Vol. 290, Issue 2, 725-730, August 1999

Studies on Maitotoxin-Induced Intracellular Ca²⁺ Elevation in Chinese Hamster Ovary Cells Stably Transfected with cDNAs Encoding for L-Type Ca²⁺ Channel Subunits^1,2

Mauro Cataldi, Agnese Secondo, Angela D'Alessio, Maurizio Taglialatela, Franz Hofmann, Norbert Klugbauer, Gianfranco Di Renzo and Lucio Annunziato

Section of Pharmacology, Department of Neuroscience, School of Medicine, University of Naples Federico II, Naples, Italy (M.C., A.S., A.D., M.T., L.A.); Institute of Pharmacology and Toxicology, Technischen Universität München, München, Germany (F.H., N.K.); and School of Pharmacy, University of Catanzaro, Italy (G. Di. R.)

The aim of the present study was to characterize the role played by different L-type Ca²⁺ channel subunits in [Ca²⁺]_i increase induced by maitotoxin (MTX). In the presence of 5 mM extracellular K⁺, MTX (0.01-0.5 ng/ml) induced a significant concentration-dependent increase in Fura-2-monitored [Ca²⁺]_i in single Chinese hamster ovary (CHO) cells expressing the _1c (CHOC9 cells) or the _1c32 (CHOC932/4 cells) subunits of voltage-gated Ca²⁺ channels (VGCCs), whereas the effect was much reduced in wild-type CHO cells lacking VGCCs. In addition, MTX effect on CHOC9, CHOC932/4, and GH₃ cells (0.01-0.1 ng/ml) was inhibited by the selective L-type Ca²⁺ channel entry-blocker nimodipine (10 µM); a nimodipine-insensitive component was still present, particularly at high (>1 ng/ml) toxin concentrations. In CHOC932/4 cells, depolarizing concentrations of extracellular K⁺ (55 mM) reinforced the [Ca²⁺]_i increase induced by MTX (0.1 ng/ml), and this effect was prevented by nimodipine (10 µM). Finally, patch-clamp experiments in CHOC932/4 cells showed that low MTX concentrations (0.03 ng/ml) induced the occurrence of an inward current at 60 mV, which was completely prevented by Cd²⁺ (100 µM) and by nimodipine (10 µM), whereas the same dihydropyridine concentration (10 µM) failed to prevent the electrophysiological effects of a higher toxin concentration (3 ng/ml). In conclusion, the results of the present study showed that MTX-induced [Ca²⁺]_i elevation involves two components: 1) an action on L-type VGCCs at the pore-forming _1c subunit level, which is responsible for the greatest rise of [Ca²⁺]_i; and 2) a VGCC-independent mechanism that is present both in excitable and in nonexcitable cells and is responsible for a lower elevation of [Ca²⁺]_i.