The effect of endurance run training on outward K⁺ currents with rapidly inactivating (I_to) and sustained or slowly inactivating (I_sus) characteristics was examined in left ventricular (LV) cardiocytes isolated from sedentary (Sed) and treadmill-trained (Tr) female Sprague-Dawley rats. Isolated LV cardiocytes were used in whole cell patch-clamp studies to characterize whole cell I_to and I_sus. Peak I_to was greatest in cells isolated from the Tr group. When I_to was corrected for cell capacitance to yield a current density, most, but not all, of the Sed vs. Tr differences in I_to magnitude were eliminated. Regardless of how I_to was expressed (e.g., I_to or I_to density), the time required to achieve a peak value was markedly shortened in the cardiocytes isolated from the Tr group. Training elicited a reduction in I_sus density. Action potential characteristics were determined in Sed and Tr cardiocytes in primary culture. Training did not affect resting membrane potential, whereas peak membrane potential was reduced and time to peak membrane potential was prolonged in the Tr group. In addition, time to 50% repolarization was significantly increased in cells from the Tr group. Collectively, these data indicate that I_to and I_sus characteristics are altered by training in isolated LV cardiocytes. These alterations in I_to and I_sus may be responsible, at least in part, for the training-induced alterations in action potential configuration in cardiocytes in primary culture.