Little is known about the effects of a reduction of mechanical load on cellular electrophysiology of the left ventricle. To address this question, hearts of male Lewis rats were unloaded by heterotopic cardiac transplantation. After two weeks of unloading action potentials (AP) were significantly prolonged in endo- and epicardial myocytes and the transmural gradient in AP duration was diminished. In parallel with AP prolongation, the magnitude of the transient outward potassium current (I_to) was significantly decreased in endo- (-80% at V_pip=+40mV) and epicardial (-64% at V_pip =+40mV) myocytes. The steady-state potassium current was solely reduced in endocardial myocytes of unloaded hearts, whereas the inward rectifying potassium current was not affected by unloading. Expression levels of the I_to encoding a-subunits Kv4.2 and Kv4.3 were significantly reduced in unloaded epi-, but not in endocardial myocytes. The expression of KChIP2 mRNA was not altered. Taken together, these results indicate that a reduction of mechanical load predominantly decreases the current density of I_to and leads to an attenuation of the transmural gradient in AP duration. Both, transcriptional and posttranscriptional mechanisms appear to contribute to the downregulation of I_to.