Exercise, capture, and handling stress in fish can elevate extracellular K+ concentration ([K+]o) with potential impact on heart function in a temperature- and frequency-dependent manner. To this end, the effects of [K+]o on the excitability of ventricular myocytes of winter-acclimatized roach (Rutilus rutilus) (4 ± 0.5°C) were examined at different test temperatures and varying pacing rates. Frequencies corresponding to in vivo heart rates at 4°C (0.37 Hz), 14°C (1.16 Hz), and 24°C (1.96 Hz) had no significant effect on the excitability of ventricular myocytes. Acute increase of temperature from 4 to 14°C did not affect excitability, but a further rise to 24 markedly decreased excitability: stimulus current and critical depolarization needed to elicit an action potential (AP) were ~25 and 14% higher, respectively, at 24°C than at 4°C and 14°C (P < 0.05). This depression could be due to temperature-related mismatch between inward Na+ and outward K+ currents. In contrast, an increase of [K+]o from 3 to 5.4 or 8 mM at 24°C reduced the stimulus current needed to trigger AP. However, other aspects of excitability were strongly depressed by high [K+]o: maximum rate of AP upstroke and AP duration were drastically (89 and 50%, respectively) reduced at 8 mM [K+]o in comparison with 3 mM (P < 0.05). As an extreme case, some myocytes completely failed to elicit all-or-none AP at 8 mM [K+]o at 24°C. Also, amplitude and overshoot of AP were reduced by elevation of [K+]o (P < 0.05). Although high [K+]oantagonizes the negative effects of high temperature on excitation threshold, the precipitous depression of the rate of AP upstroke and complete loss of excitability in some myocytes suggest that the combination of high temperature and high [K+]o will severely impair ventricular excitability in roach.