DMHPpp1r17 neurons regulate aging and lifespan in mice through hypothalamic-adipose inter-tissue communication.

Recent studies have shown that the hypothalamus functions as a control center of aging in mammals that counteracts age-associated physiological decline through inter-tissue communications. We have identified a key neuronal subpopulation in the dorsomedial hypothalamus (DMH), marked by Ppp1r17 expression (DMHPpp1r17 neurons), that regulates aging and longevity in mice. DMHPpp1r17 neurons regulate physical activity and WAT function, including the secretion of extracellular nicotinamide phosphoribosyltransferase (eNAMPT), through sympathetic nervous stimulation. Within DMHPpp1r17 neurons, the phosphorylation and subsequent nuclear-cytoplasmic translocation of Ppp1r17, regulated by cGMP-dependent protein kinase G (PKG; Prkg1), affect gene expression regulating synaptic function, causing synaptic transmission dysfunction and impaired WAT function. Both DMH-specific Prkg1 knockdown, which suppresses age-associated Ppp1r17 translocation, and the chemogenetic activation of DMHPpp1r17 neurons significantly ameliorate age-associated dysfunction in WAT, increase physical activity, and extend lifespan. Thus, these findings clearly demonstrate the importance of the inter-tissue communication between the hypothalamus and WAT in mammalian aging and longevity control.

Neurons in the Dorsomedial Hypothalamus Promote, Prolong, and Deepen Torpor in the Mouse.

Torpor is a naturally occurring, hypometabolic, hypothermic state engaged by a wide range of animals in response to imbalance between the supply and demand for nutrients. Recent work has identified some of the key neuronal populations involved in daily torpor induction in mice, in particular, projections from the preoptic area of the hypothalamus to the dorsomedial hypothalamus (DMH). The DMH plays a role in thermoregulation, control of energy expenditure, and circadian rhythms, making it well positioned to contribute to the expression of torpor. We used activity-dependent genetic TRAPing techniques to target DMH neurons that were active during natural torpor bouts in female mice. Chemogenetic reactivation of torpor-TRAPed DMH neurons in calorie-restricted mice promoted torpor, resulting in longer and deeper torpor bouts. Chemogenetic inhibition of torpor-TRAPed DMH neurons did not block torpor entry, suggesting a modulatory role for the DMH in the control of torpor. This work adds to the evidence that the preoptic area of the hypothalamus and the DMH form part of a circuit within the mouse hypothalamus that controls entry into daily torpor.SIGNIFICANCE STATEMENT Daily heterotherms, such as mice, use torpor to cope with environments in which the supply of metabolic fuel is not sufficient for the maintenance of normothermia. Daily torpor involves reductions in body temperature, as well as active suppression of heart rate and metabolism. How the CNS controls this profound deviation from normal homeostasis is not known, but a projection from the preoptic area to the dorsomedial hypothalamus has recently been implicated. We demonstrate that the dorsomedial hypothalamus contains neurons that are active during torpor. Activity in these neurons promotes torpor entry and maintenance, but their activation alone does not appear to be sufficient for torpor entry.

Role of Dorsomedial Hypothalamus GABAergic Neurons in Sleep-Wake States in Response to Changes in Ambient Temperature in Mice.

Good sleep quality is essential for maintaining the body's attention during wakefulness, which is easily affected by external factors such as an ambient temperature. However, the mechanism by which an ambient temperature influences sleep-wake behaviors remains unclear. The dorsomedial hypothalamus (DMH) has been reported to be involved in thermoregulation. It also receives projection from the preoptic area, which is an important region for sleep and energy homeostasis and the suprachiasmatic nucleus-a main control area of the clock rhythm. Therefore, we hypothesized that the DMH plays an important role in the regulation of sleep related to ambient temperatures. In this study, we found that cold exposure (24/20/16/12 °C) increased wakefulness and decreased non-rapid eye movement (NREM) sleep, while warm exposure (32/36/40/44 °C) increased NREM sleep and decreased wakefulness compared to 28 °C conditions in wild-type mice. Then, using non-specific and specific apoptosis, we found that lesions of whole DMH neurons and DMH γ-aminobutyric acid (GABA)-ergic neurons induced by caspase-3 virus aggravated the fluctuation of core body temperature after warm exposure and attenuated the change in sleep-wake behaviors during cold and warm exposure. However, chemogenetic activation or inhibition of DMH GABAergic neurons did not affect the sleep-wake cycle. Collectively, our findings reveal an essential role of DMH GABAergic neurons in the regulation of sleep-wake behaviors elicited by a change in ambient temperature.

Neuroanatomical organization and functional roles of PVN MC4R pathways in physiological and behavioral regulations.

OBJECTIVE: The paraventricular nucleus of hypothalamus (PVN), an integrative center in the brain, orchestrates a wide range of physiological and behavioral responses. While the PVN melanocortin 4 receptor (MC4R) signaling (PVNMC4R+) is involved in feeding regulation, the neuroanatomical organization of PVNMC4R+ connectivity and its role in other physiological regulations are incompletely understood. Here we aimed to better characterize the input-output organization of PVNMC4R+ neurons and test their physiological functions beyond feeding. METHODS: Using a combination of viral tools, we mapped PVNMC4R+ circuits and tested the effects of chemogenetic activation of PVNMC4R+ neurons on thermoregulation, cardiovascular control, and other behavioral responses beyond feeding. RESULTS: We found that PVNMC4R+ neurons innervate many different brain regions that are known to be important not only for feeding but also for neuroendocrine and autonomic control of thermoregulation and cardiovascular function, including but not limited to the preoptic area, median eminence, parabrachial nucleus, pre-locus coeruleus, nucleus of solitary tract, ventrolateral medulla, and thoracic spinal cord. Contrary to these broad efferent projections, PVNMC4R+ neurons receive monosynaptic inputs mainly from other hypothalamic nuclei (preoptic area, arcuate and dorsomedial hypothalamic nuclei, supraoptic nucleus, and premammillary nucleus), the circumventricular organs (subfornical organ and vascular organ of lamina terminalis), the bed nucleus of stria terminalis, and the parabrachial nucleus. Consistent with their broad efferent projections, chemogenetic activation of PVNMC4R+ neurons not only suppressed feeding but also led to an apparent increase in heart rate, blood pressure, and brown adipose tissue temperature. These physiological changes accompanied acute transient hyperactivity followed by hypoactivity and resting-like behavior. CONCLUSIONS: Our results elucidate the neuroanatomical organization of PVNMC4R+ circuits and shed new light on the roles of PVNMC4R+ pathways in autonomic control of thermoregulation, cardiovascular function, and biphasic behavioral activation.

Refeeding activates neurons in the dorsomedial hypothalamus to inhibit food intake and promote positive valence.

OBJECTIVE: The regulation of food intake is a major research area in the study of obesity, which plays a key role in the development of metabolic syndrome. Gene targeting studies have clarified the roles of hypothalamic neurons in feeding behavior, but the deletion of a gene has a long-term effect on neurophysiology. Our understanding of short-term changes such as appetite under physiological conditions is therefore still limited. METHODS: Targeted recombination in active populations (TRAP) is a newly developed method for labeling active neurons by using tamoxifen-inducible Cre recombination controlled by the promoter of activity-regulated cytoskeleton-associated protein (Arc/Arg3.1), a member of immediate early genes. Transgenic mice for TRAP were fasted overnight, re-fed with normal diet, and injected with 4-hydroxytamoxifen 1 h after the refeeding to label the active neurons. The role of labeled neurons was examined by expressing excitatory or inhibitory designer receptors exclusively activated by designer drugs (DREADDs). The labeled neurons were extracted and RNA sequencing was performed to identify genes that are specifically expressed in these neurons. RESULTS: Fasting-refeeding activated and labeled neurons in the compact part of the dorsomedial hypothalamus (DMH) that project to the paraventricular hypothalamic nucleus. Chemogenetic activation of the labeled DMH neurons decreased food intake and developed place preference, an indicator of positive valence. Chemogenetic activation or inhibition of these neurons had no influence on the whole-body glucose metabolism. The labeled DMH neurons expressed prodynorphin (pdyn), gastrin-releasing peptide (GRP), cholecystokinin (CCK), and thyrotropin-releasing hormone receptor (Trhr) genes. CONCLUSIONS: We identified a novel cell type of DMH neurons that can inhibit food intake and promote feeding-induced positive valence. Our study provides insight into the role of DMH and its molecular mechanism in the regulation of appetite and emotion.

Cholecystokinin acts in the dorsomedial hypothalamus of young male rats to suppress appetite in a nitric oxide-dependent manner.

Cholecystokinin (CCK) is an appetite-suppressing hormone that acts in the dorsomedial hypothalamus (DMH) in adult rats to suppress food intake. It remains unknown, however, whether CCK has the same affect in young animals, despite the rising prevalence of childhood obesity and drastic need for research in this area. At the synaptic level, CCK has been shown to inhibit putative orexigenic DMH neurons in young male rats by increasing GABA release onto these neurons via a CCK2 receptor and nitric oxide-dependent pathway. Whether this pathway leads to appetite suppression in young rats is not known. We therefore investigated whether intra-DMH administration of CCK, with or without inhibitors of the CCK2 receptor and nitric oxide signaling pathways, affects food intake in young, male, fasted Sprague-Dawley rats. We implanted bilateral guide cannulas into the DMH and allowed animals to recover from the surgery. Animals were then fasted for 24 h, following which they received intra-DMH microinjections of vehicle, CCK-8S, or CCK-8S combined with either LY-225910 (CCK2 receptor antagonist), L-NAME (a nitric oxide synthase inhibitor), or ODQ (a soluble guanylate cyclase inhibitor) and were given access to regular chow. Following a two hour refeeding period during which food intake, latency to feed, and body weight were measured, brains were subsequently removed to confirm cannula placement in the DMH. The effect of CCK on these parameters in rats given a high fat diet were also measured. Here we show that intra-DMH administration of CCK suppresses food intake and body weight in young rats. This effect requires activation of CCK2 receptors and nitric oxide signaling. Finally, CCK has no effect on consumption of a high fat diet when administered into the DMH. Overall, these findings demonstrate a potential pathway through which CCK might suppress appetite in young rats.

Changes of discharge properties of neurons from dorsomedial hypothalamic nuclei during aging in rats.

The hypothalamus is a vital brain center that is participated in the integration of the endocrine and nervous systems and control of the homeostasis and aging. Spontaneous firing activity from single neurons of the dorsomedial hypothalamic nucleus (DMN) was studied extracellularly in vivo in urethane-anaesthetized rats. The discharge patterns of the majority of DMN neurons were irregular, including periods of relatively stable activity interrupted by pauses. Based on the features of interval interspike histogram, we have selected neurons with an irregular arrhythmic activity (50% in young, 46% in adult and 44% in aged rats), with a constant rhythmic activity (18% of neurons in young, 19% in adult and 23% in aged rats), with a wide interspike interval distribution (22% in young, 26% in adult and 25% in aged rats) and cells with bursts of two or three spikes (10% in young, 9% in adult and 8% in aged rats). The firing rate of DMN neurons was 2.5 ± 0.12 Hz in young and 2.4 ± 0.21 Hz in adult rats and significantly decreased to 1.8 ± 0.17 Hz in aged rats.

Neuropeptide Y suppresses thermogenic and cardiovascular sympathetic nerve activity via Y1 receptors in the paraventricular nucleus and dorsomedial hypothalamus.

In hungry animals, neuropeptide Y (NPY) neurones in the arcuate nucleus (ArcN) are activated to suppress energy expenditure, in part by decreasing brown adipose tissue sympathetic nerve activity (BAT SNA); however, the NPY receptor subtype and brain neurocircuitry are unclear. In the present study, we investigated the inhibition of BAT SNA by exogenous and endogenous NPY via binding to Y1 receptors (NPY1R) in the hypothalamic paraventricular nucleus (PVN) and dorsomedial hypothalamus (DMH), in anaesthetised male rats. Downstream projections of PVN/DMH NPY1R-expressing neurones were identified using male Npy1r-cre mice and localised unilateral DMH or PVN injections of an adeno-associated virus, which allows for the cre-dependent expression of a fluorescent protein (mCherry) in the cell bodies, axon fibres and nerve terminals of NPY1R-containing neurones. Nanoinjections of NPY into the DMH of cooled rats decreased BAT SNA, as well as mean arterial pressure (MAP) and heart rate (HR), and these responses were reversed by subsequent injection of the selective NPY1R antagonist, BIBO3304. In warmed rats, with little to no BAT SNA, bilateral nanoinjections of BIBO3304 into the DMH or PVN increased BAT SNA, MAP and HR. DMH NPY1R-expressing neurones projected heavily to the raphe pallidus (RPa), which houses BAT presympathetic neurones, as well as the PVN. In anaesthetised mice, DMH BIBO3304 increased splanchnic SNA, MAP and HR, all of which were reversed by nonselective blockade of the PVN with muscimol, suggesting that DMH-to-PVN connections are involved in this DMH BIBO3304 disinhibition. PVN Y1R expressing neurones also projected to the RPa, as well as to the nucleus tractus solitarius. We conclude that NPY tonically released in the DMH and PVN suppresses BAT SNA, MAP and HR via Y1R. Downstream neuropathways for BAT SNA may utilise direct projections to the RPa. Release of tonic NPY inhibition of BAT SNA may contribute to feeding- and diet-induced thermogenesis.

Tachycardia evoked from insular stroke in rats is dependent on glutamatergic neurotransmission in the dorsomedial hypothalamus.

BACKGROUND AND PURPOSE: Damage to the insula results in cardiovascular complications. In rats, activation of N-methyl-d-aspartate receptors (NMDARs) in the intermediate region of the posterior insular cortex (iIC) results in sympathoexcitation, tachycardia and arterial pressure increases. Similarly, focal experimental hemorrhage at the iIC results in a marked sympathetic-mediated increase in baseline heart rate. The dorsomedial hypothalamic region (DMH) is critical for the integration of sympathetic-mediated tachycardic responses. Here, whether responses evoked from the iIC are dependent on a synaptic relay in the DMH was evaluated. METHODS: Wistar rats were prepared for injections into the iIC and DMH. Anatomical (tracing combined with immunofluorescence) and functional experiments (cardiovascular and sympathetic recordings) were performed. RESULTS: The iIC sends dense projections to the DMH. Approximately 50% of iIC neurons projecting to the DMH express NMDARs, NR1 subunit. Blockade of glutamatergic receptors in the DMH abolishes the cardiovascular and autonomic responses evoked by the activation of NMDARs in the iIC (change in mean arterial pressure 7 ± 1 vs. 1 ± 1 mmHg after DMH blockade; change in heart rate 28 ± 3 vs. 0 ± 3 bpm after DMH blockade; change in renal sympathetic nerve activity 23% ± 1% vs. -1% ± 4% after DMH blockade). Experimental hemorrhage at the iIC resulted in a marked tachycardia (change 89 ± 14 bpm) that was attenuated by 65% ± 5% (p = 0.0009) after glutamatergic blockade at the DMH. CONCLUSIONS: The iIC-induced tachycardia is largely dependent upon a glutamatergic relay in the DMH. Our study reveals the presence of an excitatory glutamatergic pathway from the iIC to the DMH that may be involved in the cardiovascular alterations observed after insular stroke.

Restriction of food intake by PPP1R17-expressing neurons in the DMH.

Leptin-deficient ob/ob mice eat voraciously, and their food intake is markedly reduced by leptin treatment. In order to identify potentially novel sites of leptin action, we used PhosphoTRAP to molecularly profile leptin-responsive neurons in the hypothalamus and brainstem. In addition to identifying several known leptin responsive populations, we found that neurons in the dorsomedial hypothalamus (DMH) of ob/ob mice expressing protein phosphatase 1 regulatory subunit 17 (PPP1R17) constitutively express cFos and that this is suppressed by leptin treatment. Because ob mice are hyperphagic, we hypothesized that activating PPP1R17 neurons would increase food intake. However, chemogenetic activation of PPP1R17 neurons decreased food intake and body weight of ob/ob mice while inhibition of PPP1R17 neurons increased them. Similarly, in a scheduled feeding protocol that elicits increased consumption, mice also ate more when PPP1R17 neurons were inhibited and ate less when they were activated. Finally, we found that pair-feeding of ob mice reduced cFos expression to a similar extent as leptin and that reducing the amount of food available during scheduled feeding in DMHPpp1r17 neurons also decreased cFos in DMHPpp1r17 neurons. Finally, these neurons do not express the leptin receptor, suggesting that the effect of leptin on these neurons is indirect and secondary to reduced food intake. In aggregate, these results show that PPP1R17 neurons in the DMH are activated by increased food intake and in turn restrict intake to limit overconsumption, suggesting that they function to constrain binges of eating.

Orexin-A directly depolarizes dorsomedial hypothalamic neurons, including those innervating the rostral ventrolateral medulla.

The dorsomedial hypothalamus (DMH) receives dense orexinergic innervation. Intra-DMH application of orexins increases arterial pressure and heart rate in rats. We studied the effects of orexin-A on DMH neurons, including those innervating the medullary cardiovascular center, the rostral ventrolateral medulla (RVLM), by using whole-cell recordings in brain slices. In the presence of tetrodotoxin, orexin-A (30-1000 nM) depolarized 56% of DMH neurons (EC50 82.4 ± 4.4 nM). Under voltage-clamp recording, orexin-A (300 nM) induced three types of responses characterized by different current-voltage relationships, namely unchanged, increased, and decreased slope conductance in 68%, 14%, and 18% of orexin-A-responsive neurons, respectively. The reversal potential of the decreased-conductance response was near the equilibrium potential of K+ and became more positive in a high-K+ solution, suggesting that K+ conductance blockade is the underlying mechanism. In a low-Na+ solution, unchanged-, increased-, and decreased-conductance responses were observed in 56%, 11%, and 33% of orexin-A-responsive neurons, respectively, implying that a non-selective cation current (NSCC) underlies orexin-A-induced responses in a small population of DMH neurons. KBR-7943 (70 µM), an inhibitor of Na+-Ca2+ exchanger (NCX), suppressed orexin-A-induced depolarization in 7 of 10 neurons. In the presence of KBR-7943, the majority of orexin-A-responsive neurons exhibited decreased-conductance responses. These findings suggest that NCX activation may underlie orexin-A-induced depolarization in the majority of orexin-responsive DMH neurons. Of 19 RVLM-projecting DMH neurons identified by retrograde labeling, 17 (90%) were orexin-A responsive. In conclusion, orexin-A directly excited over half of DMH neurons, including those innervating the RVLM, through decreasing K+ conductance, activating NCX, and/or increasing NSCC.

Leptin receptor neurons in the dorsomedial hypothalamus regulate diurnal patterns of feeding, locomotion, and metabolism.

The brain plays an essential role in driving daily rhythms of behavior and metabolism in harmony with environmental light-dark cycles. Within the brain, the dorsomedial hypothalamic nucleus (DMH) has been implicated in the integrative circadian control of feeding and energy homeostasis, but the underlying cell types are unknown. Here, we identify a role for DMH leptin receptor-expressing (DMHLepR) neurons in this integrative control. Using a viral approach, we show that silencing neurotransmission in DMHLepR neurons in adult mice not only increases body weight and adiposity but also phase-advances diurnal rhythms of feeding and metabolism into the light cycle and abolishes the normal increase in dark-cycle locomotor activity characteristic of nocturnal rodents. Finally, DMHLepR-silenced mice fail to entrain to a restrictive change in food availability. Together, these findings identify DMHLepR neurons as critical determinants of the daily time of feeding and associated metabolic rhythms.

Expression of calbindin and calretinin in the dorsomedial and ventromedial hypothalamic nuclei during aging.

The hypothalamus is involved in the regulation of rhythms, autonomic, endocrine, and behavioral functions and may also participate in aging development and control. The aim of this work was to study the expression of calbindin (CB) and calretinin (CR) in the ventromedial (VMH) and dorsomedial (DMH) hypothalamic nuclei in young and old rats of both sexes by immunohistochemistry and western blotting. In young animals, the largest number of CB-immunoreactive (IR) neurons was detected in the ventral part of DMH (DMHv) and smaller percentage was found in its dorsal part (DMHd), in the dorsomedial part of the VMH (VMHdm) and in the ventrolateral part of the VMH (VMHvl). In aged animals, the percentage of CB-IR neurons significantly decreased in all studied nuclei, including DMHv, DMHd, VMHdm and VMHvl. CR-IR neurons were found in moderate number in the DMHv, DMHd, VMHdm and VMHvl of young rats. In aged rats, the percentage of CR-IR neurons significantly increased in the DMHv, DMHd, VMHdm and VMHvl. Less than one third of IR neurons colocalized CB and CR in young and aged rats. The expression of CB significantly decreased, and the expression of CR significantly increased in the DMH and VMH during aging by western blot analysis. Thus, there are opposite changes of the calcium-binding proteins expression in the hypothalamic nuclei involved in the metabolic and sexual regulation during aging.

The anorexigenic effect of vasoactive intestinal polypeptide in Japanese quail is associated with molecular changes in the arcuate and dorsomedial hypothalamic nuclei.

Vasoactive intestinal polypeptide (VIP) is involved in gastric smooth muscle relaxation, vasodilation, and gastric secretions. It is also associated with appetite regulation, eliciting an anorexigenic response in mammals, birds, and fish; however, the molecular mechanism mediating this response is not well understood. The aim of the present study was thus to investigate hypothalamic mechanisms mediating VIP-induced satiety in 7-d old Japanese quail. In experiment 1, chicks that received intracerebroventricular (ICV) injection of VIP had reduced food intake for up to 180 min after injection and reduced water intake for 90 min. In experiment 2, VIP-treated chicks that were food restricted did not reduce water intake. In experiment 3, there was increased c-Fos immunoreactivity in the arcuate (ARC) and dorsomedial (DMN) nuclei of the hypothalamus in VIP-injected quail. In experiment 4, ICV VIP was associated with decreased neuropeptide Y mRNA in the ARC and DMN and an increase in corticotropin releasing factor mRNA in the DMN. In experiment 5, VIP-treated chicks displayed fewer feed pecks and locomotor behaviors. These results demonstrate that central VIP causes anorexigenic effects that are likely associated with reductions in orexigenic tone involving the ARC and DMN.

GLP-2 decreases food intake in the dorsomedial hypothalamic nucleus (DMH) through Exendin (9-39) in male Sprague-Dawley (SD) rats.

Glucagon-like peptide 2 (GLP-2), a member of Glucagon peptide family involved in regulating energy metabolism, can be produced and secreted by preproglucagonergic (PPG) neurons in the brain. GLP-2 reduces food intake but at which brain sites GLP-2 exerts its feeding-suppress effects are still unclear. In this study, we used the stereological microinjection technique and behavioral test to examine the functions of locally delivered GLP-2 into DMH on feeding behavior. We compared effects of different concentration of GLP-2 on the food intake behavior in free-feeding rats and fasted-refeeding rats. We found that GLP-2 inhibited food intake in fasted rats after a short-term intervention in a dose-dependent manner. Importantly, the effects of locally delivered GLP-2 can be blocked by specific GLP-1 receptor antagonist Exendin(9-39), but not the melanocortin-4 receptor antagonist SHU9119, indicating the involvement of specificity of GLP-2 signaling in regulating the feeding behavior. Taken together, our data revealed that GLP-2 peptide pharmacologically inhibited food intake in DMH and this effect could be blocked functionally by Exendin(9-39).

Median preoptic area neurons are required for the cooling and febrile activations of brown adipose tissue thermogenesis in rat.

Within the central neural circuitry for thermoregulation, the balance between excitatory and inhibitory inputs to the dorsomedial hypothalamus (DMH) determines the level of activation of brown adipose tissue (BAT) thermogenesis. We employed neuroanatomical and in vivo electrophysiological techniques to identify a source of excitation to thermogenesis-promoting neurons in the DMH that is required for cold defense and fever. Inhibition of median preoptic area (MnPO) neurons blocked the BAT thermogenic responses during both PGE2-induced fever and cold exposure. Disinhibition or direct activation of MnPO neurons induced a BAT thermogenic response in warm rats. Blockade of ionotropic glutamate receptors in the DMH, or brain transection rostral to DMH, blocked cold-evoked or NMDA in MnPO-evoked BAT thermogenesis. RNAscope technique identified a glutamatergic population of MnPO neurons that projects to the DMH and expresses c-Fos following cold exposure. These discoveries relative to the glutamatergic drive to BAT sympathoexcitatory neurons in DMH augment our understanding of the central thermoregulatory circuitry in non-torpid mammals. Our data will contribute to the development of novel therapeutic approaches to induce therapeutic hypothermia for treating drug-resistant fever, and for improving glucose and energy homeostasis.

Sirtuin 1 Expression in the Rat Ventromedial and Dorsomedial Hypothalamic Nuclei during Ageing.

We studied the expression of sirtuin 1 (Sirt1) in the dorsomedial and ventromedial nuclei of the hypothalamus in young (2 months) and old (2 years) rats by immunohistochemical methods and Western blotting. In aged males and females, a decrease in Sirt1 expression in dorsomedial nucleus was observed. In ventromedial nucleus, the expression of Sirt1 did not change with age. The results confirm the hypothesis that aging is associated with a decrease in the content of sirtuins in the hypothalamic nuclei.

Structural and functional connectivity from the dorsomedial hypothalamus to the ventral medulla as a chronological amplifier of sympathetic outflow.

Psychological stress activates the hypothalamus, augments the sympathetic nervous output, and elevates blood pressure via excitation of the ventral medullary cardiovascular regions. However, anatomical and functional connectivity from the hypothalamus to the ventral medullary cardiovascular regions has not been fully elucidated. We investigated this issue by tract-tracing and functional imaging in rats. Retrograde tracing revealed the rostral ventrolateral medulla was innervated by neurons in the ipsilateral dorsomedial hypothalamus (DMH). Anterograde tracing showed DMH neurons projected to the ventral medullary cardiovascular regions with axon terminals in contiguity with tyrosine hydroxylase-immunoreactive neurons. By voltage-sensitive dye imaging, dynamics of ventral medullary activation evoked by electrical stimulation of the DMH were analyzed in the diencephalon-lower brainstem-spinal cord preparation of rats. Although the activation of the ventral medulla induced by single pulse stimulation of the DMH was brief, tetanic stimulation caused activation of the DMH sustained into the post-stimulus phase, resulting in delayed recovery. We suggest that prolonged excitation of the DMH, which is triggered by tetanic electrical stimulation and could also be triggered by psychological stress in a real life, induces further prolonged excitation of the medullary cardiovascular networks, and could contribute to the pathological elevation of blood pressure. The connectivity from the DMH to the medullary cardiovascular networks serves as a chronological amplifier of stress-induced sympathetic excitation. This notion will be the anatomical and pathophysiological basis to understand the mechanisms of stress-induced sustained augmentation of sympathetic activity.

A discrete neuronal circuit induces a hibernation-like state in rodents.

Hibernating mammals actively lower their body temperature to reduce energy expenditure when facing food scarcity1. This ability to induce a hypometabolic state has evoked great interest owing to its potential medical benefits2,3. Here we show that a hypothalamic neuronal circuit in rodents induces a long-lasting hypothermic and hypometabolic state similar to hibernation. In this state, although body temperature and levels of oxygen consumption are kept very low, the ability to regulate metabolism still remains functional, as in hibernation4. There was no obvious damage to tissues and organs or abnormalities in behaviour after recovery from this state. Our findings could enable the development of a method to induce a hibernation-like state, which would have potential applications in non-hibernating mammalian species including humans.

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.