Effect of tacrine on intracellular calcium in cholinergic SN56 neuronal cells.
We have found earlier that the depolarization-induced release of acetylcholine from the brain could be inhibited by tacrine (tetrahydroaminoacridine) but the mechanism of this action of tacrine was not clarified (S. Tucek, V. Dolezal, J. Neurochem. 56 (1991) 1216). We have now investigated whether tacrine has an effect on the changes in the intracellular concentration of calcium ions ([Ca2+]i) induced by depolarization. Experiments were performed on the cholinergic SN56 neuronal cell line with Fura-2 fluorescence technique of calcium imaging. The depolarization by 71 mmol/l K+ evoked minimum increases of [Ca2+]i up to day 5 in culture. Then the response gradually increased and reached a plateau after 7 days in culture. A similar time course was observed for acetylcholinesterase activity. The effect of K+ ions was concentration-dependent and the concentration of 71 mmol/l K+ evoked maximum [Ca2+]i responses. The increases of [Ca2+]i did not occur in the absence of extracellular calcium. They were mediated by high voltage-activated calcium channels of the L-type and the N-type. Nifedipine (2 micromol/l; L-type calcium channel blocker) and omega-conotoxin GVIA (100 nmol/l; N-type calcium channel blocker) diminished the response to 71 mmol/l K+ by 53% and 39%, respectively, and their effects were additive (decrease to 8% of controls). Non-selective inorganic blocker of voltage-activated calcium channels LaCl3 (0.1 mmol/l) decreased the response by 83%. Tacrine attenuated the [Ca2+]i response in a concentration-dependent manner. At a concentration of 10 micromol/l it inhibited the [Ca2+]i response by 55% and its inhibitory effect was additive with that of omega-conotoxin GVIA but not with that of nifedipine. An equimolar concentration of paraoxon, an irreversible inhibitor of cholinesterases, had no influence on [Ca2+]i response. Tacrine exhibited the same inhibitory effect when paraoxon was present. In conclusion, our data indicate that high-voltage-activated calcium channels of the L-type and the N-type are both present in the SN56 cells but that they are fully expressed only after 6-7 days in culture. Tacrine attenuates the influx of calcium by inhibiting the L-type calcium channels. This inhibitory effect is not a consequence of the anticholinesterase activity of tacrine. The finding that low micromolar concentrations of tacrine may interfere with calcium-dependent events is likely to be of importance for the evaluation of the therapeutic potential of the drug.
Dolezal
V
V
Institute of Physiology, Academy of Sciences of the Czech Republic, Vídenská, Prague. dolezal@biomed.cas.cz
Lisá
V
V
Tucek
S
S
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D002459:Cell Fusion; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D007425:Intracellular Membranes / Q000378:metabolism*; D051379:Mice; D018721:Muscarinic Agonists / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D018722:Nicotinic Agonists / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D013619:Tacrine / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D002121:Calcium Channel Blockers; D002800:Cholinesterase Inhibitors; D018721:Muscarinic Agonists; D018722:Nicotinic Agonists; D013619:Tacrine; D011189:Potassium Chloride; D002217:Carbachol; D000109:Acetylcholine; D002118:Calcium