Metabolic sensing in the hypothalamus

• Main Author: Sjoberg, Hanna
• Published: University of Manchester 2014
• Subjects: 612.8
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.626923

The hypothalamus is an established regulatory hub with regards to energy homeostasis. While the arcuate nucleus has been researched extensively and substantial emphasis has been put on its role in energy balance, the ventromedial nucleus of the hypothalamus (VMN) is still poorly understood. However, the anorexia-inducing pituitary adenylate-cyclase activating peptide (PACAP) and brain-derived neurotrophic factor (BDNF) have both been proposed as potential candidates as VMN-produced regulators. The mRNAs of both neuropeptides is up regulated in diet-induced obesity (DIO)-resistant mice, but not in DIO-prone mice, indicating that they could be responsible for signalling in the VMN, especially in terms of countering the effects of an obesogenic diet. The initial objective of this PhD project was to evaluate the effect of metabolic manipulation, in the form of fasting or feeding with high-energy diet (HED), on the gene expression of these two possible neuronal markers in the VMN. This was done using quantitative PCR and, while our findings did not fully support the hypothesis that PACAP and BDNF have protective roles against obesity, we determined that metabolic manipulation differentially regulates pacap and bdnf transcripts in the VMN of outbred mice. The presence of several bdnf transcripts in the VMN, and their differential regulation, indicates that the transcripts play distinct roles in the response to metabolic manipulation. The findings also added further support for the role of PACAP and BDNF as important signalling molecules in the VMN. Their identification as important cellular phenotypes, allows future manipulation of specific neurons in the VMN, which should help us to rapidly increase our knowledge of the nucleus and its functions. We utilised a transgenic mouse model where leptin receptor deletion is driven by pacap expression in PACAP—IRES-Cre recombinase mice, to study whether leptin is having a physiological role through PACAP neurons. However, since we may not have achieved full expression of cre recombinase in all PACAP neurons, it was difficult to interpret our results, leaving open the question of what is the role of PACAP neurons in leptin signalling. Further, attempts have been made here to characterise this subtype of VMN neuron by using two GFP-reporter lines: Adcyap1-eGFP and PACAP-i-cre X Z/EG transgenic mouse lines. These were used to study the responsiveness of PACAP neurons to fluctuations in glucose availability, and our very promising early results indicate that PACAP VMN neurons are glucose inhibited. We studied the anorectic effects of PACAP and the peptide’s downstream targets, including the corticotrophin-releasing hormone (CRH), melanocortin and oxytocin pathways. The anorexic effect of central PACAP were maintained in VPAC1 and VPAC2 receptor (which also binds the related VIP) knockout animals, implicating the PAC1 receptor as the mediator of the hyperphagic response. We also showed that the feeding effects of PACAP are mediated by CRH, rather than by melanocortin or oxytocin pathways. Finally, we utilised a systems-genetics approach in the BXD set of recombinant inbred mouse strains, in an attempt to tease apart underlying networks of genes contributing to metabolic phenotypes. Quantitative trait analysis identified several loci, which contained genes of potential interest (for example: a QTL related to adiposity contained the orphan G-protein-coupled receptor, GPRC5C). Joint mapping revealed that the phenotypic traits we measured correlated with each other and with trait data from BXD mice recorded in other laboratories. For example, this supported the role of PACAP and BDNF in body-weight management. The candidate gene, gprc5c, was further investigated. Its mRNA was found in the hypothalamus, but was also widely distributed in peripheral tissues. Gprc5c mRNA was regulated by HED in normal C57 mice in the hypothalamus, liver and brown adipose tissue, indicating a possible role for this orphan receptor in metabolism. In fact, similar to PACAP and BDNF, in BXD mice, expression of gprc5c in the hypothalamus was inversely correlated with body-weight gain.