Cerebral blood vessels contain both sympathetic and nitric oxide (NO) synthase (NOS)-containing nerves. NO has been proposed to modulate smooth muscle function and adrenergic nerve activity, and the nature of this modulation is controversial: some data show NO inhibits norepinephrine (NE) release, whereas others suggest that NO augments release. To test the hypothesis that in cerebral arteries NO released by NOS-containing nerves augments stimulation-evoked NE release, we used direct measurement of NE and NO release in isolated sheep middle cerebral arteries. The facial artery, which has not been reported to be innervated with NOS-containing nerves, was used as an artery comparison model. HPLC and redox electrochemical detection was used to measure NE, and NO was measured by chemiluminescence. Stimulation-evoked NE release from the middle cerebral artery significantly declined in the presence of the NOS inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME). The effect of L-NAME was reversed by the addition of the NO donor S-nitroso-N-acetyl-DL-penicillamine. In contrast, in facial arteries, L-NAME had no effect on stimulation-evoked NE release, whereas S-nitroso-N-acetyl-DL-penicillamine still significantly elevated NE release. Activation of perivascular nerves significantly increased NE release in both the middle cerebral and facial arteries. However, when NO was measured in the same samples, stimulation-evoked release of NO was significantly increased compared with basal release only in middle cerebral arteries. These data support the concept that cerebral arteries in the sheep contain both adrenergic and NOS-containing nerves. Furthermore, this study provides succinct evidence that NO released from NOS nerves augments stimulation-evoked NE release.