A9 FIBROBLASTS TRANSFECTED WITH THE M3-MUSCARINIC-RECEPTOR CLONE EXPRESS A CA2+ CHANNEL ACTIVATED BY CARBACHOL, GTP AND GDP

Muscarinic m3 receptor-mediated changes in cytosolic Ca2+ concentratio n ([Ca2+](1)) occur by activation of Ca2+ release channels present in the endoplasmic reticulum membrane and Ca2+ entry pathways across the plasma membrane. In this report we demonstrate the coupling of m3 musc arinic receptors to the activation of a voltage-insensitive, cation-se lective channel of low conductance (3.2 +/- 0.6 pS; 25 mM Ca2+ as char ge carrier) in a fibroblast cell line expressing m3 muscarinic recepto r clone (A9m3 cells). Carbachol (CCh)-induced activation of the cation -selective channel occurred both in whole cell and excised membrane pa tches (CCh on the external side), suggesting that the underlying mecha nism involves receptor-channel coupling independent of intracellular m essengers. in excised inside-out membrane patches from nonstimulated A 9m3 cells GTP (10 mu M) and GDP (10 mu M) activated cation-selective c hannels with conductances of approximately 4.3 and 3.3 pS, (25 mM Ca2 as charge carrier) respectively. In contrast, ATP (10 mu M), UTP (10 mu M) or CTP (10 mu M) failed to activate the channel. Taken together, these results suggest that carbachol and guanine nucleotides regulate the activation of a cation channel that conducts calcium.