AJP - Regu AJP: Regulatory, Integrative and Comparative Physiology
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Am J Physiol Regul Integr Comp Physiol 259: R453-R460, 1990;
0363-6119/90 $5.00
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AJP - Regulatory, Integrative and Comparative Physiology, Vol 259, Issue 3 453-R460, Copyright © 1990 by American Physiological Society

ARTICLES

Ca2+ transport in myocardial sarcolemma from rainbow trout

G. F. Tibbits, H. Kashihara, M. J. Thomas, J. E. Keen and A. P. Farrell
Cardiac Membrane Research Laboratory, School of Kinesiology, Simon Fraser University, Burnaby, British Columbia, Canada.

Sacrolemmal vesicles were isolated from trout ventricles with a yield of 0.51 mg protein/g wet wt of a fraction enriched approximately 15-fold over the crude homogenate as estimated by K(+)-stimulated p-nitrophenylphosphatase (K(+)-pNPPase) activity. Although the K(+)-pNPPase specific activity compared favorably with that of the rat heart, there were some striking differences in the sodium dodecyl sulfate-polyacrylamide gel electrophoresis and specific phospholipid content (mumol/mg protein) of the sacrolemmal fractions between the two species. Two major sarcolemmal Ca2(+)-transport proteins were investigated, the Na(+)-Ca2+ exchanger and the dihydropyridine (DHP) receptor, a component of the voltage-dependent L-type Ca2+ channel. From the initial rates of Na(+)-dependent Ca2+ uptake, it was determined that the exchanger has an apparent Km for Ca2+ of 14 +/- 1 microM and a maximum velocity of 7.7 +/- 1.1 nmol.mg protein-1.s-1 at 21 degrees C. Experiments using the DHP ligand [3H] (+) PN 200-110 to characterize the equilibrium binding to the DHP receptor in the sarcolemmal fraction yielded a Kd of 0.08 nM and maximum binding sites of 3.06 +/- 0.49 pmol/mg protein. Given the smaller dimensions of the trout myocyte and the resultant higher sarcolemmal surface to cytosolic volume compared with the mammalian myocyte, these in vitro findings are consistent with the notion that Ca2+ transport across sarcolemma is a quantitatively important contributor of Ca2+ delivery to and removal from the contractile element.


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