Measurement of calcium entry and exit in quiescent rat ventricular myocytes.
The aim of this work was to obtain the first quantitative measurements of Ca2+ influx and efflux in quiescent cardiac cells. The relationship between free and total Ca2+ was obtained during a caffeine application. This buffering curve was then used to calculate changes of total Ca2+ from measurements of free cytosolic [Ca2+] ([Ca2+]i) made with Indo-1. The rate of Ca2+ removal from the cytoplasm was calculated by differentiating total Ca2+ with respect to time. The dependence of d(total Ca2+)/dt on [Ca2+]i was hyperbolic. Inhibition of either Na+-Ca2+ exchange (by addition of 10 mmol l(-1) NiCl2 or removal of external Na+) or the sarcolemmal Ca2+-activated adenosine triphosphatase (Ca2+-ATPase) (with carboxyeosin) decreased the calculated efflux. In both cases, the main effect was on the apparent maximum rate (Vmax) with little effect on the Michaelis-Menten constant (Km). These results suggest that the Na+-Ca2+ exchange and Ca2+-ATPase have very similar affinities for [Ca2+]i and that their fractional contributions do not change over the systolic range of [Ca2+]i. Ca2+ influx was quantified in two ways. The first method was to extrapolate the curve relating Ca2+ efflux to [Ca2+]i to zero [Ca2+]i. This gave a value of 4.49+/-0.54 micromol l(-1) s(-1) which was reduced to zero by either removal of external Ca2+ or addition of Ni2+. In other experiments external Ca2+ was removed and the maximum rate of fall of total Ca2+ calculated as 2.53+/-0.93 micromol l(-1) s(-1). This approach can be used to provide a quantitative analysis of the control of resting [Ca2+]i.
Choi
H S
HS
Department of Medicine, The University of Manchester, UK.
Trafford
A W
AW
Eisner
D A
DA
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes; D006352:Heart Ventricles / Q000378:metabolism; D007211:Indoles; D009206:Myocardium / Q000378:metabolism*; D009532:Nickel / Q000494:pharmacology; D051381:Rats; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors
D020746:Calcium Channels, L-Type; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D007211:Indoles; D019831:Sodium-Calcium Exchanger; D002110:Caffeine; C022838:nickel chloride; D009532:Nickel; D000252:Calcium-Transporting ATPases; C048960:indo-1; D002118:Calcium