Atrial fibrillation (AF) occurs from disordered atrial action potential conduction and is associated with reduced gap junction electrical conductance (Gj). The Ca2+ and calmodulin-dependent phosphatase, calcineurin, reduces Gj in ventricular myocardium via a protein phosphatase-1 (PP1)-dependent pathway culminating in phosphorylation of serine368 on connexin43 (pSer368-Cx43). However, characterisation of corresponding pathways in left atrial myocardium, which have a more complex connexin subtype profile, is undefined and was the aim of this study. Gj was measured in guinea-pig left atrium from the frequency-dependent variation of intracellular impedance; intracellular [Ca2+], ([Ca2+]i) in low-Na solution was measured by Fura-2 fluorescence. Phosphorylation of guinea-pig Ser368-Cx43 residues was measured by Western blot; Cx40 was immunoprecipitated and probed for serine/threonine residue phosphorylation. Low-Na solution reversibly reduced Gj, in turn attenuated or prevented by calcineurin inhibitors cyclosporin-A or CAIP, respectively. Moreover, Ser368-Cx43 phosphorylation in low-Na solution was also prevented by CAIP. Changes were partially prevented by fostreicin (FST), a protein phosphatase-2A (PP2A) inhibitor; but not by tautomycin, a PP1 inhibitor. Serine/threonine residues on Cx40 were also phosphorylated in low-Na solution; prevented by CAIP and attenuated by FST. Reduced Gj with raised [Ca2+]i is paralleled by a changed Cx43/Cx40 phosphorylation status; changes mediated by calcineurin and PP2A-dependent pathways, but not PP1. The pharmacological profile underlying changes to guinea-pig atrial gap junction electrical conductance with raised intracellular [Ca2+]i is fundamentally different from that in ventricular myocardium. This provides a targeted drug model whereby atrial and ventricular myocardium can be selectively targeted to correct conduction defects.