Neurons In Dorsomedial Hypothalamus Research Articles

Top-down brain circuits for operant bradycardia.

Heart rate (HR) can be voluntarily regulated when individuals receive real-time feedback. In a rat model of HR biofeedback, the neocortex and medial forebrain bundle were stimulated as feedback and reward, respectively. The rats reduced their HR within 30 minutes, achieving a reduction of approximately 50% after 5 days of 3-hour feedback. The reduced HR persisted for at least 10 days after training while the rats exhibited anxiolytic behavior and an elevation in blood erythrocyte count. This bradycardia was prevented by inactivating anterior cingulate cortical (ACC) neurons projecting to the ventromedial thalamic nucleus (VMT). Theta-rhythm stimulation of the ACC-to-VMT pathway replicated the bradycardia. VMT neurons projected to the dorsomedial hypothalamus (DMH) and DMH neurons projected to the nucleus ambiguus, which innervates parasympathetic neurons in the heart.

Dorsomedial hypothalamus-raphe pallidus-cardiac sympathetic pathway mediates electroacupuncture intervention of stress-induced tachycardia.

Electroacupuncture at Neiguan point (PC6) effectively ameliorates tachycardia. However, very little is known about the neural pathway mechanism underlying the effect of electroacupuncture at PC6 in stress-induced tachycardia. Here, we investigate whether there exists a dorsomedial hypothalamus (DMH)-raphe pallidus (RP)-heart pathway to mediate the effect of electroacupuncture at PC6. The virus tracing results show that the heart is innervated by the neurons in DMH and RP, and the neurons of DMH project to RP. Chemogenetic inhibition of RP projecting DMH neurons reverses the cardiac autonomic imbalance and tachycardia induced by stress. Of note, immunofluorescence results show that the neural activity of DMH and RP is inhibited by electroacupuncture at PC6 accompanied with improved cardiac autonomic imbalance and tachycardia under stress. Moreover, chemogenetic inhibition of RP projecting DMH neurons cannot affect autonomic nervous activity and heart rate of stress rats after administrating electroacupuncture at PC6.NEW & NOTEWORTHY Our study suggests that this dorsomedial hypothalamus (DMH)-raphe pallidus (RP)-cardiac sympathetic pathway involves in the improvement of cardiac dysfunction associated with stress by administrating electroacupuncture at PC6, thus providing beneficial information for the development of therapeutic strategies to prevent stress-induced cardiovascular diseases, and insight into neural pathway basis for electroacupuncture at PC6 intervention of cardiac dysfunction.

Linkage and association of rs3110045 and rs28499085 variants in the thyrotropin-releasing hormone receptor (TRHR) gene with the risk of familial type 2 diabetes

Type 2 diabetes (T2D) is a chronic and prevalent multisystemic disease that significantly increases morbidity and mortality. Dysfunction of the thyroid hormone system is common in patients with T2D, increasing their risk of both hyperthyroidism and hypothyroidism. Several components of the thyroid system are candidate risk genes for T2D. The thyrotropin-releasing hormone receptor (TRHR) gene encoding for TRHR is of particular interest since it is expressed by the dorsomedial hypothalamus neurons, which are known to regulate food intake. In humans, a variant in the TRHR gene has been previously reported in T2D patients in a population-based case-control study but not in familial T2D. We recruited 212 multigenerational families with T2D originated from the Italian peninsula with multiple cases of T2D and tested, via Pseudomarker 9 single nucleotide polymorphisms (SNPs) in the TRHR gene for linkage and linkage disequilibrium (i.e., linkage plus association) to/with T2D. We identified 2 novel risk variants (rs3110045 and rs28499085) significantly linked to and associated with the risk of T2D in the Italian families across several inheritance models. Our study is the first to confirm the previously reported association of TRHR gene with T2D and extends the risk to familial inheritance. However, functional and replication studies are still needed to confirm these results.

Leptin signaling in the dorsomedial hypothalamus couples breathing and metabolism in obesity

Mismatch between CO2 production (Vco2) and respiration underlies the pathogenesis of obesity hypoventilation. Leptin-mediated CNS pathways stimulate both metabolism and breathing, but interactions between these functions remain elusive. We hypothesized that LEPRb+ neurons of the dorsomedial hypothalamus (DMH) regulate metabolism and breathing in obesity. In diet-induced obese LeprbCre mice, chemogenetic activation of LEPRb+ DMH neurons increases minute ventilation (Ve) during sleep, the hypercapnic ventilatory response, Vco2, and Ve/Vco2, indicating that breathing is stimulated out of proportion to metabolism. The effects of chemogenetic activation are abolished by a serotonin blocker. Optogenetic stimulation of the LEPRb+ DMH neurons evokes excitatory postsynaptic currents in downstream serotonergic neurons of the dorsal raphe (DR). Administration of retrograde AAV harboring Cre-dependent caspase to the DR deletes LEPRb+ DMH neurons and abolishes metabolic and respiratory responses to leptin. These findings indicate that LEPRb+ DMH neurons match breathing to metabolism through serotonergic pathways to prevent obesity-induced hypoventilation.

0040 Leptin Signaling in the Dorsomedial Hypothalamus Couples Metabolism and Breathing in Obesity

Abstract Introduction Individuals with obesity hypoventilation syndrome (OHS) are unable to increase ventilation to match increased CO2 production (VCO2). Obstructive sleep apnea (OSA), defined by the closure of the upper airway during sleep, is present in 90% of OHS patients. Leptin, an adipocyte-produced hormone, acts in leptin receptor (LEPRb) positive neurons and was implicated in the pathophysiology of OSA and OHS. Diet-induced obese (DIO) C57BL/6J mice shows features of OHS and OSA. Leptin-mediated neural pathways may link metabolism and breathing in obesity. We hypothesized that LEPRb-positive neurons in the dorsomedial hypothalamus (DMH) are necessary and sufficient to match control of breathing to VCO2 and that the stimulation of these neurons relieves OHS in DIO mice. Methods We expressed designer receptors exclusively activated by designer drugs (DREADDs) in DMH of DIO Leprb-Cre mice and measured breathing during sleep, the hypercapnic ventilatory response (HCVR), VCO2, and O2 consumption (VO2). Results Activation of LEPRb-positive DMH neurons increased inspiratory flow and minute ventilation during REM sleep, without affecting sleep architecture. The effect was pronounced during inspiratory flow limitation, suggeting that upper airway obstruction during REM sleep was relieved. Activation of these neurons increased the HCVR and this response was blunted by a serotonin receptor antagonist, methysergide. Activation of these neurons augmented VCO2, VO2, and VE/VCO2, suggesting that breathing was stimulated out of proportion to increases in metabolism. In vitro experiments revealed that LEPRb-positive DMH neurons project to 5-HT neurons in the dorsal raphe (DR) and that optogenetic stimulation of these neurons evoked excitatory postsynaptic currents in 5-HT neurons of the DR. Administration of retrograde Cre-dependent AAV harboring caspase to the DR in Leprb-Cre mice deleted LEPRb-positive DMH neurons and abolished respiratory and metabolic effects of chemogenetic stimulation. Apoptosis of LEPRb-positive DMH neurons projecting to the DR also prevented the increase in ventilation during REM sleep induced by intranasal leptin. Conclusion Leptin increases the HCVR, improves upper airway patency during sleep, and increases VCO2 acting on LEPRb-positive DMH neurons projecting to the DR. We conclude that LEPRb-positive DMH neurons projecting to the DR couple metabolism and breathing in obesity. Support (if any) NIH (R01 HL128970, R01 HL133100, and R01 HL138932), AHA (#915167).

Tonic GABAergic signaling from prostaglandin EP3 receptor-expressing preoptic neurons bidirectionally controls body temperature

The principal controller neurons of the thermoregulatory center in the preoptic area (POA) have yet to be determined. In the present study using rats, we discovered that prostaglandin EP3 receptor-expressing POA neurons (POA EP3R neurons) are a pivotal bidirectional controller in the central thermoregulatory circuit mechanism. POA EP3R neurons were activated in response to elevated ambient temperature, but inhibited by prostaglandin E 2 , a pyrogenic mediator. In newly generated Ptger3 (EP3R)-tTA transgenic rats, chemogenetic stimulation of POA EP3R neurons at room temperature reduced body temperature by eliciting skin vasodilation (enhancing heat dissipation), whereas inhibition of them elicited hyperthermia involving brown adipose tissue thermogenesis and tachycardia, mimicking fever. Furthermore, we revealed that POA EP3R neurons innervate sympathoexcitatory neurons in the dorsomedial hypothalamus (DMH) via tonic inhibitory signaling. Although many POA EP3R neuronal cell bodies expressed a glutamatergic mRNA marker, paradoxically, most of their axons in the DMH contained GABAergic synaptic proteins. Interestingly, such GABAergic terminals of POA EP3R →DMH axons were increased by chronic heat exposure of rats, indicating a synaptic alteration to enhance heat tolerance. Consistent with the anatomical observation, slice patch-clamp recordings from DMH neurons combined with optogenetic stimulation of POA EP3R →DMH axon terminals showed that these terminals predominantly form GABAergic synapses onto DMH neurons. These findings demonstrate that tonic GABAergic inhibitory signaling from POA EP3R neurons is a fundamental determinant of body temperature for thermal homeostasis and fever. NEXT Program from JSPS (LS070 to K.N.); MEXT KAKENHI (JP21K06767, JP17K08568, JP26860159, JP23790271 to Y.N.; JP22K06844 to A.F.; JP22K06470, JP19K06954, JP16K19006 to N.K.; JP21H02592 to H.H.; JP20H03418, JP16H06276 (AdAMS), JP16H05128, JP15H05932, JP26118508, JP26713009, JP22689007 to K.N.); PRESTO (JPMJPR13M9 to K.N.), FOREST (JPMJFR204D to H.H.), Moonshot R&D (JPMJMS2024 to H.H.; JPMJMS2023 to K.N.) of JST; AMED (JP21wm0525002 to N.K.; JP21dm0207112 to H.H.; JP21gm5010002 to K.N.); Hori Sciences and Arts Foundation (to Y.N.); Kato Memorial Bioscience Foundation, Foundation of Kinoshita Memorial Enterprise (to N.K.); Takeda Science Foundation (to T.Y. and K.N.); Nakajima Foundation, Uehara Memorial Foundation, Ono Medical Research Foundation, Brain Science Foundation, Kowa Life Science Foundation (to K.N.). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Leptin signaling in the dorsomedial hypothalamus couples metabolism and breathing in obesity

Individuals with obesity hypoventilation syndrome (OHS) are unable to increase ventilation to match increased CO 2 production (VCO 2 ). Obstructive sleep apnea (OSA), defined by the closure of the upper airway during sleep, is present in 90% of OHS patients. Leptin, an adipocyte-produced hormone, acts in leptin receptor (LEPR b ) positive neurons and was implicated in the pathophysiology of OSA and OHS. Diet-induced obese (DIO) C57BL/6J mice shows features of OHS and OSA. Leptin-mediated neural pathways may link metabolism and breathing in obesity. We hypothesized that LEPR b -positive neurons in the dorsomedial hypothalamus (DMH) are necessary and sufficient to match control of breathing to VCO 2 and that the stimulation of these neurons relieves OHS in DIO mice. We examined our hypothesis using in vivo and in vitro studies. We expressed designer receptors exclusively activated by designer drugs (DREADDs) in DMH of DIO Lepr b -Cre mice and measured breathing during sleep, the hypercapnic ventilatory response (HCVR), VCO 2 , and O 2 consumption (VO 2 ). Activation of LEPR b -positive DMH neurons increased inspiratory flow and minute ventilation during REM sleep, without affecting sleep architecture. The effect was pronounced during inspiratory flow limitation, suggesting that upper airway obstruction during REM sleep was relieved. Activation of these neurons increased the HCVR and this response was blunted by a serotonin receptor antagonist, methysergide. Activation of these neurons augmented VCO 2 , VO 2 , and VE/VCO 2 , suggesting that breathing was stimulated out of proportion to increases in metabolism. In vitro experiments revealed that LEPR b -positive DMH neurons project to 5-HT neurons in the dorsal raphe (DR) and that optogenetic stimulation of these neurons evoked excitatory postsynaptic currents in 5-HT neurons of the DR. Administration of retrograde Cre-dependent AAV harboring caspase to the DR in Lepr b -Cre mice deleted LEPR b -positive DMH neurons and abolished respiratory and metabolic effects of chemogenetic stimulation. Apoptosis of LEPR b -positive DMH neurons projecting to the DR also prevented the increase in ventilation during REM sleep induced by intranasal leptin. Leptin increases the HCVR, improves upper airway patency during sleep, and increases VCO 2 acting on LEPR b -positive DMH neurons projecting to the DR. We conclude that LEPR b -positive DMH neurons projecting to the DR couple metabolism and breathing in obesity. NIH (R01 HL128970, R01 HL133100, and R01 HL138932), AHA (#915167). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Leptin is a key regulator of glucose homeostasis in obesity

Approximately 9 out of 10 people with type-2 diabetes are overweight. Leptin is a hormone secreted from white fat cells that exerts control over both food intake and energy expenditure. We described the central mechanisms underlying the hypertensive effects of hyperleptinemia in obesity. Work presented here used continuous radiotelemetric glucose monitoring in mice in combination with multiple gold standard techniques in order to determine the effects of dorsomedial hypothalamic (DMH) leptin signalling on peripheral glucose control. A leptin receptor (LepR) antagonist administered directly into the brain impaired DIO mouse glucose tolerance demonstrating that leptin retains the ability to regulate blood glucose concentration in obesity. Knockdown of DMH LepR function using short hairpin adeno-associated virus (AAV) RNA significantly reduced brown adipose tissue (BAT) thermogenesis by -0.8 ± 0.1°C and increased basal blood glucose (14.3 ± 0.6mmol/l to 9.4 ± 0.5mmol/l) and impaired glucose tolerance. Conversely, chemogenic activation of LepR-expressing DMH neurons significantly elevated BAT thermogenesis (1.5 ± 0.2 °C increase). This increased BAT temperature was immediately followed by a decrease in plasma blood glucose concentration (7.6 ± 0.5 mmol/l to 5.8 ± 0.5 mmol/l). Work presented here demonstrates that increasing the activity of LepR expressing neurons in the DMH can improve glucose tolerance in obesity though an elevation of BAT metabolic activity. This work was supported by the National Heart Foundation of Australia (SES), The National Health and Medical Research Council of Australia (SES and MAC). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Electrophysiological complexity in the rat dorsomedial hypothalamus and its susceptibility to daily rhythms and high-fat diet.

The dorsomedial hypothalamus (DMH) in amongst the most important brain structures involved in the regulation of feeding behaviour and metabolism. In contrast to other hypothalamic centres, its main role is related to the circadian rhythmicity of food intake and energy homeostasis; both reported to be disrupted in obesity. In modern world, overweight and obesity reached global epidemic proportions. Thus, not only is it important to study their negative implications but also the mechanism responsible for their development. Here, we exposed rats to short-term (2-4weeks) high-fat diet (HFD)-not long enough to induce obesity. Next, we performed electrophysiological patch-clamp recordings ex vivo from neurons in the DMH either during the day or at night. Our results showed a day-to-night change in the firing frequency of DMH cells, with higher activity during the dark phase. This was abolished by HFD consumption, resulting in a decreased threshold for action potential generation during the day and therefore increased electrical activity at this phase. We propose this electrophysiological disturbance as a mechanism for the induction of abnormal daytime feeding, previously observed for HFD-fed animals, which might in turn contribute to the development of obesity. In addition, we provide an electrophysiological characteristic of DMH neurons with a separation into three anatomically and functionally distinct subpopulations, namely, the compact part, separating the structure into the ventral and dorsal divisions. Our study is the first to show electrophysiological complexity of the DMH with its sensitivity to diet and daily rhythms.

A role of prefrontal cortico-hypothalamic projections in wake promotion.

Ventromedial prefrontal cortex (vmPFC) processes many critical brain functions, such as decision-making, value-coding, thinking, and emotional arousal/recognition, but whether vmPFC plays a role in sleep-wake promotion circuitry is still unclear. Here, we find that photoactivation of dorsomedial hypothalamus (DMH)-projecting vmPFC neurons, their terminals, or their postsynaptic DMH neurons rapidly switches non-rapid eye movement (NREM) but not rapid eye movement sleep to wakefulness, which is blocked by photoinhibition of DMH outputs in lateral hypothalamus (LHs). Chemoactivation of DMH glutamatergic but not GABAergic neurons innervated by vmPFC promotes wakefulness and suppresses NREM sleep, whereas chemoinhibition of vmPFC projections in DMH produces opposite effects. DMH-projecting vmPFC neurons are inhibited during NREM sleep and activated during wakefulness. Thus, vmPFC neurons innervating DMH likely represent the first identified set of cerebral cortical neurons for promotion of physiological wakefulness and suppression of NREM sleep.

Hindbrain catecholaminergic inputs to the paraventricular thalamus scale feeding and metabolic efficiency in stress-related contexts.

The regulation of food intake and energy balance relies on the dynamic integration of exteroceptive and interoceptive signals monitoring nutritional, metabolic, cognitive, and emotional states. The paraventricular thalamus (PVT) is a central hub that, by integrating sensory, metabolic, and emotional states, may contribute to the regulation of feeding and homeostatic/allostatic processes. However, the underlying PVT circuits still remain elusive. Here, we aimed at unravelling the role of catecholaminergic (CA) inputs to the PVT in scaling feeding and metabolic efficiency. First, using region-specific retrograde disruption of CA projections, we show that PVT CA inputs mainly arise from the hindbrain, notably the locus coeruleus (LC) and the nucleus tractus solitarius. Second, taking advantage of integrative calorimetric measurements of metabolic efficiency, we reveal that CA inputs to the PVT scale adaptive feeding and metabolic responses in environmental, behavioural, physiological, and metabolic stress-like contexts. Third, we show that hindbrainTH →PVT inputs contribute to modulating the activity of PVT as well as lateral and dorsomedial hypothalamic neurons. In conclusion, the present study, by assessing the key role of CA inputs to the PVT in scaling homeostatic/allostatic regulations of feeding patterns, reveals the integrative and converging hindbrainTH →PVT paths that contribute to whole-body metabolic adaptations in stress-like contexts. KEY POINTS: The paraventricular thalamus (PVT) is known to receive projections from the hindbrain. Here, we confirm and further extend current knowledge on the existence of hindbrainTH →PVT catecholaminergic inputs, notably from the locus coeruleus and the nucleus tractus solitarius, with the nucleus tractus solitarius representing the main source. Disruption of hindbrainTH →PVT inputs contributes to the modulation of PVT neuron activity. HindbrainTH →PVT inputs scale feeding strategies in environmental, behavioural, physiological, and metabolic stress-like contexts. HindbrainTH →PVT inputs participate in regulating metabolic efficiency and nutrient partitioning in stress-like contexts. HindbrainTH →PVT inputs, directly and/or indirectly, contribute to modulating the downstream activity of lateral and dorsomedial hypothalamic neurons.

Day/night Changes in the Dorsomedial Hypothalamus Firing Responses to Ghrelin are Modulated by High-fat Diet

Dorsomedial hypothalamus (DMH) is a part of the feeding center involved in food intake and regulation of the metabolism. DMH neurons express many receptors for different metabolic cues which can modulate its network and influence animals’ behaviour. One of the metabolic peptides deliveredto this structure is ghrelin, the only well-known hunger signal, produced mainly in the stomach. Diet-induced obesity is a physiological model of obesity widely used in research. Here we investigated how time-of-day and high-fat diet (HFD) affect neuronal networks and the sensitivity to the metabolic information received by the DMH. Our results indicate that even a short period of HFD (2–3 weeks) consumption can cause dysregulation of the DMH neuronal network, manifested as a disruption of the day/night pattern of basal activity and altered sensitivity to incoming information. We showed for the first time a day/night pattern of sensitivity to ghrelin in the DMH, with a higher level during the behaviorally active phase of animals. This day/night rhythm of sensitivity to ghrelin was reversed in HFD group, causing a stronger effect during the non-active phase. After prolongation of the HFD consumption to 7–8 weeks we observed an increase in the responsiveness to ghrelin, than during the short-term diet.

Activation of Leptin‐Receptor‐Expressing Neurons in the Dorsomedial Hypothalamus Increases Hypercapcnic Ventilatory Response and Relieves Obstructive Sleep Apnea in Obese Mice

IntroductionObesity leads to obstructive sleep apnea (OSA), characterized by loss of upper airway muscle tone during sleep leading to carbon dioxide retention and intermittent hypoxemia. OSA leads to high neurocognitive, cardiovascular, and oncologic morbidity and mortality. There is no effective pharmacotherapy for OSA. We have previously shown that an adipose‐produce hormone leptin acts in the brain to abolish hypoventilation and upper airway obstruction during sleep, and, therefore may be a promising therapeutic in obese patients with OSA. Our previous data also suggest that leptin stimulates breathing acting on the leptin receptor (LEPRb) positive neurons in the dorsomedial hypothalamus (DMH), but not in the medulla. Herein, we tested the hyphotesis that leptin stimulates control of breathing and improves upper airway patency during sleep acting on LEPRb positive DMH neurons and their projections to serotonergic neurons in raphe. Aim:In this study, we examined the contribution of LEPRb positive neurons in the DMH to control of breathing and upper airway function during sleep in diet‐induced obese mice.MethodsWe expressed designer receptors exclusively activated by designer drugs (DREADD) selectively in the LEPRb positive neurons of the DMH of Leprb‐Cre‐GFP mice with diet‐induced obesity (DIO) and examined the effect of DREADD ligand, J60, on breathing during sleep.ResultsChemogenetic activation of LEPRb positive DMH neurons increased inspiratory flow, respiratory rate, and minute ventilation during REM sleep, without significant changes in sleep architecture. The effect was particularly pronounced during inspiratory flow limitation, which suggest that upper airway obstruction during REM sleep was relieved. Activation of LEPRb positive DMH neurons with a DREADD ligand, J60, increased the hypercapnic ventilatory response and this response was blunted by a serotonin receptor antagonist, methysergide. In vitroexperiments revealed that DMH LEPRb neurons project to the serotonergic neurons in the dorsal raphe and that optogenetic stimulation of DMH LEPRb neurons evoked excitatory post‐synaptic currents in ChR2 expressing neurons.ConclusionLeptin up‐regulates the hypercapnic ventilatory response and upper airway patency during sleep acting on LEPRb neurons in the DMH. Respiratory effects of leptin are likely mediated via projections of these neurons to the dorsal raphe.

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.

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

ObjectiveThe 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. MethodsTargeted 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. ResultsFasting-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. ConclusionsWe 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.

The Role of Dorsomedial Hypothalamic LepRb‐expressing Neurons in the Control of Respiratory Motor Output

Previously we have demonstrated that leptin in the NTS activates leptin receptor (LepRb) and galanin-expressing neurons which directly increases phrenic motor output via a NALCN-dependent mechanism (Do et al, Cell Reports, 2020). We also reported that such NTS LepRb neurons are glutamatergic and project to the rostral ventral respiratory group (rVRG) and, interestingly, the dorsomedial hypothalamus (DMH). While the excitatory projections to the rVRG are consistent with the robust increase in integrated phrenic activity, projections to the DMH imply an integrative leptin-dependent circuit. The DMH includes a significant number of LepRb expressing neurons and our previous preliminary work suggests that optogenetic activation of DMH LepRb neurons increases phrenic nerve activity (Chang et al, FASEB Abstr. 2018). Here we report that selective optogenetic activation of LepRb DMH neurons induces a robust increase, on discrete timescales, in integrated phrenic activity and inspiratory activity in vagal motoneurons. These DMH LepRb neurons are dominantly GABAergic confirmed by the expression of VGAT with in situ hybridization. Lastly, viral tracing studies reveals DMH LepRb neurons project to the arcuate nucleus, periaqueductal gray (PAG), and parabrachial nucleus (PBN). Together, these findings highlight a leptin-dependent suprapontine circuit that, along with NTS LepRb neurons, is likely to participate in matching respiratory motor output to metabolism.

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 expression in the dorsomedial hypothalamus stimulates breathing during NREM sleep in db/db mice.

Obesity leads to obstructive sleep apnea (OSA), which is recurrent upper airway obstruction during sleep, and obesity hypoventilation syndrome (OHS), hypoventilation during sleep resulting in daytime hypercapnia. Impaired leptin signaling in the brain was implicated in both conditions, but mechanisms are unknown. We have previously shown that leptin stimulates breathing and treats OSA and OHS in leptin-deficient ob/ob mice and leptin-resistant diet-induced obese mice and that leptin's respiratory effects may occur in the dorsomedial hypothalamus (DMH). We hypothesized that leptin receptor LepRb-deficient db/db mice have obesity hypoventilation and that restoration of leptin signaling in the DMH will increase ventilation during sleep in these animals. We measured arterial blood gas in unanesthetized awake db/db mice. We subsequently infected these animals with Ad-LepRb or control Ad-mCherry virus into the DMH and measured ventilation during sleep as well as CO2 production after intracerebroventricular (ICV) infusions of phosphate-buffered saline or leptin. Awake db/db mice had elevated CO2 levels in the arterial blood. Ad-LepRb infection resulted in LepRb expression in the DMH neurons in a similar fashion to wildtype mice. In LepRb-DMH db/db mice, ICV leptin shortened REM sleep and increased inspiratory flow, tidal volume, and minute ventilation during NREM sleep without any effect on the quality of NREM sleep or CO2 production. Leptin had no effect on upper airway obstruction in these animals. Leptin stimulates breathing and treats obesity hypoventilation acting on LepRb-positive neurons in the DMH.