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Atrial fibrillation (AF) is characterized by decreased L-type calcium current (I Ca,L ) in atrial myocytes and decreased atrial contractility. Oxidant stress and redox modulation of calcium channels are implicated in these pathologic changes. We evaluated the relationship between glutathione content (the primary cellular reducing moiety) and I Ca,L in atrial specimens from AF patients undergoing cardiac surgery. Left atrial glutathione content was significantly lower in patients with either paroxysmal or persistent AF relative to control patients with no history of AF. Incubation of atrial myocytes from AF patients (but not controls) with the glutathione precursor N-acetylcysteine caused a marked increase in I Ca,L . To test the hypothesis that glutathione levels were mechanistically linked with the reduction in I Ca,L , dogs were treated for 48 h with buthionine sulfoximine, an inhibitor of glutathione synthesis. Buthionine sulfoximine treatment resulted in a 24% reduction in canine atrial glutathione content, a reduction in atrial contractility, and an attenuation of I Ca,L in the canine atrial myocytes. Incubation of these myocytes with exogenous glutathione also restored I Ca,L to normal or greater than normal levels. To probe the mechanism linking decreased glutathione levels to down-regulation of I Ca , the biotin switch technique was used to evaluate S-nitrosylation of calcium channels. S-Nitrosylation was apparent in left atrial tissues from AF patients; the extent of S-nitrosylation was inversely related to tissue glutathione content. S-Nitrosylation was also detectable in HEK cells expressing recombinant human cardiac calcium channel subunits following exposure to nitrosoglutathione. S-Nitrosylation may contribute to the glutathione-sensitive attenuation of I Ca,L observed in AF. Atrial fibrillation (AF)2 is characterized by rapid electrical activation of the atria. Persistent AF results in phenotypic electrical and structural remodeling of the atria. Calcium influx via I Ca triggers calcium release from intracellular stores and is an essential first step in excitation-contraction coupling and a critical determinant of atrial contractility. In atrial myocytes from patients with persistent AF (1) or from animals with pacinginduced AF (2), calcium current density is significantly decreased. The time course of contractile impairment following the onset of AF (3) or rapid atrial pacing (4) parallels the down-regulation of I Ca , and contractile recovery proceeding cardioversion follows a time course similar to that of reverse electrical remodeling (4). Reports characterizing the impact of AF on the expression of the calcium channel pore subunit in human AF and in experimental models have had conflicting results (5, 6); the mechanisms by which I Ca,L down-regulation occurs are not fully resolved.In a porcine model of rapid atrial pacing, superoxide production is increased (7), and nitric oxide production is decreased (8). Oxidant-generating pathways can shift the myocyte intracellular redox environment from...

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The canine sterile pericarditis model is characterized by impaired conduction and atrial arrhythmia vulnerability. Electrical and structural remodeling processes caused by the inflammatory response likely promote these abnormalities. In the present study, we tested the hypothesis that altered distribution of atrial connexins is associated with markedly abnormal atrial conduction, thereby contributing to vulnerability to atrial flutter (AFL) and atrial fibrillation (AF) induction and maintenance. During rapid pacing and induced, sustained AFL or AF in five sterile pericarditis (SP) and five normal (NL) dogs, epicardial atrial electrograms were recorded simultaneously from both atria (380 electrodes) or from the right atrium (RA) and Bachmann's bundle (212 electrodes). Tissues from RA sites were subjected to immunostaining and immunoblotting to assess connexin (Cx) 40 and Cx43 distribution and expression. Transmural myocyte (alpha-actinin) and fibroblast (vimentin) volume were also assessed by immunostaining. RA pacing maps showed markedly abnormal conduction in SP, with uniform conduction in NL. Total RA activation time was significantly prolonged in SP vs. NL at 300-ms and 200-ms pacing-cycle lengths. Sustained arrhythmias were only inducible in SP [total: 4/5 (AFL: 3/5; AF: 1/5)]. In NL, Cx40, Cx43, alpha-actinin, and vimentin were homogeneously distributed transmurally. In SP, Cx40, Cx43, and alpha-actinin were absent epicardially, decreased midmyocardially, and normal endocardially. SP increased epicardial vimentin expression, suggesting fibroblast proliferation. Immunoblot analysis confirmed reduced expression of Cx40 and Cx43 in SP. The transmural gradient in the volume fraction of Cx40 and Cx43 in SP is associated with markedly abnormal atrial conduction and is likely an important factor in the vulnerability to induction and maintenance of AFL/AF in SP.

The accumulation of radiolabeled arachidonic acid (AA), immunoblot analysis of subcellular fractions, and immunofluorescence tagging of proteins in intact cells were used to examine the coupling of ANG II receptors with the activity and location of a cytosolic phospholipase A2(cPLA2) in vascular smooth muscle cells (VSMC). ANG II induced the accumulation of AA, which peaked by 10 min and was downregulated by 20 min. A large proportion of the AA released in response to ANG II was due to the activation of a Ca2+-dependent lipase coupled to an AT1 receptor. However, regulation of Ca2+ availability failed to completely block AA release, and a small but significant reduction in ANG II-mediated AA release was observed in the presence of an AT2 antagonist. These findings, coupled with a 25% reduction in the ANG II-induced AA release by an inhibitor specific for a Ca2+-independent PLA2, are consistent with the presence and activation of a Ca2+-independent PLA2. In contrast, immunoblot analysis and immunofluorescence detection showed that the ANG II-mediated translocation of cPLA2to a membrane fraction was exclusively AT1 dependent and regulated by Ca2+ availability. Furthermore, the nucleus was the membrane target. We conclude that ANG II regulates the Ca2+-dependent activation and translocation of cPLA2 through an AT1 receptor and that this event is targeted at the nucleus in VSMC.