G(i) Proteins Regulate Adenylyl Cyclase Activity Independent of Receptor Activation

BACKGROUND AND PURPOSE: Despite the view that only β(2)- as opposed to β(1)-adrenoceptors (βARs) couple to G(i), some data indicate that the β(1)AR-evoked inotropic response is also influenced by the inhibition of G(i). Therefore, we wanted to determine if G(i) exerts tonic receptor-independent inhibition upon basal adenylyl cyclase (AC) activity in cardiomyocytes. EXPERIMENTAL APPROACH: We used the G(s)-selective (R,R)- and the G(s)- and G(i)-activating (R,S)-fenoterol to selectively activate β(2)ARs (β(1)AR blockade present) in combination with G(i) inactivation with pertussis toxin (PTX). We also determined the effect of PTX upon basal and forskolin-mediated responses. Contractility was measured ex vivo in left ventricular strips and cAMP accumulation was measured in isolated ventricular cardiomyocytes from adult Wistar rats. KEY RESULTS: PTX amplified both the (R,R)- and (R,S)-fenoterol-evoked maximal inotropic response and concentration-dependent increases in cAMP accumulation. The EC(50) values of fenoterol matched published binding affinities. The PTX enhancement of the G(s)-selective (R,R)-fenoterol-mediated responses suggests that G(i) regulates AC activity independent of receptor coupling to G(i) protein. Consistent with this hypothesis, forskolin-evoked cAMP accumulation was increased and inotropic responses to forskolin were potentiated by PTX treatment. In non-PTX-treated tissue, phosphodiesterase (PDE) 3 and 4 inhibition or removal of either constitutive muscarinic receptor activation of G(i) with atropine or removal of constitutive adenosine receptor activation with CGS 15943 had no effect upon contractility. However, in PTX-treated tissue, PDE3 and 4 inhibition alone increased basal levels of cAMP and accordingly evoked a large inotropic response. CONCLUSIONS AND IMPLICATIONS: Together, these data indicate that G(i) exerts intrinsic receptor-independent inhibitory activity upon AC. We propose that PTX treatment shifts the balance of intrinsic G(i) and G(s) activity upon AC towards G(s), enhancing the effect of all cAMP-mediated inotropic agents.