Muscarinic receptor activation inhibits AVP-induced water flow in rabbit cortical collecting ducts
Snyder, H.M.; Fredin, D.M.; Breyer, M.D.
American Journal of Physiology 260(6 Pt 2): F929-F936
1991
ISSN/ISBN: 0002-9513 PMID: 1647693 Document Number: 374421
We examined the effect of carbachol, an acetylcholine analogue, on hydraulic conductivity (Lp) response to 10 .mu.U/ml arginine vasopressin (AVP) in rabbit cortical collecting duct (CCD). In CCDs in which water flow had been established with AVP, subsequent addition of carbachol caused Lp (.times. 10-7 cm .cntdot. atm-1 .cntdot. s-1) to fall from 251 .+-. 32 to 146 .+-. 19. Carbachol washout resulted in recovery of Lp to 217 .+-. 38. In CCDs in which water flow had been established using 10-4 M 8-chlorophenylthioadenosine 3',5'-cyclic monophosphate (8-CPT-cAMP), addition of carbachol had no effect. These posttreatment studies suggest that carbachol's effects on modulating established water flow occur at a "pre-cAMP" step. With carbachol added first, AVP-induced Lp was reduced from 233 .+-. 24 (controls) to 105 .+-. 19 (carbachol-pretreated). Pretreatment with 10-6 M atropine, a muscarinic receptor antagonist, totally reversed the inhibitory effect of carbachol, consistent with a receptor-mediated effect of carbachol. Carbachol pretreatment also inhibited 8-CPT-cAMP-induced Lp, indicating that carbachol's effects also occur at a "post-cAMP" step. Pretreatment with 10-7 M staurosporine, a protein kinase C (PKC) inhibitor, reversed inhibitory effect of carbachol on AVP-induced Lp (193 .+-. 26), suggesting that carbachol's effects are mediated by PKC. Intracellular calcium concentration ([Ca2+]i) was measured in fura-2-loaded CCDs. Carbachol also increased [Ca2+]i from 229 .+-. 120 to 389 .+-. 160 nM. In 0 Ca2+ bath with EGTA, carbachol caused a spikelike increase in [Ca2+]i from 92 .+-. 21 to 130 .+-. 16 nM, indicating release of Ca2+ from intracellular stores consistent with carbachol's effects occurring via phosphoinositide (PI) hydrolysis. We conclude that muscarinic receptor activation by carbachol directly inhibits AVP-induced Lp and that muscarinic receptors on CCDs probably signal through PI hydrolysis.