Abstract The fate of prolactin in the cerebrospinal fluid was studied by the use of fluorescein labelled prolactin. The distribution was compared to the immunocytochemical distribution of endogenous prolactin. High intensity fluorescence was found in the ependyma of the area postrema, rostral dorsolateral cerebral aqueduct close to the subcommissural organ, and in some cells of the floor of the cerebral aqueduct. This distribution was not seen when excess unlabelled prolactin was injected. The results suggest prolactin uptake from CSF at specific sites which correspond to sites of localization of immunoreactive prolactin.