NPY neuron developmental mechanisms are disrupted by maternal obesity

Obesity in the Western world is on the rise. Both epidemiological studies and experimental studies in model organisms have demonstrated that offspring born to obese mothers are at a greater risk of developing obesity than those born to normal weight mothers. While obesity is usually thought of as a disease of the periphery, body weight is ultimately regulated by the brain, and therefore dysregulation of the neural circuitry controlling bodyweight represents one way in which an individual might become predisposed to obesity. The hypothalamus is a key area of the brain involved in the synthesis of and response to neural, endocrine, and nutritional signals concerning body weight. Interestingly, the ability of body weight regulating Neuropeptide Y (NPY) neurons in the arcuate nucleus of the hypothalamus to innervate their targets appropriately is disrupted in the offspring of obese mothers. The mechanisms behind this are unknown. However, it has recently emerged that maternal obesity is associated with increases in certain inflammatory cytokines, including interleukin-6 (IL-6), in both the maternal and fetal circulation. This led to the hypothesis that activation of the maternal immune system during pregnancy affects the expression of important developmental molecules in the fetal arcuate nucleus. This in turn impairs the way arcuate neurons develop axons and establish connections, thus disturbing their ability to communicate with other important weight regulatory regions of the brain, and ultimately leading to obesity predisposition. To in investigate this hypothesis, firstly, the normal development of NPY neuron projections to their target regions in the prenatal mouse brain was determined. I found that NPY neuron axons were already leaving the arcuate nucleus at gestational day (GD) 15, and first began to innervate one of their target regions, the paraventricular nucleus (PVN), at GD17. The initial formation of this circuitry in the prenatal period indicates that these projections may be acting as pioneers, forming a scaffold which later follower axons can elaborate upon in the postnatal period. The role of the axon growth modulator Netrin-1 in the development of these prenatal projections was then investigated. Using in situ hybridisation, I have shown that in late gestation Netrin-1 was expressed in a spatial and temporal pattern consistent with an involvement in the growth of NPY neuron axons to their targets. Additionally using a primary neuron culture model it was found NPY neurons respond to Netrin-1 treatment by increased elaboration of their growth cones when compared to controls. This data represents the first time a growth factor critical in the prenatal development of NPY circuitry has been identified and studied. Secondly, both in vitro and in vivo methods were used to investigate the effects of maternal obesity on NPY neuron development. Acute exposure of fetal mouse arcuate nucleus explants to IL-6 in vitro induced gene expression changes in the Netrin-1 receptor Dcc which were similar to those found in vivo using a mouse model of maternal obesity. In addition, IL-6 treatment of fetal NPY neurons in primary culture appeared to block the normal growth cone response to Netrin-1. This suggests that during maternal obesity the developing fetus is exposed to aberrant levels of IL-6, which may act extrinsically to disrupt Netrin-1 signalling, and therefore perturb the correct formation of prenatal NPY circuitry. Lastly, the ability of an environment of maternal obesity to alter the intrinsic capacity of NPY neurons to develop normally was assessed. Fetal NPY neurons which had been exposed in vivo to maternal obesity were more immature in culture then those from control pregnancies, and had a blunted growth cone response to Netrin-1. These data suggest that