title: Up-regulation of L-type voltage-dependent calcium channels after long term exposure to nicotine in cerebral cortical neurons.
abstract: Effects of long term (72-h) exposure to low concentration (0.1 mum) of nicotine on various types of voltage-dependent Ca(2+) channels (VDCCs) and neuronal nicotinic acetylcholine receptors (nnAChRs) were examined using primary cultures of mouse cerebral cortical neurons. High potassium (30 mm KCl)-stimulated (45)Ca(2+) influx into the neurons increased with increasing the duration of nicotine exposure and its concentrations. The maximal increase of the KCl-stimulated (45)Ca(2+) influx was found 24 h after the initiation of exposure and thereafter maintained up to 72 h. This enhancement of KCl-induced (45)Ca(2+) influx after 72-h exposure to 0.1 mum nicotine was completely abolished by concomitant exposure with mecamylamine, an inhibitor for nnAChRs. Only the component of the KCl-induced (45)Ca(2+) influx observed after long term exposure to nicotine, which was sensitive to nifedipine, an inhibitor of L-type VDCCs, was facilitated, while the (45)Ca(2+) influx through P/Q- and N-type VDCCs showed no changes. Moreover, enhanced immunoreactivity against antibody for the alpha(1C) subunit of L-type VDCCs was recognized, whereas no changes in immunoreactivities against antibodies for alpha(1A) and alpha(1B) subunits of other types of VDCCs were noted. In addition, a Western blot analysis showed an increase of immunoreactivities against antibodies for alpha(1D) and alpha(2)/delta(1), and expression of mRNA for L-type VDCC subunit, alpha(1F), was also enhanced, although beta(4) mRNA expression was not changed. Whole cell patch clamp analysis revealed that the increase of the amplitude of Ba(2+) currents was also recognized in the neurons exposed to nicotine, and nicardipine reduced this increased amplitude to the level of the amplitude detected in nontreated neurons with nicardipine. The up-regulation of alpha(4) and beta(2) subunits, but not the alpha(3) subunit of nnAChRs, was also noted after the nicotine exposure when examining by the Western blot analysis. Taken together, these results indicate that the long term exposure of the neurons to a low concentration of nicotine induces both increased (45)Ca(2+) influx through up-regulated L-type VDCCs and nnAChR up-regulation.
authors.affiliation: Department of Pharmacology, Kawasaki Medical School, Matsushima, Kurashiki 701-0192, Japan.
authors.forename: Masashi
authors.initials: M
authors.lastname: Katsura
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Mohri
authors.forename: Keijiro
authors.initials: K
authors.lastname: Shuto
authors.forename: Yan
authors.initials: Y
authors.lastname: Hai-Du
authors.forename: Taku
authors.initials: T
authors.lastname: Amano
authors.forename: Atsushi
authors.initials: A
authors.lastname: Tsujimura
authors.forename: Masashi
authors.initials: M
authors.lastname: Sasa
authors.forename: Seitaro
authors.initials: S
authors.lastname: Ohkuma
mesh_terms: D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000818:Animals; D001464:Barium / Q000378:metabolism; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000378:metabolism*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005731:Ganglionic Stimulants / Q000494:pharmacology; D015151:Immunoblotting; D007700:Kinetics; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009538:Nicotine / Q000494:pharmacology*; D011189:Potassium Chloride / Q000494:pharmacology; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D015854:Up-Regulation; D014700:Verapamil / Q000494:pharmacology
chemical_list: D002120:Calcium Channel Agonists; D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D005731:Ganglionic Stimulants; D012333:RNA, Messenger; D001464:Barium; D011189:Potassium Chloride; D009538:Nicotine; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D014700:Verapamil; D002118:Calcium