Mitogen-activated protein (MAP) kinases have been shown to be activated by various growth factors in cultured or isolated cardiomyocytes. However, little is known about the regulation of MAP kinases in vivo, especially in clinically important conditions, such as hypertension and senescence. In this study, we assessed mechanical overload–induced activation of myocardial MAP kinases in beating hearts from hypertensive or senescent rats. Fifteen minutes of left ventricular hemodynamic overload activated MAP kinase activity by 2.2-fold (P<0.05) in 4-week-old Wistar-Kyoto rats. The age-matched spontaneously hypertensive rats had greater MAP kinase activity than did Wistar-Kyoto rats both at baseline (1.4 times,P<0.05) and after the hemodynamic overload (1.7 times,P<0.05). Myocardial MAP kinase protein level, assessed by Western blot analysis, was also higher (1.6 times,P<0.01) in spontaneously hypertensive rats. In contrast, aged (18-month-old) Fischer 344 rats, which were known to have a diminished capacity of hypertrophy in response to mechanical stress, had lower MAP kinase activity both at baseline (63%,P<0.01) and after the hemodynamic overload (52%,P<0.05). Their MAP kinase protein level was lower (38%,P<0.01) than that in young (6-month-old) adults. Alterations in MAP kinase may contribute to changes in hypertrophic response in these animals.