The molecular basis for adrenergic desensitization in hamster brown adipose tissue: uncoupling of adenylate cyclase activation.

A crude membrane fraction was prepared from hamster brown adipose tissue. Extensive washing of the crude membranes was crucial for the appearance of specific beta-adrenergic receptor binding as assessed by (-)-[3H]dihydroalprenolol. Adrenergic agents competed for the specific binding sites with beta 1-specificity. Binding characteristics were very similar to those earlier found in intact cells, supporting our previous finding that a single (non-tumour) mammalian cell may contain as many as 60,000 beta-adrenergic receptors. Desensitization in situ (i.e. chronic norepinephrine stimulation due to cold acclimation) only marginally affected the number of beta 1-receptors and their affinity (Ki) for norepinephrine. Total (fluoride-stimulated) adenylate cyclase increased somewhat, but the Kact for norepinephrine slightly decreased. Thus the ratio Ki/Kact was rather unaffected by cold acclimation. However, the fraction of adenylate cyclase which could be stimulated by norepinephrine decreased drastically. GTP introduced a low-affinity form (for agonist) of the receptor. The form observed in isolated cells must primarily be the high-affinity form. The basis for desensitization must reside in a diminished ability to transfer the signal from the receptor to the cyclase. This change may be molecularly located in the N-protein or in its interaction with the receptor.