Inhibition of oxytocin and vasopressin neuron activity in rat hypothalamic paraventricular nucleus by relaxin‐3–RXFP3 signalling

Abstract

The paraventricular nucleus of the hypothalamus (PVN) plays an essential role in the control of food intake and energy expenditure by integrating multiple neural and humoral inputs. Recent studies have demonstrated that intracerebroventricular and intra-PVN injections of the neuropeptide relaxin-3 or selective relaxin-3 receptor (RXFP3) agonists produce robust feeding in satiated rats, but the cellular and molecular mechanisms of action associated with these orexigenic effects have not been identified. In the present studies, using rat brain slices, we demonstrated that relaxin-3, acting through its cognate G-protein-coupled receptor, RXFP3, hyperpolarized a majority of putative magnocellular PVN neurons (88%, 22/25), including cells producing the anorexigenic neuropeptides, oxytocin and vasopressin. Importantly, the action of relaxin-3 persisted in the presence of tetrodotoxin and glutamate/GABA receptor antagonists, indicating its direct action on PVN neurons. Similar inhibitory effects on PVN oxytocin and vasopressin neurons were produced by the RXFP3 agonist, RXFP3-A2 (82%, 80/98 cells). In situ hybridization histochemistry revealed a strong colocalization of RXFP3 mRNA with oxytocin and vasopressin immunoreactivity in rat PVN neurons. A smaller percentage of putative parvocellular PVN neurons was sensitive to RXFP3-A2 (40%, 16/40 cells). These data, along with a demonstration of abundant peri-PVN and sparse intra-PVN relaxin-3-immunoreactive nerve fibres, originating from the nucleus incertus, the major source of relaxin-3 neurons, identify a strong inhibitory influence of relaxin-3-RXFP3 signalling on the electrical activity of PVN oxytocin and vasopressin neurons, consistent with the orexigenic effect of RXFP3 activation observed in vivo.

Relaxin-3/RXFP3 signalling in mouse hypothalamus: no effect of RXFP3 activation on corticosterone, despite reduced presynaptic excitatory input onto paraventricular CRH neurons in vitro

“…Subsequently, more recent electrophysiology studies in the rat have revealed a strong inhibition of magnocellular PVN neurons by RXFP3-A2 in acute PVN slices and expression of RXFP3 mRNA by both oxytocin and vasopressin immunoreactive neurons (Kania et al 2017). Although no equivalent studies in mice have been completed, our initial analysis of mRNA expression from the Allen Brain Institute atlas reveals similar expression patterns of RXFP3 and oxytocin and vasopressin mRNAs in the mouse, suggesting that these observations may be conserved between species.…”

“…This is also in line with in vitro studies that reveal RXFP3 inhibits cAMP signalling (Liu et al 2003), which is more consistent with an inhibition of target neurons. Therefore, if CRH neurons expressed RXFP3, activation of this receptor would likely result in inhibition of their activity, resulting in a reduction, rather than an elevation, in HPA axis activity (see Kania et al 2017).…”

“…Therefore, in the rat, any RXFP3-related reduction in HPA axis activity may be obscured by a larger RXFP1-mediated drive of HPA axis activity and further comparative studies are required to test this possibility. Notably, however, we recently observed that the selective RXFP1 agonist, relaxin, failed to alter the electrical activity of rat magnocellular PVN neurons, recorded in vitro (Kania et al 2017), suggesting that any RXFP1-mediated effects on HPA axis activity may be mediated via afferent inputs rather than direct effects within the PVN (Sunn et al 2001).…”

“…Furthermore, the identity of the RXFP3-expressing neurons within mouse PVN and their physiological and behavioural influences requires further investigation. In this regard, the gene expression patterns in mouse brain (Lein et al 2007), and recent anatomical/functional studies in rats, provide data that suggest likely potential target neurons/circuits for direct RXFP3 modulation are those expressing oxytocin and/or vasopressin (Yang et al 2012;Zhang et al 2012;Ganella et al 2013;Yan et al 2014;Calvez et al 2015;Hernandez et al 2015;Kania et al 2017). …”

“…However, although anatomical studies of the rat NI have revealed that dense projections from the NI and relaxin-3-positive fibres are present in areas such as the lateral hypothalamic area and the dorsomedial hypothalamus, they are sparse in the PVN proper (Goto et al 2001;Olucha-Bordonau et al 2003;Ma et al 2007;Kania et al 2017). Consistent with these findings, a low density of relaxin-3-like immunoreactive fibres was detected in the mouse hypothalamus using a rabbit polyclonal antisera (Smith et al 2010), although the nature of relaxin-3 projections to the mouse hypothalamus and PVN has not yet be reported using a more sensitive monoclonal antiserum (Kizawa et al 2003;Tanaka et al 2005).…”