Intracellular Ca2+ stores regulate muscarinic receptor stimulation of phospholipase C in cerebellar granule cells

Muscarinic receptor activation of phosphoinositide phospholipase C (PLC) ha s been examined in rat cerebellar granule cells under conditions that modif y intracellular Ca2+ stores. Exposure of cells to medium devoid of Ca2+ for various times reduced carbachol stimulation of PLC with a substantial loss (88%) seen at 30 min. A progressive recovery of responses was observed fol lowing the reexposure of cells to Ca2+-containing medium (1,3 mM). However, these changes did not appear to result exclusively from changes in the cyt osolic Ca2+ concentration ([Ca2+](i)), which decreased to a lower steady le vel (similar to 25 nM decrease in 1-3 min after extracellular omission) and rapidly returned (within 1 min) to control values when extracellular Ca2was restored. Only after loading of the intracellular Ca2+ stores through a transient 1-min depolarization of cerebellar granule cells with 40 mM KCI, followed by washing in nondepolarizing buffer, was carbachol able to mobil ize intracellular Ca2+ However, the same treatment resulted in an 80% enhan cement of carbachol activation of PLC. In other experiments, partial deplet ion of the Ca2+ stores by pretreatment of cells with thapsigargin and caffe ine resulted in an inhibition (18 and 52%, respectively) of the PLC respons e, Furthermore, chelation of cytosolic Ca2+ with BAPTA/AM did not influence muscarinic activation of PLC in either the control or predepolarized cells . These conditions, however, inhibited both the increase in [Ca2+](i) and t he PLC activation elicited by 40 mM KCI and abolished carbachol-induced int racellular Ca2+ release in predepolarized cells. Overall, these results sug gest that muscarinic receptor activation of PLC in cerebellar granule cells can be modulated by changes in the loading state of the Ca2+ stores.