N-ethylmaleimide, elevated temperature, and digitonin solubilization eliminate guanine nucleotide but not sodium effects on human platelet alpha 2-adrenergic receptor-agonist interactions.
Three independent perturbants of the human platelet membrane were found to differentially interfere with modulation of platelet alpha 2-adrenergic receptor-agonist interactions by guanine nucleotides and Na+. Treatment of intact platelets with 1mM N-ethylmaleimide, exposure of particulate preparations to 60 degrees or digitonin solubilization of platelet membranes all eliminated guanine nucleotide regulation of alpha 2-receptor-agonist interactions without interfering with Na+-promoted decreases in receptor affinity for epinephrine. The observations with N-ethylmaleimide and elevated temperature are consistent either with separate membrane components conferring sensitivity to Na+ and guanine nucleotides or with different domains of a single protein component mediating effects of Na+ and GTP on alpha 2-receptor affinity for agonists. However, the observation that Na+ modulation of alpha 2-receptor-agonist interactions is retained in digitonin-solubilized preparations is persuasive evidence that distinct membrane components confer Na+ and GTP sensitivity to alpha 2-adrenergic receptor-agonist interactions, since previous studies (Smith, S.K. and Limbird, L.E. Proc. Natl. Acad. Sci. USA 78:4026-4030, 1981) have demonstrated that the GTP-binding protein modulating alpha 2-receptor affinity for agonists is dissociated from the alpha 2-receptor upon digitonin solubilization unless the receptor is occupied by an agonist at the time of exposure to detergent. Thus, although GTP and Na+ have similar phenomenological effects to permit or enhance alpha-adrenergic attenuation of adenylate cyclase activity and to decrease receptor affinity for agonists, these agents nonetheless appear to mediate their regulatory effects via different membrane components or at least distinct subunits of perhaps shared multisubunit proteins.
D000316:Adrenergic alpha-Agonists; D001792:Blood Platelets / Q000187:drug effects*; D002462:Cell Membrane / Q000187:drug effects; D004072:Digitonin / Q000494:pharmacology*; D005033:Ethylmaleimide / Q000494:pharmacology*; D006160:Guanosine Triphosphate / Q000494:pharmacology*; D006358:Hot Temperature; D006801:Humans; D066298:In Vitro Techniques; D011942:Receptors, Adrenergic, alpha / Q000502:physiology*; D012964:Sodium / Q000494:pharmacology
D000316:Adrenergic alpha-Agonists; D011942:Receptors, Adrenergic, alpha; D006160:Guanosine Triphosphate; D012964:Sodium; D004072:Digitonin; D005033:Ethylmaleimide