Measurement of sarcolemmal vesicle orientation by beta-adrenergic receptor binding.
To assess the orientation (inside-out vs. outside-out) of purified cardiac sarcolemmal vesicles, we developed a new method utilizing the known outward-facing binding site of the beta-adrenergic receptor. We compared the binding of the lipid-insoluble ligand 3H-CGP-12177, which binds to beta-adrenergic receptors on outside-out sarcolemmal vesicles only, to the binding of the lipid soluble ligand 125I-iodocyanopindolol, which binds to beta-adrenergic receptors in sarcolemmal vesicles of either orientation. The ratio of CGP to ICYP binding is equal to the fraction of outside-out sarcolemmal vesicles. Sidedness measurements by beta-adrenergic receptor-binding showed similar mean values but less scatter than sidedness assessments by measurement of 3H-ouabain-binding or Na+,K(+)-ATPase activity in the presence or absence of membrane permeabilizing agents.
Bersohn
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MM
Cardiology Section, Veterans Affairs Medical Center, Los Angeles, CA 90073.
Doshi
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RN
D000818:Animals; D001665:Binding Sites; D006321:Heart / Q000187:drug effects; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes; D020120:Iodocyanopindolol; D008722:Methods; D010042:Ouabain / Q000493:pharmacokinetics; D010869:Pindolol / Q000031:analogs & derivatives / Q000494:pharmacology; D011412:Propanolamines / Q000493:pharmacokinetics; D011817:Rabbits; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*; D012508:Sarcolemma / Q000737:chemistry / Q000502:physiology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis; D014316:Tritium
D007457:Iodine Radioisotopes; D011412:Propanolamines; D011943:Receptors, Adrenergic, beta; D014316:Tritium; D010042:Ouabain; D020120:Iodocyanopindolol; D010869:Pindolol; D000254:Sodium-Potassium-Exchanging ATPase; C033394:CGP 12177