Kirkpatrick CJ


Full name : Kirkpatrick Charles James

First name : Charles James

Mail : Institute of Pathology, University Medical Center, Johannes Gutenberg University Mainz, D-55101 Mainz

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Country : Germany

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References (38)

Title : Non-neuronal acetylcholine involved in reproduction in mammals and honeybees - Wessler_2017_J.Neurochem_142 Suppl 2_144
Author(s) : Wessler I , Kirkpatrick CJ
Ref : Journal of Neurochemistry , 142 Suppl 2 :144 , 2017
Abstract : Bacteria and archaea synthesize acetylcholine (ACh). Thus, it can be postulated that ACh was created by nature roughly three billion years ago. Therefore, the wide expression of ACh in nature (i.e., in bacteria, archaea, unicellular organisms, plants, fungi, non-vertebrates and vertebrates and in the abundance of non-neuronal cells of mammals) is not surprising. The term non-neuronal ACh and non-neuronal cholinergic system have been introduced to describe the auto- and paracrine, that is, local regulatory actions of ACh in cells not innervated by neuronal cholinergic fibers and to communicate among themselves. In this way non-neuronal ACh binds to the nicotinic or muscarinic receptors expressed on these local and migrating cells and modulates basic cells functions such as proliferation, differentiation, migration and the transport of ions and water. The present article is focused to the effects of non-neuronal ACh linked to reproduction; data on the expression and function of the non-neuronal cholinergic system in the following topics are summarized: (i) Sperm, granulosa cells, oocytes; (ii) Auxiliary systems (ovary, oviduct, placenta); (iii) Embryonic stem cells as first step for reproduction of a new individual after fertilization; (iv) Larval food as an example of reproduction in insects (honeybees) and adverse effects of the neonicotinoids, a class of world-wide applied insecticides. The review article will show that non-neuronal ACh is substantially involved in the regulation of reproduction in mammals and also non-mammals like insects (honeybees). There is a need to learn more about this biological role of ACh. In particular, we have to consider that insecticides like the neonicotinoids, but also carbamates and organophosphorus pesticides, interfere with the non-neuronal cholinergic system thus compromising for example the breeding of honeybees. But it is possible that other species may also be adversely affected as well, a mechanism which may contribute to the observed decline in biodiversity. This is an article for the special issue XVth International Symposium on Cholinergic Mechanisms.
ESTHER : Wessler_2017_J.Neurochem_142 Suppl 2_144
PubMedSearch : Wessler_2017_J.Neurochem_142 Suppl 2_144
PubMedID: 28072454

Title : Honeybees Produce Millimolar Concentrations of Non-Neuronal Acetylcholine for Breeding: Possible Adverse Effects of Neonicotinoids - Wessler_2016_PLoS.One_11_e0156886
Author(s) : Wessler I , Gartner HA , Michel-Schmidt R , Brochhausen C , Schmitz L , Anspach L , Grunewald B , Kirkpatrick CJ
Ref : PLoS ONE , 11 :e0156886 , 2016
Abstract : The worldwide use of neonicotinoid pesticides has caused concern on account of their involvement in the decline of bee populations, which are key pollinators in most ecosystems. Here we describe a role of non-neuronal acetylcholine (ACh) for breeding of Apis mellifera carnica and a so far unknown effect of neonicotinoids on non-target insects. Royal jelly or larval food are produced by the hypopharyngeal gland of nursing bees and contain unusually high ACh concentrations (4-8 mM). ACh is extremely well conserved in royal jelly or brood food because of the acidic pH of 4.0. This condition protects ACh from degradation thus ensuring delivery of intact ACh to larvae. Raising the pH to >/=5.5 and applying cholinesterase reduced the content of ACh substantially (by 75-90%) in larval food. When this manipulated brood was tested in artificial larval breeding experiments, the survival rate was higher with food supplemented by 100% with ACh (6 mM) than with food not supplemented with ACh. ACh release from the hypopharyngeal gland and its content in brood food declined by 80%, when honeybee colonies were exposed for 4 weeks to high concentrations of the neonicotinoids clothianidin (100 parts per billion [ppb]) or thiacloprid (8,800 ppb). Under these conditions the secretory cells of the gland were markedly damaged and brood development was severely compromised. Even field-relevant low concentrations of thiacloprid (200 ppb) or clothianidin (1 and 10 ppb) reduced ACh level in the brood food and showed initial adverse effects on brood development. Our findings indicate a hitherto unknown target of neonicotinoids to induce adverse effects on non-neuronal ACh which should be considered when re-assessing the environmental risks of these compounds. To our knowledge this is a new biological mechanism, and we suggest that, in addition to their well documented neurotoxic effects, neonicotinoids may contribute to honeybee colony losses consecutive to a reduction of the ACh content in the brood food.
ESTHER : Wessler_2016_PLoS.One_11_e0156886
PubMedSearch : Wessler_2016_PLoS.One_11_e0156886
PubMedID: 27285384

Title : Recent progress in revealing the biological and medical significance of the non-neuronal cholinergic system - Grando_2015_Int.Immunopharmacol_29(1)_1
Author(s) : Grando SA , Kawashima K , Kirkpatrick CJ , Kummer W , Wessler I
Ref : Int Immunopharmacol , 29 :1 , 2015
Abstract : This special issue of International Immunopharmacology is the proceedings of the Fourth International Symposium on Non-neuronal Acetylcholine that was held on August 28-30, 2014 at the Justus Liebig University of Giessen in Germany. It contains original contributions of meeting participants covering the significant progress in understanding of the biological and medical significance of the non-neuronal cholinergic system extending from exciting insights into molecular mechanisms regulating this system via miRNAs over the discovery of novel cholinergic cellular signaling circuitries to clinical implications in cancer, wound healing, immunity and inflammation, cardiovascular, respiratory and other diseases.
ESTHER : Grando_2015_Int.Immunopharmacol_29(1)_1
PubMedSearch : Grando_2015_Int.Immunopharmacol_29(1)_1
PubMedID: 26362206

Title : Murine embryonic stem cell line CGR8 expresses all subtypes of muscarinic receptors and multiple nicotinic receptor subunits: Down-regulation of alpha4- and beta4-subunits during early differentiation - Kaltwasser_2015_Int.Immunopharmacol_29(1)_110
Author(s) : Kaltwasser S , Schmitz L , Michel-Schmidt R , Anspach L , Kirkpatrick CJ , Wessler I
Ref : Int Immunopharmacol , 29 :110 , 2015
Abstract : Non-neuronal acetylcholine mediates its cellular effects via stimulation of the G-protein-coupled muscarinic receptors and the ligand-gated ion channel nicotinic receptors. The murine embryonic stem cell line CGR8 synthesizes and releases non-neuronal acetylcholine. In the present study a systematic investigation of the expression of nicotinic receptor subunits and muscarinic receptors was performed, when the stem cells were grown in the presence or absence of LIF, as the latter condition induces early differentiation. CGR8 cells expressed multiple nicotinic receptor subtypes (alpha3, alpha4, alpha7, alpha9, alpha10, beta1, beta2, beta3, beta4, gamma, delta, epsilon) and muscarinic receptors (M1, M3, M4, M5); M2 was detected only in 2 out of 8 cultures. LIF removal caused a down-regulation only of the alpha4- and beta4-subunit. In conclusion, more or less the whole repertoire of cholinergic receptors is expressed on the murine embryonic stem cell line CGR8 for mediating cellular signaling of non-neuronal acetylcholine which acts via auto- and paracrine pathways. During early differentiation of the murine CGR8 stem cell signaling via nicotinic receptors containing alpha4- or beta4 subunits is reduced. Thus, the so-called neuronal alpha4 nicotine receptor composed of these subunits may be involved in the regulation of pluripotency in this murine stem cell line.
ESTHER : Kaltwasser_2015_Int.Immunopharmacol_29(1)_110
PubMedSearch : Kaltwasser_2015_Int.Immunopharmacol_29(1)_110
PubMedID: 26299974

Title : Effect of LIF-withdrawal on acetylcholine synthesis in the embryonic stem cell line CGR8 is not mediated by STAT3, PI3Ks or cAMP\/PKA pathways - Michel-Schmidt_2015_Int.Immunopharmacol_29(1)_115
Author(s) : Michel-Schmidt R , Kirkpatrick CJ , Wessler I
Ref : Int Immunopharmacol , 29 :115 , 2015
Abstract : Acetylcholine (ACh) acts as a local cellular signaling molecule and is widely expressed in nature, including mammalian cells and embryonic stem cells. The murine embryonic stem cell line CGR8 synthesizes and releases substantial amounts of ACh. Particularly during early differentiation - a period associated with multiple alterations in geno-/phenotype functions - synthesis and release of ACh are increased by 10-fold. In murine stem cells second messengers of the STAT-3, PI3K and cAMP/PKA pathways are involved in maintaining self-renewal and pluripotency. The present experiments were designed to test whether blockers of these signaling pathways enhance ACh cell content in the presence of LIF, i.e. when CGR8 is pluripotent. NSC74859, an inhibitor of STAT-3, affected neither the proliferation rate nor ACh cell content, whereas the more sensitive STAT-3 inhibitor FLLL31 reduced the proliferation rate and increased ACh cell content by about 3-fold. The PI3K inhibitor LY294002 reduced the proliferation rate but did not modify the ACh cell content, whereas the PKA inhibitor H89 produced effects comparable to FLLL31. Interestingly, in control experiments a strong inverse correlation was found between cell density and ACh cell content, which could explain the 3-fold increase in the ACh cell content observed in the presence of FLLL31 and H89. Forskolin, a PKA activator, had no effect. In conclusion, it appears unlikely that the 10-fold increase in ACh cell content induced by LIF removal, i.e. during early differentiation, is mediated by second messengers of the STAT-3, PI3K and cAMP/PKA pathways. However, the PI3K pathway appears to be involved in control of the inverse relation between cell density and ACh cell content, because this correlation was significantly attenuated in the presence of LY294002.
ESTHER : Michel-Schmidt_2015_Int.Immunopharmacol_29(1)_115
PubMedSearch : Michel-Schmidt_2015_Int.Immunopharmacol_29(1)_115
PubMedID: 25887270

Title : pH-dependent hydrolysis of acetylcholine: Consequences for non-neuronal acetylcholine - Wessler_2015_Int.Immunopharmacol_29(1)_27
Author(s) : Wessler I , Michel-Schmidt R , Kirkpatrick CJ
Ref : Int Immunopharmacol , 29 :27 , 2015
Abstract : Acetylcholine is inactivated by acetylcholinesterase and butyrylcholinesterase and thereby its cellular signalling is stopped. One distinguishing difference between the neuronal and non-neuronal cholinergic system is the high expression level of the esterase activity within the former and a considerably lower level within the latter system. Thus, any situation which limits the activity of both esterases will affect the non-neuronal cholinergic system to a much greater extent than the neuronal one. Both esterases are pH-dependent with an optimum at pH above 7, whereas at pH values below 6 particularly the specific acetylcholinesterase is more or less inactive. Thus, acetylcholine is prevented from hydrolysis at such low pH values. The pH of the surface of the human skin is around 5 and therefore non-neuronal acetylcholine released from keratinocytes can be detected in a non-invasive manner. Several clinical conditions like metabolic acidosis, inflammation, fracture-related haematomas, cardiac ischemia and malignant tumours are associated with local or systemic pH values below 7. Thus, the present article describes some consequences of an impaired inactivation of extracellular non-neuronal acetylcholine.
ESTHER : Wessler_2015_Int.Immunopharmacol_29(1)_27
PubMedSearch : Wessler_2015_Int.Immunopharmacol_29(1)_27
PubMedID: 25929445

Title : Influence of organophosphate poisoning on human dendritic cells - Schafer_2013_Chem.Biol.Interact_206_472
Author(s) : Schafer M , Koppe F , Stenger B , Brochhausen C , Schmidt A , Steinritz D , Thiermann H , Kirkpatrick CJ , Pohl C
Ref : Chemico-Biological Interactions , 206 :472 , 2013
Abstract : Organophosphourus compounds (OPC, including nerve agents and pesticides) exhibit acute toxicity by inhibition of acetylcholinesterase. Lung affections are frequent complications and a risk factor for death. In addition, epidemiological studies reported immunological alterations after OPC exposure. In our experiments we investigated the effects of organophosphourus pesticides dimethoate and chlorpyrifos on dendritic cells (DC) that are essential for the initial immune response, especially in the pulmonary system. DC, differentiated from the monocyte cell line THP-1 by using various cytokines (IL-4, GM-CSF, TNF-alpha, Ionomycin), were exposed to organophosphourus compounds at different concentrations for a 24h time period. DC were characterized by flow cytometry and immunofluorescence using typical dendritic cell markers (e.g., CD11c, CD209 and CD83). After OPC exposure we investigated cell death, the secretion profile of inflammatory mediators, changes of DC morphology, and the effect on protein kinase signalling pathways. Our results revealed a successful differentiation of THP-1 into DC. OPC exposure caused a significant concentration-dependent influence on DC: Dendrites of the DC were shortened and damaged, DC-specific cell surface markers (i.e., CD83and CD209) decreased dramatically after chlorpyrifos exposure. Interestingly, the effects caused by dimethoate were in general less pronounced. The organophosphourus compounds affected the release of inflammatory cytokines, such as IL-1ss and IL-8. The anti-inflammatory cytokine IL-10 was significantly down regulated. Protein kinases like the Akt family or ERK, which are essential for cell survival and proliferation, were inhibited by both OPC. These findings indicate that the tested organophosphourus compounds induced significant changes in cell morphology, inhibited anti-inflammatory cytokines and influenced important protein signalling pathways which are involved in regulation of apoptosis. Thus our results highlight novel aspects -apparently independent of AChE inhibition- of OPC poisoning with regard to lung toxicity. Our findings contribute to the basic understanding of pulmonary complications caused by OPC poisoning.
ESTHER : Schafer_2013_Chem.Biol.Interact_206_472
PubMedSearch : Schafer_2013_Chem.Biol.Interact_206_472
PubMedID: 23994500

Title : Upregulated acetylcholine synthesis during early differentiation in the embryonic stem cell line CGR8 - Wessler_2013_Neurosci.Lett_547_32
Author(s) : Wessler I , Michel-Schmidt R , Schmidt H , Kaltwasser S , Unger R , Kirkpatrick CJ
Ref : Neuroscience Letters , 547 :32 , 2013
Abstract : Stem cells are used to generate differentiated somatic cells including neuronal cells. Synthesis and release of acetylcholine, a neurotransmitter and widely expressed signaling molecule, were investigated in the murine embryonic stem cell line CGR8 during early differentiation, i.e. in the presence of leukemia inhibitory factor (LIF) to maintain pluripotency and in the absence of LIF to induce early differentiation. CGR8 cells express choline acetyltransferase (ChAT) as demonstrated by measurement of enzyme activity and substantial inhibition by bromoacetylcholine. Pluripotent CGR8 cells showed a ChAT activity of 250 pmol acetylcholine/mg/h, contained 1.1 pmol acetylcholine/10(6) cells and released about 12.00 pmol acetylcholine/1 x 10(6) cells/6 h. Removal of LIF induced early differentiation as evidenced by reduced transcription factors Oct-4 and Nanog and a substantial slowing of the proliferation rate. Under this condition acetylcholine synthesis increased to 1640 pmol/mg/h; related to the pluripotent state the content of acetylcholine increased 10-fold and the release to about 32 pmol acetylcholine/1 x 10(6) cells/6 h. Enzyme kinetic analysis showed a significant increase of the K(m) for the precursor acetyl-CoA and of V(max) without a change of the K(m) for the precursor choline. In conclusion, early differentiation of the stem cell line CGR8 is associated with a substantial increase in ChAT activity and acetylcholine release.
ESTHER : Wessler_2013_Neurosci.Lett_547_32
PubMedSearch : Wessler_2013_Neurosci.Lett_547_32
PubMedID: 23669640

Title : Activation of muscarinic receptors by non-neuronal acetylcholine - Wessler_2012_Handb.Exp.Pharmacol_208_469
Author(s) : Wessler I , Kirkpatrick CJ
Ref : Handbook of Experimental Pharmacology , 208 :469 , 2012
Abstract : The biological role of acetylcholine and the cholinergic system is revisited based particularly on scientific research early and late in the last century. On the one hand, acetylcholine represents the classical neurotransmitter, whereas on the other hand, acetylcholine and the pivotal components of the cholinergic system (high-affinity choline uptake, choline acetyltransferase and its end product acetylcholine, muscarinic and nicotinic receptors and esterase) are expressed by more or less all mammalian cells, i.e. by the majority of cells not innervated by neurons at all. Moreover, it has been demonstrated that acetylcholine and "cholinergic receptors" are expressed in non-neuronal organisms such as plants and protists. Acetylcholine is even synthesized by bacteria and algae representing an extremely old signalling molecule on the evolutionary timescale. The following article summarizes examples, in which non-neuronal acetylcholine is released from primitive organisms as well as from mammalian non-neuronal cells and binds to muscarinic receptors to modulate/regulate phenotypic cell functions via auto-/paracrine pathways. The examples demonstrate that non-neuronal acetylcholine and the non-neuronal cholinergic system are vital for various types of cells such as epithelial, endothelial and immune cells.
ESTHER : Wessler_2012_Handb.Exp.Pharmacol_208_469
PubMedSearch : Wessler_2012_Handb.Exp.Pharmacol_208_469
PubMedID: 22222711

Title : Subcellular distribution of choline acetyltransferase by immunogold electron microscopy in non-neuronal cells: placenta, airways and murine embryonic stem cells - Wessler_2012_Life.Sci_91_977
Author(s) : Wessler I , Michel-Schmidt R , Brochhausen C , Kirkpatrick CJ
Ref : Life Sciences , 91 :977 , 2012
Abstract : AIMS: Acetylcholine is synthesized in more or less all mammalian cells. However, little is known about the subcellular location of acetylcholine synthesis. Therefore, in the present experiments the subcellular location of the synthesizing enzyme choline acetyltransferase (ChAT) was investigated by anti-ChAT immunogold electron microscopy in human placenta and airways as well as in a murine embryonic stem cell line (CGR8 cell line). MAIN
METHODS: Human tissue was obtained as so-called surplus tissue (after delivery/surgical removal because of lung tumor); the CGR8 stem cell line was cultured under standard conditions. For human tissue a monoclonal mouse anti-ChAT antibody (ab) was used and for the CGR8 cell line a polyclonal goat anti-ChAT ab. Immunogold electron microscopy was applied to identify the subcellular location of ChAT. KEY FINDINGS: In trophoblast cells (placenta) specific anti-ChAT immunogold deposition was found within the cell membrane, microvilli, and caveolae but also within the cytosol, for example associated with intermediate filaments. In addition, immunogold deposition was identified within mitochondria and the nuclear membrane. In airway epithelial cells anti-ChAT immunogold was found particularly within the apical cell membrane, cilia, submucosa, cytosol and nuclear membrane. Likewise alveolar macrophages showed positive anti-ChAT immunogold within the nucleus, nuclear membrane and granula. Also in the CGR8 cell line positive anti-ChAT immunogold was identified within the cell nucleus and cytosol. SIGNIFICANCE: The present experiments demonstrate a wide subcellular distribution of ChAT with particular preference of the cell membrane in human epithelial cells.
ESTHER : Wessler_2012_Life.Sci_91_977
PubMedSearch : Wessler_2012_Life.Sci_91_977
PubMedID: 22683430

Title : Release of acetylcholine from murine embryonic stem cells: effect of nicotinic and muscarinic receptors and blockade of organic cation transporter - Wessler_2012_Life.Sci_91_973
Author(s) : Wessler I , Michel-Schmidt R , Dohle E , Kirkpatrick CJ
Ref : Life Sciences , 91 :973 , 2012
Abstract : AIMS: The non-neuronal cholinergic system is widely expressed in nature. The present experiments were performed to characterize the non-neuronal cholinergic system in murine embryonic stem cells (CGR8 cell line). MAIN
METHODS: CGR8 cells were cultured in gelatinized flasks with Glasgow's buffered minimal essential medium (Gibco, Germany). Acetylcholine was measured by HPLC combined with bioreactor and electrochemical detection. KEY FINDINGS: CGR8 cells contained 1.08+/-0.12 pmol acetylcholine/10(6) cells (n=7) which was reduced to 0.50+/-0.06 pmol/10(6) cells (n=6; p<0.05) in the presence (4h) of 30muM bromoacetylcholine to block choline acetyltransferase. A time-dependent release of acetylcholine into the incubation medium was demonstrated, when cholinesterase activity was blocked by 10 muM physostigmine, with 97+/-13, 180+/-15 and 216+/-14 pmol being released from 65x10(6) cells after incubation periods of 2, 4 and 6h, respectively. The cumulative release corresponds to a fractional release rate of 2%/min. Blockade of nicotine or muscarine receptors did not significantly modulate the release of acetylcholine which was substantially reduced by 300 muM quinine (inhibitor of organic cation transporters). This inhibition showed considerable fading over the incubation period, indicating additional release mechanisms activated upon inhibition of organic cation transporters. SIGNIFICANCE: Murine embryonic stem cells contain and release significant amounts of acetylcholine. The high fractional release rate and the compensation for blocked organic cation transporters indicate that non-neuronal acetylcholine may play a functional role in the homeostasis of murine embryonic stem cells.
ESTHER : Wessler_2012_Life.Sci_91_973
PubMedSearch : Wessler_2012_Life.Sci_91_973
PubMedID: 22569291

Title : Blockade of nicotinic and muscarinic receptors facilitates spontaneous migration of human peripheral granulocytes: failure in cystic fibrosis - Wessler_2012_Life.Sci_91_1119
Author(s) : Wessler I , Neumann S , Razen M , Zepp F , Kirkpatrick CJ
Ref : Life Sciences , 91 :1119 , 2012
Abstract : AIMS: Circulating leucocytes express muscarinic (m) and nicotinic (n) receptors and synthesize acetylcholine (ACh) regulating various cell functions. Leucocytes from patients with cystic fibrosis contain less ACh; therefore it was tested whether the regulation of cellular functions like migration differed from healthy volunteers. MAIN
METHODS: Peripheral blood (10-20 ml) was used, leucocytes were isolated by Ficoll(R) gradient and the commercial MIGRATEST(R) combined with flow cytometric analysis was applied (pore size 3 mum). KEY FINDINGS: In the absence of test substances 4900+/-1800 (n=10) leucocytes migrated within a time period of 2 h. In the presence of tubocurarine (TC, 30 muM) the cell number increased to 7500+/-2700 [n=10] corresponding to an increase of 162+/-20% (mean of individual experiments; p<0.02). Atropine (1 muM) was not effective (120+/-17%, n=7). Simultaneous application of atropine and TC produced a slightly higher effect than TC alone (185+/-23%; n=8); a 10-fold increase of TC and atropine resulted to a somewhat stronger effect (248+/-39%; n=8). When migration time was reduced to 30 min or the chemoattractant fMLP (0.05 muM) present neither atropine nor TC affected migration. Granulocytes isolated from patients with cystic fibrosis did not respond (2h migration) to 30 muM TC (control: 5180+/-1400 cells [n=10]; TC: 5800+/-1400 [n=10]). Also in the presence of atropine (1 muM) and TC (30 muM) a significant effect was not detected (5800+/-1300 [n=10]). SIGNIFICANCE: Auto-paracrine acetylcholine limits the migration of unstimulated peripheral granulocytes. This effect is impaired in cystic fibrosis most likely because of a reduced endogenous cholinergic tone.
ESTHER : Wessler_2012_Life.Sci_91_1119
PubMedSearch : Wessler_2012_Life.Sci_91_1119
PubMedID: 22525378

Title : The non-neuronal cholinergic system: basic science, therapeutic implications and new perspectives -
Author(s) : Grando SA , Kawashima K , Kirkpatrick CJ , Meurs H , Wessler I
Ref : Life Sciences , 91 :969 , 2012
PubMedID: 23141771

Title : Acetylcholine beyond neurons: the non-neuronal cholinergic system in humans - Wessler_2008_Br.J.Pharmacol_154_1558
Author(s) : Wessler I , Kirkpatrick CJ
Ref : British Journal of Pharmacology , 154 :1558 , 2008
Abstract : Animal life is controlled by neurons and in this setting cholinergic neurons play an important role. Cholinergic neurons release ACh, which via nicotinic and muscarinic receptors (n- and mAChRs) mediate chemical neurotransmission, a highly integrative process. Thus, the organism responds to external and internal stimuli to maintain and optimize survival and mood. Blockade of cholinergic neurotransmission is followed by immediate death. However, cholinergic communication has been established from the beginning of life in primitive organisms such as bacteria, algae, protozoa, sponge and primitive plants and fungi, irrespective of neurons. Tubocurarine- and atropine-sensitive effects are observed in plants indicating functional significance. All components of the cholinergic system (ChAT, ACh, n- and mAChRs, high-affinity choline uptake, esterase) have been demonstrated in mammalian non-neuronal cells, including those of humans. Embryonic stem cells (mice), epithelial, endothelial and immune cells synthesize ACh, which via differently expressed patterns of n- and mAChRs modulates cell activities to respond to internal or external stimuli. This helps to maintain and optimize cell function, such as proliferation, differentiation, formation of a physical barrier, migration, and ion and water movements. Blockade of n- and mACHRs on non-innervated cells causes cellular dysfunction and/or cell death. Thus, cholinergic signalling in non-neuronal cells is comparable to cholinergic neurotransmission. Dysfunction of the non-neuronal cholinergic system is involved in the pathogenesis of diseases. Alterations have been detected in inflammatory processes and a pathobiologic role of non-neuronal ACh in different diseases is discussed. The present article reviews recent findings about the non-neuronal cholinergic system in humans.
ESTHER : Wessler_2008_Br.J.Pharmacol_154_1558
PubMedSearch : Wessler_2008_Br.J.Pharmacol_154_1558
PubMedID: 18500366

Title : The non-neuronal cholinergic system in peripheral blood cells: effects of nicotinic and muscarinic receptor antagonists on phagocytosis, respiratory burst and migration - Neumann_2007_Life.Sci_80_2361
Author(s) : Neumann S , Razen M , Habermehl P , Meyer CU , Zepp F , Kirkpatrick CJ , Wessler I
Ref : Life Sciences , 80 :2361 , 2007
Abstract : Peripheral blood cells express the complete non-neuronal cholinergic system. For example synthesis of acetylcholine and nicotinic as well muscarinic receptors have been demonstrated in leucocytes isolated from human peripheral blood. In the present experiments mononuclear cells and granulocytes were isolated from the peripheral blood to investigate content and synthesis of acetylcholine as well as phenotypic functions like respiratory burst, phagocytosis and migration. Mononuclear cells (T-cells and monocytes) contained 0.36 pmol/10(6) cells acetylcholine, whereas acetylcholine content in granulocytes was 100-fold lower. Acetylcholine synthesis amounted to 23.2+/-4.7 nmol/mg protein/h and 2.90+/-0.84 in CD15+ (granulocytes) and CD3+ cells (T-lymphocytes), respectively. Neither atropine (blockade of muscarinic receptors) nor tubocurarine (blockade of nicotinic receptors) exerted an effect on the respiratory burst. Tubocurarine (30 muM), alone or in combination with atropine (1 microM), reduced phagocytosis in granulocytes by 13% and 19%, respectively (p<0.05). Spontaneous transwell migration of granulocytes was doubled by tubocurarine combined with atropine (p>0.05). Also alpha-bungarotoxin (10 microg/ml) enhanced spontaneous granulocyte migration, but hexamethonium (300 microM) was without effect. The present experiments demonstrate a cholinergic modulation of immune functions in peripheral leucocytes under in vitro conditions, i.e. in the absence of a neuronal innervation. Blockade of nicotine receptors (alpha1 muscular subtype) facilitates spontaneous migration of granulocytes.
ESTHER : Neumann_2007_Life.Sci_80_2361
PubMedSearch : Neumann_2007_Life.Sci_80_2361
PubMedID: 17286990

Title : In vivo release of non-neuronal acetylcholine from human skin by dermal microdialysis: effects of sunlight, UV-A and tactile stimulus - Schlereth_2007_Life.Sci_80_2239
Author(s) : Schlereth T , Schonefeld S , Birklein F , Kirkpatrick CJ , Wessler I
Ref : Life Sciences , 80 :2239 , 2007
Abstract : Non-neuronal acetylcholine (ACh) is expressed in epithelial, endothelial and immune cells. For example, the in vivo release of ACh from the human skin pretreated with botulinum toxin has recently been demonstrated. In the present experiments the effects of light (sunlight and solar radiation by a commercial UV-A applier) and of a tactile stimulus on the release of non-neuronal ACh were investigated. Release of ACh from the proximal and distal shin, i.e. anterior tibial region, was measured by dermal microdialysis in 20 min samples over a time period of at least 140 min. Control experiments were performed in a dark room throughout. In some experiments volunteers were exposed to sunshine (80-140 min) or the shin region was illuminated (80-95 min) by a commercial UV-A lamp (400 W at a distance of 50 cm). In control experiments ACh release between 20 and 80 min (B1) amounted to 118+/-32 pmol (n=17) and gradually declined between 80 and 140 min (B2) to 112+/-34 pmol, resulting in a B2/B1 ratio of 0.95. When the skin was exposed to sunlight ACh release increased from 205+/-58 pmol (B1) to 349+/-122 pmol resulting in a B2/B1 ratio of 1.70. UV-A radiation, however, had no significant effect on the B2/B1 ratio. When very smooth tactile stimuli were applied by a cotton wool tip for 20 min to the skin close to the microdialysis membranes in a dark room, ACh release was increased from 9+/-2 pmol/20 min to 52+/-36 (n=7). In conclusion, the in vivo release of ACh from the human skin appears to be regulated by external stimuli like sunlight and tactile stimuli.
ESTHER : Schlereth_2007_Life.Sci_80_2239
PubMedSearch : Schlereth_2007_Life.Sci_80_2239
PubMedID: 17276464

Title : Dysfunction of the non-neuronal cholinergic system in the airways and blood cells of patients with cystic fibrosis - Wessler_2007_Life.Sci_80_2253
Author(s) : Wessler I , Bittinger F , Kamin W , Zepp F , Meyer E , Schad A , Kirkpatrick CJ
Ref : Life Sciences , 80 :2253 , 2007
Abstract : The non-neuronal cholinergic system is widely expressed in human airways, skin and immune cells. Choline acetyltransferase (ChAT), acetylcholine and nicotine/muscarine receptors are demonstrated in epithelial surface cells, submucosal glands, airway smooth muscle fibres and immune cells. Moreover, acetylcholine is involved in the regulation of cell functions like proliferation, differentiation, migration, organization of the cytoskeleton, cell-cell contact, secretion and transport of ions and water. Cystic fibrosis (CF), the most frequent genetic disorder, is known to be caused by a mutation of the CF-gene coding for the cystic fibrosis transmembrane regulator protein (CFTR). CFTR represents a regulating transport protein for ion channels and processes involving endo- and exocytosis. Despite the identification of the genetic mutation knowledge of the underlying cellular pathways is limited. In the present experiments the cholinergic system was investigated in the peripheral blood and in the lung of CF patients undergoing lung transplantation (n=7). Acetylcholine content in bronchi and lung parenchyma of CF was reduced by 70% compared to controls (tumor-free tissue obtained from patients with lung tumor; n=13). In contrast, ChAT activity was elevated to some extent (p>0.05) in CF, and esterase activity did not differ from control. Acetylcholine content extracted from peripheral leucocytes (30 ml) was also reduced by 70% in CF (n=13) compared to healthy volunteers (n=9). Double labelling experiments with anti-CF antibodies and anti-ChAT antibodies showed a co-localization in peripheral lymphocytes, giving first evidence that CFTR may be linked with the intracellular storage/transport of non-neuronal acetylcholine. It is concluded that the non-neuronal cholinergic system is involved in the pathogenesis of CF. A reduced content of non-neuronal acetylcholine could contribute to the deleterious changes of epithelial ion and water movements in CF, because acetylcholine stimulates apical Cl(-) secretion, inhibits apical Na(+) and water absorption and therewith facilitates mucociliary clearance.
ESTHER : Wessler_2007_Life.Sci_80_2253
PubMedSearch : Wessler_2007_Life.Sci_80_2253
PubMedID: 17346753

Title : Release of non-neuronal acetylcholine from the isolated human placenta is affected by antidepressants - Wessler_2007_Life.Sci_80_2210
Author(s) : Wessler I , Herschel S , Bittinger F , Kirkpatrick CJ
Ref : Life Sciences , 80 :2210 , 2007
Abstract : Non-neuronal acetylcholine (ACh) is released from the human placenta into the extracellular space via organic cation transporters (OCTs). The present experiments investigated whether ACh release from epithelial cells is affected by drugs which are substrates of OCTs. The antidepressant drugs amitriptyline and doxepine were tested as both substances are not approved for pregnant women but frequently used. Release of ACh was measured in 10 min intervals over a period of 100 min. Test substances were added from t=50 min of incubation onwards. The effect was calculated by comparing the ACh release of the last three samples (t=70-100 min; B2) with that immediately before the application of the test substances (t=20-50 min; B1). The baseline ACh release amounted to 2.07+/-0.17 nmol/10 min (n=29; villus). Under control conditions a B2/B1 ratio of 0.78+/-0.02 was obtained. The following B2/B1 ratios were found, when therapeutic drugs were added: 0.54+/-0.04 (n=7; P<0.05) in the presence of 10 microM amitriptyline; 0.44+/-0.04 (10; P<0.01) in the presence of 10 microM doxepin; 0.73+/-0.04 (13) in the presence of 10 microM metformin; 0.76+/-0.06 (7) in the presence of 10 microM minoxidil; 0.63+/-0.03 (10) in the presence of 1 microM theophylline. The results demonstrate that antidepressants reduce the release of non-neuronal ACh at least in the human placenta, most likely by intracellular substrate competition at the polyspecific organic cation transporters (OCTs) but only at concentrations roughly 30-fold above the therapeutic range. Theophylline may also interfere with the release of non-neuronal ACh.
ESTHER : Wessler_2007_Life.Sci_80_2210
PubMedSearch : Wessler_2007_Life.Sci_80_2210
PubMedID: 17275854

Title : Recent progress in understanding the non-neuronal cholinergic system in humans -
Author(s) : Grando SA , Kawashima K , Kirkpatrick CJ , Wessler I
Ref : Life Sciences , 80 :2181 , 2007
PubMedID: 17467010

Title : The non-neuronal cholinergic system of human skin - Kurzen_2007_Horm.Metab.Res_39_125
Author(s) : Kurzen H , Wessler I , Kirkpatrick CJ , Kawashima K , Grando SA
Ref : Hormone & Metabolic Research , 39 :125 , 2007
Abstract : In human skin both resident and transiently residing cells are part of the extra- or non-neuronal cholinergic system, creating a highly complex and interconnected cosmos in which acetylcholine (ACh) and choline are the natural ligands of nicotinic and muscarinic receptors with regulatory function in both physiology and pathophysiology. ACh is produced in keratinocytes, endothelial cells and most notably in immune competent cells invading the skin at sites of inflammation. The cholinergic system is involved in basic functions of the skin through autocrine, paracrine, and endocrine mechanisms, like keratinocyte proliferation, differentiation, adhesion and migration, epidermal barrier formation, pigment-, sweat- and sebum production, blood circulation, angiogenesis, and a variety of immune reactions. The pathophysiological consequences of this complex cholinergic "concert" are only beginning to be understood. The present review aims at providing insight into basic mechanisms of this highly complex system.
ESTHER : Kurzen_2007_Horm.Metab.Res_39_125
PubMedSearch : Kurzen_2007_Horm.Metab.Res_39_125
PubMedID: 17326008

Title : Dysfunctional inhibitory muscarinic receptors mediate enhanced histamine release in isolated human bronchi - Wessler_2007_Life.Sci_80_2294
Author(s) : Wessler I , Holper B , Kortsik C , Buhl R , Kilbinger H , Kirkpatrick CJ
Ref : Life Sciences , 80 :2294 , 2007
Abstract : In human airways mucosal mast cells are under the control of inhibitory muscarinic receptors. The described experiments tested, whether the inhibitory potency of two muscarinic receptor agonists (oxotremorine, acetylcholine) becomes impaired in advanced chronic obstructive pulmonary disease (COPD). Isolated human bronchi obtained from 26 patients with lung cancer were separated into two groups. Group 1 patients suffered from moderate COPD (mean FEV1 56%; range 34-71%; mean pack years of cigarette smoking 50, range 20-96; one non-smoker). Group 2 patients had no or only a mild form of COPD; mean FEV1 was 82% (62-97%) and the number of pack years 22 (6-45; 3 non-smoker). The calcium ionophore A23187 induced a maximal histamine release of 4100+/-870 pmol/g/5 min in group 1 bronchi, in contrast to only 1730+/-240 pmol/g/5 min in group 2 bronchi (p<0.02). Oxotremorine (1 nmol/L) reduced the stimulated histamine release by 81+/-5% in group 2 bronchi, but did not produce a significant effect in group 1 bronchi (11+/-14%). In conclusion, the present experiments show an enhanced histamine release in advanced COPD, which can be explained by a dysfunction of inhibitory muscarinic receptors.
ESTHER : Wessler_2007_Life.Sci_80_2294
PubMedSearch : Wessler_2007_Life.Sci_80_2294
PubMedID: 17320912

Title : In vivo release of non-neuronal acetylcholine from the human skin as measured by dermal microdialysis: effect of botulinum toxin - Schlereth_2006_Br.J.Pharmacol_147_183
Author(s) : Schlereth T , Birklein F , an Haack K , Schiffmann S , Kilbinger H , Kirkpatrick CJ , Wessler I
Ref : British Journal of Pharmacology , 147 :183 , 2006
Abstract : 1.--Acetylcholine is synthesized in the majority of non-neuronal cells, for example in human skin. In the present experiments, the in vivo release of acetylcholine was measured by dermal microdialysis. 2.--Two microdialysis membranes were inserted intradermally at the medial shank of volunteers. Physiological saline containing 1 muM neostigmine was perfused at a constant rate of 4 microl min(-1) and the effluent was collected in six subsequent 20 min periods. Acetylcholine was measured by high-pressure liquid chromatography (HPLC) combined with bioreactors and electrochemical detection. 3.--Analysis of the effluent by HPLC showed an acetylcholine peak that disappeared, when the analytical column was packed with acetylcholine-specific esterase, confirming the presence of acetylcholine. 4.--In the absence of neostigmine, 71+/-51 pmol acetylcholine (n=4) was found during a 120 min period. The amount increased to 183+/-43 pmol (n=34), when the perfusion medium contained 1 microM neostigmine. 5.--Injection of 100 MU botulinum toxin subcutaneously blocked sweating completely, but the release of acetylcholine was not affected (botulinum toxin treated skin: 116+/-70 pmol acetylcholine/120 min; untreated skin: 50+/-20 pmol; n=4). 6.--Quinine (1 mM), inhibitor of organic cation transporters, and carnitine (0.1 mM), substrate of the Na(+)-dependent carnitine transporter OCTN2, tended to reduce acetylcholine release (by 40%, not significant). 7.--Our experiments demonstrate, for the first time, the in vivo release of non-neuronal acetylcholine in human skin. Organic cation transporters are not predominantly involved in the release of non-neuronal acetylcholine from the human skin.
ESTHER : Schlereth_2006_Br.J.Pharmacol_147_183
PubMedSearch : Schlereth_2006_Br.J.Pharmacol_147_183
PubMedID: 16273117

Title : Proliferative effect of acetylcholine on rat trachea epithelial cells is mediated by nicotinic receptors and muscarinic receptors of the M1-subtype - Metzen_2003_Life.Sci_72_2075
Author(s) : Metzen J , Bittinger F , Kirkpatrick CJ , Kilbinger H , Wessler I
Ref : Life Sciences , 72 :2075 , 2003
Abstract : Acetylcholine (ACh), synthesized in mammalian non-neuronal cells such as epithelial cells of the airways, digestive tract and skin, is involved in the regulation of basic cell functions (so-called non-neuronal cholinergic system). In the present experiments rat trachea epithelial cells have been cultured to study the proliferative effect of applied ACh by [3H]thymidine incorporation. ACh (exposure time 24 h) caused a concentration-dependent increase in cell proliferation with a doubling of the [3H]thymidine incorporation at a concentration of 0.1 microM. This effect was partly reduced by 30 microM tubocurarine and completely abolished by the additional application of 1 microM atropine. The stimulatory effect of acetylcholine, remaining in the presence of tubocurarine, was prevented by 1 microM pirenzepine (preferentially acting at M1-receptors), but neither by 1 microM AFDX 116 (preferentially acting at M2-receptors) nor by 1 microM hexahydrosiladifenidol (preferentially acting at M3-receptors). The combination of tubocurarine and pirenzepine halved the basal [3H]thymidine incorporation. In conclusion, ACh produces a proliferative effect in rat trachea epithelial cells, the effect being mediated by both nicotinic receptors and muscarinic receptors of the M1-subtype.
ESTHER : Metzen_2003_Life.Sci_72_2075
PubMedSearch : Metzen_2003_Life.Sci_72_2075
PubMedID: 12628459

Title : Expression and function of the non-neuronal cholinergic system in endothelial cells - Kirkpatrick_2003_Life.Sci_72_2111
Author(s) : Kirkpatrick CJ , Bittinger F , Nozadze K , Wessler I
Ref : Life Sciences , 72 :2111 , 2003
Abstract : Increasing evidence has shown the expression of the non-neuronal cholinergic system in endothelial cells. In the present experiments the expression of choline acetyltransferase (ChAT) was investigated in human endothelial cells by anti-ChAT immunohistochemistry and anti-ChAT immunofluorescence. Positive ChAT immunoreactivity was found in cultures of human umbilical endothelial cells (HUVEC) and a human angiosarcoma cell line (HAEND). In HUVEC and HAEND choline acetyltransferase activity and small amounts of acetylcholine were also detected. Positive ChAT-immunoreactivity was demonstrated in situ in endothelial cells of the human umbilical cord. In addition, in experiments with confocal laser scanning microscopy positive anti-ChAT immunoreactivity was found in situ in endothelial cells of human skin blood vessels. In the first functional experiments with HUVEC acetylcholine appeared to mediate a small facilitatory effect on the expression of intracellular adhesion molecule-1. The present experiments demonstrate the wide existence of ChAT in human endothelial cells. Further experiments are addressed to elucidate the biological role of acetylcholine in the endothelium and possible differences between the different subtypes of endothelial cells.
ESTHER : Kirkpatrick_2003_Life.Sci_72_2111
PubMedSearch : Kirkpatrick_2003_Life.Sci_72_2111
PubMedID: 12628465

Title : The non-neuronal cholinergic system in humans: expression, function and pathophysiology - Wessler_2003_Life.Sci_72_2055
Author(s) : Wessler I , Kilbinger H , Bittinger F , Unger R , Kirkpatrick CJ
Ref : Life Sciences , 72 :2055 , 2003
Abstract : Acetylcholine, a prime example of a neurotransmitter, has been detected in bacteria, algae, protozoa, and primitive plants, indicating an extremely early appearance in the evolutionary process (about 3 billion years). In humans, acetylcholine and/or the synthesizing enzyme, choline acetyltransferase (ChAT), have been found in epithelial cells (airways, alimentary tract, urogenital tract, epidermis), mesothelial (pleura, pericardium), endothelial, muscle and immune cells (mononuclear cells, granulocytes, alveolar macrophages, mast cells). The widespread expression of non-neuronal acetylcholine is accompanied by the ubiquitous presence of cholinesterase and receptors (nicotinic, muscarinic). Thus, the non-neuronal cholinergic system and non-neuronal acetylcholine, acting as a local cellular signaling molecule, has to be discriminated from the neuronal cholinergic system and neuronal acetylcholine, acting as neurotransmitter. In the human placenta anti-ChAT immunoreactivity is found in multiple subcellular compartments like the cell membrane (microvilli, coated pits), endosomes, cytoskeleton, mitochondria and in the cell nucleus. These locations correspond with the results of experiments where possible functions of non-neuronal acetylcholine have been identified (proliferation, differentiation, organization of the cytoskeleton and the cell-cell contact, locomotion, migration, ciliary activity, immune functions). In the human placenta acetylcholine release is mediated by organic cation transporters. Thus, structural and functional differences are evident between the non-neuronal and neuronal cholinergic system. Enhanced levels of acetylcholine are detected in inflammatory diseases. In conclusion, it is time to revise the role of acetylcholine in humans. Its biological and pathobiological roles have to be elucidated in more detail and possibly, new therapeutical targets may become available.
ESTHER : Wessler_2003_Life.Sci_72_2055
PubMedSearch : Wessler_2003_Life.Sci_72_2055
PubMedID: 12628456

Title : Increased acetylcholine levels in skin biopsies of patients with atopic dermatitis - Wessler_2003_Life.Sci_72_2169
Author(s) : Wessler I , Reinheimer T , Kilbinger H , Bittinger F , Kirkpatrick CJ , Saloga J , Knop J
Ref : Life Sciences , 72 :2169 , 2003
Abstract : Recent experimental evidence indicates that non-neuronal acetylcholine is involved in the regulation of basic cell functions. Here we investigated the cholinergic system in the skin of healthy volunteers and patients with atopic dermatitis (AD). The synthesizing enzyme, choline-acetyltransferase (ChAT), was studied by anti-ChAT immunohistochemistry and enzyme assay. Skin biopsies taken from healthy volunteers and from AD patients were separated into the 2 mm superfical (epidermis and upper dermis) and 3 mm underlying portion (deeper dermis and subcutis). ChAT enzyme activity was detected in homogenized skin and subcutaneous fat (about 13 nmol/mg protein/h). ChAT immunoreactivity was expressed in keratinocytes, hair papilla, sebaceous and eccrine sweat glands, endothelial cells and mast cells. In healthy volunteers the superficial and underlying portion of skin biopsies contained 130 +/- 30 and 550 +/- 170 pmol/g acetylcholine (n = 12), respectively. In AD patients (n = 7) acetylcholine was increased 14-fold in the superficial and 3-fold in the underlying biopsy portion. The present study demonstrates the widespread expression of ChAT protein in the vast majority of human skin cells. Tissue levels of acetylcholine are greatly (14-fold) enhanced in the superficial 2 mm skin of AD patients.
ESTHER : Wessler_2003_Life.Sci_72_2169
PubMedSearch : Wessler_2003_Life.Sci_72_2169
PubMedID: 12628475

Title : Effects of sex hormones, forskolin, and nicotine on choline acetyltransferase activity in human isolated placenta - Wessler_2003_Neurochem.Res_28_489
Author(s) : Wessler I , Schwarze S , Brockerhoff P , Bittinger F , Kirkpatrick CJ , Kilbinger H
Ref : Neurochem Res , 28 :489 , 2003
Abstract : The activity of choline acetyltransferase (ChAT) was investigated in the human placenta before and after long-term incubation (24 h) to test the effects of sex hormones, nicotine and forskolin. ChAT activity differed considerably between the amnion (0.03 micromol/mg protein/h) and the villus (0.56). After long-term incubation, ChAT activity persisted in the latter but declined in the amnion. Neither sex hormones (beta-estradiol, testosterone, progesterone; 10 or 100 nM each) nor follicle stimulating hormone and luteinizing hormone (FSH/LH; 8.4 U/ml each) modified ChAT activity. Also nicotine (1 nM-100 microM) did not affect ChAT activity. Forskolin, an activitor of adenylyl cyclase, reduced ChAT activity in the villus but not in amnion. The present model offers the possibility to investigate ChAT regulation in intact tissue under long-term incubation. The risks of maternal smoking during pregnancy cannot be attributed to an effect of nicotine on placental ChAT activity. Differences in the regulation of ChAT appear to exist between neuronal and nonneuronal cells.
ESTHER : Wessler_2003_Neurochem.Res_28_489
PubMedSearch : Wessler_2003_Neurochem.Res_28_489
PubMedID: 12675136

Title : The Non-neuronal cholinergic system: an emerging drug target in the airways - Wessler_2001_Pulm.Pharmacol.Ther_14_423
Author(s) : Wessler I , Kirkpatrick CJ
Ref : Pulm Pharmacol Ther , 14 :423 , 2001
Abstract : The non-neuronal cholinergic system is widely expressed in human airways. Choline acetyltransferase (ChAT) and/or acetylcholine are demonstrated in more or less all epithelial surface cells (goblet cells, ciliated cells, basal cells), submucosal glands and airway smooth muscle fibres. Acetylcholine is also demonstrated in the effector cells of the immune system (lymphocytes, macrophages, mast cells). Epithelial, endothelial and immune cells express nicotinic and muscarinic receptors. Thus the cytomolecule acetylcholine can contribute to the regulation of basic cell functions via auto-/paracrine mechanisms (proliferation, differentiation, ciliary activity, secretion of water, ions and mucus, organization of the cytoskeleton, cell-cell contact). Acetylcholine also modulates immune functions (release of cytokines; proliferation, activation and inhibition of immune cells). Preliminary experimental evidence suggests that mucosal inflammation may be associated with raised acetylcholine levels, impairing cell and organ homeostasis. It should be considered that anti-muscarinic drugs which are applied for the treatment of chronic airway diseases antagonize the effect of both neuronal and non-neuronal acetylcholine. Non-neuronal acetylcholine, however, is still active, possibly directly within the cell cytosol and also via nicotinic receptors localized on various non-neuronal cells. It is an essential task to clarify the pathophysiological role of the non-neuronal cholinergic system in more detail to develop new drugs which can target the synthesis, release, inactivation and cellular activity of non-neuronal acetylcholine.
ESTHER : Wessler_2001_Pulm.Pharmacol.Ther_14_423
PubMedSearch : Wessler_2001_Pulm.Pharmacol.Ther_14_423
PubMedID: 11782122

Title : The non-neuronal cholinergic system in the endothelium: evidence and possible pathobiological significance - Kirkpatrick_2001_Jpn.J.Pharmacol_85_24
Author(s) : Kirkpatrick CJ , Bittinger F , Unger RE , Kriegsmann J , Kilbinger H , Wessler I
Ref : Japanese Journal of Pharmacology , 85 :24 , 2001
Abstract : An increasing body of knowledge indicates that the cholinergic system is not confined to the nervous system, but is practically ubiquitous. The present paper will address the question of the non-neuronal cholinergic system in vascular endothelial cells (EC). In tissue sections of human skin, immunohistochemical studies using confocal laser scanning microscopy showed ChAT (choline acetyltransferase) activity in the EC of dermal blood vessels. Positive ChAT immunoreactivity was also demonstrated in monolayer cultures of human umbilical vein EC (HUVEC) and a human angiosarcoma EC line (HAEND). That the synthesizing enzyme is not only present in EC, but also active was shown by measuring ChAT activity. Thus, in HUVEC cultures, ChAT activity amounted to 0.78 +/- 0.15 nmol x mg protein(-1) x h(-1) (n = 3), but was only partially (about 50%) inhibited by the ChAT inhibitor bromoacetylcholine (30 microM). In HPLC measurements, a concentration of 22 +/- 2 pmol acetylcholine (ACh) per 10(6) cells was found (n = 6). However, using a cholinesterase-packed analytical column to check the identity of the acetylcholine peak, the peak height was found to be reduced, although a significant peak still remained, indicating the existence of a compound closely related to ACh. Further immunocytochemical experiments indicated that EC in vitro also express the vesicular acetylcholine transporter (VAChT) system. Preliminary immunoelectron microscopic studies suggest a topographical association of VAChT with endothelial endocytotic vesicles. The presented experiments clearly demonstrate the existence of essential elements of the cholinergic system (ChAT, VAChT, ACh) in the human endothelium. The biological functions of ACh synthesized by endothelial cells are the focus of ongoing research activity.
ESTHER : Kirkpatrick_2001_Jpn.J.Pharmacol_85_24
PubMedSearch : Kirkpatrick_2001_Jpn.J.Pharmacol_85_24
PubMedID: 11243570

Title : The biological role of non-neuronal acetylcholine in plants and humans - Wessler_2001_Jpn.J.Pharmacol_85_2
Author(s) : Wessler I , Kilbinger H , Bittinger F , Kirkpatrick CJ
Ref : Japanese Journal of Pharmacology , 85 :2 , 2001
Abstract : Acetylcholine, one of the most exemplary neurotransmitters, has been detected in bacteria, algae, protozoa, tubellariae and primitive plants, suggesting an extremely early appearance in the evolutionary process and a wide expression in non-neuronal cells. In plants (Urtica dioica), acetylcholine is involved in the regulation of water resorption and photosynthesis. In humans, acetylcholine and/or the synthesizing enzyme, choline acetyltransferase, have been demonstrated in epithelial (airways, alimentary tract, urogenital tract, epidermis), mesothelial (pleura, pericardium), endothelial, muscle and immune cells (granulocytes, lymphocytes, macrophages, mast cells). The widespread expression of non-neuronal acetylcholine is accompanied by the ubiquitous expression of cholinesterase and acetylcholine sensitive receptors (nicotinic, muscarinic). Both receptor populations interact with more or less all cellular signalling pathways. Thus, non-neuronal acetylcholine can be involved in the regulation of basic cell functions like gene expression, proliferation, differentiation, cytoskeletal organization, cell-cell contact (tight and gap junctions, desmosomes), locomotion, migration, ciliary activity, electrical activity, secretion and absorption. Non-neuronal acetylcholine also plays a role in the control of unspecific and specific immune functions. Future experiments should be designed to analyze the cellular effects of acetylcholine in greater detail and to illuminate the involvement of the non-neuronal cholinergic system in the pathogenesis of diseases such as acute and chronic inflammation, local and systemic infection, dementia, atherosclerosis, and finally cancer.
ESTHER : Wessler_2001_Jpn.J.Pharmacol_85_2
PubMedSearch : Wessler_2001_Jpn.J.Pharmacol_85_2
PubMedID: 11243568

Title : Release of non-neuronal acetylcholine from the isolated human placenta is mediated by organic cation transporters - Wessler_2001_Br.J.Pharmacol_134_951
Author(s) : Wessler I , Roth E , Deutsch C , Brockerhoff P , Bittinger F , Kirkpatrick CJ , Kilbinger H
Ref : British Journal of Pharmacology , 134 :951 , 2001
Abstract : 1. The release of acetylcholine was investigated in the human placenta villus, a useful model for the characterization of the non-neuronal cholinergic system. 2. Quinine, an inhibitor of organic cation transporters (OCT), reduced acetylcholine release in a reversible and concentration-dependent manner with an IC(50) value of 5 microM. The maximal effect, inhibition by 99%, occurred at a concentration of 300 microM. 3. Procaine (100 microM), a sodium channel blocker, and vesamicol (10 microM), an inhibitor of the vesicular acetylcholine transporter, were ineffective. 4. Corticosterone, an inhibitor of OCT subtype 1, 2 and 3 reduced acetylcholine in a concentration-dependent manner with an IC(50) value of 2 microM. 5. Substrates of OCT subtype 1, 2 and 3 (amiloride, cimetidine, guanidine, noradrenaline, verapamil) inhibited acetylcholine release, whereas carnitine, a substrate of subtype OCTN2, exerted no effect. 6. Long term exposure (48 and 72 h) of villus strips to anti-sense oligonucleotides (5 microM) directed against transcription of OCT1 and OCT3 reduced the release of acetylcholine, whereas OCT2 anti-sense oliogonucleotides were ineffective. 7. It is concluded that the release of non-neuronal acetylcholine from the human placenta is mediated via organic cation transporters of the OCT1 and OCT3 subtype.
ESTHER : Wessler_2001_Br.J.Pharmacol_134_951
PubMedSearch : Wessler_2001_Br.J.Pharmacol_134_951
PubMedID: 11682442

Title : Release of non-neuronal acetylcholine from the human placenta: difference to neuronal acetylcholine - Wessler_2001_Naunyn.Schmiedebergs.Arch.Pharmacol_364_205
Author(s) : Wessler I , Roth E , Schwarze S , Weikel W , Bittinger F , Kirkpatrick CJ , Kilbinger H
Ref : Naunyn Schmiedebergs Arch Pharmacol , 364 :205 , 2001
Abstract : The synthesis and release of non-neuronal acetylcholine, a widely expressed signaling molecule, were investigated in the human placenta. This tissue is free of cholinergic neurons, i.e. a contamination of neuronal acetylcholine can be excluded. The villus showed a choline acetyltransferase (ChAT) activity of 0.65 nmol/mg protein per h and contained 500 nmol acetylcholine/g dry weight. In the absence of cholinesterase inhibitors the release of acetylcholine from isolated villus pieces amounted to 1.3 nmol/g wet weight per 10 min corresponding to a fractional release rate of 0.13% per min. The following substances did not significantly modify the release of acetylcholine: oxotremorine (1 microM), scopolamine (1 microM), (+)-tubocurarine (30 microM), forskolin (30 microM), ouabain (10 microM), 4alpha-phorbol 12,13-didecanoate (1 microM) and tetrodotoxin (1 microM). Removal of extracellular calcium, phorbol 12,13-dibutyrate (1 microM) and colchicine (100 microM) reduced the acetylcholine release between 30% and 50%. High potassium chloride (54 mM and 108 mM) increased the acetylcholine release slightly (by about 30%). A concentration of 10 microM nicotine was ineffective, but 100 microM nicotine enhanced acetylcholine release gradually over a 50-min period without desensitization of the response. The facilitatory effect of nicotine was prevented by 30 microM (+)-tubocurarine. Inhibitors of cholinesterase (physostigmine, neostigmine; 3 microM) facilitated the efflux of acetylcholine about sixfold, and a combination of both (+)-tubocurarine (30 microM) and scopolamine (1 microM) halved the enhancing effect. In conclusion, release mechanisms differ between non-neuronal and neuronal acetylcholine. Facilitatory nicotine receptors are present which are activated by applied nicotine or by blocking cholinesterase. Thus, cholinesterase inhibitors increase assayed acetylcholine by two mechanisms, protection of hydrolysis and stimulation of facilitatory nicotine receptors.
ESTHER : Wessler_2001_Naunyn.Schmiedebergs.Arch.Pharmacol_364_205
PubMedSearch : Wessler_2001_Naunyn.Schmiedebergs.Arch.Pharmacol_364_205
PubMedID: 11521162

Title : Airway epithelium: more than just a barrier! -
Author(s) : Wessler I , Kirkpatrick CJ , Racke K
Ref : Trends in Pharmacological Sciences , 20 :52 , 1999
PubMedID: 10101963

Title : The cholinergic 'pitfall': acetylcholine, a universal cell molecule in biological systems, including humans - Wessler_1999_Clin.Exp.Pharmacol.Physiol_26_198
Author(s) : Wessler I , Kirkpatrick CJ , Racke K
Ref : Clinical & Experimental Pharmacology & Physiology , 26 :198 , 1999
Abstract : 1. Acetylcholine (ACh) represents one of the most exemplary neurotransmitters. In addition to its presence in neuronal tissue, there is increasing experimental evidence that ACh is widely expressed in pro- and eukaryotic non-neuronal cells. Thus, ACh has been detected in bacteria, algae, protozoa, tubellariae and primitive plants, suggesting an extremely early appearance of ACh in the evolutionary process. 2. In humans, ACh and/or the synthesizing enzyme, choline acetyltransferase, has been demonstrated in epithelial cells (airways, alimentary tract, urogenital tract, epidermis), mesothelial (pleura, pericardium) and endothelial and muscle cells. In addition, immune cells express the non-neuronal cholinergic system (i.e. the synthesis of ACh can be detected in human leucocytes (granulocytes, lymphocytes and macrophages)), as well as in rat microglia in vitro. 3. The widespread expression of non-neuronal ACh is accompanied by the ubiquitous expression of cholinesterase activity, which prevents ACh from acting as a classical hormone. 4. Non-neuronal ACh mediates its cellular actions in an auto- and paracrine manner via the activation of the widely expressed nicotinic and muscarinic acetylcholine receptors, which can interfere with virtually all cellular signalling pathways (ion channels and key enzymes). 5. Non-neuronal ACh appears to be involved in the regulation of basic cell functions, such as mitosis, cell differentiation, organization of the cytoskeleton, cell-cell contact, secretion and absorption. Non-neuronal ACh also plays a role in the regulation of immune functions. All these qualities together may mediate the so-called 'trophic property' of ACh. 6. Future experiments should be designed to analyse the cellular effects of ACh in greater detail. The involvement of the non-neuronal cholinergic system in the pathogenesis of chronic inflammatory diseases should be investigated to open up new therapeutic strategies.
ESTHER : Wessler_1999_Clin.Exp.Pharmacol.Physiol_26_198
PubMedSearch : Wessler_1999_Clin.Exp.Pharmacol.Physiol_26_198
PubMedID: 10081614

Title : Glucocorticoids mediate reduction of epithelial acetylcholine content in the airways of rats and humans - Reinheimer_1998_Eur.J.Pharmacol_349_277
Author(s) : Reinheimer T , Munch M , Bittinger F , Racke K , Kirkpatrick CJ , Wessler I
Ref : European Journal of Pharmacology , 349 :277 , 1998
Abstract : The cholinergic system in rat and human airways and the effects of glucocorticoids were investigated by assay of choline acetyltransferase activity, by high-pressure liquid chromatography measurement of acetylcholine, and by anti-choline acetyltransferase immunocyto-/histochemistry. Human bronchi were obtained at surgery from patients with lung cancer. Group 1 patients did not suffer from additional lung diseases and had not been treated with glucocorticoids. Group 2 patients, who suffered in addition to lung cancer from chronic obstructive bronchitis, had been treated for at least 6 weeks before surgery with four puffs of flusinolid daily. Isolated bronchial epithelial cells as well as intact surface epithelium of human bronchi expressed choline acetyltransferase immunoreactivity and choline acetyltransferase enzyme activity (3 +/- 1 nmol/mg protein per h). Ciliated epithelial cells showed strong choline acetyltransferase immunoreactivity at the basal body and the roolet of cilia. Surface epithelium in group 1 and 2 bronchi contained 23 +/- 6 (n = 14) and 1.8 +/- 0.3 pmol/g acetylcholine) (n = 7, P < 0.001), respectively, whereas the transmural acetylcholine content did not differ significantly between both groups. The amount of choline acetyltransferase immunoreactivity appeared similar in the surface epithelium of both groups. In an animal (rat) study the effects of oral dexamethasone (3 mg/day, 1 week) on choline acetyltransferase activity and acetylcholine levels were investigated. Dexamethasone treatment reduced epithelial acetylcholine in the airways and small intestine by about 80% and inhibited epithelial choline acetyltransferase activity. In conclusion, epithelial cells of human airways possess components of the cholinergic system, i.e., contain the synthesizing enzyme choline acetyltransferase and store acetylcholine. The data obtained from the animal study indicate that glucocorticoids can inhibit epithelial acetylcholine.
ESTHER : Reinheimer_1998_Eur.J.Pharmacol_349_277
PubMedSearch : Reinheimer_1998_Eur.J.Pharmacol_349_277
PubMedID: 9671108

Title : Non-neuronal acetylcholine, a locally acting molecule, widely distributed in biological systems: expression and function in humans - Wessler_1998_Pharmacol.Ther_77_59
Author(s) : Wessler I , Kirkpatrick CJ , Racke K
Ref : Pharmacol Ther , 77 :59 , 1998
Abstract : Acetylcholine acts as a neurotransmitter in the central and peripheral nervous systems in humans. However, recent experiments demonstrate a widespread expression of the cholinergic system in non-neuronal cells in humans. The synthesizing enzyme choline acetyltransferase, the signalling molecule acetylcholine, and the respective receptors (nicotinic or muscarinic) are expressed in epithelial cells (human airways, alimentary tract, epidermis). Acetylcholine is also found in mesothelial, endothelial, glial, and circulating blood cells (platelets, mononuclear cells), as well as in alveolar macrophages. The existence of non-neuronal acetylcholine explains the widespread expression of muscarinic and nicotinic receptors in cells not innervated by cholinergic neurons. Non-neuronal acetylcholine appears to be involved in the regulation of important cell functions, such as mitosis, trophic functions, automaticity, locomotion, ciliary activity, cell-cell contact, cytoskeleton, as well as barrier and immune functions. The most important tasks for the future will be to clarify the multiple biological roles of non-neuronal acetylcholine in detail and to identify pathological conditions in which this system is up- or down-regulated. This could provide the basis for the development of new therapeutic strategies to target the non-neuronal cholinergic system.
ESTHER : Wessler_1998_Pharmacol.Ther_77_59
PubMedSearch : Wessler_1998_Pharmacol.Ther_77_59
PubMedID: 9500159

Title : Day-night rhythm of acetylcholine in the rat pineal gland - Wessler_1997_Neurosci.Lett_224_173
Author(s) : Wessler I , Reinheimer T , Bittinger F , Kirkpatrick CJ , Schenda J , Vollrath L
Ref : Neuroscience Letters , 224 :173 , 1997
Abstract : Using high-performance-liquid-chromatography (HPLC) measurement of acetylcholine, choline acetyltransferase (ChAT) enzyme assay and anti-ChAT immunohistochemistry, we have investigated the expression of the cholinergic system in pineal glands of male rats. Glands procured during the day period (1200 h) contained significant amounts of acetylcholine (0.5 pmol/gland). A similar content was found in pineal glands after a 48 h culture period, i.e. when the intrapineal nerve fibres have degenerated. This strongly indicates that the pinealocytes are the cells which contain acetylcholine. To confirm this conclusion we demonstrate substantial ChAT-like immunoreactivity in pinealocytes. ChAT enzyme activity measured in homogenized glands (day period) was 7 +/- 3 nmol/mg per h. Acetylcholine content as well as ChAT enzyme activity increased about 10-fold in pineal glands during the night period (2400 h). The present study demonstrates for the first time the presence of a day-night rhythm of ChAT and acetylcholine in rat pinealocytes. The function of pineal acetylcholine is not clear, but there are indications that acetylcholine may depress noradrenaline release from intrapineal sympathetic fibres and hence melatonin synthesis.
ESTHER : Wessler_1997_Neurosci.Lett_224_173
PubMedSearch : Wessler_1997_Neurosci.Lett_224_173
PubMedID: 9131664

Title : Non-neuronal acetylcholine, a signalling molecule synthezised by surface cells of rat and man - Klapproth_1997_Naunyn.Schmiedebergs.Arch.Pharmacol_355_515
Author(s) : Klapproth H , Reinheimer T , Metzen J , Munch M , Bittinger F , Kirkpatrick CJ , Hohle KD , Schemann M , Racke K , Wessler I
Ref : Naunyn Schmiedebergs Arch Pharmacol , 355 :515 , 1997
Abstract : Acetylcholine acts as a prominent transmitter in the central and peripheral nervous system. The aim of the present study was to investigate whether mammalian non-neuronal cells can synthesize and store acetylcholine. A cotton tipped applicator (Q-tip) was used to collect surface cells from airways and alimentary tract. Histological inspection indicated that rubbing of the luminal surface of human bronchi did not penetrate the basal membrane. Acetylcholine was measured by an HPLC-method using substrate-specific enzyme reactor-columns. Non-neuronal acetylcholine was found in cells covering inner and outer surfaces of rat and man. For example, acetylcholine was detected in the surface epithelium of human bronchi (33 pmol/g), mouth (female 0.7 and male 8 pmol/sample), small and large intestine (800 and 16 pmol/g, respectively), gall bladder (12 pmol/g), vagina (6 pmol/sample), skin 1000 (pmol/g) and in pulmonary pleura (5 pmol/sample). Somewhat higher amounts of acetylcholine were found in rat tracheal and intestinal epithelium and in rat skin. The synthesizing enzyme choline acetyltransferase (ChAT) was demonstrated in human surface epithelium by immunohistochemistry and by Western blot analysis. Enzymatic ChAT activity was demonstrated in isolated epithelial cells of human bronchi and small intestine (3.5 and 28 nmol/mg protein/h, respectively). Applied acetylcholine (in nM concentrations) increased, whereas inhibition of ChAT activity by bromoacetylcholine (10 microM) reduced the growth of cultured human bronchial epithelial cells. Inhibition of cell growth occurred also in the presence of atropine (1 microM) together with (+/-)-tubocurarine (30 microM). In conclusion, the present experiments demonstrate a widespread existence of non-neuronal acetylcholine in surface cells of man. Non-neuronal acetylcholine may act as a local signalling molecule.
ESTHER : Klapproth_1997_Naunyn.Schmiedebergs.Arch.Pharmacol_355_515
PubMedSearch : Klapproth_1997_Naunyn.Schmiedebergs.Arch.Pharmacol_355_515
PubMedID: 9109369