Excitation-inhibition imbalance in medial preoptic area circuits underlies chronic stress-induced depression-like states.

Dysregulation of brain homeostasis is associated with neuropsychiatric conditions such as major depressive disorder. However, underlying neural-circuit mechanisms remain not well-understood. We show in mice that chronic restraint stress (CRS) and social defeat stress (SDS) are both associated with disruption of excitation (E)-inhibition (I) balance, with increased E/I ratios, in medial preoptic area (MPOA) circuits, but through affecting different neuronal types. CRS results in elevated activity in glutamatergic neurons, and their suppression mitigates CRS-induced depressive-like behaviors. Paraventricular hypothalamic input to these neurons contributes to induction but not expression of depressive-like behaviors. Their projections to ventral tegmental area and periaqueductal gray/dorsal raphe suppress midbrain dopaminergic and serotonergic activity, respectively, and mediate expression of divergent depressive-like symptoms. By contrast, SDS results in reduced activity of GABAergic neurons, and their activation alleviates SDS-induced depressive-like behaviors. Thus, E/I imbalance with relatively increased excitation in MPOA circuits may be a general mechanism underlying depression caused by different etiological factors.

Central Mechanisms of Thermoregulation and Fever in Mammals.

Thermoregulation is a fundamental homeostatic function in mammals mediated by the central nervous system. The framework of the central circuitry for thermoregulation lies in the hypothalamus and brainstem. The preoptic area (POA) of the hypothalamus integrates cutaneous and central thermosensory information into efferent control signals that regulate excitatory descending pathways through the dorsomedial hypothalamus (DMH) and rostral medullary raphe region (rMR). The cutaneous thermosensory feedforward signals are delivered to the POA by afferent pathways through the lateral parabrachial nucleus, while the central monitoring of body core temperature is primarily mediated by warm-sensitive neurons in the POA for negative feedback regulation. Prostaglandin E2, a pyrogenic mediator produced in response to infection, acts on the POA to trigger fever. Recent studies have revealed that this circuitry also functions for physiological responses to psychological stress and starvation. Master psychological stress signaling from the medial prefrontal cortex to the DMH has been discovered to drive a variety of physiological responses for stress coping, including hyperthermia. During starvation, hunger signaling from the hypothalamus was found to activate medullary reticular neurons, which then suppress thermogenic sympathetic outflows from the rMR for energy saving. This thermoregulatory circuit represents a fundamental mechanism of the central regulation for homeostasis.

Divergent neural nodes are species- and hormone-dependent in the brood parasitic brain.

Avian brood parasitism is an evolutionarily derived behavior for which the neurobiological mechanisms are mostly unexplored. We aimed to identify brain regions that have diverged in the brood-parasitic brain using relative transcript abundance of social neuropeptides and receptors. We compared behavioral responses and transcript abundance in three brain regions in the brown-headed cowbird (BHCO), a brood parasite, and a closely related parental species, the red-winged blackbird (RWBL). Females of both species were treated with mesotocin (MT; avian homolog of oxytocin) or saline prior to exposure to nest stimuli. Results reveal that MT promotes approach toward nests with eggs rather than nests with begging nestlings in both species. We also examined relative transcript abundance of the five social neuropeptides and receptors in the brain regions examined: preoptic area (POA), paraventricular nucleus (PVN) and bed nucleus of the stria terminalis (BST). We found that MT-treated cowbirds but not blackbirds exhibited lower transcript abundance for two receptors, corticotropin-releasing factor 2 (CRFR2) and prolactin receptor (PRLR) in BST. Additionally, MT-treated cowbirds had higher PRLR in POA, comparable to those found in blackbirds, regardless of treatment. No other transcripts of interest exhibited significant differences as a result of MT treatment, but we found a significant effect of species in the three regions. Together, these results indicate that POA, PVN, and BST represent neural nodes that have diverged in avian brood parasites and may serve as neural substrates of brood-parasitic behavior.

Sedum kamtschaticum Exerts Hypnotic Effects via the Adenosine A2A Receptor in Mice.

Insomnia is a common sleep disorder with significant societal and economic impacts. Current pharmacotherapies for insomnia are often accompanied by side effects, necessitating the development of new therapeutic drugs. In this study, the hypnotic effects and mechanisms of Sedum kamtschaticum 30% ethanol extract (ESK) and one of its active compounds, myricitrin, were investigated using pentobarbital-induced sleep experiments, immunohistochemistry (IHC), receptor binding assays, and enzyme-linked immunosorbent assay (ELISA). The pentobarbital-induced sleep experiments revealed that ESK and myricitrin reduced sleep latency and prolonged total sleep time in a dose-dependent manner. Based on c-Fos immunostaining, ESK, and myricitrin enhanced the GABAergic neural activity in sleep-promoting ventrolateral preoptic nucleus (VLPO) GABAergic. By measuring the level of GABA released from VLPO GABAergic neurons, ESK and myricitrin were found to increase GABA release in the hypothalamus. These effects were significantly inhibited by SCH. Moreover, ESK exhibited a concentration-dependent binding affinity for the adenosine A2A receptors (A2AR). In conclusion, ESK and myricitrin have hypnotic effects, and their underlying mechanisms may be related to the activation of A2AR.

Opposing actions of co-released GABA and neurotensin on the activity of preoptic neurons and on body temperature.

Neurotensin (Nts) is a neuropeptide acting as a neuromodulator in the brain. Pharmacological studies have identified Nts as a potent hypothermic agent. The medial preoptic area, a region that plays an important role in the control of thermoregulation, contains a high density of neurotensinergic neurons and Nts receptors. The conditions in which neurotensinergic neurons play a role in thermoregulation are not known. In this study, optogenetic stimulation of preoptic Nts neurons induced a small hyperthermia. In vitro, optogenetic stimulation of preoptic Nts neurons resulted in synaptic release of GABA and net inhibition of the preoptic pituitary adenylate cyclase-activating polypeptide (Adcyap1) neurons firing activity. GABA-A receptor antagonist or genetic deletion of Slc32a1 (VGAT) in Nts neurons unmasked also an excitatory effect that was blocked by a Nts receptor 1 antagonist. Stimulation of preoptic Nts neurons lacking Slc32a1 resulted in excitation of Adcyap1 neurons and hypothermia. Mice lacking Slc32a1 expression in Nts neurons presented changes in the fever response and in the responses to heat or cold exposure as well as an altered circadian rhythm of body temperature. Chemogenetic activation of all Nts neurons in the brain induced a 4-5°C hypothermia, which could be blocked by Nts receptor antagonists in the preoptic area. Chemogenetic activation of preoptic neurotensinergic projections resulted in robust excitation of preoptic Adcyap1 neurons. Taken together, our data demonstrate that endogenously released Nts can induce potent hypothermia and that excitation of preoptic Adcyap1 neurons is the cellular mechanism that triggers this response.

Adaptation to photoperiod via dynamic neurotransmitter segregation.

Changes in the amount of daylight (photoperiod) alter physiology and behaviour1,2. Adaptive responses to seasonal photoperiods are vital to all organisms-dysregulation associates with disease, including affective disorders3 and metabolic syndromes4. The circadian rhythm circuitry is implicated in such responses5,6, yet little is known about the precise cellular substrates that underlie phase synchronization to photoperiod change. Here we identify a brain circuit and system of axon branch-specific and reversible neurotransmitter deployment that are critical for behavioural and sleep adaptation to photoperiod. A type of neuron called mrEn1-Pet17 in the mouse brainstem median raphe nucleus segregates serotonin from VGLUT3 (also known as SLC17A8, a proxy for glutamate) to different axonal branches that innervate specific brain regions involved in circadian rhythm and sleep-wake timing8,9. This branch-specific neurotransmitter deployment did not distinguish between daylight and dark phase; however, it reorganized with change in photoperiod. Axonal boutons, but not cell soma, changed neurochemical phenotype upon a shift away from equinox light/dark conditions, and these changes were reversed upon return to equinox conditions. When we genetically disabled Vglut3 in mrEn1-Pet1 neurons, sleep-wake periods, voluntary activity and clock gene expression did not synchronize to the new photoperiod or were delayed. Combining intersectional rabies virus tracing and projection-specific neuronal silencing, we delineated a preoptic area-to-mrEn1Pet1 connection that was responsible for decoding the photoperiodic inputs, driving the neurotransmitter reorganization and promoting behavioural synchronization. Our results reveal a brain circuit and periodic, branch-specific neurotransmitter deployment that regulates organismal adaptation to photoperiod change.

RNA-sequencing reveals a shared neurotranscriptomic profile in the medial preoptic area of highly social songbirds and rats.

Rough-and-tumble play in juvenile rats and song in flocks of adult songbirds outside a breeding context (gregarious song) are two distinct forms of non-sexual social behavior. Both are believed to play roles in the development of sociomotor skills needed for later life-history events, including reproduction, providing opportunities for low-stakes practice. Additionally, both behaviors are thought to be intrinsically rewarded and are associated with a positive affective state. Given the functional similarities of these behaviors, this study used RNA-sequencing to identify commonalities in their underlying neurochemical systems within the medial preoptic area. This brain region is implicated in multiple social behaviors, including song and play, and is highly conserved across vertebrates. DESeq2 and rank-rank hypergeometric overlap analyses identified a shared neurotranscriptomic profile in adult European starlings singing high rates of gregarious song and juvenile rats playing at high rates. Transcript levels for several glutamatergic receptor genes, such as GRIN1, GRIN2A, and GRIA1, were consistently upregulated in highly gregarious (i.e., playful/high singing) animals. This study is the first to directly investigate shared neuromodulators of positive, non-sexual social behaviors across songbirds and mammals. It provides insight into a conserved brain region that may regulate similar behaviors across vertebrates.

Re-wiring of the bonded brain: Gene expression among pair bonded female prairie voles changes as they transition to motherhood.

Motherhood is a costly life-history transition accompanied by behavioral and neural plasticity necessary for offspring care. Motherhood in the monogamous prairie vole is associated with decreased pair bond strength, suggesting a trade-off between parental investment and pair bond maintenance. Neural mechanisms governing pair bonds and maternal bonds overlap, creating possible competition between the two. We measured mRNA expression of genes encoding receptors for oxytocin (oxtr), dopamine (d1r and d2r), mu-opioids (oprm1a), and kappa-opioids (oprk1a) within three brain areas processing salience of sociosensory cues (anterior cingulate cortex; ACC), pair bonding (nucleus accumbens; NAc), and maternal care (medial preoptic area; MPOA). We compared gene expression differences between pair bonded prairie voles that were never pregnant, pregnant (~day 16 of pregnancy), and recent mothers (day 3 of lactation). We found greater gene expression in the NAc (oxtr, d2r, oprm1a, and oprk1a) and MPOA (oxtr, d1r, d2r, oprm1a, and oprk1a) following the transition to motherhood. Expression for all five genes in the ACC was greatest for females that had been bonded for longer. Gene expression within each region was highly correlated, indicating that oxytocin, dopamine, and opioids comprise a complimentary gene network for social signaling. ACC-NAc gene expression correlations indicated that being a mother (oxtr and d1r) or maintaining long-term pair bonds (oprm1a) relies on the coordination of different signaling systems within the same circuit. Our study suggests the maternal brain undergoes changes that prepare females to face the trade-off associated with increased emotional investment in offspring, while also maintaining a pair bond.

Chronic Astrocytic TNFα Production in the Preoptic-Basal Forebrain Causes Aging-like Sleep-Wake Disturbances in Young Mice.

Sleep disruption is a frequent problem of advancing age, often accompanied by low-grade chronic central and peripheral inflammation. We examined whether chronic neuroinflammation in the preoptic and basal forebrain area (POA-BF), a critical sleep-wake regulatory structure, contributes to this disruption. We developed a targeted viral vector designed to overexpress tumor necrosis factor-alpha (TNFα), specifically in astrocytes (AAV5-GFAP-TNFα-mCherry), and injected it into the POA of young mice to induce heightened neuroinflammation within the POA-BF. Compared to the control (treated with AAV5-GFAP-mCherry), mice with astrocytic TNFα overproduction within the POA-BF exhibited signs of increased microglia activation, indicating a heightened local inflammatory milieu. These mice also exhibited aging-like changes in sleep-wake organization and physical performance, including (a) impaired sleep-wake functions characterized by disruptions in sleep and waking during light and dark phases, respectively, and a reduced ability to compensate for sleep loss; (b) dysfunctional VLPO sleep-active neurons, indicated by fewer neurons expressing c-fos after suvorexant-induced sleep; and (c) compromised physical performance as demonstrated by a decline in grip strength. These findings suggest that inflammation-induced dysfunction of sleep- and wake-regulatory mechanisms within the POA-BF may be a critical component of sleep-wake disturbances in aging.

Identification of hypothermia-inducing neurons in the preoptic area and activation of them by isoflurane anesthesia and central injection of adenosine.

Hibernation and torpor are not passive responses caused by external temperature drops and fasting but are active brain functions that lower body temperature. A population of neurons in the preoptic area was recently identified as such active torpor-regulating neurons. We hypothesized that the other hypothermia-inducing maneuvers would also activate these neurons. To test our hypothesis, we first refined the previous observations, examined the brain regions explicitly activated during the falling phase of body temperature using c-Fos expression, and confirmed the preoptic area. Next, we observed long-lasting hypothermia by reactivating torpor-tagged Gq-expressing neurons using the activity tagging and DREADD systems. Finally, we found that about 40-60% of torpor-tagged neurons were activated by succeeding isoflurane anesthesia and by icv administration of an adenosine A1 agonist. Isoflurane-induced and central adenosine-induced hypothermia is, at least in part, an active process mediated by the torpor-regulating neurons in the preoptic area.

Effects of acute inhibition of dopamine ß-hydroxylase on neural responses to pups in adult virgin male California mice (Peromyscus californicus).

The neural mechanisms underlying paternal care in biparental mammals are not well understood. The California mouse (Peromyscus californicus) is a biparental rodent in which virtually all fathers are attracted to pups, while virgin males vary widely in their behavior toward unrelated infants, ranging from attacking to avoiding to huddling and grooming pups. We previously showed that pharmacologically inhibiting the synthesis of the neurotransmitter norepinephrine (NE) with the dopamine ß-hydroxylase inhibitor nepicastat reduced the propensity of virgin male and female California mice to interact with pups. The current study tested the hypothesis that nepicastat would reduce pup-induced c-Fos immunoreactivity, a cellular marker of neural activity, in the medial preoptic area (MPOA), medial amygdala (MeA), basolateral amygdala (BLA), and bed nucleus of the stria terminalis (BNST), brain regions implicated in the control of parental behavior and/or anxiety. Virgin males were injected with nepicastat (75 mg/kg, i.p.) or vehicle 2 hours prior to exposure to either an unrelated pup or novel object for 60 minutes (n = 4-6 mice per group). Immediately following the 60-minute stimulus exposure, mice were euthanized and their brains were collected for c-Fos immunohistochemistry. Nepicastat reduced c-Fos expression in the MeA and MPOA of pup-exposed virgin males compared to vehicle-injected controls. In contrast, nepicastat did not alter c-Fos expression in any of the above brain regions following exposure to a novel object. Overall, these results suggest that the noradrenergic system might influence MeA and MPOA function to promote behavioral interactions with pups in virgin males.

Microglia Caspase11 non-canonical inflammasome drives fever.

AIM: Animals exhibit physiological changes designed to eliminate the perceived danger, provoking similar symptoms of fever. However, a high-grade fever indicates poor clinical outcomes. Caspase11 (Casp11) is involved in many inflammatory diseases. Whether Casp11 leads to fever remains unclear. In this study, we investigate the role of the preoptic area of the hypothalamus (PO/AH) microglia Casp11 in fever. METHODS: We perform experiments using a rat model of LPS-induced fever. We measure body temperature and explore the functions of peripheral macrophages and PO/AH microglia in fever signaling by ELISA, immunohistochemistry, immunofluorescence, flow cytometry, macrophage depletion, protein blotting, and RNA-seq. Then, the effects of macrophages on microglia in a hyperthermic environment are observed in vitro. Finally, adeno-associated viruses are used to knockdown or overexpress microglia Casp11 in PO/AH to determine the role of Casp11 in fever. RESULTS: We find peripheral macrophages and PO/AH microglia play important roles in the process of fever, which is proved by macrophage and microglia depletion. By RNA-seq analysis, we find Casp11 expression in PO/AH is significantly increased during fever. Co-culture and conditioned-culture simulate the induction of microglia Casp11 activation by macrophages in a non-contact manner. Microglia Casp11 knockdown decreases body temperature, pyrogenic factors, and inflammasome, and vice versa. CONCLUSION: We report that Casp11 drives fever. Mechanistically, peripheral macrophages transmit immune signals via cytokines to microglia in PO/AH, which activate the Casp11 non-canonical inflammasome. Our findings identify a novel player, the microglia Casp11, in the control of fever, providing an explanation for the transmission and amplification of fever immune signaling.

Homeostatic regulation of rapid eye movement sleep by the preoptic area of the hypothalamus.

Rapid eye movement sleep (REMs) is characterized by activated electroencephalogram (EEG) and muscle atonia, accompanied by vivid dreams. REMs is homeostatically regulated, ensuring that any loss of REMs is compensated by a subsequent increase in its amount. However, the neural mechanisms underlying the homeostatic control of REMs are largely unknown. Here, we show that GABAergic neurons in the preoptic area of the hypothalamus projecting to the tuberomammillary nucleus (POAGAD2→TMN neurons) are crucial for the homeostatic regulation of REMs in mice. POAGAD2→TMN neurons are most active during REMs, and inhibiting them specifically decreases REMs. REMs restriction leads to an increased number and amplitude of calcium transients in POAGAD2→TMN neurons, reflecting the accumulation of REMs pressure. Inhibiting POAGAD2→TMN neurons during REMs restriction blocked the subsequent rebound of REMs. Our findings reveal a hypothalamic circuit whose activity mirrors the buildup of homeostatic REMs pressure during restriction and that is required for the ensuing rebound in REMs.

Single-nucleus RNA-seq identifies one galanin neuronal subtype in mouse preoptic hypothalamus activated during recovery from sleep deprivation.

The preoptic area of the hypothalamus (POA) is essential for sleep regulation. However, the cellular makeup of the POA is heterogeneous, and the molecular identities of the sleep-promoting cells remain elusive. To address this question, this study compares mice during recovery sleep following sleep deprivation to mice allowed extended sleep. Single-nucleus RNA sequencing (single-nucleus RNA-seq) identifies one galanin inhibitory neuronal subtype that shows upregulation of rapid and delayed activity-regulated genes during recovery sleep. This cell type expresses higher levels of growth hormone receptor and lower levels of estrogen receptor compared to other galanin subtypes. single-nucleus RNA-seq also reveals cell-type-specific upregulation of purinergic receptor (P2ry14) and serotonin receptor (Htr2a) during recovery sleep in this neuronal subtype, suggesting possible mechanisms for sleep regulation. Studies with RNAscope validate the single-nucleus RNA-seq findings. Thus, the combined use of single-nucleus RNA-seq and activity-regulated genes identifies a neuronal subtype functionally involved in sleep regulation.

Thermoregulatory pathway underlying the pyrogenic effects of prostaglandin E2 in the lateral parabrachial nucleus of male rats.

It has been shown that prostaglandin (PG) E2 synthesized in the lateral parabrachial nucleus (LPBN) is involved in lipopolysaccharide-induced fever. But the neural mechanisms of how intra-LPBN PGE2 induces fever remain unclear. In this study, we investigated whether the LPBN-preoptic area (POA) pathway, the thermoafferent pathway for feed-forward thermoregulatory responses, mediates fever induced by intra-LPBN PGE2 in male rats. The core temperature (Tcore) was monitored using a temperature radiotelemetry transponder implanted in rat abdomen. We showed that microinjection of PGE2 (0.28 nmol) into the LPBN significantly enhanced the density of c-Fos-positive neurons in the median preoptic area (MnPO). The chemical lesioning of MnPO with ibotenate or selective genetic lesioning or inhibition of the LPBN-MnPO pathway significantly attenuated fever induced by intra-LPBN injection of PGE2. We demonstrated that EP3 receptor was a pivotal receptor for PGE2-induced fever, since microinjection of EP3 receptor agonist sulprostone (0.2 nmol) or EP3 receptor antagonist L-798106 (2 nmol) into the LPBN mimicked or weakened the pyrogenic action of LPBN PGE2, respectively, but this was not the case for EP4 and EP1 receptors. Whole-cell recording from acute LPBN slices revealed that the majority of MnPO-projecting neurons originating from the external lateral (el) and dorsal (d) LPBN were excited and inhibited, respectively, by PGE2 perfusion, initiating heat-gain and heat-loss mechanisms. The amplitude but not the frequency of spontaneous and miniature glutamatergic excitatory postsynaptic currents (sEPSCs and mEPSCs) in MnPO-projecting LPBel neurons increased after perfusion with PGE2; whereas the frequency and amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs) and the A-type potassium (IA) current density did not change. In MnPO-projecting LPBd neurons, neither sEPSCs nor sIPSCs responded to PGE2; however, the IA current density was significantly increased by PGE2 perfusion. These electrophysiological responses and the thermoeffector reactions to intra-LPBN PGE2 injection, including increased brown adipose tissue thermogenesis, shivering, and decreased heat dissipation, were all abolished by L-798106, and mimicked by sulprostone. These results suggest that the pyrogenic effects of intra-LPBN PGE2 are mediated by both the inhibition of the LPBd-POA pathway through the EP3 receptor-mediated activation of IA currents and the activation of the LPBel-POA pathway through the selective enhancement of glutamatergic synaptic transmission via EP3 receptors.

Role of the median preoptic nucleus in the development of chronic deoxycorticosterone acetate (DOCA)-salt hypertension.

We have previously reported that the subfornical organ (SFO) does not contribute to the chronic hypertensive response to DOCA-salt in rats, and yet the organum vasculosum of the lamina terminalis (OVLT) plays a significant role in the development of deoxycorticosterone acetate (DOCA)-salt hypertension. Since efferent fibers of the OVLT project to and through the median preoptic nucleus (MnPO), the present study was designed to test the hypothesis that the MnPO is necessary for DOCA-salt hypertension in the rat. Male Sprague-Dawley rats underwent SHAM (MnPOsham; n = 5) or electrolytic lesion of the MnPO (MnPOx; n = 7) followed by subsequent unilateral nephrectomy and telemetry instrumentation. After recovery and during the experimental protocol, rats consumed a 0.1% NaCl diet and 0.9% NaCl drinking solution. Mean arterial pressure (MAP) was recorded telemetrically 5 days before and 21 days after DOCA implantation (100 mg/rat; SQ). The chronic pressor response to DOCA was attenuated in MnPOx rats by Day 11 of treatment and continued such that MAP increased 25 ± 3 mmHg in MnPOsham rats by Day 21 of DOCA compared to 14 ± 3 mmHg in MnPOx rats. These results support the hypothesis that the MnPO is an important brain site of action and necessary for the full development of DOCA-salt hypertension in the rat.

Astrocytes in preoptic area regulate acute nociception-induced hypothermia through adenosine receptors.

AIMS: The preoptic area (POA) of the hypothalamus, crucial in thermoregulation, has long been implicated in the pain process. However, whether nociceptive stimulation affects body temperature and its mechanism remains poorly studied. METHODS: We used capsaicin, formalin, and surgery to induce acute nociceptive stimulation and monitored rectal temperature. Optical fiber recording, chemical genetics, confocal imaging, and pharmacology assays were employed to confirm the role and interaction of POA astrocytes and extracellular adenosine. Immunofluorescence was utilized for further validation. RESULTS: Acute nociception could activate POA astrocytes and induce a decrease in body temperature. Manipulation of astrocytes allowed bidirectional control of body temperature. Furthermore, acute nociception and astrocyte activation led to increased extracellular adenosine concentration within the POA. Activation of adenosine A1 or A2A receptors contributed to decreased body temperature, while inhibition of these receptors mitigated the thermo-lowering effect of astrocytes. CONCLUSION: Our results elucidate the interplay between acute nociception and thermoregulation, specifically highlighting POA astrocyte activation. This enriches our understanding of physiological responses to painful stimuli and contributes to the analysis of the anatomical basis involved in the process.

Ventromedial hypothalamic nucleus subset stimulates tissue thermogenesis via preoptic area outputs.

OBJECTIVE: Hypothalamic signals potently stimulate energy expenditure by engaging peripheral mechanisms to restore energy homeostasis. Previous studies have identified several critical hypothalamic sites (e.g. preoptic area (POA) and ventromedial hypothalamic nucleus (VMN)) that could be part of an interconnected neurocircuit that controls tissue thermogenesis and essential for body weight control. However, the key neurocircuit that can stimulate energy expenditure has not yet been established. METHODS: Here, we investigated the downstream mechanisms by which VMN neurons stimulate adipose tissue thermogenesis. We manipulated subsets of VMN neurons acutely as well as chronically and studied its effect on tissue thermogenesis and body weight control, using Sf1Cre and Adcyap1Cre mice and measured physiological parameters under both high-fat diet and standard chow diet conditions. To determine the node efferent to these VMN neurons, that is involved in modulating energy expenditure, we employed electrophysiology and optogenetics experiments combined with measurements using tissue-implantable temperature microchips. RESULTS: Activation of the VMN neurons that express the steroidogenic factor 1 (Sf1; VMNSf1 neurons) reduced body weight, adiposity and increased energy expenditure in diet-induced obese mice. This function is likely mediated, at least in part, by the release of the pituitary adenylate cyclase-activating polypeptide (PACAP; encoded by the Adcyap1 gene) by the VMN neurons, since we previously demonstrated that PACAP, at the VMN, plays a key role in energy expenditure control. Thus, we then shifted focus to the subpopulation of VMNSf1 neurons that contain the neuropeptide PACAP (VMNPACAP neurons). Since the VMN neurons do not directly project to the peripheral tissues, we traced the location of the VMNPACAP neurons' efferents. We identified that VMNPACAP neurons project to and activate neurons in the caudal regions of the POA whereby these projections stimulate tissue thermogenesis in brown and beige adipose tissue. We demonstrated that selective activation of caudal POA projections from VMNPACAP neurons induces tissue thermogenesis, most potently in negative energy balance and activating these projections lead to some similar, but mostly unique, patterns of gene expression in brown and beige tissue. Finally, we demonstrated that the activation of the VMNPACAP neurons' efferents that lie at the caudal POA are necessary for inducing tissue thermogenesis in brown and beige adipose tissue. CONCLUSIONS: These data indicate that VMNPACAP connections with the caudal POA neurons impact adipose tissue function and are important for induction of tissue thermogenesis. Our data suggests that the VMNPACAP → caudal POA neurocircuit and its components are critical for controlling energy balance by activating energy expenditure and body weight control.

Activation of lateral preoptic neurons is associated with nest-building in male mice.

Nest-building behavior is a widely observed innate behavior. A nest provides animals with a secure environment for parenting, sleep, feeding, reproduction, and temperature maintenance. Since animal infants spend their time in a nest, nest-building behavior has been generally studied as parental behaviors, and the medial preoptic area (MPOA) neurons are known to be involved in parental nest-building. However, nest-building of singly housed male mice has been less examined. Here we show that male mice spent longer time in nest-building at the early to middle dark phase and at the end of the dark phase. These two periods are followed by sleep-rich periods. When a nest was removed and fresh nest material was introduced, both male and female mice built nests at Zeitgeber time (ZT) 6, but not at ZT12. Using Fos-immunostaining combined with double in situ hybridization of Vgat and Vglut2, we found that Vgat- and Vglut2-positive cells of the lateral preoptic area (LPOA) were the only hypothalamic neuron population that exhibited a greater number of activated cells in response to fresh nest material at ZT6, compared to being naturally awake at ZT12. Fos-positive LPOA neurons were negative for estrogen receptor 1 (Esr1). Both Vgat-positive and Vglut2-positive neurons in both the LPOA and MPOA were activated at pup retrieval by male mice. Our findings suggest the possibility that GABAergic and glutamatergic neurons in the LPOA are associated with nest-building behavior in male mice.

Computer vision analysis of mother-infant interaction identified efficient pup retrieval in V1b receptor knockout mice.

Close contact between lactating rodent mothers and their infants is essential for effective nursing. Whether the mother's effort to retrieve the infants to their nest requires the vasopressin-signaling via V1b receptor has not been fully defined. To address this question, V1b receptor knockout (V1bKO) and control mice were analyzed in pup retrieval test. Because an exploring mother in a new test cage randomly accessed to multiple infants in changing backgrounds over time, a computer vision-based deep learning analysis was applied to continuously calculate the distances between the mother and the infants as a parameter of their relationship. In an open-field, a virgin female V1bKO mice entered fewer times into the center area and moved shorter distances than wild-type (WT). While this behavioral pattern persisted in V1bKO mother, the pup retrieval test demonstrated that total distances between a V1bKO mother and infants came closer in a shorter time than with a WT mother. Moreover, in the medial preoptic area, parts of the V1b receptor transcripts were detected in galanin- and c-fos-positive neurons following maternal stimulation by infants. This research highlights the effectiveness of deep learning analysis in evaluating the mother-infant relationship and the critical role of V1b receptor in pup retrieval during the early lactation phase.