Effect of prorenin peptide vaccine on the early phase of diabetic retinopathy in a murine model of type 2 diabetes.

Prorenin is viewed as an ideal target molecule in the prevention of diabetic retinopathy. However, no drugs are available for inhibiting activation of prorenin. Here, we tested the effect of a prorenin peptide vaccine (VP) in the retina of a murine model of type 2 diabetes (T2D). To choose the optimal vaccine, we selected three different epitopes of the prorenin prosegment (E1, E2, and E3) and conjugated them to keyhole limpet hemocyanin (KLH). We injected C57BL/6J mice twice with KLH only (as a control vaccine), E1 conjugated with KLH (E1-KLH), E2-KLH, or E3-KLH and compared antibody titers. E2-KLH showed the highest antibody titer and specific immunoreactivity of anti-sera against prorenin, so we used E2-KLH as VP. Then, we administered injections to the non-diabetic db/m and diabetic db/db mice, as follows: db/m + KLH, db/db + KLH, and db/db + VP. Retinal blood flow measurement with laser speckle flowgraphy showed that the impaired retinal circulation response to both flicker light and systemic hyperoxia in db/db mice improved with VP. Furthermore, the prolonged implicit time of b-wave and oscillatory potentials in electroretinography was prevented, and immunohistochemical analysis showed reduced microglial activation, gliosis, and vascular leakage. The enzyme-linked immunosorbent spot assay confirmed vaccinated mice had no auto-immune response against prorenin itself. The present data suggest that vaccination against prorenin is an effective and safe measure against the early pathological changes of diabetic retinopathy in T2D.

Sustained Release of Insulin-Like Growth Factor-1 from Bombyx mori L. Silk Fibroin Delivery for Diabetic Wound Therapy.

The goals of this study are to develop a high purity patented silk fibroin (SF) film and test its suitability to be used as a slow-release delivery for insulin-like growth factor-1 (IGF-1). The release rate of the SF film delivering IGF-1 followed zero-order kinetics as determined via the Ritger and Peppas equation. The release rate constant was identified as 0.11, 0.23, and 0.09% h-1 at 37 °C for SF films loaded with 0.65, 6.5, and 65 pmol IGF-1, respectively. More importantly, the IGF-1 activity was preserved for more than 30 days when complexed with the SF film. We show that the IGF-1-loaded SF films significantly accelerated wound healing in vitro (BALB/3T3) and in vivo (diabetic mice), compared with wounds treated with free IGF-1 and an IGF-1-loaded hydrocolloid dressing. This was evidenced by a six-fold increase in the granulation tissue area in the IGF-1-loaded SF film treatment group compared to that of the PBS control group. Western blotting analysis also demonstrated that IGF-1 receptor (IGF1R) phosphorylation in diabetic wounds increased more significantly in the IGF-1-loaded SF films group than in other experimental groups. Our results suggest that IGF-1 sustained release from SF films promotes wound healing through continuously activating the IGF1R pathway, leading to the enhancement of both wound re-epithelialization and granulation tissue formation in diabetic mice. Collectively, these data indicate that SF films have considerable potential to be used as a wound dressing material for long-term IGF-1 delivery for diabetic wound therapy.

Leptin system loss of function in the absence of obesity in zebrafish.

The leptin system plays a crucial role in the regulation of appetite and energy homeostasis in vertebrates. While the phenotype of morbid obesity due to leptin (Lep) or leptin receptor (LEPR) loss of function is well established in mammals, evidence in fish is controversial, questioning the role of leptin as the vertebrate adipostat. Here we report on three (Lepr) loss of function (LOF) and one leptin loss of function alleles in zebrafish. In order to demonstrate that the Lepr LOF alleles cannot transduce a leptin signal, we measured socs3a transcription after i.p. leptin which is abolished by Lepr LOF. None of the Lepr/Lepa LOF alleles leads to obesity/a body growth phenotype. We explore possible reasons leading to the difference in published results and find that even slight changes in background genetics such as inbreeding siblings and cousins can lead to significant variance in growth.

Leptin receptor-expressing pericytes mediate access of hypothalamic feeding centers to circulating leptin.

Knowledge of how leptin receptor (LepR) neurons of the mediobasal hypothalamus (MBH) access circulating leptin is still rudimentary. Employing intravital microscopy, we found that almost half of the blood-vessel-enwrapping pericytes in the MBH express LepR. Selective disruption of pericytic LepR led to increased food intake, increased fat mass, and loss of leptin-dependent signaling in nearby LepR neurons. When delivered intravenously, fluorescently tagged leptin accumulated at hypothalamic LepR pericytes, which was attenuated upon pericyte-specific LepR loss. Because a paracellular tracer was also preferentially retained at LepR pericytes, we pharmacologically targeted regulators of inter-endothelial junction tightness and found that they affect LepR neuronal signaling and food intake. Optical imaging in MBH slices revealed a long-lasting, tonic calcium increase in LepR pericytes in response to leptin, suggesting pericytic contraction and vessel constriction. Together, our data indicate that LepR pericytes facilitate localized, paracellular blood-brain barrier leaks, enabling MBH LepR neurons to access circulating leptin.

Three-dimensional remodeling of functional cerebrovascular architecture and gliovascular unit in leptin receptor-deficient mice.

The cerebrovascular sequelae of diabetes render victims more susceptible to ischemic stroke, vascular cognitive impairment, and Alzheimer's disease. However, limited knowledge exists on the progressive changes in cerebrovascular structure and functional remodeling in type 2 diabetes. To ascertain the impact of diabetes on whole-brain cerebrovascular perfusion, leptin-receptor-deficient mice were transcardially injected with tomato-lectin before sacrifice. The whole brain was clarified by the Fast free-of-acrylamide clearing tissue technique. Functional vascular anatomy of the cerebrum was visualized by light-sheet microscopy, followed by analysis in Imaris software. We observed enhanced neovascularization in adult db/db mice, characterized by increased branch level and loop structures. Microvascular hypoperfusion was initially detected in juvenile db/db mice, suggesting early onset of insufficient microcirculation. Furthermore, gliovascular unit remodeling was verified by loss of pericytes and overactivation of microglia and astrocytes in adult diabetic mice. However, the integrity of the blood-brain barrier (BBB) was fundamentally preserved, as shown by a lack of extravasation of IgG into the brain parenchyma. In summary, we, for the first time, reveal that functional cerebrovascular remodeling occurs as early as four weeks in db/db mice and the deficit in gliovascular coupling may play a role in cerebral hypoperfusion before BBB breakdown in 16-week-old db/db mice.

Surgical ablation of whitened interscapular brown fat ameliorates cardiac pathology in salt-loaded metabolic syndrome rats.

Brown adipose tissue (BAT) is an endocrine organ that contributes to thermogenesis and energy consumption. We investigated the effects of salt loading and surgical removal of whitened interscapular BAT (iBAT) on cardiac and adipose tissue pathology in DahlS.Z-Leprfa /Leprfa (DS/obese) rats, an animal model of metabolic syndrome (MetS). DS/obese rats were subjected to surgical removal of iBAT or sham surgery at 8 weeks of age and were provided with drinking water containing or not containing 0.3% NaCl for 4 weeks beginning at 9 weeks of age. Removal of iBAT suppressed the salt-induced exacerbation of left ventricular inflammation, fibrosis, and diastolic dysfunction, but not that of hypertension development, in DS/obese rats. Salt loading attenuated adipocyte hypertrophy but enhanced inflammation in both visceral white adipose tissue (WAT) and iBAT. Although iBAT removal did not affect visceral WAT pathology in salt-loaded DS/obese rats, it attenuated the elevation of circulating interleukin-6 levels in these animals. Downregulation of uncoupling protein-1 expression in iBAT of DS/obese rats was not affected by salt loading. Our results suggest that the conversion of iBAT to WAT-like tissue contributes to a salt-induced elevation of circulating proinflammatory cytokine levels that leads to exacerbation of cardiac pathology in this model of MetS.

Leptin Receptors in RIP-Cre25Mgn Neurons Mediate Anti-dyslipidemia Effects of Leptin in Insulin-Deficient Mice.

Leptin is a potent endocrine hormone produced by adipose tissue and regulates a broad range of whole-body metabolism such as glucose and lipid metabolism, even without insulin. Central leptin signaling can lower hyperglycemia in insulin-deficient rodents via multiple mechanisms, including improvements of dyslipidemia. However, the specific neurons that regulate anti-dyslipidemia effects of leptin remain unidentified. Here we report that leptin receptors (LEPRs) in neurons expressing Cre recombinase driven by a short fragment of a promoter region of Ins2 gene (RIP-Cre25Mgn neurons) are required for central leptin signaling to reverse dyslipidemia, thereby hyperglycemia in insulin-deficient mice. Ablation of LEPRs in RIP-Cre25Mgn neurons completely blocks glucose-lowering effects of leptin in insulin-deficient mice. Further investigations reveal that insulin-deficient mice lacking LEPRs in RIP-Cre25Mgn neurons (RIP-CreΔLEPR mice) exhibit greater lipid levels in blood and liver compared to wild-type controls, and that leptin injection into the brain does not suppress dyslipidemia in insulin-deficient RIP-CreΔLEPR mice. Leptin administration into the brain combined with acipimox, which lowers blood lipids by suppressing triglyceride lipase activity, can restore normal glycemia in insulin-deficient RIP-CreΔLEPR mice, suggesting that excess circulating lipids are a driving-force of hyperglycemia in these mice. Collectively, our data demonstrate that LEPRs in RIP-Cre25Mgn neurons significantly contribute to glucose-lowering effects of leptin in an insulin-independent manner by improving dyslipidemia.

Leptin signalling on arcuate NPY neurones controls adiposity independent of energy balance or diet composition.

Central action of the adipocyte hormone leptin via the neuropeptide Y (NPY) system is considered critical for energy homeostatic control. However, the precise mechanisms for this control are still not clear. To specifically investigate how leptin signalling on the NPY neurone contributes to the control of energy homeostasis, we generated an inducible adult-onset NPY neurone-specific leptin receptor (Lepr) knockout model and performed a comprehensive metabolic phenotyping study. Here, we show that the NPY neurone subpopulation that is directly responsive to leptin is not required for the inhibition of fasting-induced hyperphagia by leptin, although it is essential for the regulation of adiposity independent of changes in energy balance or diet composition. Furthermore, under obesogenic conditions such as a high-fat diet, a lack of Lepr signalling on NPY neurones results in significant increases in food intake and concomitant reductions in energy expenditure, leading to accelerated accumulation of fat mass. Collectively, these findings support the notion that Lepr-expressing NPY neurones act as the key relay point where peripheral adipose storage information is sensed, and corresponding responses are initiated to protect adipose reserves.

Fatty acid transport protein-2 regulates glycemic control and diabetic kidney disease progression.

Kidney disease is one of the most devastating complications of diabetes, and tubular atrophy predicts diabetic kidney disease (DKD) progression to end-stage renal disease. We have proposed that fatty acids bound to albumin contribute to tubular atrophy by inducing lipotoxicity, after filtration across damaged glomeruli, and subsequent proximal tubule reabsorption by a fatty acid transport protein-2-dependent (FATP2-dependent) mechanism. To address this possibility, genetic (Leprdb/db eNOS-/-) and induced (high-fat diet plus low-dose streptozotocin) mouse models of obesity and DKD were bred with global FATP2 gene-deleted mice (Slc27a2) and then phenotyped. DKD-prone mice with the Slc27a2-/- genotype demonstrated normalization of glomerular filtration rate, reduced albuminuria, improved kidney histopathology, and longer life span compared with diabetic Slc27a2+/+ mice. Genetic and induced DKD-prone Slc27a2-/- mice also exhibited markedly reduced fasting plasma glucose, with mean values approaching euglycemia, despite increased obesity and decreased physical activity. Glucose lowering in DKD-prone Slc27a2-/- mice was accompanied by ß cell hyperplasia and sustained insulin secretion. Together, our data indicate that FATP2 regulates DKD pathogenesis by a combined lipotoxicity and glucotoxicity (glucolipotoxicity) mechanism.

Effects of leptin on HPG axis and reproductive function in male rat in simulated altitude of 5500 m hypoxia environment.

High altitude hypobaric hypoxia environment impairs male's reproductive function. Leptin is an adipose tissue-derived hormone which regulates body weight homeostasis. Its receptor (LepR) has been found in all levels of male reproductive axis, indicating that it can affect male reproductive system in a direct or (and) indirect way. However, the role of leptin signaling in hypobaric hypoxia induced male reproductive dysfunction remains to be elucidated. In this study, we investigated the changes of leptin levels in male SD rats in stimulated altitude of 5500 m hypobaric hypoxia environment and their effects on the hypothalamus-pituitary-gonad axis (HPG axis). A hypoxia animal model was established using a hypobaric hypoxia chamber. Rats were divided randomly into 1, 7, 14, 28-day hypoxia group, recovery group (14 days hypoxia+14 days normoxia) and their control groups. Hypoxia groups displayed obvious changes of testicular and epididymis index compared to control groups. The total number of sperm and sperm motility rate decreased dramatically, while sperm deformity rate increased in hypoxia groups. The flow cytometry analysis showed that the percentage of haploid in 1-day, 7-day and 28-day hypoxia groups increased while the proportion of diploid decreased in 14-day and 28-day hypoxia group. TUNEL staining showed that the testis cells apoptosis index (AI) of hypoxia groups increased significantly, and the apoptosis of cells mainly focus on spermatogonia and spermatocytes. The expression of GnRH in hypothalamus decreased dramatically under hypoxia condition, accompanied with the reduction of serum testosterone (T) level in 1-day and 28-day hypoxia groups and free-testosterone level (FT) in 1-day and 14-day hypoxia groups. Importantly, ELISA analysis showed that serum leptin level decreased in 7-day hypoxia groups and acylated-ghrelin, gastrin also changed, accompanying with reduction of LepR in hypothalamus in hypoxia groups. Immunohistochemical staining exhibited increased leptin and LepR in testis under hypobaric hypoxia conditions. Our results suggested that simulated high altitude hypobaric hypoxia environment decreased male reproductive function, depressed HPG axis activity and altered the serum concentration of hormones related to food intake in adult male rats. Additionally, hypobaric hypoxia induced the leptin-LepR expression in adult male rats' testis, suggesting leptin-LepR signaling may mediate hypoxia-induced impairment in male rats' reproductive system.

Leptin increases sympathetic nerve activity via induction of its own receptor in the paraventricular nucleus.

Whether leptin acts in the paraventricular nucleus (PVN) to increase sympathetic nerve activity (SNA) is unclear, since PVN leptin receptors (LepR) are sparse. We show in rats that PVN leptin slowly increases SNA to muscle and brown adipose tissue, because it induces the expression of its own receptor and synergizes with local glutamatergic neurons. PVN LepR are not expressed in astroglia and rarely in microglia; instead, glutamatergic neurons express LepR, some of which project to a key presympathetic hub, the rostral ventrolateral medulla (RVLM). In PVN slices from mice expressing GCaMP6, leptin excites glutamatergic neurons. LepR are expressed mainly in thyrotropin-releasing hormone (TRH) neurons, some of which project to the RVLM. Injections of TRH into the RVLM and dorsomedial hypothalamus increase SNA, highlighting these nuclei as likely targets. We suggest that this neuropathway becomes important in obesity, in which elevated leptin maintains the hypothalamic pituitary thyroid axis, despite leptin resistance.

A Novel Indoline Derivative Ameliorates Diabesity-Induced Chronic Kidney Disease by Reducing Metabolic Abnormalities.

Both diabetes and obesity (diabesity) contribute significantly to the development of chronic kidney disease (CKD). In search of new remedies to reverse or arrest the progression of CKD, we examined the therapeutic potential of a novel compound, AN1284, in a mouse model of CKD induced by type 2 diabetes with obesity. Six-week-old BKS Cg-Dock 7m+/+ Leprdb/J mice with type 2 diabetes and obesity were treated with AN1284 (2.5 or 5 mg kg-1 per day) via micro-osmotic pumps implanted subcutaneously for 3 months. Measures included renal, pancreatic, and liver assessment as well as energy utilization. AN1284 improved kidney function in BSK-db/db animals by reducing albumin and creatinine and preventing renal inflammation and morphological changes. The treatment was associated with weight loss, decreased body fat mass, increased utilization of body fat toward energy, preservation of insulin sensitivity and pancreatic ß cell mass, and reduction of dyslipidemia, hepatic steatosis, and liver injury. This indoline derivative protected the kidney from the deleterious effects of hyperglycemia by ameliorating the metabolic abnormalities of diabetes. It could have therapeutic potential for preventing CKD in human subjects with diabesity.

Interaction between genes involved in energy intake regulation and diet in obesity.

Obesity is a multifactorial, complex, and public health problem worldwide. Interaction between genes and environment as associated with diet may predispose an individual to obesity. In this sense, nutrigenetics appears to be a strategy that can improve understanding of the gene-diet interaction. The aim of this literature review was to summarize data from studies of genes involved in the regulation of energy intake (melanocortin 4 receptor [MC4R], fat mass and obesity-associated [FTO], ghrelin [GHRL], leptin [LEP], and cholecystokinin [CCK]) and diet interaction in obesity. The presence of polymorphisms in MC4R, FTO, leptin, and the respective receptor appear to be associated with higher energy and total lipid consumption. Polymorphisms in FTO, leptin, and leptin receptor are also related to increased intake of saturated fatty acids. Individuals with the MC4R, FTO, and ghrelin polymorphisms, who submitted themselves for weight loss intervention, appeared to achieve weight loss similar to individuals without polymorphisms in these genes. Additionally, protein seems to interact with these genes, which increases or decreases appetite, or to drive or lessen body weight recovery. Additionally, polymorphisms in these genes were found to be associated with inappropriate eating behaviors, such as increased consumption of sweets and snacks, consumption of large food portions, desire to eat, and eating associated with emotional issues. Preliminary data has supported the gene-diet interaction in determining weight loss and gain in individuals with polymorphisms in the genes involved in energy intake. Despite the advent of nutrigenetics in obesity, it is still too early to define the dietary management for weight loss based on the presence or absence of obesity polymorphisms.

Mesenchymal stromal cells lower platelet activation and assist in platelet formation in vitro.

The complex process of platelet formation originates with the hematopoietic stem cell, which differentiates through the myeloid lineage, matures, and releases proplatelets into the BM sinusoids. How formed platelets maintain a low basal activation state in the circulation remains unknown. We identify Lepr+ stromal cells lining the BM sinusoids as important contributors to sustaining low platelet activation. Ablation of murine Lepr+ cells led to a decreased number of platelets in the circulation with an increased activation state. We developed a potentially novel culture system for supporting platelet formation in vitro using a unique population of CD51+PDGFRα+ perivascular cells, derived from human umbilical cord tissue, which display numerous mesenchymal stem cell (MSC) properties. Megakaryocytes cocultured with MSCs had altered LAT and Rap1b gene expression, yielding platelets that are functional with low basal activation levels, a critical consideration for developing a transfusion product. Identification of a regulatory cell that maintains low baseline platelet activation during thrombopoiesis opens up new avenues for improving blood product production ex vivo.

Leptin receptor deficiency induces early, transient and hyperglycaemia-independent blood-brain barrier dysfunction.

Diabetes mellitus (DM) significantly increases susceptibility to central nervous system (CNS) pathologies, including stroke, vascular dementia, cognitive deficits and Alzheimer's disease. Previous studies (mostly using the streptozotocin model) suggested that blood-brain barrier (BBB) disruption is involved in these conditions. Here, we examined the integrity of brain capillaries and BBB permeability in Leprdb/db obesity-related diabetic mice. Surprisingly, significant BBB leakage was observed only in young mice at the pre-hyperglycemic stage. Thorough examination of barrier permeability at later diabetic stages showed no evidence for significant BBB leakage during the hyperglycemic state. Electron microscopy imaging of mice with short-term hyperglycaemia supported normal BBB permeability but indicated other stress-related changes in capillary ultrastructure, such as mitochondrial degeneration. Based on our study with this mouse genetic model of obesity-related DM, we suggest that previously reported hyperglycaemia-induced BBB leakage is most likely not the underlying mechanism of DM-related CNS pathologies. Finally we propose that BBB hyper-permeability might be an early and transient phenomenon while stress-related endothelial pathologies do correlate with a short-term diabetic state.

The LepRdb/db mice model for studying glycation in the context of diabetes.

BACKGROUND: Early (furosine) and advanced (carboxymethyllysine, CML) products of glycation (AGEs) have been reported as increased in plasma, tissues, and organs of diabetic people, indicating a direct link between glycation and type 2 diabetes (T2D). While murine models present some of the characteristics observed in diabetic humans, their pertinence as models of glycation, particularly for T2D, remains poorly described. The aim of this study was to characterize and compare glycation in several organs of two commonly studied murine models of T2D using stable isotope dilution liquid chromatography tandem mass spectrometry (LC-MS/MS). METHODS: Defining parameters of type 2 diabetes including body weight, fasting glycaemia, and glucose intolerance were measured in three different C57BL6 mouse models of T2D-the genetic LepRdb/db (db/db) model and two diet-induced obesity (DIO) models-and their respective controls. Furosine, free, and protein-bound CML were quantified in kidneys, lungs, heart, and liver by LC-MS/MS. RESULTS: The obesity, hyperglycaemia, and glucose intolerance in db/db mice was accompanied by an increase of furosine and protein-bound CML levels in all organs relative to controls. The DIO models took several months to become obese, exhibited less severe hyperglycaemia and glucose intolerance, while glycation products were not significantly different between these groups (with the exception of furosine in liver and CML in lungs). CONCLUSIONS: The db/db model better reflected the characteristics of human T2D compared with the DIO models and exhibited greater formation and accumulation of both furosine and protein-bound CML in all of the organs tested here.

Leptin acts in the carotid bodies to increase minute ventilation during wakefulness and sleep and augment the hypoxic ventilatory response.

KEY POINTS: Leptin is a potent respiratory stimulant. A long functional isoform of leptin receptor, LepRb , was detected in the carotid body (CB), a key peripheral hypoxia sensor. However, the effect of leptin on minute ventilation (VE ) and the hypoxic ventilatory response (HVR) has not been sufficiently studied. We report that LepRb is present in approximately 74% of the CB glomus cells. Leptin increased carotid sinus nerve activity at baseline and in response to hypoxia in vivo. Subcutaneous infusion of leptin increased VE and HVR in C57BL/6J mice and this effect was abolished by CB denervation. Expression of LepRb in the carotid bodies of LepRb deficient obese db/db mice increased VE during wakefulness and sleep and augmented the HVR. We conclude that leptin acts on LepRb in the CBs to stimulate breathing and HVR, which may protect against sleep disordered breathing in obesity. ABSTRACT: Leptin is a potent respiratory stimulant. The carotid bodies (CB) express the long functional isoform of leptin receptor, LepRb , but the role of leptin in CB has not been fully elucidated. The objectives of the current study were (1) to examine the effect of subcutaneous leptin infusion on minute ventilation (VE ) and the hypoxic ventilatory response to 10% O2 (HVR) in C57BL/6J mice before and after CB denervation; (2) to express LepRb in CB of LepRb -deficient obese db/db mice and examine its effects on breathing during sleep and wakefulness and on HVR. We found that leptin enhanced carotid sinus nerve activity at baseline and in response to 10% O2 in vivo. In C57BL/6J mice, leptin increased VE from 1.1 to 1.5 mL/min/g during normoxia (P < 0.01) and from 3.6 to 4.7 mL/min/g during hypoxia (P < 0.001), augmenting HVR from 0.23 to 0.31 mL/min/g/Δ FIO2 (P < 0.001). The effects of leptin on VE and HVR were abolished by CB denervation. In db/db mice, LepRb expression in CB increased VE from 1.1 to 1.3 mL/min/g during normoxia (P < 0.05) and from 2.8 to 3.2 mL/min/g during hypoxia (P < 0.02), increasing HVR. Compared to control db/db mice, LepRb transfected mice showed significantly higher VE throughout non-rapid eye movement (20.1 vs. -27.7 mL/min respectively, P < 0.05) and rapid eye movement sleep (16.5 vs 23.4 mL/min, P < 0.05). We conclude that leptin acts in CB to augment VE and HVR, which may protect against sleep disordered breathing in obesity.

The Leptin Signaling.

Leptin plays a critical role in the regulation of energy balance and metabolic homeostasis. Impairment of leptin signaling is closely involved in the pathogenesis of obesity and metabolic diseases, including diabetes, cardiovascular disease, etc. Leptin initiates its intracellular signaling in the leptin-receptor-expressing neurons in the central nervous system to exert physiological function, thereby leading to a suppression of appetite, a reduction of food intake, a promotion of mitochondrial oxidation, an enhancement of thermogenesis, and a decrease in body weight. In this review, the studies on leptin neural and cellular pathways are summarized with an emphasis on the progress made during the last 10 years, for better understanding the molecular mechanism of obesity and other metabolic diseases.

To eat or not to eat: ontogeny of hypothalamic feeding controls and a role for leptin in modulating life-history transition in amphibian tadpoles.

Many animal life histories entail changing feeding ecology, but the molecular bases for these transitions are poorly understood. The amphibian tadpole is typically a growth and dispersal life-history stage. Tadpoles are primarily herbivorous, and they capitalize on growth opportunities to reach a minimum body size to initiate metamorphosis. During metamorphic climax, feeding declines, at which time the gastrointestinal (GI) tract remodels to accommodate the carnivorous diet of the adult frog. Here we show that anorexigenic hypothalamic feeding controls are absent in the tadpole, but develop during metamorphosis concurrent with the production of the satiety signal leptin. Before metamorphosis there is a large increase in leptin mRNA in fat tissue. Leptin receptor mRNA increased during metamorphosis in the preoptic area/hypothalamus, the key brain region involved with the control of food intake and metabolism. This corresponded with an increase in functional leptin receptor, as evidenced by induction of socs3 mRNA and phosphorylated STAT3 immunoreactivity, and suppression of feeding behaviour after injection of recombinant frog leptin. Furthermore, we found that immunoneutralization of leptin in tadpoles at metamorphic climax caused them to resume feeding. The absence of negative regulation of food intake in the tadpole allows the animal to maximize growth prior to metamorphosis. Maturation of leptin-responsive neural circuits suppresses feeding during metamorphosis to facilitate remodelling of the GI tract.