The actions of the cholinergic agonists acetylcholine (ACh), nicotine (NIC) and carbamylcholine (CARB) were studied in the supraoptic nucleus with extracellular recordings from coronal slices of the hypothalamus of the rat. Agonists were either applied to the bath or pressure-ejected from micropipettes. Acetylcholine, NIC and CARB produced short-latency increases in discharge rate of about one-half of the cells of the supraoptic nucleus, particularly if the cell was spontaneously active. Excitations produced by NIC were typically brief and the firing rate often declined during prolonged exposure to the agonist. A vigorous excitation frequently occurred in normal medium immediately after cessation of prolonged application of NIC. Local application of glutamate (GLUT) reliably excited cells of the supraoptic nucleus. In some of the cells that were unresponsive to cholinergic agonists, ACh and NIC enhanced responses to electrical stimulation in the dorsolateral area. The nicotinic cholinergic antagonists, hexamethonium (HEX) and d-tubocurarine (dTC), reduced or blocked excitations induced by NIC, but not those from GLUT. This study provides further evidence that nicotinic receptors can mediate cholinergic influences on cells of the supraoptic nucleus. However, these effects were not observed in all cells in the region of the supraoptic nucleus and the excitations by NIC showed densensitization. Acetylcholine may subserve a modulatory role to other transmitters in the supraoptic nucleus.