Acetylcholine increases the free intracellular calcium concentration in podocytes in intact rat glomeruli via muscarinic M-5 receptors

The effects of acetylcholine (ACh) on the free intracellular calcium concen tration ([Ca2+](i)) of microdissected glomeruli were investigated using fur a-2 fluorescence digital imaging and two-photon confocal microscopy. ACh ca used a concentration-dependent [Ca2+](i) increase with an initial peak foll owed by a sustained plateau, which was suppressed by reduced extracellular Ca2+ concentrations. The [Ca2+](i) plateau was not affected by the L-type C a2+ channel blocker nicardipine, whereas gadolinium and lanthanum (both at 1 muM) blocked the plateau. Diphenylacetoxy-N-methylpiperidine methiodide ( 100 nM), an M-3/M-5 receptor antagonist, and pirenzepine (1 muM), an M-1 re ceptor antagonist, completely inhibited The effect of ACh. [Ca2+](i) measur ements using two-photon excitation of fluo-3 and staining of the cells with calcein/ arcetoxymethyl ester, for observation of the capillary network to gether with the glomerular cells, showed that [Ca2+](i) was increased in si ngle podocytes. Immunohistochemical studies did not demonstrate M-3 recepto r expression in glomerular cells. M-1 receptors could be detected only in t he parietal sheet of Bowman's capsule, whereas M-5 receptors were found onl y in podocytes. The data show that Ach increases [Ca2+](i) in podocytes of intact glomeruli, most likely via muscarinic M-5 receptors.