Differential coupling of G-protein-linked receptors to Ca2+ mobilization through inositol(1,4,5)trisphosphate or ryanodine receptors in cerebellar granule cells in primary culture

Rat cerebellar granule cells in primary culture possess muscarinic, metabotropic glutamatergic, histaminergic and alpha-adrenergic receptors which couple to phosphoinositide-specific phospholipase C. We have determined the ability of these receptors to elevate inositol(1,4,5)trisphosphate and to release intracellular calcium, in order to establish the correlation between these two responses. In resting cerebellar granule cells, only the muscarinic agonist carbachol evoked significant increases in both inositol(1,4,5)trisphosphate and cytoplasmic free Ca2+. Mild depolarization (20 mM KCl) enhanced inositol(1,4,5)trisphosphate elevation by carbachol and histamine, but not by noradrenaline or the metabotropic glutamate agonist 1S,3R ACPD. In contrast, Ca2+-release responses were modified differently by 20 mM KCl-depolarization: the responses to carbachol, histamine and 1S,3R ACPD, but not the responses to noradrenaline, were markedly enhanced. The contribution of ryanodine-sensitive Ca2+-release channels (ryanodine receptors) to the calcium release signal in depolarized cells was determined. Ryanodine (10 mu M) inhibited most effectively the cytoplasmic Ca2+ elevation evoked by 1S,3R ACPD (> 90%), while Ca2+ release upon stimulation by carbachol and histamine was only inhibited by approximate to 60% and remained larger than in the absence of KCl. Our data are consistent with a specific coupling between metabotropic glutamate receptors and ryanodine-sensitive Ca2+-release channels which may not require generation of inositol(1,4,5)trisphosphate.