Early Life Origins of Obesity: Role of Hypothalamic Programming

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1 Journal of Pediatric Gastroenterology and Nutrition 48:S31 S38 # 2009 by European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition Invited Review Early Life Origins of Obesity: Role of Hypothalamic Programming ysebastien G. Bouret The Saban Research Institute, Neuroscience Program, Children s Hospital Los Angeles, University of Southern California, Los Angeles, CA, USA, and { Inserm, Jean-Pierre Aubert Research Center, U837, University Lille 2, Lille, France ABSTRACT The incidence of obesity is increasing at an alarming rate and this worldwide epidemic represents an ominous predictor of increases in diseases such as type 2 diabetes and metabolic syndrome. Epidemiological and animals studies suggest that maternal obesity and alterations in postnatal nutrition are associated with increased risks for obesity, hypertension, and type 2 diabetes in the offspring. Furthermore, there is also growing appreciation that developmental programming of neuroendocrine systems by the perinatal environment represents a possible cause for these diseases. This review article provides a synthesis of recent evidence concerning the actions of perinatal hormones and nutrition in programming the development and organization of hypothalamic circuits that regulate body weight and energy balance. Particular attention is given to the neurodevelopmental actions of insulin and leptin. JPGN 48:S31 S38, Key Words: Development Hormones Hypothalamus Leptin Programming. # 2009 by European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition A principal goal of brain development is to produce the necessary neural architecture for integrating information from the external environment with internal cues that reflect important aspects of an animal s physiological state. This integration allows the elaboration of adaptive behavioral and physiological responses that are essential for survival. However, disorders can arise when an individual is confronted with environmental conditions that differ markedly from those present during perinatal development. For example, epidemiological evidence has indicated that alterations in perinatal nutrition could predispose an individual toward obesity and other associated diseases such as type 2 diabetes, particularly in an environment with high availability of energy dense foods. Paradoxically, both maternal obesity and maternal energy deprivation during pregnancy may increase the incidence of obesity and type 2 diabetes in the offspring (1 4). Mothers who are obese or have type 2 diabetes during pregnancy also have an increased incidence of obese progeny. Similarly, maternal malnutrition during Address correspondence and reprint requests to Dr Sebastien G. Bouret, PhD, The Saban Research Institute, Neuroscience Program, Childrens Hospital Los Angeles, University of Southern California, 4650 Sunset Boulevard, MS#135, Los Angeles, CA 90027, USA ( sbouret@chla.usc.edu). The author reports no conflicts of interest. gestation produces offspring obesity and diabetes (1,2). Data from a variety of animal models have supported a link between the perinatal nutritional environment and the programming of energy balance set points. Interestingly, both energy restriction and overfeeding could cause lasting perturbations in energy balance (1,2,5). In this review, we will attempt to examine these observations within the neurobiological aspects of hypothalamic programming. NEURAL CONTROL OF ENERGY BALANCE Signals Communicating Nutrient Availability in the Environment to the Developing Brain Peripheral hormones represent important signals that regulate adiposity as well as central nervous system (CNS) circuits that control food intake. The bestcharacterized hormonal signals of adiposity are insulin and leptin (Fig. 1) (6). Insulin is secreted by the Islets of Langerhans in the pancreas to promote energy storage, and increased circulating insulin is observed in response to nutrient repletion and in states of obesity. Insulin receptors are expressed in the CNS (7) and injections of small amounts of insulin into the brain of insulin-deficient animals can eliminate hyperphagia (8). Deletion of the insulin receptor from the CNS S31

2 S32 BOURET FIG. 1. Organization of hypothalamic circuits regulating energy balance. These simplified schematic diagrams illustrate the possible routes and the neuronal populations relaying hormonal and nutrient signals from the periphery to the brain. Two distinct populations of neurons in the arcuate nucleus (ARH) one coexpressing neuropeptide Y (NPY) and agouti-related protein (AgRP) and the other containing proopiomelanocortin (POMC)-derived peptides and cocaine- and amphetamine-regulated transcript (CART) represent major routes for the regulation of body weight by peripheral hormones such as leptin. These neurons send direct projections to discrete populations of neurons located in the dorsomedial nuclei of the hypothalamus (DMH) and paraventicular nuclei of the hypothalamus (PVH) and in the lateral hypothalamic area (LHA). Each of these regions plays a major role in the control of ingestive behavior. Projections originating from the DMH and the ventromedial nucleus of the hypothalamus (VMH) and projecting to the PVH also represent important routes for the action of leptin at the hypothalamic level. me, median eminence; V3, third ventricle. The schematics have been modified with permission (6). resulted in obesity and insulin resistance (9), thus further adding support to the importance of insulin action in appetite regulation by the brain. Insulin levels are elevated and known to mediate compensatory responses (such as macrosomia) in the offspring of diabetic mothers (10). Leptin, a hormone secreted by fat cells, is a crucial signal of body energy stores and acts to downregulate feeding behavior and promote energy expenditure through a variety of neural and endocrine mechanisms. These include regulation of the autonomic nervous system and the synthesis of thyroid hormones (11). Thus, mice lacking leptin (Lep ob /Lep ob mice) are obese, diabetic, cold intolerant, and hypoactive (12,13). Similarly, mutations that affect the long form of the leptin receptor (LepRb) or its downstream signaling pathways result in a diabetic phenotype and infertility (14 17). Importantly, this receptor is highly expressed in regions of the CNS involved in energy balance, particularly the hypothalamus (18), and leptin acts directly on the CNS to mediate most of its action (19 21). Thus, leptin and insulin are particularly well suited to communicate nutrient availability in the environment to the hypothalamus during development.

3 EARLY LIFE ORIGINS OF OBESITY S33 Hypothalamic Circuits Controlling Energy Balance The hypothalamus is well known to regulate feeding and energy balance. Neurons in the arcuate nucleus of the hypothalamus (ARH) play an important role in this regulation. The ARH resides above the median eminence and shares connections with circumventricular organs, making it appropriate to receive and integrate signals from peripheral hormones such as leptin and insulin (Fig. 1). The ARH has long been associated with obesity (22), and it contains numerous leptin-sensitive neurons (18,23 26). Moreover, recent genetic studies have specifically demonstrated the importance of leptin receptor signaling in ARH neurons. Restoring ARH neurons leptin receptor signaling in leptin-deficient mice ameliorated body weight gain by reducing food intake and decreasing adipose tissue mass (21,27). These data indicate that the ARH is an important site of action for the central regulatory effects of leptin on energy balance. The ARH contains 2 populations of neurons that play particularly important roles in distributing leptin signals centrally. One subpopulation of ARH neurons coexpresses neuropeptide Y (NPY) and agouti-related peptide (AgRP) and acts as a major orexigenic signal (ie, promotes feeding). A separate subpopulation of ARH neurons expresses proopiomelanocortin (POMC)-derived peptides, such as alpha-melanocyte-stimulating hormone, and represents an important anorectic regulator (ie, inhibits feeding). These anatomically distinct populations of ARH neurons provide overlapping projections to other key parts of the hypothalamus that are implicated in the control of feeding. These hypothalamic parts include the paraventricular nuclei of the hypothalamus (PVH) and dorsomedial nuclei of the hypothalamus (DMH), as well as the lateral hypothalamic area (LHA). In addition to playing an important role in regulating food intake and body weight, a number of studies have suggested that the hypothalamus is a key component of peripheral glucose homeostasis. Infusion of insulin into the medio-basal hypothalamus (a region comprising the ARH and ventromedial nucleus of the hypothalamus, VMH) reduced hepatic gluconeogenesis by increasing hepatic insulin sensitivity (28). Furthermore, downregulation of insulin receptor signaling in the medio-basal hypothalamus induced insulin resistance in rats (28 30). Altogether these data show that hypothalamic insulin signaling is important for the regulation of glucose homeostasis. Recent data have also indicated that most of the effects of leptin on glucose homeostasis are mediated by its effects on the hypothalamus. Restoration of functional leptin receptors exclusively in the ARH of leptin-receptor-deficient animals significantly improved glucose homeostasis and insulin sensitivity (27,31). Thus, the hypothalamus appears to play a major role in the regulation of energy balance and glucose homeostasis and its core circuitry appears to mediate many of the metabolic effects of leptin and insulin (Fig. 1). DEVELOPMENT OF HYPOTHALAMIC FEEDING CIRCUITS Rodent Studies Hypothalamic development is initiated by cell proliferation in the neuroepithelium of the third ventricle. This involves generation of neuronal progenitors that ultimately produce postmitotic neurons (Fig. 2). These postmitotic neurons migrate to their appropriate location in various parts of the hypothalamus. Neuronal birthdating studies in rats revealed that ARH neurons are generated between embryonic day (E) 12 and E17, VMH neurons between E13 and E16, DMH neurons between E12 and E16, PVH neurons between E13 and E15, and LHA neurons between E12 and E14 [see (32)]. Although hypothalamic neuronal proliferation occurs primarily during mid-gestation, the development of neural projections from these neurons to their downstream target sites is initiated primarily postnatally (Fig. 2). Axonal tract tracing experiments performed in mice revealed that ARH projections reached their target nuclei within distinct temporal domains; innervation of the DMH occurred first on postnatal day (P) 6, followed by innervation of the PVH on P8-P10 (33). Projections to the LHA were established later in P12 (33). Immunohistochemical studies in rats showed that development of axonal projections from ARH NPY/AgRP neurons follow similar temporal domains (34). It is also interesting to note that a significant proportion of NPY could be produced by neurons in the DHM and LHA during the first weeks of postnatal life, in addition to what is already produced by ARH neurons (35). However, the precise role and function of these transient populations of NPY neurons remain unclear. Nevertheless, we should note that NPY is permanently induced in the DMH of obese FIG. 2. Critical periods of hypothalamic development in rodents. Schematic drawings summarizing major neurobiological events governing hypothalamic development. The growth of the hypothalamus after organogenesis can be described as occurring in 2 major phases: a phase of neuronal proliferation (neurogenesis) that takes place between embryonic days 12 and 16, followed by a phase of axonal extension that occurs between postnatal days 6 and 12.

4 S34 BOURET animals (specifically in the melanocortin 4 [MC4] receptor knockout and the lethal yellow [A(y)] mice), raising the possibility that this ectopic population of NPY neurons may play an important role in the development of obesity (36). Together, these data reveal the existence of 2 major critical periods (ie, mid-gestation and early postnatal life) during which alterations in the intrauterine environment may affect hypothalamic neurogenesis and/ or axonal outgrowth and, therefore, will have long-term consequences on nutrition and metabolism. Human and Non-human Primate Studies Limited information is available on how the hypothalamus develops in humans. Much of what we know about the development of hypothalamic neural pathways in primates has been inferred from studies in nonhuman primates. Hypothalamic neurogenesis in these animals occurs in the first quarter of gestation (37,38). Limited reports on human fetal chemoarchitecture and cytoarchitecture have suggested that early hypothalamic neurogenesis is limited to the 9th and 10th weeks of gestation (39 43). Although many of the hypothalamic feeding circuits develop during the first 2 weeks of life in rodents, these circuits appear to develop in utero in primates, including humans. In Japanese macaques, NPY/AgRP fibers innervate the PVH as early as gestational day 100 (ie, late second trimester of gestation) and a mature pattern of projections is apparent at gestational day 170 (44). Similarly, in human fetuses, NPY immunoreactive fibers are detected in the ARH and in the PVH as early as at 21 weeks of gestation (45). Thus, development of neural projection in humans occurs significantly later than neurogenesis. Whether the same developmental factors influence both neuronal proliferation and axonal extension is unknown. However, it is notable that these 2 developmental events occur during distinct temporal periods. Developmental Regulation of Leptin s Action on Metabolism In addition to its effects on appetite regulation in adults, leptin also regulates appetite-related neuropeptides in the hypothalamus during early development. Administration of leptin to P10 rats increased suppressor of cytokine signaling 3 (SOCS-3) and POMC mrna levels, but decreased NPY mrna levels in the ARH (46). Moreover, chronic neonatal leptin administration downregulated all subtypes of leptin receptor mrna and increased corticotropin-releasing factor receptor-2 mrna levels in the VMH (46). Furthermore, leptin induced cfos expression in ARH neurons (specifically in POMC neurons) as early as P6 (33). However, these transcriptional changes were not matched by a corresponding reduction in food intake in neonatal mice, because administration of leptin in lean or Lep ob /Lep ob mice did not affect milk/food intake, oxygen consumption, body weight, or adiposity until after weaning (46,47). Therefore, despite its regulatory action on hypothalamic neuropeptide expression, leptin does not appear to regulate food intake during early development. This decreased anorectic action of leptin before weaning may help the animals maximize food intake to support growth and to maintain high thermoregulatory metabolic rates to optimize survival until weaning. PERINATAL FACTORS INFLUENCING DEVELOPMENT OF HYPOTHALAMIC FEEDING PATHWAYS As noted above, a plethora of data from rodent to human studies have suggested that nutritional status during early development affects the later metabolic fate of the organism. Insulin and leptin thus likely represent the hormonal mediators for these environmental nutrient sensing systems that control this program (Fig. 3). Leptin It is now clear that in addition to playing an important role in the regulation of energy balance and neuroendocrine functions in mature animals, leptin also acts early in life as a developmental signal that promotes the formation of metabolic pathways. Elevated leptin levels are found particularly during the first 2 weeks of life in rodents (48 50) at a time when leptin is largely ineffective in altering body weight or food intake. Limited information is available on the origin of the neonatal leptin surge. It is probable that neonatal leptin is produced, at least in part, by fetal adipose tissue, as revealed by the elevated neonatal leptin mrna expression in white and brown adipose tissues that mirrors the circulating hormone concentrations (50). Alternatively, perinatal leptin may also be produced by other organs such as the stomach (51). Moreover, several studies have shown the important contribution of maternal milk to serum leptin levels in newborns (52,53). Interestingly, the neonatal leptin surge (48) appeared to coincide with the development of major hypothalamic feeding circuits (54). Neuroanatomical experiments further revealed that instead of regulating food intake and body weight, neonatal leptin is an important trophic factor for the development of hypothalamic circuits that control energy homeostasis. Injections of anterograde axonal tracers into the ARH of leptin-deficient mice demonstrated that leptin deficiency induced profound disruption in the formation of ARH circuits (55). The density of axons from arcuate nucleus neurons that innervate other hypothalamic sites involved in the control of energy homeostasis (such as the PVH, DMH, and LHA) is severely reduced in Lep ob /Lep ob neonates and remains diminished throughout life (55).

5 EARLY LIFE ORIGINS OF OBESITY S35 FIG. 3. Developmental programming of hypothalamic metabolic pathways. The developmental programming of hypothalamic neuroendocrine systems by the perinatal environment represents a possible mechanism by which maternal obesity, gestational diabetes, and alterations in perinatal nutrition predispose the offspring to develop metabolic syndrome and altered energy balance. The hormones insulin and leptin are well positioned to signal alterations in the nutritional environment to the central nervous system and alter the development and activity of these neural circuits, which may result in abnormal regulation of metabolism later in life. Similar disruptions were observed in other animal models of leptin receptor deficiency, such as in Zucker rats (54). Both orexigenic (NPY/AgRP) and anorexigenic (POMC) projections appeared to be affected by leptin deficiency (55), suggesting a widespread developmental effect of leptin on arcuate neurons involved in the regulation of metabolism. In vitro experiments also revealed that leptin could act directly on ARH neurons to induce axonal outgrowth (55). Furthermore, leptin appeared to exert its effects on axonal formation primarily during a restricted postnatal period, because daily injections of P4 to P12 Lep ob /Lep ob mice with leptin rescued a normal pattern of innervation by arcuate neurons of the PVH. On the contrary, injections of the hormone in mature animals remained largely ineffective in restoring a normal pattern of ARH projections (55). These data indicate that there is a critical period for the neurodevelopmental actions of leptin that seems to be restricted to the first few weeks of life. The existence of a critical period for the developmental effects of leptin suggests that changes in leptin levels during key periods of hypothalamic development may induce long-lasting and potentially irreversible effects on metabolism in adults. Leptin levels are directly regulated by nutritional factors, thus this hormone is well positioned to participate in developmental responses to nutritional changes. In support of this hypothesis, recent data have indicated that an ill-timed neonatal leptin surge may cause lasting effects on metabolism. Using a mouse model of intrauterine energy restriction, Yura et al (56) found that prenatal underfeeding resulted in an earlier leptin surge that was accompanied by deleterious effects on body weight regulation and glucose homeostasis. The premature leptin surge observed in the offspring of undernourished dams was also associated with a reduced anorectic effect of leptin and an altered hypothalamic response to leptin, as evidenced by a decreased leptininduced cfos immunoreactivity in the PVH (56). Similarly, the blunted postnatal leptin surge, as induced by the administration of a specific leptin antagonist from P2 to P13, was associated with long-term leptin insensitivity and increased susceptibility to diet-induced obesity (DIO) in rats (57). Taken together, these studies have suggested that the postnatal leptin surge is an important trophic factor for the development of hypothalamic feeding circuits and is critical for normal energy balance and hypothalamic regulation later in life. Insulin In addition to leptin, insulin also appears to exert important influences on the development of hypothalamic circuits that regulate energy homeostasis. Maternal injections of insulin between gestational day 15 and 20, a critical period for hypothalamic development, induced obesity in the offspring (58). The metabolic abnormalities observed in the offspring of insulin-injected dams

6 S36 BOURET were also accompanied by increased hypothalamic norepinephrine levels (58) and increased density of norepinephrine-containing fibers innervating the PVH (59). Similarly, maternal diabetes induced by streptozotocin injections resulted in hyperinsulinism associated with hypothalamic alterations such as decreased brain NPY mrna and protein expression (60), as well as altered neuronal morphology in the arcuate nucleus in the fetus (10). Furthermore, postnatal injections of insulin have been associated with morphological changes in the VMH (61). Together, these data suggest that changes in insulin levels (specifically hyperinsulinism) during pregnancy could induce alterations in hypothalamic organization that may affect metabolism of the offspring later in life. Polygenic Obesity It is increasingly accepted that obesity results from a combination of genetic and environmental factors. Dietinduced obesity in rats is a useful model to study the pathogenesis of human obesity because DIO rats, like humans, have a polygenic mode of inheritance. Moreover, these rats develop metabolic syndrome when a moderate amount of fat is added to the diet (62,63). One of the particular traits of DIO rats is that they exhibit leptin resistance characterized by elevated serum leptin and a decreased anorectic and thermogenic response to exogenous leptin (63 65). The body of evidence suggests that leptin resistance observed in DIO rats is mediated by central leptin insensitivity. For example, DIO rats have decreased expression of LepRb associated with attenuated leptin receptor signaling in the hypothalamus, particularly in the ARH (63,65,66). Interestingly, this reduction in hypothalamic leptin sensitivity occurred before the animals became obese and was established during early postnatal life (67). The diminished responsiveness of hypothalamic neurons to leptin appeared to impact the development of hypothalamic circuits. Thus, the density of axons emanating from the ARH and innervating the PVH appeared severely reduced in the progeny of DIO mothers compared with the offspring of diet-resistant (DR) dams (67). Moreover, ARH neurons derived from DIO rats were significantly less responsive to the neurotrophic action of leptin than ARH neurons in explants derived from DR rats (67). Thus, polygenic obesity appears to induce the abnormal organization of neural pathways involved in energy homeostasis; this may be the result of the diminished responsiveness of ARH neurons to the trophic actions of leptin during critical periods of postnatal development. Perinatal Nutrition It has been known for decades that changes in perinatal nutrition have long-term effects on metabolism (1,4,68). Previous studies have suggested that neonatal nutrition may also play an important role in the programming of hypothalamic feeding systems (Fig. 3). Using an animal model of divergent litter size, Plagemann et al (2) demonstrated that animals that were raised in small litters (3 pups per litter) showed increased body weight and adiposity during adult life. These metabolic abnormalities were associated with altered responsiveness of ARH and VMH neurons to insulin and leptin. Although leptin is a major stimulatory signal on VMH neurons in normal animals, it mainly inhibits VMH neurons in rats that are raised in small litters (69). Similarly, postnatal overnutrition induced a reduction in the inhibitory effect of leptin on ARH neurons (70). Moreover, ARH neurons of rats exposed to early postnatal overfeeding were less inhibited by insulin when compared with controls (71). Changes in postnatal nutrition also modified the response of VMH neurons to orexigenic peptides (NPY and AgRP) and of PVH neurons to anorexigenic neuropeptides (amsh) and cocaine- and amphetamine-regulated transcript (CART) (72 74). Taken together, these data indicate that alteration in nutrition during critical periods of postnatal development may induce permanent changes in the responsiveness of hypothalamic neurons to hormonal and peptidergic cues. Nutrition during prenatal life also appears to influence the programming of hypothalamic appetite networks. Maternal protein restriction induced hypoinsulinemia in the offspring and was associated with increased NPY levels in the PVH and LHA at weaning (75). Importantly, exposure to high doses of NPY during early postnatal development was linked to permanent changes in food intake in adults (76). Similarly, increased maternal nutrition in late pregnancy resulted in persistent changes in the hypothalamic expression of appetite-regulating genes in sheep (77). Offspring of dams fed with 40% excess nutrient intake in late pregnancy had a permanent increase in hypothalamic POMC mrna expression when compared with that of control animals (77). In addition to altering gene expression, maternal overnutrition also affected central leptin sensitivity, as demonstrated by the attenuated levels of leptin-induced phosphorylation of the signal transducer and activator of transcription 3 (pstat3, a key intracellular signaling pathway of LepRb) in offspring born from dams fed with a high-fat diet (78). CONCLUSIONS Previous studies during the past decade have indicated that normal metabolic regulation during adulthood not only requires a good matching of energy intake with energy expenditure but also is influenced by optimal fetal and postnatal environments. Although the mechanisms underlying this metabolic imprinting require further elucidation, the evidence accumulated to date indicates that perinatal hormones (particularly insulin and leptin)

7 EARLY LIFE ORIGINS OF OBESITY S37 represent key signals that program CNS (hypothalamic) development and function and exert lasting effects on body weight regulation and glucose homeostasis. A better understanding of how these metabolic hormones exert their neurotrophic effects may open new avenues for understanding pre- and perinatally acquired predisposition to obesity and diabetes. Furthermore, a more detailed determination of whether hypothalamic misprogramming can be reversed, and the definition of the precise limits of the critical period for plasticity may provide new preventive and/or therapeutic opportunities. REFERENCES 1. Levin B. Metabolic imprinting: critical impact of the perinatal environment on the regulation of energy homeostasis. Phil Trans R Soc Lond B 2006;361: Plagemann A. Perinatal nutrition and hormone-dependent programming of food intake. Horm Res 2006;65: Martin-Gronert MS, Ozanne SE. Programming of appetite and type 2 diabetes. Early Hum Dev 2005;81: Taylor PD, Poston L. Developmental programming of obesity in mammals. Exp Physiol 2007;92: Bouret SG, Simerly RB. Developmental programming of hypothalamic feeding circuits. Clin Genet 2006;70: Swanson LW. Brain Maps: Structure of the Rat Brain. 2nd revised ed. Amsterdam: Elsevier; Adamo MRM, LeRoith D. Insulin and insulin-like growth factor receptors in the nervous system. Mol Neurobiol 1989;3: Sipols AJ, Baskin DG, Schwartz MW. Effect of intracerebroventricular insulin infusion on diabetic hyperphagia and hypothalamic neuropeptide gene expression. Diabetes 1995;44: Bruning JC, Gautam D, Burks DJ, et al. Role of brain insulin receptor in control of body weight and reproduction. Science 2000;289: Plagemann A, Harder T, Janert U, et al. Malformations of hypothalamic nuclei in hyperinsulinemic offspring of rats with gestational diabetes. Dev Neurosci 1999;21: Ahima RS, Flier JS. Leptin. Annu Rev Physiol 2000;62: Zhang Y, Proenca R, Maffei M, et al. Positional cloning of the mouse obese gene and its human homologue. Nature 1994; 372: Tartaglia LA, Dembski M, Weng X, et al. Identification and expression cloning of a leptin receptor, OB-R. Cell 1995; 83: Chen H, Charlat O, Tartaglia LA, et al. Evidence that the diabetes gene encodes the leptin receptor: identification of a mutation in the leptin receptor gene in db/db mice. Cell 1996;84: Lee G-H, Proenca R, Montez JM, et al. Abnormal splicing of the leptin receptor in diabetic mice. Nature 1996;379: Chua SC Jr, Chung WK, Wu-Peng XS, et al. Phenotypes of mouse diabetes and rat fatty due to mutations in the OB (leptin) receptor. Science 1996;271: Bates SH, Stearns WH, Dundon TA, et al. STAT3 signalling is required for leptin regulation of energy balance but not reproduction. Nature 2003;421: Elmquist J, Bjorbaek C, Ahima R, et al. Distributions of leptin receptor mrna isoforms in the rat brain. J Comp Neurol 1998; 395: Cohen P, Zhao C, Cai X, et al. Selective deletion of leptin receptor in neurons leads to obesity. J Clin Invest 2001; 108: McMinn JE, Liu S-M, Liu H, et al. Neuronal deletion of Lepr elicits diabesity in mice without affecting cold tolerance or fertility. Am J Physiol Endocrinol Metab 2005;289:E Balthasar N, Coppari R, McMinn J, et al. Leptin receptor signaling in POMC neurons is required for normal body weight homeostasis. Neuron 2004;42: Elmquist JK, Elias CF, Saper CB. From lesions to leptin: hypothalamic control of food intake and body weight. Neuron 1999; 22: Elias C, Saper C, Maratos-Flier E, et al. Chemically defined projections linking the mediobasal hypothalamus and the lateral hypothalamic area. J Comp Neurol 1998;402: Elmquist JK, Ahima RS, Maratos-Flier E, et al. Leptin activates neurons in ventrobasal hypothalamus and brainstem. Endocrinology 1997;138: Hubschle T, Thom E, Watson A, et al. Leptin-induced nuclear translocation of STAT3 immunoreactivity in hypothalamic nuclei involved in body weight regulation. J Neurosci 2001; 21: Cowley M, Smart J, Rubinstein M, et al. Leptin activates anorexigenic POMC neurons through a neural network in the arcuate nucleus. Nature 2001;411: Coppari R, Ichinose M, Lee CE, et al. The hypothalamic arcuate nucleus: a key site for mediating leptin s effects on glucose homeostasis and locomotor activity. Cell Metab 2005;1: Obici S, Zhang BB, Karkanias G, et al. Hypothalamic insulin signaling is required for inhibition of glucose production. Nat Med 2002;8: Obici S, Feng Z, Karkanias G, et al. Decreasing hypothalamic insulin receptors causes hyperphagia and insulin resistance in rats. Nat Neurosci 2002;5: Gelling RW, Morton GJ, Morrison CD, et al. Insulin action in the brain contributes to glucose lowering during insulin treatment of diabetes. Cell Metab 2006;3: Morton GJ, Gelling RW, Niswender KD, et al. Leptin regulates insulin sensitivity via phosphatidylinositol-3-oh kinase signaling in mediobasal hypothalamic neurons. Cell Metab 2005;2: Markakis EA. Development of the neuroendocrine hypothalamus. Front Neuroendocrinol 2002;23: Bouret SG, Draper SJ, Simerly RB. Formation of projection pathways from the arcuate nucleus of the hypothalamus to hypothalamic regions implicated in the neural control of feeding behavior in mice. J Neurosci 2004;24: Grove KL, Allen S, Grayson BE, et al. Postnatal development of the hypothalamic neuropeptide Y system. Neuroscience 2003; 116: Singer LK, Kuper J, Brogan RS, et al. Novel expression of hypothalamic neuropeptide Y during postnatal development in the rat. Neuroreport 2000;11: Kesterson RA, Huszar D, Lynch CA, et al. Induction of neuropeptide Y gene expression in the dorsal medial hypothalamic nucleus in two models of the agouti obesity syndrome. Mol Endocrinol 1997;11: Keyser A. Development of the Hypothalamus in Mammals. New York: Marcel Dekker Inc; van Eerdenburg FJ, Rakic P. Early neurogenesis in the anterior hypothalamus of the rhesus monkey. Brain Res Dev Brain Res 1994;79: Bugnon C, Fellmann D, Bresson JL, et al. Immunocytochemical study of the ontogenesis of the CRH-containing neuroglandular system in the human hypothalamus. C R Acad Sci 1982;294: Burford GD, Robinson IC. Oxytocin, vasopressin and neurophysins in the hypothalamo-neurohypophysial system of the human fetus. J Endocrinol 1982;95: Ackland J, Ratter S, Bourne GL, et al. Characterization of immunoreactive somatostatin in human fetal hypothalamic tissue. Regul Pept 1983;5: Mai JK, Lensing-Hohn S, Ende AA, et al. Developmental organization of neurophysin neurons in the human brain. J Comp Neurol 1997;385:

8 S38 BOURET 43. Koutcherov Y, Mai JK, Ashwell KW, et al. Organization of human hypothalamus in fetal development. J Comp Neurol 2002;446: Grayson BE, Allen SE, Billes SK, et al. Prenatal development of hypothalamic neuropeptide systems in the nonhuman primate. Neuroscience 2006;143: Koutcherov Y. Organization of human hypothalamus in fetal development. J Comp Neurol 2002;446: Proulx K, Richard D, Walker CD. Leptin regulates appetite-related neuropeptides in the hypothalamus of developing rats without affecting food intake. Endocrinology 2002;143: Mistry A, Swick A, Romsos D. Leptin alters metabolic rates before acquisition of its anorectic effect in developing neonatal mice. Am J Physiol 1999;277:R Ahima R, Prabakaran D, Flier J. Postnatal leptin surge and regulation of circadian rhythm of leptin by feeding. Implications for energy homeostasis and neuroendocrine function. J Clin Invest 1998;101: Smith JT, Waddell BJ. Developmental changes in plasma leptin and hypothalamic leptin receptor expression in the rat: peripubertal changes and the emergence of sex differences. J Endocrinol 2003;176: Devaskar S, Ollesch C, Rajakumar R, et al. Developmental changes in ob gene expression and circulating leptin peptide concentration. Biochem Biophys Res Commun 1997;238: Oliver P, Picó C, De Matteis R, et al. Perinatal expression of leptin in rat stomach. Dev Dyn 2002;223: Casabiell X, Pineiro V, Tome MA, et al. Presence of leptin in colostrum and/or breast milk from lactating mothers: a potential role in the regulation of neonatal food intake. J Clin Endocrinol Metab 1997;82: McFadin EL, Morrison CD, Buff PR, et al. Leptin concentrations in periparturient ewes and their subsequent offspring. J Anim Sci 2002;80: Bouret S, Simerly RB. Development of leptin-sensitive circuits. J Neuroendocrinol 2007;19: Bouret SG, Draper SJ, Simerly RB. Trophic action of leptin on hypothalamic neurons that regulate feeding. Science 2004; 304: Yura S, Itoh H, Sagawa N, et al. Role of premature leptin surge in obesity resulting from intrauterine undernutrition. Cell Metab 2005;1: Attig L, Solomon G, Ferezou J, et al. Early postnatal leptin blockage leads to a long-term leptin resistance and susceptibility to diet-induced obesity in rats. Int J Obes 2008;32: Jones AP, Pothos EN, Rada P, et al. Maternal hormonal manipulations in rats cause obesity and increase medial hypothalamic norepinephrine release in male offspring. Dev Brain Res 1995; 88: Jones A, Olster D, States B. Maternal insulin manipulations in rats organize body weight and noradrenergic innervation of the hypothalamus in gonadally intact male offspring. Dev Brain Res 1996;97: Singh BS, Westfall TC, Devaskar SU. Maternal diabetes-induced hyperglycemia and acute intracerebral hyperinsulinism suppress fetal brain neuropeptide Y concentrations. Endocrinology 1997; 138: Plagemann A, Harder T, Rake A, et al. Morphological alterations of hypothalamic nuclei due to intrahypothalamic hyperinsulinism in newborn rats. Int J Dev Neurosci 1999;17: Levin BE, Dunn-Meynell AA, Balkan B, et al. Selective breeding for diet-induced obesity and resistance in Sprague- Dawley rats. Am J Physiol Regul Integr Comp Physiol 1997; 273:R Levin BE, Dunn-Meynell AA, Banks WA. Obesity-prone rats have normal blood-brain barrier transport but defective central leptin signaling before obesity onset. Am J Physiol Regul Integr Comp Physiol 2004;286:R Gorski JN, Dunn-Meynell AA, Levin BE. Maternal obesity increases hypothalamic leptin receptor expression and sensitivity in juvenile obesity-prone rats. Am J Physiol Regul Integr Comp Physiol 2007;292:R Levin BE, Dunn-Meynell AA. Reduced central leptin sensitivity in rats with diet-induced obesity. Am J Physiol Regul Integr Comp Physiol 2002;283:R Irani BG, Dunn-Meynell AA, Levin BE. Altered hypothalamic leptin, insulin, and melanocortin binding associated with moderatefat diet and predisposition to obesity. Endocrinology 2007; 148: Bouret SG, Gorski JN, Patterson CM, et al. Hypothalamic neural projections are permanently disrupted in diet-induced obese rats. Cell Metab 2008;7: McMillen IC, Robinson JS. Developmental origins of the metabolic syndrome: prediction, plasticity, and programming. Physiol Rev 2005;85: Davidowa H, Plagemann A. Different responses of ventromedial hypothalamic neurons to leptin in normal and early postnatally overfed rats. Neurosci Lett 2000;293: Davidowa H, Plagemann A. Decreased inhibition by leptin of hypothalamic arcuate neurons in neonatally overfed young rats. Neuroreport 2000;11: Davidowa H, Plagemann A. Insulin resistance of hypothalamic arcuate neurons in neonatally overfed rats. Neuroreport 2007; 18: Davidowa H, Li Y, Plagemann A. Altered responses to orexigenic (AGRP, MCH) and anorexigenic (alpha-msh, CART) neuropeptides of paraventricular hypothalamic neurons in early postnatally overfed rats. Eur J Neurosci 2003;18: Li Y, Plagemann A, Davidowa H. Increased inhibition by agoutirelated peptide of ventromedial hypothalamic neurons in rats overweight due to early postnatal overfeeding. Neurosci Lett 2002; 330: Heidel E, Plagemann A, Davidowa H. Increased response to NPYof hypothalamic VMN neurons in postnatally overfed juvenile rats. Neuroreport 1999;10: Plagemann A, Waas T, Harder T, et al. Hypothalamic neuropeptide Y levels in weaning offspring of low-protein malnourished mother rats. Neuropeptides 2000;34: Varma A, He J, Weissfeld L, et al. Postnatal intracerebroventricular exposure to neuropeptide Y causes weight loss in female adult rats. Am J Physiol Regul Integr Comp Physiol 2003; 284:R Muhlhausler BS, Adam CL, Findlay PA, et al. Increased maternal nutrition alters development of the appetite-regulating network in the brain. FASEB J 2006;20: Ferezou-Viala J, Roy A-F, Serougne C, et al. Long-term consequences of maternal high-fat feeding on hypothalamic leptin sensitivity and diet-induced obesity in the offspring. Am J Physiol Regul Integr Comp Physiol 2007;293:R