Investigating the dose-response relationship of PYY on appetite and visceral illness in rodents. Eleanor Spreckley, BSc, Amin Alamshah, MSc, Katherine Banks, MRes, James Kinsey-Jones, PhD, Mohammad A. Ghatei, PhD, Stephen R. Bloom, FRS and Kevin G. Murphy, PhD Results Introduction Peptide YY (PYY) is a 36 amino acid peptide, synthesised by the enteroendocrine L-cells of the distal small intestine, that is released postprandially in proportion to the calorie content of a meal. PYY1-36 is post-translationally cleaved to its bioactive form, PYY3-36, which acts via central Y2 receptors to reduce food intake. Acute exogenous PYY3-36 administration physiologically inhibits food intake in obese humans (Batterham et. al. 2003). Supraphysiological doses of PYY cause nausea with no additional benefit on food intake reduction in humans, and produce a conditioned taste aversion (CTA) in rodents (le Roux 2008, Chelikani 2005, Reidelberger 2006). There may be a threshold dose at which exogenous PYY administration is able to reduce food intake. Central aversive circuits may be activated by a further threshold dose of PYY, with nausea representing one extreme of the satiety spectrum. We therefore investigated the doses of PYY3-36 required to suppress food intake and to produce conditioned taste aversion following intra-peritoneal administration in rats, and identified the brain regions activated by specific doses of PYY3-36. Fig 1. The dose-response effect of intra-peritoneal PYY3-36 on food intake in rats. Combination of results from four feeding studies expressed as % of control. Doses of 1.5, 2.5, 7.5, 25, 50 and 300 nmol/kg PYY significantly reduced food intake compared to saline control. *p<0.05, n=13-38. Fig 4. The effect of intra-peritoneal administration of specific doses of PYY3-36 on neuronal activation in rats. An overview of the areas studied for c-Fos positive cells, following IP injections of PYY at 3 doses. Doses of 25 and 300 nmol/kg PYY resulted in significantly increased c-Fos positivity in cells of the prelimbic cortex. Significantly increased c-Fos positivity was observed in cells of the shell of the nucleus accumbens following administration of 300 nmol/kg PYY3-36. *p<0.05, n=4-5. Discussion Peripheral administration of PYY significantly reduced food intake at doses of 1.5, 2.5, 7.5, 25, 50 and 300 nmol/kg. All doses studied resulted in supraphysiological circulating levels of PYY when compared to fed controls. A significant CTA was produced following administration of 7.5 nmol/kg PYY compared to a saline control, indicating that this dose results in feelings of visceral illness in rodents. Immunohistochemical staining for c-Fos, a marker of neuronal activation, revealed increased activity in nuclei of the mesocortical pathway, specifically the prelimbic cortex and shell of the nucleus accumbens. Higher doses of PYY may exert their effects on appetite by activating specific aversive circuits downstream of the mesocorticolimbic pathways. We plan to determine the effects of doses of PYY3-36 that result in conditioned taste aversion compared to those that don’t on cardiovascular function, energy expenditure and specific neuronal pathways. Identifying a recognizable pattern in these physiological responses to varying doses of PYY may identify a model that could replace the use of CTA protocols in the future, and may determine whether aversion represents the activation of pathways distinct to those that mediate satiation. Fig 2. The effects of intra-peritoneal administration of 2.5 and 7.5 nmol/kg PYY3-36 on conditioned taste aversion in rats. A significant CTA was induced by 7.5 nmol/kg PYY compared to saline control. *p<0.05, n= 15 (Saline, PYY), n= 5 (LiCl). Methods and Materials Feeding studies – Male Wistar rats were randomised into treatment groups and fasted 16 h prior to the study. Food intake was measured 1, 2, 4, 8 and 24 h post administration of PYY3-36 at various doses. CTA - Male Wistar rats were subjected to a one-bottle CTA protocol (adapted from Lachey 2005), in which they were trained to consume their daily fluid intake within a one hour period for 1 week. They were introduced to a novel flavour (grape KoolAid) during fluid access on three occasions, with the third immediately followed by an IP injection of 2.5 or 7.5 nmol/kg PYY3-36, saline or lithium chloride (LiCl). Plasma PYY levels were measured using a sensitive and specific in-house radioimmunoassay. Brains were dissected and analysed for c-Fos staining using immunohistochemistry. Fig 3. The effect of intra-peritoneal administration of specific doses of PYY3-36 on circulating PYY-immunoreactivity in rats. 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