A monoclonal antibody (Ab8) to choline acetyltransferase (ChAT) was used to locate structures showing ChAT-like immunoreactivity (ChAT-IR) in the brain of a teleost fish, the midshipman (Porichthys notatus). ChAT is the synthetic enzyme for acetylcholine found in neurons using that neurotransmitter; thus ChAT-IR may be interpreted as indicating putative cholinergic activity. Robust staining is seen in all cranial nerve motor nuclei. In addition, the brainstem of Porichthys is distinguished by two other expansive ChAT-IR zones: a sonic motor nucleus, which innervates swimbladder "drum" muscles, and an octavolateralis efferent nucleus, which innervates acoustic, vestibular, and lateral line end organs. Scattered labeled cells are found in several cranial sensory nuclei--the vagal lobe, and the main and descending trigeminal nuclei. ChAT-IR cells form restricted subpopulations in other noncranial nerve nuclei, including the granule cell layer of the cerebellum; superior, medial, and inferior divisions of the reticular formation; the stratum periventriculare of the midbrain's optic tectum; and the nucleus isthmi in the midbrain tegmentum. In the telencephalon, a dense population of ChAT-IR cells is found in the ventral nucleus of area ventralis; terminals and fine fibers are found in the dorsal, medial, and central nuclei of area dorsalis. Together, the data represent the first complete report of ChAT-IR cell bodies in the brain of any nonmammal with the monoclonal antibody Ab8, which has already been extensively used on a variety of vertebrate brains. The results are thus discussed from a comparative viewpoint, considering reports of ChAT-IR in different taxa.