Medial septal-nucleus of the diagonal band of Broca area (MS-nDBB) neurons, identified by their antidromic response to the electrical stimulation of the fimbria and/or hippocampus, were studied in the rat under various conditions of anesthesia. These septo-hippocampal neurons (SHNs) were classified into 4 groups on the basis of: (i) their antidromic latency; and (ii) the presence or absence of a rhythmically bursting pattern of spontaneous discharge. The rhythmically bursting activity (43.5% of the SHNs) was highly dependent on the anesthetic conditions. The groups of SHNs differed in their mean conduction velocity and rate of spontaneous activity. In contrast, irrespective of their classification in a particular group, the large majority of the SHNs could be excited by the iontophoretic application of cholinergic agonists. Beside the SHNs, two other populations of MS-nDBB neurons could be identified by electrical antidromic stimulation: neurons projecting to the amygdala (Am) and neurons projecting through the medial forebrain bundle (MFB). Half of the MS-nDBB neurons projecting to Am were also antidromically driven from the fimbria. The axonal branch projecting to Am had a slower conduction velocity than that projecting to hippocampus. In contrast MS-nDBB neurons projecting through the MFB were never antidromically driven from the fimbria, although they received orthodromic inputs. They had a slower conduction velocity than the other groups of MS-nDBB neurons.