Our previous study demonstrated that FSH-induced immediate Ca2+ influx in rat Sertoli cells (SCs) is mediated by the Gαh/phospholipase C-δ1 (PLC-δ1) signaling pathway. As to which Ca2+ channel is responsible for such Ca2+ influx was not understood. In this study, thapsigargin triggered an instore calcium release and evoked a 1.5-fold elevation of intracellular Ca2+ in Ca2+-free media, whereas FSH exhibited no effect. The readdition of CaCl2 (2.5 mM) to FSH-pretreated or thapsigargin-sensitized SCs in Ca2+-free media immediately elicited a rapid Ca2+ influx or a 2-fold increase of second intracellular Ca2+ elevation, respectively. The addition of Ca2+ chelator EGTA (0.2 mM) reduced the FSH-induced elevation of intracellular Ca2+ in SCs incubated with CaCl2. However, pretreatment with dantrolene (25 μM), which inhibits in-store calcium release, did not affect the FSH-induced elevation of intracellular Ca 2+. NiCl2 (10 μM), a T-type calcium channel blocker, abolished the FSH-induced SC Ca2+ influx. Furthermore, mibefradil (10 and 100 μM), another specific blocker for T-type Ca2+ channels, dose-dependently suppressed the FSH-induced Ca2+ influx. In contrast, nifedipine (10 and 50 μM) or ω-conotoxin GVIA (100 and 500 nM), blocker of L- or N-type Ca2+ channels, respectively, did not affect the FSH-induced SC Ca2+ influx. On the other hand, FSH-induced Ca2+ influx was significantly reduced by pretreatment of SCs with myristoylated synthetic peptide (0.1 and 1 μM) of PLC-δ1 fragment TIPWNSLKQGYRHVHLL but not affected by 2′,5′-dideoxyadenosine (3 and 15 μM), a selective inhibitor of adenylate cyclase. In conclusion, the FSH-induced Gαh/PLC-δ1 pathway-dependent Ca2+ influx of rat SCs is mediated by T-type Ca2+ channels and independent of in-store calcium release.
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