PDE8 Mediates L-type Ca²⁺ Channels Dephosphorylation in Human Atrial Fibrillation

Objectives: Atrial fibrillation (AF) is associated with reduced L-type Ca²⁺ current (I_Ca,L) and altered cAMP-dependent signaling. Phosphodiesterases (PDEs) degrade cAMP and regulate cAMP-mediated PKA-dependent phosphorylation of various proteins, including I_Ca,L channel subunits. PDE1-4 is the main PDE isozymes hydrolyzing cAMP in heart, but recent studies demonstrate the existence of a novel isoform PDE8 in ventricle. Here we assess the expression, localization and function of PDE8 in human atria of patients with sinus rhythm (SR), paroxysmal AF (pAF) and longstanding persistent (chronic) AF (cAF).

Methods: mRNA (RT-qPCR) and protein (western blot) levels of PDE8A and PDE8B isoforms were assessed in right atria of SR, pAF, and cAF patients. Localization of PDE8A and PDE8B in human atrial cardiomyocytes was determined by immunofluorescence. Protein–protein interaction between I_Ca,L channel and PDE8B was studied by coimmunoprecipitation in the three rhythm groups. I_Ca,L and cAMP were measured in isolated human myocytes using whole-cell perforated patch-clamp technique and FRET. Action potential was assessed in right atrial trabeculae with sharp electrode.

Results: PDE8A mRNA and protein expression levels were twofold higher in pAF, but unaltered in cAF patients. Conversely, PDE8B protein abundance was increased by ~77% in cAF only. Immunostaining confirmed the presence of PDE8A and PDE8B in human atrial cardiomyocytes, with PDE8A being localized in the cytosol, and PDE8B preferentially localized at the plasma membrane. Immunoprecipitation of I_Ca,L channel resulted in strongly enhanced coimmunoprecipitation of PDE8B in cAF but not pAF, identifying PDE8B as part of the I_Ca,L channel complex and pointing to potential contribution of PDE8B to reduced I_Ca,L in cAF. Pharmacological inhibition of PDE8 increased I_Ca,L in cAF up to SR densities and the plateau potential to more positive values with a slight prolongation of the APD at 50% of repolarization by increasing cAMP levels.

Conclusion: Our results show for the first time that PDE8A and B are expressed in human atrium. PDE8B localizes at the plasma membrane of human atrial cardiomyocytes, and upregulates and accumulates in the I_Ca,L channel of cAF patients. PDE8 inhibitor increase cAMP levels restoring I_Ca,L in cAF. PDE8B may constitute a novel regulator of atrial I_Ca,L, with potential implications for AF pathophysiology. PDE8B inhibition could provide a novel therapeutic approach in chronic AF.