Rat Hypothalamus Research Articles

Involvement of <i>Bdnf</i>, <i>Ntrk2</i> and <i>Pi3k</i> in the mechanism of binge eating after psychogenic stressors in ontogenesis

BACKGROUND: The study of the neurochemical mechanisms of food addiction provides experimental modeling of some of its clinical manifestations. AIM: This study aimed to examine the effect of binge eating after maternal deprivation or after rearing in social isolation on the expression of Bdnf, Ntrk2, and Pi3k in the hypothalamus of rats. MATERIALS AND METHODS: Animals aged 2–12 days were weaned from their mother for 10 days at 180 min, and males aged 90–100 days were used in the experiments. Another group of animals was reared in individual cages from day 21 after birth, and males aged 90–100 days were used in the experiments. To induce binge eating, the animals received a high-carbohydrate feed (chocolate spread) for 1 h every day or every third day within 30 days. Fifteen minutes before feeding, the paste was placed 5 cm within visual contact. RESULTS: In groups with intermittent exposure to high-calorie food (the animals received pasta every third day), polymerase chain reaction analysis revealed the expression of the Bdnf, Ntrk2, and Pi3k in the hypothalamus. The expression level of Bdnf was higher in the maternal deprivation group than in the control group. The expression levels of Ntrk2 and Pi3k in rats taking a high-carbohydrate feed were higher in animals reared in isolation than in those reared in the community. CONCLUSIONS: The results present new pathways for the synthesis of peptide drugs associated with the PI3K/AKT/mTOR signaling pathway for the correction of food addiction caused by psychogenic stress in ontogenesis.

Moderate aerobic training enhances the effectiveness of insulin therapy through hypothalamic IGF1 signaling in rat model of Alzheimer's disease

Alzheimer's disease (AD) is a neurological condition that is connected with a decline in a person's memory as well as their cognitive ability. One of the key topics of AD research has been the exploration of metabolic causes. We investigated the effects of treadmill exercise and intranasal insulin on learning and memory impairment and the expression of IGF1, BDNF, and GLUT4 in hypothalamus. The animals were put into 9 groups at random. In this study, we examined the impact of insulin on spatial memory in male Wistar rats and analyzed the effects of a 4-week pretreatment of moderate treadmill exercise and insulin on the mechanisms of improved hypothalamic glucose metabolism through changes in gene and protein expression of IGF1, BDNF, and GLUT4. We discovered that rat given Aβ25–35 had impaired spatial learning and memory, which was accompanied by higher levels of Aβ plaque burden in the hippocampus and lower levels of IGF1, BDNF, and GLUT4 mRNA and protein expression in the hypothalamus. Additionally, the administration of exercise training and intranasal insulin results in the enhancement of spatial learning and memory impairments, the reduction of plaque burden in the hippocampus, and the enhancement of the expression of IGF1, BDNF, and GLUT4 in the hypothalamus of rats that were treated with Aβ25–35. Our results show that the improvement of learning and spatial memory due to the improvement of metabolism and upregulation of the IGF1, BDNF, and GLUT4 pathways can be affected by pretreatment exercise and intranasal insulin.

Transgenerational effect of prenatal stress on behavior and lipid peroxidation in brain structures of female rats during the estral cycle.

The effect of stress in pregnant female Wistar rats on the behavior and lipid peroxidation (LP) in the neocortex, hippocampus and hypothalamus in the female F2 generation during the ovarian cycle was investigated. We subjected pregnant females to daily 1-hour immobilization stress from the 15th to the 19th days of pregnancy. Further, family groups were formed from prenatally stressed and control male and female rats of the F1 generation: group 1, the control female and male; group 2, the control female and the prenatally stressed male; group 3, the prenatally stressed female and the control male; group 4, the prenatally stressed female and male. The females of the F2 generation born from these couples were selected into four experimental groups in accordance with the family group. At the age of 3 months, behavior of rats was studied in the "open field" test in two stages of the ovarian cycle - estrus and diestrus. After 7-10 days, the rats were decapitated and the neocortex, hypothalamus and hippocampus were selected to determine the level of diene and triene conjugates, Schiff bases and the degree of lipid oxidation (Klein index). In F2 females with one prenatally stressed parent, there was no interstage difference in locomotor-exploratory activity and anxiety. If both F1 parents were prenatally stressed, female F2 rats retained interstage differences similar to the control pattern, while their locomotor-exploratory activity and time spent in the center of an "open field" decreased in absolute values. In the neocortex of F2 females in groups with prenatally stressed mothers, the level of primary LP products decreased and the level of Schiff bases increased in the estrus stage. In the hippocampus of F2 females in the groups with prenatally stressed fathers, the level of Schiff bases decreased in the estrus stage, and the level of primary LP products increased in group 2 and decreased in group 4. In the hypothalamus of F2 females in the groups with prenatally stressed mothers, the level of Schiff bases increased in the estrus stage and decreased in the diestrus; in addition, in group 3, the level of primary LP products in the estrus stage increased. Thus, we demonstrated the influence of prenatal stress of both F1 mother and F1 father on the behavior and the level of LP in the neocortex, hippocampus and hypothalamus in female rats of the F2 generation in estrus and diestrus.

Morphological Signatures of Neurogenesis and Neuronal Migration in Hypothalamic Vasopressinergic Magnocellular Nuclei of the Adult Rat.

The arginine vasopressin (AVP)-magnocellular neurosecretory system (AVPMNS) in the hypothalamus plays a critical role in homeostatic regulation as well as in allostatic motivational behaviors. However, it remains unclear whether adult neurogenesis exists in the AVPMNS. By using immunoreaction against AVP, neurophysin II, glial fibrillar acidic protein (GFAP), cell division marker (Ki67), migrating neuroblast markers (doublecortin, DCX), microglial marker (Ionized calcium binding adaptor molecule 1, Iba1), and 5'-bromo-2'-deoxyuridine (BrdU), we report morphological evidence that low-rate neurogenesis and migration occur in adult AVPMNS in the rat hypothalamus. Tangential AVP/GFAP migration routes and AVP/DCX neuronal chains as well as ascending AVP axonal scaffolds were observed. Chronic water deprivation significantly increased the BrdU+ nuclei within both the supraaoptic (SON) and paraventricular (PVN) nuclei. These findings raise new questions about AVPMNS's potential hormonal role for brain physiological adaptation across the lifespan, with possible involvement in coping with homeostatic adversities.

Effect of Short-Term Restraint Stress on the Hypothalamic Transcriptome Profiles of Rats with Inherited Stress-Induced Arterial Hypertension (ISIAH) and Normotensive Wistar Albino Glaxo (WAG) Rats.

Emotional stress is one of the health risk factors in the modern human lifestyle. Stress exposure can provoke the manifestation of various pathological conditions, one of which is a sharp increase in the blood pressure level. In the present study, we analyzed changes in the transcriptome profiles of the hypothalamus of hypertensive ISIAH and normotensive WAG rats exposed to a single short-term restraint stress (the rat was placed in a tight wire-mesh cage for 2 h). This type of stress can be considered emotional stress. The functional annotation of differentially expressed genes allowed us to identify the most significantly altered biological processes in the hypothalamus of hypertensive and normotensive rats. The study made it possible to identify a group of genes that describe a general response to stress, independent of the rat genotype, as well as a hypothalamic response to stress specific to each strain. The alternatively changing expression of the Npas4 (neuronal PAS domain protein 4) gene, which is downregulated in the hypothalamus of the control WAG rats and induced in the hypothalamus of hypertensive ISIAH rats, is suggested to be the key event for understanding inter-strain differences in the hypothalamic response to stress. The stress-dependent ISIAH strain-specific induction of Fos and Jun gene transcription may play a crucial role in neuronal activation in this rat strain. The data obtained can be potentially useful in the selection of molecular targets for the development of pharmacological approaches to the correction of stress-induced pathologies related to neuronal excitability, taking into account the hypertensive status of the patients.

Reinforcing systems of the brain and quantification of their work

BACKGROUND: The reinforcing systems of the brain are represented by the ventral forbrain dopaminergic bundle, which innervates the emotiogenic structures of the limbic system. Their study shows the reproduction of unconditioned (self-stimulation, self-administration) and conditioned reflex (preference for place, temperature, color) reactions. The quantitative assessment of the brain’s reinforcing systems remains unclear. For self-stimulation of brain structures, the change of the pedal presses in the Skinner chamber and some calculated coefficients are used, for example, the “mismatch coefficient”, which characterizes the temporal characteristics of the pedal pressings. AIM: To develop, test, and substantiate an additional objective quantitative method for assessing the reinforcing systems of the brain, called the “addiction coefficient”, based on an analysis of the effect of three psychoactive compounds (amphetamine, morphine and ethanol) in different doses on self-stimulation of the lateral hypothalamus in rats. MATERIALS AND METHODS: The main method for studying the reinforcing systems of the brain was the reaction of self-stimulation of the lateral hypothalamus in Wistar rats, which was modulated by the administration of psychoactive substances. The psychomotor stimulant amphetamine (phenamine) hydrochloride (0.5, 1, 2, and 4 mg/kg), narcotic analgesic morphine hydrochloride (1, 2, 4, and 8 mg/kg), and ethanol (0.5, 1, 2, and 4 g/kg) administered intraperitoneally were used as inductors of reinforcing. The control was the administration of of 0.9% NaCl solution (0.1, 0.2, 0.4, and 0.8 ml/rat). RESULTS: The use of different controls, characterized by an increase or decrease in the self-stimulation reaction in response to the introduction of 0.9% NaCl solution, showed that calculated coefficients, including the “mismatch coefficient”, can change in different directions and do not objectively reflect the reinforcing effects of pharmacological substances. The proposed “addiction coefficient”, which reflected the component of psychic dependence, changed unidirectionally toward an increase. The degree of this increase can be tens and hundreds of percent of the control and is significantly independent of the initial values of self-stimulation. As expected, the “addiction coefficient” increased most clearly after amphetamine administration and less significantly after morphine and ethanol injections. CONCLUSIONS: The “addiction coefficient” of a psychoactive substance, calculated as the ratio of the increase in pedal presses to the value of the “mismatch coefficient”, is a clear quantitative indicator when assessing the reinforcing properties of psychoactive substances in the self-stimulation reaction of the lateral hypothalamus. The “addiction coefficient” does not significantly depend on the initial level of self-stimulation and is recommended for a comparative assessment of the reinforcing properties of primarily related psychoactive compounds.

Quantitative Proteomics Provided Insights into the Protective Effects of Heat Acclimation on the Rat Hypothalamus after Exertional Heatstroke

Quantitative Proteomics Provided Insights into the Protective Effects of Heat Acclimation on the Rat Hypothalamus after Exertional Heatstroke

SARS-CoV-2 Spike Protein Enhances Carboxypeptidase Activity of Angiotensin-Converting Enzyme 2.

In this study, we investigated whether severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) spike protein may modify angiotensin-converting enzyme 2 (ACE2) activity in the plasma, heart, kidney, liver, lung, and six brain regions (amygdala, brain stem, cortex, hippocampus, hypothalamus, and striatum) of diabetic and hypertensive rats. We determine ACE2 activity in the plasma and lysates of heart, kidney, liver, lung, and six brain regions. MLN-4760 inhibits ACE2 activity in the plasma and all organs. On the other hand, soluble ACE2 (sACE2) activity increased in the plasma of diabetic rats, and there was no change in the plasma of hypertensive rats. ACE2 activity was augmented in the liver, brain stem, and striatum, while it decreased in the kidney, amygdala, cortex, and hippocampus of diabetic rats. ACE2 activity increased in the kidney, liver, and lung, while it decreased in the heart, amygdala, cortex, and hypothalamus of hypertensive rats. We measured the ACE2 content via enzyme-linked immunosorbent assay and found that ACE2 protein levels increased in the heart, while it decreased in the plasma, kidney, brain stem, cortex, hippocampus, hypothalamus, and striatum of diabetic rats. ACE2 protein levels decreased in the brain stem, cortex, hippocampus, and hypothalamus of hypertensive rats. Our data showed that the spike protein enhanced ACE2 activity in the liver and lungs of diabetic rats, as well as in the heart and three of the brain regions (cortex, hypothalamus, and striatum) of hypertensive rats.

Effects of preventative moxibustion at Shenque (CV8) and Guanyuan (CV4) on BDNF-TrkB pathway in the spinal dorsal horn and hypothalamus in rats with dysmenorrhea

Effects of preventative moxibustion at Shenque (CV8) and Guanyuan (CV4) on BDNF-TrkB pathway in the spinal dorsal horn and hypothalamus in rats with dysmenorrhea

Butyrate attenuates sympathetic activation in rats with chronic heart failure by inhibiting microglial inflammation in the paraventricular nucleus.

Sympathetic activation is a hallmark of heart failure and the underlying mechanism remains elusive. Butyrate is generated by gut microbiota and influences numerous physiological and pathological processes in the host. The present study aims to investigate whether the intestinal metabolite butyrate reduces sympathetic activation in rats with heart failure (HF) and the underlying mechanisms involved. Sprague-Dawley rats (220‒250 g) are anaesthetized with isoflurane, and the left anterior descending artery is ligated to model HF. Then, the rats are treated with or without butyrate sodium (NaB, a donor of butyrate, 10 g/L in water) for 8 weeks. Blood pressure and renal sympathetic nerve activity (RSNA) are recorded to assess sympathetic outflow. Cardiac function is improved (mean ejection fraction, 22.6%±4.8% vs 38.3%±5.3%; P<0.05), and sympathetic activation is decreased (RSNA, 36.3%±7.9% vs 23.9%±7.6%; P<0.05) in HF rats treated with NaB compared with untreated HF rats. The plasma and cerebrospinal fluid levels of norepinephrine are decreased in HF rats treated with NaB. The infusion of N-methyl-D-aspartic acid (NMDA) into the paraventricular nucleus (PVN) of the hypothalamus of HF model rats increases sympathetic nervous activity by upregulating the NMDA receptor. Microglia polarized to the M2 phenotype and inflammation are markedly attenuated in the PVN of HF model rats after NaB administration. In addition, HF model rats treated with NaB exhibit enhanced intestinal barrier function and increased levels of GPR109A, zona occludens-1 and occludin, but decreased levels of lipopolysaccharide-binding protein and zonulin. In conclusion, butyrate attenuates sympathetic activation and improves cardiac function in rats with HF. The improvements in intestinal barrier function, reductions in microglia-mediated inflammation and decreases in NMDA receptor 1 expression in the PVN are all due to the protective effects of NaB.

The evolutionary conserved miR-137/325 tandem mediates obesity-induced hypogonadism and metabolic comorbidities by repressing hypothalamic kisspeptin

BackgroundObesity-induced hypogonadism (OIH) is a prevalent, but often neglected condition in men, which aggravates the metabolic complications of overweight. While hypothalamic suppression of Kiss1-encoded kisspeptin has been suggested to contribute to OIH, the molecular mechanisms for such repression in obesity, and the therapeutic implications thereof, remain unknown. MethodsA combination of bioinformatic, expression and functional analyses was implemented, assessing the role of the evolutionary-conserved miRNAs, miR-137 and miR-325, in mediating obesity-induced suppression of hypothalamic kisspeptin, as putative mechanism of central hypogonadism and metabolic comorbidities. The implications of such miR-137/325-kisspeptin interplay for therapeutic intervention in obesity were also explored using preclinical OIH models. ResultsMiR-137/325 repressed human KISS1 3’-UTR in-vitro and inhibited hypothalamic kisspeptin content in male rats, while miR-137/325 expression was up-regulated, and Kiss1/kisspeptin decreased, in the medio-basal hypothalamus of obese rats. Selective over-expression of miR-137 in Kiss1 neurons reduced Kiss1/ kisspeptin and partially replicated reproductive and metabolic alterations of OIH in lean mice. Conversely, interference of the repressive actions of miR-137/325 selectively on Kiss1 3’-UTR in vivo, using target-site blockers (TSB), enhanced kisspeptin content and reversed central hypogonadism in obese rats, together with improvement of glucose intolerance, insulin resistance and cardiovascular and inflammatory markers, despite persistent exposure to obesogenic diet. Reversal of OIH by TSB miR-137/325 was more effective than chronic kisspeptin or testosterone treatments in obese rats. ConclusionsOur data disclose that the miR-137/325-Kisspeptin repressive interaction is a major player in the pathogenesis of obesity-induced hypogonadism and a putative druggable target for improved management of this condition and its metabolic comorbidities in men suffering obesity. Significance statementUp to half of the men suffering obesity display also central hypogonadism, an often neglected complication of overweight that can aggravate the clinical course of obesity and its complications. The mechanisms for such obesity-induced hypogonadism remain poorly defined. We show here that the evolutionary conserved miR137/miR325 tandem centrally mediates obesity-induced hypogonadism via repression of the reproductive-stimulatory signal, kisspeptin; this may represent an amenable druggable target for improved management of hypogonadism and other metabolic complications of obesity.

Exploring the metabolic implications of blue light exposure during daytime in rats

Excessive exposure to light is a global issue. Artificial light pollution has been shown to disrupt the body's natural circadian rhythm. To investigate the impacts of light on metabolism, we studied Sprague-Dawley rats chronically exposed to red or blue light during daytime or nighttime. Rats in the experimental group were exposed to extended light for 4 hours during daytime or nighttime to simulate the effects of excessive light usage. Strikingly, we found systemic metabolic alterations only induced by blue light during daytime. Furthermore, we conducted metabolomic analyses of the cerebrospinal fluid, serum, heart, liver, spleen, adrenal, cerebellum, pituitary, prostate, spermatophore, hypothalamus and kidney from rats in the control and blue light exposure during daytime. Significant changes in metabolites have been observed in cerebrospinal fluid, serum, hypothalamus and kidney of rats exposed to blue light during daytime. Metabolic alterations observed in rats encompassing pyruvate metabolism, glutathione metabolism homocysteine degradation, phosphatidylethanolamine biosynthesis, and phospholipid biosynthesis, exhibit analogous patterns to those inherent in specific physiological processes, notably neurodevelopment, cellular injury, oxidative stress, and autophagic pathways. Our study provides insights into tissue-specific metabolic changes in rats exposed to blue light during the daytime and may help explain potential mechanisms of photopathogenesis.

Aerobic training reduces microglial activation and TNFα release in the hypothalamus of hypertensive rats

Spontaneously hypertensive rats (SHR) exhibit autonomic imbalance and blood-brain barrier (BBB) dysfunction, a pathophysiological condition that can be corrected by exercise training. Although microglia cells are not part of the structure of the BBB itself, they modulate it function through the synthesis and release of cytokines. In hypertension condition, for instance, microglia are activated releasing pro-inflammatory cytokines, such as tumor necrosis factor-alpha (TNFα), which is able to break down different BBB constituents. Therefore, here we hypothesized that the benefit of exercise training to ameliorate BBB integrity of hypertensive rats could be due to the reduction in the release of TNFα by microglia at the hypothalamus level. Thus, our aim is to evaluate whether microglial activation, and consequent TNFα release, in the hypothalamus of SHR can be affected by aerobic exercise training. Wistar and SHR male rats (CEUA ICB/USP #3407080618) aged 90 days were divided into four groups: sedentary (W-S n=6) and trained Wistar (W-T n=6), sedentary (SHR-S n=6) and trained SHR (SHR-T n=6). Exercise trained animals were submitted to moderate intensity of aerobic training on a treadmill (50%-60% of maximum capacity, 1 h/day, 5 days/week, for 4 weeks). At the end of the 4th week of training, the hypothalamus, brainstem, and cortex of all groups were collected and processed for isolation of mononuclear cells, followed by flow cytometry analysis to evaluate the microglial labeling by CD45lowCD11b+, gated for TNFα+. All data as analyzed by two-way ANOVA followed by Bonferroni’s post-test. Trained animals showed performance gains compared to their control groups (Velocity: W-S: 1.07±0.23km/h vs. W-T: 1.7±0.36km/h, p&lt;0.0001 and SHR-S: 1.25±0.17km/h vs. SHT-T: 1.95±0.16km/h, p&lt;0.0001; Distance: W-S: 85±42m vs. W-T: 218±95m, p=0.0042 and SHR-S: 124±44m vs. SHT-T: 315±21m, p&lt;0.001). Hypertensive animals present an elevated neuroinflammation degree compared to normotensive subjects, by increased microglia activation (W-S: 0.65±0.39% vs. SHR-S: 7.0±1.4%, p&lt;0.0001) and TNFα release (W-S: 33.9±24.6% vs. SHR-S: 95.2±3.6%, p=0.0007). After training, hypertensive rats exhibited a reduction in hypothalamic microglial activation in a similar way to sedentary and trained Wistar (CD45lowCD11b+, SHR-T: 0.55±0.26%: vs. W-S:0.65±0.39% p=0.99; vs. W-T: 0.41±0.28% p=0.98; vs. SHR-S: 7.0±1.4% p&lt;0.0001). TNFα release also reduced in hypothalamus of SHR-T (CD11b+TNFα+, SHR-T: 59.4±17.0%: vs. W-S: 33.9±24.6% p=0.15; vs. W-T: 41.9±4.6% p=0.6; vs. SHR-S: 95.2±3.6% p&lt;0.01). SHR-S exhibit higher microglial activation and TNFα release in brainstem compared to W-S, but 4 weeks of training was not enough to reduce this condition. No difference was observed in cortex. In conclusion, exercise training of moderate intensity is effcient to reduce microglia activation and consequently TNFα release in the hypothalamus of hypertensive animals, which could be one of the causes affecting the integrity of the BBB in hypertensive animals. FAPESP #2018/14544-6; #2022/07465-8. This is the full abstract presented at the American Physiology Summit 2024 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.

A potential role for GPR107 in the development of Hypoglycemia Unawareness

Diabetes remains one of the largest health burdens in modern society and while an estimated 37.3 million people suffer from either type 1 or type 2 diabetes worldwide, a further 96 million people are currently estimated to be prediabetic. So far, most efforts for diabetic therapies have been focused on insulin-replacement therapies and secretagogues; however, a common risk associated with these therapies is hypoglycemia. Hypoglycemia is defined as low plasma glucose levels (&lt; 70 mg/dl) and can lead to loss of consciousness and death. Recently, it has been established that repeated exposure to bouts of hypoglycemia in diabetic individuals likely impairs sympathoadrenal counter regulation, and shifts the glycemic threshold for symptoms to lower plasma glucose concentrations, close to levels which result in cognitive failure. This pathology has been titled hypoglycemia unawareness (HU) and constitutes a major risk factor for developing severe hypoglycemia. The development of severe hypoglycemia in diabetics accounts for a six-fold increase in mortality, compared to diabetic individuals who do not experience severe hypoglycemia. Thus, it is important to understand how to prevent and reverse its development. We have developed a novel rat model of HU in which rats are exposed to insulin-induced recurrent hypoglycemia for five consecutive days. In this model of HU, we found that GPR107 expression is dramatically increased in the hypothalamus of rats exhibiting loss of hypoglycemia-induced food intake. Our group previously identified GPR107 as a receptor for neuronostatin (NST), a 13 amino acid peptide derived from the somatostatin prepro-hormone, which can exert anorexigenic actions in the hypothalamus. Using a GPR107-flox rat model we are currently evaluating the effect of hypothalamic knock-down of GPR107 on the development of HU and basal feeding behavior. We anticipate that if knock-down of GPR107 reverses HU, the development of NST antagonists could potentially be useful therapeutically for diabetic patients who experience recurrent hypoglycemia. Our GPR107-flox rat model will enable us to identify upstream and downstream interacting partners of GPR107/NST. This could reveal hypothalamic pathways involved in the development of HU that also could serve as potential therapeutic targets for this devastation consequence of iatrogenic hypoglycemia. Internal Funds St. Louis University. This is the full abstract presented at the American Physiology Summit 2024 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.

Acute hyperosmotic stimulus of the Organum Vasculosum of the Lamina Terminalis leads to increase in the blood pressure dependent on release of ATP into the Paraventricular Nucleus of the Hypothalamus of rats

The neural network involved in hyperosmolality-induced hypertension includes the activation of the organum vasculosum of the lamina terminalis (OVLT) with monosynaptic connections to the paraventricular nucleus of the hypothalamus (PVN) that expresses an abundance of purinergic receptors including P2 receptors. Previously, we have shown that hyperosmotic-induced sympathoexcitation at the level of the PVN depends on the P2 receptors. More recently, we have shown that hyperosmotic stimulus induces the release of ATP in the PVN, at least in part, from astrocytes. Here we hypothesize that the increase in the blood pressure (BP) induced by acute hyperosmotic stimulus in the OVLT involves the release of ATP within the PVN neurons, acting on P2 receptors. Thus, we sought to determine (1) whether P2X7-expressing OVLT and PVN neurons are activated by hyperosmotic challenge, and (2) what effects P2 receptor antagonism on changes in the BP mediated by an acute hyperosmotic stimulus in the OVLT. Adult Wistar rats (10-12 weeks old; CEUA ICB/USP: #4895140819 and #1521230221) were used in two protocols: 1) Double-labeling immunofluorescence of P2X7 (anti-P2X7 receptor, 1:500) expressing neurons with FOS (anti-cfos, 1:3000) animals received i.v administration of hypertonic (NaCl, 3M, n=4) or isotonic saline (NaCl, 0.154M, n=4), followed by evaluation of immunostaining to FOS and P2X7 receptors in the OVLT and PVN; 2) OVLT-PVN microinjections and BP monitoring: under anesthesia rats (n=5), received catheters in the femoral vein and artery for the administration of urethane (1g/kg) and BP recording signals, respectively. Microinjections of 3M NaCl (50nL) were administered into the OVLT, before and after bilateral antagonism of P2 purinergic receptors (PPADS, 10uM-100nL on each side) in the PVN with simultaneous BP recording. Data were expressed as the mean ± SD, and statistical significance level of p&lt;0.05 by Student’s T-test. Immunoreactive neurons for the P2X7 receptor subunit were observed in all compartments of the OVLT and PVN. Hyperosmotic stimulus significantly increased the number of Fos+ neurons in both the OVLT and PVN. While a small number of P2X7 receptors expressing OVLT and PVN neurons was observed in rats treated with isotonic saline, significantly more P2X7-expressing neurons expressed Fos+ in the OVLT with hypertonic saline. The stimulus of the OVLT with 3M NaCl elicited a significant increase in the BP. This effect was attenuated after bilateral injection of PPADS in the PVN (ΔMAP: NaCl OVLT:+10±1mmHg vs. NaCl OVLT post-PPADS PVN: +2.4±1.3mmHg, p&lt;0.0001). Collectively, our findings indicate that hyperosmotic stimulation activates a subset of P2X7 expressing OVLT and PVN neurons and that the elevation in the BP induced by hyperosmotic stimulus in the OVLT involves the endogenous release of ATP within the PVN, likely as part of the neural control of circulation during osmotic challenges. Financial Support: FAPESP #2019/19894-8, CAPES #88887.364432/2019-00. This is the full abstract presented at the American Physiology Summit 2024 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.

Cobalt protoporphyrin modulates antioxidant enzyme activity in the hypothalamus and motor cortex of female rats.

Cobalt protoporphyrin (CoPP) is a synthetic heme analog that has been observed to reduce food intake and promote sustained weight loss. While the precise mechanisms responsible for these effects remain elusive, earlier research has hinted at the potential involvement of nitric oxide synthase in the hypothalamus. This study aimed to delve into CoPP's impact on the activities of crucial antioxidant enzymes: superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione-S-transferase (GST) across seven distinct brain regions (hippocampus, hypothalamus, prefrontal cortex, motor cortex, striatum, midbrain, and cerebellum), as well as in the liver and kidneys. Female Wistar rats weighing 180 to 200 grams received a single subcutaneous dose of 25 μmol/kg CoPP. After six days, brain tissue was extracted to assess the activities of antioxidant enzymes and quantify malondialdehyde levels. Our findings confirm that CoPP administration triggers the characteristic effects of decreased food intake and reduced body weight. Moreover, it led to an increase in SOD activity in the hypothalamus, a pivotal brain region associated with food intake regulation. Notably, CoPP-treated rats exhibited elevated enzymatic activity of catalase, GR, and GST in the motor cortex without concurrent signs of heightened oxidative stress. These results underscore a strong connection between the antioxidant system and food intake regulation. They also emphasize the need for further investigation into the roles of antioxidant enzymes in modulating food intake and the ensuing weight loss, using CoPP as a valuable research tool.

Long-duration leptin transgene expression in dorsal vagal complex does not alter bone parameters in female Sprague Dawley rats

The hypothalamus and dorsal vagal complex (DVC) are both important for integration of signals that regulate energy balance. Increased leptin transgene expression in either the hypothalamus or DVC of female rats was shown to decrease white adipose tissue and circulating levels of leptin and adiponectin. However, in contrast to hypothalamus, leptin transgene expression in the DVC had no effect on food intake, circulating insulin, ghrelin and glucose, nor on thermogenic energy expenditure. These findings imply different roles for hypothalamus and DVC in leptin signaling. Leptin signaling is required for normal bone accrual and turnover. Leptin transgene expression in the hypothalamus normalized the skeletal phenotype of leptin-deficient ob/ob mice but had no long-duration (≥10 weeks) effects on the skeleton of leptin-replete rats. The goal of this investigation was to determine the long-duration effects of leptin transgene expression in the DVC on the skeleton of leptin-replete rats. To accomplish this goal, we analyzed bone from three-month-old female rats that were microinjected with recombinant adeno-associated virus encoding either rat leptin (rAAV-Leptin, n = 6) or green fluorescent protein (rAAV-GFP, control, n = 5) gene. Representative bones from the appendicular (femur) and axial (3rd lumbar vertebra) skeleton were evaluated following 10 weeks of treatment. Selectively increasing leptin transgene expression in the DVC had no effect on femur cortical or cancellous bone microarchitecture. Additionally, increasing leptin transgene expression had no effect on vertebral osteoblast-lined or osteoclast-lined bone perimeter or marrow adiposity. Taken together, the findings suggest that activation of leptin receptors in the DVC has minimal specific effects on the skeleton of leptin-replete female rats.

Calming effect of traditional Chinese medicine cinnabar based on neurotransmitter content and metal elements concentration in PCPA-induced insomnia rats

IntroductionCinnabar is a commonly utilized mineral traditional Chinese medicine (TCM) in China. It has the functions of clearing the heart and calming the mind. However, there remains a dearth of pharmacological experiments on the metal elements of cinnabar on animal models. Therefore, this study aims to investigate the effects of cinnabar on neurotransmitter levels in p-chlorophenylalanine (PCPA) induced insomnia rats and the content of metal elements in plasma, heart, liver, spleen, lung and kidney of rats. MethodsTissue hematoxylin and eosin (H&E) staining were used to assess the histological changes. The 5-HT and GABA in plasma and hypothalamus of rats were determined by enzyme-linked immunosorbent to evaluate the therapeutic effect. The content of metal elements in plasma, heart, liver, spleen, lung and kidney of rats was determined by ICP-MS. ResultsAfter cured with cinnabar and its processed for fourteen days, the level of 5-HT in the plasma and hypothalamus increased (P<0.05), and the level of GABA decreased (P<0.05). Furthermore, both low dose (1 mg/kg) and high dose (10 mg/kg) of cinnabar resulted in an increase in plasma 5-HT levels;, however, the high dose of cinnabar increased the level of 5-HT more than low dose of cinnabar. The content of metal elements in plasma, hemocyte and heart, liver, spleen, lung and kidney of rats ranged from less than 1 µg.g−1 to hundreds of µg.g−1, among which the highest content was Fe in rat's spleen of cinnabar sedative pill group, and the content is 919 µg.g−1. The content of most metal elements in plasma and hemocyte of rats was significantly lower than that in heart, liver, spleen, lung and kidney. There is no obvious difference of Hg in plasma and tissue of rats between the control group fed normally and rats fed with drugs. DiscussionCinnabar and its products can improve insomnia symptoms in rats by regulating the neurotransmitters 5-HT and GABA. Cinnabar is basically non-toxic in short-term (less than 14 days) use accordance with the dosage prescribed by pharmacopoeia.

Quercetin alleviates chronic unpredictable mild stress-induced depression-like behavior by inhibiting NMDAR1 with α2δ-1 in rats.

Depression is a serious mental disorder and the most prevalent cause of disability and suicide worldwide. Chronic unpredictable mild stress (CUMS) can lead to a significant acceleration of depression development. Quercetin (Que) is a flavonoid compound with a wide range of pharmacological effects. Recent studies have shown that quercetin can improve CUMS-induced depression-like behavior, but the mechanism of its improvement is still unclear. α2δ-1 is a regulatory subunit of voltage-gated calcium channel, which can interact with N-methyl-D-aspartate receptor (NMDAR) to form a complex. In this study, we found that Que could inhibit the increase of α2δ-1 and NMDAR expression in rat hypothalamus induced by CUMS. In pain, chronic hypertension and other studies have shown that α2δ-1 interacts with the NMDAR to form a complex, which subsequently affects the expression level of NMDAR. Consequently, the present study aimed to investigate the antidepressant effect of Que invivo and invitro and to explore its mechanism of action in terms of the interaction between α2δ-1 and NMDAR. Rats were randomly exposed to two stressors every day for 4 weeks to establish a CUMS rat model, then sucrose preference test (SPT), forced swimming test (FST), tail suspension test (TST), and open field test (OFT) were performed to detect the behavior of CUMS rats, so as to evaluate whether the CUMS rat model was successfully established and the improvement effect of Que on CUMS-induced depression-like behavior in rats. Experimental techniques such as serum enzyme-linked immunosorbent assay (ELISA), immunofluorescence, Western blot, and co-immunoprecipitation, as well as invitro experiments, were used to investigate the mechanisms by which Que exerts its antidepressant effects. Behavioral and ELISA test results showed that Que could produce a reduction in the excitability of the hypothalamic-pituitary-adrenal (HPA) axis in CUMS rats and lead to significant improvements in their depressive behavior. Western blot, immunofluorescence, and co-immunoprecipitation experiments showed that Que produced a decrease in NMDAR1 and α2δ-1 expression levels and interfered with α2δ-1 and NMDAR1 binding. In addition, the neural regulation mechanism of Que on antidepressant effect in PC12 cells knocked out α2δ-1 gene was further verified. Cellular experiments demonstrated that Que led to a reversal of up-regulation of NMDAR1 and α2δ-1 expression levels in corticosterone-injured PC12 cells, while Que had no effects on NMDAR1 expression in PC12 cells with the α2δ-1 gene knockout. Que has a good antidepressant effect and can significantly improve the depression-like behavior caused by CUMS. It exerts antidepressant effects by inhibiting the expression level of α2δ-1, interfering with the interaction between α2δ-1 and NMDAR, and then reducing the excitability of the HPA axis.

Hypothalamic Gene Expression in a Rat Model of Chronic Unpredictable Mild Stress Treated with Electroacupuncture.

Depression is characterized by the loss of pleasure and a depressed mood, and it is a common mental disorder in the twenty-first century. Multiple gene imbalances, which are considered pathological factors in depression, were detected in the brain. Electroacupuncture is an effective therapeutic approach for depression that has minimal side effects. As a crucial structure in the hypothalamus-pituitary-adrenal, the hypothalamus plays a key role in depression. Our study focused on the transcriptome level in the hypothalamus of depressive rats. After chronic unpredictable mild stress, the rats exhibited depressive-like behaviors, such as decreased sucrose consumption in the SPT, increased time in the central area of the OFT and increased immobility in the FST. Moreover, electroacupuncture alleviated depressive behaviors. Because of the importance of the hypothalamus in depression, we next detected gene expression in the hypothalamus. A total of 510 genes (125 upregulated genes and 385 downregulated genes) were detected in the hypothalamus of depressive rats. 15 of the 125 upregulated genes and 63 of the 385 downregulated genes could be altered by electroacupuncture, which suggests the antidepressant effect of electroacupuncture. Our study also provided the evidence that regulation of transcriptome in the hypothalamus might be a potential mechanism of electroacupuncture treatment.