The entorhinal cortex (EC) of the rat has been divided into medial (MEA) and lateral (LEA) subdivisions. In order to analyze its afferent connections, small deposits of horseradish peroxidase (HRP) were placed at various loci within EC. The patterns of retrograde cell-labeling charted in 18 such cases suggested that EC is projected upon by several allocortical and subcortical structures and that there are differences in the afferent connections of the two subdivisions. Thus, although HRP injection of either division of EC led to cell-labeling in the hippocampal formation, most in ammonic field CA1 and the subiculum, several cells of the presubiculum were preferentially labeled by injection of MEA. Injections of LEA, but not those in MEA, resulted in substantial cell-labeling in the anterior piriform cortex of both hemispheres. Regardless of the location of its injection site within EC, the enzyme labeled cells in the diagonal band nucleus of Broca, amygdala and claustrum. The pattern of cell-labeling in the diagonal band nucleus extended into the ventrolaterally contiguous nucleus basalis after injection of LEA and into the dorsally contiguous medial septal nucleus after injection of MEA Whereas HRP deposits in either division of EC resulted in cell-labeling in the cortical and medial nuclei of the amygdala, only those deposits which involved LEA led to cell-labeling in the posterior part of the lateral nucleus. In the thalamus, labeled cells were found in the rostral part of the paratenial, periventricular and reuniens nuclei. Finally, at midbrain levels, numerous labeled cells appeared in the dorsal raphe nucleus, a few in the median raphe and locus coeruleus, and, only after rostral EC injection, in the ventral tegmental area.