Role of G Proteins in α₁-Adrenergic Inhibition of the β-Adrenergically Activated Chloride Current in Cardiac Myocytes

Abstract

α₁-Adrenergic receptor stimulation can inhibit the Cl⁻ current activated by β-adrenergic receptor agonists in guinea-pig ventricular myocytes. We investigated the role of G proteins in mediating this type of α-adrenergic response. The combined α- and β-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 α₁-adrenergic receptor antagonist prazosin did not cause any further change in sensitivity to NE. These results suggest that the α₁-adrenergic inhibition of β-adrenergic responses is mediated through a PTX-sensitive G protein. However, PTX pretreatment also increased the sensitivity of the Cl⁻ current to the selective β-adrenergic agonist isoproterenol (Iso), which indicates that the PTX treatment increases the sensitivity to β-adrenergic stimulation alone and that this could account for the PTX-induced change in sensitivity to NE. Consistent with this idea, the selective α₁-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 nmIso. These data suggest that the effects of PTX on α₁-adrenergic responses can actually be explained by changes in the sensitivity to β-adrenergic stimulation. To verify the role for a G protein in mediating the inhibitory α₁-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 α₁-adrenergic inhibition of β-adrenergic responses is mediated by a G protein-dependent mechanism that appears to be PTX-insensitive.