Identification of two populations of cardiac microsomes with nitrendipine receptors: correlation of the distribution of dihydropyridine receptors with organelle specific markers.
Conventional sarcolemma and microsome preparations from rabbit and cat ventricular muscle were fractionated on continuous linear sucrose gradients. The distribution of nitrendipine receptors was compared with the distribution of organelle specific markers. For the conventional sarcolemma preparation, the dihydropyridine receptor distribution matched the pattern for external membrane markers in position and shape. The number of nitrendipine receptors was three times the number of muscarine binding sites (approximately 1.0 pmol/mg protein) at the isopycnic point of the vesicles. In contrast, two populations of vesicles with nitrendipine receptors were found in the microsome preparations. One population banded with the external membrane vesicles at a mean buoyant density of 24% (w/w) sucrose. The specific content of dihydropyridine receptors (0.2 pmol/mg) was 1/5 that for the muscarine receptors. The second and major population followed the distribution of an Mr 300K polypeptide, a marker for the junctional cisternae of the sarcoplasmic reticulum (SR). Muscarine receptors, however, were also present throughout that band, albeit at a reduced specific content (approximately 0.1 pmol/mg) compared to the light vesicles. The nitrendipine specific content increased over threefold from that of the light vesicles such that the relative content (nitrendipine/muscarine) was twice that determined for the conventional sarcolemma preparation. Nitrendipine receptors were not associated with nonjunctional SR or mitochondria. The light and heavy microsome populations were incubated with 0.2 mg digitonin/mg protein, a treatment which preferentially perturbs the isopycnic point of external membrane vesicles. For the light vesicles, the membranes with muscarine and nitrendipine receptors became heavier than the bulk of the SR. In contrast, after digitonin treatment of the heavy vesicle population, the nitrendipine and muscarine receptors and the SR marker appeared to comigrate into a sharpened band at 39% sucrose. The possibility that the dihydropyridine binding sites in the heavy microsome population are on external membrane vesicles physically linked to the junctional SR is discussed.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels; D002415:Cats; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D004072:Digitonin / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D008274:Magnesium / Q000378:metabolism; D008861:Microsomes / Q000032:analysis* / Q000187:drug effects; D008970:Molecular Weight; D009206:Myocardium / Q000648:ultrastructure*; D009940:Organoids / Q000032:analysis; D011188:Potassium / Q000378:metabolism; D011817:Rabbits; D011976:Receptors, Muscarinic / Q000032:analysis; D011978:Receptors, Nicotinic / Q000032:analysis*; D014018:Tissue Distribution
D015220:Calcium Channels; D011976:Receptors, Muscarinic; D011978:Receptors, Nicotinic; D008274:Magnesium; D004072:Digitonin; D011188:Potassium; D002118:Calcium