Ventromedial Hypothalamus Of Mice Research Articles

7753 Anoctamin 4 in the Area Postrema Regulates Glucose and Energy Homeostasis

Abstract Disclosure: L. Tu: None. Y. Xu: None. Located on the floor of the 4th ventricle within the dorsal surface of the medulla oblongata, the area postrema (AP) is a circumventricular organ where a proper blood-brain barrier is missing. Numerous studies reveal that neurons in the AP are implicated into suppression of food intake (e.g., glucagon-like peptide 1 (GLP-1)-mediated anorexia), but the role of AP neurons in whole-body glucose homeostasis is less investigated. Here we identified and fully characterized the physiological functions of anoctamin-4 (Ano4, a calcium activated chloride channel)1 in the AP. CRISPR-mediated deletion of Ano4 in the AP caused a lower body weight and reduced blood glucose, concurrent with an improved glucose tolerance and increased insulin sensitivity. Chemogenetic activation of Ano4-expressing neurons in the AP (APAno4) increased food intake and blood glucose, and did not induce conditioned flavor avoidance. Both Ano4 neurons and non-Ano4 neurons (i.e., neurons do not express Ano4) sent projections to the lateral parabrachial nucleus (LPBN). Optogenetic stimulation of APAno4 ◊ LPBN circuit promoted feeding behavior and increased blood glucose, whereas stimulation of APnon-Ano4 ◊ LPBN circuit inhibited food intake and did not influence glucose homeostasis. Finally, RNAscope studies reveal that 45.7% of APAno4 neurons are glutamatergic neurons, and 41.8% of them are GABAergic neurons. Furthermore, APAno4 neurons showed only a 2.6% overlap with Glp-1r-expressing neurons in the AP, but had a 60.8% co-localization with protein tyrosine phosphatase receptor δ2, which serves as orexigenic asprosin receptor. Together, our results indicate that Ano4 neurons in the AP are the first reported orexigenic neurons, and represent potential therapeutic targets for human diseases with glucose imbalance and abnormal feeding behavior. Reference: (1) Tu, L. et al. Anoctamin 4 channel currents activate glucose-inhibited neurons in the mouse ventromedial hypothalamus during hypoglycemia. J Clin Invest 133,14. (2023). (2) Mishra, I. et al. Protein tyrosine phosphatase receptor δ serves as the orexigenic asprosin receptor. Cell Metab 34, 549-563. (2022). Presentation: 6/2/2024

9207 Anoctamin 4 in the Area Postrema Regulates Glucose and Energy Homeostasis

Abstract Disclosure: L. Tu: None. Y. Xu: None. Located on the floor of the 4th ventricle within the dorsal surface of the medulla oblongata, the area postrema (AP) is a circumventricular organ where a proper blood-brain barrier is missing. Numerous studies reveal that neurons in the AP are implicated into suppression of food intake (e.g., glucagon-like peptide 1 (GLP-1)-mediated anorexia), but the role of AP neurons in whole-body glucose homeostasis is less investigated. Here we identified and fully characterized the physiological functions of anoctamin-4 (Ano4, a calcium activated chloride channel)1 in the AP. CRISPR-mediated deletion of Ano4 in the AP caused a lower body weight and reduced blood glucose, concurrent with an improved glucose tolerance and increased insulin sensitivity. Chemogenetic activation of Ano4-expressing neurons in the AP (APAno4) increased food intake and blood glucose, and did not induce conditioned flavor avoidance. Both Ano4 neurons and non-Ano4 neurons (i.e., neurons do not express Ano4) sent projections to the lateral parabrachial nucleus (LPBN). Optogenetic stimulation of APAno4 ◊ LPBN circuit promoted feeding behavior and increased blood glucose, whereas stimulation of APnon-Ano4 ◊ LPBN circuit inhibited food intake and did not influence glucose homeostasis. Finally, RNAscope studies reveal that 45.7% of APAno4 neurons are glutamatergic neurons, and 41.8% of them are GABAergic neurons. Furthermore, APAno4 neurons showed only a 2.6% overlap with Glp-1r-expressing neurons in the AP, but had a 60.8% co-localization with protein tyrosine phosphatase receptor δ2, which serves as orexigenic asprosin receptor. Together, our results indicate that Ano4 neurons in the AP are the first reported orexigenic neurons, and represent potential therapeutic targets for human diseases with glucose imbalance and abnormal feeding behavior. Reference: (1) Tu, L. et al. Anoctamin 4 channel currents activate glucose-inhibited neurons in the mouse ventromedial hypothalamus during hypoglycemia. J Clin Invest 133,14. (2023). (2) Mishra, I. et al. Protein tyrosine phosphatase receptor δ serves as the orexigenic asprosin receptor. Cell Metab 34, 549-563. (2022). Presentation: 6/2/2024

Anoctamin 4 channel currents activate glucose-inhibited neurons in the mouse ventromedial hypothalamus during hypoglycemia.

Glucose is the basic fuel essential for maintenance of viability and functionality of all cells. However, some neurons - namely, glucose-inhibited (GI) neurons - paradoxically increase their firing activity in low-glucose conditions and decrease that activity in high-glucose conditions. The ionic mechanisms mediating electric responses of GI neurons to glucose fluctuations remain unclear. Here, we showed that currents mediated by the anoctamin 4 (Ano4) channel are only detected in GI neurons in the ventromedial hypothalamic nucleus (VMH) and are functionally required for their activation in response to low glucose. Genetic disruption of the Ano4 gene in VMH neurons reduced blood glucose and impaired counterregulatory responses during hypoglycemia in mice. Activation of VMHAno4 neurons increased food intake and blood glucose, while chronic inhibition of VMHAno4 neurons ameliorated hyperglycemia in a type 1 diabetic mouse model. Finally, we showed that VMHAno4 neurons represent a unique orexigenic VMH population and transmit a positive valence, while stimulation of neurons that do not express Ano4 in the VMH (VMHnon-Ano4) suppress feeding and transmit a negative valence. Together, our results indicate that the Ano4 channel and VMHAno4 neurons are potential therapeutic targets for human diseases with abnormal feeding behavior or glucose imbalance.

Aggression: The dark side of mirror neurons sheds light on their functions

Mirror neurons have been found mainly in neocortical structures of primates and rodents; however, their functions are still debated. A new study has discovered mirror neurons for aggressive behaviors in the ventromedial hypothalamus of mice, an evolutionarily ancient structure, highlighting a new function key for survival.

LBSUN99 Combination Of Drosophila And Mammalian Genetics Identifies Lcorl As A Novel Regulator Of Metabolism

Abstract Obesity and adult-onset diabetes result from a complex interaction of multiple genetic, environmental, and behavioral factors. The hypothalamus contains multiple nuclei required for the regulation of metabolic homeostasis. One nucleus, termed the ventromedial hypothalamus (VMH), facilitates glucose homeostasis and metabolic adaptations to challenges such as high fat diet (HFD) and exercise. Using a combination of Drosophila and mouse genetics, we uncovered a novel gene network whose function is required for VMH-regulated whole body metabolism. Using Drosophila to rapidly screen orthologs of genes enriched in the mouse VMH, we identified the gene encoding the Ecdysone Induced Protein 93F (E93; human ortholog, Ligand dependent corepressor-like, Lcorl). Adult flies with neural knockdown of E93 are obese and hyperphagic, with increased energy stores, reduced exercise endurance, and dampened circadian amplitude. These findings reveal a novel role of E93 in metabolism. We found that the knockdown of E93 specifically in myoinhibitory peptide (MIP) and GABAergic neurons are sufficient to recapitulate the phenotype seen in a pan-neuronal E93 knockdown. | In mice, we found that Lcorl, the mammalian orthologue of E93, is highly expressed throughout the entire VMH. We used CRISPR/Cas9 to generate mice harboring both a Lcorl KO and a floxed allele to investigate its function in the whole-body and in a tissue specific manner. Lcorl KO mice are viable and fertile. However, they have disruptions in growth, lipid metabolism, and circadian rhythm. Taken together these data reveal that Lcorl is as a novel regulator of energy metabolism and circadian rhythm. Additionally, this study shows the power of combined drosophila and mammalian genetics in uncovering novel regulators of metabolism. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m.

Sensory-thresholded switch of neural firing states in a computational model of the ventromedial hypothalamus.

The mouse ventromedial hypothalamus (VMH) is both necessary and sufficient for defensive responses to predator and social threats. Defensive behaviors typically involve cautious approach toward potentially threatening stimuli aimed at obtaining information about the risk involved, followed by sudden avoidance and flight behavior to escape harm. In vivo neural recording studies in mice have identified two major populations of VMH neurons that either increase their firing activity as the animal approaches the threat (called Assessment+ cells) or increase their activity as the animal flees the threat (called Flight+ cells). Interestingly, Assessment+ and Flight+ cells abruptly decrease and increase their firing activity, respectively, at the decision point for flight, creating an escape-related “switch” in functional state. This suggests that the activity of the two cell types in VMH is coordinated and could result from local circuit interactions. Here, we used computational modeling to test if a local inhibitory feedback circuit could give rise to key features of the neural activity seen in VMH during the approach-to-flight transition. Starting from a simple dual-population inhibitory feedback circuit receiving repeated trains of monotonically increasing sensory input to mimic approach to threat, we tested the requirement for balanced sensory input, balanced feedback, short-term synaptic plasticity, rebound excitation, and inhibitory feedback exclusivity to reproduce an abrupt, sensory-thresholded reciprocal firing change that resembles Assessment+ and Flight+ cell activity seen in vivo. Our work demonstrates that a relatively simple local circuit architecture is sufficient for the emergence of firing patterns similar to those seen in vivo and suggests that a reiterative process of experimental and computational work may be a fruitful avenue for better understanding the functional organization of mammalian instinctive behaviors at the circuit level.

Ciliary Type III Adenylyl Cyclase in the VMH Is Crucial for High-Fat Diet-Induced Obesity Mediated by Autophagy.

Neuronal primary cilia are crucial for body weight maintenance. Type III adenylyl cyclase (AC3) is abundantly enriched in neuronal cilia, and mice with global AC3 ablation are obese. However, whether AC3 regulates body weight through its ciliary expression and the mechanism underlying this potential regulation are not clear. In this study, humanized AC3 knock‐in mice that are resistant to high‐fat diet (HFD)‐induced obesity are generated, and increases in the number and length of cilia in the ventromedial hypothalamus (VMH) are shown. It is demonstrated that mice with specifically knocked down ciliary AC3 expression in the VMH show pronounced HFD‐induced obesity. In addition, in vitro and in vivo analyses of the VMH show that ciliary AC3 regulates autophagy by binding an autophagy‐related gene, gamma‐aminobutyric acid A receptor‐associated protein (GABARAP). Mice with GABARAP knockdown in the VMH exhibit exacerbated HFD‐induced obesity. Overall, the findings may reveal a potential mechanism by which ciliary AC3 expression regulates body weight in the mouse VMH.

Dynamic encoding of social threat and spatial context in the hypothalamus.

Social aggression and avoidance are defensive behaviors expressed by territorial animals in a manner appropriate to spatial context and experience. The ventromedial hypothalamus controls both social aggression and avoidance, suggesting that it may encode a general internal state of threat modulated by space and experience. Here, we show that neurons in the mouse ventromedial hypothalamus are activated both by the presence of a social threat as well as by a chamber where social defeat previously occurred. Moreover, under conditions where the animal could move freely between a home and defeat chamber, firing activity emerged that predicted the animal's position, demonstrating the dynamic encoding of spatial context in the hypothalamus. Finally, we found that social defeat induced a functional reorganization of neural activity as optogenetic activation could elicit avoidance after, but not before social defeat. These findings reveal how the hypothalamus dynamically encodes spatial and sensory cues to drive social behaviors.

Hypothalamic estrogen receptor alpha establishes a sexually dimorphic regulatory node of energy expenditure.

Estrogen receptor a (ERa) signaling in the ventromedial hypothalamus (VMH) contributes to energy homeostasis by modulating physical activity and thermogenesis. However, the precise neuronal populations involved remain undefined. Here, we describe six neuronal populations in the mouse VMH by using single-cell RNA transcriptomics and in situ hybridization. ERa is enriched in populations showing sex biased expression of reprimo (Rprm), tachykinin 1 (Tac1), and prodynorphin (Pdyn). Female biased expression of Tac1 and Rprm is patterned by ERa-dependent repression during male development, whereas male biased expression of Pdyn is maintained by circulating testicular hormone in adulthood. Chemogenetic activation of ERa positive VMH neurons stimulates heat generation and movement in both sexes. However, silencing Rprm gene function increases core temperature selectively in females and ectopic Rprm expression in males is associated with reduced core temperature. Together these findings reveal a role for Rprm in temperature regulation and ERa in the masculinization of neuron populations that underlie energy expenditure.

The effect of alloxan-induced diabetes on triamine lesions in the ventromedial hypothalamus

It has been demonstrated that the ventromedial hypothalamus (VMH) of alloxan-induced diabetic mice is protected from subsequent gold thioglucose (GTG)-induced lesions. Another compound, 3,3′-methyliminobis-(N-methylpropylamine) (MIMPA), a triamine structurally unrelated to GTG, has been shown to cause similar VMH lesions in mice. We chose to investigate the effect of alloxan-induced diabetes on VMH lesion formation in MIMPA-treated mice. In this study CF-1 female mice were made diabetic by a simple intravenous (IV) injection of alloxan and subsequently treated with MIMPA by subcutaneous injection (SC). Contrary to studies which showed that GTG-induced VMH lesions are insulin dependent, an insulin deficiency did not inhibit MIMPA-induced lesions in the VMH of mice. Our data suggests, albeit GTG is suspected to induce VMH necrosis by attaching to glucoreceptors and insulin sensitive neurons, MIMPA works by a different and as yet unknown mechanism. We conclude that MIMPA-induced lesions in the VMH of mice are not insulin dependent.

Beta-thioglucose inhibits gold thioglucose lesions in the ventromedial hypothalamus

Gold thioglucose (GTG) has been known to be an obesity causing agent for over 40 years. GTG works by affecting dendrites in the mouse ventromedial hypothalamus (VMH) producing a permanent VMH lesion and subsequent hyperphagia and obesity. We have investigated the effect of beta-thioglucose (BTG), a glucose antimetabolite, on GTG-induced lesions in the VMH of mice. Twenty-eight female CF-1 mice were used in this study. Seven micron sections were made of the mouse VMH, mounted on glass slides, and stained with hematoxylin and eosin. A previous report of BTG action on GTG-induced lesions has not supported a competitive inhibition between these two drugs. Our data demonstrate that at 1 2 hour, 6 hours, and 12 hours post BTG, BTG completely inhibited GTG-induced lesions in the VMH.

An ultrastructural study of gold thioglucose lesion formation in the ventromedial hypothalamus: subnecrotic gold thioglucose dose.

In an ultrastructural study of gold thioglucose (GTG) lesion formation in the mouse ventromedial hypothalamus (VMH), a single intraperitoneal subnecrotic dose of GTG (100 mg/kg) was used to induce a VMH microlesion, visible only by electron microscopy. Definitive neuronal alterations were not evident until eight hours after GTG injection. Large pericapillary processes, probably dendrites, displayed a loss of cytoplasmic density and distention, while the capillary endothelium appeared normal. Some intercellular distention also was apparent. By 24 hours after GTG administration, degenerating neurons were visible. Swollen and disrupted synaptic profiles were seen. However, no vascular damage was evident. These results support the evidence that GTG is a neurotoxin for structures in the mouse VMH.

Glucose analog effects on glucose oxidation in the ventromedial hypothalamus.

The effect of the glucose analogs 2-deoxyglucose (2DG), glucosamine (GLA), mannoheptulose (MAN), and gold thioglucose (GTG) on glucose utilization in the mouse ventromedial hypothalamus (VMH) was assessed by using an in vitro incubation system. The amount of 14C-CO2 generated from uniformly labelled 14C-glucose was measured in VMH tissue cubes. MAN and GTG did not alter glucose oxidation at any of the concentrations compared with controls. GLA inhibited glucose utilization but only when the amount of GLA was 3 times that of glucose in the medium. However, this difference was not significant. The 2DG inhibited glucose oxidation significantly (50%; P less than 0.005), but only when 2DG was used in a quantity three-fold greater than glucose. These results imply that the VMH, a reputed central area for controlling feeding behavior, may possess a high affinity for glucose, which is not effectively altered by competitive inhibitors.