To analyze the effect of hyperthermia on the vascular response, the isometric response of isolated rabbit femoral artery segments was recorded at 37°C and hyperthermia (41 and 44°C). Contraction to potassium (5 × 10³-5 × 10² M) was significantly greater at 41 and 44 than at 37°C and increased by inhibition of nitric oxide (NO) synthesis with N-nitro-L-arginine (L-NNA; 10⁴ M) or endothelium removal at 37°C but not at 41 or 44°C. Norepinephrine (10⁹-10⁴ M) produced a concentration-dependent contraction greater at 41 or 44 than at 37°C and not modified by endothelium removal or L-NNA at either temperature. Phenylephrine (10⁹-10⁴ M) produced a contraction increased by warming to 44°C but not to 41°C. The specific ₂-adrenoceptor agonist BHT-920 produced a weak contraction, reduced by the ₁-adrenoceptor antagonist prazosin (10⁶ M) and increased at 44°C but not at 41°C. The concentration-dependent contraction to endothelin-1 (ET-1; 10¹¹-10⁷ M) was increased by warming to 41 and 44°C and by endothelium removal or L-NNA at 37°C but not at 41 or 44°C. Response to ET-1 was reduced by endothelin ET_A-receptor antagonist BQ-123 (10⁵ M) and ET_B-receptor antagonist BQ-788 (10⁵ M). In arteries precontracted with ET-1 (10⁸-3 × 10⁸ M), relaxation to sodium nitroprusside (10⁸-10⁴ M) was increased at 41 and 44°C vs. at 37°C, but that of ACh (10⁸-10⁴ M) or adenosine (10⁸-10⁴ M) was not different at all temperatures studied. Relaxation to ACh, but not adenosine, was reduced similarly by L-NNA at all temperatures studied. These results suggest hyperthermia in muscular arteries may inhibit production of, and increase dilatation to, NO, resulting in unchanged relaxation to ACh and increased constriction to KCl and ET-1, and may increase constriction to stimulation of ₁-adrenoceptors by NO-independent mechanisms.