Caffeine-stimulated GTH-II release involves Ca²⁺stores with novel properties

Abstract

Modulation of Ca(2+) stores with 10 mM caffeine stimulates robust secretion of gonadotropin (GTH-II) from goldfish gonadotropes. Although both endogenous forms of gonadotropin-releasing hormone (GnRH) utilize a common intracellular Ca(2+) store, sGnRH, but not cGnRH-II, uses an additional caffeine-sensitive mechanism. We examined caffeine signaling by using Ca(2+) imaging, electrophysiology, and cell-column perifusion. Although caffeine inhibited K+ channels, this action appeared to be unrelated to caffeine-induced GTH-II release, because the latter was insensitive to tetraethylammonium. The effects of caffeine also were not mediated by the cAMP/protein kinase A pathway. Instead, caffeine-evoked GTH-II responses were Ca(2+) signal dependent because they were abolished by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid loading. Caffeine generated localized Ca(2+) signals that began near secretory granules. Surprisingly, caffeine-stimulated GTH-II release was insensitive to 100 microM ryanodine and, unlike GnRH action, was unaffected by inhibitors of voltage-gated Ca(2+) channels or sarco(endo)plasmic reticulum Ca(2+)-ATPases. Collectively, these data indicate that caffeine-stimulated GTH-II release is not mediated by typical agonist-sensitive Ca(2+) stores found in endoplasmic reticulum.

Ryanodine receptors in human pancreatic β cells: localization and effects on insulin secretion

“…The second assumption is challenged by the finding that ryanodine stimulates insulin release under basal conditions and not when β cells are stimulated with glucose or KCl. The observation that nanomolar concentrations of ryanodine led to an increase in cytosolic Ca 2+ concentrations sufficient to release insulin from human β cells supports the controversial hypothesis that strategically located intracellular Ca 2+ stores can stimulate secretion from endocrine cells (15,22,(32)(33)(34). The ability of RyRs to generate large enough Ca 2+ release events to drive exocytosis is consistent with the large conductance of these channels (8,17,35).…”

“…In pituitary cells, increased hormone release was seen with nanomolar ryanodine concentrations, but not micromolar ryanodine concentrations (22,23,39). Although a Ca 2+ store associated with the secretory granules can control exocytosis in pituitary gonadotropes, it appears to be ryanodineinsensitive (15). In other cell types, Ca 2+ flux in or around secretory granules is thought to control the rapid alkalinization and decondensation of secretory granule contents that immediately precedes exocytosis (40), either directly or by acting on a Ca 2+ -activated vesicle cation channel similar to synaptophysin (41).…”

“…Ca 2+ mobilization by GLP-1 in human β cells and rodent cell lines is completely abolished by inhibiting RyR (7,14). The majority of studies examining the RyR pathway have relied on caffeine, which reportedly modulates IP 3 receptors, phosphodiesterase, phospholipase C, voltage-gated Ca 2+ channels, voltage-gated K + channels, and K ATP channels (15)(16)(17). By definition, the plant alkaloid ryanodine is the most specific probe for RyR.…”

A Gonadotropin‐Releasing Hormone Insensitive, Thapsigargin‐Sensitive Ca2+ Store Reduces Basal Gonadotropin Exocytosis and Gene Expression: Comparison with Agonist‐Sensitive Ca2+ Stores

“…The possibility that long‐term changes in [Ca 2+ ] L may be generated by our treatments, along with the known roles for [Ca 2+ ] L in the secretory pathway, suggests that [Ca 2+ ] L signalling should be considered in further experiments. Future studies should directly examine organelle Ca 2+ levels in the presence of Tg, Ry and GnRH, as well as the relative subcellular localization of the various pharmacologically distinct Ca 2+ stores shown here and in previous studies (22, 23, 36). Regardless if ER [Ca 2+ ] L is involved, this is likely compartmentalized in the goldfish gonadotropes to allow for the differential Tg and Ry actions observed in this study.…”

“…2). Similarly, Tg did not affect the GTH‐II release evoked by 10 m m caffeine, which in goldfish gonadotropes mobilizes intracellular Ca 2+ directly from stores that are preferentially involved in sGnRH signalling (22, 36). These data demonstrate that Tg‐sensitive Ca 2+ stores exist in goldfish gonadotropes, but that they are not involved in acute GnRH‐ or caffeine‐stimulated GTH‐II release.…”