title: Voltage-dependent calcium channels in skeletal muscle transverse tubules. Measurements of calcium efflux in membrane vesicles.
abstract: Transverse tubule membranes isolated from rabbit skeletal muscle consist mainly of sealed vesicles that are oriented primarily inside out. These membranes contain a high density of binding sites for 1,4-dihydropyridine calcium channel antagonists. The presence of functional voltage-dependent calcium channels in these membranes has been demonstrated by their ability to mediate 45Ca2+ efflux in response to changes in membrane potential. Fluorescence changes of the voltage-sensitive dye, 3,3'-dipropyl-2,2'-thiadicarbocyanine, have shown that transverse tubule vesicles may generate and maintain membrane potentials in response to establishing potassium gradients across the membrane in the presence of valinomycin. A two-step procedure has been developed to measure voltage-dependent calcium fluxes. Vesicles loaded with 45Ca2+ are first diluted into a buffer designed to generate a membrane potential mimicking the resting state of the cell and to reduce the extravesicular Ca2+ to sub-micromolar levels. 45Ca2+ efflux is then measured upon subsequent depolarization. Flux responses are modulated with appropriate pharmacological specificity by 1,4-dihydropyridines and are inhibited by other calcium channel antagonists such as lanthanum and verapamil.
authors: Department of Physiology and Biophysics, University of Iowa, Iowa City 52242.
authors: S M
authors: SM
authors: Dunn
mesh_terms: D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000818:Animals; D052160:Benzothiazoles; D002121:Calcium Channel Blockers; D015220:Calcium Channels / Q000378:metabolism*; D002132:Calcium Radioisotopes / Q000378:metabolism; D002232:Carbocyanines; D002462:Cell Membrane / Q000378:metabolism*; D004095:Dihydropyridines / Q000378:metabolism / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D005456:Fluorescent Dyes; D017275:Isradipine; D007811:Lanthanum / Q000494:pharmacology; D008564:Membrane Potentials; D009132:Muscles / Q000187:drug effects / Q000378:metabolism*; D009568:Nitrendipine / Q000378:metabolism; D010069:Oxadiazoles / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D011817:Rabbits; D014700:Verapamil / Q000494:pharmacology
chemical_list: D052160:Benzothiazoles; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D002132:Calcium Radioisotopes; D002232:Carbocyanines; D004095:Dihydropyridines; D005456:Fluorescent Dyes; D010069:Oxadiazoles; C012944:3,3'-dipropyl-2,2'-thiadicarbocyanine; D007811:Lanthanum; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D009568:Nitrendipine; D004492:Edetic Acid; D014700:Verapamil; D011188:Potassium; D017275:Isradipine