Dorsomedial Nucleus Of The Hypothalamus Research Articles

The temporal and spatial pattern of leptin receptor-expressing cells in the developing mouse hypothalamus.

The arcuate nucleus is a crucial hypothalamic brain region involved in regulating body weight homeostasis. Neurons within the arcuate nucleus respond to peripheral metabolic signals, such as leptin, and relay these signals via neuronal projections to brain regions both within and outside the hypothalamus, ultimately causing changes in an animal's behaviour and physiology. There is a substantial amount of evidence to indicate that leptin is intimately involved with the postnatal development of arcuate nucleus melanocortin circuitry. Further, it is clear that leptin signalling directly in the arcuate nucleus is required for circuitry development. However, as leptin receptor long isoform (Leprb) mRNA is expressed in multiple nuclei within the developing hypothalamus, including the postsynaptic target regions of arcuate melanocortin projections, this raises the possibility that leptin also signals in these nuclei to promote circuitry development. Here, we used RT-qPCR and RNAscope® to reveal the spatio-temporal pattern of Leprb mRNA in the early postnatal mouse hypothalamus. We found that Leprb mRNA expression increased significantly in the arcuate nucleus, ventromedial nucleus and paraventricular nucleus of the hypothalamus from P8, in concert with the leptin surge. In the dorsomedial nucleus of the hypothalamus, increases in Leprb mRNA were slightly later, increasing significantly from P12. Using duplex RNAscope®, we found Leprb co-expressed with Sim1, Pou3f2, Mc4r and Bdnf in the paraventricular nucleus at P8. Together, these data suggest that leptin may signal in a subset of neurons postsynaptic to arcuate melanocortin neurons, as well as within the arcuate nucleus itself, to promote the formation of arcuate melanocortin circuitry during the early postnatal period.

Low-dose peripheral leptin infusion produces selective activation of ventromedial hypothalamic and hindbrain STAT3.

Previous studies have shown that very low dose, acute, single peripheral leptin injections fully activate arcuate nucleus signal transducer and activator of transcription 3 (STAT3), but ventromedial hypothalamus (VMH) pSTAT3 continues to increase with higher doses of leptin that inhibit food intake. The lowest dose that inhibited intake increased circulating leptin 300-fold whereas food intake is inhibited by chronic peripheral leptin infusions that only double circulating leptin. This study examined whether the pattern of hypothalamic pSTAT3 was the same in leptin-infused rats as in leptin-injected rats. Male Sprague-Dawley rats received intraperitoneal infusions of 0, 5, 10, 20, or 40 µg leptin/day for 9 days. The highest dose of leptin increased serum leptin by 50-100%, inhibited food intake for 5 days, but inhibited weight gain and retroperitoneal fat mass for 9 days. Energy expenditure, respiratory exchange ratio, and brown fat temperature did not change. pSTAT3 was quantified in hypothalamic nuclei and the nucleus of the solitary tract (NTS) when food intake was inhibited and when it had returned to control levels. There was no effect of leptin on pSTAT3 in the medial or lateral arcuate nucleus or in the dorsomedial nucleus of the hypothalamus. VMH pSTAT3 was increased only at day 4 when food intake was inhibited, but NTS pSTAT3 was increased at both 4 and 9 days of infusion. These results suggest that activation of leptin VMH receptors contributes to the suppression of food intake, but that hindbrain receptors contribute to a sustained change in metabolism that maintains a reduced weight and fat mass.NEW & NOTEWORTHY Low-dose, chronic peripheral infusions of leptin produced an initial, transient inhibition of food intake that correlated with signal transducer and activator of transcription 3 (STAT3) activation in the ventromedial hypothalamus (VMH) and nucleus of the solitary tract (NTS). When intake normalized, but weight remained suppressed, the NTS was the only area that remained activated. These data suggest that leptin's primary function is to reduce body fat, that hypophagia is a means of achieving this and that different areas of the brain are responsible for the progressive response.

Sex-dependent and -independent regulation of thyrotropin-releasing hormone expression in the hypothalamic dorsomedial nucleus by negative energy balance, exercise, and chronic stress

The dorsomedial nucleus of the hypothalamus (DMH) is part of the brain circuits that modulate organism responses to the circadian cycle, energy balance, and psychological stress. A large group of thyrotropin-releasing hormone (Trh) neurons is localized in the DMH; they comprise about one third of the DMH neurons that project to the lateral hypothalamus area (LH). We tested their response to various paradigms. In male Wistar rats, food restriction during adulthood, or chronic variable stress (CVS) during adolescence down-regulated adult DMH Trh mRNA levels compared to those in sedentary animals fed ad libitum; two weeks of voluntary wheel running during adulthood enhanced DMH Trh mRNA levels compared to pair-fed rats. Except for their magnitude, female responses to exercise were like those in male rats; in contrast, in female rats CVS did not change DMH Trh mRNA levels. A very strong negative correlation between DMH Trh mRNA levels and serum corticosterone concentration in rats of either sex was lost in CVS rats. CVS canceled the response to food restriction, but not that to exercise in either sex. TRH receptor 1 (Trhr) cells were numerous along the rostro-caudal extent of the medial LH. In either sex, fasting during adulthood reduced DMH Trh mRNA levels, and increased LH Trhr mRNA levels, suggesting fasting may inhibit the activity of TRHDMH->LH neurons. Thus, in Wistar rats DMH Trh mRNA levels are regulated by negative energy balance, exercise and chronic variable stress through sex-dependent and -independent pathways.

Immunoreactive distribution of gonadotropin-inhibitory hormone precursor, RFRP, in a temperate bat species (Eptesicus fuscus).

Gonadotropin-inhibitory hormone (GnIH, also known RFRP-3 in mammals) is an important regulator of the hypothalamic-pituitary-gonadal axis and downstream reproductive physiology. Substantial species differences exist in the localization of cell bodies producing RFRP-3 and patterns of fiber immunoreactivity in the brain, raising the question of functional differences. Many temperate bat species exhibit unusual annual reproductive patterns. Male bats upregulate spermatogenesis in late spring which is asynchronous with periods of mating in the fall, while females have the physiological capacity to delay their reproductive investment over winter via sperm storage or delayed ovulation/fertilization. Neuroendocrine mechanisms regulating reproductive timing in male and female bats are not well-studied. We provide the first description of RFRP-precursor peptide of GnIH -expression and localization in the brain of any bat using a widespread temperate species (Eptesicus fuscus, big brown bat) as a model. RFRP mRNA expression was detected in the hypothalamus, testes, and ovaries of big brown bats. Cellular RFRP-immunoreactivity was observed within the periventricular nuclei, dorsomedial nucleus of the hypothalamus, arcuate nucleus (Arc), and median eminence (ME). As in other vertebrates, RFRP fiber immunoreactivity was widespread, with the greatest density observed in the hypothalamus, preoptic area, Arc, ME, midbrain, and thalamic nuclei. Putative interactions between RFRP-ir fibers and gonadotropin-releasing hormone (GnRH) cell bodies were observed in 16% of GnRH-immunoreactive cells, suggesting direct regulation of GnRH via RFRP signaling. This characterization of RFRP distribution contributes to a deeper understanding of bat neuroendocrinology, which serves as foundation for manipulative approaches examining changes in reproductive neuropeptide signaling in response to environmental and physiological challenges within, and among, bat species.

Developmental genes controlling neural circuit formation are expressed in the early postnatal hypothalamus and cellular lining of the third ventricle.

The arcuate nucleus of the hypothalamus is central in the regulation of body weight homeostasis through its ability to sense peripheral metabolic signals and relay them, through neural circuits, to other brain areas, ultimately affecting physiological and behavioural changes. The early postnatal development of these neural circuits is critical for normal body weight homeostasis, such that perturbations during this critical period can lead to obesity. The role for peripheral regulators of body weight homeostasis, including leptin, insulin and ghrelin, in this postnatal development is well described, yet some of the fundamental processes underpinning axonal and dendritic growth remain unclear. Here, we hypothesised that molecules known to regulate axonal and dendritic growth processes in other areas of the developing brain would be expressed in the postnatal arcuate nucleus and/or target nuclei where they would function to mediate the development of this circuitry. Using state-of-the-art RNAscope® technology, we have revealed the expression patterns of genes encoding Dcc/Netrin-1, Robo1/Slit1 and Fzd5/Wnt5a receptor/ligand pairs in the early postnatal mouse hypothalamus. We found that individual genes had unique expression patterns across developmental time in the arcuate nucleus, paraventricular nucleus of the hypothalamus, ventromedial nucleus of the hypothalamus, dorsomedial nucleus of the hypothalamus, median eminence and, somewhat unexpectedly, the third ventricle epithelium. These observations indicate a number of new molecular players in the development of neural circuits regulating body weight homeostasis, as well as novel molecular markers of tanycyte heterogeneity.

Presynaptic inputs to vasopressin neurons in the hypothalamic supraoptic nucleus and paraventricular nucleus in mice

Arginine vasopressin (AVP) neurons in the hypothalamic supraoptic nucleus (SON) and paraventricular nucleus (PVN) are involved in important physiological behaviors, such as controling osmotic stability and thermoregulation. However, the presynaptic input patterns governing AVP neurons have remained poorly understood due to their heterogeneity, as well as intermingling of AVP neurons with other neurons both in the SON and PVN. In the present study, we employed a retrograde modified rabies-virus system to reveal the brain areas that provide specific inputs to AVP neurons in the SON and PVN. We found that AVP neurons of the SON and PVN received similar input patterns from multiple areas of the brain, particularly massive afferent inputs from the diencephalon and other brain regions of the limbic system; however, PVNAVP neurons received relatively broader and denser inputs compared to SONAVP neurons. Additionally, SONAVP neurons received more projections from the median preoptic nucleus and organum vasculosum of the lamina terminalis (a circumventricular organ), compared to PVNAVP neurons, while PVNAVP neurons received more afferent inputs from the bed nucleus of stria terminalis and dorsomedial nucleus of the hypothalamus, both of which are thermoregulatory nuclei, compared to those of SONAVP neurons. In addition, both SONAVP and PVNAVP neurons received direct afferent projections from the bilateral suprachiasmatic nucleus, which is the master regulator of circadian rhythms and is concomitantly responsible for fluctuations in AVP levels. Taken together, our present results provide a comprehensive understanding of the specific afferent framework of AVP neurons both in the SON and PVN, and lay the foundation for further dissecting the diverse roles of SONAVP and PVNAVP neurons.

Fasting reduces the number of TRH immunoreactive neurons in the hypothalamic paraventricular nucleus of male rats, but not in mice

During fasting or weight loss, the fall in leptin levels leads to suppression of thyrotropin-releasing hormone (TRH) expression in the paraventricular nucleus of the hypothalamus (PVH) and, consequently, inhibition of the hypothalamic-pituitary-thyroid (HPT) axis. However, differently than rats, just few PVHTRH neurons express the leptin receptor in mice. In the present study, male adult rats and mice were submitted to 48 -h fasting to evaluate the consequences on proTRH peptide expression at the PVH level. Additionally, the proTRH peptide expression was also assessed in the brains of leptin-deficient (Lepob/ob) mice. We observed that approximately 50 % of PVHTRH neurons of leptin-injected rats exhibited phosphorylation of the signal transducer and activator of transcription 3 (pSTAT3), a marker of leptin receptor activation. In contrast, very few PVHTRH neurons of leptin-injected mice exhibited pSTAT3. Rats submitted to 48 -h fasting showed a significant reduction in the number of PVHTRH immunoreactive neurons, as compared to fed rats. On the other hand, no changes in the number of PVHTRH immunoreactive neurons were observed between fasted and fed mice. Next, the number of TRH immunoreactive cells was determined in the PVH, dorsomedial nucleus of the hypothalamus and nucleus raphe pallidus of Lepob/ob and wild-type mice and no significant differences were observed, despite reduced plasma T4 levels in Lepob/ob mice. Taken together, these findings provide additional evidence of the important species-specific differences in the mechanisms used by fasting and/or leptin to regulate the HPT axis.

Peculiar protrusions along tanycyte processes face diverse neural and nonneural cell types in the hypothalamic parenchyma.

Tanycytes are highly specialized ependymal cells that line the bottom and the lateral walls of the third ventricle. In contact with the cerebrospinal fluid through their cell bodies, they send processes into the arcuate nucleus, the ventromedial nucleus, and the dorsomedial nucleus of the hypothalamus. In the present work, we combined transgenic and immunohistochemical approaches to investigate the neuroanatomical associations between tanycytes and neural cells present in the hypothalamic parenchyma, in particular in the arcuate nucleus. The specific expression of tdTomato in tanycytes first allowed the observation of peculiar subcellular protrusions along tanycyte processes and at their endfeet such as spines, swelling, en passant boutons, boutons, or claws. Interestingly, these protrusions contact different neural cells in the brain parenchyma including blood vessels and neurons, and in particular NPY and POMC neurons in the arcuate nucleus. Using both fluorescent and electron microscopy, we finally observed that these tanycyte protrusions contain ribosomes, mitochondria, diverse vesicles, and transporters, suggesting dense tanycyte/neuron and tanycyte/blood vessel communications. Altogether, our results lay the neuroanatomical basis for tanycyte/neural cell interactions, which will be useful to further understand cell‐to‐cell communications involved in the regulation of neuroendocrine functions.

Nitric Oxide Acts in the Rat Dorsomedial Hypothalamus to Increase High Fat Food Intake and Glutamate Transmission

The dorsomedial nucleus of the hypothalamus (DMH) plays an important role in the regulation of energy intake and expenditure. Numerous appetite-regulatory signals present in the DMH, including nitric oxide (NO) and endogenous cannabinoids (eCBs), act to regulate food intake, but whether these signals are involved in regulating high fat food intake remains unknown. We therefore asked whether NO and eCBs, administered alone or in combination, would influence the consumption of high fat food in rats. We implanted bilateral guide cannulas in the DMH of young, male Sprague-Dawley rats for microinjection of vehicle, NO (via the precursor l-arginine), the eCB 2-arachidonylglycerol (2-AG), or a combination of the two signals. Following the intrahypothalamic injections, both high fat food intake and body weight were measured for two hours at which point brains were removed and sectioned to confirm cannula placement in the DMH. Here we show that l-arginine significantly increases high fat food intake when administered into the DMH. This effect is abolished in the presence of 2-AG, which alone has no effect on high fat food intake or body weight. The l-arginine-induced increase in high fat food intake is dependent on NO synthesis, as it is prevented with the NO synthase inhibitor, l-NAME. We also demonstrate that l-arginine increases glutamate transmission onto DMH neurons, an effect that also requires NO synthesis and is abolished with 2-AG. Together, these data indicate that NO acts in the DMH to regulate the consumption of high fat food, possibly by enhancing glutamate signaling at DMH synapses.

Significance of medial preoptic area among the subcortical and cortical areas that are related to pain regulation in the rats with stress-induced hyperalgesia

Psychophysical stresses frequently increase sensitivity and response to pain, which is termed stress-induced hyperalgesia (SIH). However, the mechanism remains unknown. The subcortical areas such as medial preoptic area (MPO), dorsomedial nucleus of the hypothalamus (DMH), basolateral (BLA) and central nuclei of the amygdala (CeA), and the cortical areas such as insular (IC) and anterior cingulate cortices (ACC) play an important role in pain control via the descending pain modulatory system. In the present study we examined the expression of phosphorylated -cAMP-response element binding protein (pCREB) and the acetylation of histone H3 in these subcortical and cortical areas after repeated restraint stress to reveal changes in the subcortical and cortical areas that affect the function of descending pain modulatory system in the rats with SIH. The repeated restraint stress for 3 weeks induced a decrease in mechanical threshold in the rat hindpaw, an increase in the expression of pCREB in the MPO and an increase in the acetylation of histone H3 in the MPO, BLA and IC. The MPO was the only area that showed an increase in both the expression of pCREB and the acetylation of histone H3 among these examined areas after the repeated restraint stress. Furthermore, the number of pCREB-IR or acetylated histone H3-IR cells in the MPO was negatively correlated with the mechanical threshold. Together, our data represent the importance of the MPO among the subcortical and cortical areas that control descending pain modulatory system under the condition of SIH.

Vulnerability to stress consequences induced by repeated social defeat in rats: Contribution of the angiotensin II type 1 receptor in cardiovascular alterations associated to low brain derived neurotrophic factor

After social stress, rats become vulnerable to depression, and this state is characterized by persistent low blood levels of brain-derived neurotrophic factor (BDNF). The aim of this study was to determine whether low BDNF levels are associated with long term autonomic changes. Defeated animals were subjected to four daily episodes of social defeats. Twenty five days later, defeated rats with low BDNF levels (Dlow) still displayed elevated sympathetic tone (as indicated by an elevated low frequency to high frequency ratio (LF/HF) in heart rate) and elevated blood pressure, as well as reduced baroreflex sensitivity (BRS). In contrast, those with higher BDNF levels (Dhigh) similar to controls, did not. Dlow animals persistent cardiovascular changes were abolished by acute inhibition of the dorsomedial nucleus of the hypothalamus (DMH). These cardiovascular changes were also prevented by chronic sub-cutaneous osmotic infusion of losartan, an angiotensin II type 1 receptor (AT1) receptor antagonist, started immediately after social defeat. In conclusion, the results show that greater vulnerability to stress consequences following a traumatic event is associated with an elevated LF/HF ratio, a persistent high blood pressure and a low BRS, all due to an AT1 receptor activation.

2-arachidonylglycerol interacts with nitric oxide in the dorsomedial hypothalamus to increase food intake and body weight in young male rats

The dorsomedial nucleus of the hypothalamus (DMH) is an important appetite regulatory center in the brain. In young rats, neural communication in the DMH is modulated by two interacting signals: endocannabinoids (eCBs) and nitric oxide (NO), both of which are known to modulate appetite. It remains unknown, however, whether eCBs and NO interact in the DMH to regulate food intake and body weight in young rats. We developed stereotaxic coordinates for the DMH in young, male Sprague-Dawley rats and conducted surgeries to implant bilateral guide cannulas for microinjection of vehicle, eCBs [2-arachidonylglycerol (2-AG) or anandamide]; NO (via the precursor l-arginine), or a combination of the two, with and without prior subcutaneous injections of drugs to block cannabinoid receptors or NO synthesis. Food intake and body weight of animals were measured two hours following the injection and brains were subsequently removed and sliced to verify placement of the cannulas relative to the DMH. Here we show that 2-AG, when administered in combination with l-arginine, significantly increased food intake and body weight, an effect that required type I cannabinoid receptors and NO synthesis. 2-AG and l-arginine had no effect on food intake or body weight when administered into the DMH independently. Anandamide also failed to affect these parameters when administered alone or with l-arginine. Together, these data suggest that 2-AG and NO interact in the DMH to increase food intake in young male rats and provide insight into a possible mechanism by which 2-AG increases appetite.

Temporal and regional onset of leptin resistance in diet-induced obese mice.

In common forms of obesity, leptin fails to convey its regulatory effect. This so called "leptin resistance" is not well understood, and solving this puzzle is a key to understanding how obesity develops. In the present study, we investigated the temporal and regional onset of leptin resistance in response to a diet enriched with long-chain saturated fatty acids (high-fat diet; HFD) in mice. Mice were exposed to either a low-fat diet (LFD) or a HFD for 4hours, 24hours, 10days and 28days. Mice in each group received an i.p. injection of either phosphate-buffered saline or leptin and the number of phosphorylated signal transducer and activator of transcription-3 (pSTAT3) immunoreactive (-IR) cells in the arcuate nucleus (ARC), ventromedial nucleus of the hypothalamus (VMH) and dorsomedial nucleus of the hypothalamus (DMH) was analysed 30 or 120minutes after treatment. In the ARC, as soon as 24hours of HFD, the molecular leptin response was reduced by 40% (P≤.01). Compared to at 24hours, after 10days, the number of leptin-induced pSTAT3-IR cells was elevated after 120minutes, suggesting a sustained response and a partial return of leptin sensitivity. After 28days, leptin failed to induce the number of pSTAT3-IR over control levels, suggesting a markedly reduced sensitivity to leptin. In the VMH after 24hours, we observed a 50% reduction in leptin-induced pSTAT-3-IR cells, followed by a further decline after 10days. However, after 28days, there was a significant increase in pSTAT-3-IR cells (P≤.05), indicating partial recovery of leptin sensitivity. By contrast to these two regions, in the DMH, no loss of leptin sensitivity was observed at any time-point. These findings demonstrate that a loss of sensitivity to leptin occurs rapidly after exposure to HFD in the ARC and VMH but not the DMH. However, there appears to be a biphasic pattern of leptin responsiveness, with a partial return of leptin sensitivity occurring after 10days in the arcuate nucleus, and after 28days in the VMH. By 28days, the response to leptin in the arcuate nucleus was completely lost. These findings suggest that the molecular responses to leptin are altered after high-fat feeding in a time- and region-specific manner.

Endocannabinoid signaling in hypothalamic circuits regulates arousal from general anesthesia in mice.

Consciousness can be defined by two major attributes: awareness of environment and self, and arousal, which reflects the level of awareness. The return of arousal after general anesthesia presents an experimental tool for probing the neural mechanisms that control consciousness. Here we have identified that systemic or intracerebral injection of the cannabinoid CB1 receptor (CB1R) antagonist AM281 into the dorsomedial nucleus of the hypothalamus (DMH) - but not the adjacent perifornical area (Pef) or the ventrolateral preoptic nucleus of the hypothalamus (VLPO) - accelerates arousal in mice recovering from general anesthesia. Anesthetics selectively activated endocannabinoid (eCB) signaling at DMH glutamatergic but not GABAergic synapses, leading to suppression of both glutamatergic DMH-Pef and GABAergic DMH-VLPO projections. Deletion of CB1R from widespread cerebral cortical or prefrontal cortical (PFC) glutamatergic neurons, including those innervating the DMH, mimicked the arousal-accelerating effects of AM281. In contrast, CB1R deletion from brain GABAergic neurons or hypothalamic glutamatergic neurons did not affect recovery time from anesthesia. Inactivation of PFC-DMH, DMH-VLPO, or DMH-Pef projections blocked AM281-accelerated arousal, whereas activation of these projections mimicked the effects of AM281. We propose that decreased eCB signaling at glutamatergic terminals of the PFC-DMH projection accelerates arousal from general anesthesia through enhancement of the excitatory DMH-Pef projection, the inhibitory DMH-VLPO projection, or both.

Gsα deficiency in the dorsomedial hypothalamus underlies obesity associated with Gsα mutations.

Gsα, encoded by Gnas, mediates hormone and neurotransmitter receptor-stimulated cAMP generation. Heterozygous Gsα-inactivating mutations lead to obesity in Albright hereditary osteodystrophy (AHO) patients, but only when the mutations occur on the maternal allele. This parent-of-origin effect is due to Gsα imprinting in the CNS, although the relevant CNS regions are unknown. We have now shown that mice with a Gnas gene deletion disrupting Gsα expression on the maternal allele, but not the paternal allele, in the dorsomedial nucleus of the hypothalamus (DMH) developed obesity and reduced energy expenditure without hyperphagia. Although maternal Gnas deletion impaired activation of brown adipose tissue (BAT) in mice, their responses to cold environment remained intact. Similar findings were observed in mice with DMH-specific deficiency of melanocortin MC4R receptors, which are known to activate Gsα. Our results show that Gsα imprinting in the DMH underlies the parent-of-origin metabolic phenotype that results from Gsα mutations and that DMH MC4R/Gsα signaling is important for regulation of energy expenditure and BAT activation, but not the metabolic response to cold.

Molecular characterization of GPR50 gene and study of its comparative genetic variability in sheep breeds adapted to different thermo-contrasting climatic regimens.

GPR50, formerly known as a melatonin-related receptor, is one of the three subtypes of melatonin receptor subfamily, together with MTNR1A and MTNR1B. GPR50, despite its high identity with the melatonin receptor family, does not bind melatonin and is considered to be an ortholog of MTNR1C in mammals. GPR50-expressing cells have been found in the dorsomedial nucleus of the hypothalamus, the periventricular nucleus, and the median eminence. Genetic and functional evidence have been recently investigated linking GPR50 to adaptive thermogenesis and torpor, but still, it is an orphan receptor and is yet to be studied conclusively. The aims of the study were to characterize the GPR50 gene of sheep and to study the sequence variability of the gene in Indian sheep breeds of two different thermo-varied agroclimatic conditions. Genomic DNA isolation was done and a 791-bp sequence was amplified using self-designed primers and SNP profiling done out of samples of all the breeds to study the relative frequency of SNPs in each of the breed. Five important non-synonymous mutations were observed in the various breeds studied. T698G, G1097A, G1270A, G1318A, and C1334G lead to the following substitution: valine by glycine, arginine by glutamine, threonine by alanine, isoleucine by valine, and serine by cytosine, respectively. Two synonymous mutations (T663G and C888T) were also observed in some of the studied breeds. G1270A and C888T were the most prevalent SNPs observed in nearly all of the breeds. C888T SNPs were observed in higher prevalence in Chokla, Marwari, and Magra in comparison to Gaddi and Bharat Merino. A PolyPhen-2 analysis, which is used to assess the potential damaging nature of an SNP, revealed that mutation T698G and G1270A were benign while G1097A, G1318A, and C1334G were damaging with a score of 0.987, 0.993, and 0.739, respectively. A 3-D homology model of the protein was prepared using c4zwjA (UniProt sequence ID) as a template using the online version of Phyre2 protein modeling software. The structure demonstrated closed similarity with other G-coupled receptor and it had a 45% α-helical content. G1270A and C888T may be taken up for SNP correlation in a larger population study for their association with heat stress protection.

Role of the Suprachiasmatic and Arcuate Nuclei in Diurnal Temperature Regulation in the Rat.

When circadian and metabolic systems are not well synchronized, individuals may develop metabolic diseases. The underlying mechanisms are unknown. Here, we demonstrate that the balance between the releases of neuropeptides derived from the biological clock and from a metabolic sensory organ as the arcuate nucleus, are essential for an adequate temperature control. These observations show that brain areas involved in circadian and metabolic functions of the body need to interact to produce a coherent arrangement of physiological processes associated with temperature control.

Galanin-Mediated Behavioural Hyperalgesia from the Dorsomedial Nucleus of the Hypothalamus Involves Two Independent Descending Pronociceptive Pathways.

Activation of the dorsomedial nucleus of the hypothalamus (DMH) by galanin (GAL) induces behavioural hyperalgesia. Since DMH neurones do not project directly to the spinal cord, we hypothesized that the medullary dorsal reticular nucleus (DRt), a pronociceptive region projecting to the spinal dorsal horn (SDH) and/or the serotoninergic raphe-spinal pathway acting on the spinal 5-HT3 receptor (5HT3R) could relay descending nociceptive facilitation induced by GAL in the DMH. Heat-evoked paw-withdrawal latency (PWL) and activity of SDH neurones were assessed in monoarthritic (ARTH) and control (SHAM) animals after pharmacological manipulations of the DMH, DRt and spinal cord. The results showed that GAL in the DMH and glutamate in the DRt lead to behavioural hyperalgesia in both SHAM and ARTH animals, which is accompanied particularly by an increase in heat-evoked responses of wide-dynamic range neurons, a group of nociceptive SDH neurones. Facilitation of pain behaviour induced by GAL in the DMH was reversed by lidocaine in the DRt and by ondansetron, a 5HT3R antagonist, in the spinal cord. However, the hyperalgesia induced by glutamate in the DRt was not blocked by spinal ondansetron. In addition, in ARTH but not SHAM animals PWL was increased after lidocaine in the DRt and ondansetron in the spinal cord. Our data demonstrate that GAL in the DMH activates two independent descending facilitatory pathways: (i) one relays in the DRt and (ii) the other one involves 5-HT neurones acting on spinal 5HT3Rs. In experimental ARTH, the tonic pain-facilitatory action is increased in both of these descending pathways.

Prolonged hyperglycemia & hyperinsulinemia increases BDNF mRNA expression in the posterior ventromedial hypothalamus and the dorsomedial hypothalamus of fed female rats.

Brain-derived neurotrophic factor (BDNF) plays a key role in neuronal development, synaptic plasticity, and the central control of energy homeostasis. Peripheral metabolic signals such as leptin and glucose regulate hypothalamic BDNF gene expression. However, the effects of long-term hyperglycemia and/or hyperinsulinemia on BDNF mRNA levels in the hypothalamus and other brain regions where BDNF regulates physiological functions have not been investigated. Therefore, using in situ hybridization we examined whether high glucose, high insulin, or both affected BDNF gene expression in vivo. Ovariectomized, estrogen-replaced adult rats were fitted with indwelling jugular catheters and infused for 48 h with: saline (control), glucose (hyperglycemia-hyperinsulinemia), glucose with insulin (hyperinsulinemia only), diazoxide (Dzx) (control), or glucose with Dzx (hyperglycemia only). Glucose infusion (Hyperglycemia and hyperinsulinemia) significantly increased BDNF mRNA expression in the posterior ventromedial nucleus of the hypothalamus (pVMH) and in the dorsomedial nucleus of the hypothalamus (DMH). Unexpectedly, infusion of the KATP channel opener Dzx also increased BDNF mRNA expression in the pVMH and DMH. In contrast, no significant changes in BDNF mRNA expression were observed in the groups that were hyperinsulinemic only or hyperglycemic only. BDNF mRNA expression did not differ as a function of treatment in the anterior VMH, paraventricular nucleus of the hypothalamus, the hippocampus, or the amygdala. Hyperglycemia with and without hyperinsulinemia decreased BDNF mRNA levels in the pituitary. Plasma BDNF concentrations were not changed by any of the treatments. Our results suggest that hyperinsulinemia alone does not affect BDNF mRNA expression in the hypothalamus, hippocampus, or pituitary. Our study is the first to distinguish that within the hypothalamus, prolonged high glucose levels in non-fasted rats regulates BDNF gene expression in a brain nuclei-specific fashion.

A role of supraspinal galanin in behavioural hyperalgesia in the rat.

IntroductionIn chronic pain disorders, galanin (GAL) is able to either facilitate or inhibit nociception in the spinal cord but the contribution of supraspinal galanin to pain signalling is mostly unknown. The dorsomedial nucleus of the hypothalamus (DMH) is rich in galanin receptors (GALR) and is involved in behavioural hyperalgesia. In this study, we evaluated the contribution of supraspinal GAL to behavioural hyperalgesia in experimental monoarthritis.MethodsIn Wistar-Han males with a four week kaolin/carrageenan-induced monoarthritis (ARTH), paw-withdrawal latency (PWL) was assessed before and after DMH administration of exogenous GAL, a non-specific GALR antagonist (M40), a specific GALR1 agonist (M617) and a specific GALR2 antagonist (M871). Additionally, the analysis of c-Fos expression after GAL injection in the DMH was used to investigate the potential involvement of brainstem pain control centres. Finally, electrophysiological recordings were performed to evaluate whether pronociceptive On- or antinociceptive Off-like cells in the rostral ventromedial medulla (RVM) relay the effect of GAL.ResultsExogenous GAL in the DMH decreased PWL in ARTH and SHAM animals, an effect that was mimicked by a GALR1 agonist (M617). In SHAM animals, an unselective GALR antagonist (M40) increased PWL, while a GALR2 antagonist (M871) decreased PWL. M40 or M871 failed to influence PWL in ARTH animals. Exogenous GAL increased c-Fos expression in the RVM and dorsal raphe nucleus (DRN), with effects being more prominent in SHAM than ARTH animals. Exogenous GAL failed to influence activity of RVM On- or Off-like cells of SHAM and ARTH animals.ConclusionsOverall, exogenous GAL in the DMH had a pronociceptive effect that is mediated by GALR1 in healthy and arthritic animals and is associated with alterations of c-Fos expression in RVM and DRN that are serotonergic brainstem nuclei known to be involved in the regulation of pain.