The role of Ca2+ stores in the muscarinic inhibition of the K+ current IK(SO) in neonatal rat cerebellar granule cells

1. Cerebellar granule neurons (CGNs) possess a standing outward potassium current (I(K(SO))) which shares many similarities with current through the two-pore domain potassium channel TASK-1 and which is inhibited following activation of muscarinic acetylcholine receptors. 2. The action of muscarine on I(K(SO)) was unaffected by the M(2) receptor antagonist methoctramine (100 nM) but was blocked by the M(3) antagonist zamifenacin, which, at a concentration of 100 nM, shifted the muscarine concentration-response curve to the right by around 50-fold. 3. Surprisingly, M(3) receptor activation rarely produced a detectable increase in [Ca(2+)](i) unless preceded by depolarization of the cells with 25 mM K(+). Experiments with thapsigargin and ionomycin suggested that the endoplasmic reticulum Ca(2+) stores in CGNs were depleted at rest. In contrast, cerebellar glial cells in the same fields of cells possessed substantial endoplasmic reticulum Ca(2+) stores at rest. 4. Pretreatment of the cells with BAPTA AM, thapsigargin or the phospholipase C (PLC) inhibitor U-73122 all blocked the muscarine-induced Ca(2+) signal but had little or no effect on muscarinic inhibition of I(K(SO)) Raising [Ca(2+)](i) directly with ionomycin caused a small but significant inhibition of I(K(SO)). 5. It is concluded that muscarine acts on M(3) muscarinic acetylcholine receptors both to inhibit I(K(SO)) and to mobilize Ca(2+) from intracellular stores in CGNs. While the mobilization of Ca(2+) occurs through activation of PLC, this does not seem to be the primary mechanism underlying muscarinic inhibition of I(K(SO)).