Activation of the R-type and L-type voltage-gated Ca²⁺ channels contributes to Ca²⁺-dependent GnRH release in immortalized hypothalamic GnRH neurons

Abstract

Cellular functions of the GnRH neuron have been widely studied by using its clonal cell line GT1-7. To further characterize GT1-7 cells, we investigated the diversity of voltage-gated Ca²⁺ channels in GT1-7 cells with perforated-patch clamp and RT-PCR. An R-type Ca²⁺ channel blocker SNX-482 inhibited the Ca²⁺ currents by 76% in all cells examined (n=9). A T-type Ca²⁺ channel blocker Ni²⁺ inhibited the Ca²⁺ currents by 13% in all cells examined (n=9). An L-type Ca²⁺ channel blocker nimodipine inhibited the Ca²⁺ currents by 18% in 5 out of 11 cells examined. An N-type Ca²⁺ channel blocker ω-conotoxin-GVIA inhibited the Ca²⁺ currents by 14% in 3 out of 20 cells examined. A P/Q-type Ca²⁺ channel blocker ω-agatoxin-IVA had no effect on the currents (n=9). The contribution of the L-type was much greater in the current elicited by ramp depolarization than in that evoked by a square pulse. On the contrary, the contribution of the R-type was less in the current by ramp depolarization. RT-PCR revealed that GT1-7 cells expressed the α_1B, _1D, _1E and _1H subunit mRNA. Furthermore, SNX-482 and nifedipine inhibited the high K⁺ induced increase in the intracellular Ca²⁺ concentration and GnRH release. These results suggest that GT1-7 cells express R-, L-, N- and T-type voltage-gated Ca²⁺ channels and the R-type is a major current component, and the L- N- and T-type are minor ones. The R- and L-type Ca²⁺ channels play a critical role in the regulation of Ca²⁺-dependent GnRH release. [Jpn J Physiol 54 Suppl:S219 (2004)]