title: Alpha3* and alpha 7 nAChR-mediated Ca2+ transient generation in IMR-32 neuroblastoma cells.
abstract: Alpha3-containing (alpha 3*) and alpha 7 nicotinic acetylcholine receptors (nAChRs) are expressed in human IMR-32 neuroblastoma cells and implicated in Ca(2+) signaling. In this study, we investigated the intracellular Ca(2+) transient generation evoked by selective activation of alpha 3* (agonist potency rank order: epibatidine>varenicline>nicotine approximately cytisine) and alpha 7 (rank order in the presence of alpha 7 positive allosteric modulator or PAM: A-795723>NS6784 approximately PNU-282987) using, respectively, varenicline and NS6784 (+alpha 7 PAM) by Ca(2+) imaging. Effects of inhibitors of nAChRs (MLA and mecamylamine), ER Ca(2+) ATPase pump (CPA and thapsigargin), Ca(2+)-induced Ca(2+) release (ryanodine and dantrolene), Ca(2+) channels (nitrendipine, diltiazem, and Cd(2+)), and removal of extracellular Ca(2+) were examined. alpha 7 PAMs, when tested in the presence of NS6784, were more active when added first, followed by the agonist, than in the reverse order. Removal of extracellular Ca(2+) - but not CPA, thapsigargin, ryanodine, dantrolene, nitrendipine, diltiazem, or Cd(2+) - diminished the alpha 7 agonist-evoked Ca(2+) transients. In contrast, only diltiazem and nitrendipine and removal of extracellular Ca(2+) inhibited the alpha 3*-mediated Ca(2+) transients. The differential effect of diltiazem and nitrendipine versus Cd(2+) was due to direct inhibition of alpha 3* nAChRs as revealed by Ca(2+) imaging in HEK-293 cells expressing human alpha 3 beta 4 nAChRs and patch clamp in IMR-32 cells. In summary, this study provides evidence that alpha 3* and alpha 7 nAChR agonist-evoked global Ca(2+) transient generation in IMR-32 cells does not primarily involve voltage-dependent Ca(2+) channels, intracellular Ca(2+) stores, or Ca(2+)-induced Ca(2+) release. These mechanisms may, however, be still involved in other forms of nAChR-mediated Ca(2+) signaling.
authors.affiliation: Neuroscience Research, Global Pharmaceutical Research and Development, Abbott, Oslo Research Park, Gaustadaleen 21, Oslo, Norway.
authors.forename: Hilde
authors.initials: H
authors.lastname: Ween
authors.forename: Kirsten
authors.initials: K
authors.lastname: Thorin-Hagene
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Andersen
authors.forename: Jens Halvard
authors.initials: JH
authors.lastname: Grønlien
authors.forename: Chih-Hung
authors.initials: CH
authors.lastname: Lee
authors.forename: Murali
authors.initials: M
authors.lastname: Gopalakrishnan
authors.forename: John
authors.initials: J
authors.lastname: Malysz
mesh_terms: D001549:Benzamides / Q000494:pharmacology; D001932:Brain Neoplasms / Q000378:metabolism*; D001643:Bridged Bicyclo Compounds / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020013:Calcium Signaling / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D045744:Cell Line, Tumor; D003627:Data Interpretation, Statistical; D004110:Diltiazem / Q000494:pharmacology; D004594:Electrophysiology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006801:Humans; D009447:Neuroblastoma / Q000378:metabolism*; D018722:Nicotinic Agonists / Q000494:pharmacology; D009568:Nitrendipine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011758:Pyrroles / Q000494:pharmacology; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000378:metabolism*; D013449:Sulfonamides / Q000494:pharmacology; D064569:alpha7 Nicotinic Acetylcholine Receptor; D000096926:Benzenesulfonamides
chemical_list: D001549:Benzamides; D001643:Bridged Bicyclo Compounds; D002121:Calcium Channel Blockers; D000252:Calcium-Transporting ATPases; D004110:Diltiazem; D018722:Nicotinic Agonists; D009568:Nitrendipine; D011758:Pyrroles; D011978:Receptors, Nicotinic; D013449:Sulfonamides; D064569:alpha7 Nicotinic Acetylcholine Receptor; C542089:4-(5-(4-chlorophenyl)-2-methyl-3-propionyl-1H-pyrrol-1-yl)benzenesulfonamide; D000096926:Benzenesulfonamides; C579167:Chrna7 protein, human; C498513:PNU-282987