Journal of the American Society of Nephrology, Vol 4, 195-205, Copyright © 1993 by American Society of Nephrology

REGULAR ARTICLES

Cholinergic inhibition of Na-K-ATPase via activation of protein kinase C in Madin-Darby canine kidney cells

LC Garg, PK Saha and D Mohuczy-Dominiak
Department of Pharmacology and Therapeutics, University of Florida College of Medicine, Gainesville.

Recently, it was reported that muscarinic-type cholinergic receptors coupled to the phosphoinositide messenger system are present in the rabbit inner medullary collecting duct and Madin-Darby canine kidney (MDCK) cells. The receptor density in MDCK cells is 50 times more than that in inner medullary collecting duct cells. To examine if muscarinic receptor activation influences Na-K-ATPase, the effects of a cholinergic agonist, carbachol, on Na-K-ATPase activity in MDCK cells were measured. Carbachol inhibited Na-K-ATPase activity in a time- and concentration-dependent manner. A maximum of approximately 80% of the enzyme activity was inhibited in 160 min with an EC50 of 5 microM carbachol. The inhibition of Na-K-ATPase activity was reversible; up to 80% of the enzyme activity was recovered within 4 h after carbachol was removed. The inhibitory effect of carbachol was blocked by a muscarinic antagonist atropine and by inhibitors of protein kinase C (PKC), 1-(5- isoquinolinesulfonyl)-2-methyl-piperazine HCl, and N-(2- (methylamino)ethyl)-5-isoquinoline sulfonamide HCl. Direct activators of PKC, phorbol 12-myristate 13-acetate, N(n-heptyl)-5-chloro-1- naphthalene sulfonamide, and phosphatidyl serine, also inhibited Na-K- ATPase activity in MDCK cells, and their effect was also blocked by PKC inhibitors. These results indicate that cholinergic agonists inhibit Na- K-ATPase activity in MDCK cells by the activation of PKC. It is concluded that the inhibition of Na-K-ATPase by PKC may, in part, be responsible for the natriuretic action of cholinergic agonists, which have been shown to stimulate phosphoinositide hydrolysis in renal collecting duct cells.

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