Prolactin (Prl) receptor (Prlr) gene is expressed in various brain regions, with the highest level present in the choroid plexus, a site for receptor-mediated PRL transport from the blood to cerebrospinal fluid. We investigated the regulatory mechanism ofPrlrgene expression by PRL in the murine choroid plexus. We first examined the organization of the alternative first exons in murinePrlrgene. In addition to the three known first exons, mE11, mE12, and mE13, two first exons, mE14and mE15, were newly identified by cDNA cloning. Each first exon variant ofPrlrmRNA exhibited tissue-specific or generic expression. In the choroid plexus of mice, the expression levels ofmE13-,mE14-, andmE15-PrlrmRNAs were increased in the lactating mice compared with those in the diestrus mice. Furthermore, the expression level ofmE14-PrlrmRNA was decreased in the PRL-deficient (Prl−/−) mice compared with the PRL-normal (Prl+/+andPrl+/−) mice. In the ovariectomizedPrl−/−mice, the expression level ofmE14-PrlrmRNA was significantly increased by PRL administration but not by 17β-estradiol administration. The expression levels of the two last exon variants ofPrlrmRNAs, encoding the long and short cytoplasmic regions of PRLR, were also increased in the lactating mice and decreased in thePrl−/−mice. These findings suggest that PRL stimulates thePrlrgene expression through the transcriptional activation of mE14first exon, leading to increases in the long- and short-form variants ofPrlrmRNA in the murine choroid plexus.