The Effects Of Rewarding System On High-fat Diet-induced Obese C57BL/6 Mice Following Different Dietary Intervention

Obesity, a serious metabolic disorder, is defined medically as a state of increased body weight, more specifically adipose tissue. Obesity has reached epidemic proportions worldwide with 1 billion adults estimated to be overweight and 300 million of them obese (WHO,1999). The lack of effective treatments for weight reduction and the extremely high rate of relapse in obesity highlight the importance of understanding the development and reversal of this condition. Only some human obesities can be explained by genetic defects. Most obesity is thought to be caused by the interaction of genetic susceptibility and environment. Furthermore, the contributions of genetic and environmental factors are not simply additive; rather, they interact in complex and sometimes counterintuitive ways. Nutrition is a key environmental factor involved in the development of obesity. Diets high in saturated fats are obesogenic in humans and rodents. This fact is important to consider as the obesity rate climbs steadily higher in recent 30 years.Fat is also more palatable than carbohydrate and protein, due to its caloric density and texture, and cause over energy consumption. The food, like drugs, can activate brain circuitry involved reward system. Furthermore, obesity and drugs addiction shared the same neurochemical pathway. Obesity, like drugs addiction, is strongly linked with exposure to reinforecers. The palatable foods produce the more hedonic sense, which may be powerful stimulator in rewarding system in brain. It is possible that the rewarding system play the important role in the high-fat diet induced obesity. The neurotransmitters that are known to be involved in the regulation of mesolimbic (rewarding) circuit of the brain are dopamine and endogenous opioid peptide. The drug addiction activates the dopamine releasing neurons that project from the ventral tegmental area (VTA) to the nucleus accumbens (Acb). The dopaminergic neuron in the VTA is inhibited by GABA neurons,μ- opioid receptors locate on GABA interneurons. The agonist binds toμ- opioid receptors, which would inhibit activation of GABA and reduce GABA release. This disinherits the dopamine neurons and induces dopamine release, which take part in the rewarding effect. It is the most important that dopamine and the endogenous opioid peptide participate in the food intake and energy balance regulation. The dopamine is essential to feeding as the dopamine-deficient mice die of starvation. The endogenous opioid peptide has been found to be involved in the modulation of feeding behavior in both human and rats; the agonist of opioid receptor increase food intake. The adipocyte hormone leptin and pancreatic hormone insulin are important regulators of food intake and energy balance. Plasma leptin and insulin levels are positively correlated with the body weight and adipose mass. Leptin and insulin enter into brain from the plasma and bind to their receptors. In recent years, new evidence is forthcoming in support of a role for insulin and leptin in the modulation of reward and motivated behaviors.This project aims to investigate the involvement of rewarding system in the development of obesisty. In order to explore the interplay of gentic and environmental factor on the development of obesity, we firstly establish the C57BL/6 mice model of diet-induced obese (DIO) and diet-resistant (DR), and then vary food density and restriction of energy intake. Base on dietary interventional model, I have investigated the influence of macronutrient on the dopamine and opioid receptor, and the relationship between them and plasma leptin and insulin in DIO and DR mice. The main results are summarized as follows:In the first study, Forty-five C57BL/6 mice were fed with a high-fat diet for 8 weeks. Fifteen mice with highest body gain were classified as diet-induced obese (DIO) and while lowest fifteen mice were classified as obese-resistant (DR) mice. The DIO and DR mice were then placed on diet with different levels of fat including a high-fat diet (-H), a low-fat diet (-L) and an energy-restricted high-fat diet (-P) for six weeks. The control group was fed a low-fat diet (LF). During the experiment, we examine the weekly body weight and energy intake, white fat mass, plasma leptin and insulin levels. The results reveal the body weight, white fat mass, ratio of visceral fat to body weight (RVB), energy intake and plasma leptin levels are higher in DIO-H than DR-H mice, and DIO-L than DR-L mice. The energy efficiency, plasma insulin levels and HOMAIR increase in DIO-H than DR-H mice; and leptin sensitive decrease in DIO-L than DR-L mice. After switching to low-fat (-L) or energy-restricted diet (-P), the plasma leptin levels and energy efficiency of both DIO and DR mice decrease; and simultaneously the leptin sensitive increase. Moreover the body weight and white fat mass of DIO mice also decrease. By the foundation of diet-interventional model, we conclude that the genetic predisposition of DIO mice is prone to obesity. In both DIO and DR mice, the high-fat diet raises the energy efficiency and plasma leptin levels and reduces the leptin sensitivity. When changing to low-fat diet or energy restriction, the energy efficiency, plasma leptin levels and leptin sensitive can be reinstated.In the second study, Dopamine regulates energy balance primarily by modulating food intake via the mesolimbic (rewarding) and mesohypothalamic (satiety) circuits of the brain. Tyrosine hydroxylase (TH) is the rate-limiting enzyme in the synthesis of dopamine, as a marker of dopamine. Using the quantitative in situ hybridization techniques, this study examined TH mRNA expression in the brain of diet-interventional model established in the first study. The results revealed that TH mRNA expression was significantly decreased in the ventral tegmental area (VTA), ventromedial hypothalamic nucleus (VMH), and substantia nigra (SN) of the high-fat diet-induced obese (-29%, -26% and -26%) and obese-resistant mice (-21%, -24% and -18%) compared to controls. After switching the diet from high to low fat diet, there were the higher TH mRNA expressions in the VMH and VTA of DR-L (23% and 28%) than DR-H mice and the SN of DIO-L (24%) than DIO-H mice. Energy restriction, even with high-fat feeding, reduced TH mRNA expression in the VMH, SN and VTA compared to LF mice. In addition, TH mRNA expression in VMH, SN and VTA showed a significant negative correlation with plasma leptin levels. But TH mRNA is irrespective with the phenotype of DIO and DR. This study suggested that high-fat diet can reduce the levels of TH mRNA expression in VTA, VMH and SN. Moreover, the altered levels of leptin according to the nutritional condition facilitated the variation of TH mRNA expression. Take together, my study indicated that the up-or down-regulation of TH mRNA expression in VMH, SN and VTA is mainly due to the intake of macronutrient type rather than body weight, energy intake and obesity phenotype.In the third study, opioid is a key regulator forming an interconnected brain network and participates in the food intake regulation, especially in the rewarding effect of food. Mu (μ) opioid agonists preferentially increase the intake of diet in high saturated fat. Theμ-receptor binding density are measured in brain slices of the diet-interventional model established in the first study, using [H]-DAMGO (D-Ala~2, N-Me-Phe~4, Gly-ol~5) and quantitative autoradiography. The results revealed thatμ-opioid receptor binding density was significantly decreased in the accumbens nucleus (Acb), and amygdaloid nucleus (AMG) of the high-fat diet-induced obese (-32% and -38%) and obese-resistant mice (-28% and -33%) compared to controls (LF). After switching from the high-fat to low fat diet, DIO-L and DR-L have the higherμ-opioid receptor binding density in the Acb (49% and 68%) and AMG (44% and 56%) compare to the respective DIO-H and DR-H mice. Energy restriction of amount of high-fat diet, cause the decreased trend ofμ-opioid receptor binding density in the Acb and AMG compare to the LF. Butμ-opioid receptor binding is irrespective with the phenotype of DIO and DR. In the Acb and AMG,μ-opioid receptor binding is significantly negatively correlation with plasma leptin levels and positive correlation with the TH mRNA expression in VTA. In conclusion, this study show that high-fat diet can significantly lower theμ-opioid receptor binding in Acb and AMG related to rewarding circuit. When changing to low-fat diet,μ-opioid receptor in Acb and AMG can be up-regulated. This suggest that the alteredμ-opioid receptor binding in Acb and AMG is mainly due to the intake of macronutrient type, rather thanthe body weight, energy intake and obesity phenotype.