Gi Proteins Regulate Adenylyl Cyclase Activity Independent of Receptor Activation

Background and purpose

Despite the view that only β₂- as opposed to β₁-adrenoceptors (βARs) couple to G_i, some data indicate that the β₁AR-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 β₂ARs (β₁AR 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 EC₅₀ 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.