Nucleus Tractus Solitarii Of Rats Research Articles

Effect of sex on chronic stress induced alterations in hindbrain catecholaminergic system and autonomic dysfunction resulting in gastrointestinal dysmotility

It has been reported that the clinical symptoms of functional dyspepsia (FD) exacerbate upon stress while the gender-related factors have been incompletely understood. This study aims to investigate the role of sex in chronic heterotypic stress (CHS)-induced autonomic and gastric motor dysfunction. For CHS, the rats were exposed to the combination of different stressors for 7 consecutive days. Subsequently, electrocardiography was recorded in anesthetized rats to evaluate heart rate variability (HRV) for the determination of autonomic outflow and sympathovagal balance. Solid gastric emptying (GE) was measured in control and CHS-loaded male and female rats. The immunoreactivities of catecholaminergic cell marker tyrosine hydroxylase (TH), choline acetyltransferase (ChAT), corticotropin releasing factor (CRF), and estrogen receptor (ER-α/β) were evaluated in medullary and pontine brainstem sections by immunohistochemistry. Compared with the controls, CHS significantly delayed GE in males but not in females. There was no significant sex-related difference in parasympathetic indicator HF under either control or CHS conditions. Sympathetic indicator LF was significantly higher in control females compared to the males. The higher sympathetic output in females was found to be attenuated upon CHS; in contrast, the elevated sympathetic output was detected in CHS-loaded males. No sex- or stress-related effect was observed on ChAT immunoreactivity in the dorsal motor nucleus of N.vagus (DMV). In males, greater number of TH-ir cells was observed in the caudal locus coeruleus (LC), while they were more densely detected in the rostral LC of females. Regardless of sex, CHS elevated immunoreactivity of TH throughout the LC. Under basal conditions, greater number of TH-ir cells was detected in the rostral ventrolateral medulla (RVLM) of females. In contrast, CHS remarkably increased the number of TH-ir cells in the RVLM of males which was found to be decreased in females. There was no sex-related alteration in TH immunoreactivity in the nucleus tractus solitarius (NTS) of control rats, while CHS affected both sexes in a similar manner. Compared with females, CRF immunoreactivity was prominently observed in control males, while both of which were stimulated by CHS. ER-α/β was found to be co-expressed with TH in the NTS and LC which exhibit no alteration related to either sex or stress status. These results indicate a sexual dimorphism in the catecholaminergic and the CRF system in brainstem which might be involved in the CHS-induced autonomic and visceral dysfunction occurred in males.

Sleep deprivation reduces the baroreflex sensitivity through elevated angiotensin (Ang) II subtype 1 receptor expression in the nucleus tractus solitarii.

Sleep insufficiency has been linked to an increased risk of high blood pressure and cardiovascular diseases. Emerging studies have demonstrated that impaired baroreflex sensitivity (BRS) is involved in the adverse cardiovascular effects caused by sleep deprivation, however, the underlying mechanisms remain unknown. Therefore, the present study aims to clarify the role of abnormal renin-angiotensin system in the nucleus tractus solitarii (NTS) in impaired BRS induced by sleep deprivation. Rats were randomly divided into two groups: normal sleep (Ctrl) and chronic sleep deprivation (CSD) group. Rats were sleep deprived by an automated sleep deprivation system. The blood pressure, heart rate, BRS, the number of c-Fos positive cells and the expression of angiotensin (Ang) II subtype 1 receptors (AT1R) in the NTS of rats were assessed. Compared to Ctrl group, CSD group exhibited a higher blood pressure, heart rate, and reduced BRS. Moreover, the number of c-Fos positive cells and local field potential in the NTS in CSD group were increased compared with the Ctrl group. It was shown that the expression of the AT1R and the content of Ang II and the ratio of Ang II to Ang-(1-7) were increased in the NTS of rats in CSD group compared to Ctrl group. In addition, microinjection of losartan into the NTS significantly improved the impaired BRS caused by sleep deprivation. In conclusion, these data suggest that the elevated AT1R expression in the NTS mediates the reduced BRS induced by chronic sleep deprivation.

Effect of high-frequency repetitive transcranial magnetic stimulation on swallowing function and pneumonia in poststroke dysphagia in rats

BackgroundPost-stroke dysphagia (PSD) is a common symptom of stroke. Clinical complications of PSD include malnutrition and pneumonia. Clinical studies have shown that high-frequency repetitive transcranial magnetic stimulation (HF-rTMS) can improve the swallowing function in stroke patients. However, few studies have elucidated the underlying molecular mechanisms. MethodsA PSD rat model was established using transient middle cerebral artery occlusion (tMCAO). Rats were randomly divided into sham-operated groups, PSD groups, PSD + sham-rTMS groups, PSD + 5 Hz-rTMS groups, PSD + 10 Hz-rTMS groups and PSD + 20 Hz-rTMS groups. Rats were weighed and videofluoroscopic swallowing studies were conducted. Pulmonary inflammation, levels of substance P (SP) and calcitonin gene-related peptide (CGRP) in the serum, lung, and nucleus tractus solitarius (NTS), brain-derived neurotrophic factor (BDNF) and 5-hydroxytryptamine (5HT) in NTS were evaluated. ResultsRats in the PSD group experienced weight loss, reduced bolus area and pharyngeal bolus speed, and increased pharyngeal transit time (PTT) and inter-swallow interval (ISI) on day 7 and day 14 after operation. Moreover, PSD rats showed pulmonary inflammation, reduced levels of SP in the lung and serum, increased levels of CGRP in the lung and NTS, reduced levels of BDNF and 5HT in the NTS. There was no significant difference between the PSD group and the PSD + sham-rTMS group in the results of weight and VFSS. Comparing with the PSD group, there significant increases in the bolus area, decreases in PTT of rats following 5 Hz rTMS intervention. HF-rTMS at 10 Hz significantly increased the weight, bolus area, pharyngeal bolus speed and decreased the PTT and ISI of rats. There were also significant increases in the bolus area (p < 0.01) and pharyngeal bolus speed, decreases in PTT and ISI of rats following 20 Hz rTMS intervention. Furthermore, compared with the PSD + 5 Hz-rTMS group, there were significant increases in the bolus area and pharyngeal bolus speed, decreases in ISI in the swallowing function of rats in the PSD + 10 Hz-rTMS group. Besides, compared with the PSD + 5 Hz-rTMS group, there were significant decreases in ISI in the swallowing function of rats in the PSD + 20 Hz-rTMS group. HF-rTMS at 10 Hz alleviated pulmonary inflammation, increased the levels of SP in the lung, serum, and NTS, CGRP in the serum and NTS, 5HT in the NTS of PSD rats. ConclusionCompared with 5 Hz and 20 Hz rTMS, 10 Hz rTMS more effectively improved the swallowing function of rats with PSD. HF-rTMS at 10 Hz improved the swallowing function and alleviated pneumonia in PSD rats. The mechanism may be related to increased levels of SP in the lung, serum and NTS, levels of CGRP in the serum and NTS, 5HT in the NTS after HF-rTMS treatment.

Glucagon-like peptide 1 receptor mediated modulation of excitatory synaptic transmission in the rat nucleus tractus solitarius

Glucagon-like peptide 1 receptor mediated modulation of excitatory synaptic transmission in the rat nucleus tractus solitarius

Distinct effects of orexin A on spontaneous and evoked synaptic currents in the rat nucleus tractus solitarius

Orexins are produced in hypothalamic areas and orexin-containing neurons are distributed in widespread areas of the central nervous system. Orexins regulate several physiological functions such as arousal, food intake and autonomic control. The presence of orexin-containing neuron terminals and orexin receptors has been confirmed in the nucleus tractus solitarius (NTS), which receives primary afferent fibers from peripheral organs including baroreceptors. However, the neuronal effects of orexin-1 receptor (OX1R) activation were not examined. Here, we aimed to determine the effects of OX1R activation on excitatory synaptic transmission. OX1R activation increased the frequency of spontaneous excitatory synaptic currents (sEPSCs). This effect was blocked by the prior application of L-NAME. In contrast, the amplitude of evoked excitatory postsynaptic currents (eEPSCs) was suppressed by OX1R activation, and this effect was prevented by a cannabinoid receptor 1 blocker, AM251, but not by the pretreatment with L-NAME. Altogether, these results suggest that OX1R activation increases sEPSCs frequency by stimulating NO production, whereas it inhibits eEPSCs by releasing endocannabinoids in the NTS. Thus, OX1R activation had distinct effects on spontaneous and evoked excitatory synaptic transmissions in the NTS.

Glut5 Knockdown in the Nucleus Tractus Solitarii Alleviates Fructose-Induced Hypertension in Rats

BackgroundSeveral studies have suggested mechanisms whereby excessive fructose intake increases blood pressure (BP). Glucose transporter 5 (GLUT5) is a fructose transporter expressed on enterocytes, and its involvement in the nucleus tractus solitarius (NTS)–modulated increase in BP following fructose intake remains unclear. ObjectivesHerein, we investigated whether NTS Glut5 knockdown (KD) can alleviate fructose-induced hypertension in rat models. MethodsMale Wistar-Kyoto rats (6–8 weeks old; average weight: 230 g) were randomly assigned into 4 groups [control (Con), fructose (Fru), fructose + scrambled (Fru + S), and Fru + KD]. The Con group rats had ad libitum access to regular water, and the other 3 groups were provided 10% fructose water ad libitum for 4 weeks (2 weeks before lentiviral transfection in the Fru + S and Fru + KD groups). Glut5 short hairpin RNA was delivered into the NTS of rats using a lentivirus system. Fructose-induced hypertension was assessed via the tail-cuff technique, a noninvasive blood pressure measurement approach. GLUT5-associated and other insulin signaling pathways in the NTS of rats were assessed using immunofluorescence and immunoblotting analyses. We evaluated between-group differences using the Mann-Whitney U test or Kruskal-Wallis 1-way ANOVA. ResultsCompared with the Fru + S group, the Fru + KD group had reduced sympathetic nerve hyperactivity (48.8 ± 3.2 bursts/min; P < 0.05), improved central insulin signaling, upregulated protein kinase B (AKT; 3.0-fold) and neuronal NO synthase (nNOS; 2.78-fold) expression, and lowered BP (17 ± 1 mmHg, P < 0.05). Moreover, Glut5 KD restored signaling dependent on adenosine 5′-monophosphate–activated protein kinase and reduced fructose-induced oxidative stress 2.0-fold, and thus decreased NAD(P)H oxidase in p67-phox 1.9-fold within the NTS. ConclusionsFructose-induced reactive oxygen species generates in the NTS of rats through GLUT5 and receptor for advanced glycation end products signaling, thus impairing the AKT–nNOS–NO signaling pathway and ultimately causing hypertension.

Effects of orexin A on excitatory synaptic transmission in the rat nucleus tractus solitarius

Effects of orexin A on excitatory synaptic transmission in the rat nucleus tractus solitarius

Single nuclei RNA sequencing of the rat AP and NTS following GDF15 treatment

ObjectiveGrowth differentiation factor 15 (GDF15) is known to play a role in feeding, nausea, and body weight, with action through the GFRAL-RET receptor complex in the area postrema (AP) and nucleus tractus solitarius (NTS). To further elucidate the underlying cell type-specific molecular mechanisms downstream of GDF15 signaling, we used a single nuclei RNA sequencing (snRNAseq) approach to profile AP and NTS cellular subtype-specific transcriptomes after systemic GDF15 treatment. MethodsAP and NTS micropunches were used for snRNAseq from Sprague Dawley rats 6 h following GDF15 or saline injection, and Seurat was used to identify cellular subtypes and cell type-specific alterations in gene expression that were due to the direct and secondary effects of systemic GDF15 treatment. ResultsUsing the transcriptome profile of ∼35,000 individual AP/NTS nuclei, we identified 19 transcriptomically distinct cellular subtypes, including a single population Gfral and Ret positive excitatory neurons, representing the primary site of action for GDF15. A total of ∼600 cell type-specific differential expression events were identified in neurons and glia, including the identification of transcriptome alterations specific to the direct effects of GDF15 in the Gfral-Ret positive excitatory neurons and shared transcriptome alterations across neuronal and glial cell types. Downstream analyses identified shared and cell type-specific alterations in signaling pathways and upstream regulatory mechanisms of the observed transcriptome alterations. ConclusionsThese data provide a considerable advance in our understanding of AP and NTS cell type-specific molecular mechanisms associated with GDF15 signaling. The identified cellular subtype-specific regulatory mechanism and signaling pathways likely represent important targets for future pharmacotherapies.

Single Cell Scale Neuronal and Glial Gene Expression and Putative Cell Phenotypes and Networks in the Nucleus Tractus Solitarius in an Alcohol Withdrawal Time Series.

Alcohol withdrawal syndrome (AWS) is characterized by neuronal hyperexcitability, autonomic dysregulation, and severe negative emotion. The nucleus tractus solitarius (NTS) likely plays a prominent role in the neurological processes underlying these symptoms as it is the main viscerosensory nucleus in the brain. The NTS receives visceral interoceptive inputs, influences autonomic outputs, and has strong connections to the limbic system and hypothalamic-pituitary-adrenal axis to maintain homeostasis. Our prior analysis of single neuronal gene expression data from the NTS shows that neurons exist in heterogeneous transcriptional states that form distinct functional subphenotypes. Our working model conjectures that the allostasis secondary to alcohol dependence causes peripheral and central biological network decompensation in acute abstinence resulting in neurovisceral feedback to the NTS that substantially contributes to the observed AWS. We collected single noradrenergic and glucagon-like peptide-1 (GLP-1) neurons and microglia from rat NTS and measured a subset of their transcriptome as pooled samples in an alcohol withdrawal time series. Inflammatory subphenotypes predominate at certain time points, and GLP-1 subphenotypes demonstrated hyperexcitability post-withdrawal. We hypothesize such inflammatory and anxiogenic signaling contributes to alcohol dependence via negative reinforcement. Targets to mitigate such dysregulation and treat dependence can be identified from this dataset.

Investigation of vascular endothelial growth factor receptor-dependent neuroplasticity on rat nucleus tractus solitarius and phrenic nerve after chronic sustained hypoxia.

The neuronal system that controls respiration creates plasticity in response to physiological changes. Chronic sustained hypoxia causes neuroplasticity that contributes to ventilatory acclimatization to hypoxia (VAH). The purpose of this study is to explain the potential roles of the VAH mechanism developing because of chronic sustained hypoxia on respiratory neuroplasticity of vascular endothelial growth factor (VEGF) receptor activationon the nucleus tractus solitarius (NTS) and phrenic nerve. In this study 24 adult male Sprague-Dawley rats were used. Subjects were separated into four groups, amoderate-sham (mSHAM), severed-sham (sSHAM), moderate chronic sustained hypoxia (mCSH), and severed chronic sustained hypoxia (sCSH). Normoxic group (mSHAM and sSHAM) rats were exposed to 21% O₂ level (7 days) in the normobaric room while hypoxia group (mCSH and sCSH) rats were exposed to 13% and 10% O₂ level (7 days). Different protocols were applied for normoxic and hypoxia groups and ventilation, respiratory frequency, and tidal volume measurements were made with whole-body plethysmography. After the test HIF-1α, erythropoietin (EPO), and VEGFR-2 expressions on the NTS region in the medulla oblongata and phrenic nerve motor neurons in spinal cord tissue were analyzed using the immunohistochemical stain method. Examinations on the medulla oblongata and spinal cord tissues revealed that HIF-1α, EPO, and VEGFR-2 expressions increased inhypoxia groups compared to normoxic groups while a similar increase was also seen when respiratory parameters were assessed. Consequently, learning about VAH-related neuroplasticity mechanisms developed as a result of chronic continuous hypoxia will contribute to developing new therapeutical approaches to various diseases causing respiratory failure using brain plasticity without recourse to medicines.

Increased neuronal activation in sympathoregulatory regions of the brain and spinal cord in type 2 diabetic rats.

Increased cardiac sympathetic nerve activity in type 2 diabetes mellitus (DM) suggests impaired autonomic control of the heart. However, the central regions that contribute to the autonomic cardiac pathologies in type 2 DM are unknown. Therefore, we tested the hypothesis that neuronal activation would be increased in central sympathoregulatory areas in a pre-clinical type 2 DM animal model. Immunohistochemistry in 20-week-old male Zucker diabetic fatty (ZDF) rats revealed an increased number of neurones expressing ΔFosB (a marker of chronic neuronal activation) in the intermediolateral column (IML) of the spinal cord in DM compared to non-diabetic (non-DM) rats (P<0.05). Rostral ventrolateral medulla (RVLM) neurones activate IML neurones and receive inputs from the hypothalamic paraventricular nucleus (PVN), as well as the nucleus tractus solitarius (NTS) and area postrema (AP), in the brainstem. We observed more ΔFosB-positive noradrenergic RVLM neurones (P<0.001) and corticotrophin-releasing hormone PVN neurones (P<0.05) in DM compared to non-DM rats. More ΔFosB-positive neurones were also observed in the NTS (P<0.05) and AP (P<0.01) of DM rats compared to non-DM rats. Finally, because DM ZDF rats are obese, we also expected increased activation of pro-opiomelanocortin (POMC) arcuate nucleus (ARC) neurones in DM rats; however, fewer ΔFosB-positive POMC ARC neurones were observed in DM compared to non-DM rats (P<0.01). In conclusion, increased neuronal activation in the IML of type 2 DM ZDF rats might be driven by RVLM neurones that are possibly activated by PVN, NTS and AP inputs. Elucidating the contribution of central sympathoexcitatory drive in type 2 DM might improve the effectiveness of pharmacotherapies for diabetic heart disease.

Electroacupuncture at "Taichong"(LR3) improves baroreflex sensitivity and α2-adrenergic receptor activity in nucleus tractus solitarii of hypertensive rats

To investigate the effect of electroacupuncture (EA) at "Taichong"(LR3) on blood pressure, sympathetic nerve activity, baroreflex sensitivity, and α 2-adrenergic receptor (α2AR) expression in nucleus tractus solitarii (NTS) in hypertensive rats, so as to reveal its mechanisms underlying improvement of hypertension. Male Sprague Dawley rats were randomly divided into 4 groups: sham-operation, model, EA, and sham-EA (non-acupoint) groups, with 12 rats in each group. The hypertension model was established by occlusion of the right renal aorta (two-kidney-one clip method). Rats of the sham-operation group received the same surgery but without occlusion of the renal artery. EA (2 Hz/15 Hz, 2 mA) was applied to bilateral LR3 for 30 min, once a day for 28 days, and sham EA was applied to the skin of the rat tail near the buttock on both sides. The mean arterial pressure (MAP) of the abdominal aorta and heart rate (HR) were recorded. The autonomic nerve function was assessed by using frequency domain analysis of heart rate variability (HRV), and the baroreflex sensitivity detected by sequential method. Plasma norepinephrine (NE) level was measured by ELISA, and the α2AR positive neurons and α2AR protein expression in NTS were detected by using fluorescence immunohistochemistry and Western blot, respectively. The functions of α2AR within the NTS in modulating MBP and HR were verified by microinjection of its agonist (clonidine) and antagonist (yohimbine) separately. Compared to the sham operation rats, the hypertension rats displayed significant increases in the MAP (P<0.01), plasma norepinephrine content (P<0.01), ratios of low frequency/total power (LF/TP) and low frequency/high frequency (LF/HF) (P<0.01), and significant reduction in the overall gain, uplink sequence gain and downlink sequence gain of baroreflex (P<0.01), the number α2AR positive neurons and α2AR protein expression level in NTS (P<0.01). The rats in the EA group (rather than in the sham-EA group) showed significant reduction in MAP at the 3rd and 4th week, plasma NE content, LF/TP and LF/HF (P<0.01), and obvious increase in the overall gain, uplink sequence gain and downlink sequence gain of baroreflex (P<0.01), and the number of α2AR positive neurons and α2AR protein expression in comparison (P<0.05) with those of the model group. Microinjection of clonidine into NTS induced an evident decrease in both MAP and HR in the model group relevant to the sham operation group (P<0.01), while the MAP and HR changes of the EA (not sham EA) group were considerably bigger than those of the model group (P<0.05), being similar to those of the sham-operation group (P>0.05), which suggested an elimination of the BP-lowering effect of clonidine after EA. EA at LR3 can reduce MBP, sympathetic activities, improve baroreflex sensitivity in renovascular hypertensive rats, which may be associated with its effects in up-regulating the decreased NTS α2AR expression and functional activities.

The baroreflex afferent pathway plays a critical role in H2S-mediated autonomic control of blood pressure regulation under physiological and hypertensive conditions.

Hydrogen sulfide (H2S), which is closely related to various cardiovascular disorders, lowers blood pressure (BP), but whether this action is mediated via the modification of baroreflex afferent function has not been elucidated. Therefore, the current study aimed to investigate the role of the baroreflex afferent pathway in H2S-mediated autonomic control of BP regulation. The results showed that baroreflex sensitivity (BRS) was increased by acute intravenous NaHS (a H2S donor) administration to renovascular hypertensive (RVH) and control rats. Molecular expression data also showed that the expression levels of critical enzymes related to H2S were aberrantly downregulated in the nodose ganglion (NG) and nucleus tractus solitarius (NTS) in RVH rats. A clear reduction in BP by the microinjection of NaHS or L-cysteine into the NG was confirmed in both RVH and control rats, and a less dramatic effect was observed in model rats. Furthermore, the beneficial effects of NaHS administered by chronic intraperitoneal infusion on dysregulated systolic blood pressure (SBP), cardiac parameters, and BRS were verified in RVH rats. Moreover, the increase in BRS was attributed to activation and upregulation of the ATP-sensitive potassium (KATP) channels Kir6.2 and SUR1, which are functionally expressed in the NG and NTS. In summary, H2S plays a crucial role in the autonomic control of BP regulation by improving baroreflex afferent function due at least in part to increased KATP channel expression in the baroreflex afferent pathway under physiological and hypertensive conditions.

CX3CR1-microglia mediates neuroinflammation and blood pressure regulation in the nucleus tractus solitarii of fructose-induced hypertensive rats

BackgroundInflammation is a common pathophysiological trait found in both hypertension and cardiac vascular disease. Recent evidence indicates that fractalkine (FKN) and its receptor CX3CR1 have been linked to inflammatory response in the brain of hypertensive animal models. Here, we investigated the role of CX3CR1-microglia in nitric oxide (NO) generation during chronic inflammation and systemic blood pressure recovery in the nucleus tractus solitarii (NTS).MethodsThe hypertensive rat model was used to study the role of CX3CR1-microglia in NTS inflammation following hypertension induction by oral administration of 10% fructose water. The systolic blood pressure was measured by tail-cuff method of non-invasive blood pressure. The CX3CR1 inhibitor AZD8797 was administered intracerebroventricularly (ICV) in the fructose-induced hypertensive rat. Using immunoblotting, we studied the nitric oxide synthase signaling pathway, NO concentration, and the levels of FKN and CX3CR1, and pro-inflammatory cytokines were analyzed by immunohistochemistry staining.ResultsThe level of pro-inflammatory cytokines IL-1β, IL-6, TNF-α, FKN, and CX3CR1 were elevated two weeks after fructose feeding. AZD8797 inhibited CX3CR1-microglia, which improved the regulation of systemic blood pressure and NO generation in the NTS. We also found that IL-1β, IL-6, and TNF-α levels were recovered by AZD8797 addition.ConclusionWe conclude that CX3CR1-microglia represses the nNOS signaling pathway and promotes chronic inflammation in fructose-induced hypertension. Collectively, our results reveal the role of chemokines such as IL-1β, IL-6, and TNF-α in NTS neuroinflammation with the involvement of FKN and CX3CR1.

Chronic kidney disease‐induced insulin resistance is mediated by increased phosphorylation of SGK1 and enhanced SGLT2 expression in NTS

BackgroundThe serum and glucocorticoid inducible‐kinase‐1 (SGK1) expression is regulated during discrete developmental stages and pathological conditions, such as hypertension and diabetic neuropathy. In the present study, chronic kidney disease (CKD) in rats enhanced SGK1 function and regulation within the nucleus tractus solitarii (NTS). This study aimed to assess the regulatory mechanism of SGK1 in the NTS in rats presenting CKD‐induced insulin resistance.MethodsEight‐week‐ and nine‐week‐old male Wistar–Kyoto (WKY) rats underwent nephrectomy of the left and right kidneys, respectively. After subjecting the rats to CKD‐induced hypertension for 2 weeks, they were treated with SGK1 inhibitor (GSK650394, 100 mM) for another 2 weeks. Furthermore, the rats underwent 5/6 nephrectomy for 4 weeks; thereafter, they were tested for blood pressure, blood and urine chemistry, renal function, and protein expression. Mann–Whitney U test was used to compare the mean rank differences in scales of the general parameters between the CKD (sham, CKD, and CKD + GSK650394) groups, and the values were presented as mean ± SEM.ResultsRats with CKD revealed proteinuria, lower urine sodium and chloride levels and hyperglycemia, while treatment with SGK1 inhibitor significantly decreased these effects. Immunoblotting analysis further revealed that phospho‐SGK1 (Ser78), phosphoinositide‐dependent kinase‐1 (PDK1), and sodium‐glucose co‐transporter 2 (SGLT2) expressions increased in the NTS of CKD rats. Furthermore, inhibition of phospho‐SGK1 (Ser78) by GSK650394 significantly decreased the expression of PDK1 and SGLT2 in the NTS of CKD rats.ConclusionOur results suggest that inhibition of SGK1 expression in the NTS by intracerebroventricularly injected SGK1 inhibitor decreases the fasting serum glucose by negatively regulating CKD‐induced SGK1 signaling, reducing SGK1 (Ser78) phosphorylation, and by inhibiting the PDK1 and SGLT2 expressions in the NTS. These results indicate that SGK1 regulates the cardiometabolic syndrome in the NTS of CKD rats.Support or Funding Information107‐2320‐B‐075B‐002‐MY2、107‐2320‐B‐075B‐001

Time Course of Astrocyte Activation in Mouse Nucleus Tractus Solitarius During Exposure to Hypoxia

Chronic sustained hypoxia (CH) occurs in populations living at high altitude and in patients with chronic pulmonary disease. Exposure to CH produces ventilatory acclimatization to hypoxia (VAH) and increases the hypoxic ventilatory response (HVR) by mechanisms that involve areas of the brainstem that control breathing. The nucleus tractus solitarius (NTS) is a sensory integrative center in the medulla receiving carotid body afferents and known to be crucial for VAH. Our previous results demonstrated that glia cells in the rat NTS contribute to VAH but these mechanisms have not been studied in mice exposed to CH beyond 24 hours. We hypothesized that CH produces an early transient activation of astrocytes and microglia in the mouse brainstem as observed in rats. To study the activation of glial cells with CH, we exposed mice to normobaric hypoxia (10% FiO2) for 0.5, 1 and 4 hours and 1 and 7 days. Mice were perfused with 4% paraformaldehyde and brainstem sections were obtained. We quantified astrocyte activation by measuring glial fibrillary acidic protein (GFAP) intensity with immunofluorescence, and microglial activation by measuring branch morphology using Iba‐1 marker. Exposure to CH significantly increased GFAP intensity after 30 minutes and 1day while microglia morphology did not change significantly. The results differ from those in rats (J. Neurophysiol.117: 1625–1635, 2017) and suggest that time‐dependent astrocyte activation in the NTS may contribute to VAH.Support or Funding InformationSupported by NIH RO1 HL‐081823

A preliminary study of the effect of a high-salt diet on transcriptome dynamics in rat hypothalamic forebrain and brainstem cardiovascular control centers.

BackgroundHigh dietary salt intake is strongly correlated with cardiovascular (CV) diseases and it is regarded as a major risk factor associated with the pathogenesis of hypertension. The CV control centres in the brainstem (the nucleus tractus solitarii (NTS) and the rostral ventrolateral medulla (RVLM)) and hypothalamic forebrain (the subfornical organ, SFO; the supraoptic nucleus, SON and the paraventricular nucleus, PVN) have critical roles in regulating CV autonomic motor outflows, and thus maintaining blood pressure (BP). Growing evidence has implicated autonomic regulatory networks in salt-sensitive HPN (SSH), but the genetic basis remains to be delineated. We hypothesized that the development and/ or maintenance of SSH is reliant on the change in the expression of genes in brain regions controlling the CV system.MethodologyWe used RNA-Sequencing (RNA-Seq) to describe the differential expression of genes in SFO, SON, PVN, NTS and RVLM of rats being chronically fed with high-salt (HS) diet. Subsequently, a selection of putatively regulated genes was validated with quantitative reverse transcription polymerase chain reaction (qRT-PCR) in both Spontaneously Hypertensive rats (SHRs) and Wistar Kyoto (WKY) rats.ResultsThe findings enabled us to identify number of differentially expressed genes in SFO, SON, PVN, NTS and RVLM; that are either up-regulated in both strains of rats (SON- Caprin2, Sctr), down-regulated in both strains of rats (PVN- Orc, Gkap1), up-regulated only in SHRs (SFO- Apopt1, Lin52, AVP, OXT; SON- AVP, OXT; PVN- Caprin2, Sclt; RVLM- A4galt, Slc29a4, Cmc1) or down-regulated only in SHRs (SON- Ndufaf2, Kcnv1; PVN- Pi4k2a; NTS- Snrpd2l, Ankrd29, St6galnac6, Rnf157, Iglon5, Csrnp3, Rprd1a; RVLM- Ttr, Faim).ConclusionsThese findings demonstrated the adverse effects of HS diet on BP, which may be mediated via modulating the signaling systems in CV centers in the hypothalamic forebrain and brainstem.

Binge-like palatable food intake in rats reduces preproglucagon in the nucleus tractus solitarius

Binge eating involves eating larger than normal quantities of food within a discrete period of time. The neurohormonal controls governing binge-like palatable food intake are not well understood. Glucagon-like peptide-1 (GLP-1), a hormone produced peripherally in the intestine and centrally in the nucleus tractus solitarius (NTS), reduces food intake. Given that the NTS plays a critical role in integrating peripheral and central signals relevant for food intake, as well as the role of GLP-1 in motivated feeding, we tested the hypothesis that expression of the GLP-1 precursor preproglucagon (PPG) would be reduced in the NTS of rats with a history of binge-like palatable food intake. Adult male rats received access to fat for 1 h shortly before lights off, either every day (Daily, D) or only 3d/week (Intermittent, INT). INT rats ate significantly more fat than did D rats in sessions where all rats had fat access. After ~8.5 weeks of diet maintenance, we measured plasma GLP-1 as well as NTS PPG and GLP-1 receptor expression. INT rats had significantly lower NTS PPG mRNA expression compared to D rats. However, plasma GLP-1 was significantly increased in the INT group versus D rats. No significant differences were observed in NTS GLP-1 receptor expression. We also measured plasma insulin levels, fasted blood glucose, and plasma corticosterone but no differences were detected between groups. These results support the hypothesis that binge-like eating reduces NTS GLP-1 expression, and furthermore, demonstrate divergent impacts of binge-like eating on peripheral (plasma) versus central GLP-1.

Roux-en-Y gastric bypass surgery triggers rapid DNA fragmentation in vagal afferent neurons in rats

Previous studies have shown that Roux‑en‑Y gastric bypass (RYGB), one of the most effective weight loss treatments for obesity, results in neurodegenerative responses in vagal afferent gut‑brain connection reflected by microglia activation and reduced sensory input to the nucleus tractus solitarius (NTS). However, it is not known whether RYGB‑induced microglia activation is the cause or an effect of the reported neuronal damage. Therefore, the aim of this study was to establish the order of neurodegenerative responses in vagal afferents after RYGB in the nodose ganglia (NG) and NTS in male and female rats. Sprague‑Dawley rats were fed regular chow or an energy‑dense diet for two weeks followed by RYGB or sham surgery. Twenty‑four hours later, animals were sacrificed and NG and NTS were collected. Neuronal cell damage was determined by TUNEL assay. Microglia activation was determined by quantifying the fluorescent staining against the ionizing calcium adapter‑binding molecule 1. Reorganization of vagal afferents was evaluated by fluorescent staining against isolectin 4. Results of the study revealed significantly increased DNA fragmentation in vagal neurons in the NG when observed at 24 h after RYGB. The surgery did not produce rapid changes in the density of vagal afferents and microglia activation in the NTS. These data indicate that decreased density of vagal afferents and increased microglia activation in the NTS likely ensue as a res ult of RYGB‑induced neuronal damage.

Acute myocardial infarction activates magnocellular vasopressin and oxytocin neurones.

Myocardial infarction (MI) is a leading cause of death worldwide. For those who survive the acute insult, the progressive dilation of the ventricle associated with chronic heart failure is driven by an adverse increase in circulating levels of the antidiuretic hormone, vasopressin, which is secreted from hypothalamic supraoptic (SON) and paraventricular nuclei (PVN) nerve terminals. Although increased vasopressin neuronal activity has been demonstrated in the latter stages of chronic heart failure, we hypothesised that vasopressin neurones become activated immediately following an acute MI. Male Sprague-Dawley rats were anaesthetised and an acute MI was induced by ligation of the left anterior descending coronary artery. After 90minutes of myocardial ischaemia, brains were collected. Dual-label immunohistochemistry was used to quantify the expression of Fos protein, a marker of neuronal activation, within vasopressin- or oxytocin-labelled neurones of the hypothalamic PVN and SON. Fos protein and tyrosine hydroxylase within the brainstem were also quantified. The results obtained show that the expression of Fos in both vasopressin and oxytocin neurones of the PVN and SON was significantly elevated as soon as 90minutes post-MI compared to sham rats. Moreover, Fos protein was also elevated in tyrosine hydroxylase neurones in the nucleus tractus solitarius and rostral ventrolateral medulla of MI rats than sham rats. We conclude that magnocellular vasopressin and oxytocin neuronal activation occurs immediately following acute MI, rather than in the later stages of chronic heart failure. Therefore, prompt vasopressin antagonist therapy as an adjunct treatment for acute MI may impede the progression of ventricular dilatation, which remains a key adverse hallmark of chronic heart failure.