High expression of the R-type voltage-gated Ca2+ channel and its involvement in Ca2+-dependent gonadotropin-releasing hormone release in GT1-7 cells.
The GT1 cell has been widely used as a model cell to study cellular functions of GnRH neurons. Despite the importance of Ca(2+) channels, little is known except for L- and T-type Ca(2+) channels in GT1 cells. Therefore, we studied the diversity of voltage-gated Ca(2+) channels in GT1-7 cells with perforated-patch clamp and RT-PCR. An R-type Ca(2+) channel blocker, SNX-482, inhibited the Ca(2+) currents by 75.6% in all cells examined (n = 9). A T-type Ca(2+) channel blocker, Ni(2+), inhibited the Ca(2+) currents by 12.6% in all cells examined (n = 9). An L-type Ca(2+) channel blocker, nimodipine, inhibited the Ca(2+) currents by 17.9% in five of 11 cells examined. When using Ba(2+) as a charge carrier, another dihydropyridine antagonist, nifedipine, clearly inhibited the currents by 12.1% in all cells examined (n = 16). An N-type Ca(2+) channel blocker, omega-conotoxin-GVIA, inhibited the Ca(2+) currents by 13.8% in three of 20 cells examined. A P/Q type Ca(2+) channel blocker, omega-agatoxin-IVA, had no effect on the currents (n = 9). RT-PCR revealed that GT1-7 cells expressed the alpha(1B), alpha(1D), alpha(1E), and alpha(1H) subunit mRNA. Furthermore, SNX-482 and nifedipine inhibited the high K(+)-induced increase in the intracellular Ca(2+) concentration and GnRH release. omega-Conotoxin-GVIA and omega-agatoxin-IVA had no effect. These results suggest that GT1-7 cells express R-, L-, N-, and T-type voltage-gated Ca(2+) channels; the R-type was a major current component, and the L-, N-, and T-types were minor ones. The R- and L-type Ca(2+) channels play a critical role in the regulation of Ca(2+)-dependent GnRH release.
Watanabe
Miho
M
Department of Physiology, Nippon Medical School, Bunkyo, Tokyo 113-8602 Japan.
Sakuma
Yasuo
Y
Kato
Masakatsu
M
D000818:Animals; D001464:Barium / Q000378:metabolism; D002118:Calcium / Q000032:analysis / Q000378:metabolism / Q000494:pharmacology*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020908:Calcium Channels, R-Type / Q000235:genetics* / Q000502:physiology*; D002461:Cell Line, Transformed; D004553:Electric Conductivity; D015870:Gene Expression; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D009532:Nickel / Q000494:pharmacology; D009543:Nifedipine / Q000494:pharmacology; D009553:Nimodipine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013111:Spider Venoms / Q000494:pharmacology; D020866:omega-Conotoxin GVIA / Q000494:pharmacology
D002121:Calcium Channel Blockers; D020908:Calcium Channels, R-Type; D012333:RNA, Messenger; C115521:SNX 482; D013111:Spider Venoms; D001464:Barium; D007987:Gonadotropin-Releasing Hormone; D009553:Nimodipine; D009532:Nickel; D020866:omega-Conotoxin GVIA; D009543:Nifedipine; D011188:Potassium; D002118:Calcium