Mechanism of Ca release from sarcoplasmic reticulum: Effect of adenine nucleotide on magnesium binding to fragmented sarcoplasmic reticulum from bullfrog skeletal muscle

We have reported that caffeine reduces both the apparent affinity for Mg (K) and the number of Mg binding sites (n) of fragmented sarcoplasmic reticulum (FSR) from bullfrog skeletal muscle which are accompanied by Ca release (Jpn. J. Pharmacol. 32, 81P, 1982). The effect may be the basis of Ca release by caffeine. To get further insight into this problem, we here examine the effect of β,γ-methylene adenosine triphosphate (AMPOPCP) on FSR. First of all, the apparent binding constants of AMPOPCP for Mg and Ca should be determined to obtain Mg ion concentration after correction for Mg-AMPOPCP complex in the presence of 40μM free Ca ion. Under the experimental conditions, they were determined to be 3.28±0.07(3) for Mg and 2.97±0.02(3) for Ca in logarithm at pH 6.50 at 20°C using ion selective electrode. In the presence of about 40μM Ca ion concentration, 0.1mM AMPOPCP reduced K and n to 0.58±0.10(3) and 0.80±0.10(3), respectively, giving rise to Ca release, whereas lOmM caffeine reduced them to 0.52±0.10(8) and 0.46±0.04(8), respectively. In contrast to caffeine, AMPOPCP reduced Z to a larger extent than n. These effects of AMPOPCP were prevented by procaine. AMPOPCP in combination with caffeine reduced n markedly, while it increased K on the contrary. Correspondingly, the accompanying Ca release was not affected by high concentrations of Mg up to 4mM.