We investigated whether physical exercise can affect platelet L-arginine - nitric oxide pathway in spontaneously hypertensive rats (SHR). Sixteen male SHR and 16 Wistar Kyoto rats (WKY) were divided among exercise (EX) and sedentary (SED) groups. After 20 weeks of treadmill training, systolic blood pressure (mm Hg) was significantly lower in exercised spontaneously hypertensive rats (SHR/EX; 138 ± 8) than in sedentary spontaneously hypertensive rats (SHR/SED; 214 ± 9). Exercise significantly increased platelet L-arginine transport (pmol L-arginine·(10(9) cells)(-1)·min(-1)), assessed by incubation with L-[(3)H]-arginine, in both WKY (SED, 0.196 ± 0.054 compared with EX, 0.531 ± 0.052) and SHR (SED, 0.346 ± 0.076 compared with EX, 0.600 ± 0.049). Nitric oxide synthase (NOS) activity (pmol L-citrulline·(10(8) cells)(-1)), measured by the conversion of L-[(3)H]-arginine to L-[(3)H]-citrulline, was significantly increased in SHR/EX (0.072 ± 0.007) compared with SHR/SED (0.038 ± 0.007), but no changes were observed in WKY. The iNOS and eNOS protein levels assessed by Western blot were not affected by exercise. This upregulation of the platelet L-arginine-NO pathway may attenuate the risk of thromboembolic events, supporting the role of exercise in hypertension management.