The distribution of serotonin (5-HT)-like immunoreactive (5-HT-LI) nerves and the potential role of 5-HT as a vasoconstrictor transmitter in large cerebral arteries of the rabbit were examined. 5-HT-LI fibers with weak immunofluorescence were observed in the anterior cerebral, middle cerebral, and basilar arteries when fixed by immersion after dissection from exsanguinated animals. The 5-HT-LI fibers, however, were not detected in these arteries when fixed either in vitro or in situ after first being perfused with Krebs solution in situ to flush the blood component from the lumen prior to dissection. In these arteries, 5-HT-LI nerve fibers with intense immunofluorescence, however, reappeared following incubation with 5-HT in vitro. The intensity of the 5-HT-LI fibers seemed to be proportional to the duration and 5-HT concentration during incubation. Following chronic surgical sympathectomy, 5-HT-LI fibers were not detected in arteries before or after incubation with 5-HT. Transmural nerve stimulation elicited constriction in 50% of the control arterial segments examined. The constriction was not affected by ketanserin but was prevented by guanethidine and chronic surgical sympathectomy. The remaining arterial segments that did not respond on transmural nerve stimulation, however, became constrictive on transmural nerve stimulation following incubation with 5-HT in vitro. The constriction was blocked by ketanserin and clonidine. These results demonstrate that the large cerebral artery of the rabbit brain has extremely sparse or no authentic 5-HT-LI nerves. The intense 5-HT-LI nerves observed following incubation with 5-HT are primarily due to uptake of 5-HT into sympathetic nerves, and on transmural nerve stimulation 5-HT can be released to induce vasocon-striction. It is suggested that 5-HT can act as an alternative sympathetic vasoconstrictor transmitter in cerebral arteries.