Aronstam RS

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Full name : Aronstam Robert S

First name : Robert S

Mail : Department of Biological Sciences, Missouri University of Science & Technology, 400 W 11th St, Rolla MO 65409

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Country : USA

Email : aronstam@mst.edu

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References (100)

Title : Oxidative stress disruption of receptor-mediated calcium signaling mechanisms - Tang_2013_J.Biomed.Sci_20_48
Author(s) : Tang TH , Chang CT , Wang HJ , Erickson JD , Reichard RA , Martin AG , Shannon EK , Martin AL , Huang YW , Aronstam RS
Ref : J Biomed Sci , 20 :48 , 2013
Abstract : BACKGROUND: Oxidative stress increases the cytosolic content of calcium in the cytoplasm through a combination of effects on calcium pumps, exchangers, channels and binding proteins. In this study, oxidative stress was produced by exposure to tert-butyl hydroperoxide (tBHP); cell viability was assessed using a dye reduction assay; receptor binding was characterized using [3H]N-methylscopolamine ([3H]MS); and cytosolic and luminal endoplasmic reticulum (ER) calcium concentrations ([Ca2+]i and [Ca2+]L, respectively) were measured by fluorescent imaging.
RESULTS: Activation of M3 muscarinic receptors induced a biphasic increase in [Ca2+]i: an initial, inositol trisphosphate (IP3)-mediated release of Ca2+ from endoplasmic reticulum (ER) stores followed by a sustained phase of Ca2+ entry (i.e., store-operated calcium entry; SOCE). Under non-cytotoxic conditions, tBHP increased resting [Ca2+]i; a 90 minute exposure to tBHP (0.5-10 mM ) increased [Ca2+]i from 26 to up to 127 nM and decreased [Ca2+]L by 55%. The initial response to 10 muM carbamylcholine was depressed by tBHP in the absence, but not the presence, of extracellular calcium. SOCE, however, was depressed in both the presence and absence of extracellular calcium. Acute exposure to tBHP did not block calcium influx through open SOCE channels. Activation of SOCE following thapsigargin-induced depletion of ER calcium was depressed by tBHP exposure. In calcium-free media, tBHP depressed both SOCE and the extent of thapsigargin-induced release of Ca2+ from the ER. M3 receptor binding parameters (ligand affinity, guanine nucleotide sensitivity, allosteric modulation) were not affected by exposure to tBHP.
CONCLUSIONS: Oxidative stress induced by tBHP affected several aspects of M3 receptor signaling pathway in CHO cells, including resting [Ca2+]i, [Ca2+]L, IP3 receptor mediated release of calcium from the ER, and calcium entry through the SOCE. tBHP had little effect on M3 receptor binding or G protein coupling. Thus, oxidative stress affects multiple aspects of calcium homeostasis and calcium dependent signaling.
ESTHER : Tang_2013_J.Biomed.Sci_20_48
PubMedSearch : Tang_2013_J.Biomed.Sci_20_48
PubMedID: 23844974

Title : Honokiol blocks store operated calcium entry in CHO cells expressing the M3 muscarinic receptor: honokiol and muscarinic signaling - Wang_2013_J.Biomed.Sci_20_11
Author(s) : Wang HJ , Martin AG , Chao PK , Reichard RA , Martin AL , Huang YW , Chan MH , Aronstam RS
Ref : J Biomed Sci , 20 :11 , 2013
Abstract : BACKGROUND: Honokiol, a cell-permeable phenolic compound derived from the bark of magnolia trees and present in Asian herbal teas, has a unique array of pharmacological actions, including the inhibition of multiple autonomic responses. We determined the effects of honokiol on calcium signaling underlying transmission mediated by human M3 muscarinic receptors expressed in Chinese hamster ovary (CHO) cells. Receptor binding was determined in radiolabelled ligand binding assays; changes in intracellular calcium concentrations were determined using a fura-2 ratiometric imaging protocol; cytotoxicity was determined using a dye reduction assay.
RESULTS: Honokiol had a potent (EC50 approximately 5 mumol/l) inhibitory effect on store operated calcium entry (SOCE) that was induced by activation of the M3 receptors. This effect was specific, rapid and partially reversible, and was seen at concentrations not associated with cytotoxicity, inhibition of IP3 receptor-mediated calcium release, depletion of ER calcium stores, or disruption of M3 receptor binding.
CONCLUSIONS: It is likely that an inhibition of SOCE contributes to honokiol disruption of parasympathetic motor functions, as well as many of its beneficial pharmacological properties.
ESTHER : Wang_2013_J.Biomed.Sci_20_11
PubMedSearch : Wang_2013_J.Biomed.Sci_20_11
PubMedID: 23432810

Title : Zinc oxide nanoparticle disruption of store-operated calcium entry in a muscarinic receptor signaling pathway - Wang_2010_Toxicol.In.Vitro_24_1953
Author(s) : Wang HJ , Growcock AC , Tang TH , O'Hara J , Huang YW , Aronstam RS
Ref : Toxicol In Vitro , 24 :1953 , 2010
Abstract : The influences of ZnO nanoparticles on cellular responses to activation of muscarinic receptors were studied in Chinese hamster ovary cells expressing the human M3 muscarinic acetylcholine receptor. ZnO particles (20 nm) induced cytotoxicity in a time and concentration-dependent manner: following a 24h exposure, toxicity was minimal at concentrations below 20 mug/ml but virtually complete at concentrations above 28 mug/ml. ZnO particles did not affect antagonist binding to M3 receptors or allosteric ligand effects, but increased agonist binding affinity while eliminating guanine nucleotide sensitivity. At a noncytotoxic concentration (10 mug/ml), ZnO increased resting [Ca(2+)](i) from 40 to 130 nM without compromising calcium homeostatic mechanisms. ZnO particles had minimal effects on IP3- or thapsigargin-mediated release of intracellular calcium from the endoplasmic reticulum, but strongly inhibited store-operated calcium entry (capacitive calcium entry). The latter effect was seen as (1) a decrease in the plateau phase of the response and (2) a decrease in Ca(2+) entry upon introduction of calcium to the extracellular medium following thapsigargin-induced depletion of calcium from the endoplasmic reticulum (EC50's approximately 2 mug/ml). Thus, ZnO nanoparticles interfere with two specific aspects of the M3 signaling pathway, agonist binding and store-operated calcium entry.
ESTHER : Wang_2010_Toxicol.In.Vitro_24_1953
PubMedSearch : Wang_2010_Toxicol.In.Vitro_24_1953
PubMedID: 20708676

Title : Involvement of protein kinase C and protein kinase A in the muscarinic receptor signalling pathways mediating phospholipase C activation, arachidonic acid release and calcium mobilisation - May_1999_Cell.Signal_11_179
Author(s) : May LG , Johnson S , Krebs S , Newman A , Aronstam RS
Ref : Cell Signal , 11 :179 , 1999
Abstract : The involvement of protein kinase C (PKC) and protein kinase A (PKA) in cholinergic signalling in CHO cells expressing the M3 subtype of the muscarinic acetylcholine receptor was examined. Muscarinic signalling was assessed by measuring carbachol-induced activation of phospholipase C (PLC), arachidonic acid release, and calcium mobilisation. Carbachol activation of PLC was not altered by inhibition of PKC with chelerythrine chloride, bisindolylmaleimide or chronic treatment with phorbol myristate acetate (PMA). Activation of PKC by acute treatment with PMA was similarly without effect. In contrast, inhibition of PKC blocked carbachol stimulation of arachidonic acid release. Likewise, PKC inhibition resulted in a decreased ability of carbachol to mobilise calcium, whereas PKC activation potentiated calcium mobilisation. Inhibition of PKA with H89 or Rp-cAMP did not alter the ability of carbachol to activate PLC. Similarly, PKA activation with Sp-cAMP or forskolin had no effect on PLC stimulation by carbachol. Carbachol-mediated release of arachidonic acid was decreased by H89 but only slightly increased by forskolin. Forskolin also increased calcium mobilisation by carbachol. These results suggest a function for PKC and PKA in M3 stimulation of arachidonic acid release and calcium mobilisation but not in PLC activation.
ESTHER : May_1999_Cell.Signal_11_179
PubMedSearch : May_1999_Cell.Signal_11_179
PubMedID: 10353692

Title : M3 muscarinic acetylcholine receptors regulate cytoplasmic myosin by a process involving RhoA and requiring conventional protein kinase C isoforms - Strassheim_1999_J.Biol.Chem_274_18675
Author(s) : Strassheim D , May LG , Varker KA , Puhl HL , Phelps SH , Porter RA , Aronstam RS , Noti JD , Williams CL
Ref : Journal of Biological Chemistry , 274 :18675 , 1999
Abstract : Although muscarinic acetylcholine receptors (mAChR) regulate the activity of smooth muscle myosin, the effects of mAChR activation on cytoplasmic myosin have not been characterized. We found that activation of transfected human M3 mAChR induces the phosphorylation of myosin light chains (MLC) and the formation of myosin-containing stress fibers in Chinese hamster ovary (CHO-m3) cells. Direct activation of protein kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA) also induces myosin light chain phosphorylation and myosin reorganization in CHO-m3 cells. Conventional (alpha), novel (delta), and atypical (iota) PKC isoforms are activated by mAChR stimulation or PMA treatment in CHO-m3 cells, as indicated by PKC translocation or degradation. mAChR-mediated myosin reorganization is abolished by inhibiting conventional PKC isoforms with Go6976 (IC50 = 0.4 microM), calphostin C (IC50 = 2.4 microM), or chelerythrine (IC50 = 8.0 microM). Stable expression of dominant negative RhoAAsn-19 diminishes, but does not abolish, mAChR-mediated myosin reorganization in the CHO-m3 cells. Similarly, mAChR-mediated myosin reorganization is diminished, but not abolished, in CHO-m3 cells which are multi-nucleate due to inactivation of Rho with C3 exoenzyme. Expression of dominant negative RhoAAsn-19 or inactivation of RhoA with C3 exoenzyme does not affect PMA-induced myosin reorganization. These findings indicate that the PKC-mediated pathway of myosin reorganization (induced either by M3 mAChR activation or PMA treatment) can continue to operate even when RhoA activity is diminished in CHO-m3 cells. Conventional PKC isoforms and RhoA may participate in separate but parallel pathways induced by M3 mAChR activation to regulate cytoplasmic myosin. Changes in cytoplasmic myosin elicited by M3 mAChR activation may contribute to the unique ability of these receptors to regulate cell morphology, adhesion, and proliferation.
ESTHER : Strassheim_1999_J.Biol.Chem_274_18675
PubMedSearch : Strassheim_1999_J.Biol.Chem_274_18675
PubMedID: 10373480

Title : Characterization of ethyl (3-quinuclidinyl) acetate (EQA) as a ligand for acetylcholine receptors - Canney_1998_Life.Sci_63_PL329
Author(s) : Canney DJ , Zhang M , Doukas PH , Massakowski C , Aronstam RS , Gattu M , Buccafusco JJ , Aronstam R
Ref : Life Sciences , 63 :PL329 , 1998
Abstract : Recent studies suggest that neuronal nicotinic acetylcholine receptors (nAChRs) may play a role in several CNS disorders and that subtype selective nicotinic ligands may be useful in the treatment of these disorders. Ethyl (3-quinuclidinyl)acetate (EQA) is a bulky, reverse-ester analog of ACh, that produces signs of cholinergic stimulation that may be nicotinic in origin The objective of the present study was to further evaluate EQA as a potential cholinergic ligand. Behavioral studies, smooth muscle assays, and radioligand binding assays were performed on this novel ligand. The effects of EQA on blood pressure and acetylcholinesterase activity were also evaluated. The results of the study suggest that EQA is an agonist at peripheral nicotinic acetylcholine receptors and may have antagonist properties at central nicotinic receptors.
ESTHER : Canney_1998_Life.Sci_63_PL329
PubMedSearch : Canney_1998_Life.Sci_63_PL329
PubMedID: 9851313

Title : Ethanol disrupts carbamylcholine-stimulated release of arachidonic acid from Chinese hamster ovary cells expressing different subtypes of human muscarinic receptor - Stair_1998_Alcohol.Clin.Exp.Res_22_409
Author(s) : Stair SM , May LG , Puhl HL , Phelps SH , Williams CL , Aronstam RS
Ref : Alcohol Clin Exp Res , 22 :409 , 1998
Abstract : Ethanol disrupts signal transduction mediated by a variety of G-protein coupled receptors. We examined the effects of ethanol on arachidonic acid release mediated by muscarinic acetylcholine receptors. Chinese hamster ovary (CHO) cells transfected with the different subtypes of human muscarinic receptors (M1 to M5) were incubated with [3H]arachidonic acid ([3H]AA) for 18 hr, washed, and exposed to the cholinergic agonist carbamylcholine for 15 min. Carbamylcholine induced [3H]AA release from CHO cells expressing M1, M3, or M5, but not M2 or M4, muscarinic receptors. Dose response curves revealed that carbamylcholine stimulated [3H]AA release by up to 12-fold with an ECo of approximately 0.4 microM; maximal responses were obtained with 10 microM carbamylcholine. Exposure of M1-, M3-, or M5-expressing cells to ethanol for 5 min before stimulating with carbamylcholine reduced [3H]AA release by 40 to 65%; 50% of the maximal inhibition was obtained with an ethanol concentration of 30 to 50 mM. Ethanol did not affect basal [3H]AA release measured in the absence of carbamylcholine. Dose response curves suggest that ethanol acts as a noncompetitive inhibitor of muscarinic receptor-induced [3H]AA release insofar as maximal [3H]AA release was depressed in the presence of ethanol with no apparent change in the EC50 for stimulation by carbamylcholine. Exposure of CHO cells to 38 mM ethanol for 48 hr increased [3H]AA release induced by carbamylcholine without affecting basal [3H]AA release or altering the EC50 for carbamylcholine. These results indicate that ethanol acutely inhibits muscarinic receptor signaling through the arachidonic acid pathway in a noncompetitive manner, but chronically enhances muscarinic signaling through the same pathway.
ESTHER : Stair_1998_Alcohol.Clin.Exp.Res_22_409
PubMedSearch : Stair_1998_Alcohol.Clin.Exp.Res_22_409
PubMedID: 9581647

Title : Influence of acute and chronic ethanol treatment on muscarinic responses and receptor expression in Chinese hamster ovary cells - Chang_1997_Biochem.Pharmacol_54_833
Author(s) : Chang ZL , Puhl HL , May LG , Williams CL , Aronstam RS
Ref : Biochemical Pharmacology , 54 :833 , 1997
Abstract : The influence of ethanol on the muscarinic receptor-mediated release of inositol phosphate from Chinese hamster ovary (CHO) cells stably transfected with one of the five subtypes of muscarinic acetylcholine receptor was determined. In CHO cells expressing M3 muscarinic receptors (CHO-M3), carbamylcholine increased muscarinic receptor-induced release of inositol phosphate by 150-350% following a 15-min incubation with an EC50 of approximately 30 microM. Maximal responses were obtained with 1 mM carbamylcholine, while responses to 10 mM carbamylcholine were somewhat less than maximal. Preincubation with atropine for 10 min inhibited the response with an IC50 of approximately 30 nM. CHO cells transfected with M1, M3, and M5 receptors displayed a similar pattern of activity; CHO cells transfected with M2 and M4, as well as untransfected cells, were unresponsive to carbamylcholine. Ethanol acutely inhibited the response of CHO-M3 cells to carbamylcholine by 15% at 18 mM and by 47% at 180 mM (the highest concentration examined). CHO-M3 cells were incubated with 50 mM ethanol for 48 hr. This treatment did not affect the number of cells or their protein content (113 pg/cell). The expression of M3 muscarinic receptors (determined using [3H]N-methylscopolamine) increased from 1.34 +/- 0.23 to 1.75 +/- 0.16 pmol/mg protein (P < 0.05). In contrast, carbamylcholine-stimulated release of inositol phosphate was depressed by 40-70% in four experiments. Concentration-response analyses indicated a non-competitive inhibitory mechanism. This dissociation of muscarinic receptor expression and muscarinic signaling suggests a compensatory increase in receptor expression in response to chronic inhibition of muscarinic signaling by ethanol.
ESTHER : Chang_1997_Biochem.Pharmacol_54_833
PubMedSearch : Chang_1997_Biochem.Pharmacol_54_833
PubMedID: 9353138

Title : Inhibition of M3 muscarinic acetylcholine receptor-mediated Ca2+ influx and intracellular Ca2+ mobilization in neuroblastoma cells by the Ca2+\/calmodulin-dependent protein kinase inhibitor 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-trosyl]-4-phenylpiperazin e (KN-62) - Puhl_1997_Biochem.Pharmacol_53_1107
Author(s) : Puhl HL , Raman PS , Williams CL , Aronstam RS
Ref : Biochemical Pharmacology , 53 :1107 , 1997
Abstract : The role of Ca2+/calmodulin-dependent protein kinase (CaM kinase; EC 2.7.1.123) in the generation of Ca2+ signals by muscarinic acetylcholine receptors (mAChR) was studied. Changes in intracellular Ca2+ concentrations ([Ca2+]i) induced by mAChR activation were monitored in SK-N-SH human neuroblastoma cells using the dye Fura-2. SK-N-SH cells express M3 mAChR, as well as CaM kinase types II and IV, which are specifically inhibited by the CaM kinase antagonist KN-62 (1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazi ne). Carbamylcholine (100 microM) elicited an initial transient peak in [Ca2+]i due to mobilization of Ca2+ from internal stores, followed by a sustained elevation in [Ca2+]i that depended on the influx of extracellular Ca2+ and which was inhibited by EGTA and Ni2+. These mAChR-induced Ca2+ signals were diminished to an equal extent by preincubating the cells with 0.01 to 100 microM KN-62. KN-62 inhibited mAChR-induced Ca2+ influx and mobilization from internal stores by about 25-30%, producing a half-maximal effect at approximately 1 microM. In contrast, KN-62 (25 microM) almost completely abolished carbamylcholine-stimulated entry of divalent cations through Mn2+-permeant channels, as revealed by Mn2+ quenching of Fura-2 fluorescence. KN-62 also almost completely abolished Ca2+ influx induced by depolarization of the cells with 25 mM K+ (IC50 = 3 microM). These results suggest that CaM kinases regulate both the mobilization of intracellular Ca2+ and the stimulation of Ca2+ influx that are induced by mAChR activation, and indicate that the mAChR-induced influx of Ca2+ occurs through Ca2+ channels other than, or in addition to, the voltage-gated calcium channels or Mn2+-permeant channels which are inhibited by KN-62.
ESTHER : Puhl_1997_Biochem.Pharmacol_53_1107
PubMedSearch : Puhl_1997_Biochem.Pharmacol_53_1107
PubMedID: 9175715

Title : Characterization of the muscarinic receptor subtypes in the bovine corneal epithelial cells - Socci_1996_J.Ocul.Pharmacol.Ther_12_259
Author(s) : Socci RR , Tachado SD , Aronstam RS , Reinach PS
Ref : J Ocul Pharmacol Ther , 12 :259 , 1996
Abstract : Muscarinic receptor subtypes in the bovine corneal epithelial cells (BCE) were characterized on the basis of their: 1) ligand binding properties, 2) linkage to Ca2+ and cAMP cell signaling pathways, and 3) gene transcripts. Receptor subtypes, m1 and m2, are indicated by competition experiments using subtype-selective muscarinic receptor ligands. [3H]N-methylscopolamine ([3H]-MS) binding was displaced with IC50s of: 1) 1 microM for the m1 antagonist, pirenzipine; 2) 51 microM for the competitive m2 antagonist, AFDX-116; 3) 100 microM for the competitive m3 antagonist, 4-DAMP. In fural2 loaded BCE, carbachol (0.001 - 100 microM) increased intracellular Ca2+ concentration ([Ca2+]i), and these responses were significantly suppressed if they were preincubated with either atropine (1 microM) or 1 microM pirenzipine. In the absence of extracellular Ca2+, these carbachol-induced increases in [Ca2+]i were depressed. A considerable fall occurred with the presence of extracellular Ca2+ and 1 microM verapamil, an L-type Ca2+ channel blocker. These responses suggest that carbachol increases Ca2+ influx through an L-type Ca2+ channel in the plasma membrane, in addition to mobilizing Ca2+ from an intracellular store. BCE also possessed muscarinic receptors which were negatively linked to cAMP production insofar as: 1) preincubation with 10 microM carbachol significantly suppressed the increases in cAMP accumulation induced by isoproterenol (1 - 25 microM); 2) this blunting effect of carbachol on cAMP production was eliminated when the BCE were preincubated with either 1 microM AFDX-116, or 100 ng/ml pertussis toxin. The results of probing for muscarinic receptor gene expression are partially consistent with the ligand binding and functional assays. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed the presence of m2 but not m1, m3 or m4 gene transcripts. In summary, we obtained pharmacological and functional evidence for m1 and m2 receptors in BCE. However, only the m2 gene transcript could be detected.
ESTHER : Socci_1996_J.Ocul.Pharmacol.Ther_12_259
PubMedSearch : Socci_1996_J.Ocul.Pharmacol.Ther_12_259
PubMedID: 8875332

Title : S-nitrosylation of m2 muscarinic receptor thiols disrupts receptor-G-protein coupling -
Author(s) : Aronstam RS , Martin DC , Dennison RL , Cooley HG
Ref : Annals of the New York Academy of Sciences , 757 :215 , 1995
PubMedID: 7611675

Title : A novel agonist binding site on nicotinic acetylcholine receptors - Pereira_1993_J.Recept.Res_13_413
Author(s) : Pereira EF , Alkondon M , Tano T , Castro NG , Froes-Ferrao MM , Rozental R , Aronstam RS , Schrattenholz A , Maelicke A , Albuquerque EX
Ref : J Recept Res , 13 :413 , 1993
Abstract : This report provides evidence that physostigmine (Phy) and benzoquinonium (BZQ) are able to activate nicotinic acetylcholine receptors (nAChRs) through binding site(s) distinct from those of the natural transmitter, ACh. Such findings are in agreement with a second pathway of activation of nAChRs. Receptor activation may be modulated through the novel site, and, consequently, physiological processes involving nicotinic synapses could be controlled. Using patch clamp techniques, single channel currents activated by ACh and anatoxin were recorded from frog interosseal muscle fibers under cell-attached condition and outside-out patches excised from cultured rat hippocampal neurons. Whole cell nicotinic currents were also studied in the cultured neurons. In most of the neurons, nicotinic responses were blocked by the nicotinic antagonists methyllycaconitine (MLA) and alpha-bungarotoxin (alpha-BGT). Evaluation of the effects of Phy and BZQ on the muscle and on the alpha-BGT- and MLA-sensitive neuronal nAChRs demonstrated that both compounds were open channel blockers at these receptors. Furthermore, at low micromolar concentrations, Phy and BZQ activated the nAChRs of all preparations tested, such an effect being unexpectedly resistant to alpha-BGT or MLA. Thus, the nAChRs could be activated via two distinct binding sites: one for ACh and the other for Phy and BZQ. These findings and previous biochemical results led us to suggest that a putative endogenous ligand could bind to the new site and thereby regulate the activation of nAChRs in nicotinic synapses.
ESTHER : Pereira_1993_J.Recept.Res_13_413
PubMedSearch : Pereira_1993_J.Recept.Res_13_413
PubMedID: 8450498

Title : Retinoic acid-induced differentiation of a human neuroblastoma cell line alters muscarinic receptor expression - Baumgartner_1993_Brain.Res.Dev.Brain.Res_72_305
Author(s) : Baumgartner MK , Wei J , Aronstam RS
Ref : Brain Research Developmental Brain Research , 72 :305 , 1993
Abstract : Muscarinic receptor density increased by approximately 36% after differentiation induced by retinoic acid (Bmax, control = 126 +/- 13 fmol/mg protein; Bmax, retinoic acid-treated = 170 +/- 17 fmol/mg protein; P < 0.05), corresponding to a 170% increase in receptor content per cell. The affinity of [3H]NMS for the receptors was somewhat lower in the retinoic acid-treated cells (Kd, control = 0.14 +/- 0.04 nM; Kd, retinoic acid-treated = 0.25 +/- 0.04 nM; P < 0.05). Reverse transcriptase/polymerase chain reaction analysis using subtype-specific primers revealed that undifferentiated Sk-N-SH cells transcribed mRNA for all 5 receptor subtypes; this pattern was not affected by retinoic acid treatment. [3H]NMS displacement curves with subtype selective receptor ligands (pirenzepine, m1; AFDX-116, m2; 4-DAMP, m3) indicated the predominant expression of m3 and m1 receptor subtypes, and differentiation did not affect the pharmacological profile of the expressed receptor populations. The present results indicate that differentiation induces selective changes in the expression and activity of muscarinic receptors in a neuronal cell line.
ESTHER : Baumgartner_1993_Brain.Res.Dev.Brain.Res_72_305
PubMedSearch : Baumgartner_1993_Brain.Res.Dev.Brain.Res_72_305
PubMedID: 8485852

Title : Role of prostanoids in the regulation of central cholinergic receptor sensitivity - Buccafusco_1993_J.Pharmacol.Exp.Ther_266_314
Author(s) : Buccafusco JJ , Lapp CA , Aronstam RS , Hays AC , Shuster LC
Ref : Journal of Pharmacology & Experimental Therapeutics , 266 :314 , 1993
Abstract : Tachyphylaxis develops to the hypertensive response to central (i.c.v.) injection of carbachol in conscious rats. This pressor response exhibits tachyphylaxis if the injection is repeated within 8 hr of the first injection. Blockade of brain prostaglandin synthesis with indomethacin does not inhibit the pressor response to carbachol in naive rats, but eliminates the pressor response to carbachol when the muscarinic agonist is repeated within a few hours of the first injection. If the time interval is extended to permit return of the full response (i.e., 24 hr later), indomethacin no longer inhibits the pressor response. The related cyclooygenase inhibitor meclofenamate produced effects which were identical to those of indomethacin, but at approximately 10-fold higher doses. When shorter acting drugs (duration of action < 30 min), physostigmine or arecoline, were used according to the same paradigm, indomethacin was less effective at inhibiting the pressor response to the second injection, even when the two agonist injections were spaced only 30 min apart. The ability of indomethacin to enhance central muscarinic receptor tachyphylaxis was also observed in carbachol-induced hypothermia. The density of diencephalic muscarinic receptors was estimated by using N-[3H]methylscopolamine as a probe. Carbachol-induced a down-regulation of muscarinic receptors, and indomethacin increased the extent of this down regulation. These findings suggest that prostaglandins play a role in the development of tachyphylaxis to brain muscarinic receptor stimulation: activation of prostaglandin synthesis may decelerate the development of desensitization to muscarinic agonists.
ESTHER : Buccafusco_1993_J.Pharmacol.Exp.Ther_266_314
PubMedSearch : Buccafusco_1993_J.Pharmacol.Exp.Ther_266_314
PubMedID: 8331566

Title : Pyrazole, an alcohol dehydrogenase inhibitor, has dual effects on N-methyl-D-aspartate receptors of hippocampal pyramidal cells: agonist and noncompetitive antagonist - Pereira_1992_J.Pharmacol.Exp.Ther_261_331
Author(s) : Pereira EF , Aracava Y , Aronstam RS , Barreiro EJ , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 261 :331 , 1992
Abstract : Electrophysiological and biochemical studies demonstrated that pyrazole, an inhibitor of alcohol dehydrogenase and a proposed therapeutic agent for treatment of alcoholic intoxication, activated and blocked the N-methyl-D-aspartate (NMDA) receptor and did not interact significantly with the end-plate nicotinic acetylcholine receptor (AChR). Pyrazole, at concentrations as low as 0.5 microM, applied to outside-out patches excised from the membrane of cultured rat hippocampal neurons, elicited single-channel currents of 48 pS which were blocked by DL-2-amino-5-phosphorovaleric acid, a competitive antagonist of NMDA. In addition, binding studies showed that pyrazole displaced 1-(cis-2-carboxypiperidine-4-yl)methyl-1-phosphoric acid from the agonist recognition site of the NMDA receptor in a concentration-dependent manner and enhanced the binding of (+)-5-methyl-10,11-dihydro-5H- dibenzo[a,d]cyclohepten-5,10-imine to this complex. These data indicate that pyrazole is an agonist at NMDA receptors. However, at higher concentrations, open and burst times as well as the frequency of single-channel currents activated by pyrazole were reduced significantly, a finding which suggests that this compound is also an open channel blocker. In agreement with these results, it was shown biochemically that pyrazole was able to stimulate influx of Ca++ into rat brain microsomes via NMDA receptors and on the other hand to block the influx of Ca++ induced by NMDA. Pyrazole was unable to affect the neuromuscular transmission of frog sartorius muscle-sciatic nerve preparations. Additionally, pyrazole did not interact either with the agonist recognition site or with noncompetitive sites of the AChR. However, this drug had a very weak agonist-like action on the AChR of the Torpedo electric organ, most likely via binding sites different from those described previously for acetylcholine. Therefore, the therapeutic efficacy of pyrazole may be related at least in part to its effects on the NMDA receptor. Furthermore, this compound, because of the small size and rigidity of its molecular structure, becomes a promising drug for the study of the NMDA receptor. Indeed its use may allow a better understanding of the physiological and pathological processes involving this receptor.
ESTHER : Pereira_1992_J.Pharmacol.Exp.Ther_261_331
PubMedSearch : Pereira_1992_J.Pharmacol.Exp.Ther_261_331
PubMedID: 1313873

Title : Muscarinic binding sites on bovine pulmonary arterial endothelial cells in culture - Aronstam_1992_Pharmacology_44_324
Author(s) : Aronstam RS , Ryan US , Catravas JD
Ref : Pharmacology , 44 :324 , 1992
Abstract : We have investigated the presence and nature of muscarinic binding sites on membranes from cultured bovine pulmonary arterial endothelial cells (BPAE). BPAE were harvested and subcultured nonenzymatically; experiments were performed 3-5 days postconfluence and between 10 and 25 passage numbers. Utilizing radioligand binding techniques with the muscarinic receptor antagonists [3H]3-quinuclidinyl benzilate ([3H]QNB) and [3H]N-methylscopolamine ([3H]MS) as probes, we identified a small population of atropine-sensitive muscarinic sites (1,800-2,000 sites/cell or 7-8 fmol/mg protein). Muscarinic binding sites on BPAE membranes resembled classical muscarinic receptors in that (a) the binding of 2 nM [3H]QNB was inhibited by muscarinic agonists and antagonists, (b) [3H]QNB binding was 30 times more sensitive to R(-)- than to S(+)-QNB, (c) binding of the muscarinic receptor agonist carbamylcholine involved high and low affinity components, (d) the stable GTP analog, Gpp(NH)p (100 microM) shifted agonist binding curves to the right by a factor of three, and (e) the high affinity binding of the agonist [3H]oxotremorine-M to muscarinic receptors was depressed by Gpp(NH)p. On the other hand, gallamine, which allosterically regulates muscarinic receptor binding in other tissues, did not affect the rates of dissociation of [3H]QNB, [3H]MS or [3H]oxotremorine-M from BPAE binding sites. We concluded that BPAE in culture exhibit muscarinic binding sites which possess many but not all of the properties associated with classical muscarinic receptors.
ESTHER : Aronstam_1992_Pharmacology_44_324
PubMedSearch : Aronstam_1992_Pharmacology_44_324
PubMedID: 1508962

Title : Nicotinic receptor-elicited sodium flux in rat pheochromocytoma PC12 cells: effects of agonists, antagonists, and noncompetitive blockers - Daly_1991_Neurochem.Res_16_489
Author(s) : Daly JW , Nishizawa Y , Edwards MW , Waters JA , Aronstam RS
Ref : Neurochem Res , 16 :489 , 1991
Abstract : Nicotinic agonists stimulate 22Na flux in rat pheochromocytoma PC12 cells. The stimulatory effect of carbamylcholine is maximal at 1 mM, while the stimulatory effect of nicotine and anatoxin maximize at the same level at 100 microM and 10 microM, respectively. The tertiary amines arecolone and isoarecolone have no effect on flux at 100 microM, while the methiodides at 100 microM stimulate flux to an extent similar to 1 mM carbamylcholine. Dihydro and alcohol analogues of isoarecolone methiodide have markedly smaller effects on flux. A preincubation for 2 to 20 min with carbamylcholine (2 mM), nicotine (300 microM), anatoxin (30 microM) or isoarecolone methiodide (100 microM) causes marked desensitization to a subsequent carbamylcholine-elicited stimulation of flux. d-Tubocurarine, mecamylamine, hexamethonium, and chlorisondamine inhibit carbamylcholine-elicited flux with IC50 values of 1.0, 0.8, 43, and 0.020 microM, respectively. Atropine has no effect at 1 microM, but reduces the response to carbamylcholine by 50% at 8.6 microM, presumably as a noncompetitive blocker. Other noncompetitive blockers of nicotinic acetylcholine-receptors, such as histrionicotoxins, gephyrotoxin, pumiliotoxin C, phencyclidine, bupivacaine and piperocaine, inhibit carbamylcholine-elicited stimulation of 22Na flux with IC50 values from 0.3 to 1.8 microM. In contrast to d-tubocurarine, which inhibits carbamylcholine-elicited desensitization, and mecamylamine, which has no apparent effect on desensitization, chlorisondamine and certain noncompetitive blockers appear to enhance desensitization. The effects of agonists, antagonists and noncompetitive blockers at the neuronal nicotinic acetylcholine receptor-channel of PC12 cells are compared to their effects on binding of [125I]alpha-bungarotoxin to agonist-recognition sites and of [3H]perhydrohistrionicotoxin to noncompetitive blocker sites of the nicotinic acetylcholine receptor-channel of electric ray (Torpedo) electroplax membranes. There are marked differences in relative potencies for the two types of nicotinic acetylcholine receptor-channel.
ESTHER : Daly_1991_Neurochem.Res_16_489
PubMedSearch : Daly_1991_Neurochem.Res_16_489
PubMedID: 1922660

Title : Decahydroquinoline alkaloids: noncompetitive blockers for nicotinic acetylcholine receptor-channels in pheochromocytoma cells and Torpedo electroplax - Daly_1991_Neurochem.Res_16_1207
Author(s) : Daly JW , Nishizawa Y , Padgett WL , Tokuyama T , McCloskey PJ , Waykole L , Schultz AG , Aronstam RS
Ref : Neurochem Res , 16 :1207 , 1991
Abstract : In pheochromocytoma PC12 cells, (+)-cis-decahydroquinoline 195A (5-methyl-2-propyl-cis-decahydroquinoline) and (+)-perhydro-cis-decahydroquinoline 219A (2,5-dipropyl-cis-decahydroquinoline) inhibit carbamylcholine-elicited sodium flux with IC50 values of 1.0 and 1.5 microM, respectively. Both of these decahydroquinolines appear to enhance desensitization, although apparent lack of complete removal of (+)-perhydro-cis-219A by washing complicates interpretation of the effects of that agent. A series of cis- and trans-decahydroquinolines with substituents in the 2- and 5-position also exhibit structure-dependent inhibition of carbamylcholine-elicited sodium flux in PC12 cells and all of the decahydroquinolines inhibit binding of the noncompetitive blocking agent [3H]perhydrohistrionicotoxin to muscle-type nicotinic acetylcholine receptor-channels in membranes from Torpedo electroplax. The Ki values in electroplax membranes range from 1.4 to 7.9 microM, making these alkaloids comparable in potencies to the histrionicotoxins. Potencies are increased 2- to 3-fold in the presence of an agonist, carbamylcholine. The profile of activities are similar in PC12 cells and electroplax membranes. The cis- and trans-decahydroquinolines represent another class of noncompetitive blockers for acetylcholine receptor-channels with similar activity for both muscle-type and ganglionic type nicotinic receptors.
ESTHER : Daly_1991_Neurochem.Res_16_1207
PubMedSearch : Daly_1991_Neurochem.Res_16_1207
PubMedID: 1815136

Title : 5,8-disubstituted indolizidines: a new class of noncompetitive blockers for nicotinic receptor-channels - Daly_1991_Neurochem.Res_16_1213
Author(s) : Daly JW , Nishizawa Y , Padgett WL , Tokuyama T , Smith AL , Holmes AB , Kibayashi C , Aronstam RS
Ref : Neurochem Res , 16 :1213 , 1991
Abstract : A series of 8-methyl-5-substituted indolizidines inhibit binding of the noncompetitive blocking agent [3H]perhydrohistrionicotoxin to muscle-type nicotinic acetylcholine receptor-channels in membranes from Torpedo electroplax. The Ki values range from 0.16 to 1.12 microM, making these alkaloids among the most potent ligands for this site. Unlike most noncompetitive blockers, the potencies of the 8-methyl-5-substituted indolizidines are reduced in the presence of carbamylcholine. Indolizidine 205A (8-methyl-5-(4-pentynyl)indolizidine) is unique in enhancing binding of [3H]perhydrohistrionicotoxin by 1.5-fold. The enhancement is at a maximum at 0.01 to 0.1 microM, followed by progressive inhibition with an IC50 of about 20 microM. In the presence of carbamylcholine, which itself enhances binding of [3H]perhydrohistrionicotoxin, indolizidine 205A causes only an inhibition of binding with an IC50 of about 10 microM. Indolizidines with a hydroxy substituent on the 8-methyl group have very low activity. None of the indolizidines affect binding of [125I]alpha-bungarotoxin to acetylcholine recognition sites. In pheochromocytoma PC12 cells, indolizidine 205A has no agonist activity, but only inhibits carbamylcholine-elicited 22Na+ influx. The profile of potencies for the 8-methyl-5-substituted indolizidines is similar in electroplax membranes and PC12 cells. Indolizidines 205A and 209B (8-methyl-5-pentylindolizidine) have no apparent effect on desensitization of receptors in PC12 cells. The 5,8-disubstituted indolizidines appear to represent an atypical and potent class of noncompetitive blockers for muscle-type and ganglionic nicotinic receptor-channels.
ESTHER : Daly_1991_Neurochem.Res_16_1213
PubMedSearch : Daly_1991_Neurochem.Res_16_1213
PubMedID: 1815137

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 : Acetylcholine stimulates GTP binding to G proteins in rat striatum - Ravindra_1991_Neuroreport_2_127
Author(s) : Ravindra R , Aronstam RS
Ref : Neuroreport , 2 :127 , 1991
Abstract : To evaluate the role of G proteins in acetylcholine (ACh)-mediated signal transduction, a novel assay to measure [35S]GTP gamma S binding to rat striatal membranes was standardized. ACh and carbamylcholine (CCh) stimulated the high affinity [35S]GTP gamma S binding to striatal membranes by up to 45% in a concentration-dependent manner. A time course study revealed that 0.1 microM CCh stimulated [35S]GTP gamma S binding by 26% at 4 min and 20% at 15 min. CCh-stimulated [35S]GTP gamma S binding was inhibited by atropine, N-methylscopolamine and pirenzepine. Oxotremorine (1 microM), a partial muscarinic agonist, maximally stimulated the binding to striatal membranes by only 25%.
ESTHER : Ravindra_1991_Neuroreport_2_127
PubMedSearch : Ravindra_1991_Neuroreport_2_127
PubMedID: 1768854

Title : Colchicine inhibits acetylcholine receptor stimulation of G protein GTPase activity in rat striatum - Ravindra_1991_Pharmacol.Toxicol_69_259
Author(s) : Ravindra R , Aronstam RS
Ref : Pharmacol Toxicol , 69 :259 , 1991
Abstract : Colchicine, which is known to influence tubulin function, was used to delineate a possible role of tubulin in signal transduction in the rat striatal membranes. Low Km GTPase activity (EC 3.6.1.-) was assayed in 10 micrograms membrane protein using [gamma-32P]GTP at 37 degrees in an ATP-regenerating buffer containing 1 microM unlabeled GTP. At 10 and 100 microM, colchicine inhibited the GTPase activity by 18% and 40%, respectively. Colchicine (100 microM) inhibited the enzymatic activity by 30-40% at all the time points the reaction was monitored. Acetylcholine (ACh) stimulated the low Km GTPase activity in a concentration-dependent manner, by up to 57%. ACh-stimulated activity was accepted as reflecting GTP hydrolysis catalyzed by receptor-coupled transducer G proteins. In the presence of 100 microM colchicine, the ability of ACh to stimulate G protein GTPase activity was inhibited. For example, at 10 microM ACh the enzyme activity was stimulated up to 52%; in the presence of 100 microM colchicine, 10 microM ACh stimulated the activity by only up to 33%. These results suggest that colchicine disrupts ACh receptor-G protein coupling as a result of its interaction with tubulin or G protein(s) or both.
ESTHER : Ravindra_1991_Pharmacol.Toxicol_69_259
PubMedSearch : Ravindra_1991_Pharmacol.Toxicol_69_259
PubMedID: 1956877

Title : Selective blockade of NMDA-activated channel currents may be implicated in learning deficits caused by lead - Alkondon_1990_FEBS.Lett_261_124
Author(s) : Alkondon M , Costa AC , Radhakrishnan V , Aronstam RS , Albuquerque EX
Ref : FEBS Letters , 261 :124 , 1990
Abstract : The effect of Pb2+ on glutamate receptor activity in rat hippocampal neurons was investigated with a view of explaining the cognitive and learning deficits produced by this heavy metal. Pb2+ (2.5-50 microM) selectively inhibited N-methyl-D-aspartate (NMDA)-induced whole-cell and single-channel currents in a concentration-dependent but voltage-independent manner, without significantly altering currents induced by either quisqualate or kainate. The frequency of NMDA-induced channel activation was decreased by Pb2+. Neither glycine (10-100 microM), nor Ca2+ (10 mM) reversed the effect of Pb2+. Pb2+ also inhibited the [3H]MK-801 binding to rat hippocampal membranes in vitro. The elucidation of the actions of Pb2+ on the NMDA receptor ion channel complex provides important insights into the clinical and toxic effects of this cation.
ESTHER : Alkondon_1990_FEBS.Lett_261_124
PubMedSearch : Alkondon_1990_FEBS.Lett_261_124
PubMedID: 1689669

Title : Activation and blockade of the acetylcholine receptor-ion channel by the agonists (+)-anatoxin-a, the N-methyl derivative and the enantiomer - Kofuji_1990_J.Pharmacol.Exp.Ther_252_517
Author(s) : Kofuji P , Aracava Y , Swanson KL , Aronstam RS , Rapoport H , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 252 :517 , 1990
Abstract : The effects of (+)- and (-)-anatoxin-a (AnTX) and N-methylanatoxin (M-AnTX) on peripheral nicotinic ion channel activity were studied using high micromolar concentrations. Whereas (+)-AnTX is an effective agonist at nanomolar concentrations, (-)-AnTX and M-AnTX were effective at low micromolar concentrations. The binding of [3H]perhydrohistrionicotoxin to the nicotinic acetylcholine receptor-ion channel was stimulated by the above agonist concentrations, but [3H]perhydrohistrionicotoxin binding was inhibited at high micromolar concentrations of each of the toxins. In single channel recordings, these toxins exhibited ion channel blocking properties; the concentration- and voltage-dependent kinetics of each were essentially the same. In the case of (+)-AnTX, desensitization was also present at micromolar concentrations. These data show that ion channel blockade may be a property of many anatoxin-a analogs, and that in the particular case of analogs with low agonist potency, ion channel blockade may be a concomitant primary effect of the toxins. Stereospecificity and number of amine moieties did not influence the ion channel blocking characteristics in this series of molecules, although these factors strongly modified agonist potency.
ESTHER : Kofuji_1990_J.Pharmacol.Exp.Ther_252_517
PubMedSearch : Kofuji_1990_J.Pharmacol.Exp.Ther_252_517
PubMedID: 1690292

Title : Molecular effects of dimethylanatoxin on the peripheral nicotinic acetylcholine receptor - Costa_1990_J.Pharmacol.Exp.Ther_252_507
Author(s) : Costa AC , Swanson KL , Aracava Y , Aronstam RS , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 252 :507 , 1990
Abstract : N,N-dimethylanatoxin (DMAnTX), the quaternary derivative of the potent nicotinic agonist (+)-anatoxin-a (AnTX), has been evaluated for potency and efficacy at nicotinic acetylcholine receptors of frog motor endplates and Torpedo electric organs. DMAnTX was only weakly effective in eliciting contracture of the frog rectus abdominis and was orders of magnitude less potent than AnTX. Biochemical assay showed that DMAnTX was a weak inhibitor of [125I]alpha-bungarotoxin binding to the receptors in frog muscle and Torpedo electroplaque membranes: the IC50 values were 60 and 14 microM, respectively. A low frequency of single channel currents recorded from isolated interosseal fibers at concentrations from 20 to 100 microM of DMAnTX and the stimulation of [3H]perhydrohistrionicotoxin [( 3H]H12-HTX) binding (half-maximal at 0.3 microM) confirmed the weak activation of the receptor. DMAnTX also exhibited antagonist effects. In muscle twitch assays, 100 microM of DMAnTX effectively decreased the tension induced by nerve stimulation, although DMAnTX did not affect muscle membrane action potentials. The binding of [3H] perhydrohistrionicotoxin was also inhibited at high micromolar concentrations of DMAnTX. Combination of DMAnTX with acetylcholine in single channel current experiments demonstrated that DMAnTX possesses ion channel blocking properties, which become apparent at low micromolar concentrations, and DMAnTX enhances the desensitization induced by acetylcholine above 10 microM AnTX. The difference in agonist potency between AnTX and DMAnTX may be attributed to a change in conformation of the molecular skeleton induced by the N-methyl groups.
ESTHER : Costa_1990_J.Pharmacol.Exp.Ther_252_507
PubMedSearch : Costa_1990_J.Pharmacol.Exp.Ther_252_507
PubMedID: 1690291

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 : Increased muscarinic responsiveness and decreased muscarinic receptor content in ileal smooth muscle in diabetes - Carrier_1990_J.Pharmacol.Exp.Ther_254_445
Author(s) : Carrier GO , Aronstam RS
Ref : Journal of Pharmacology & Experimental Therapeutics , 254 :445 , 1990
Abstract : Longitudinal muscles from the ileum of rats rendered diabetic for 10 to 12 weeks by injection of streptozotocin (STZ) developed greater contractile force in response to muscarinic agonists, (acetylcholine, carbamylcholine and bethanechol) than muscles from age-matched control rats. There was no change, however, in the sensitivity of longitudinal smooth muscles to muscarinic agonists as reflected by the EC50 values for stimulation of muscle contraction. This increased responsiveness of the muscles was accompanied by a 32% reduction in the density of muscarinic receptors (381 +/- 93 vs. 560 +/- 74 fmol/mg of membrane protein) in muscles from STZ-diabetic rats. There was no change in agonist or antagonist binding affinities in muscles from diabetic rats, and there was no alteration in the distribution of receptors between the states characterized by high and low affinity agonist binding. There was also no change in acetylcholinesterase activity in muscle membranes isolated from STZ-diabetic rats. The origin of the increased responsiveness of ileal smooth muscles in this model of diabetes is not clear, but may involve changes in the muscarinic signal transduction pathway beyond the receptor level, or in the contractile apparatus itself.
ESTHER : Carrier_1990_J.Pharmacol.Exp.Ther_254_445
PubMedSearch : Carrier_1990_J.Pharmacol.Exp.Ther_254_445
PubMedID: 2384882

Title : Influence of volatile anesthetics on muscarinic regulation of adenylate cyclase activity -
Author(s) : Anthony BL , Dennison RL , Aronstam RS
Ref : Biochemical Pharmacology , 40 :376 , 1990
PubMedID: 2375772

Title : Effects of forskolin and analogues on nicotinic receptor-mediated sodium flux, voltage-dependent calcium flux, and voltage-dependent rubidium efflux in pheochromocytoma PC12 cells - Nishizawa_1990_Cell.Mol.Neurobiol_10_351
Author(s) : Nishizawa Y , Seamon KB , Daly JW , Aronstam RS
Ref : Cellular Molecular Neurobiology , 10 :351 , 1990
Abstract : 1. Forskolin, a naturally occurring diterpene that activates adenylate cyclase, HL706, a water-soluble derivative of forskolin (6 beta-[(piperidino)acetoxy]-7-desacetylforskolin) that is less potent than forskolin in activating adenylate cyclase, and 1,9-dideoxyforskolin, an analogue that does not activate adenylate cyclase, were examined for effects on the nicotinic receptor-mediated 22Na+ flux, a high potassium-induced 45Ca2+ flux through L-type calcium channels, and a high potassium-induced 86Rb+ efflux through a calcium-dependent potassium channels in PC12 cells. 2. Forskolin and analogues at 30 microM completely blocked carbamylcholine-elicited flux of 22Na+ through the nicotinic receptor-gated channel. 1,9-Dideoxyforskolin had an IC50 value of 1.6 microM with forskolin and HL706 being two- to three fold less potent. 3. Forskolin and its analogues appear to be noncompetitive blockers of the neuronal nicotinic receptor-channel complex in PC12 cells, but unlike many noncompetitive blockers, did not markedly enhance desensitization. Instead, forskolin, but not HL706 or 1,9-dideoxyforskolin, slightly antagonized the desensitization evoked by high concentrations of carbamylcholine. N-Ethylcarboxamidoadenosine, an adenosine analogue that elevates cyclic AMP and 8-bromo-cyclic AMP had no effect on desensitization. 4. Forskolin, HL706, and 1,9-dideoxyforskolin in the presence of carbamylcholine inhibited the binding of a noncompetitive blocker, [3H]perhydrohistrionicotoxin, to the muscle-type nicotinic receptor-channel complex in Torpedo electroplax membranes with IC50 values of 20 microM. Forskolin had no effect on [3H]perhydrohistrionicotoxin binding in the absence of carbamylcholine, while HL706 and 1,9-dideoxyforskolin still inhibited binding in the absence of carbamylcholine. 5. Forskolin, but not HL706 or 1,9-dideoxyforskolin had a slight inhibitory effect on the binding of [125I]alpha-bungarotoxin to acetylcholine recognition sites in Torpedo membranes. 1,9-Dideoxyforskolin at 30 microM, but not forskolin or HL706, markedly inhibited depolarization-evoked 45Ca+ flux and 86Rb+ efflux in PC12 cells, suggesting that 1,9-dideoxyforskolin has nonspecific inhibitory effects on a variety of ion channels.
ESTHER : Nishizawa_1990_Cell.Mol.Neurobiol_10_351
PubMedSearch : Nishizawa_1990_Cell.Mol.Neurobiol_10_351
PubMedID: 1701359

Title : Influence of anti-tubulin antibodies on muscarinic receptor modulation of G protein GTPase activity in rat striatum - Ravindra_1990_Biochem.Pharmacol_40_457
Author(s) : Ravindra R , Aronstam RS
Ref : Biochemical Pharmacology , 40 :457 , 1990
Abstract : To understand the role of tubulin, an integral component of neural membranes, in signal transduction processes, the influence of anti-tubulin antibodies on the low Km GTPase activity associated with transducer G proteins was examined in rat striatum. Membranes were prepared from striatum by conventional procedures, and the low Km GTPase activity (EC 3.6.1.-) was determined using [gamma-32P]GTP at 37 degrees in an ATP-regenerating buffer containing 0.2 to 2.0 microM unlabeled GTP. GTPase activity was linear for up to 30 min and was directly proportional to protein concentration. Polyclonal anti-tubulin antibodies, anti-alpha-tubulin antibodies, and anti-beta-tubulin antibodies (10 micrograms) stimulated G protein GTPase activity. Anti-beta-tubulin antibody (10 micrograms) stimulated GTPase activity by about 60% at each time point, while 10 micrograms of either anti-alpha-tubulin or polyclonal anti-tubulin antibodies stimulated GTPase activity by only 20-30% at each time point. The Vmax/Km ratio, an index of the enzyme-substrate interaction, increased by only 26% with the anti-alpha-tubulin antibody and by 52% with anti-beta-tubulin antibody; polyclonal anti-tubulin antibodies did not affect this ratio. GTPase activity was stimulated by acetylcholine in an atropine-sensitive manner. At 100 microM, acetylcholine stimulated GTPase activity by about 50%. Polyclonal anti-tubulin, anti-alpha-tubulin, or anti-beta-tubulin antibodies (10 micrograms) potentiated acetylcholine stimulation of GTPase activity. Two possible mechanisms by which anti-tubulin antibodies could stimulate low Km GTPase activity and potentiate the stimulatory effects of acetylcholine are: (1) by inhibiting GTP binding to beta-tubulin, and (2) by eliminating a chronic inhibitory effect of tubulin on G protein or receptor-G protein interaction.
ESTHER : Ravindra_1990_Biochem.Pharmacol_40_457
PubMedSearch : Ravindra_1990_Biochem.Pharmacol_40_457
PubMedID: 2166514

Title : N-methylanatoxinol isomers: derivatives of the agonist (+)-anatoxin-a block the nicotinic acetylcholine receptor ion channel - Swanson_1989_Mol.Pharmacol_35_223
Author(s) : Swanson KL , Aracava Y , Sardina FJ , Rapoport H , Aronstam RS , Albuquerque EX
Ref : Molecular Pharmacology , 35 :223 , 1989
Abstract : Using biochemical and patch-clamp techniques, we investigated the pharmacology of S- and R-epimers of N-methylanatoxinol, which are analogs of the semi-rigid, stereoselective, nicotinic agonist (+)-anatoxin-a. In contrast to (+)-anatoxin-a, both isomers had poor ability to inhibit the binding of 125I-alpha-bungarotoxin or to open acetylcholine channels, and they were unable to elicit contracture of frog rectus abdominis muscles. However, both isomers were able to demonstrate significant concentration-dependent blockade of the nicotinic acetylcholine receptor ion channel. The R-isomer was approximately 4-fold more potent in causing inhibition of [3H]H12HTX binding than was the S-isomer, in the absence of carbamylcholine. In the presence of carbamylcholine, the affinity of the R-isomer of N-methylanatoxinol for the ion channel sites was further enhanced, so that its affinity became much greater than that of the S-isomer. Refinement of voltage- and concentration-dependent terms for the ion channel blocking and unblocking rates yielded functions that were able to predict the channel open times and short closed times well. The S-isomer bound and dissociated from the ion channel site of the nicotinic acetylcholine receptor more rapidly and with greater voltage sensitivity than the R-isomer. The present characterization of the antagonistic properties of these new analogs of (+)-anatoxin-a introduces a new aspect to the molecular pharmacology of (+)-anatoxin-a analogs; the semi-rigid compounds could be useful in describing the allosteric binding sites of the acetylcholine receptor, as well as in delimiting the agonist binding site.
ESTHER : Swanson_1989_Mol.Pharmacol_35_223
PubMedSearch : Swanson_1989_Mol.Pharmacol_35_223
PubMedID: 2465486

Title : Protection afforded by clonidine from the acute and chronic behavioral toxicity produced by the cholinesterase inhibitor soman - Buccafusco_1989_Neurotoxicol.Teratol_11_39
Author(s) : Buccafusco JJ , Graham JH , VanLingen J , Aronstam RS
Ref : Neurotoxicology & Teratology , 11 :39 , 1989
Abstract : Studies in our laboratory have demonstrated that alpha 2-adrenergic agonists such as clonidine inhibit acetylcholine synthesis and release. The effect of clonidine is so marked, that pretreatment reduces both the accumulation of brain acetylcholine, and the toxicity caused by cholinesterase inhibitors such as soman. In this study rats were pretreated with regimens which included known protective doses of clonidine or atropine. The dose of soman was adjusted to allow for the 24 hr survival of 50% of the animals in each group. Behavioral toxicity was assessed by observational techniques and by employing an automated animal activity monitor. Clonidine pretreatment resulted in a significant degree of protection from the lethal effects of soman in the rat. In addition, the ability of soman to inhibit normal ongoing behavior as well to elicit the expression of several abnormal behaviors, including convulsive behavior were inhibited by clonidine pretreatment. While clonidine pretreatment resulted in a similar degree of protection as atropine pretreatment against the acute phase of soman toxicity, only clonidine was effective in preventing the expression of chronic behavioral toxic manifestations to soman. Thus, animals pretreated with clonidine exhibited a significantly improved prognosis 24 or 48 hr following soman injection as compared with saline- or atropine-pretreated animals.
ESTHER : Buccafusco_1989_Neurotoxicol.Teratol_11_39
PubMedSearch : Buccafusco_1989_Neurotoxicol.Teratol_11_39
PubMedID: 2725439

Title : Disruption of muscarinic receptor-G protein coupling is a general property of liquid volatile anesthetics - Anthony_1989_Neurosci.Lett_99_191
Author(s) : Anthony BL , Dennison RL , Aronstam RS
Ref : Neuroscience Letters , 99 :191 , 1989
Abstract : The influences of 3 volatile anesthetics, chloroform, enflurane and isoflurane, on muscarinic acetylcholine receptors in rat brainstem were determined. Each of the volatile anesthetics increased [3H]methylscopolamine [( 3H]MS) binding affinity, but did not affect the number of [3H]MS binding sites. Carbamylcholine affinity for brainstem muscarinic receptors was not altered after equilibration of brainstem membranes with any of these anesthetics. The ability of guanine nucleotides to depress the high affinity binding of two agonists, carbamylcholine and [3H]oxotremorine-M, was decreased or eliminated after equilibration of brainstem membranes with any of the anesthetics. In each of these actions, these anesthetics resemble halothane and diethyl ether. These results indicate that interference with muscarinic receptor-G protein interactions is a common property of liquid volatile anesthetics and may represent a general mechanism for the disruption of signal transmission between cells during anesthesia.
ESTHER : Anthony_1989_Neurosci.Lett_99_191
PubMedSearch : Anthony_1989_Neurosci.Lett_99_191
PubMedID: 2501717

Title : Anesthetic effects on muscarinic signal transduction - Aronstam_1989_Int.Anesthesiol.Clin_27_265
Author(s) : Aronstam RS , Dennison RL, Jr.
Ref : Int Anesthesiol Clin , 27 :265 , 1989
Abstract : A wealth of pharmacological and physiological evidence has established that anesthetics disrupt synaptic transmission at muscarinic and other synapses. The sequence of molecular events precipitated by agonist binding to the receptors is under intense scrutiny. It appears that at the majority of synapses G proteins serve to mediate the transfer of information from receptors to intracellular mechanisms. The major exception to this scheme is the situation in which an ion channel is incorporated directly in the receptor structure. Binding of an agonist to these receptors produces a conformational change in the receptors which opens an intrinsic ion channel. This situation occurs in nicotinic acetylcholine gamma-amino butyric acid type A (GABAA, and 5-hydroxytryptamine type 3 (5-HT3) receptors). Assays have been developed to evaluate several steps in the cascade of events involved in synaptic signal transduction, and these assays have been employed to determine the step at which anesthetics act to disrupt synaptic transmission. We have demonstrated that several volatile anesthetics alter the interaction of muscarinic receptors with transducer G proteins. Ligand-binding experiments suggest that receptor-G protein complexes are stabilized, thereby disrupting G protein GTPase activity and muscarinic control of cellular activity. This "stabilization" does not appear to involve an inhibition of guanine nucleotide binding, the proximal event in receptor-G protein dissociation. Two possibilities warrant further consideration: (1) that GDP release from inactive G protein trimers, which is normally catalyzed by the receptor, is inhibited, and (2) that receptor-G protein complexes fail to dissociate even in response to GTP binding. We are currently examining these possibilities using purified G proteins and receptors in reconstituted systems.
ESTHER : Aronstam_1989_Int.Anesthesiol.Clin_27_265
PubMedSearch : Aronstam_1989_Int.Anesthesiol.Clin_27_265
PubMedID: 2512258

Title : Binding of semirigid nicotinic agonists to nicotinic and muscarinic receptors - Spivak_1989_Mol.Pharmacol_36_177
Author(s) : Spivak CE , Waters JA , Aronstam RS
Ref : Molecular Pharmacology , 36 :177 , 1989
Abstract : Isoarecolone methiodide (1-methyl-4-acetyl-1,2,3,6-tetrahydropyridine methiodide) was previously shown to be among the most potent agonists tested at the frog neuromuscular junction. Because nicotinic receptors from different sources vary in their selectivities, isoarecolone methiodide as well as 19 additional congeners, most of which were also previously tested at the frog neuromuscular junction, were studied in binding assays. Torpedo nobiliana was the tissue source for nicotinic receptors. Two types of experiments were conducted. The first evaluated the affinities of the agonists (including acetylcholine and carbamylcholine) for the recognition site by allowing the agonists to compete for that site with 125I-alpha-bungarotoxin. The inhibition potencies obtained correlated strongly (Spearman's correlation coefficient,-0.91) with the potency obtained at the frog neuromuscular junction. The second type of experiment evaluated the agonists for their ability to activate the receptor. The binding of [3H]perhydrohistrionicotoxin, which was employed as an indicator of the activation of the receptor, was measured in the presence of each of the agonists. Isoarecolone methiodide was the most potent of all. A few of the agonists (partial agonists) were incapable of fully enhancing this binding. For the full agonists, the concentration that produced half of the maximum binding of [3H]perhydrohistrionicotoxin was defined as the EC50. The correlation coefficient (Spearman's) for EC50 versus potency at the frog neuromuscular junction was -0.73, indicating innate differences between Torpedo and frog receptors. In addition, these compounds were tested for their affinity at muscarinic receptors from rat brain. Competition experiments were carried out using [3H]N-methylscopolamine. The affinity of isoarecolone methiodide was only about 7-fold lower than that of acetylcholine and less than 2-fold lower than that of carbamylcholine. In contrast, 1-methyl-4-acetylpiperazine methiodide was much more selective for nicotinic receptors. Its activity was similar to isoarecolone methiodide at the nicotinic receptor, but it was among the weakest compounds in its affinity for the muscarinic receptor.
ESTHER : Spivak_1989_Mol.Pharmacol_36_177
PubMedSearch : Spivak_1989_Mol.Pharmacol_36_177
PubMedID: 2747625

Title : Insulin prevention of altered muscarinic receptor-G protein coupling in diabetic rat atria - Aronstam_1989_Diabetes_38_1611
Author(s) : Aronstam RS , Carrier GO
Ref : Diabetes , 38 :1611 , 1989
Abstract : Right atria from rats rendered diabetic by injection of streptozocin (STZ-D) for 8-10 wk are supersensitive to the negative chronotropic effects of muscarinic agonists but have decreased levels of muscarinic receptors and acetylcholinesterase activity. Insulin treatment completely prevents the development of these changes. The proportion of atrial muscarinic receptors displaying high-affinity agonist binding is lower in STZ-D rats; however, the sensitivity of high-affinity agonist binding to regulation by a guanine nucleotide (5'-guanylylimidodiphosphate) is greater in atria from diabetic rats. Again, insulin treatment eliminates these differences. These findings indicate that alterations in atrial muscarinic systems in STZ-D rats are a consequence of the elaboration of the diabetic state and suggest that an alteration of functional muscarinic receptor-G protein coupling contributes to the altered physiological responsiveness of the heart in diabetes.
ESTHER : Aronstam_1989_Diabetes_38_1611
PubMedSearch : Aronstam_1989_Diabetes_38_1611
PubMedID: 2573555

Title : Precursors of cholinergic false transmitters: central effects on blood pressure and direct interactions with cholinergic receptors - Buccafusco_1988_Neuropharmacol_27_227
Author(s) : Buccafusco JJ , Aronstam RS
Ref : Neuropharmacology , 27 :227 , 1988
Abstract : The purpose of this study was to determine whether a series of analogs of choline depress central cholinergic function in a manner consistent with the activity of their acetylated derivatives. Intracerebroventricular infusion of monoethylcholine (MECh), diethylcholine (DECh) and triethylcholine (TECh) inhibited the pressor response of unanesthetized rats to the subsequent intravenous injection of physostigmine (a well-characterized muscarinic response). The order of blocking potency was TECh greater than DECh greater than MECh greater than choline, directly opposite to the order of potency for elicitation of a central pressor response by their associated acetylated derivative (i.e. ACh greater than AMECh greater than ADECh = ATECh; Aronstam, Marshall and Buccafusco, 1988). In contrast, there was little selectivity between the analogs of choline in terms of their affinity for muscarinic receptors in the brainstem or cortex; the Ki's for inhibition of the binding of [3H]quinuclidinyl benzilate ranged from 0.33 to 0.95 mM). In terms of their affinity for nicotinic receptors (from the electric organ of Torpedo californica) the following order of potency was obtained: choline greater than MECh = DECh greater than TECh. Choline and MEC stimulated the binding of [3H]phencyclidine to the nicotinic ion channel (EC50's = 79 and 115 microM, respectively). At greater concentrations, all of the analogs inhibited ligand binding to the channel (Ki's from 0.2 to 10 mM), with the following order of potency: TECh greater than DECh greater than MECh greater than choline. These findings suggest that the inhibitory actions of these analogs of choline are related to their synthesis and release as false cholinergic neurotransmitters.
ESTHER : Buccafusco_1988_Neuropharmacol_27_227
PubMedSearch : Buccafusco_1988_Neuropharmacol_27_227
PubMedID: 2836749

Title : Role of central biogenic amines on the protection afforded by clonidine against the toxicity of soman, an irreversible cholinesterase inhibitor - Buccafusco_1988_Toxicol.Lett_42_291
Author(s) : Buccafusco JJ , Aronstam RS , Graham JH
Ref : Toxicol Lett , 42 :291 , 1988
Abstract : Studies in our laboratory have demonstrated that alpha 2-adrenergic agonists such as clonidine offer protection against the toxicity caused by cholinesterase inhibitors such as soman. Experiments were designed to determine whether central catecholaminergic systems are implicated in the toxic and lethal manifestations of soman toxicity and whether the protection afforded by clonidine involves such pathways. Clonidine pretreatment resulted in a significant degree of protection from the lethal effects of soman in the mouse. Depletion of brain catecholamines did not alter soman-induced lethality or behavioral toxicity. Furthermore, catecholamine depletion was not effective in blocking clonidine-induced protection against soman toxicity. In contrast, elevation of brain catecholamines using the monoamine oxidase inhibitor, pargyline, resulted in significant protection against soman toxicity which was additive with that of clonidine.
ESTHER : Buccafusco_1988_Toxicol.Lett_42_291
PubMedSearch : Buccafusco_1988_Toxicol.Lett_42_291
PubMedID: 3176058

Title : Behavioral effects of toxic doses of soman, an organophosphate cholinesterase inhibitor, in the rat: protection afforded by clonidine - Buccafusco_1988_Pharmacol.Biochem.Behav_29_309
Author(s) : Buccafusco JJ , Graham JH , Aronstam RS
Ref : Pharmacol Biochem Behav , 29 :309 , 1988
Abstract : Atropine, a postsynaptic muscarinic antagonist, and clonidine, a presynaptic inhibitor of acetylcholine release, protect mice from the lethal effects of soman, a potent and irreversible cholinesterase inhibitor. The purpose of this study was to determine the effects of atropine (6 mg/kg) and clonidine (0.2 mg/kg) on soman-induced lethality and behavioral changes in the rat. Soman produced a dose-dependent increase in lethality over a narrow concentration range (50-200 micrograms/kg, SC). Soman produced time- and dose-dependent increases in tremor, salivation, hind limb extension, convulsions and chewing behaviors, as well as decreases in three normal stereotyped behaviors, sniffing, locomotion and rearing. Atropine and clonidine were equally effective at limiting soman-induced lethality and behavioral changes. The protective effects of clonidine and atropine were synergistic, even though clonidine antagonizes some of the stereotyped behaviors elicited by atropine. Simultaneous pretreatment with clonidine and atropine completely eliminated the lethality and behavioral changes produced by injection of 200 micrograms/kg soman.
ESTHER : Buccafusco_1988_Pharmacol.Biochem.Behav_29_309
PubMedSearch : Buccafusco_1988_Pharmacol.Biochem.Behav_29_309
PubMedID: 3362926

Title : Interactions of piperidine derivatives with the nicotinic cholinergic receptor complex from Torpedo electric organ - Aronstam_1988_Neurochem.Res_13_171
Author(s) : Aronstam RS , Edwards MW , Daly JW , Albuquerque EX
Ref : Neurochem Res , 13 :171 , 1988
Abstract : The interactions of eight piperidine derivatives with nicotinic receptor complexes from Torpedo californica electric organ were studied using [125I] alpha-bungarotoxin [( 125I]BGT) as a probe for the acetylcholine binding site and [3H]perhydrohistrionicotoxin [( 3H]H12-HTX) as a probe for a site associated with the receptor-gated ion channel. Cis- and trans-2-methyl-6-n-undecanyl piperidines (MUP), major constituents of fire ant venom, had a high-affinity for [3H]H12-HTX binding sites (Ki = 0.08-0.24 microM), but had no affect on receptor binding. MUP affinity for [3H]H12-HTX binding sites was approximately doubled in the presence of 1 microM carbamylcholine. Introduction of a 2'-hydroxyl group to the undecanyl side channel had little effect on activity of the alkaloid. The analog 2,6- (but not 3,5-) dimethylpiperidine was a moderately active inhibitor of [3H]H12-HTX binding (Ki = 8.8 microM). 2-Methylpiperidine was considerably less active (Ki = 600 microM), although it was more potent than either 3- or 4-methylpiperidine. The affinities of 2,6-dimethylpiperidine and 2-methylpiperidine for [3H]H12-HTX binding sites were decreased in the presence of 1 microM carbamylcholine. Carbamylcholine affinity for the receptor was increased by up to 7 fold in the presence of 10 and 32 microM MUP, but was decreased in the presence of 2,6-dimethylpiperidine and 2-methylpiperidine. The cis- and trans-isomers of MUP were equipotent in producing each of its effects. In these actions, MUP resembles a variety of other compounds derived from 2,6-disubstituted piperidines, including histrionicotoxins, gephyrotoxins and pumiliotoxins.(ABSTRACT TRUNCATED AT 250 WORDS)
ESTHER : Aronstam_1988_Neurochem.Res_13_171
PubMedSearch : Aronstam_1988_Neurochem.Res_13_171
PubMedID: 2452357

Title : Protection by clonidine from the cardiovascular changes and lethality following soman administration in the rat: role of brain acetylcholine - Magri_1988_Toxicol.Appl.Pharmacol_95_464
Author(s) : Magri V , Buccafusco JJ , Aronstam RS
Ref : Toxicol Appl Pharmacol , 95 :464 , 1988
Abstract : The organophosphate cholinesterase inhibitor soman produced a dramatic increase in arterial blood pressure of up to 60 mm Hg in the unanesthetized rat with variable changes in heart rate. Pretreatment with clonidine resulted in a dose-related inhibition of soman-induced cardiovascular changes and enhanced the time of survival as well as the number of animals surviving 24 hr after soman injection. High doses of clonidine itself produced a transient pressor response; however, the peripheral actions of clonidine were not important for its protective actions. The paradigm of repeated injections of a sublethal dose of soman at regular intervals produced the most reliable cardiovascular changes and degree of lethality. The development of the pressor response under these conditions paralleled the increase in regional brain acetylcholine levels. Pretreatment with a protective dose of clonidine did not alter steady-state levels of acetylcholine, but did inhibit the soman-induced increase in cerebral cortex, medulla-pons, and midbrain, but not that in the striatum. These results are consistent with the ability of clonidine to offer protection against centrally acting acetylcholinesterase inhibitors as a result of marked inhibition of cholinergic function in certain brain regions.
ESTHER : Magri_1988_Toxicol.Appl.Pharmacol_95_464
PubMedSearch : Magri_1988_Toxicol.Appl.Pharmacol_95_464
PubMedID: 3188010

Title : Diethyl ether effects on muscarinic acetylcholine receptor complexes in rat brainstem - Anthony_1988_Biochem.Pharmacol_37_4041
Author(s) : Anthony BL , Dennison RL, Jr. , Narayanan TK , Aronstam RS
Ref : Biochemical Pharmacology , 37 :4041 , 1988
Abstract : The influence of diethyl ether on muscarinic acetylcholine receptor-G protein interactions was studied using membranes isolated from rat brainstem. Membranes were equilibrated with diethyl ether (0.5 to 10%) for 20 min before, and then during, the binding assay. The affinity, but not the number, of [3H]N-methylscopolamine [( 3H]MS) binding sites was increased in the presence of diethyl ether (KD in air = 0.41 nM, KD in 2% diethyl ether = 0.21 nM). This increase in affinity reflected a decrease in the rapid dissociation rate constant (air k-1 = 13 X 10(-3) min-1, 2% diethyl ether k-1 = 7 X 10(-4) min-1) rather than a change in the association rate constant. Diethyl ether had no effect on the binding affinity of the muscarinic agonist carbamylcholine. However, the binding of a radiolabeled muscarinic agonist, [3H]oxotremorine-M [( 3H]Oxo-M), to high affinity binding sites decreased about 25% in the presence of 2% diethyl ether. The ability of a guanine nucleotide to depress the high affinity binding of both carbamylcholine and [3H]Oxo-M was decreased or eliminated by diethyl ether. Diethyl ether appears to interfere with muscarinic receptor-G protein interactions, perhaps by stabilizing receptor-G protein complexes or inhibiting the binding of guanine nucleotides.
ESTHER : Anthony_1988_Biochem.Pharmacol_37_4041
PubMedSearch : Anthony_1988_Biochem.Pharmacol_37_4041
PubMedID: 3142483

Title : Halothane attenuation of muscarinic inhibition of adenylate cyclase in rat heart - Narayanan_1988_Biochem.Pharmacol_37_1219
Author(s) : Narayanan TK , Confer RA , Dennison RL, Jr. , Anthony BL , Aronstam RS
Ref : Biochemical Pharmacology , 37 :1219 , 1988
Abstract : Halothane stimulated basal adenylate cyclase activity in rat cardiac membranes. Maximal stimulation (54%) was obtained after equilibrating the membranes with 2% halothane. Halothane did not affect the fractional stimulation of adenylate cyclase activity produced by either forskolin or isoproterenol. However, halothane decreased carbamylcholine inhibition of adenylate cyclase activity stimulated by both forskolin and isoproterenol. Maximal depression of carbamylcholine inhibition of stimulated cyclase activity was obtained after equilibration with 1% halothane. These results are consistent with evidence from ligand binding studies and indicate that halothane disrupts muscarinic receptor-G-protein interactions.
ESTHER : Narayanan_1988_Biochem.Pharmacol_37_1219
PubMedSearch : Narayanan_1988_Biochem.Pharmacol_37_1219
PubMedID: 3128298

Title : Cholinergic false transmitters: physiological and biochemical actions in central and peripheral systems - Aronstam_1988_Neuropharmacol_27_217
Author(s) : Aronstam RS , Marshall DC , Buccafusco JJ
Ref : Neuropharmacology , 27 :217 , 1988
Abstract : Three N-ethyl substituted analogs of acetylcholine (ACh) were evaluated for potential use as false neurotransmitters to decrease cholinergic transmission. This evaluation included (1) the elevation of arterial blood pressure upon central administration, (2) depression of blood pressure upon intravenous injection and (3) interactions with central muscarinic and peripheral nicotinic receptors. With respect to the central pressor response, ACh, acetylmonoethylcholine (AMECh) and acetyldiethylcholine (ADECh) were full agonists of decreasing potency; acetyltriethylcholine (ATECh) was a partial agonist of considerably lower potency. The duration of response paralleled potency. With respect to the peripheral depressor response, ACh and AMECh were full agonists of equal potency, and ADECh and ATECh were partial agonists of at least 100-fold lower potency. In terms of their affinity for central muscarinic receptors (brainstem and cerebral cortex), the following series was obtained: ACh greater than AMECh much greater than ADECh = ATECh. All of the agents had a greater affinity for muscarinic receptors in the brainstem compared to cortex. Acetylcholine and AMECh recognized multiple receptor binding conformations; the binding of ADECh and ATECh indicated interaction with a single set of equivalent sites. The affinity for nicotinic ACh receptors from the Torpedo electric organ was ACh = AMECh much greater than ADECh; ATECh had little affinity for these receptors. Acetylcholine, AMECh and ADECh stimulated the binding of [3H]phencyclidine to the ion channel of nicotinic receptor (potency series = ACh greater than AMECh = ADECh); ATECh was inactive. Acetylcholine, AMECh and ADECh also induced receptor conversion to a desensitized conformation; ATECh did not.(ABSTRACT TRUNCATED AT 250 WORDS)
ESTHER : Aronstam_1988_Neuropharmacol_27_217
PubMedSearch : Aronstam_1988_Neuropharmacol_27_217
PubMedID: 2836748

Title : Temperature effect on the detection of muscarinic receptor-G protein interactions in ligand binding assays - Aronstam_1988_Biochem.Pharmacol_37_1045
Author(s) : Aronstam RS , Narayanan TK
Ref : Biochemical Pharmacology , 37 :1045 , 1988
Abstract : The ability of guanine nucleotides to lower agonist binding affinity provides a convenient indication of receptor-G protein coupling: guanine nucleotides convert muscarinic receptors from high-affinity states for agonists to low-affinity states. We studied the influence of assay temperature on the demonstration of this coupling in rat brainstem and atrium. Agonist affinity of brainstem receptors increased as temperature was lowered, reflecting a greater proportion of receptors in high-affinity conformations. The influence of 5'-guanylylimidodiphosphate, a stable analog of GTP, on agonist binding, determined directly (using [3H]oxotremorine-M) or indirectly (in [3H]N-methylscopolamine/carbamylcholine competition studies), was greatest from 16 to 20 degrees. Guanine nucleotide sensitivity was much reduced at 0-4 degrees and 37 degrees. Brainstem and atrial muscarinic receptors were similarly affected by temperature. We suggest that high-affinity receptor-G protein complexes are unstable at high temperatures, thereby decreasing agonist affinity and masking the guanine nucleotide effect. At low temperatures, the receptor-G protein complex is stabilized and fails to dissociate in the presence of guanine nucleotides. The optimum temperature for monitoring receptor-G protein interactions in binding assays was 16-20 degrees.
ESTHER : Aronstam_1988_Biochem.Pharmacol_37_1045
PubMedSearch : Aronstam_1988_Biochem.Pharmacol_37_1045
PubMedID: 3128292

Title : Adrenergic agonists protect against soman, an irreversible acetylcholinesterase inhibitor - Buccafusco_1987_Toxicol.Lett_38_67
Author(s) : Buccafusco JJ , Aronstam RS
Ref : Toxicol Lett , 38 :67 , 1987
Abstract : Adrenergic drugs from several chemical classes were compared with clonidine as protectants against soman toxicity. All agents tested produced some degree of protection. Alpha-adrenergic agonists such as methyldopa, xylazine and clonidine were among the most effective drugs, while a dopaminergic agonist (bromocriptine) and beta-receptor antagonist (pindolol) were the least effective. Potency for protection was related to affinity for alpha-adrenergic binding sites labelled with [3H]clonidine. Atropine acted synergistically with adrenergic agonists to potentiate protection.
ESTHER : Buccafusco_1987_Toxicol.Lett_38_67
PubMedSearch : Buccafusco_1987_Toxicol.Lett_38_67
PubMedID: 2820088

Title : Effects of halothane on high affinity agonist binding and guanine nucleotide sensitivity of muscarinic acetylcholine receptors from brainstem of rat - Dennison_1987_Neuropharmacol_26_1201
Author(s) : Dennison RL, Jr. , Anthony BL , Narayanan TK , Aronstam RS
Ref : Neuropharmacology , 26 :1201 , 1987
Abstract : The influence of halothane on muscarinic receptors with a high affinity for agonists was studied using [3H]oxotremorine-M. [3H]Oxotremorine-M bound with high affinity (KD = 2.8 nM) to a subpopulation of muscarinic receptors in the brainstem of rat, representing 32% of the total receptor pool. Agonist affinity for binding sites for [3H]oxotremorine-M was not affected by a guanine nucleotide (5'-guanylylimidodidiphosphate; Gpp(NH)p), although the level of binding was decreased, presumably due to the conversion of receptors to lower affinity conformations. However, only 58% of 3 nM binding of [3H]oxotremorine-M was sensitive to Gpp(NH)p. Halothane had two effects on the binding of [3H]oxotremorine-M: halothane (1) decreased the level of binding of [3H]oxotremorine-M without affecting agonist affinity for the surviving sites, and (2) lowered the sensitivity of the binding of [3H]oxotremorine-M to Gpp(NH)p by a factor of 120. The decrease in binding of [3H]oxotremorine-M binding was nonselective with regard to the sensitivity of the receptors to the guanine nucleotide, insofar as Gpp(NH)p inhibited the binding of [3H]oxotremorine-M to the same extent in the presence and absence of halothane. These results suggest that halothane (1) converts both G protein-coupled and -uncoupled muscarinic receptors to states of lower agonist affinity and (2) lowers the affinity of receptor-G protein complexes for guanine nucleotides.
ESTHER : Dennison_1987_Neuropharmacol_26_1201
PubMedSearch : Dennison_1987_Neuropharmacol_26_1201
PubMedID: 3116449

Title : Altered muscarinic receptor properties and function in the heart in diabetes - Carrier_1987_J.Pharmacol.Exp.Ther_242_531
Author(s) : Carrier GO , Aronstam RS
Ref : Journal of Pharmacology & Experimental Therapeutics , 242 :531 , 1987
Abstract : The cardiac cholinergic system was studied in streptozotocin (STZ)-diabetic and age-matched control rats. STZ-diabetic rats (8-10 weeks) were supersensitive to the negative chronotropic effects of acetylcholine, carbamylcholine and bethanechol; inotropic responses to these muscarinic agonists were unaltered. This phenomenon was associated with a decrease in acetylcholinesterase activity but no change in the rate and extent of neuronal choline uptake. [3H]N-methylscopolamine bound to muscarinic receptors in atria from both groups of rats with the same high affinity. The density of [3H]N-methylscopolamine binding sites, however, was 34% lower in atria from STZ-diabetic rats. Agonist binding affinity was lower in diabetes; carbamylcholine had a lower affinity for both the high- and low-affinity receptors. These results indicate that cardiac cholinergic supersensitivity in right atria in diabetes occurs before the development of autonomic neuropathy insofar as neuronal [3H]choline uptake is unaltered at this stage of STZ diabetes. Changes in agonist binding conformation, without a concomitant change in antagonist binding affinity, suggest that supersensitivity of right atria to muscarinic agonist may be a consequence of altered coupling of muscarinic receptor to transduction mechanisms involved in chronotropism in diabetes.
ESTHER : Carrier_1987_J.Pharmacol.Exp.Ther_242_531
PubMedSearch : Carrier_1987_J.Pharmacol.Exp.Ther_242_531
PubMedID: 2956413

Title : Clonidine prevents the short-term down regulation of muscarinic receptors in mouse brain induced by the acetylcholinesterase inhibitor soman - Aronstam_1987_Neurosci.Lett_78_107
Author(s) : Aronstam RS , Smith MD , Buccafusco JJ
Ref : Neuroscience Letters , 78 :107 , 1987
Abstract : The influence of clonidine on the down regulation of muscarinic acetylcholine receptors induced by the irreversible acetylcholinesterase inhibitor soman was studied in mouse brain (cortex and brainstem). Muscarinic receptor binding was measured using [N-methyl-3H]scopolamine ([3H]MS) as the probe in a filtration assay. Thirty min after subcutaneous injection of soman at a dose which killed 80-90% of the mice, the concentration of [3H]MS binding sites was reduced by 25-28% in surviving mice. [3H]MS binding affinity (Kd = 0.36-0.42 nM), agonist binding state, and receptor coupling to guanine nucleotide-dependent transducer proteins (G proteins) were not affected by soman. Administration of clonidine (1 mg/kg, s.c.) 5 min prior to soman prevented the decrease in receptor number and decreased the extent of acetylcholinesterase inhibition caused by the soman. It is possible that a reversible inhibition of acetylcholinesterase by clonidine protects the enzyme from irreversible inactivation by soman, thereby decreasing the extent of chronic cholinergic overstimulation with its attendant down regulation of muscarinic receptors.
ESTHER : Aronstam_1987_Neurosci.Lett_78_107
PubMedSearch : Aronstam_1987_Neurosci.Lett_78_107
PubMedID: 3614769

Title : Clonidine protection from the toxicity of soman, an organophosphate acetylcholinesterase inhibitor, in the mouse - Buccafusco_1986_J.Pharmacol.Exp.Ther_239_43
Author(s) : Buccafusco JJ , Aronstam RS
Ref : Journal of Pharmacology & Experimental Therapeutics , 239 :43 , 1986
Abstract : Mice pretreated with the centrally active alpha-2 adrenergic agonist, clonidine, were protected from several of the toxic manifestations of soman, an organophosphate acetylcholinesterase inhibitor. The protection resulted in increased survival rates and a reduction in centrally mediated symptoms of soman, including tremor and straub tail, as well as one peripheral muscarinic symptom, excessive salivation. Doses of clonidine between 0.1 and 1 mg/kg, administered between 5 and 40 min before LD80 to LD90 doses of soman, produced significant protection. Pretreatment with atropine (25 mg/kg) also protected against soman toxicity. When atropine was combined with clonidine, the degree of protection afforded by the combination was greater than that predicted for a simple additive effect. Mice protected by atropine from the initial toxicity of soman frequently died within 24 h; no such delayed lethality was observed with protective doses of clonidine. Clonidine noncompetitively inhibited acetylcholinesterase activity in vitro and after in vivo administration at protective doses. At brain concentrations obtained after in vivo administration in protective doses, clonidine also inhibited ligand binding to cortical muscarinic receptors in vitro. The protective effects of clonidine are likely to involve multiple effects, including blockade of acetylcholine release and postsynaptic muscarinic receptors and transient inhibition of acetylcholinesterase.
ESTHER : Buccafusco_1986_J.Pharmacol.Exp.Ther_239_43
PubMedSearch : Buccafusco_1986_J.Pharmacol.Exp.Ther_239_43
PubMedID: 3761196

Title : Interaction of gephyrotoxin and indolizidine alkaloids with the nicotinic acetylcholine receptor-ion channel complex of Torpedo electroplax - Aronstam_1986_Neurochem.Res_11_1227
Author(s) : Aronstam RS , Daly JW , Spande TF , Narayanan TK , Albuquerque EX
Ref : Neurochem Res , 11 :1227 , 1986
Abstract : The interactions of eighteen natural and synthetic gephyrotoxin and indolizidine alkaloids with binding sites on nicotinic acetylcholine receptor channel (AChR) complex from Torpedo californica electric organ were investigated using two radiolabeled probes, [3H]perhydrohistrionicotoxin and [3H]phencyclidine. Both gephyrotoxins and indolizidines were moderately active inhibitors of the binding of these probes (Ki's = 0.1-20 microM), but did not interact with the acetylcholine binding site. Structure-activity relationships indicate an important contribution of hydrophobic interactions to both gephyrotoxin and indolizidine binding. The stereoconfiguration of the alkaloids had little effect on binding. Carbamylcholine enhanced the affinity of certain alkaloids up to 6 to 8-fold suggesting that interactions with open or desensitized conformations of the AChR complex are favored over interactions with resting conformations.
ESTHER : Aronstam_1986_Neurochem.Res_11_1227
PubMedSearch : Aronstam_1986_Neurochem.Res_11_1227
PubMedID: 2431336

Title : Effect of pH on muscarinic acetylcholine receptors from rat brainstem - Anthony_1986_J.Neurochem_46_556
Author(s) : Anthony BL , Aronstam RS
Ref : Journal of Neurochemistry , 46 :556 , 1986
Abstract : The effect of hydrogen ion concentration on ligand binding to muscarinic acetylcholine receptors was studied in membranes isolated from rat brainstem. The binding of [3H]methylscopolamine was constant between pH 7 and 10. The affinity, but not the number, of [3H]methylscopolamine binding sites decreased below pH 7; at pH 4 little binding was detected. When brainstem membranes were incubated at various pH levels from 3 to 11 for 1 h and then returned to pH 8, [3H]methylscopolamine binding affinity was restored to control levels. Carbamylcholine binding affinity was also depressed in media of low pH. However, this decrease was permanent after a 1-h incubation at pH 4 (i.e. carbamylcholine affinity was not restored on raising the pH to 8). The capacity of a guanine nucleotide to affect carbamylcholine was also abolished by a 1-h incubation at pH 4, and was not restored by raising the pH. The guanine nucleotide-dependent regulatory protein may be irreversibly inactivated or dissociated from the receptor at low pH. The receptor's binding subunit, on the other hand, appears to be much less sensitive to hydrogen ion concentration.
ESTHER : Anthony_1986_J.Neurochem_46_556
PubMedSearch : Anthony_1986_J.Neurochem_46_556
PubMedID: 3079819

Title : High-affinity agonist binding to rat brainstem muscarinic receptors is eliminated by low pH - Anthony_1986_Neurosci.Lett_69_84
Author(s) : Anthony BL , Aronstam RS
Ref : Neuroscience Letters , 69 :84 , 1986
Abstract : The effects of hydrogen ion concentration on high-affinity agonist binding to muscarinic receptors were determined in rat brainstem membranes using [3H]oxotremorine-M as a probe. [3H]Oxotremorine-M bound with high affinity to a small subpopulation of brainstem muscarinic receptors (10% of the receptors labelled by [3H]methylscopolamine). [3H]Oxotremorine-M binding was constant between pH 7.0 and 9.0; the number of high-affinity sites decreased below pH 7.0 and at pH 5.0 no binding was detected. This decrease was irreversible; when brainstem membranes were incubated for 1 h at low pH and then returned to pH 8.0, agonist binding was not restored. In contrast, the total number of receptors, i.e. the number of [3H]antagonist binding sites, was not affected by prolonged incubation at low pH. Agonist affinity for the surviving [3H]oxotremorine binding sites and the sensitivity of agonist binding to guanine nucleotides were not altered in media of low pH (pH 5.5). These findings suggest that [3H]oxotremorine-M binds only to receptors which are functionally coupled to guanine nucleotide-dependent regulatory proteins and that this coupling is irreversibly inactivated in media of low pH.
ESTHER : Anthony_1986_Neurosci.Lett_69_84
PubMedSearch : Anthony_1986_Neurosci.Lett_69_84
PubMedID: 3748469

Title : Clonidine protection from soman and echothiophate toxicity in mice - Aronstam_1986_Life.Sci_39_2097
Author(s) : Aronstam RS , Smith MD , Buccafusco JJ
Ref : Life Sciences , 39 :2097 , 1986
Abstract : The influence of clonidine on the toxicity produced by two irreversible, organophosphate cholinesterase inhibitors, soman and echothiophate, was studied in mice. At lethal doses, soman produced whole body tremor but no muscle fasciculation; at lethal doses, echothiophate produced muscle fasciculations but no whole body tremor. Pretreatment with clonidine protected against several toxic manifestations of soman, but had little effect on echothiophate toxicity. In addition to its documented effects on acetylcholine metabolism, clonidine was found to be a weak inhibitor of acetylcholinesterase. At certain concentrations, clonidine protected the enzyme from permanent inactivation by soman. These findings indicate that the toxicity of soman and echothiophate reflect primarily central and peripheral actions, respectively, and that clonidine has a much greater protective effect versus the centrally-acting agent. Moreover, direct interactions with acetylcholinesterase may contribute to clonidine protection from cholinesterase inhibitor toxicity.
ESTHER : Aronstam_1986_Life.Sci_39_2097
PubMedSearch : Aronstam_1986_Life.Sci_39_2097
PubMedID: 3784771

Title : Selective antagonism by clonidine of the stereotyped and non-stereotyped motor activity elicited by atropine - Molloy_1986_Pharmacol.Biochem.Behav_25_985
Author(s) : Molloy AG , Aronstam RS , Buccafusco JJ
Ref : Pharmacol Biochem Behav , 25 :985 , 1986
Abstract : The effects of clonidine, an indirectly-acting cholinergic antagonist, on 5 behaviors elicited by atropine (locomotion, rearing, sniffing, grooming and gnawing) were studied in rats. Clonidine did not alter the prevalence or magnitude of atropine-elicited locomotion and rearing. In contrast, clonidine suppressed the occurrence and degree of 3 stereotyped behaviors, namely, sniffing, grooming and gnawing. This selectivity of clonidine suggests differences in the neural pathways subserving the various stereotyped motor activities.
ESTHER : Molloy_1986_Pharmacol.Biochem.Behav_25_985
PubMedSearch : Molloy_1986_Pharmacol.Biochem.Behav_25_985
PubMedID: 3786370

Title : Allosteric effect of gallamine on muscarinic cholinergic receptor binding: influence of guanine nucleotides and conformational state - Narayanan_1986_Neurochem.Res_11_1397
Author(s) : Narayanan TK , Aronstam RS
Ref : Neurochem Res , 11 :1397 , 1986
Abstract : Gallamine interacts with an allosteric site on muscarinic acetylcholine receptor complexes in rat brain membranes, thereby slowing the dissociation of a radiolabelled ligand ([3H]N-methylscopolamine) from the receptor complex. This effect involves the elimination of the fast component of the biphasic dissociation curve. The allosteric effect of gallamine is equally prominent in membranes containing predominantly M1 (cerebral cortex) and M2 (brainstem) subtypes of muscarinic receptor. Gallamine's action is not affected by a variety of treatments which influence the conformational state of the receptor as reflected by agonist binding affinity, including treatments with heat, N-ethylmaleimide and trypsin. A guanine nucleotide (5'-guanylylimidodiphosphate), however, moderates the effects of gallamine on muscarinic receptors in brainstem, but not in cortical, membranes.
ESTHER : Narayanan_1986_Neurochem.Res_11_1397
PubMedSearch : Narayanan_1986_Neurochem.Res_11_1397
PubMedID: 3785535

Title : Halothane effects on muscarinic acetylcholine receptor complexes in rat brain - Aronstam_1986_Biochem.Pharmacol_35_667
Author(s) : Aronstam RS , Anthony BL , Dennison RL, Jr.
Ref : Biochemical Pharmacology , 35 :667 , 1986
Abstract : Muscarinic acetylcholine receptors in membranes from rat cerebral cortex or brainstem were equilibrated with halothane (0.5 to 5%). Halothane did not affect the number of [3H]methylscopolamine [( 3H]MS) binding sites. [3H]MS binding affinity, however, was increased in the presence of halothane (KD, air = 0.41 nM; KD, 2% halothane = 0.26 nM). This increase reflected a decrease in the dissociation rate constant (from 13 X 10(-3) min-1 to 6.5 X 10(-3) min-1) rather than a change in the bimolecular rate constant of association (1.8 and 1.9 X 10(7) M-1 min-1 in the absence and presence of 2% halothane respectively). Carbamylcholine affinity for brainstem or cortical muscarinic receptors was not affected by halothane. The ability of a guanine nucleotide to lower carbamylcholine affinity for brainstem receptors, however, was eliminated after equilibration with 2% halothane.
ESTHER : Aronstam_1986_Biochem.Pharmacol_35_667
PubMedSearch : Aronstam_1986_Biochem.Pharmacol_35_667
PubMedID: 3947397

Title : Interactions of chlordecone (kepone) and mirex with the nicotinic acetylcholine receptor--ion channel complex - Aronstam_1986_Toxicol.Lett_30_247
Author(s) : Aronstam RS , Hong JS
Ref : Toxicol Lett , 30 :247 , 1986
Abstract : Interactions of chlordecone (kepone) and mirex with nicotinic acetylcholine (ACh) receptor complexes from Torpedo californica electric organs were studied using biochemical probes for ACh and ion-channel binding sites. Neither compound inhibited the binding of [125I] alpha-bungarotoxin (BGT) to the receptor; chlordecone, however, enhanced carbamylcholine affinity for the receptor 5-fold. Chlordecone, but not mirex, also inhibited the binding of [3H]perhydrohistrionicotoxin and [3H]phencyclidine to sites associated with the receptor-gated ion channel. Ion-channel inhibition by chlordecone was enhanced in the presence of carbamylcholine. These results indicate that chlordecone, but not mirex, interacts with the ion-translocation mechanism associated with nicotinic ACh receptors, where it may sterically block ion flux as well as stabilize a desensitized conformation of the receptor complex.
ESTHER : Aronstam_1986_Toxicol.Lett_30_247
PubMedSearch : Aronstam_1986_Toxicol.Lett_30_247
PubMedID: 2422792

Title : Molecular mechanisms of the potent and stereospecific nicotinic receptor agonist (+)-anatoxin-a - Swanson_1986_Mol.Pharmacol_29_250
Author(s) : Swanson KL , Allen CN , Aronstam RS , Rapoport H , Albuquerque EX
Ref : Molecular Pharmacology , 29 :250 , 1986
Abstract : Anatoxin-a (AnTX) was shown to be a highly potent and stereospecific agonist at nicotinic synapses in frog skeletal muscle and Torpedo electric organs. AnTX binds to the nicotinic-acetylcholine receptor with a higher affinity than for acetylcholine (ACh) but does not bind to sites in the receptor-gated ionic channel. (+)AnTX caused receptor desensitization, i.e., the loss of agonist-stimulated binding of histrionicotoxin to an allosteric site with time, at a rate significantly slower than that of ACh. Single channel patch clamp recordings indicated that the conductance of channels activated by (+)AnTX (28 pS) and ACh (27 pS) were similar. The (+)AnTX-activated channels contained rapid closing events, the burst times caused by the toxin were shorter than those caused by ACh but had similar voltage dependencies, and the number of short closures per burst was constant at all potentials with both agonists. The bursts of rapid openings and rapid closures (tau = 0.4 msec) appear to result from repetitive opening and closing of the (+)AnTX-bound receptor-ion channel. It is concluded that the semirigid molecule and secondary amine (+)AnTX is a more potent agonist than ACh or carbamylcholine because of a higher affinity for the receptor. At various concentrations the toxin activates the appearance of channels with the same conductances as ACh-induced channels but with a shorter channel lifetime.
ESTHER : Swanson_1986_Mol.Pharmacol_29_250
PubMedSearch : Swanson_1986_Mol.Pharmacol_29_250
PubMedID: 2419745

Title : Poster: Differential inactivation of muscarinic receptor-G protein interactions in rat brainstems by physical and enzymatic means -
Author(s) : Aronstam RS , Anthony SL , Carrier GO
Ref : Trends in Pharmacological Sciences , Suppl :78 , 1986
PubMedID:

Title : Biphasic regulation of detergent-solubilized muscarinic acetylcholine receptors from rat cerebral cortex by N-ethylmaleimide - Dennison_1985_Res.Commun.Chem.Pathol.Pharmacol_47_465
Author(s) : Dennison RL, Jr. , Wenger DA , Aronstam RS
Ref : Res Commun Chem Pathol Pharmacol , 47 :465 , 1985
Abstract : Muscarinic acetylcholine receptors were solubilized from rat cerebral cortex with 1% digitonin. Treatment with N-ethylmaleimide decreased or increased agonist affinity for the receptor, depending on the extent of alkylation. This indicates that certain determinants of receptor state do not depend on interactions with other membrane structures.
ESTHER : Dennison_1985_Res.Commun.Chem.Pathol.Pharmacol_47_465
PubMedSearch : Dennison_1985_Res.Commun.Chem.Pathol.Pharmacol_47_465
PubMedID: 3992023

Title : Changes in cholinergic parameters associated with failure of conotruncal septation in embryonic chick hearts after neural crest ablation - Kirby_1985_Circ.Res_56_392
Author(s) : Kirby ML , Aronstam RS , Buccafusco JJ
Ref : Circulation Research , 56 :392 , 1985
Abstract : Cells from the neural crest over occipital somites migrate to the heart, where they give rise to parasympathetic postganglionic neurons as well as ectomesenchymal elements which contribute to conotruncal septation. With a microcautery needle, the neural crest over occipital somites was ablated bilaterally in chicken embryos at an early stage of development. Histological examination on incubation day 15 revealed conotruncal malformations, involving malformation or absence of the conotruncal septum in all embryos. Two peaks of embryo mortality were observed. One peak (incubation days 6-8) occurred at the same time as conotruncal septal closure; the second peak (incubation days 11-13) was concurrent with the onset of functional parasympathetic innervation. A disruption of parasympathetic innervation was indicated by: (1) a decrease in acetylcholinesterase staining, (2) a decrease (27%) in the number of ganglion cells in the conotruncus, (3) decreases in the acetylcholine content of atrium (31%) and ventricle (39%), and (4) a decrease (21%) in muscarinic acetylcholine receptor density on incubation day 15. Radiolabeled ligand-binding studies revealed no change in the affinity of cardiac muscarinic receptors for [3H]methylscopolamine (KD = 0.17-0.21 nM). Agonist-binding affinity and sensitivity to guanine nucleotides were similarly unaffected. The reasons for the limited extent of the parasympathetic lesion are unclear, but may involve recruitment of precursor cells from other regions of the neural crest, partial regeneration of the neural crest following surgical removal, or an alteration in the contribution of incoming sympathetic or preganglionic parasympathetic elements. No such plasticity was associated with neural crest contributions to the structural development of the conotruncus. Malformations were observed in all lesioned embryos.
ESTHER : Kirby_1985_Circ.Res_56_392
PubMedSearch : Kirby_1985_Circ.Res_56_392
PubMedID: 3971512

Title : Interactions of microtubule-active agents with nicotinic acetylcholine receptors. Relationship to their inhibition of catecholamine secretion by adrenal chromaffin cells - McKay_1985_Mol.Pharmacol_28_10
Author(s) : McKay DB , Aronstam RS , Schneider AS
Ref : Molecular Pharmacology , 28 :10 , 1985
Abstract : Several microtubule-active drugs block cholinergically mediated catecholamine secretion from adrenal chromaffin cells without affecting secretion induced by other secretagogues. Interactions of these agents with nicotinic acetylcholine receptor-ion channel complexes from Torpedo californica electric organs were studied using radiolabeled probes for receptor and associated ion channel-binding sites. Colchicine, taxol, and the Vinca alkaloids had minimal affinity for cholinergic receptor-binding sites (nicotinic or muscarinic). The Vinca alkaloids (vinblastine, vincristine, vindesine) and colchicine inhibited [3H]perhydrohistrionicotoxin ([3H]H12-HTX) binding to the receptor-gated ion channel with IC50 values of 2-32 microM and 6 mM, respectively. The ability of the microtubule-active drugs to inhibit [3H]H12-HTX binding was increased by up to 5-fold in the presence of 1 microM carbamylcholine. The IC50 values for inhibition of [3H]H12-HTX binding by colchicine and three Vinca alkaloids were closely correlated with their abilities to inhibit acetylcholine-induced catecholamine secretion from cultured bovine adrenal chromaffin cells. As a consequence of its interaction (direct or indirect) with the ion channel, at least one Vinca alkaloid (vinblastine) stabilized a high agonist affinity conformation of the nicotinic receptor complex. beta-Lumicolchicine, an analog of colchicine devoid of microtubule activity, also blocked ion channel binding. On the other hand, taxol, a microtubule-stabilizing agent which also selectively blocks cholinergically mediated secretion, did not affect receptor or ion channel binding. The present results indicate that interactions with the nicotinic receptor-ion channel complex may underlie the actions of certain microtubule-active agents on catecholamine secretion by adrenal chromaffin cells.
ESTHER : McKay_1985_Mol.Pharmacol_28_10
PubMedSearch : McKay_1985_Mol.Pharmacol_28_10
PubMedID: 2410767

Title : Binding of [3H]perhydrohistrionicotoxin and [3H]phencyclidine to the nicotinic receptor-ion channel complex of Torpedo electroplax. Inhibition by histrionicotoxins and derivatives - Aronstam_1985_Biochem.Pharmacol_34_3037
Author(s) : Aronstam RS , King CT, Jr. , Albuquerque EX , Daly JW , Feigl DM
Ref : Biochemical Pharmacology , 34 :3037 , 1985
Abstract : Histrionicotoxin, a spiropiperidine alkaloid, and twenty-two analogs inhibited binding of [3H]perhydrohistrionicotoxin [( 3H]H12-HTX) and of [3H]phencyclidine [( 3H]PCP) to sites on the acetylcholine receptor-ion complex of Torpedo electroplax membranes. Structural alterations to the nitrogen (secondary amine) or oxygen (alcohol) functions or to the five carbon and four carbon side chain of histrionicotoxin altered the potency versus [3H]H12-HTX and [3H]PCP binding measured in the presence or absence of a receptor agonist, carbamylcholine. Histrionicotoxin itself was 3-fold more potent versus [3H]PCP binding than versus [3H]H12-HTX binding. N-Methylation or O-acetylation increased this difference, while alterations to the side chains either slightly decreased or markedly increased this difference. Histrionicotoxin was some 3.5-fold more potent versus [3H]H12-HTX binding in the presence of carbamylcholine than in its absence. O-Acetylation increased this selectivity for the carbamylcholine-activated state of the receptor channel complex, while alterations in the side chains either reduced or increased the selectivity. Histrionicotoxin was some 2.2-fold more potent versus [3H]PCP binding in the presence of carbamylcholine than in its absence. N-Methylation of O-acetyl-histrionicotoxin greatly increased this selectivity, while alterations in the side chains either reduced or had no effect on selectivity.
ESTHER : Aronstam_1985_Biochem.Pharmacol_34_3037
PubMedSearch : Aronstam_1985_Biochem.Pharmacol_34_3037
PubMedID: 2412560

Title : Muscarinic acetylcholine receptors in chick heart: influence of heat and N-ethylmaleimide on receptor conformations and interactions with guanine nucleotide-dependent regulatory proteins - Aronstam_1985_Neurosci.Lett_54_289
Author(s) : Aronstam RS , Kirby ML , Smith MD
Ref : Neuroscience Letters , 54 :289 , 1985
Abstract : Several conformations of muscarinic acetylcholine receptors in chick heart are revealed by the energetics of agonist binding. Heating cardiac membranes at 50 degrees C for 5 min decreased agonist affinity, steepened agonist binding curves and eliminated receptor sensitivity to guanine nucleotides. N-ethylmaleimide (NEM) steepened agonist binding curves and eliminated receptor sensitivity to guanine nucleotides without decreasing agonist binding affinity. NEM prevented and reversed the effect of heat on agonist affinity. Pirenzepine and N-methylscopolamine binding were unaffected by either treatment. NEM and heat stabilize muscarinic receptors in conformations unrelated to those associated with naturally occurring receptor populations.
ESTHER : Aronstam_1985_Neurosci.Lett_54_289
PubMedSearch : Aronstam_1985_Neurosci.Lett_54_289
PubMedID: 3838808

Title : Regional heterogeneity of muscarinic acetylcholine receptors from rat brain is retained after detergent solubilization - Wenger_1985_Neurosci.Lett_54_65
Author(s) : Wenger DA , Parthasarthy N , Aronstam RS
Ref : Neuroscience Letters , 54 :65 , 1985
Abstract : Muscarinic acetylcholine receptors were extracted from rat cerebral cortex, hippocampus and brainstem membranes with 1% digitonin. Solubilized receptors retained both multiple receptor affinities and their regional selectivity in agonist binding (in terms of their affinity for carbamylcholine, brainstem greater than cortex greater than hippocampus). The affinity for carbamylcholine was increased 2-8-fold by solubilization; this increase was largely accounted for by an increase in the proportion of receptors displaying high affinity binding. No differences were observed in the size of receptor-detergent complexes from cortex or brainstem in gel filtration chromatography using a variety of gels and solubilizing agents. These findings indicate that not all of the factors underlying receptor heterogeneity and conformational state are removed or disrupted by solubilization.
ESTHER : Wenger_1985_Neurosci.Lett_54_65
PubMedSearch : Wenger_1985_Neurosci.Lett_54_65
PubMedID: 3974946

Title : Interactions of bupivacaine with ionic channels of the nicotinic receptor. Electrophysiological and biochemical studies - Ikeda_1984_Mol.Pharmacol_26_293
Author(s) : Ikeda SR , Aronstam RS , Daly JW , Aracava Y , Albuquerque EX
Ref : Molecular Pharmacology , 26 :293 , 1984
Abstract : The actions of the tertiary local anesthetic bupivacaine were studied on the nicotinic receptor-ionic channel complex (AChR) using electrophysiological and biochemical methods. Voltage clamp studies of the frog sartorius and cutaneous pectoris neuromuscular junction revealed a concentration-dependent depression of the decay time constant of the end-plate (tau EPC) and spontaneous miniature end-plate (tau MEPC) currents. The relationship of the reciprocal of either tau EPC or tau MEPC and bupivacaine concentration up to 100 microM was linear. Voltage dependence of EPC over the range +60 to -150 mV was reduced, whereas both EPC and MEPC decays were adequately described by a single exponential function at all concentrations tested. Peak MEPC and EPC amplitudes were also depressed in a concentration-dependent manner such that 100 microM bupivacaine reduced peak amplitude by about 50%. The current-voltage relationship remained linear under all conditions tested. Nerve-evoked responses were difficult to study at concentrations greater than 100 microM because of apparent blockade of nerve conduction. Extracellular recording of the MEPC afforded results similar to those obtained with EPCs. The tau MEPC could be reduced to less than 300 mu sec at a bupivacaine concentration of 400 microM. Fluctuation analysis showed that bupivacaine at concentrations of 10 and 25 microM did not change channel conductance but decreased single-channel lifetime to 76% and 39% of control values, respectively. Biochemical studies were performed on Torpedo californica membrane fragments using [3H]phencyclidine ([3H]PCP) and [3H]perhydrohistrionicotoxin ([3H]H12-HTX) as channel probes. Bupivacaine inhibited the binding of [3H]PCP and [3H]H12-HTX with inhibition constants (Ki) of 32 and 25 microM, respectively. The corresponding inhibition constants for bupivacaine methiodide were 1.8 and 3.2 microM. The preincubation of the membranes with carbamylcholine increased the affinity of bupivacaine for the ionic channel sites 5- to 8-fold and the affinity of bupivacaine methiodide 3- to 4-fold. Bupivacaine, however, had no affinity for the agonist recognition site as determined by [3H]ACh and [125I]alpha-bungarotoxin bindings. The electrophysiological and biochemical studies indicate that bupivacaine reacts primarily with the ionic channel of the nicotinic AChR. The results are consistent with a sequential model in which the drug interacts with the sites at the ionic channel of AChR in its open conformation, producing species with little or no conductance. From the present studies there is no evidence for an interaction of bupivacaine with the agonist binding site or closed states of AChR.
ESTHER : Ikeda_1984_Mol.Pharmacol_26_293
PubMedSearch : Ikeda_1984_Mol.Pharmacol_26_293
PubMedID: 6090884

Title : The nature of the interactions of pyridostigmine with the nicotinic acetylcholine receptor-ionic channel complex. I. Agonist, desensitizing, and binding properties - Pascuzzo_1984_Mol.Pharmacol_25_92
Author(s) : Pascuzzo GJ , Akaike A , Maleque MA , Shaw KP , Aronstam RS , Rickett DL , Albuquerque EX
Ref : Molecular Pharmacology , 25 :92 , 1984
Abstract : The actions of pyridostigmine (Pyr), an anticholinesterase agent, were studied on the acetylcholine (ACh) receptor-ion channel complex and on the electrically excitable membrane of the frog cutaneous pectoris and sartorius muscles and the chronically denervated soleus muscle of the rat. Pyr at concentrations of 0.2-0.4 mM potentiated the indirect evoked muscle twitch and at concentrations greater than or equal to 0.8 mM depressed the indirect twitch with an IC50 of about 2 mM. Twitch depression produced by Pyr was reversed slowly, and after a 60-min wash only 59% of the control muscle twitch had returned. Pyr did not affect either the membrane potential or the muscle action potential. Pyr had several effects at the neuromuscular junction of the frog and rat. It decreased the peak amplitude of the end-plate current (EPC) in a voltage- and concentration-dependent manner. In contrast to diisopropylfluorophosphate, which depresses the EPC amplitude and induces a double exponential decay of the EPC and miniature end-plate current (MEPC), Pyr produced a marked prolongation of the time constants of EPC and MEPC decay while maintaining a single exponential decay. The decrease caused by Pyr of indirect twitch tension, EPC amplitude, and ACh sensitivity indicates mechanisms which limit the number and/or properties of conducting channels. The drug decreased channel conductance and prolonged channel lifetime as revealed by Fourier analysis of ACh-induced end-plate current fluctuations. An altered form of the conducting species induced by Pyr appears to be responsible for either the apparent agonist-induced depolarization or its ability to increase the affinity of ACh for its recognition site. Pyr was also found to inhibit the binding of ACh and alpha-bungarotoxin to receptor-rich membrane from the electric organ of Torpedo nobiliana, and to have a higher affinity for the receptor than for the ion channel binding sites. These actions are distinct from acetylcholinesterase inhibition caused by the agent. Strong evidence suggests that the direct influences of the agent on neuromuscular transmission involve at least three distinct, although possibly interacting, mechanisms: (a) a weak agonist action, (b) the formation of desensitized receptor-complex intermediates, and (c) the alteration of the conductance properties of active channels.
ESTHER : Pascuzzo_1984_Mol.Pharmacol_25_92
PubMedSearch : Pascuzzo_1984_Mol.Pharmacol_25_92
PubMedID: 6323955

Title : Cholinergic supersensitivity and decreased number of muscarinic receptors in atria from short-term diabetic rats - Carrier_1984_J.Mol.Cell.Cardiol_16_963
Author(s) : Carrier GO , Edwards AD , Aronstam RS
Ref : Journal of Molecular & Cellular Cardiology , 16 :963 , 1984
Abstract : Autonomic neuropathy is a major complication of chronic diabetes and is responsible for disturbances in the cardiovascular system and other organs. Early cardiac disturbances have been attributed to defective vagal control of the heart. The heart rates of rats with chemically-induced diabetes are depressed and an increase in blood pressure produces a greater reflex bradycardia in diabetic rats. Tomlinson and Yusof found that isolated, stimulated left atria from rats made diabetic with alloxan are supersensitive to the negative inotropic influence of acetylcholine. On the other hand, Rao, et al. found that perfused working-heart preparations from streptozotocin- and alloxan-diabetic rats have a reduced sensitivity to carbamylcholine. In the present study, we measured chronotropic responses to cholinergic agonists of isolated, spontaneously-beating atria, as well as muscarinic receptor populations, in cardiac tissue from short-term (8 to 9 weeks) diabetic and age-matched control rats.
ESTHER : Carrier_1984_J.Mol.Cell.Cardiol_16_963
PubMedSearch : Carrier_1984_J.Mol.Cell.Cardiol_16_963
PubMedID: 6512866

Title : Interactions of gephyrotoxin with the acetylcholine receptor-ionic channel complex. II. Enhancement of desensitization - Souccar_1984_Mol.Pharmacol_25_395
Author(s) : Souccar C , Varanda WA , Aronstam RS , Daly JW , Albuquerque EX
Ref : Molecular Pharmacology , 25 :395 , 1984
Abstract : The actions of the tricyclic alkaloid gephyrotoxin ( GyTX ) on the extrajunctional and junctional acetylcholine (ACh) sensitivity and desensitization were studied in the chronically denervated rat soleus muscle and cutaneous pectoris muscle of the frog. At low concentrations, GyTX greatly depressed the extrajunctional ACh sensitivity of the chronically denervated soleus muscles. In addition, GyTX produced a strong inhibition of junctional end-plate potentials evoked by ACh. Junctional and extrajunctional desensitizations induced by microiontophoretically applied ACh were greatly enhanced by the alkaloid in a frequency-dependent manner. These effects were readily reversible. The interaction of GyTX with binding sites on the acetylcholine receptor-channel (AChR) complex was studied on electroplax membranes from Torpedo californica. GyTX binds to the AChR complex at a site distinct from the ACh binding site, as revealed by its lack of inhibition of [125I]alpha-bungarotoxin ( [125I]BGT) binding. On the other hand, GyTX at a concentration range between 1 microM and 100 microM significantly increased the potency of the agonist carbamylcholine as an antagonist of binding of [125I]BGT. At low micromolar concentrations, GyTX inhibited the binding of [3H]perhydrohistrionicotoxin and [3H] phencyclidine to sites associated with the ionic channel of the AChR complex. The affinity of GyTX for these sites was increased 3- to 5-fold by carbamylcholine. Results of electrophysiological and binding studies indicate that GyTX not only blocks the open channel of the AChR but also enhances desensitization of the AChR complex by increasing receptor affinity for agonists.
ESTHER : Souccar_1984_Mol.Pharmacol_25_395
PubMedSearch : Souccar_1984_Mol.Pharmacol_25_395
PubMedID: 6328265

Title : Influence of phospholipase C on muscarinic acetylcholine receptor binding in rat brain - Parthasarathy_1984_Neurochem.Res_9_709
Author(s) : Parthasarathy N , Pickard C , Aronstam RS
Ref : Neurochem Res , 9 :709 , 1984
Abstract : Treatment of neural membranes from rat cerebral cortex with phospholipase C (phosphatidylcholine cholinephosphohydrolase) inhibited the binding of radiolabelled antagonists to muscarinic acetylcholine receptors. This inhibition was incomplete, was not competitive, and did not appear to be related to the production of inhibitory products. The affinity of carbamylcholine for cortex muscarinic receptors was increased by phospholipase C action. The distribution of receptors between states of high and low affinity was not affected by phospholipase C; rather, the affinity for carbamylcholine of the lowest affinity receptors was selectively increased. This suggests that membrane lipids influence the interaction of the receptor binding subunit with other structures in the synaptic membrane.
ESTHER : Parthasarathy_1984_Neurochem.Res_9_709
PubMedSearch : Parthasarathy_1984_Neurochem.Res_9_709
PubMedID: 6472570

Title : Guanine nucleotide sensitivity of muscarinic acetylcholine receptors from rat brainstem is eliminated by endogenous proteolytic activity - Aronstam_1984_Neurosci.Lett_47_131
Author(s) : Aronstam RS , Greenbaum LM
Ref : Neuroscience Letters , 47 :131 , 1984
Abstract : The sensitivity to guanine nucleotides of agonist binding to muscarinic acetylcholine receptors was eliminated by incubating rat brainstem membranes at 37 degrees C for 30 min. Pretreatment with any of a variety of proteinase inhibitors prevented this loss of sensitivity. In contrast to other treatments which inactivate guanine nucleotide regulatory mechanisms of muscarinic receptors, incubation at 37 degrees C did not alter agonist binding measured in the absence of guanine nucleotides. Endogenous proteolytic activity appears to inactivate the nucleotide regulatory subunit without engendering its dissociation from the receptor binding subunit.
ESTHER : Aronstam_1984_Neurosci.Lett_47_131
PubMedSearch : Aronstam_1984_Neurosci.Lett_47_131
PubMedID: 6379516

Title : Electrophysiological and biochemical studies on enhancement of desensitization by phenothiazine neuroleptics - Carp_1983_Proc.Natl.Acad.Sci.U.S.A_80_310
Author(s) : Carp JS , Aronstam RS , Witkop B , Albuquerque EX
Ref : Proc Natl Acad Sci U S A , 80 :310 , 1983
Abstract : The actions of the phenothiazines chlorpromazine, prochlorperazine, and trifluoperazine were studied on the acetylcholine receptor-ionic channel complex of frog and rat skeletal muscle and of Torpedo californica to determine their role in pharmacological desensitization and their interactions with different states of the receptor-ionic channel complex. The phenothiazines depressed the peak amplitude of spontaneous and evoked endplate currents while having negligible effect on the decay time constants. Mean channel lifetime and single channel conductance were not altered by these drugs. They also produced a frequency-dependent depression of the peak amplitude of endplate potentials evoked by repetitive microiontophoresis at the extrajunctional region. In addition, these drugs enhanced the ability of carbamoylcholine to displace 125I-labeled alpha-bungarotoxin from receptor-rich membrane preparations of T. californica when used in concentrations that had no effect on 125I-labeled alpha-bungarotoxin binding alone (10 microM). Similarly, the phenothiazines inhibited the binding of tritiated ionic channel ligands, such as phencyclidine and perhydrohistrionicotoxin, a process also enhanced by the presence of carbamoylcholine. These data suggest that the phenothiazines augment agonist-induced desensitization primarily by interacting with the receptor-ionic channel complex prior to channel opening.
ESTHER : Carp_1983_Proc.Natl.Acad.Sci.U.S.A_80_310
PubMedSearch : Carp_1983_Proc.Natl.Acad.Sci.U.S.A_80_310
PubMedID: 6130531

Title : Atropine-induced alterations of normal development of muscarinic receptors in the embryonic chick heart - Kirby_1983_J.Mol.Cell.Cardiol_15_685
Author(s) : Kirby ML , Aronstam RS
Ref : Journal of Molecular & Cellular Cardiology , 15 :685 , 1983
Abstract : The development of muscarinic acetylcholine receptors in chick heart was studied from incubation days 5 through 20. There is a parallel increase in receptor density in atrium and ventricle until the last half of incubation, when the atrial, but not the ventricular, receptor density increases. This increase is blocked by exposure to atropine on incubation days 11 through 14, but not on days 16 through 19. This specific regional increase is coincident with the appearance of functional cholinergic innervation of the heart. During this same period there is an alteration in muscarinic receptor binding properties in both atrium and ventricle that is characterized by an increase in the proportion of receptors displaying high affinity agonist binding. This increase is blocked in the atrium, but not the ventricle, by atropine exposure on incubation days 11 through 14. Thus, there is a critical period in the development of atrial muscarinic receptors during which the receptors are susceptible to modulation by exposure to an antagonist.
ESTHER : Kirby_1983_J.Mol.Cell.Cardiol_15_685
PubMedSearch : Kirby_1983_J.Mol.Cell.Cardiol_15_685
PubMedID: 6644827

Title : Calcium uptake by intestinal smooth muscle: influence of alkylation -
Author(s) : Carrier GO , Marshall DC , Aronstam RS
Ref : European Journal of Pharmacology , 89 :173 , 1983
PubMedID: 6861885

Title : Cholinergic actions of false neurotransmitters: acetyldiethylcholine - Aronstam_1982_Neurosci.Lett_28_51
Author(s) : Aronstam RS , Buccafusco JJ
Ref : Neuroscience Letters , 28 :51 , 1982
Abstract : Acetyldiethylcholine (AcDECh), a false transmitter at cholinergic synapses, binds to central muscarinic receptors with 14-fold lower affinity than acetylcholine (AcCh). The properties of this binding, including the limited extent of regional specificity and binding activation by N-ethylmaleimide, are those associated with weak agonists and antagonists. Intracerebroventricular injection of AcDECh or AcCh produces an increase in arterial blood pressure which is blocked by prior administration of atropine (10 nmol). While AcDECh is 17-fold less potent tha AcCh in producing the pressor response, the maximum pressure changes and time courses are comparable. AcDECh also binds to nicotinic receptors of Torpedo electric organs thereby inducing conformational changes in the receptor-ion channel complex that are associated with postsynaptic activation. In these nicotinic actions, however, AcDECh is 320-fold less active than AcCh.
ESTHER : Aronstam_1982_Neurosci.Lett_28_51
PubMedSearch : Aronstam_1982_Neurosci.Lett_28_51
PubMedID: 6278372

Title : Ketamine inhibition of ligand binding to cholinergic receptors and ion channels - Aronstam_1982_Eur.J.Pharmacol_78_367
Author(s) : Aronstam RS , Narayanan L , Wenger DA
Ref : European Journal of Pharmacology , 78 :367 , 1982
Abstract : Ketamine inhibited the binding of [3H]perhydrohistrionicotoxin and [3H]phencyclidine to the ion channel associated with Torpedo acetylcholine receptors with ID50 values between 11 and 84 micro M, but had not effect on [3H]acetylcholine or [3H]d-tubocurarine binding to the receptor itself. Ketamine's affinity for the ion channel was increased 3-5 fold by receptor agonists. Ketamine also inhibited muscarinic cholinergic receptors with ID50 between 28 and 38 micro M. These results are consistent with biophysical evidence that ketamine interferes with neuromuscular transmission through blockade of the ion channel.
ESTHER : Aronstam_1982_Eur.J.Pharmacol_78_367
PubMedSearch : Aronstam_1982_Eur.J.Pharmacol_78_367
PubMedID: 6279413

Title : Influence of N-ethylmaleimide on cholinoceptors and responses in longitudinal muscles from guinea-pig ileum - Aronstam_1982_Br.J.Pharmacol_77_89
Author(s) : Aronstam RS , Carrier GO
Ref : British Journal of Pharmacology , 77 :89 , 1982
Abstract : 1 The binding of carbamylcholine to membranes prepared from the longitudinal muscle of guinea-pig ileum was determined from its inhibition of the binding of [3H]-3-quinuclidinyl benzilate. Carbamylcholine binding was resolved into high and low affinity components with apparent dissociation constants of 0.11 +/- 0.02 and 11 +/- 1 microM; 42% of the receptors displayed high affinity carbamylcholine binding. 2 Alkylation of longitudinal muscle membranes with N-ethylmaleimide increased muscarinic receptor affinity for carbamylcholine in a manner consistent with a conversion of low affinity to high affinity receptors. After exposure the muscle membrane fragments to 1 mM N-ethylmaleimide for 20 min at 35 degrees C, carbamylcholine binding was resolved into two components with apparent dissociation constants of 0.11 +/- 0.01 and 9 +/- 2 microM, with 74% of the receptors displaying the higher affinity. 3 Exposure of longitudinal membranes mounted in an organ chamber to 1 mM N-ethylmaleimide for 30s depressed isometric contractions in response to acetylcholine by 80%, while contractions induced by K+ and Ba2+ were reduced by less than 20% and 10%, respectively. Acetylcholine dose-response curves were shifted to the right while Ba2+ curves were unaffected. 4 It is suggested that N-ethylmaleimide has a selective effect on muscarinic responses in the longitudinal muscle by disrupting processes occurring after receptor occupancy but before the induction of phospholipid turnover or calcium influx in the postsynaptic membrane.
ESTHER : Aronstam_1982_Br.J.Pharmacol_77_89
PubMedSearch : Aronstam_1982_Br.J.Pharmacol_77_89
PubMedID: 7126999

Title : Anatoxin-a interactions with cholinergic synaptic molecules - Aronstam_1981_Proc.Natl.Acad.Sci.U.S.A_78_4639
Author(s) : Aronstam RS , Witkop B
Ref : Proc Natl Acad Sci U S A , 78 :4639 , 1981
Abstract : Anatoxin-a, a bicyclic amine isolated from blue-green alga, binds to the nicotinic acetylcholine receptor of Torpedo electric tissue, thereby inducing conformational changes in the postsynaptic receptor--ion channel complex as evidenced by alterations in the binding of radiolabeled ligands to the complex. Anatoxin-a binds to the acetylcholine recognition site (Kd = 0.1--0.2 microM) as indicated by its competitive inhibition of specific [3H]acetylcholine and d-[3H]tubocurarine binding, Anatoxin-a stimulates the binding of three physiologically identified "ion channel blockers," [3H]perhydrohistrionicotoxin, [3H]phencyclidine, and [3H]phencyclidine methiodide. The 50% effective doses for these effects range from 0.14 to 0.28 microM. Incubation of Torpedo membranes with anatoxin-a before addition of a radiolabeled channel probe produces a time- and concentration-dependent attenuation of the binding compared to the situation in which anatoxin-a and the probe are added simultaneously. The time course for the elaboration of this decrease corresponds to electrophysiological measurements of anatoxin-a-induced desensitization of neuromuscular junction responses. In these nicotinic actions, anatoxin-a is about as potent as acetylcholine. Anatoxin-a has relatively low affinity for the muscarinic acetylcholine receptors of rat brain, inhibiting 3-[3H]quinuclidinyl benzilate binding (10(-10) M) by 50% at concentrations between 10 and 20 microM. In contrast to classical muscarinic agonists, anatoxin-a displays little regional selectivity in its binding, and its receptor affinity is unaltered by alkylation of the neural membranes with N-ethylmaleimide.
ESTHER : Aronstam_1981_Proc.Natl.Acad.Sci.U.S.A_78_4639
PubMedSearch : Aronstam_1981_Proc.Natl.Acad.Sci.U.S.A_78_4639
PubMedID: 6270690

Title : Cholinergic actions of false neurotransmitters: acetylpyrrolidinecholine - Buccafusco_1981_Neurosci.Lett_23_319
Author(s) : Buccafusco JJ , Aronstam RS
Ref : Neuroscience Letters , 23 :319 , 1981
Abstract : Acetylpyrrolidinecholine (AcPyCh), a false transmitter at cholinergic synapses, produces an increase in arterial blood pressure upon intraventricular injection that is comparable in extent and time course to that produced by equivalent doses of acetylcholine. AcPyCh binds to central muscarinic receptors with affinities and properties associated with potent muscarinic agonists. AcPyCh also binds to nicotinic receptors of Torpedo electric organs thereby inducing conformational changes in the receptor-ion channel complex, including certain changes associated with postsynaptic activation and desensitization processes. In all of these cholinergic actions AcPyCh is as potent as acetylcholine.
ESTHER : Buccafusco_1981_Neurosci.Lett_23_319
PubMedSearch : Buccafusco_1981_Neurosci.Lett_23_319
PubMedID: 7266932

Title : Interactions of tricyclic antidepressants with a synaptic ion channel -
Author(s) : Aronstam RS
Ref : Life Sciences , 28 :59 , 1981
PubMedID: 6261057

Title : Regulation of [3H]perhydrohistrionicotoxin binding to Torpedo ocellata electroplax by effectors of the acetylcholine receptor - Aronstam_1981_J.Biol.Chem_256_2843
Author(s) : Aronstam RS , Eldefrawi AT , Pessah IN , Daly JW , Albuquerque EX , Eldefrawi ME
Ref : Journal of Biological Chemistry , 256 :2843 , 1981
Abstract : The specific binding of [3H]perhydrohistrionicotoxin ([3H]H12-HTX) to the ionic channel of the nicotinic receptor in membranes from the electric organ of the electric ray Torpedo ocellata was studied by use of a rapid filter assay. The time course of the binding was monitored and the initial rate of binding (i.e. within 30 s) was found to be increased up to several hundredfold by the presence of several receptor agonists in a dose-dependent manner. Receptor antagonists blocked this agonist-increased initial rate of binding. The presence of receptor antagonists, with the exception of alpha-bungarotoxin, also increased the initial rate of [3H]H12-HTX binding, though to a much lesser degree than agonists. Preincubation of the membranes with carbamylcholine reduced the initial rate of binding in a time-dependent manner. This was proposed to be due to receptor desensitization. Thus, it is suggested that the time course of [3H]H12-HTX binding to the ionic channel sites is influenced by the conformational state of the receptor-channel complex such that receptor activation, inactivation, and desensitization appear to be reflected in [3H]H12-HTX binding.
ESTHER : Aronstam_1981_J.Biol.Chem_256_2843
PubMedSearch : Aronstam_1981_J.Biol.Chem_256_2843
PubMedID: 7204378

Title : 3-Chlorodibenzazepine binding to cholinergic receptors and ion channels - Aronstam_1981_Res.Commun.Chem.Pathol.Pharmacol_34_551
Author(s) : Aronstam RS , Narayanan L
Ref : Res Commun Chem Pathol Pharmacol , 34 :551 , 1981
Abstract : The interactions of 3-chlorodihydrodibenzazepines with nicotinic receptors and ion channels from Torpedo californica electric organ and muscarinic receptors from rat brain were revealed by their inhibition of the binding of specific radiolabelled probes. Desmethylchlorimipramine (DCI) was twice as active as chlorimipramine (CI) in inhibiting [3H]phencyclidine and [3H]perhydrohistrionicotoxin binding to nicotinic ion channels; inhibition constants were 0.13 and 0.28 micro M in the presence and 1.2 and 3.4 micro M in the absence of carbamylcholine. Both of the drugs had marginal affinity for the nicotinic receptor (i.e., the [3H]3-quinuclidinyl benzilate binding to muscarinic receptors from rat brain stem and cerebral cortex with inhibitions constants below 0.1 micro M. Little regional specificity or activation by N-ethylmaleimide was apparent in the muscarinic binding of CI and DCI, properties associated with the binding of receptor antagonists.
ESTHER : Aronstam_1981_Res.Commun.Chem.Pathol.Pharmacol_34_551
PubMedSearch : Aronstam_1981_Res.Commun.Chem.Pathol.Pharmacol_34_551
PubMedID: 6275473

Title : Phencyclidine interactions with the ionic channel of the acetylcholine receptor and electrogenic membrane - Albuquerque_1980_Proc.Natl.Acad.Sci.U.S.A_77_1224
Author(s) : Albuquerque EX , Tsai MC , Aronstam RS , Witkop B , Eldefrawi AT , Eldefrawi ME
Ref : Proc Natl Acad Sci U S A , 77 :1224 , 1980
Abstract : The effects of phencyclidine (PCP) were studied on the electrogenic and chemosensitive properties of the neuromuscular junction of skeletal muscle as well as on the binding sites on the acetylcholine (AcCho) receptor and its ionic channel in the electric organ membranes of the electric ray. The directly elicited muscle twitch was markedly potentiated by prolonging the falling phase of the muscle action potential and blocking delayed rectification. The indirectly elicited muscle twitch was transiently potentiated and then blocked by PCP at concentrations below 60 muM. PCP blocked miniature endplate potentials and AcCho sensitivities at the junctional region of innervated muscle, blocked the extrajunctional sensitivity of the chronically denervated muscle, and significantly depressed the peak amplitude of the endplate current (EPC) in a voltage- and time-dependent manner. PCP also caused acceleration of the time course of EPC decay and shortening of the mean life-time of the open ionic channel. The effects of PCP were not due to inhibition of AcCho receptor sites because PCP did not protect against the quasi-irreversible inhibition of receptor sites by alpha-bungarotoxin, nor did it inhibit binding of [(3)H]AcCho or [(125)I-labeled alpha-bungarotoxin to the receptor sites. On the other hand, PCP blocked the binding of [(3)H]perhydrohistrionicotoxin to the sites of the ionic channel of the AcCho receptor. The data suggest that PCP reacts with the electrogenic K(+) channel and the ionic channel associated with the AcCho receptor in the open as well as the closed conformation.
ESTHER : Albuquerque_1980_Proc.Natl.Acad.Sci.U.S.A_77_1224
PubMedSearch : Albuquerque_1980_Proc.Natl.Acad.Sci.U.S.A_77_1224
PubMedID: 6928673

Title : Activation, inactivation, and desensitization of acetylcholine receptor channel complex detected by binding of perhydrohistrionicotoxin - Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_2309
Author(s) : Eldefrawi ME , Aronstam RS , Bakry NM , Eldefrawi AT , Albuquerque EX
Ref : Proc Natl Acad Sci U S A , 77 :2309 , 1980
Abstract : The effects of receptor activation were studied on the interaction of perhydrohistrionicotoxin (H(12)-HTX) with the ionic channel of the nicotinic acetylcholine (AcCho) receptor in membranes from the electric organ of Torpedo ocellata and with the endplate region of the soleus muscle of the rat. In Torpedo membranes, the initial rate (i.e., within 30 sec) of [(3)H]H(12)-HTX bindings to the ionic channel of the AcCho receptor was accelerated 10(2)- to 10(3)-fold in the presence of carbamoylcholine (Carb). H(12)-HTX also inhibited Carb-activated (22)Na(+) influx, over 95% inhibition at 10 muM H(12)-HTX. At this concentration H(12)-HTX did not inhibit [(3)H]AcCho binding to the AcCho-receptor sites. There was good correspondence between the degree of acceleration of [(3)H]H(12)-HTX binding and the stimulation of (22)Na(+) influx over a wide range of Carb concentrations (up to 100 muM). Preincubation of Torpedo membranes with Carb decreased the initial rate of [(3)H]H(12)-HTX binding, as well as the rate of (22)Na(+) influx, which may reflect desensitization of the AcCho-receptor. d-Tubocurarine inhibited the agonist-mediated acceleration of [(3)H]H(12)-HTX binding and (22)Na(+) influx. In the soleus muscle endplate, H(12)-HTX inhibited the transient depolarization induced by microiontophoretic application of AcCho; the more receptors activated and channels opened, the stronger was the inhibition by H(12)-HTX. These findings suggest that H(12)-HTX binds to closed and open ionic channels, with a preference for the latter conformation. It is also suggested that the conformational changes associated with activation or desensitization of the receptor can be monitored by studying binding of [(3)H]H(12)-HTX to the ionic channel sites as well as by the AcCho-receptor-regulated (22)Na(+) influx.
ESTHER : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_2309
PubMedSearch : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_2309
PubMedID: 6246539

Title : Nature of regional and chemically-induced differences in the binding properties of muscarinic acetylcholine receptors from rat brain -
Author(s) : Ikeda SR , Aronstam RS , Eldefrawi ME
Ref : Neuropharmacology , 19 :575 , 1980
PubMedID: 7402447

Title : Sites of action of phencyclidine. II. Interaction with the ionic channel of the nicotinic receptor -
Author(s) : Albuquerque EX , Tsai MC , Aronstam RS , Eldefrawi AT , Eldefrawi ME
Ref : Molecular Pharmacology , 18 :167 , 1980
PubMedID: 6252436

Title : Similarities in the binding sites of the muscarinic receptor and the ionic channel of the nicotinic receptor -
Author(s) : Aronstam RS , Eldefrawi AT , Eldefrawi ME
Ref : Biochemical Pharmacology , 29 :1311 , 1980
PubMedID: 6249329

Title : Sites of action of phencyclidine. III. Interactions with muscarinic receptors -
Author(s) : Aronstam RS , Eldefrawi ME , Eldefrawi AT , Albuquerque EX , Jim KF , Triggle DJ
Ref : Molecular Pharmacology , 18 :179 , 1980
PubMedID: 7421791

Title : Sites of action of phencyclidine. I. Effects on the electrical excitability and chemosensitive properties of the neuromuscular junction of skeletal muscle -
Author(s) : Tsai MC , Albuquerque EX , Aronstam RS , Eldefrawi AT , Eldefrawi ME , Triggle DJ
Ref : Molecular Pharmacology , 18 :159 , 1980
PubMedID: 6968397

Title : [3H]Phencyclidine: a probe for the ionic channel of the nicotinic receptor - Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_7458
Author(s) : Eldefrawi ME , Eldefrawi AT , Aronstam RS , Maleque MA , Warnick JE , Albuquerque EX
Ref : Proc Natl Acad Sci U S A , 77 :7458 , 1980
Abstract : To evaluate [3H]phencyclidine ([3H]PCP)as a probe for the ionic channel of the nicotinic receptor, the characteristics of its binding to electric organ membranes od Torpedo ocellata and its effects on frog sartorius muscle were studied. Similar to PCP, [3H]PCP depressed the peak amplitude of endplate current, caused nonlinearity in the voltage-current relationship at negative potentials, accelerated the decay time of the end-plate current, and shortened the channel lifetime. Thus, [3H]PCP interacted with the ionic channel of the nicotinic receptor, although there were a few differences between its effect and that of PCP. Binding of [3H]PCP to Torpedo membranes was to sites on the ionic channel of acetylcholine (AcCho) receptor because it was saturable, dependent upon protein concentration, and inhibited by drugs that interact with the ionic channel, and the initial rate of binding was potentiated by receptor agonists. Equilibrium binding of [3H]PCP to Torpedo membranes was with two affinities, but in the presence of AcCho, [3H]PCP binding was with a single affinity. The affinities of channel drugs obtained by inhibition of binding of [3H]PCP and [3H[perhydrohistrionicotoxin to Torpedo membranes were different, with correlation coefficients of 0.52 and 0.82 in the absence and presence of a receptor agonist, respectively; this suggests differences in their binding sites on the ionic channel of the AcCho receptor.
ESTHER : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_7458
PubMedSearch : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_7458
PubMedID: 6261260

Title : Reversible conversion between affinity states for agonists of the muscarinic acetylcholine receptor from rat brain -
Author(s) : Aronstam RS , Eldefrawi ME
Ref : Biochemical Pharmacology , 28 :701 , 1979
PubMedID: 444257

Title : Transition and heavy metal inhibition of ligand binding to muscarinic acetylcholine receptors from rat brain -
Author(s) : Aronstam RS , Elderfrawi ME
Ref : Toxicol Appl Pharmacol , 48 :489 , 1979
PubMedID: 473192

Title : Development of muscarinic cholinergic receptors in inbred strains of mice: identification of receptor heterogeneity and relation to audiogenic seizure susceptibility - Aronstam_1979_Brain.Res_162_231
Author(s) : Aronstam RS , Kellogg C , Abood LG
Ref : Brain Research , 162 :231 , 1979
Abstract : The concentrations and biochemical properties of muscarinic acetylcholine receptors in the brains of two highly inbred strains of mice, DBA/2J and C57BL/6J, have been studied using [3H]quinuclidinyl benzilate (QNB), a potent and specific receptor antagonist. As is the case with rat brain, murine muscarinic receptors exist in at least two forms, which differ in their affinities for receptor agonists but which have the same high affinity for receptor antagonists. Carbamylcholine binding to mouse neural membranes can be resolved into two components with KDs of 5.2 times 10(-7) and 7.9 times 10(-5) M. There is a regional heterogeneity of brain receptors with respect to their distribution between these high and low agonist affinity forms. Brain stem and hypothalamus receptors display binding properties that would be expected if over 60% of their receptors were in the high affinity state, while only 30-40% of cortex, striatum and thalamus receptors appear to be in the high affinity form. Hippocampal receptors display the least amount of high agonist affinity character. Saturation curves and Scatchard plots of QNB binding at 2, 14, 21 and 42 days postnatal age in both strains indicate no differences or changes in the affinity or nature of the binding with age. Significant increases in QNB binding per mg membrane protein were observed between 14 and 42 days in the cortex, hippocampus, striatum, thalamus and hypothalamus, but not in the midbrain-pons-medulla region. In the hippocampus the DBA mice had significantly more QNB binding. In the hypothalamus decreases with age in total binding were noted in DBA, while slight increases were noted in C57. Compared to C57, hippocampal receptors in DBA displayed lower agonist affinity at 14 and 21 days, a trait which was not apparent when DBA had outgrown their audiogenic seizure sensitivity at 42 days. The differences in receptor density and agonist state distribution between the two strains may be related to audiogenic seizure sensitivity.
ESTHER : Aronstam_1979_Brain.Res_162_231
PubMedSearch : Aronstam_1979_Brain.Res_162_231
PubMedID: 761088

Title : Structural and stereochemical requirements for muscarinic receptor binding -
Author(s) : Aronstam RS , Triggle DJ , Eldefrawi ME
Ref : Molecular Pharmacology , 15 :227 , 1979
PubMedID: 470929

Title : Influence of sulfhydryl reagents and heavy metals on the functional state of the muscarinic acetylcholine receptor in rat brain -
Author(s) : Aronstam RS , Abood LG , Hoss WP
Ref : Molecular Pharmacology , 14 :575 , 1978
PubMedID: 683175

Title : Solubilization of muscarinic acetylcholine receptors of bovine brain -
Author(s) : Aronstam RS , Schuessler DC, Jr. , Eldefrawi ME
Ref : Life Sciences , 23 :1377 , 1978
PubMedID: 723431

Title : Role of phospholipids in muscarinic binding by neural membranes -
Author(s) : Aronstam RS , Abood LG , Baumgold J
Ref : Biochemical Pharmacology , 26 :1689 , 1977
PubMedID: 20109

Title : Muscarinic cholinergic binding in human erythrocyte membranes -
Author(s) : Aronstam RS , Abood LG , MacNeil MK
Ref : Life Sciences , 20 :1175 , 1977
PubMedID: 850469

Title : Conversion between configurational states of the muscarinic receptor in rat brain - Aronstam_1977_Eur.J.Pharmacol_46_279
Author(s) : Aronstam RS , Hoss WP , Abood LG
Ref : European Journal of Pharmacology , 46 :279 , 1977
Abstract : Reductive alkylation of neural membranes by N-ethyl maleimide (NEM) converts muscarinic acetylcholine receptors from a state of low to high affinity for receptor agonists. Interactions of muscarinic antagonists with the receptor are unaffected by this treatment. Muscarinic receptors from the rat telencephalon in the high agonist affinity state are increased from 34.2 to 53.4% of the total receptor population and the Ki for carbamylcholine inhibition of 3-quinuclidinyl benzilate binding is decreased from 1.2 X 10(-4) to 6.9 X 10(-5) M by NEM treatment.
ESTHER : Aronstam_1977_Eur.J.Pharmacol_46_279
PubMedSearch : Aronstam_1977_Eur.J.Pharmacol_46_279
PubMedID: 590337