Cation exchange and glycoside binding in cultured rat heart cells

Abstract

The Na/K-exchange characteristics, ouabain-binding kinetics, and Na pump turnover rates of synchronously contracting monolayers of neonatal rat myocardial cells were studied. The cells exchange Na rapidly (T1/2 = 35 s) with a mean Na flux of approximately 25 (pmol/cm2)/s. The half time (T1/2) of K exchange is much longer (12 min); the mean K flux is 13 (pmol/cm2)/s. Active Na/K transport, as measured by K influx, is relatively ouabain sensitive, and 10(-6) M ouabain produces half-maximal inhibition. Ouabain (10(-2)M) inhibits 60% of the Na efflux and 75% of the K influx. The cells bind [3H]ouabain rapidly (T1/2 = 8 min), but release it very slowly (T1/2 = 11 h), and both the amount bound and the rate of binding were inversely proportional to extracellular K. Specific [3H]ouabain binding demonstrates saturation reaching a maximum of 1.6 x 10(6) molecules per cell at 2 x 10(-7) M [3H]ouabain. From cell surface area and ouabain-sensitive flux measurements, the Na pump density was calculated at 720/micrometer2 with an individual pump turnover rate of 50/s. Thus the studies indicate that despite their neonatal origin, the behavior of the Na pump in these cells is very similar to that in other mammalian tissues.

Cardiac Basal Metabolism.

“…Thus Gibbs [2], using the Na ϩ -flux data of McCall [146] for the rat heart, calculated that the basal demand of the Na ϩ -pump could be as high as 4 mW g Ϫ1 in that species. An earlier experimental study, in which pharmacological inhibition of the pump was achieved in papillary muscles while the rate of heat production was being measured [93], returned null results.…”

Cardiac glycoside receptors in cultured heart cells—I. characterization of one single class of high affinity receptors in heart muscle cells from chick embryos

“…Ouabain binding sites have also been described in eultured heart muscle cells from neonalal rats 114, 24,39). Idcntifieation of these binding sites as cardiae glyeoside receptors .…”

Cardiac glycoside receptors in cultured heart cells—II

“…11Ie binding to bolh types of binding siles is depressed by K+ and aboIished after heat denaturation of the ee Us. Tbe kinetics of [lH]ouabain binding 10 rat heart muscl e eells (association an d dissociation rate constanlS, K '-and temperaturedependence of association and dissociation processes) have been eha raeterized.…”

“…Despite this evidence of the existence of two discernible classes of cardiac glycoside receptors in membrane preparations and intact tissue of rat heart, only one dass of saturable binding sites for ouabain has been described in cultured heart musc1e cells from neonatal rats: McCall [10] has characlerized a high affinity binding site for ouabain (K D = 2 X 10-7 M) , with ca 1.6 x 1()6 binding sites/celL…”