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MOLECULAR PHARMACOLOGY 51:853-860 (1997).

Role of G Proteins in ₁-Adrenergic Inhibition of the -Adrenergically Activated Chloride Current in Cardiac Myocytes

Livia C. Hool, Lisa M. Oleksa, and Robert D. Harvey

Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio, 44106.

₁-Adrenergic receptor stimulation can inhibit the Cl current activated by -adrenergic receptor agonists in guinea-pig ventricular myocytes. We investigated the role of G proteins in mediating this type of -adrenergic response. The combined - and -adrenergic agonist norepinephrine (NE) activated the Cl current with an EC₅₀ value of 53 nM. Preincubation of myocytes with PTX decreased the EC₅₀ value for NE activation of the Cl current to 5.9 nM, and addition of the ₁-adrenergic receptor antagonist prazosin did not cause any further change in sensitivity to NE. These results suggest that the ₁-adrenergic inhibition of -adrenergic responses is mediated through a PTX-sensitive G protein. However, PTX pretreatment also increased the sensitivity of the Cl current to the selective -adrenergic agonist isoproterenol (Iso), which indicates that the PTX treatment increases the sensitivity to -adrenergic stimulation alone and that this could account for the PTX-induced change in sensitivity to NE. Consistent with this idea, the selective ₁-adrenergic receptor agonist methoxamine was still able to inhibit the Cl current activated by Iso in PTX-treated myocytes. However, the sensitivity to methoxamine was significantly decreased. In control cells, the Cl current activated by 30 nM Iso was inhibited by methoxamine with an EC₅₀ value of 8.3 µM, but in PTX-treated cells, the EC₅₀ value was 284 µM. The EC₅₀ for methoxamine inhibition was similarly increased when the Cl current was activated by 300 nM Iso. These data suggest that the effects of PTX on ₁-adrenergic responses can actually be explained by changes in the sensitivity to -adrenergic stimulation. To verify the role for a G protein in mediating the inhibitory ₁-adrenergic response, we examined the effect of methoxamine on the Cl current activated in cells dialyzed with the nonhydrolyzable GTP analogue guanosine-5-O-(3-thio)triphosphate. Pre-exposure to methoxamine resulted in an attenuated response upon subsequent exposure to Iso alone. We conclude that ₁-adrenergic inhibition of -adrenergic responses is mediated by a G protein-dependent mechanism that appears to be PTX-insensitive.