Centrally acting prolactin has been shown to have anti-stress effects by modulating the activity of the hypothalamic–pituitary–adrenal axis. We tested the hypothesis that prolactin directly targets hypothalamic corticotropin-releasing hormone (CRH) neurons. In situ hybridisation confirmed expression of mRNA encoding the long, but not the short, isoform of the prolactin receptor (PRLR) within the paraventricular nucleus (PVN) of the virgin rat; however, only 6% of CRH neurons expressed long-form Prlr mRNA. Examination of the functional response of CRH neurons to intracerebroventricular prolactin (500 ng) showed that these neurons did not respond with activation of phosphorylated signal transducer and activator of transcription 5 (pSTAT5), a marker of long-form PRLR activation. However, as only a subset of neurons expressing Crh mRNA could be detected using immunohistochemistry, we utilised a transgenic mouse model to label CRH neurons with a fluorescent reporter (CRH-Cre-tdTomato). In lactating animals, chronically elevated prolactin levels resulted in significantly increased pSTAT5 expression in the PVN. Overall, few tdTomato-labelled CRH neurons were double-labelled, although a small subset of CRH neurons in the caudal PVN were pSTAT5 positive (approximately 10% of tdTomato neurons at this level, compared to 1% in the rostral PVN). These data suggest that most CRH neurons do not respond directly to prolactin. To confirm that prolactin was not activating another signalling pathway, we used a transgenic mouse line to label PRLR-expressing neurons with Cre-dependent green fluorescent protein (GFP) expression (CRH-Cre-Prlrlox/lox). No GFP-expressing cells were evident in the PVN, indicating that in the mouse, as in the rat, the CRH neurons do not express either PRLR isoform. Together these data showed that the anti-stress effects of prolactin are not the result of prolactin directly regulating CRH neurons.
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This work was supported by a Programme Grant from the Health Research Council of New Zealand (14–568). PG was supported by a University of Otago PhD Scholarship.
Compliance with Ethical Standards
Conflict of interest
The authors declare that they have no conflict of interest.
All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.
Augustine RA, Grattan DR (2008) Induction of central leptin resistance in hyperphagic pseudopregnant rats by chronic prolactin infusion. Endocrinology 149:1049–1055CrossRefPubMedGoogle Scholar
Augustine RA, Kokay IC, Andrews ZB, Ladyman SR, Grattan DR (2003) Quantitation of prolactin receptor mRNA in the maternal rat brain during pregnancy and lactation. J Mol Endocrinol 31:221–232CrossRefPubMedGoogle Scholar
Bloom FE, Battenberg EL, Rivier J, Vale W (1982) Corticotropin releasing factor (CRF): immunoreactive neurones and fibers in rat hypothalamus. Regul Pept 4:43–48CrossRefPubMedGoogle Scholar
Blume A, Torner L, Liu Y, Subburaju S, Aguilera G, Neumann ID (2009) Prolactin activates mitogen-activated protein kinase signaling and corticotropin releasing hormone transcription in rat hypothalamic neurons. Endocrinology 150:1841–1849. doi:10.1210/en.2008-1023CrossRefPubMedGoogle Scholar
Bole-Feysot C, Goffin V, Edery M, Binart N, Kelly PA (1998) Prolactin (PRL) and its receptor: actions, signal transduction pathways and phenotypes observed in PRL receptor knockout mice. Endocr Rev 19:225–268CrossRefPubMedGoogle Scholar
Bridges RS, Numan M, Ronsheim PM, Mann PE, Lupini CE (1990) Central prolactin infusions stimulate maternal behavior in steroid-treated, nulliparous female rats. Proc Natl Acad Sci USA 87:8003–8007CrossRefPubMedPubMedCentralGoogle Scholar
Casanova E, Fehsenfeld S, Mantamadiotis T, Lemberger T, Greiner E, Stewart AF, Schutz G (2001) A CamKIIalpha iCre BAC allows brain-specific gene inactivation. Genesis 31:37–42CrossRefPubMedGoogle Scholar
da Costa AP, Wood S, Ingram CD, Lightman SL (1996) Region-specific reduction in stress-induced c-fos mRNA expression during pregnancy and lactation. Brain Res 742:177–184CrossRefPubMedGoogle Scholar
Drago F, Continella G, Conforto G, Scapagnini U (1985) Prolactin inhibits the development of stress-induced ulcers in the rat. Life Sci 36:191–197CrossRefPubMedGoogle Scholar
Feher P, Olah M, Bodnar I et al (2010) Dephosphorylation/inactivation of tyrosine hydroxylase at the median eminence of the hypothalamus is required for suckling-induced prolactin and adrenocorticotrop hormone responses. Brain Res Bull 82:141–145. doi:10.1016/j.brainresbull.2010.02.006CrossRefPubMedGoogle Scholar
Forsyth IA, Wallis M (2002) Growth hormone and prolactin–molecular and functional evolution. J Mammary Gland Biol Neoplasia 7:291–312CrossRefPubMedGoogle Scholar
Freeman ME, Kanyicska B, Lerant A, Nagy G (2000) Prolactin: structure, function, and regulation of secretion. Physiol Rev 80:1523–1631PubMedGoogle Scholar
Fujikawa T, Soya H, Yoshizato H, Sakaguchi K, Doh-Ura K, Tanaka M, Nakashima K (1995) Restraint stress enhances the gene expression of prolactin receptor long form at the choroid plexus. Endocrinology 136:5608–5613. doi:10.1210/endo.136.12.7588315CrossRefPubMedGoogle Scholar
Fujikawa T, Tamura K, Kawase T et al (2005) Prolactin receptor knockdown in the rat paraventricular nucleus by a morpholino-antisense oligonucleotide causes hypocalcemia and stress gastric erosion. Endocrinology 146:3471–3480. doi:10.1210/en.2004-1528CrossRefPubMedGoogle Scholar
Kokay IC, Bull PM, Davis RL, Ludwig M, Grattan DR (2006) Expression of the long form of the prolactin receptor in magnocellular oxytocin neurons is associated with specific prolactin regulation of oxytocin neurons. Am J Physiol Regul Integr Comp Physiol 290:R1216–R1225. doi:10.1152/ajpregu.00730.2005CrossRefPubMedGoogle Scholar
Larsen CM, Grattan DR (2010) Prolactin-induced mitogenesis in the subventricular zone of the maternal brain during early pregnancy is essential for normal postpartum behavioral responses in the mother. Endocrinology 151:3805–3814. doi:10.1210/en.2009-1385CrossRefPubMedGoogle Scholar
Lightman SL, Young WS 3rd (1989) Lactation inhibits stress-mediated secretion of corticosterone and oxytocin and hypothalamic accumulation of corticotropin-releasing factor and enkephalin messenger ribonucleic acids. Endocrinology 124:2358–2364CrossRefPubMedGoogle Scholar
Madisen L, Zwingman TA, Sunkin SM et al (2010) A robust and high-throughput Cre reporting and characterization system for the whole mouse brain. Nat Neurosci 13:133–140. doi:10.1038/nn.2467CrossRefPubMedGoogle Scholar
Neumann ID, Johnstone HA, Hatzinger M et al (1998) Attenuated neuroendocrine responses to emotional and physical stressors in pregnant rats involve adenohypophysial changes. J Physiol 508(Pt 1):289–300CrossRefPubMedPubMedCentralGoogle Scholar
Neumann ID, Kromer SA, Toschi N, Ebner K (2000a) Brain oxytocin inhibits the (re)activity of the hypothalamo-pituitary-adrenal axis in male rats: involvement of hypothalamic and limbic brain regions. Regul Pept 96:31–38CrossRefPubMedGoogle Scholar
Neumann ID, Torner L, Wigger A (2000b) Brain oxytocin: differential inhibition of neuroendocrine stress responses and anxiety-related behaviour in virgin, pregnant and lactating rats. Neuroscience 95:567–575CrossRefPubMedGoogle Scholar
Torner L, Toschi N, Pohlinger A, Landgraf R, Neumann ID (2001) Anxiolytic and anti-stress effects of brain prolactin: improved efficacy of antisense targeting of the prolactin receptor by molecular modeling. J Neurosci 21:3207–3214PubMedGoogle Scholar
Torner L, Toschi N, Nava G, Clapp C, Neumann ID (2002) Increased hypothalamic expression of prolactin in lactation: involvement in behavioural and neuroendocrine stress responses. Eur J Neurosci 15:1381–1389CrossRefPubMedGoogle Scholar
Walker CD, Tilders FJ, Burlet A (2001) Increased colocalization of corticotropin-releasing factor and arginine vasopressin in paraventricular neurones of the hypothalamus in lactating rats: evidence from immunotargeted lesions and immunohistochemistry. J Neuroendocrinol 13:74–85CrossRefPubMedGoogle Scholar
Wamsteeker Cusulin JI, Fuzesi T, Watts AG, Bains JS (2013) Characterization of corticotropin-releasing hormone neurons in the paraventricular nucleus of the hypothalamus of Crh-IRES-Cre mutant mice. PLoS One 8:e64943CrossRefPubMedPubMedCentralGoogle Scholar
Weber RF, Calogero AE (1991) Prolactin stimulates rat hypothalamic corticotropin-releasing hormone and pituitary adrenocorticotropin secretion in vitro. Neuroendocrinology 54:248–253CrossRefPubMedGoogle Scholar
Windle RJ, Wood S, Shanks N et al (1997) Endocrine and behavioural responses to noise stress: comparison of virgin and lactating female rats during non-disrupted maternal activity. J Neuroendocrinol 9:407–414CrossRefPubMedGoogle Scholar