Prolactin regulation of the HPA axis is not mediated by a direct action upon CRH neurons: evidence from the rat and mouse

Abstract

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-Prlr ). 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.

Elevated Prolactin during Pregnancy Drives a Phenotypic Switch in Mouse Hypothalamic Dopaminergic Neurons

“…Images were captured using an Olympus BX61 light microscope fitted with a QImaging Micropublisher 5.0RTV camera (QImaging, Surrey, Canada). Quantification and analysis were performed as described previously (Gustafson et al, 2017) In vitro measurement of met-enkephalin and TH expression in the median eminence Lactating mice (day 10 of lactation) were killed by rapid decapitation, following isoflurane anesthesia. The brain was quickly removed and placed in ice-cold artificial cerebrospinal fluid (aCSF) containing 118 mM NaCl, 3 mM KCl, 11 mM D-glucose, 10 mM HEPES, 25 mM NaHCO 3 , 6 mM MgCl 2 and 0.5 mM CaCl 2 , osmolarity between 295 and 305 mOsm, pH 7.2 when gassed with 5% CO 2 and 95% O 2 .…”

The subfornical organ: A novel site for prolactin action

“…Although the PVN expresses PRL‐R, whether the PRL responses are direct or indirect has not been reported. In the PVN, corticotrophin‐releasing hormone (CRH) neurones are PRL‐responsive but do not co‐express the marker for direct PRL activation, pSTAT5 (phosphorylated signal transducer and activator of transcription 5) . It is therefore possible that the SFO projections to the PVN increase the sensitivity of PVN oxytocin and CRH neurones or may directly signal to these neurones to carry out PRL functions (eg, the PRL‐induced suppression of the stress response).…”

The median eminence detects and responds to circulating prolactin in the male mouse

“…Transgenic animals lacking prolactin receptors in neurones expressing Cam Kinase IIα (Prlr lox/lox /CamK‐Cre mice) or lacking prolactin receptors in GABAergic cells (Prlr lox/lox /VGat‐Cre mice) or glutamatergic neurones (Prlr lox/lox /VGlut‐Cre mice) were generated and genotyped as described previously . Mice expressing td‐Tomato within corticotrophin‐releasing hormone (CRH) neurones (CRH‐Cre‐tdTomato) were generated and genotyped as described previously …”