Gonadotropin-Releasing Hormone Stimulates Luteinizing Hormone Secretion By Extracellular Calcium-Dependent and -Independent Mechanisms*

Abstract

The dependence of LH responses to GnRH on extracellular calcium was investigated in cultured rat pituitary cells exposed to GnRH for 3 h in static culture or for 2 min during column perifusion. During static culture in normal medium, LH release was stimulated by GnRH with an ED50 of 0.3 nM and by K+ with an ED50 of 32 mM. Incubation in Ca2+-deficient (no added Ca2+) or Ca2+-free medium (containing 100 microM EGTA) substantially decreased, but did not abolish, the LH responses to 10 and 100 nM GnRH, whereas K+-induced LH release was almost completely abolished in Ca2+-deficient medium. The Ca2+ channel agonist (BK 8644) and antagonists (nifedipine, nicardipine, verapamil, and Co2+) respectively enhanced or reduced the LH responses to both GnRH and K+. However, the calcium antagonists completely abolished the LH response to depolarization by K+, but only partially inhibited the LH response to GnRH, confirming the existence of a significant component of GnRH action that is not dependent on extracellular Ca2+. In perifused pituitary cells, exposure to Ca2+-deficient medium or normal medium containing 5 mM EGTA or 5 mM EDTA, reduced the initial rapid LH response to 2-min pulses of 10 nM GnRH and abolished the second phase of LH release. Reintroduction of Ca2+-containing medium at the end of the GnRH pulse caused recovery of the second phase of LH secretion, demonstrating that influx of extracellular Ca2+ is not required for the early phase of the LH response to GnRH but, rather, appears to be essential for its prolongation. The release of LH in response to arachidonic acid, which has been implicated in the mechanism of the secretory action of GnRH, was completely independent of extracellular Ca2+ and unaffected by addition of 10 nM BK 8644. These observations indicate that the initiation of the secretory response to GnRH is largely independent of calcium entry, whereas the prolongation of gonadotropin secretion is maintained by calcium influx, in part through voltage-sensitive calcium channels. The role of arachidonic acid metabolites in GnRH action is probably related to the calcium-independent component of GnRH-induced LH secretion. Since GnRH is secreted episodically and for short periods, much of its physiological action on pulsatile gonadotropin release could be independent of calcium influx from the extracellular fluid.

Dependence of hormone secretion on activation-inactivation kinetics of voltage-sensitive Ca2+ channels in pituitary gonadotrophs.

“…The relationships between the activation status of voltage-sensitive Ca2+ channels and secretory responses were analyzed in perifused rat gonadotrophs during stimulation by high extraceflular K+ concentration ([K+] entry by at least two pathways, the best-defined of which is through VSCCs that are modulated by dihydropyridine agonist and antagonist analogs (1,3,9). The residual, dihydropyridine-insensitive entry ofCa2+ may occur through another type of VSCC or through receptor-operated Ca2" channels.…”

“…Agonist stimulation during the steady-state phase of the [K+] pulse or after repetitive stimulation by high [ (1,2), and the sensitivity of these effects to dihydropyridine agonist and antagonist analogs (1-3), have provided indirect evidence for the presence of voltage-sensitive Ca2" channels (VSCCs) in pituitary gonadotrophs. Electrophysiological studies (4) have confirmed that pituitary gonadotrophs, like corticotrophs (5) and pituitary tumor cells (6), possess a fast, transient (T-type) channel and a slower channel that is dihydropyridinesensitive.…”

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ABSTRACTThe relationships between the activation status of voltage-sensitive Ca2+ channels and secretory responses were analyzed in perifused rat gonadotrophs during stimulation by high extraceflular K+ concentration ([K+] entry by at least two pathways, the best-defined of which is through VSCCs that are modulated by dihydropyridine agonist and antagonist analogs (1,3,9). The residual, dihydropyridine-insensitive entry ofCa2+ may occur through another type of VSCC or through receptor-operated Ca2" channels.

To further delineate the properties and the role of these channels in stimulus-secretion coupling, we have defined the temporal course of LH secretion evoked by high [K+] tTo whom reprint requests should be addressed at: Building 10, Room B1-L400, National Institutes ofHealth, Bethesda, MD 20892.

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Involvement of voltage-dependent calcium channels (VDCC) in the action of GnRH on GtH release in common carp (Cyprinus carpio L): comparison with K+ action

“…It is well established that calcium ions (Ca 2 +) play the role of second messenger in GnRH action in mammals ; Huckle and Conn, 1988 However, the spike phase of the secretory response to GnRH is independent of Ca 2 + influx through VDCC (Hansen et al, 1987;Chang et al, 1986Chang et al, , 1988Smith et al, 1987;Davidson et al, 1988;Smith et al, 1989). There could be an influx of Ca 2 + through channels other than VDCC, the so-called receptor operated channels (ROC) Davidson et al, 1988;Smith et al, 1989) or mobilization of Ca 2+ from intracellular stores Hansen et al, 1987 (1989) found that D-600 had no effect on spontaneous and GnRH-stimulated GtH release in African catfish, while such an inhibition was observed when nifedipine (another class of VDCC blockers) was used.…”

“…Chang et al (1988) (Hansen et al, 1987;Smith et al, 1987;Chang et al, 1988;Davidson et al, 1988;Smith et al, 1989). If the first (spike) phase of the secretory response to GnRH is concerned there is some controversy about the pathways as well as the sources of the Ca 2+ .…”