Nerve Growth Factor (NGF) is a neurotrophin involved in growth and differentiation of central cholinergic neurons. In this study a maternal separation paradigm was used to test whether levels of NGF might be affected by brief manipulations of rat pups early during ontogeny. The expression of NGF mRNA was examined in 3-day-old rat pups following 45 min maternal separation using in situ hybridization. Early maternal separation in neonatal rats resulted in increased expression of NGF mRNA in the dentate gyrus and the hilus of the hippocampus. NGF protein levels measured (by means of a sensitive ELISA assay) in the whole hippocampus the day following the separation procedure did not differ in separated vs. nonseparated pups. These data indicate that brief manipulations performed early during development can affect hippocampal NGF expression.