Fat cell adenylate cyclase system: Enhanced inhibition by adenosine and GTP in the hypothyroid rat

Hypothyroidism is associated with an enhanced sensitivity of rat fat cells to the inhibitory action of adenosine and adenosine agonists. The sensitivity of the forskolin-stimulated cyclic AMP response of rat fat cells to the adenosine agonist N⁶-phenylisopropyladenosine is amplified 3-fold by hypothyroidism. Forskolin-stimulated adenylate cyclase activity is more sensitive to inhibition by this adenosine agonist in membranes of fat cells isolated from hypothyroid as compared to euthyroid rats. Hypothyroidism does not significantly alter the number or affinity of binding sites for N⁶-cyclohexyl[³H]adenosine or N⁶-phenylisopropyladenosine in membranes of rat fat cells. GTP-induced inhibition of forskolin-stimulated adenylate cyclase was markedly enhanced in the hypothyroid state, suggesting an alteration in the inhibitory regulatory component (N(i)-mediated control of adenylate cyclase. Incubating membranes with [α-³²P]NAD⁺ and preactivated pertussis toxin results in the radiolabeling of two peptides with M(r) = 40,000 and 41,000 as visualized in autoradiograms of polyacrylamide gels run in sodium dodecyl sulfate. The amount of label incorporated by pertussis toxin into these two peptides (putative subunits of N(i)) per mg of protein of membrane is increased 2-3-fold in the hypothyroid state. The amount of the stimulatory regulatory component, N(s), in fat cell membranes is not altered by hypothyroidism. The amplified response of hypothyroid rat fat cells to the inhibitory action of adenosine appears to reflect a specific increase in the activity and abundance of N(i).