Effects of chelators and calcium ions on the response of renal adenosine triphosphatase activity to hormones and concanavalin A.

Experiments with plasma membrane from rat liver have been interpreted as showing that the activity of the (Na++K+)-dependent ATPase* is subject to modulation by the action of hormones, such as L-adrenaline (epinephrine) and insulin, through the latter's effects on adenylate cyclase activity and consequent changes in cyclic AMP concentration (Luly et af. , 1972; Tria et af . , 1974; Luly & Emmelot, 1975). Because of the obvious physiological importance of such hormonal control of the ATPase activity, if the interpretation is correct, we have examined this system further. Rat kidney tissue was used instead of liver because the former is a better source of (Na++K+)-dependent ATPase. When the same method for preparation of a membrane fraction (Ray, 1970) was applied to kidney and the (Na++K+)-dependent ATPase activity was assayed under the conditions used by Bonting (1970), results closely paralleling those of Luly & Emmelot (1975) were obtained (Tables 1 and 2). However, no such effects were observed when the (Na++K+)-dependent ATPase activity was assayed under the conditions described by Wheeler et al. (1975). Comparison of the differences between the two assay conditions revealed that the presence of 0.5mM-EGTA in place of 0.lmM-EDTA abolished the effects of the hormones and concanavalin A. The EGTA appeared to mimic the effect of concanavalin A and adrenaline. For example, an (Na+ + K+)-dependent ATPase activity (in pmol of PJmin per mg of protein) of 0.13 was decreased to 0.04 by concanavalin A, and to 0.06 by EGTA. Similarly, when Ca2+ was omitted from the solution used for the initial homogenization of the tissue, the final preparation had much lower (Na++K+)dependent ATPase activity than usual (0.07unit). Moreover, that low activity was no longer sensitive to adrenaline or concanavalin A.