Characterisation of the metabolic response to hypothalamic neuropeptides involved in energy balance

Kirsty Smith, Nina Semjonous, James Parkinson, David Gunner, Yong-Ling Liu, Kevin Murphy, Mohammad Ghatei, Stephen Bloom, Caroline Small

Imperial College London, London, UK

Obesity is a leading cause of morbidity and premature mortality in the UK and costs the NHS in excess of £2 billion per year. The hypothalamus is an important CNS region in the regulation of energy balance. Understanding the mechanisms involved in the regulation of energy balance is important for the identification of novel targets for anti-obesity therapies. The hypothalamic control of energy balance is regulated by a complex network of neuropeptide-releasing neurones. Whilst the effect of these neuropeptides on individual aspects of energy homeostasis has been studied, the coordinated response of these effects has not been comprehensively investigated. The Comprehensive Laboratory Animal Monitoring System (CLAMS) (Columbus Instruments) allows continuous and simultaneous measurement of a number of metabolic parameters including food intake, activity and energy expenditure

We have used the CLAMS to establish baseline metabolic profiles in normal chow fed and fasted male Wister rats (200-250g), n=8 per group. In order to characterise the metabolic response following intra-hypothalamic administration of hypothalamic neuropeptides with established roles in the regulation of feeding, activity and energy expenditure, male Wistar rats were cannulated into the third cerebral ventricle using methods previously described (Smith et al, Endocrinology 2006 147(7):3510-8). Following a 7 day recovery period, animals were injected intracerebroventricularly (ICV) with either a 1 or 3nmol dose of hypothalamic neuropeptide or saline for control animals. These doses were chosen because they have previously been shown to have a robust effect on food intake and allow direct comparison between the peptides. Ad libitum fed rats received a single ICV injection of an orexigenic neuropeptide, either neuropeptide Y (NPY), agouti-related protein (AgRP), melanin-concentrating hormone (MCH) or orexin-A. Overnight fasted rats received an ICV injection of the anorectic peptides α-MSH, corticotrophin releasing factor (CRF) or neuromedin U (NMU). Animals were then returned to the CLAMS and food intake, activity and energy expenditure measured for 24 hours post injection

Our results reveal the temporal sequence of the effects of these neuropeptides on both energy intake and expenditure, highlighting key differences in their function as mediators of energy balance. NPY significantly increased food intake for up to 17 hours post injection whilst the effects of AgRP on food intake were more prolonged, lasting in excess of 24 hours. Furthermore, NPY caused a significant increase in locomotor activity and a significant decrease in oxygen compared with saline injected controls.

Figure 1: Effect of single ICV injection of NPY (1 or 3nmol) on (a) food intake, (b) oxygen consumption and (c) activity in male Wistar rats. *p< 0.05 1nmol NPY; #p<0.05 3nmol NPY vs saline (n = 8 per group). Black bar at bottom indicates dark phase.

In contrast, orexin-A increased both feeding and oxygen consumption, consistent with an observed increase in activity. The potent anorexigenic effects of CRF were accompanied by a prolonged increase in activity whilst NMU injection resulted in significant but short-lasting effects on food intake, activity and oxygen consumption. Interestingly, α-MSH injection resulted in significant increases in activity and oxygen consumption with both doses whilst a reduction in food intake was only seen following administration of the 3nmol dose. This study furthers our understanding of these neuropeptides by generating a temporal and quantitative analysis of their effects on metabolic parameters. Furthermore, the CLAMS is a useful tool for characterising the metabolic response to pharmacological agents for the treatment of obesity