Identification, characterization, and photoaffinity labeling of the dihydropyridine receptor associated with the L-type calcium channel from bovine adrenal medulla.
The dihydropyridine receptor associated with the L-type Ca2+ channel in adrenal medulla membranes has been identified and characterized. [3H]PN200-110 binds in a stereoselective, saturable manner to a single class of high affinity sites in adrenal medulla membranes, with a Kd of 0.1 nM and a Bmax of 141 fmol/mg of protein. Dihydropyridines inhibited [3H]PN200-110 binding with the rank order (+)-PN200-110 greater than nifedipine greater than nimodipine greater than usoldipine greater than or equal to nitrendipine greater than BayK8644 greater than (-)-PN200-110. [3H] PN200-110 binding was sensitive to divalent cations, as examined by the effects of Ca2+, Mg2+, and the chelators ethylene glycol bis-(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid and EDTA. [3H]PN200-110 binding was modulated by various classes of L-type Ca2+ channel effectors. Benzothiazepines modulated binding of [3H]PN200-110 in a negative or positive manner that was temperature dependent, whereas phenylalkylamines weakly inhibited [3H]PN200-110 binding. Bepridil stimulated [3H] PN200-110 binding, whereas phencyclidine was without effect. The photoaffinity probe [3H]azidopine labeled a single polypeptide that migrated with an apparent molecular weight of 185,000-190,000 in sodium dodecyl sulfate gel electrophoresis. The dihydropyridine receptor was found to bind specifically to wheat germ agglutinin columns. These results demonstrate the presence of a Ca2+ channel blocker complex in adrenal medulla. The drug receptor sites reside on a glycoprotein complex in which a polypeptide analogous to the alpha 1-subunit of the L-type Ca2+ channel from skeletal muscle has been identified.
Murphy
B J
BJ
Department of Pharmacology, University of Ottawa, Ontario, Canada.
Rogers
C A
CA
Sunahara
R K
RK
Lemaire
S
S
Tuana
B S
BS
D000313:Adrenal Medulla / Q000378:metabolism*; D000345:Affinity Labels; D000494:Allosteric Regulation; D000818:Animals; D001386:Azides; D001667:Binding, Competitive; D002121:Calcium Channel Blockers / Q000378:metabolism; D015220:Calcium Channels / Q000378:metabolism*; D002413:Cations, Divalent / Q000494:pharmacology; D002417:Cattle; D004095:Dihydropyridines; D006358:Hot Temperature; D017275:Isradipine; D010069:Oxadiazoles / Q000378:metabolism; D010777:Photochemistry; D011869:Radioligand Assay; D011978:Receptors, Nicotinic / Q000032:analysis* / Q000187:drug effects; D013237:Stereoisomerism; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D000345:Affinity Labels; D001386:Azides; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D002413:Cations, Divalent; D004095:Dihydropyridines; D010069:Oxadiazoles; D011978:Receptors, Nicotinic; D013439:Sulfhydryl Reagents; C041130:azidopine; D017275:Isradipine