-bungarotoxin-induced pseudoirreversible blockade of nicotinic currents, and shifted the concentration-response curve of acetylcholine to the right in fetal rat hippocampal neurons, suggesting a possible competitive mode of action for this toxin. Remarkably low concentrations of methyllycaconitine (1-1000 fM) decreased the frequency of anatoxin-induced single-channel openings, with no detectable decrease in the mean channel open time. These actions of methyllycaconitine commend this neurotoxin for the characterization of the alpha-bungarotoxin-sensitive subclass of neuronal nicotinic receptors, which has hitherto eluded functional demonstration.
ESTHER : Alkondon_1992_Mol.Pharmacol_41_802
PubMedSearch : Alkondon_1992_Mol.Pharmacol_41_802
PubMedID: 1569927

Title : Characterization of nicotinic receptor-mediated [3H]dopamine release from synaptosomes prepared from mouse striatum - Grady_1992_J.Neurochem_59_848
Author(s) : Grady S , Marks MJ , Wonnacott S , Collins AC
Ref : Journal of Neurochemistry , 59 :848 , 1992
Abstract : This study establishes that presynaptic nicotinic receptors modulate dopamine release in the mouse striatum. Nicotinic agonists elicit a dose-dependent increase in the release of [3H]dopamine from synaptosomes prepared from mouse striatum. At low concentrations, this release is Ca2+ dependent, whereas at higher concentrations Ca(2+)-independent, mecamylamine-insensitive release was also observed. The Ca(2+)-dependent nicotine-evoked release was not blocked by alpha-bungarotoxin but was effectively blocked by neuronal bungarotoxin as well as several other nicotinic receptor antagonists. The relationship between potency for stimulation of release for agonists and potency for inhibition of release for antagonists was compared to the affinity of these compounds for the [3H]nicotine binding site. The overall correlation between release and binding potency was not high, but the drugs may be classified into separate groups, each of which has a high correlation with binding. This finding suggests either that more than one nicotinic receptor regulates dopamine release or that not all agonists interact with the same receptor in an identical fashion.
ESTHER : Grady_1992_J.Neurochem_59_848
PubMedSearch : Grady_1992_J.Neurochem_59_848
PubMedID: 1494911

Title : Nicotinic pharmacology of anatoxin analogs. I. Side chain structure-activity relationships at peripheral agonist and noncompetitive antagonist sites - Swanson_1991_J.Pharmacol.Exp.Ther_259_377
Author(s) : Swanson KL , Aronstam RS , Wonnacott S , Rapoport H , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 259 :377 , 1991
Abstract : Anatoxin analogs were designed to evaluate the importance of H-bonding, planarity, size and steric configuration of the anatoxin side chain moiety with regard to nicotinic potency and efficacy. This report examines the actions of these analogs on the somatic nicotinic acetylcholine receptor at two different loci: the agonist recognition site and the ion channel site. Agonist effects were evaluated using stimulation of contracture and radioligand binding competition for [125I]alpha bungarotoxin sites in Rana pipiens muscle, and stimulation of [3H]perhydrohistrionicotoxin binding and competition for [125I]alpha bungarotoxin sites in Torpedo californica electric organ. Antagonist effects were evident in the inhibition of neurally evoked twitch of the frog sciatic nerve-sartorius muscle preparation and in inhibition of [3H]perhydrohistrionicotoxin binding to Torpedo receptors. The affinity of these analogs for the agonist locus was consistently associated with activation of the AChR. Our results show that side chain steric configuration has an important role in affinity of the (+)-anatoxin-a analogs for the nicotinic acetylcholine receptor ion channel sites. Several analogs also revealed stereospecific noncompetitive actions. The (+)-anatoxin-a-related structures are important probes for characterizing both agonist and ion channel target sites on the peripheral nicotinic receptor.
ESTHER : Swanson_1991_J.Pharmacol.Exp.Ther_259_377
PubMedSearch : Swanson_1991_J.Pharmacol.Exp.Ther_259_377
PubMedID: 1920124

Title : Nicotinic pharmacology of anatoxin analogs. II. Side chain structure-activity relationships at neuronal nicotinic ligand binding sites - Wonnacott_1991_J.Pharmacol.Exp.Ther_259_387
Author(s) : Wonnacott S , Jackman S , Swanson KL , Rapoport H , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 259 :387 , 1991
Abstract : Eighteen analogs of (+)-anatoxin-a were evaluated for nicotinic potency at two putative nicotinic acetylcholine receptor sites in the central nervous system. The affinities of the analogs for [3H]nicotine and [125I]alpha-bungarotoxin binding sites were compared. This series of analogs, with modifications to the side chain moieties of the parent structure, enables the importance (for nicotinic binding) of hydrogen bonding strength, planarity, size and steric configuration of this region of the molecule to be assessed. These studies confirm the importance of the side chain stereochemistry and the subordinate role of H-bonding strength of anatoxin analogs. Of all the analogs tested, the parent compound (+)-anatoxin-a is the most potent competitor of ligand binding. Although all analogs have higher affinity at the [3H](-)-nicotine site compared to the alpha-[125I]bungarotoxin site, the rank order of potency is generally the same at both central nervous system sites, and agrees with the order at the muscle nicotinic receptor. However, the simple methoxyamide and the isoxazolidide analogs appear more selective for the neuronal nicotinic receptor subtype identified by [3H](-)-nicotine, indicative of structural differences among the agonist recognition sites.
ESTHER : Wonnacott_1991_J.Pharmacol.Exp.Ther_259_387
PubMedSearch : Wonnacott_1991_J.Pharmacol.Exp.Ther_259_387
PubMedID: 1920126

Title : Isolation of hippocampal synaptosomes on Percoll gradients: cholinergic markers and ligand binding sites - Thorne_1991_J.Neurochem_56_479
Author(s) : Thorne B , Wonnacott S , Dunkley PR
Ref : Journal of Neurochemistry , 56 :479 , 1991
Abstract : The S1 Percoll procedure, devised empirically for cortical tissue, provides highly purified, functionally viable synaptosomes on a four-step Percoll gradient. Here, for the first time, the procedure has been applied to rat hippocampus, and the gradient fractions have been analysed with respect to cholinergic markers and the synaptosomal index, lactate dehydrogenase. The presynaptic cholinergic markers choline acetyltransferase and [3H]choline uptake were most enriched in fraction 4. In contrast, acetylcholinesterase activity was broadly distributed across the gradient, consistent with the separation of synaptic plasma membranes (in fractions 1 and 2) from synaptosomes (in fractions 3 and 4). This is supported by the recovery of muscarinic binding sites labelled with [3H]quinuclidinylbenzilate in fractions 1 and 2. (-)-[3H]-Nicotine binding sites, however, were most enriched in fraction 4, consistent with their predominantly presynaptic localisation in the CNS. These results demonstrate the applicability of the S1 Percoll method to discrete brain regions for the recovery of homogeneous and viable synaptosome fractions. The separation of presynaptic terminals from post-synaptic membranes is a further advantage of this technique.
ESTHER : Thorne_1991_J.Neurochem_56_479
PubMedSearch : Thorne_1991_J.Neurochem_56_479
PubMedID: 1846398

Title : Neuronal nicotinic receptors: functional correlates of ligand binding sites -
Author(s) : Wonnacott S
Ref : Biochemical Society Transactions , 19 :121 , 1991
PubMedID: 2037134

Title : The relevance of receptor binding studies to tobacco research - Wonnacott_1991_Br.J.Addict_86_537
Author(s) : Wonnacott S
Ref : Br J Addict , 86 :537 , 1991
Abstract : The initial event in the process by which nicotine acts on the nervous system is its interaction with specific receptor molecules in the neuronal membrane. This interaction can be characterized in radioligand binding assays, using [3H]nicotine. While this approach has generated a detailed description of recognition sites for nicotine in the brain, caution is necessary in correlating these binding proteins with physiologically relevant sites of action of nicotine. Of particular interest is the disparity between effective nicotine concentrations in binding and functional assays, which may reflect receptor desensitization and could be pertinent to the development of dependence. Receptor heterogeneity, reflecting subtypes of nicotine receptor having different radioligand binding specificities and differing properties, introduces a new dimension in the consideration of nicotine's actions. Thus, receptor binding assays have an important role to play, in conjunction with other approaches, in unravelling the complex mechanisms that can lead to nicotine dependence.
ESTHER : Wonnacott_1991_Br.J.Addict_86_537
PubMedSearch : Wonnacott_1991_Br.J.Addict_86_537
PubMedID: 1650270

Title : Presynaptic nicotinic receptors and the modulation of transmitter release - Wonnacott_1990_Ciba.Found.Symp_152_87
Author(s) : Wonnacott S , Drasdo A , Sanderson E , Rowell P
Ref : Ciba Found Symp , 152 :87 , 1990
Abstract : Nicotine is increasingly recognized to promote transmitter release in the brain by a direct action on presynaptic terminals. Pharmacological evidence indicates that this action is mediated by nicotinic receptors. From their sensitivity to mecamylamine, neosurugatoxin and neuronal bungarotoxin these presynaptic receptors can be distinguished from alpha-bungarotoxin-sensitive muscle-type nicotinic receptors, and can be correlated with [3H] nicotine binding sites in the brain. The release of many transmitters in different brain regions is susceptible to stimulation by nicotine, but this effect is not ubiquitous. However, lesioning and subcellular fractionation studies suggest that the majority of brain nicotine receptors are located presynaptically, so that a direct influence of nicotine on transmitter release assumes considerable importance. Although the sensitivity of presynaptic receptors is such that they are likely to be partially activated by doses of nicotine obtained by smoking, the desensitization-induced up-regulation of nicotinic binding sites that follows chronic nicotine treatment raises questions about their functional status during tobacco usage. Chronic administration of the agonist (+)anatoxin-a also up-regulated [3H] nicotine binding sites, and led to increased nicotine-evoked transmitter release in vitro. This could have implications for the involvement of these receptors during withdrawal.
ESTHER : Wonnacott_1990_Ciba.Found.Symp_152_87
PubMedSearch : Wonnacott_1990_Ciba.Found.Symp_152_87
PubMedID: 1976493

Title : Separation of pre- and post-synaptic receptors on Percoll gradients -
Author(s) : Wonnacott S , Thorne B
Ref : Biochemical Society Transactions , 18 :885 , 1990
PubMedID: 1964650

Title : The anticonvulsant MK-801 interacts with peripheral and central nicotinic acetylcholine receptor ion channels - Ramoa_1990_J.Pharmacol.Exp.Ther_254_71
Author(s) : Ramoa AS , Alkondon M , Aracava Y , Irons J , Lunt GG , Deshpande SS , Wonnacott S , Aronstam RS , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 254 :71 , 1990
Abstract : The effects of MK-801 [( +]-5-methyl-10,11-dihydro-5H-di-benzo[a, d]cyclohepten-5,10-imine) on peripheral and central nicotinic receptors were studied using electrophysiological and biochemical techniques. MK-801 depressed the peak amplitude and accelerated the decay of end-plate currents. The drug (1-10 microM) decreased the frequency of activation of acetylcholine (ACh)-induced single-channel currents in addition to shortening the mean open and burst times of channels activated by either ACh or (+)anatoxin-a (AnTX). MK-801 (10-40 microM) depressed the single potentials and trains of ACh and AnTX-induced potentials in chronically denervated rat soleus muscles. MK-801 blocked the twitch responses (20-100 microM) of both frog sartorius and rat diaphragm muscles evoked by stimulation of their respective nerves. Also this drug (less than 1 microM) decreased the frequency of channels activated by AnTX or ACh in outside-out patch membranes of rat retinal ganglion cells with minimal changes in the channel open time. MK-801 (10-25 microM) depressed (-)nicotine-evoked gamma-amino[2,3-3H]butyric acid release from rat hippocampal synaptosomes; however, it failed to affect the binding of [3H](-)nicotine to brain membranes and also failed to interfere with the binding of [125I]alpha-bungarotoxin to either frog muscle or Torpedo membranes. On the other hand, MK-801 inhibited the binding of [3H]perhydrohistrionicotoxin to Torpedo membranes and such an effect was more pronounced in the presence of carbamylcholine. Neither AnTX nor any other nicotinic agonist increased the binding of [3H]MK-801 to the N-methyl-D-aspartate receptor ion channel complex. The actions of MK-801 were evident at concentrations comparable with those needed to block N-methyl-D-aspartate receptors. These results demonstrate the existence of at least three different types of nicotinic AChR, all of which were blocked noncompetitively by MK-801.
ESTHER : Ramoa_1990_J.Pharmacol.Exp.Ther_254_71
PubMedSearch : Ramoa_1990_J.Pharmacol.Exp.Ther_254_71
PubMedID: 1694895

Title : Nicotinic modulation of [3H]dopamine release from striatal synaptosomes: pharmacological characterisation - Rapier_1990_J.Neurochem_54_937
Author(s) : Rapier C , Lunt GG , Wonnacott S
Ref : Journal of Neurochemistry , 54 :937 , 1990
Abstract : Presynaptic nicotinic acetylcholine receptors on striatal nerve terminals modulate the release of dopamine. We have compared the effects of a number of nicotinic agonists and antagonists on a perfused synaptosome preparation preloaded with [3H]dopamine. (-)-Nicotine, acetylcholine, and the nicotinic agonists cytisine and 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), at micromolar concentrations, stimulated the release of [3H]dopamine from striatal nerve terminals. Carbamylcholine was a much weaker agonist. The actions of (-)-nicotine, cytisine, and DMPP were inhibited by low concentrations of the nicotinic antagonists dihydro-beta-erythroidine, mecamylamine, pempidine, and neosurugatoxin; alpha-bungarotoxin was without effect, and extending the time of exposure to this toxin resulted in only very modest inhibition. This pharmacology points to a specific nicotinic receptor mechanism that is clearly distinct from that at the neuromuscular junction. Atropine failed to antagonise the effects of acetylcholine and carbamylcholine, suggesting that no muscarinic component is involved. The nicotinic receptor ligands (-)-[3H]nicotine and 125I-alpha-bungarotoxin bound to specific sites enriched in the synaptosome preparation. Drugs tested on the perfused synaptosomes were examined for their ability to interact with these two ligand binding sites in brain membranes. The differential sensitivity to the neurotoxins alpha-bungarotoxin and neosurugatoxin of the 125I-alpha-bungarotoxin and (-)-[3H]nicotine binding sites, respectively, leads to a tentative correlation of the (-)-[3H]nicotine site with the presynaptic nicotinic receptor on striatal nerve terminals.
ESTHER : Rapier_1990_J.Neurochem_54_937
PubMedSearch : Rapier_1990_J.Neurochem_54_937
PubMedID: 2303820

Title : Evidence for functional activity of up-regulated nicotine binding sites in rat striatal synaptosomes - Rowell_1990_J.Neurochem_55_2105
Author(s) : Rowell PP , Wonnacott S
Ref : Journal of Neurochemistry , 55 :2105 , 1990
Abstract : A number of studies have found that the chronic administration of nicotine causes an increase in the density of nicotinic binding sites in the brain, but it is not known whether these additional binding sites are functionally active receptors. In this study, the effects of 1-week administration of the potent nicotinic agonist, (+)-anatoxin-a (96 nmol/day via osmotic minipumps), was assessed on [3H]nicotine binding and [3H]dopamine uptake and release in rat striatal synaptosomes. Chronic (+)-anatoxin-a treatment resulted in a 32% increase in the Bmax of [3H]nicotine binding in anatoxin-treated animals compared to control. There was a 43% increase in the activity of 3 microM nicotine to release [3H]dopamine from synaptosomes of anatoxin-treated animals, but the release induced by 20 mM K+ depolarization was unaffected. There was no effect of chronic (+)-anatoxin-a treatment on the uptake of [3H]dopamine. A strong positive correlation (r = 0.64) was found between the density of [3H]nicotine binding sites and the nicotine-induced stimulation of [3H]dopamine release in individual animals. These results indicate that (+)-anatoxin-a, like nicotine, produces an up-regulation of nicotine binding sites following chronic administration, and that these additional sites are functional receptors capable of mediating the release of dopamine from striatal synaptosomes.
ESTHER : Rowell_1990_J.Neurochem_55_2105
PubMedSearch : Rowell_1990_J.Neurochem_55_2105
PubMedID: 2230812

Title : Methyllycaconitine: a selective probe for neuronal alpha-bungarotoxin binding sites - Ward_1990_FEBS.Lett_270_45
Author(s) : Ward JM , Cockcroft VB , Lunt GG , Smillie FS , Wonnacott S
Ref : FEBS Letters , 270 :45 , 1990
Abstract : The ability of methyllycaconitine (MLA) to inhibit the binding of [125I]alpha-bungarotoxin to rat brain membranes, frog and human muscle extracts and the human muscle cell line TE671 has been measured. MLA showed a markedly higher affinity for the rat brain site (Ki 1.4 x 10(-9) M) than for the muscle receptors (Ki 10(-5)-10(-6) M). Structure modelling techniques were used to fit the structure of MLA to a nicotinic pharmacophore model. MLA is the first low molecular weight ligand to be shown to discriminate between muscle nicotinic receptors and their alpha-bungarotoxin-binding counterpart in the brain, and as such may be a useful structural probe for pursuing the structural and functional properties of the neuronal protein.
ESTHER : Ward_1990_FEBS.Lett_270_45
PubMedSearch : Ward_1990_FEBS.Lett_270_45
PubMedID: 2226787

Title : The paradox of nicotinic acetylcholine receptor upregulation by nicotine -
Author(s) : Wonnacott S
Ref : Trends in Pharmacological Sciences , 11 :216 , 1990
PubMedID: 2200178

Title : Presynaptic modulation of transmitter release by nicotinic receptors -
Author(s) : Wonnacott S , Irons J , Rapier C , Thorne B , Lunt GG
Ref : Prog Brain Res , 79 :157 , 1989
PubMedID: 2573910

Title : Agonist recognition site of the peripheral acetylcholine receptor ion channel complex differentiates the enantiomers of nicotine - Rozental_1989_J.Pharmacol.Exp.Ther_251_395
Author(s) : Rozental R , Aracava Y , Scoble GT , Swanson KL , Wonnacott S , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 251 :395 , 1989
Abstract : The multiple actions of nicotine enantiomers at the peripheral nicotinic acetylcholine receptor were evaluated using electrophysiological and biochemical techniques. The alpha-bungarotoxin binding site showed a 6-fold greater affinity for (-)-nicotine than for the (+)-isomer, and this stereoselectivity was reflected in differences in the ability of the alkaloids to activate physiological responses in the forms of single ion channel currents, endplate depolarizations and muscle contractures. (-)-Nicotine was also more potent to induce slow desensitization. In contrast, both (-)- and (+)-nicotine were equipotent as ion channel blockers. Ion channel blockade occurred at effective agonist concentrations for (+)-nicotine but above the effective concentration for (-)-nicotine. The rapid and reversible interaction of nicotine enantiomers with the ion channel occurred at concentrations which implicate a significant contribution of channel blockade to the inhibition of indirect muscle twitch. The agonistic and ion channel blocking effects of the nicotine enantiomers provide important clues regarding the mechanisms by which nicotine may affect central nervous system nicotinic receptors.
ESTHER : Rozental_1989_J.Pharmacol.Exp.Ther_251_395
PubMedSearch : Rozental_1989_J.Pharmacol.Exp.Ther_251_395
PubMedID: 2478693

Title : Nicotinic involvement in ACh-DA interaction? -
Author(s) : Wonnacott S
Ref : Trends in Pharmacological Sciences , 10 :395 , 1989
PubMedID: 2617664

Title : Methyllycaconitine and (+)-anatoxin-a differentiate between nicotinic receptors in vertebrate and invertebrate nervous systems - Macallan_1988_FEBS.Lett_226_357
Author(s) : Macallan DR , Lunt GG , Wonnacott S , Swanson KL , Rapoport H , Albuquerque EX
Ref : FEBS Letters , 226 :357 , 1988
Abstract : Specific high-affinity binding sites for 125I-alpha-bungarotoxin and (-)-[3H]nicotine have been measured in rat brain and locust (Schistocerca gregaria) ganglia. The binding sites for 125I-alpha-bungarotoxin had similar Kd values of 1.5 x 10(-9) and 0.8 x 10(-9) M for rat and locust preparations, respectively; the corresponding values for the (-)-[3H]nicotine-binding site were 9.3 x 10(-9) and 1.7 x 10(-7) M. Methyllycaconitine (MLA) potently inhibited 125I-alpha-bungarotoxin binding in both rat and locust. MLA was a less effective inhibitor of (-)-[3H]nicotine binding whereas (+)-anatoxin-a was a very potent inhibitor at this site in the rat but not in the locust. These data suggest that (+)-anatoxin-a is a useful probe for the high-affinity nicotine-binding receptor in vertebrate brain, whereas MLA is a preferential probe for the subclass of receptor that binds alpha-bungarotoxin.
ESTHER : Macallan_1988_FEBS.Lett_226_357
PubMedSearch : Macallan_1988_FEBS.Lett_226_357
PubMedID: 3338564

Title : Stereoselective nicotine-induced release of dopamine from striatal synaptosomes: concentration dependence and repetitive stimulation - Rapier_1988_J.Neurochem_50_1123
Author(s) : Rapier C , Lunt GG , Wonnacott S
Ref : Journal of Neurochemistry , 50 :1123 , 1988
Abstract : Using a sensitive perfusion system we have studied the nicotine-induced release of [3H]dopamine ([( 3H]DA) from striatal synaptosomes. Nicotine-evoked release was concentration dependent with an EC50 of 3.8 microM. The response to 1 microM nicotine was comparable to that to 16 mM K+; 10 microM veratridine evoked a larger response. All three stimuli were Ca2+ dependent but only the response to veratridine was blocked by tetrodotoxin. Repetitive stimulations by 1 microM (-)-nicotine (100 microliters) at 30-min intervals resulted in similar levels of [3H]DA release; higher concentrations of (-)-nicotine resulted in an attenuation of the response particularly following the third stimulation. This may reflect desensitisation or tachyphylaxis of the presynaptic nicotinic receptor. The action of nicotine was markedly stereoselective: a 100-fold higher concentration of (+)-nicotine was necessary to evoke the same level of response as 1 microM (-)-nicotine. It is proposed that these presynaptic nicotinic receptors on striatal terminals are equivalent to high-affinity nicotine binding sites described in mammalian brain.
ESTHER : Rapier_1988_J.Neurochem_50_1123
PubMedSearch : Rapier_1988_J.Neurochem_50_1123
PubMedID: 3346670

Title : [3H](-)nicotine binding sites in fetal human brain - Cairns_1988_Brain.Res_475_1
Author(s) : Cairns NJ , Wonnacott S
Ref : Brain Research , 475 :1 , 1988
Abstract : The development of putative nicotinic binding sites in brains from human fetuses of 12-19 weeks gestation was studied. The binding of [3H](-)nicotine to fetal human brain membranes, using a rapid filtration method, was saturable and stereospecific. Scatchard analysis revealed a single class of high affinity sites with a Kd of 1.5 +/- 0.5 nM and a Bmax of 4.5 +/- 1.9 fmol/mg protein (n = 11). [3H](-)nicotine binding increased between the ages of 12 and 19 weeks in human fetal brain (r = 0.63, n = 20, P less than 0.01). In competition studies nicotinic agonists were the most effective in inhibiting [3H](-)nicotine binding whereas antagonists were relatively ineffective. Ki values for displacing ligands in the presence of [3H](-)nicotine were: cytisine, 1.6 nM; (-)nicotine, 16 nM; (+) nicotine, 510 nM; dihydro-beta-erythroidine, 1.9 microM; dimethyl-4-phenylpiperazinium, 6.5 microM; choline chloride, 25 microM. Atropine and alpha-bungarotoxin failed to inhibit binding up to 50 microM. Comparison of dissected brain regions revealed regional variations in the density of nicotinic binding sites: specific binding of [3H](-)nicotine was greatest in the nucleus basalis of Meynert, globus pallidus, caudate-putamen and thalamus, and lowest in the medulla. These results are interpreted in relation to the development of functional cholinergic transmission in human fetal brain, and the potential vulnerability of this system to maternal tobacco usage.
ESTHER : Cairns_1988_Brain.Res_475_1
PubMedSearch : Cairns_1988_Brain.Res_475_1
PubMedID: 3214718

Title : The neurotoxin histrionicotoxin interacts with the putative ion channel of the nicotinic acetylcholine receptors in the central nervous system - Rapier_1987_FEBS.Lett_212_292
Author(s) : Rapier C , Wonnacott S , Lunt GG , Albuquerque EX
Ref : FEBS Letters , 212 :292 , 1987
Abstract : Perhydrohistrionicotoxin at micromolar concentrations blocked the nicotine-evoked transmitter release from perfused striatal (dopaminergic) and hippocampal (cholinergic) nerve terminals. Perhydrohistrionicotoxin failed to compete with [3H]nicotine for its high-affinity binding site in rat brain, suggesting that the action of this toxin on central nicotinic receptors is noncompetitive. From the dose-response curve, 50% inhibition of nicotine-evoked striatal dopamine release occurred at 5 microM perhydrohistrionicotoxin, a value similar to that obtained in frog sartorius muscle and Electrophorus electroplax. This close agreement may suggest that the ionic channel of the presynaptic nicotinic acetylcholine receptor of brain neurons has similar properties to those of the peripheral receptor.
ESTHER : Rapier_1987_FEBS.Lett_212_292
PubMedSearch : Rapier_1987_FEBS.Lett_212_292
PubMedID: 2434360

Title : Nicotinic acetylcholine receptors in cultured neurons from the hippocampus and brain stem of the rat characterized by single channel recording - Aracava_1987_FEBS.Lett_222_63
Author(s) : Aracava Y , Deshpande SS , Swanson KL , Rapoport H , Wonnacott S , Lunt G , Albuquerque EX
Ref : FEBS Letters , 222 :63 , 1987
Abstract : Single channel recording techniques have been applied to neurons cultured from the hippocampus and the respiratory area of the brain stem of fetal rats in order to search for nicotinic acetylcholine receptors (nAChR) in the central nervous system. In addition to acetylcholine (ACh), the potent and specific agonist (+)-anatoxin-a was also used to characterize nicotinic channels. nAChRs were concentrated on the somal surface near the base of the apical dendrite, and in some patches their density was sufficient to record 2 or more channel openings simultaneously. Although a multiplicity of conductance states was also evident, the predominant population showed a single channel conductance of 20 pS at 10 degrees C. Thus, these neuronal nAChRs resembled the embryonic or denervated-type nAChRs in muscle. However, channel opening and closing kinetics were faster than reported for similar conductance channels in muscle. Therefore the nicotinic channels described here are similar but not identical to those of the well-characterized muscle nAChR, in agreement with biochemical, pharmacological, and molecular genetic studies on brain AChR.
ESTHER : Aracava_1987_FEBS.Lett_222_63
PubMedSearch : Aracava_1987_FEBS.Lett_222_63
PubMedID: 2443390

Title : Brain nicotine binding sites -
Author(s) : Wonnacott S
Ref : Hum Toxicol , 6 :343 , 1987
PubMedID: 3315965

Title : Subcellular fractionation and distribution of cholinergic binding sites in fetal human brain - Whyte_1986_Neurochem.Res_11_1011
Author(s) : Whyte J , Harrison R , Lunt GG , Wonnacott S
Ref : Neurochem Res , 11 :1011 , 1986
Abstract : Conventional subcellular fractionation techniques have been applied to human fetal brain (13-15 weeks gestation) and the fractions have been characterized by assaying for marker enzymes, cholinergic binding sites and electron microscopy. Fractionation of the homogenate resulted in a nuclear pellet (P1), a crude mitochondrial pellet (P2) and a supernatant (S2). Further resolution of the P2 fraction by density gradient centrifugation resulted in two bands at the gradient interfaces and a pellet. The P2 and subsequently the P2B fraction contained intact plasma membrane profiles as judged by the predominance of adenylate cyclase activity and the presence of occluded lactate dehydrogenase which constituted over 70% of the total activity in these fractions. Morphological examination of the gradient fractions revealed that the P2B fraction contains membrane bound structures which resemble synaptosomes prepared from neonatal rat brain. These structures have a granular matrix in which mitochondria and frequently, neurofilaments were observed. Very few synaptic vesicles were present and there was no evidence for post synaptic attachments. The cholinergic markers choline acetyltransferase, acetylcholinesterase and receptor sites defined by quinuclidinyl benzilate and alpha-bungarotoxin binding were enriched in fractions P2 and P2B which contained the bulk of nerve ending particles. This enriched preparation of fetal synaptosomes may be valuable for functional studies on pre-synaptic terminals in developing brain.
ESTHER : Whyte_1986_Neurochem.Res_11_1011
PubMedSearch : Whyte_1986_Neurochem.Res_11_1011
PubMedID: 3748272

Title : alpha-Bungarotoxin binds to low-affinity nicotine binding sites in rat brain - Wonnacott_1986_J.Neurochem_47_1706
Author(s) : Wonnacott S
Ref : Journal of Neurochemistry , 47 :1706 , 1986
Abstract : Reported differences in the pharmacology and distribution of [3H]nicotine and [125I]alpha-bungarotoxin binding sites in mammalian brain suggest that these ligands label separate receptor sites. Affinity purification of an alpha-bungarotoxin binding protein from rat brain failed to copurify the high-affinity nicotine binding site, which remained in the nonbound soluble fraction after the affinity chromatography step. This confirms the independence of these putative receptor sites. Nevertheless, the binding of [125I]alpha-bungarotoxin to P2 membranes was inhibited by (-)-nicotine (Ki = 9 X 10(-6) M), and this sensitivity was preserved after affinity purification. It is proposed that alpha-bungarotoxin binds to a population of low-affinity nicotine binding sites. Comparison of the enantiomers of nicotine in competition studies at both radioligand binding sites revealed an 80-fold preference for the (-) form at the high-affinity [3H]nicotine binding site, whereas the site labelled by [125I]alpha-bungarotoxin displayed little stereoselectivity. In this respect, the brain alpha-bungarotoxin binding site resembles the nicotinic acetylcholine receptor from Torpedo electric organ.
ESTHER : Wonnacott_1986_J.Neurochem_47_1706
PubMedSearch : Wonnacott_1986_J.Neurochem_47_1706
PubMedID: 3772372

Title : Neosurugatoxin blocks nicotinic acetylcholine receptors in the brain - Rapier_1985_Neurochem.Int_7_389
Author(s) : Rapier C , Harrison R , Lunt GG , Wonnacott S
Ref : Neurochem Int , 7 :389 , 1985
Abstract : Neosurugatoxin, a neurotoxin isolated from the Japanese ivory mollusc (Babylonia japonica ) is a nicotinic antagonist with a specificity towards ganglionic nicotinic receptors. At low concentration (5 x 10(?8) M) neosurugatoxin inhibited the release of [(3)H]dopamine evoked by 1,1-dimethyl-4-phenylpiperazinium (DMPP) from rat striatal nerve terminals, without affecting the response to K(+)-depolarisation. In contrast, ?bungarotoxin did not antagonise the action of DMPP. Neosurugatoxin also inhibited [(3)H] nicotine binding to rat brain membranes but had no effect on [(125)I]?bungarotoxin binding to the same tissue preparation. These results support the view that functional nicotinic receptors in the CNS resemble ganglionic nicotinic receptors. Neosurugatoxin has considerable potential as a useful probe for such receptors in the brain.
ESTHER : Rapier_1985_Neurochem.Int_7_389
PubMedSearch : Rapier_1985_Neurochem.Int_7_389
PubMedID: 20492940

Title : Properties of alpha-bungarotoxin binding sites in foetal human brain - Whyte_1985_Neurochem.Int_7_515
Author(s) : Whyte J , Harrison R , Lunt GG , Wonnacott S
Ref : Neurochem Int , 7 :515 , 1985
Abstract : Maximum levels of binding of alpha-bungarotoxin to foetal human brain membranes were found to remain essentially constant at 30-50 fmol/mg protein (1.1-1.5 pmol/g wet weight in whole brain) between gestational ages of 10 and 24 weeks. Equilibrium binding of alpha-bungarotoxin to both membranes and to detergent extracts showed saturable specific binding to a single class of sites with K(d) (app) values of 3.5 x 10(?9) M and 2.4 x 10(?9) M respectively. Association rate constants, determined from time courses of binding of alpha-bungarotoxin to membranes and detergent extracts, were 2.3 x 10(5) M(?1) sec(?1) and 2.6 x 10(5) M(?1) sec(?1) respectively. Dissociation of alpha-bungarotoxin from both membrane and detergent extracts showed a rapid initial rate with T (1 2 ) approx 15 min which, in the case of the detergent extract, was followed by a slower dissociation accounting for the remaining 20% of the bound ligand. Competition studies with a number of cholinergic ligands indicated that the alpha-bungarotoxin-binding sites in foetal brain display a predominantly nicotinic profile.
ESTHER : Whyte_1985_Neurochem.Int_7_515
PubMedSearch : Whyte_1985_Neurochem.Int_7_515
PubMedID: 20492956

Title : Antibodies to nicotinic acetylcholine receptors used to probe the structural and functional relationships between brain alpha-bungarotoxin binding sites and nicotinic receptors - Mills_1984_Neurochem.Int_6_249
Author(s) : Mills A , Wonnacott S
Ref : Neurochem Int , 6 :249 , 1984
Abstract : Antibodies against peripheral nicotinic acetylcholine receptors (nAChR) were used to determine the proportion of brain alpha-bungarotoxin binding sites that are immunologically related to the peripheral nAChR. The alpha-bungarotoxin binding component partially purified from rat brain was labelled with [(125)I]alpha-bungarotoxin and reacted with increasing concentrations of rabbit anti(nAChR) antisera. At least 75% of the brain protein could be immunoprecipitated by rabbit anti(rat muscle junctional nAChR) antiserum (M) whereas an antiserum against Torpedo nAChR (J) was without effect and clearly failed to cross-react with the brain component. Both antisera precipitated 100% of [(125)I]alpha-bungarotoxin-labelled nAChR from Torpedo marmorata. The lower precipitation of the brain protein was not a consequence of [(125)I]alpha-bungarotoxin dissociating during the precipitation. We conclude that the majority of alpha-bungarotoxin binding sites in brain are clearly recognised by the crossreacting antiserum. Release of [(3)H]dopamine from striatal synaptosomes could be elicited by nicotine in a dose-dependent manner and the response was prevented by the ganglionic blocker mecamylamine, although antagonism by alpha-bungarotoxin was less clearcut. Preincubation of the synaptosomes with antiserum M resulted in a statistically significant decrease in the [(3)H]dopamine response to nicotine at all agonist concentrations tested. Antiserum J, however, had no consistent effect on the response. Thus the actions of the antisera parallel their ability to recognise the brain alpha-bungarotoxin binding component. We conclude that the cholinergic regulation of dopamine release is in part mediated through a nAChR that is immunologically related to the nAChR of the neuromuscular junction and to the alpha-bungarotoxin binding component that can be isolated from rat brain.
ESTHER : Mills_1984_Neurochem.Int_6_249
PubMedSearch : Mills_1984_Neurochem.Int_6_249
PubMedID: 20488045

Title : Immunological cross-reactivity between the alpha-bungarotoxin-binding component from rat brain and nicotinic acetylcholine receptor - Wonnacott_1982_J.Neuroimmunol_3_1
Author(s) : Wonnacott S , Harrison R , Lunt G
Ref : Journal of Neuroimmunology , 3 :1 , 1982
Abstract : An alpha-bungarotoxin-binding protein was partially purified from rat brain and, when complexed with [125I] alpha-bungarotoxin, was shown to behave as a single radiolabelled protein that is distinct from the similarly complexed nAChR from Torpedo marmorata. The alpha-bungarotoxin-binding protein was used as antigen in radioimmunoassays for rabbit anti-(rat muscle nAChR) and rabbit anti-(Torpedo nAChR) antibodies, giving titres approximately 5% and 0.5%, respectively, of those obtained by using homologous antigen in the same assay.
ESTHER : Wonnacott_1982_J.Neuroimmunol_3_1
PubMedSearch : Wonnacott_1982_J.Neuroimmunol_3_1
PubMedID: 7096564

Title : The effect of acetylcholine release on choline fluxes in isolated synaptic terminals - Marchbanks_1981_J.Neurochem_36_379
Author(s) : Marchbanks RM , Wonnacott S , Rubio MA
Ref : Journal of Neurochemistry , 36 :379 , 1981
Abstract : As in intact tissues, choline influx into synaptosomes is enhanced after a period of depolarization induced release of acetylcholine. The activation of uptake is dependent on the presence of Ca2+ and inhibited by high Mg2+ concentrations in the medium during depolarization. Choline transport in erythrocytes was not activated by prior treatment with potassium. The permeability constant of the synaptosome membrane to choline was found to be 2.7 x 10(-8) cm . s-1 and to acetylcholine 1.8 x 10(-8) cm . s-1. Choline influx has been studied after pre-loading synaptosomes with choline. Different radiolabels were used to measure efflux of preloaded choline and influx simultaneously. Isotopic dilution in flux studies was estimated and corrected for. Influx was stimulated by high internal concentrations of choline, and efflux similarly stimulated by high outside concentrations of choline. The maximal influx and efflux at saturating opposite concentrations of choline were equal with a value of about 500 pmol . min-1 per mg synaptosomal protein. A reciprocating carrier would explain the equality of the maximal influx and efflux. Acetylcholine competes with choline for binding to the carrier but is itself hardly transported. Increased acetylcholine concentrations were shown to inhibit both choline influx and efflux from the trans position. Raising intrasynaptosomal acetylcholine concentrations by pre-loading abolished the stimulation of influx by prior depolarization. It is proposed that high concentrations of acetylcholine immobilize the carrier on the inside of the synaptic membrane. The stimulation of choline influx consequent upon depolarization is caused by release of ACh which results in relief of this immobilisation. The enhanced supply of choline achieved by this mechanism is likely to be important in maintaining stores of the acetylcholine in vivo.
ESTHER : Marchbanks_1981_J.Neurochem_36_379
PubMedSearch : Marchbanks_1981_J.Neurochem_36_379
PubMedID: 7463066

Title : Interrelationship of carbohydrate and the alpha-toxin binding site on the acetylcholine receptor from Torpedo marmorata -
Author(s) : Wonnacott S , Harrison R , Lunt GG
Ref : Life Sciences , 27 :1769 , 1980
PubMedID: 6257994

Title : Interrelationship of concanavalin-A-binding and antigenic sites on the acetylcholine receptor from Torpedo marmorata - Wonnacott_1980_Eur.J.Biochem_108_621
Author(s) : Wonnacott S , Harrison R , Lunt GG , Barkas T
Ref : European Journal of Biochemistry , 108 :621 , 1980
Abstract : A preparation of purified 125I-labelled acetylcholine receptor was shown to bind to concanavalin A and to be totally bound by rabbit antiserum to Torpedo acetylcholine receptor. Pre-incubation of the receptor with F(ab')2 and Fab fragments from antibodies against Torpedo acetylcholine receptor, or with corresponding fragments from control immunoglobulin G showed that subsequent binding of the receptor to concanavalin A was specifically inhibited to a maximum of approximately 25% by the immune fragments. Treatment of acetylcholine receptor with periodate or with glycosidases apparently destroyed or removed carbohydrate residues without affecting the antigenicity of the receptor as assessed by radioimmunoassay. These results suggest that although there is a steric interrelatonship between the antigenic and concanavalin-A-binding sites of the receptor the latter sites do not contain its major antigenic determinants.
ESTHER : Wonnacott_1980_Eur.J.Biochem_108_621
PubMedSearch : Wonnacott_1980_Eur.J.Biochem_108_621
PubMedID: 6157534

Title : Depolarisation-induced release of ATP from cortical synaptosomes is not associated with acetylcholine release -
Author(s) : White T , Potter P , Wonnacott S
Ref : Journal of Neurochemistry , 34 :1109 , 1980
PubMedID: 6246199

Title : Inhibition by botulinum toxin of acetylcholine release from synaptosomes: latency of action and the role of gangliosides -
Author(s) : Wonnacott S
Ref : Journal of Neurochemistry , 34 :1567 , 1980
PubMedID: 7381483

Title : Relationship of choline uptake to acetylcholine synthesis and release -
Author(s) : Marchbanks RM , Wonnacott S
Ref : Prog Brain Res , 49 :77 , 1979
PubMedID: 390614

Title : Ca2+ uptake by synaptosomes and its effect on the inhibition of acetylcholine release by botulinum toxin -
Author(s) : Wonnacott S , Marchbanks RM , Fiol C
Ref : Journal of Neurochemistry , 30 :1127 , 1978
PubMedID: 351143

Title : Inhibition by botulinum toxin of depolarization-evoked release of (14C)acetylcholine from synaptosomes in vitro - Wonnacott_1976_Biochem.J_156_701
Author(s) : Wonnacott S , Marchbanks RM
Ref : Biochemical Journal , 156 :701 , 1976
Abstract : 1. Cerebral-cortex synaptosomes were shown to synthesize (14C)acetylcholine after incubation with (14C)choline, and 25mM-KCl released (14C)acetylcholine (but not (14C)choline) into the medium by a Ca2+-dependent and Mg2+-sensitive process. 2. The K+-stimulated release of (14C)acetylcholine was inhibited by more than 80% after preincubation of the synaptosomes with 10(5) mouse lethal doses of botulinum toxin/ml. (14C)choline uptake, (14C)acetylcholine synthesis, intrasynaptosomal K+ and occluded lactate dehydrogenase were unaffected by the toxin. It also failed to prevent the K+-stimulated release of (3H)noradrenaline and (14C)glycine from synaptosomes. 3. Fractionation of hypo-osmotically shocked synaptosomes revealed that more than 75% of the radioactive acetylcholine was in the cytoplasmic compartment, although the vesicle pellet contained more total acetylcholine than the cytoplasmic pool. Consequently the specific radioactivity of acetylcholine in the cytoplasmic pool was almost 5 times that of the vesicles. This distribution was unaffected by preincubation with botulinum toxin. It is concluded that the toxin acts directly on the release of acetylcholine, rather than influencing its storage. 4. After K+-stimulation, toxin-inhibited synaptosomes contained increased amounts of total acetylcholine, which suggests that its rate of synthesis is controlled by depolarization rather than release.
ESTHER : Wonnacott_1976_Biochem.J_156_701
PubMedSearch : Wonnacott_1976_Biochem.J_156_701
PubMedID: 949350

Title : A radioenzymic assay for acetylcholine and the problems encountered during its development and application -
Author(s) : Wonnacott S , Marchbanks RM
Ref : Biochemical Society Transactions , 3 :102 , 1975
PubMedID: 236201