Clinical trials suggest females experience less heart failure (HF) progression, mortality, and arrhythmia frequency. HF increases Na/Ca exchanger (NCX) expression and activity contributing to both depressed contractility and ventricular arrhythmias, but whether gender modifies this effect is unknown. Left ventricular myocytes were isolated from control and from tachycardic pacing-induced failing swine hearts of both sexes. The Ni-sensitive NCX current (INCX) was measured in voltage clamp after blocking other channels. In control myocytes there is no difference in basal INCX and β-adrenergic responsiveness between male and female animals. HF greatly increased INCX and reduced β-adrenergic responsiveness in males compared to females, an effect that was eliminated by PP1. Diuretic therapy (furosemide, 1 mg/kg/day) further enhanced INCX and reduced β-adrenergic responsiveness in females and eliminated the gender difference. Gender-specific differences in calcium handling may contribute to improved survival of females in HF.