To examine the contribution of sarcoplasmic reticulum Ca²⁺ ATPase (SERCA2a) to early heart failure, we subjected transgenic (TG) mice expressing SERCA2a gene and wild-type (WT) mice to aortic stenosis (AS) for 7 weeks. At an early stage of hypertrophy (4-week AS), in vivo hemodynamic and echocardiographic indices were similar in TG and WT mice. By 7 weeks of AS, which is the stage of early failure in this model, TG mice with AS had lower mortality than WT mice with AS (6.7% versus 29%). The magnitude of left ventricular (LV) hypertrophy was similar in WT and TG 7-week AS mice. In vivo LV systolic function was higher in TG than in WT 7-week AS mice. In LV myocytes loaded with fluo-3, fractional cell shortening and the amplitude of the [Ca²⁺]_i transients were higher in TG than in WT 7-week AS mice under baseline conditions (0.5 Hz, 1.5 mmol/L [Ca²⁺]_o, 25°C). The rates of relengthening and decay in [Ca²⁺]_i were faster in TG than in WT 7-week AS myocytes. In myocytes from WT 7-week AS compared with sham-operated WT mice, contractile reserve in response to rapid pacing was depressed with impaired augmentation of both peak-systolic [Ca²⁺]_i and the SR Ca²⁺ load. In contrast, contractile reserve and the capacity to augment SR Ca²⁺ load were maintained in TG 7-week AS mice. SERCA2a protein levels were depressed in WT 7-week AS mice, but were preserved in TG 7-week AS mice. These data suggest that defective SR Ca²⁺ loading contributes to the onset of contractile failure in animals with chronic pressure overload.