1. Guanine nucleotide-binding proteins (G-proteins) play a central role in signal transduction between a wide variety of cell-surface receptors and intracellular second messenger systems. Recently, we and others have demonstrated that cross-regulation can occur between a variety of G-protein-linked receptors in human heart. Chronic β 1 -adrenoceptor blockade gives rise to sensitization of β 2 -adrenoceptor and of 5HT 4 -receptor responses, both of which are mediated via stimulation of adenylate cyclase through stimulatory G-proteins (G s ), and also gives rise to desensit-ization of muscarinic M 2 -receptor responses, which inhibit adenylate cyclase through inhibitory G-proteins (G i ). 2. In order to investigate whether these effects are due to quantitative changes in cardiac G-protein isoforms, we measured their abundance in right atrial appendage from patients taking or not taking β 1 -adrenoceptor antagonists, by immunoblotting. 3. Samples of right atrial appendage homogenate were subjected to SDS/PAGE, and proteins were electroblotted on to nitrocellulose membranes. These were then probed with specific anti-G protein anti-sera, and binding was revealed by means of a secondary antibody labelled with alkaline phosphatase and using a chromogenic substrate. The resulting bands were quantified by laser densitometry. 4. No quantitative differences were detected, between these two groups of patients, in the amounts of α-subunit of ‘long’ or ‘short’ G s isoforms (G s αL and G s αS), or in the amounts of G i 1 + 2 α-subunit (G i α1 + 2). Nor was any difference found in the abundance of the β-subunit of G-proteins. No ‘other’ G-protein (G o ) was detectable in these samples by immunoblotting. 5. We conclude that the phenomenon of receptor cross-regulation which we have previously observed in human right atrial appendage is unlikely to be explained by quantitative changes at the G-protein level.
1. The mechanisms underlying control of cyclic AMP responses to isoprenaline were studied in primary cultures of human airway smooth muscle cells. In these cells, isoprenaline induced concentration-related cyclic AMP formation via β 2 -adrenoceptor stimulation. 2. Prior incubation of cells with varying concentrations of isoprenaline (1–16 h), forskolin, prostaglandin E 2 or a stable analogue of cyclic AMP all produced concentration-related desensitization of cyclic AMP responses to subsequent challenge with isoprenaline (maximum reduction with 1 μmol/l isoprenaline, 85% after 16 h). The desensitization induced over 2 h (44%) by a concentration of prostaglandin E 2 which gave a similar rise in cyclic AMP levels to 1 μmol/l isoprenaline was significantly less ( P < 0.05) than the desensitization (62%) induced over 2 h by 1 μmol/l isoprenaline itself. 3. Isoprenaline-induced desensitization of β 2 -adrenoceptor-induced cyclic AMP formation was insensitive to prior exposure of cells to dexamethasone. 4. These findings suggest that isoprenaline-induced desensitization of β 2 -adrenoceptor-induced cyclic AMP formation in primary cultures of human airway smooth muscle cells is mediated through both a cyclic AMP-dependent and probably an additional cyclic AMP-independent pathway, and that these pathways are insensitive to inhibition by glucocorticoids.