The purpose of the present study was to characterize in detail the 24 h blood pressure (BP) phenotype of mice lacking the gene for endothelial nitric oxide synthase (eNOS-/-) and the corresponding control strain (C57Bl/6J). Twenty-four hour BP recordings were made in conscious 12- to 16-week-old male mice 10 days following the implantation of a BP telemeter (n = 9 per group). The BP and heart rate of both strains were markedly affected by brief locomotor activity cycles, resulting in bimodal distributions of BP and heart rate within both light and dark periods. Data from active periods were associated with the higher of the two modes, whereas data from inactive periods were associated with the lower of the two modes. In eNOS-/- mice, the 24 h average BP level was significantly elevated (+15 %, 104 +/- 2 vs. 119 +/- 1 mmHg), as was its daily range (+44 %), its coefficient of variation (+26 %), dark-light difference (+48 %), and the separation of the two modes of its distribution (+41 %). Pulse pressure was also significantly greater (+23 %) in eNOS-/- mice. The 24 h heart rate level did not differ between control and eNOS-/- mice. Considerable variation was noted among previously published values of BP in eNOS-/- mice, but not in the corresponding control mice. Our results indicate that eNOS-/- mice have mild hypertension that is accompanied by more pronounced increases in BP lability and/or reactivity. Our results also demonstrate a marked effect of locomotor activity on BP in mice, which may confound short-term measurements of BP.