PRL receptor gene expression was visualized in various tissues by in situ hybridization, using 35S-labeled probes unique to each form of receptor. Tissues were removed rapidly from adult male and female rats and placed in liquid nitrogen. Cryostat sections (10 microns) were prepared, fixed, pretreated, and dehydrated before incubation with the various probes. Hybridization was performed overnight, after which the slides were first exposed to autoradiographic film and then dipped in nuclear emulsion and exposed for 1-2 weeks. The specificity of the signal was studied by competition and using radiolabeled heterologous probes. Some tissues show no expression of either form of receptor mRNA, such as olfactory bulb and penis. Macroautoradiogram signals (optical density) were compared to a standard curve to observe the variation in mRNA expression, which was expressed in arbitrary units. Sex differences in the expression of PRL receptors were seen in a number of tissues, such as adrenal gland and pituitary. Expression of mRNAs specific to the long form of PRL receptor was predominant in adrenal gland, pituitary, thymus, spleen, skin, heart, and skeletal muscle, whereas the short form was expressed to a greater extent in kidney and lung. At the light microscopic level, the silver grains observed by epipolarization or light field were seen in the specific regions or cells that express PRL receptor mRNAs. In conclusion, the long form transcript predominates in most tissues, except kidney and lung. The advantage of in situ hybridization is that it allows the identification of specific regions or cells expressing mRNAs to be identified. The actions of PRL have not been identified in all tissues expressing PRL receptor transcripts, nor has any definitive correlation been made with the expression of short and long forms of PRL receptor and function.

Prolactin receptor gene expression was visualized in various tissues by in situ hybridization. Probes specific to the intracellular domains of the short and long form of receptor were prepared. The specificity of these signals was controlled by competition with excess unlabeled homologous probes or heterologous probes; moreover, some tissues, such as penis and vagina, show no expression of either form of receptor mRNA. Macroautoradiogram signals (optical density) were quantified and expressed in arbitrary units. The long form of receptor mRNA was preferentially expressed in testis, epididymis, prostate, seminal vesicle, and mammary gland from lactating animals, whereas the expression of the two forms of mRNA was equivalent in ovary, uterus, and pregnant mammary gland. Signals were also localized at the light microscopic level to individual cells. This approach has permitted the precise localization of prolactin receptor mRNAs in reproductive tissues. Actions of prolactin have not been demonstrated in all tissues expressing receptor transcripts; thus it will be interesting to correlate the expression of long and short forms of receptor with specific functions.

Prolactin (PRL) receptors have been identified in both classical and nonclassical target organs. Actions of PRL on the secretion of anterior pituitary hormones have been reported. In order to confirm the presence of PRL receptors on specific cell types of the anterior pituitary, immunocytology on ultrathin frozen sections was carried out using monoclonal antibodies specific to the rat PRL receptor. Anterior pituitary glands were removed and fixed in 4% paraformaldehyde or 2.5% glutaraldehyde and postfixed in 1% osmium tetroxide. Tissues were frozen and ultrathin sections were made by cryoultramicrotomy. In anterior pituitary cells, all endocrine cell populations showed PRL-R-like immunoreactivity. The relative labeling frequency based on immunolabeling studies was somatotrophs > lactotrophs > thyrotrophs > corticotrophs > gonadotrophs. Within all endocrine cell types, no apparent differences could be observed in the subcellular localization of PRL-R-like immunoreactivity. At the subcellular level, gold particles, either isolated or in groups of two, were seen at the level of the plasma membrane, in the cytoplasmic matrix, in the vicinity of the endoplasmic reticulum and in the nucleus.

Several effects of prolactin have been characterized in various tissues of the gastrointestinal tract. In the present study, the expression of short and long forms of prolactin receptor was explored and quantified in the digestive tract and correlated to the prolactin specific functions. Sections of all digestive tissues were analyzed by in situ hybridization, using 35S-labeled oligoprobes unique to each form of receptor. Macroautoradiogram signals were quantified and expressed in arbitrary units. In rat liver, prolactin receptor mRNAs are expressed to a much greater degree in females than in males. The short-form transcript is significantly expressed to a greater degree in liver, whereas the long form predominates in the pancreas and esophagus. In the remainder of the gastrointestinal tract, there is an equivalent distribution of short- and long-form transcripts. Relatively high signal intensities are seen in the stomach, duodenum, jejunum, ileum, and colon, whereas the rectum is essentially negative. The identification of prolactin receptor gene expression to limited regions should help establish specific functions associated with this hormone in the digestive tissues.

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