Arcuate Neurons Research Articles

Age- and sex-related differences of hypothalamic neuroendocrine center response to α-tocopherol acetate and thymalin preparation

The effect of thymalin preparation, α-tocopherol acetate, and stress on hypothalamic centers related to the regulation of the gonadotropic function and age-specific features of such influence have been investigated in experiment. Experimental results indicate the presence of the reaction of neurons of the rostral preoptic area and arcuate nucleus for the experimental effects. The age-specific feature of the response of neurons of the rostral preoptic area and arcuate nucleus of the hypothalamus of white rats to α-tocopherol acetate injection is a reduction in the degree of response. Thymalin reduces the threshold of sensitivity of arcuate center neurons to the action of stress.

Integration Between Different Hypothalamic Nuclei Involved in Stress and GnRH Secretion in the Ewe

This study investigated possible integrated links in the neuroanatomical pathways through which the activity of neurones in the paraventricular nucleus and arcuate nucleus may modulate suppression of gonadotrophin-releasing hormone (GnRH) secretion during stressful situations. Double-label immunofluorescence and laser scanning confocal microscopy were used to examine the hypothalamic sections from the follicular phase ewes. Noradrenergic terminals were in close contact with 65.7 ± 6.1% corticotrophin-releasing hormone (CRH) and 84.6 ± 3.2% arginine vasopressin (AVP) cell bodies in the paraventricular nucleus but not with β-endorphin cell bodies in the arcuate nucleus. Furthermore, γ-amino butyric acid (GABA) terminals were close to 80.9 ± 3.5% CRH but no AVP cell bodies in the paraventricular nucleus, as well as 60.8 ± 4.1%β-endorphin cell bodies in the arcuate nucleus. Although CRH, AVP and β-endorphin cell terminals were identified in the medial pre-optic area, no direct contacts with GnRH cell bodies were observed. Within the median eminence, abundant CRH but not AVP terminals were close to GnRH cell terminals in the external zone; whereas, β-endorphin cells and terminals were in the internal zone. In conclusion, neuroanatomical evidence is provided for the ewe supporting the hypothesis that brainstem noradrenergic and hypothalamic GABA neurones are important in modulating the activity of CRH and AVP neurones in the paraventricular nucleus, as well as β-endorphin neurones in the arcuate nucleus. These paraventricular and arcuate neurones may also involve interneurones to influence GnRH cell bodies in medial pre-optic area, whereas the median eminence may provide a major site for direct modulation of GnRH release by CRH terminals.

Evidence of hypothalamic degeneration in the anorectic anx/anx mouse

Mice homozygous for the anorexia (anx) mutation are characterized by poor food intake and death by three to five weeks after birth. By P21 these mice display lower density of hypothalamic neuropeptides, including Agouti gene-related protein (AGRP). The AGRP/neuropeptide Y (NPY) system of the anx/anx mice develops normally until postnatal day (P) 12, then the normal increase in fiber density ceases, in some areas even distinctly decreases. This overlaps with activation of microglia, indicating an inflammatory and/or degenerative process. Here we studied, by in situ hybridization and immunohistochemistry (IHC), the expression of major histocompatibility complex (MHC) class I-related molecules and markers for cellular reactivity in hypothalamus of anx/anx mice. MHC class I transcript and -related proteins were found in arcuate nucleus (Arc), presumably both in neurons and glia, the latter also in areas innervated by AGRP (NPY) neurons. In the anx/anx hypothalamus, using TUNEL labeling, significantly higher number of apoptotic cells were found compared with +/+ mice, and active caspase 6 immunoreactivity was detected in degenerating NPY-fibers as well as signs of "microglia-associated cell death". In addition, Y1 receptor-labeled processes and soma of pro-opiomelanocortin (POMC) neurons, were markedly decreased at P21. These results support the hypothesis of degeneration of hypothalamic arcuate neuron populations in the anx/anx mice, whereby the AGRP system may be first affected, the changes in the POMC system being secondary in this process.

An arcuate-ventrolateral periaqueductal gray reciprocal circuit participates in electroacupuncture cardiovascular inhibition

Electroacupuncture (EA) suppresses elevated blood pressure (BP) by activating the arcuate nucleus, ventrolateral periaqueductal gray (vlPAG), and inhibiting cardiovascular sympathetic neurons in the rostral ventrolateral medulla. This study investigated the reciprocal neural circuit between arcuate and vlPAG during EA inhibition of reflex increases in blood pressure. In α-chloralose anesthetized cats the gallbladder or splanchnic nerve was stimulated to induce cardiovascular sympathoexcitatory reflexes. Electrophysiological recordings showed that EA facilitates the arcuate neural response to splanchnic nerve stimulation. Bilateral vlPAG microinjection of d, l-homocysteic acid (DLH) facilitated the arcuate response to splanchnic nerve stimulation, while microinjection of kainic acid blocked EA (P 5-6 acupoints on pericardial meridian, overlying the median nerves) excitation of arcuate neurons. Retrograde microsphere tracer labeling in the arcuate or vlPAG perikarya was found after respective microinjection of the tracer in the vlPAG or arcuate of rats, demonstrating reciprocal direct connections between the two nuclei. EA inhibition of reflex-induced BP elevation was blocked by injection of glutamate or cholinergic receptor antagonist, kynurenic acid or atropine, into the arcuate. Excitation of vlPAG neurons during EA was blocked by arcuate microinjection of glutamate NMDA and non-NMDA receptor antagonists, AP-5 and CNQX, or the cholinergic receptor antagonist, atropine. Microinjection of DLH or acetylcholine (ACh) into the arcuate facilitated EA excitation of vlPAG neurons. Microinjection of AP5 and CNQX, but not atropine, into the vlPAG blocked EA excitation of arcuate neurons. Thus, a reciprocal excitatory glutamatergic neural circuit between the arcuate and vlPAG contributes to long-lasting EA cardiovascular inhibition. ACh in the arcuate but not in the vlPAG participates in the reciprocal excitation.

ATP-Sensitive Potassium Channel-Mediated Lactate Effect on Orexin Neurons: Implications for Brain Energetics during Arousal

Active neurons have a high demand for energy substrate, which is thought to be mainly supplied as lactate by astrocytes. Heavy lactate dependence of neuronal activity suggests that there may be a mechanism that detects and controls lactate levels and/or gates brain activation accordingly. Here, we demonstrate that orexin neurons can behave as such lactate sensors. Using acute brain slice preparations and patch-clamp techniques, we show that the monocarboxylate transporter blocker alpha-cyano-4-hydroxycinnamate (4-CIN) inhibits the spontaneous activity of orexin neurons despite the presence of extracellular glucose. Furthermore, fluoroacetate, a glial toxin, inhibits orexin neurons in the presence of glucose but not lactate. Thus, orexin neurons specifically use astrocyte-derived lactate. The effect of lactate on firing activity is concentration dependent, an essential characteristic of lactate sensors. Furthermore, lactate disinhibits and sensitizes these neurons for subsequent excitation. 4-CIN has no effect on the activity of some arcuate neurons, indicating that lactate dependency is not universal. Orexin neurons show an indirect concentration-dependent sensitivity to glucose below 1 mm, responding by hyperpolarization, which is mediated by ATP-sensitive potassium channels composed of Kir6.1 and SUR1 subunits. In conclusion, our study suggests that lactate is a critical energy substrate and a regulator of the orexin system. Together with the known effects of orexins in inducing arousal, food intake, and hepatic glucose production, as well as lactate release from astrocytes in response to neuronal activity, our study suggests that orexin neurons play an integral part in balancing brain activity and energy supply.

Reduced fasting-induced activation of hypothalamic arcuate neurons is associated with hyperleptinemia and increased leptin sensitivity in obese mice

Fasting increases c-Fos expression in neuropeptide Y (NPY) neurons of the hypothalamic arcuate nucleus (ARC) in lean, but not in hyperleptinemic mice with late-onset obesity (LOO). Although obesity is associated with leptin resistance, we hypothesized that under fasting conditions, leptin sensitivity might be restored and that hyperleptinemia may counteract the neuronal response to fasting. We investigated whether the reduced fasting response of ARC neurons in LOO is paralleled by an increase in leptin sensitivity, as measured by leptin-induced STAT-3 phosphorylation. To assess leptin's role in the modulation of the fasting-induced ARC activation, we investigated c-Fos responses and hormone and metabolite levels in hyperleptinemic diet-induced obese (DIO) and in leptin-deficient ob/ob mice. Leptin induced a stronger STAT-3 phosphorylation in fasted LOO and lean mice than in ad libitum-fed animals. Similar to LOO, hyperleptinemic DIO mice showed no c-Fos response after fasting, while ob/ob mice showed a stronger response than lean control mice. Mimicking hyperleptinemia by repeated leptin injections in lean mice during fasting attenuated the fasting-induced c-Fos expression. Our findings indicate that high leptin levels prevent the fasting-induced activation of ARC neurons in mice. Moreover, leptin sensitivity is dynamic in obese subjects and depends on the feeding status. During short-term increases in leptin sensitivity, e.g., during fasting, leptin signaling appears to be effective, even in hyperleptinemic obesity. As reflected by the blockade of the fasting-induced ARC activation, fasting seems to interfere with the responsiveness of the ARC to signals related to the status of energy intake.

Altered meal patterns in CCK1R−/− mice. Role of ghrelin receptors expressed on vagal afferents and activation of arcuate neurons

Altered meal patterns in CCK1R−/− mice. Role of ghrelin receptors expressed on vagal afferents and activation of arcuate neurons

The role of the NOP receptor in regulating food intake, meal pattern, and the excitability of proopiomelanocortin neurons

We evaluated the role of the nociceptin/orphanin FQ (NOP) receptor in regulating food intake, meal pattern and the activity of hypothalamic arcuate (ARC) neurons. The microstructural analysis of food intake and meal pattern was performed under both food-deprived and ad libitum conditions. Whole-cell patch clamp recordings were obtained using the in vitro hypothalamic slice preparation and biocytin-filled electrodes. NOP receptor knockout mice exhibited significantly reduced body weight. Fasting-induced hyperphagia was diminished for the first 2h of a 6-h re-feeding period, and was associated with decreased meal duration and size, as well as a biphasic effect on meal frequency. The genotype effect observed under ad libitum conditions was comparatively unremarkable. Orphanin FQ/nociceptin (OFQ/N) was able to decrease evoked excitatory postsynaptic current amplitude, increase the S(2):S(1) ratio via the paired-pulse paradigm, and decrease miniature excitatory postsynaptic current frequency in ARC neurons from wild type animals but not NOP receptor knockouts. In addition OFQ/N activated a reversible outward current that was antagonized by the G-protein activated, inwardly-rectifying K(+) (GIRK) channel blocker tertiapin in wild type but not NOP knockout animals. Both the presynaptic and postsynaptic actions of OFQ/N were observed in ARC neurons subsequently determined to be immunopositive for characteristic phenotypic markers of anorexigenic proopiomelanocortin (POMC) neurons. Taken together, these results demonstrate the contribution of the NOP receptor in controlling food intake and meal pattern, as well as glutamate release and GIRK1 channel activity at POMC synapses.

Direct and Indirect Effects of Cannabinoids on in vitro GABA Release in the Rat Arcuate Nucleus

Within the hypothalamic arcuate nucleus, two neuronal subpopulations play particularly important roles in energy balance; neurones expressing neuropeptide Y (NPY), agouti-related peptide (AgRP) and GABA are orexigenic, whereas neurones expressing pro-opiomelanocortin and CART are anorexigenic. The pivotal role of these neuropeptides in energy homeostasis is well-known, although GABA may also be an important signal because targeted knockout of the GABA transporter in NPY/AgRP/GABA neurones results in a lean, obesity-resistant phenotype. In the present study, we describe an in vitro model of K(+)-evoked GABA release from the hypothalamus and determine the effects of cannabinoid receptor activation. K(+)-evoked GABA release was sensitive to leptin, insulin and PYY(3-36), indicating that GABA was released by arcuate NPY/AgRP/GABA neurones. In the presence of tetrodotoxin (TTX), the cannabinoid CB1 receptor agonist WIN 55,212-2 inhibited K(+)-evoked GABA release. This was prevented by the CB1 receptor inverse agonist rimonabant. Rimonabant had no effect when applied alone. In the absence of TTX, however, the opposite effects were observed: WIN 55,212-2 had no effect while rimonabant inhibited GABA release. This indicates that GABA release can involve an indirect, TTX-sensitive mechanism. The most parsimonious explanation for the inhibition of GABA release by a CB receptor inverse agonist is via the disinhibition of an cannabinoid-sensitive inhibitory input onto GABAergic neurones. One local source of an inhibitory neurotransmitter is the opioidergic arcuate neurones. In our in vitro model, K(+)-evoked GABA release was inhibited by the endogenous opioid peptide beta-endorphin in a naloxone-sensitive manner. The inhibitory effect of rimonabant was also prevented by naloxone and a kappa-opioid receptor selective antagonist, suggesting that GABA release from arcuate NPY/AgRP/GABA neurones can be inhibited by endogenous opioid peptides, and that the release of opioid peptides is sensitive to cannabinoids.

Electrophysiological effect of ghrelin and somatostatin on rat hypothalamic arcuate neurons in vitro

Growth hormone (GH) secretion from the pituitary gland is partly regulated by GH releasing hormone (GHRH)-containing neurons located in the hypothalamic arcuate nucleus (ARC). GHRH-containing neurons express the GH secretagogue (GHS) receptor (GHS-R) and the somatostatin (SRIF) receptor. Recently, an endogenous ligand for the GHS-R named ghrelin was found. Therefore, it seems that both ghrelin and SRIF are involved in the hypothalamic regulation of GH release via GHRH-containing neurons in the ARC. In extracellular single unit recordings from in vitro hypothalamic slice preparations from rats, application of 100 nM ghrelin substantially excited ARC neurons (82.5%), whereas 1 μM SRIF substantially inhibited them (81.8%). The ghrelin-induced excitatory and SRIF-induced inhibitory effects on ARC neurons were dose-dependent and persisted during synaptic blockade using low-Ca 2+/high-Mg 2+ solution. In addition, the effects were antagonized by [D-Lys 3]-GHRP-6, a GHS-R antagonist, and CYN154806, a SRIF receptor subtype sst2 antagonist, respectively. When ghrelin and SRIF were sequentially applied to ARC neurons, 95.2% were excited by ghrelin and inhibited by SRIF. Similarly, 85.0% of ARC neuroendocrine cells that project to the median eminence were excited by ghrelin and inhibited by SRIF. These results indicate that ARC neuroendocrine cells projecting to the median eminence are dose-dependently, postsynaptically and oppositely regulated by ghrelin through GHS-R and SRIF via the SRIF sst2 receptor subtype. Our results also suggest that most of these ARC neuroendocrine cells are presumably GHRH-containing neurons and are involved in the cellular processes through which ghrelin and SRIF participate in the hypothalamic regulation of GH release.

Effects of glucagon-like peptide 1 and oxyntomodulin on neuronal activity of ghrelin-sensitive neurons in the hypothalamic arcuate nucleus

Glucagon-like peptide 1 (GLP-1) and oxyntomodulin (OXM) are structurally related gastrointestinal hormones that are secreted in response to food intake. They reduce food intake and body weight and exert partly overlapping actions on glucose homeostasis and gastrointestinal function. The hypothalamic arcuate (ARC) nucleus is among the central structures expressing a high density of GLP-1 receptors (GLP-1R), which are known to be activated by both peptides. It was the aim of our electrophysiological studies to characterize the effects of GLP-1 and OXM on functionally defined ghrelin-sensitive ARC neurons. GLP-1 and OXM (10(-7) M) exerted excitatory effects in about two-thirds of ghrelin-inhibited neurons and in approximately one-third of ghrelin-excited cells. In addition, a minor fraction of the ghrelin-excited cells was inhibited by both peptides. There was a high degree of cosensitivity to GLP-1 and OXM, and the effects of both hormones were blocked by the GLP-1R antagonist exendin(9-39). The GLP-1R-mediated excitations and inhibitions persisted under synaptic blockade, indicating a direct postsynaptic mode of action. Our results demonstrate that GLP-1 and OXM directly and similarly alter neuronal activity in the ARC, probably via a common GLP-1R-mediated mechanism. Ghrelin-antagonistic effects on neuronal activity, which might be implicated in ghrelin-antagonistic in vivo actions, resulting from GLP-1R stimulation (e.g., GLP-1R dependent supression of food intake), predominated in ghrelin-inhibited ARC neurons. However, a subset of ghrelin-excited ARC neurons showed responses to OXM or GLP-1, suggesting the existence of a common mode of action for these hormones; the functional relevance of this effect remains to be elucidated.

Glut2‐dependent glucose‐sensing controls thermoregulation by enhancing the leptin sensitivity of NPY and POMC neurons

The physiological contribution of glucose in thermoregulation is not completely established nor whether this control may involve a regulation of the melanocortin pathway. Here, we assessed thermoregulation and leptin sensitivity of hypothalamic arcuate neurons in mice with inactivation of glucose transporter type 2 (Glut2)-dependent glucose sensing. Mice with inactivation of Glut2-dependent glucose sensors are cold intolerant and show increased susceptibility to food deprivation-induced torpor and abnormal hypothermic response to intracerebroventricular administration of 2-deoxy-d-glucose compared to control mice. This is associated with a defect in regulated expression of brown adipose tissue uncoupling protein I and iodothyronine deiodinase II and with a decreased leptin sensitivity of neuropeptide Y (NPY) and proopiomelanocortin (POMC) neurons, as observed during the unfed-to-refed transition or following i.p. leptin injection. Sites of central Glut-2 expression were identified by a genetic tagging approach and revealed that glucose-sensitive neurons were present in the lateral hypothalamus, the dorsal vagal complex, and the basal medulla but not in the arcuate nucleus. NPY and POMC neurons were, however, connected to nerve terminals from Glut2-expressing neurons. Thus, our data suggest that glucose controls thermoregulation and the leptin sensitivity of NPY and POMC neurons through activation of Glut2-dependent glucose-sensing neurons located outside of the arcuate nucleus.

Synchronized Network Oscillations in Rat Tuberoinfundibular Dopamine Neurons: Switch to Tonic Discharge by Thyrotropin-Releasing Hormone

The pituitary hormone, prolactin, triggers lactation in nursing mothers. Under nonlactating conditions, prolactin secretion is suppressed by powerful inhibition from hypothalamic tuberoinfundibular dopamine (TIDA) neurons. Although firing pattern has been suggested as integral to neuroendocrine control, the electrical behavior of TIDA cells remains unknown. We demonstrate that rat TIDA neurons discharge rhythmically in a robust 0.05 Hz oscillation. The oscillation is phase locked between neurons, and while it persists during chemical synaptic transmission blockade, it is abolished by gap junction antagonists. Thyrotropin-releasing hormone (TRH) potently stimulates prolactin release, an effect assumed to take place in the pituitary. In TIDA cells, TRH caused a transition from phasic to tonic firing through combined pre- and postsynaptic effects. These findings suggest a model for prolactin regulation where a TIDA network switch from oscillations to sustained discharge converts dopamine from an antagonist at high concentrations to a functional agonist as dopamine output from the network decreases.

Forebrain projections of arcuate neurokinin B neurons demonstrated by anterograde tract-tracing and monosodium glutamate lesions in the rat

Neurokinin B (NKB) and kisspeptin receptor signaling are essential components of the reproductive axis. A population of neurons resides within the arcuate nucleus of the rat that expresses NKB, kisspeptin, dynorphin, NK3 receptors and estrogen receptor α (ERα). Here we investigate the projections of these neurons using NKB-immunocytochemistry as a marker. First, the loss of NKB-immunoreactive (ir) somata and fibers was characterized after ablation of the arcuate nucleus by neonatal injections of monosodium glutamate. Second, biotinylated dextran amine was injected into the arcuate nucleus and anterogradely labeled NKB-ir fibers were identified using dual-labeled immunofluorescence. Four major projection pathways are described: (1) local projections within the arcuate nucleus bilaterally, (2) projections to the median eminence including the lateral palisade zone, (3) projections to a periventricular pathway extending rostrally to multiple hypothalamic nuclei, the septal region and BNST and dorsally to the dorsomedial nucleus and (4) Projections to a ventral hypothalamic tract to the lateral hypothalamus and medial forebrain bundle. The diverse projections provide evidence that NKB/kisspeptin/dynorphin neurons could integrate the reproductive axis with multiple homeostatic, behavioral and neuroendocrine processes. Interestingly, anterograde tract-tracing revealed NKB-ir axons originating from arcuate neurons terminating on other NKB-ir somata within the arcuate nucleus. Combined with previous studies, these experiments reveal a bilateral interconnected network of sex-steroid responsive neurons in the arcuate nucleus of the rat that express NKB, kisspeptin, dynorphin, NK3 receptors and ERα and project to GnRH terminals in the median eminence. This circuitry provides a mechanism for bilateral synchronization of arcuate NKB/kisspeptin/dynorphin neurons to modulate the pulsatile secretion of GnRH.

228: Brain slice electrophysiology of arcuate neurons: evidence for the neonatal development of appetite pathways

The adipocyte-derived hormone leptin is a potent neurotrophic factor during fetal and neonatal development. In mature adults, leptin acts acutely on the brain to reduce food intake by regulating the activity of hypothalamic neurons, primarily in the arcuate nucleus (ARC). However, the functional impact of leptin on specific ARC neurons during developmental periods is not clear. We investigated the effects of leptin on ARC neurons during neonatal rats.

Endogenous ACTH, not only α-melanocyte-stimulating hormone, reduces food intake mediated by hypothalamic mechanisms

ACTH and alpha-melanocyte-stimulating hormone (alpha-MSH) are both consecutively processed from proopiomelanocortin (POMC), which is synthesized in hypothalamic arcuate neurons innervating the paraventricular nuclei (PVN). POMC secretion/synthesis is regulated by energy availability. ACTH and alpha-MSH bind with equal affinity to melanocortin-4 receptors and elicit similar effects on signal transduction in-vitro. Endogenous alpha-MSH thus far is believed to be the major physiological agonist and to act in an anorexigenic manner. Until now, it was fully unknown whether endogenous ACTH is also involved in the regulation of appetite and food intake. In this study in rats, we now show that icv ACTH as well as alpha-MSH possess anorexigenic effects in the PVN or areas in close proximity in vivo and that the effect of ACTH is direct and not mediated via alpha-MSH. We investigated the roles of endogenous ACTH and alpha-MSH by PVN application of the respective antibodies under different physiological conditions. In satiated rats with high levels of ACTH and alpha-MSH in the PVN, antibody administration increased food intake and body weight gain; hungry animals were unaffected. Finally, repeated injections of ACTH antibodies into PVN resulted in persistently increased food intake during the light period. These data now provide robust evidence that endogenous ACTH without further processing acts in the PVN or areas in close proximity to reduce food intake under conditions of feeding-induced satiety.

A direct neuronal connection between the subparafascicular and ventrolateral arcuate nuclei in non-lactating female rats. Could this pathway play a role in the suckling-induced prolactin release?

The neuronal pathways, through which prolactin secretion is regulated during lactation, have still not been fully explored. Studies indicate that the suckling stimulus travels through the spinal cord, the brain stem, and then reaches the hypothalamus. The focus of this present experiment is to further explore the neuronal connections between the brain stem and the arcuate nucleus that may be involved in suckling-induced prolactin release. Ante- and retrograde tracing techniques were used. To chemically characterize the explored neurons neuropeptide immunohistochemistry was applied. Previous studies have indicated that the peripeduncular nucleus is a relay of the suckling stimulus in the midbrain, conveying the information to the hypothalamus. In our experiments, we have found an additional cell group in the subparafascicular parvocellular nucleus located just behind the posterior thalamus that projects to the arcuate neurons. The injection of the retrograde tracer into the ventrolateral part of the arcuate nucleus labeled cells in the lateral subdivision of the subparafascicular parvocellular nucleus. Anterograde tracing from the subparafascicular parvocellular nucleus resulted in fiber labeling in the arcuate nucleus in close apposition with dynorphin immunopositive neurons. Double labeling revealed that a subpopulations of the subparafascicular parvocellular neurons projecting to the arcuate nucleus contained tuberoinfundibular peptide of 39 residues or calcitonin gene-related peptide. The presented findings suggest that the ascending fibers from the subparafascicular parvocellular nucleus might be in the pathway involved in the suckling-induced prolactin release.

Effect of anorexinergic peptides, cholecystokinin (CCK) and cocaine and amphetamine regulated transcript (CART) peptide, on the activity of neurons in hypothalamic structures of C57Bl/6 mice involved in the food intake regulation

The hypothalamus plays an important role in food consumption, receiving information about energy balance via hormonal, metabolic, and neural inputs. Its neurons produce neuropeptides influencing energy balance. Especially important to regulation of food consumption are certain hypothalamic structures, including the arcuate (ARC) and ventromedial (VMN) nuclei and the lateral hypothalamic area (LHA). We determined the impact of cholecystokinin (CCK) and cocaine and amphetamine regulated transcript (CART) peptide, on activity of ARC and VMN neurons and hypocretin (Hcrt) synthesizing neurons in LHA. ARC is an integrative nucleus regulating food consumption, VMN is considered to be a satiety centre, and LHA a hunger sensing centre. After overnight fasting, male C57Bl/6 mice received intraperitoneal injection of (i.p.) saline (SAL) or CCK (4μg/kg) or intracerebroventricular injection of (i.c.v.) CART peptide (0.1μg/mice) or CCK (i.p.) followed by CART peptide (i.c.v.) 5min later. Sixty minutes later, the presence of Fos or Fos/Hcrt immunostaining indicated activity of ARC and VMN neurons, as well as of Hcrt cells in LHA. CCK alone did not influence neuronal activity in any of the nuclei studied. CART peptide stimulated neurons in ARC and VMN (p<0.01) but decreased Hcrt neuronal activity in LHA (p<0.05). Co-administration of both peptides synergistically stimulated ARC neurons (p<0.01) and asynergistically inhibited LHA Hcrt neurons (p<0.01). Results indicate that CCK may modify the effect of CART peptide and thus substantially influence activity of neurons in hypothalamic structures involved in regulation of food intake.

Estrogen rapidly attenuates cannabinoid-induced changes in energy homeostasis

We examined whether estrogen negatively modulates cannabinoid-induced regulation of food intake, core body temperature and neurotransmission at proopiomelanocortin (POMC) synapses. Food intake was evaluated in ovariectomized female guinea pigs abdominally implanted with thermal DataLoggers and treated s.c. with the cannabinoid CB 1/CB 2 receptor agonist WIN 55,212-2, the CB 1 receptor antagonist AM251 or their cremephor/ethanol/0.9% saline vehicle, and with estradiol benzoate (EB) or its sesame oil vehicle. Whole-cell patch clamp recordings were performed in slices through the arcuate nucleus. WIN 55,212-2 produced dose- and time-dependent increases in food intake. EB decreased food intake 8–24 h after administration, but rapidly and completely blocked the increase in consumption caused by WIN 55,212-2. EB also attenuated the WIN 55,212-2-induced decrease in core body temperature. The AM251-induced decrease in food intake was unaffected. The diminution of the WIN 55,212-2-induced increase in food intake caused by EB correlated with a marked attenuation of cannabinoid receptor-mediated decreases in glutamatergic miniature excitatory postsynaptic current frequency occurring within 10–15 min of steroid application. Furthermore, EB completely blocked the depolarizing shift in the inactivation curve for the A-type K + current caused by WIN 55,212-2. The EB-mediated, physiologic antagonism of these presynaptic and postsynaptic actions elicited upon cannabinoid receptor activation was observed in arcuate neurons immunopositive for phenotypic markers of POMC neurons. These data reveal that estrogens negatively modulate cannabinoid-induced changes in appetite, body temperature and POMC neuronal activity. They also impart insight into the neuroanatomical substrates and effector systems upon which these counter-regulatory factors converge in the control of energy homeostasis.

A Serotonin-Dependent Mechanism Explains the Leptin Regulation of Bone Mass, Appetite, and Energy Expenditure

Leptin inhibition of bone mass accrual requires the integrity of specific hypothalamic neurons but not expression of its receptor on these neurons. The same is true for its regulation of appetite and energy expenditure. This suggests that leptin acts elsewhere in the brain to achieve these three functions. We show here that brainstem-derived serotonin (BDS) favors bone mass accrual following its binding to Htr2c receptors on ventromedial hypothalamic neurons and appetite via Htr1a and 2b receptors on arcuate neurons. Leptin inhibits these functions and increases energy expenditure because it reduces serotonin synthesis and firing of serotonergic neurons. Accordingly, while abrogating BDS synthesis corrects the bone, appetite and energy expenditure phenotypes caused by leptin deficiency, inactivation of the leptin receptor in serotonergic neurons recapitulates them fully. This study modifies the map of leptin signaling in the brain and identifies a molecular basis for the common regulation of bone and energy metabolisms. For a video summary of this article, see the PaperFlick file with the Supplemental Data available online.