Direct binding of verapamil to the ryanodine receptor channel of sarcoplasmic reticulum.
Radioligand binding experiments and single channel recordings demonstrate that verapamil interacts with the ryanodine receptor Ca2+ release channel of the sarcoplasmic reticulum of rabbit skeletal muscle. In isolated triads, verapamil decreased binding of [3H]Ryanodine with an IC50 of approximately 8 microM at an optimal pH 8.5 and pCa 4.3. Nitrendipine and d-cis-diltiazem did not interfere with binding of [3H]Ryanodine to triads, suggesting that the action of verapamil does not involve the dihydropyridine receptor. Single channel recordings showed that verapamil blocked Ca2+ release channels by decreasing open probability, duration of open events, and number of events per unit time. A direct interaction of verapamil with the ryanodine receptor peptide was demonstrated after purification of the approximately 400 kDa receptor protein from Chaps-solubilized triads. The purified receptor displayed high affinity for [3H]Ryanodine with a Kd of approximately 5 nM and a Bmax of approximately 400 pmol/mg. Verapamil and D600 decreased [3H]Ryanodine binding noncompetitively by reducing the Bmax. Thus the presence of binding sites for phenylalkylamines in the Ca2+ release channel was confirmed. Verapamil blockade of Ca2+ release channels may explain some of the paralyzing effects of phenylalkylamines observed during excitation-contraction coupling of skeletal muscle.
Valdivia
H H
HH
Department of Physiology, University of Wisconsin School of Medicine, Madison 53706.
Valdivia
C
C
Ma
J
J
Coronado
R
R
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008051:Lipid Bilayers; D009132:Muscles / Q000378:metabolism / Q000502:physiology; D011817:Rabbits; D011950:Receptors, Cholinergic / Q000187:drug effects / Q000378:metabolism*; D012433:Ryanodine / Q000378:metabolism*; D019837:Ryanodine Receptor Calcium Release Channel; D012519:Sarcoplasmic Reticulum / Q000502:physiology*; D014700:Verapamil / Q000378:metabolism* / Q000494:pharmacology
D015220:Calcium Channels; D008051:Lipid Bilayers; D011950:Receptors, Cholinergic; D019837:Ryanodine Receptor Calcium Release Channel; D012433:Ryanodine; D014700:Verapamil; D002118:Calcium