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

1. Cerebellar granule neurons (CGNs) possess a standing outward potassium c urrent (I-K(SO)) which shares many similarities with current through the tw o-pore domain potassium channel TASK-1 and which is inhibited following act ivation of muscarinic acetylcholine receptors. 2. The action of muscarine on I-K(SO) was unaffected by the M-2 receptor an tagonist methoctramine (100 nM) but was blocked by the M-3 antagonist zamif enacin, which, at a concentration of 100 nM, shifted the muscarine concentr ation-response curve to the right by around 50-fold. 3. Surprisingly, M-3 receptor activation rarely produced a detectable incre ase in [Ca2+](i) unless preceded by depolarization of the cells with 25 mM K+. Experiments with thapsigargin and ionomycin suggested that the endoplas mic reticulum Ca2+ stores in CGNs were depleted at rest. In contrast, cereb ellar glial cells in the same fields of cells possessed substantial endopla smic reticulum Ca2+ stores at rest. 4. Pretreatment of the cells with BAPTA AM, thapsigargin or the phospholipa se C (PLC) inhibitor U-73122 all blocked the muscarine-induced Ca2+ 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 rece ptors both to inhibit I-K(SO) and to mobilize Ca2+ from intracellular store s in CGNs. While the mobilization of Ca2+ occurs through activation of PLC, this does not seem to be the primary mechanism underlying muscarinic inhib ition of I-K(SO).