Adenosine regulates a chloride channel via protein kinase C and a G protein in a rabbit cortical collecting duct cell

We examined the regulation by adenosine of a 305-pS chloride (Cl-) channel in the apical membrane of a continuous cell line derived from rabbit cortical collecting duct (RCCT-28A) using the patch clamp technique. Stimulation of A1 adenosine recep-tors by N'-cyclohexyladenosine (CHA) activated the channel in cell-attached patches. Phorbol 12,13-didecanoate and 1-oleoyl 2-acetylglycerol, activators of protein kinase C (PKC), mim-icked the effect of CHA, whereas the PKC inhibitor H7 blocked the action ofCHA. Stimulation ofAl adenosine recep-tors also increased the production of diacylglycerol, an activa-tor of PKC. Exogenous PKC added to the cytoplasmic face of inside-out patches also stimulated the Cl- channel. Alkaline phosphatase reversed PKC activation. These results show that stimulation of Al adenosine receptors activates a 305-pS Cl0 channel in the apical membrane by a phosphorylation-depen-dent pathway involving PKC. In previous studies, we showed that the protein G,_3 activated the 305-pS Cl- channel (Schwie-bert et al. 1990. J. Biol. Chem. 265:7725-7728). We, therefore, tested the hypothesis that PKC activates the channel by a G protein-dependent pathway. In inside-out patches, pertussis toxin blocked PKC activation of the channel. In contrast, H7 did not prevent G protein activation of the channel. We con-clude that adenosine activates a 305-pS Cl- channel in the api-cal membrane of RCCT-28A cells by a membrane-delimited pathway involving an Al adenosine receptor, phospholipase C, diacylglycerol, PKC, and a G protein. Because we have shown, in previous studies, that this Cl- channel participates in the regulatory volume decrease subsequent to cell swelling, adeno-sine release during ischemic cell swelling may activate the ClO channel and restore cell volume. (J. Clin. Invest. 1992.89:834-841.) Key words: cell culture * intercalated cells * ion channels. RCCI-28A cells * regulatory volume decrease signal trans-ductio