Inhibition of adenylyl cyclase 1 or exchange protein activated by cAMP restores ATP-sensitive potassium channel activity after chronic opioid exposure

Background and Purpose: Prolonged exposure to Gαi/o-linked receptor agonists such as opioids can lead to a sensitization of adenylyl cyclases (ACs), resulting in heterologous sensitization or cyclic AMP (cAMP) overshoot. The molecular consequences of cAMP overshoot are not well understood, but this adaptive response is suggested to play a critical role in the development of opioid tolerance and withdrawal. Experimental Approach: Genetic reduction of AC1 and simultaneous upregulation of ATP-sensitive potassium channel (K_ATP) subunits, SUR1 or Kir6.2 were performed using viral vectors in mice. In vitro models utilizing an EPAC2-GFP-cAMP biosensor investigated sensitization of AC in SH-SY5Y neuroblastoma cells and HEK-ACΔ3/6 knockout cells. Key Results: Reduction of AC1 and upregulation of K_ATP channels significantly attenuated morphine tolerance and reduced precipitated withdrawal. Acute application of DAMGO decreased the cAMP signal from the EPAC2-GFP-cAMP biosensor, while chronic DAMGO administration resulted in enhanced cAMP production. Inhibition of cAMP overshoot was observed with naloxone (NAL) or pertussis toxin (PTX), as well as co-expression of β-adrenergic receptor kinase C-terminus (βARK-CT). Inhibition of AC1 or exchange protein directly activated by cAMP (EPAC) enhanced potassium channel activity after chronic morphine treatment in a thallium-based assay in SH-SY5Y cells and mouse dorsal root ganglia (DRG) after chronic morphine treatment. Conclusion and Implications: This study presents evidence for investigating further AC1 signalling as a target for opioid tolerance and withdrawal, by increasing EPAC activity and affecting potassium channels downstream of opioid receptors.