(-)-[H-3] DESMETHOXYVERAPAMIL LABELS MULTIPLE CALCIUM-CHANNEL MODULATOR RECEPTORS IN BRAIN AND SKELETAL-MUSCLE MEMBRANES - DIFFERENTIATION BY TEMPERATURE AND DIHYDROPYRIDINES

  • 1 June 1986
    • journal article
    • research article
    • Vol. 237  (3) , 731-738
Abstract
The verapamil-like calcium channel modulator, (.sbd.)-[3H]desmethoxyverapamil binds to multiple sites in microsomal membrane preparations from brain and skeletal muscle. In brain the Kd values of the sites are 0.55 .+-. 0.1 and 61.8 .+-. 18 nM for the high and low affinity sites and the maximum binding values are 0.22 .+-. 0.04 and 4.6 .+-. 1.0 pmol/mg of protein, respectively. Equilibrium analysis of saturation data in skeletal muscle membranes shows only one site with an affinity of 7.2 .+-. 0.8 nM and a maximum binding of 2.96 .+-. 0.32 pmol/mg of protein. However, a low affinity site with an estimated Kd of 152 nM is indicated in dissociation kinetic studies. Dihydropyridine calcium channel modulators regulate the binding of desmethoxyverapamil in a temperature-dependent fashion with (+)-PN 200110 decreasing (.sbd.)-[3H]desmethoxyverapamil binding more at 0.degree. C than at higher temperatures and, at 37.degree. C, enhancing binding in skeletal muscle. The influence of (+)-desmethoxyverapamil on (+)-[3H]PN 200110 binding is unchanged by temperature variations, whereas interactions of the (.sbd.)-enantiomer are altered markedly with more inhibiton at 0.degree. C than at higher temperatures and, in skeletal muscle, stimulation of binding at 37.degree. C. Dissociation studies indicate that the two sites labeled by (.sbd.)-[3H]-desmethoxyverapamil in skeletal muscle interact in a negative heterotropic cooperative fashion. Dihydropyridines appear to slow the dissociation of ligand from the low affinity site, whereas diltiazem accelerates the dissociation of (.sbd.)-[3H]-desmethoxyverapamil from the high affinity site. These results suggest that the high and low affinity sites labeled by (.sbd.)-[3H]desmethoxyverapamil, respectively, represent the verapamil and diltiazem receptors in brain and skeletal muscle.

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