title: Blockade of nicotinic receptors of bovine adrenal chromaffin cells by nanomolar concentrations of atropine.
abstract: Nanomolar concentrations of atropine have been considered up to now to be selective for blockade of muscarinic receptors for acetylcholine. A collateral finding indicated to us that these low concentrations of atropine could also target the neuronal nicotinic receptors. We report here a detailed study on this novel property of atropine. Catecholamine release, measured on-line with amperometry in chromaffin cells stimulated with acetylcholine pulses was blocked by atropine in a competitive manner. To corroborate a direct action of atropine on nicotinic receptors, we have employed N,N-dimethyl-N'-phenyl-piperazinium (DMPP), a pure nicotinic receptor agonist; atropine blocked its secretory responses with an IC50 of 2.04 nM. Nicotinic currents, recorded with the whole cell configuration of the patch-clamp technique were blocked by atropine in a concentration-dependent manner (IC50 of 11 nM), also showing a competitive nature. Nicotinic receptor currents in oocytes expressing bovine alpha7 and alpha3beta4 nicotinic receptors were blocked by atropine with an IC50 of 11.2 and 46.8 nM, respectively. Atropine (30 nM) also decreased the increment of the cytosolic calcium concentrations after stimulation with 30 microM DMPP in bovine chromaffin cells. However, action potentials evoked by DMPP were not modified by atropine. Our results demonstrate that nicotinic currents and their downstream consequences (i.e. cytosolic calcium elevations and catecholamine release) were blocked by nanomolar concentrations of atropine; although the blockade was partial, it must be considered when using atropine to study cholinergic neurotransmission, particularly at synapses where both nicotinic and muscarinic receptors are present i.e., the adrenal medulla and autonomic ganglia.
authors.affiliation: Instituto Teófilo Hernando, Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Avenida Arzobispo Morcillo 4, E-28029 Madrid, Spain.
authors.forename: Juana Maria
authors.initials: JM
authors.lastname: González-Rubio
authors.forename: Antonio M
authors.initials: AM
authors.lastname: García de Diego
authors.forename: Javier
authors.initials: J
authors.lastname: Egea
authors.forename: Román
authors.initials: R
authors.lastname: Olivares
authors.forename: Jonathan
authors.initials: J
authors.lastname: Rojo
authors.forename: Luis
authors.initials: L
authors.lastname: Gandía
authors.forename: Antonio G
authors.initials: AG
authors.lastname: García
authors.forename: Jesús M
authors.initials: JM
authors.lastname: Hernández-Guijo
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000313:Adrenal Medulla / Q000166:cytology; D000818:Animals; D001285:Atropine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000502:physiology; D002395:Catecholamines / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004246:Dimethylphenylpiperazinium Iodide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005260:Female; D015870:Gene Expression; D008564:Membrane Potentials / Q000187:drug effects; D008832:Microchemistry; D018727:Muscarinic Antagonists / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D018733:Nicotinic Antagonists / Q000494:pharmacology*; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D011976:Receptors, Muscarinic / Q000502:physiology; D011978:Receptors, Nicotinic / Q000235:genetics / Q000502:physiology*; D014982:Xenopus laevis; D064569:alpha7 Nicotinic Acetylcholine Receptor
chemical_list: D015220:Calcium Channels; D002395:Catecholamines; D018727:Muscarinic Antagonists; D018722:Nicotinic Agonists; D018733:Nicotinic Antagonists; D011976:Receptors, Muscarinic; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor; C403530:nicotinic receptor alpha3beta4; D004246:Dimethylphenylpiperazinium Iodide; D001285:Atropine; D000109:Acetylcholine; D002118:Calcium