Expression and activity of cardiac Na+/Ca2+ exchanger (NCX1) are altered in many disease states. We engineered mice in which the phosphomimetic phospholemman S68E mutant (inhibits NCX1 but not Na+-K+-ATPase) was constitutively overexpressed in a cardiac-specific manner (conS68E). At 4–6 wk, conS68E mice exhibited severe bradycardia, ventricular arrhythmias, increased left ventricular (LV) mass, decreased cardiac output (CO), and ∼50% mortality compared with wild-type (WT) littermates. Protein levels of NCX1, calsequestrin, ryanodine receptor, and α1- and α2-subunits of Na+-K+-ATPase were similar, but sarco(endo)plasmic reticulum Ca2+-ATPase was lower, whereas L-type Ca2+ channels were higher in conS68E hearts. Resting membrane potential and action potential amplitude were similar, but action potential duration was dramatically prolonged in conS68E myocytes. Diastolic intracellular Ca2+ ([Ca2+]i) was higher, [Ca2+]i transient and maximal contraction amplitudes were lower, and half-time of [Ca2+]i transient decline was longer in conS68E myocytes. Intracellular Na+ reached maximum within 3 min after isoproterenol addition, followed by decline in WT but not in conS68E myocytes. Na+/Ca2+ exchange, L-type Ca2+, Na+-K+-ATPase, and depolarization-activated K+ currents were decreased in conS68E myocytes. At 22 wk, bradycardia and increased LV mass persisted in conS68E survivors. Despite comparable baseline CO, conS68E survivors at 22 wk exhibited decreased chronotropic, inotropic, and lusitropic responses to isoproterenol. We conclude that constitutive overexpression of S68E mutant was detrimental, both in terms of depressed cardiac function and increased arrhythmogenesis.