Modulation of the calmodulin-induced inhibition of sarcoplasmic reticulum calcium release channel (ryanodine receptor) by sulfhydryl oxidation in single channel current recordings and [H-3]ryanodine binding

The modulation of the calmodulin-induced inhibition of the calcium release channel (ryanodine receptor) by two sulfhydryl oxidizing compounds, 4-(chlo ro-mercuri)phenyl-sulfonic acid (4-CMPS) and 4,4'-dithiodipyridine (4,4'-DT DP) was determined by single channel current recordings with the purified a nd reconstituted calcium release channel from rabbit skeletal muscle sarcop lasmic reticulum (HSR) and [H-3]ryanodine binding to HSR vesicles. 0.1 mu M CaM reduced the open probability (P-o) of the calcium release channel at m aximally activating calcium concentrations (50-100 mu M) from 0.502 +/- 0.0 2 to 0.137 +/- 0.022 (n = 28), with no effect on unitary conductance. 4-CMP S (10-40 mu M) and 4,4'-DTDP (0.1-0.3 mM) induced a concentration dependent increase in P-o (> 0.9) and caused the appearance of longer open states. C aM shifted the activation of the calcium release channel by 4-CMPS or 4,4'- DTDP to higher concentrations in single channel recordings and [H-3]ryanodi ne binding. 40 mu M 4-CMPS induced a near maximal (P-o > 0.9) and 0.3 mM 4, 4'-DTDP a submaximal (P-o = 0.74) channel opening in the presence of CaM, w hich was reversed by the specific sulfhydryl reducing agent DTT. Neither 4- CMPS nor 4,4'-DTDP affected Ca-[I-125]calmodulin binding to HSR. 1 mM MgCl2 reduced P-o from 0.53 to 0.075 and 20-40 mu M 4-CMPS induced a near maxima l channel activation (P-o > 0.9). These results demonstrate that the inhibi tory effect of CaM or magnesium in a physiological concentration is diminis hed or abolished at high concentrations of 4-CMPS or 4,4'-DTDP through oxid ation of activating sulfhydryls on cysteine residues of the calcium release channel.