Introduction: The association between sudden cardiac death and heart failure, ischemia and arterial hypertension is well established. However, the role cardiac edema plays in sudden cardiac death, a state common to the aforementioned pathophysiologies, is less well understood. We hypothesized that larger distances between myocytes caused by larger extracellular volumes (VECS) leads to increased conduction velocity anisotropy (ARθ), a well established predisposing factor for arrhythmogenesis. Methods: Electrophysiology was quantified in guinea pig by optical voltage mapping during anterior left (LV) or right ventricular (RV) epicardial pacing. VECS was either reduced by perfusing albumin (3g/L) or increased by perfusing mannitol (26.1 g/l) . Results: ARθ was significantly greater in the LV compared to RV under control conditions. While, LV and RV myocytes have similar widths, LV myocytes are significantly shorter than RV myocytes (129±25 and 153±30 ï­m respectively, p<0.05). ARθ is inconsistent with cell size anisotropy between the LV and RV. Decreasing VECS increased LV and RV transverse conduction velocity (θT) by 33±4 and 71±10% respectively (p<0.05) while longitudinal conduction velocity (θL) did not change. Both LV and RV ARθ were significantly reduced compared to control (p<0.05). Conversely, increasing VECS decreased only θT in LV by 16±5% (p<0.05) but both θT (24±4%, p<0.05) and θL (9±2%, p<0.05) in RV. ARθ was increased relative to control in the RV (p<0.05) but not in the LV. These data demonstrate that VECS changes evoke regionally heterogeneous changes in both θ and ARθ. Conclusions: Since, the link between increased ARθ and conduction failure is well established, these data suggest that increasing myocyte separation by edema may be an important pro-arrhythmic factor. Therefore, myocardial fluid balance and myocyte separation may offer novel targets to reduce the risk of sudden cardiac death.