Modulation of the calmodulin-induced inhibition of sarcoplasmic reticulum calcium release channel (ryanodine receptor) by sulfhydryl oxidation in single channel current recordings and [(3)H]ryanodine binding.
The modulation of the calmodulin-induced inhibition of the calcium release channel (ryanodine receptor) by two sulfhydryl oxidizing compounds, 4-(chloromercuri)phenyl-sulfonic acid (4-CMPS) and 4, 4'-dithiodipyridine (4,4'-DTDP) was determined by single channel current recordings with the purified and reconstituted calcium release channel from rabbit skeletal muscle sarcoplasmic reticulum (HSR) and [(3)H]ryanodine binding to HSR vesicles. 0.1 microm CaM reduced the open probability (P(o)) of the calcium release channel at maximally activating calcium concentrations (50-100 microm) from 0.502 +/- 0.02 to 0.137 +/- 0.022 (n = 28), with no effect on unitary conductance. 4-CMPS (10-40 microm) 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. CaM shifted the activation of the calcium release channel by 4-CMPS or 4,4'-DTDP to higher concentrations in single channel recordings and [(3)H]ryanodine binding. 40 microm 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, which was reversed by the specific sulfhydryl reducing agent DTT. Neither 4-CMPS nor 4,4'-DTDP affected Ca-[(125)I]calmodulin binding to HSR. 1 mm MgCl(2) reduced P(o) from 0.53 to 0.075 and 20-40 microm 4-CMPS induced a near maximal channel activation (P(o) > 0.9). These results demonstrate that the inhibitory effect of CaM or magnesium in a physiological concentration is diminished or abolished at high concentrations of 4-CMPS or 4,4'-DTDP through oxidation of activating sulfhydryls on cysteine residues of the calcium release channel.
Suko
J
J
Institute of Pharmacology, University of Vienna, Währingerstr. 13a, A-1090 Vienna, Austria.
Hellmann
G
G
Drobny
H
H
D000818:Animals; D002147:Calmodulin / Q000378:metabolism* / Q000494:pharmacology; D004220:Disulfides / Q000378:metabolism* / Q000494:pharmacology; D007457:Iodine Radioisotopes; D009941:Organomercury Compounds / Q000378:metabolism* / Q000494:pharmacology; D010084:Oxidation-Reduction; D011725:Pyridines / Q000378:metabolism* / Q000494:pharmacology; D011817:Rabbits; D012433:Ryanodine / Q000378:metabolism*; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D013439:Sulfhydryl Reagents / Q000378:metabolism* / Q000494:pharmacology; D014316:Tritium
C400327:4-CMPS; D002147:Calmodulin; D004220:Disulfides; D007457:Iodine Radioisotopes; D009941:Organomercury Compounds; D011725:Pyridines; D019837:Ryanodine Receptor Calcium Release Channel; D013439:Sulfhydryl Reagents; D014316:Tritium; D012433:Ryanodine; C007542:4,4'-dipyridyl disulfide