Ca-induced K transport in human red blood cell ghosts containing arsenazo III. Transmembrane interactions of Na, K, and Ca and the relationship to the functioning Na-K pump.

Open Access
Publisher Website
Google Scholar
Abstract
Increasing free intracellular Ca (Cai) from < 0.1 to 10 .mu.M by means of A23187 [calcimycin] activated Ca-stimulated K transport and inhibited the Na-K pump in resealed human red cell ghosts. These ghosts contained 2 mM ATP, which was maintained by a regenerating system, and arsenazo III to measure Cai. Ca-stimulated KP transport was activated 50% at 2-3 .mu.M free Cai and the Na-K pump was inhibited 50% by 5-10 .mu.M free Cai. Free Cai from 1-8 .mu.M stimulated K efflux before it inhibited the Na-K pump, dissociating the effect of Ca on the 2 systems; 3 .mu.M trifluoperazine inhibited Ca-stimulated K efflux and 0.5 mM quinidine reduced Na-K pumping by 50%. Incubating fresh intact cells in solutions containing Ca and 0.5 .mu.M A23187 caused the cells to lose K heterogeneously. Under the same conditions, increasing A23187 to 10 .mu.M initiated a homogeneous loss of K. In ATP-deficient ghosts containing Cai equilibrated with A23187, K transport was activated at the same free Cai as in the ghosts containing 2 mM ATP. Neither Cao nor the presence of an inward Ca gradient altered the effect of free Cai on the permeability to K. In these ghosts, transmembrane interactions of Na and K influenced the rate of Ca-stimulated K efflux independent of Na- and K-induced changes in free Cai or sensitivity to Cai. At constant free Cai, increasing Ko from 0.1 to 3 mM stimulated K efflux, whereas further increasing Ko inhibited it. Increasing Nai at constant Ki and free Cai markedly decreased the rate of efflux at 2 mM Ko, but had no effect when Ko was .gtoreq. 20 mM. These transmembrane interactions indicate that the mechanism underlying Ca-stimulated K transport is mediated. Since these interactions from either side of the membrane are independent of free Cai, activation of the transport mechanism by Cai must be at a site that is independent of those responsible for the interaction of Na and K. In the presence of A23187, this activating site is half-maximally stimulated by .apprx. 2 .mu.M free Ca and is not influenced by the concentration of ATP. The partial inhibition of Ca-stimulated K efflux by trifluoperazine in ghosts containing ATP suggests that calmodulin could be involved in the activation of K transport by Cai. Part of the Na-K pump apparatus could be the mechanism that underlies Ca-stimulated K transport because both processes are inhibited by the same drugs, including quinidine, and the pump could account for the transmembrane interactions of Na and K described above. Functioning Na-K pumps are not altered by Cai to transport K selectively, because K transport is activated by Cai before the Na-K pump becomes inhibited.