Ventrolateral Medulla Research Articles

Fos expression in A1/C1 neurons of rats exposed to hypoxia, hypercapnia, or hypercapnic hypoxia

The distribution of Fos expression in catecholaminergic neurons with immunoreactivity for dopamine β-hydroxylase (DBH) of the ventrolateral medulla was compared between rats exposed to hypoxia (10 % O2), hypercapnia (8 % CO2), and hypercapnic hypoxia (8 % CO2 and 10 % O2) for 2 h. Among the experimental groups, hypoxia-exposed rats had more Fos/DBH double-immunoreactive neurons than the control group (20 % O2) in the rostral area of the ventrolateral medulla, specifically in the range of + 150 μm to + 2,400 μm from the caudal end of the facial nerve nucleus. On the other hand, Fos/DBH double-immunoreactive neurons were scarcely observed in the ventrolateral medullary region of hypercapnia-exposed rats. The number of double-immunoreactive neurons in hypercapnic hypoxia-exposed rats was comparable to that in the control group. The present results suggest that adrenergic C1 neurons are specifically activated by hypoxia and are involved in the regulation of respiratory and circulatory functions.

Tyrosine hydroxylase-positive neurons in the rostral ventrolateral medulla mediate sympathetic activation in sepsis

AimSepsis results in high mortality and is associated with organ dysfunction caused by infection. The present study aimed to elucidate whether early-stage sympathetic activation is associated with the prognosis of sepsis and its possible mechanisms. MethodsPatients with sepsis and healthy controls were included. Sepsis in rats was induced by lipopolysaccharide. Dexmedetomidine, a α2-adrenergic receptor agonist, was used in patients and rats with sepsis to evaluate the role of the sympathetic nervous system in sepsis. Holter monitoring was used to detect heart rate variability, while plasma samples were obtained to measure levels of norepinephrine and inflammatory markers. Mean arterial pressure, heart rate, and renal sympathetic nerve activity were recorded. Immunofluorescence was used to detect the activation of neurons in the rostral ventrolateral medulla (RVLM). ResultsIn patients with sepsis, plasma levels of norepinephrine and interleukin-1β were higher compared with those in controls and positively correlated with acute physiology and chronic health evaluation (APACHEII). SDNN and SDANN were significantly reduced as well as negatively correlated with APACHEII. Meanwhile, rats with sepsis showed increased of sympathetic outflow and plasma levels of norepinephrine, with increased c-fos levels in the RVLM. Treatment with dexmedetomidine could improve prognosis. Lesion of tyrosine hydroxylase-positive neurons in the RVLM attenuated sympathetic activation and target organs damage in septic rats as well as improved survival. ConclusionThe results suggest that tyrosine hydroxylase-positive neurons in the RVLM might contribute to the prognosis of sepsis via activation of the sympathetic nervous system, while dexmedetomidine could ameliorate sepsis via inhibiting sympathetic activation.

Methamphetamine-Induced Blood Pressure Sensitization Correlates with Morphological Alterations within A1/C1 Catecholamine Neurons

Methamphetamine (METH) is a drug of abuse, which induces behavioral sensitization following repeated doses. Since METH alters blood pressure, in the present study we assessed whether systolic and diastolic blood pressure (SBP and DBP, respectively) are sensitized as well. In this context, we investigated whether alterations develop within A1/C1 neurons in the vasomotor center. C57Bl/6J male mice were administered METH (5 mg/kg, daily for 5 consecutive days). Blood pressure was measured by tail-cuff plethysmography. We found a sensitized response both to SBP and DBP, along with a significant decrease of catecholamine neurons within A1/C1 (both in the rostral and caudal ventrolateral medulla), while no changes were detected in glutamic acid decarboxylase. The decrease of catecholamine neurons was neither associated with the appearance of degeneration-related marker Fluoro-Jade B nor with altered expression of α-synuclein. Rather, it was associated with reduced free radicals and phospho-cJun and increased heat shock protein-70 and p62/sequestosome within A1/C1 cells. Blood pressure sensitization was not associated with altered arterial reactivity. These data indicate that reiterated METH administration may increase blood pressure persistently and may predispose to an increased cardiovascular response to METH. These data may be relevant to explain cardiovascular events following METH administration and stressful conditions.

Anatomic and functional evidence for renal autonomic innervation in normotensive and hypertensive rats.

Renal denervation (RDN) has been used for treating resistant hypertension. A few recent studies have shown vagal innervation of kidneys causing confusion. This study aimed to provide anatomical and functional evidence for renal autonomic innervation. Experiments were performed in male Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Pseudorabies virus (PRV) in the paraventricular nucleus and rostral ventrolateral medulla was prevented by bilateral RDN, but not subdiaphragmatic vagotomy. PRV did not appear in the dorsal motor nucleus of the vagus and nucleus tractus solitarii 72 h after renal injection of PRV. Adrenergic fibers were approximately seven times more than cholinergic fibers in the main renal artery (MRA) and its first (1RA) and second grade (2RA) branches. Adrenergic fibers in 1RA were more than those in MRA and 2RA. Tyrosine hydroxylase immunoreactivity in these arteries was higher in SHR than in WKY. Norepinephrine (NE) increased and α-receptor antagonist reduced vascular ring tension of renal arteries. The effect of NE was greater in 1RA and 2RA than in MRA, which was prevented by α-receptor antagonist. Acetylcholine (ACh) or blockage of β-receptors, M receptors, or N receptors had no significant effects on vascular ring tension and the effect of NE. Renal blood flow was reduced by electrical stimulation of renal nerves but not affected by stimulation of the subdiaphragmatic vagus. These results provide anatomical and functional evidence that kidneys are innervated and renal blood flow is regulated by renal sympathetic nerves rather than the vagus. Renal vasoconstriction is regulated by NE and adrenergic fibers rather than ACh or cholinergic fibers in WKY and SHR.NEW & NOTEWORTHY Kidneys are innervated by renal nerves rather than the vagus. Adrenergic fibers in renal arteries are about seven times more than cholinergic fibers. Renal vasoconstriction is regulated by norepinephrine and adrenergic fibers rather than acetylcholine or cholinergic fibers. Renal blood flow is regulated by renal sympathetic nerves and is not affected by the vagus. These findings provide anatomical and functional evidence for renal autonomic innervation in normotensive and hypertensive rats.

Effects of acupuncture on endothelial active factors and related autonomic neurotransmitters in spontaneous hypertension rats

To observe the effects of acupuncture at "antihypertensive acupoint prescription" on endothelial active factors and related autonomic neurotransmitters in spontaneous hypertension rats, and explore the vascular regulation and central regulation mechanisms of acupuncture for anti-hypertension. Thirty SPF grade male spontaneous hypertension rats were randomly divided into a model group (15 rats) and an acupuncture group (15 rats). Besides, 15 Wistar Kyoto rats were collected as a blank control group (normal group). In the acupuncture group, acupuncture was delivered at the "antihypertensive acupoint prescription" (bilateral "Renying" [ST 9], "Quchi" [LI 11], "Zusanli" [ST 36], "Taichong" [LR 3] and "Neiguan" [PC 6]), with needles retained for 30 min, once daily. The duration of intervention was 28 days. Every week, using the the irritation scale, the sign of sympathetic irritation was evaluated dynamically. The arterial blood pressure of the rats tail was determined, using non-invasive blood pressure measurement system. ELISA was adopted to detect the levels of calcitonin gene-related peptide (CGRP), nitric oxide (NO), endothelin-1 (ET-1), neuropeptide Y (NPY) in the serum. DAB chromogenic in situ hybridization (CISH) was provided to detect the mRNA expression of endothelial nitric oxide synthase (eNOS) in the internal carotid artery and the arcuate nucleus (ARC), and that of CGRP in the paraventricular nucleus posterior (PVP) and the ventrolateral medulla (VLM). Liquid chromatography-mass spectrometry (LC-MS) was used to detect the levels of epinephrine (E) and norepinephrine (NE) in the paraventricular nucleus anterior (PVA). Compared with the normal group, the irritation scores, systolic blood pressure and diastolic blood pressure were increased at each time point in the model group (P<0.05). When compared with the model group, the irritation scores after the intervention for 3 and 4 weeks, and systolic and diastolic blood pressure after intervention for 2, 3 and 4 weeks were reduced in the acupuncture group (P<0.05). In comparison with the normal group, the serum CGRP and NO levels of the rats were decreased (P<0.05), and the serum ET-1 and NPY levels, as well as E and EN levels in PVA were increased (P<0.05) in the model group. The levels of serum CGRP and NO were elevated (P<0.05), and the serum ET-1 and NPY levels, as well as E and EN levels of PVA were reduced (P<0.05) in the acupuncture group when compared with those of the model group. In the model group, the media of internal carotid artery exhibited thickening and remodeling, while the neuron volume in ARC was small. In the acupuncture group, every layer of internal carotid artery was acceptably arranged, and the parvicellular neuron of ARC was moderate in volume. For the in situ hybridization of eNOS mRNA for the rats of each group, the smooth muscle cells were predominantly expressed in each layer of the internal carotid artery, whereas the expression of parvicellular neurons was dominated in ARC. In the model group, the large and small neurosecretory cells were distributed sparsely in the nerves of PVP; in the acupuncture group, the cells of these two species were distributed regularly; and there were few species of glial cell in the VLM of either the model group or the acupuncture group. In each group, for the in situ hybridization of CGRP mRNA, the small neurosecretory cells were expressed predominately in the PVP, while, the expression of glial cell nuclei and the cell cytoplasm was dominated in the VLM. Compared with the normal group, the mRNA expression of eNOS in the internal carotid artery and ARC and that of CGRP mRNA in the PVP and VLM was decreased in the model group (P<0.05). In the acupuncture group, when compared with the model group, the mRNA expression of eNOS in the internal carotid artery and ARC and that of CGRP in the PVP and VLM was increased in the acupuncture group (P<0.05). Acupuncture at "antihypertensive acupoint prescription" can upregulate the level of vascular relaxing factors, downregulate the level of contracting factors, enhance the response of relaxing factors in targeting blood vessels and regulating the center. The mechanism may be related to the modulation of the sympathetic-adrenergic autonomic neurotransmitters in the paraventricular nucleus in spontaneous hypertension rats.

Electroacupuncture alleviates myocardial ischemia-reperfusion injury by inhibiting hypothalamic paraventricular nucleus neurons projecting to the rostral ventrolateral medulla.

Accumulating evidence suggests that electroacupuncture (EA) has obvious therapeutic effects and unique advantages in alleviating myocardial ischemia-reperfusion injury (MIRI), while the underlying neuromolecular mechanisms of EA intervention for MIRI have not been fully elucidated. The aim of the study is to investigate the role of the neural pathway of hypothalamic paraventricular nucleus (PVN) neurons projecting to the rostral ventrolateral medulla (RVLM) in the alleviation of MIRI rats by EA preconditioning. MIRI models were established by ligating the left anterior descending coronary artery for 30 min followed by reperfusion for 2h. Electrocardiogram recording, chemogenetics, enzyme-linked immunosorbent assay, multichannel physiology recording and haematoxylin-eosin and immunofluorescence staining methods were conducted to demonstrate that the firing frequencies of neurons in the PVN and the expression of c-Fos decreased by EA pretreatment. Meanwhile, EA preconditioning significantly reduced the levels of creatine kinase isoenzymes (CK-MB), cardiac troponin I (cTnI) and lactic dehydrogenase (LDH). Virus tracing showed a projection connection between PVN and RVLM. The inhibition of the PVN-RVLM neural pathway could replicate the protective effect of EA pretreatment on MIRI rats. However, the activation of the pathway weakened the effect of EA preconditioning. EA pretreatment alleviated MIRI by regulating PVN neurons projecting to RVLM. This work provides novel evidence of EA pretreatment for alleviating MIRI.

Gestationally administered RAS modulators reprogram endotoxic cardiovascular and inflammatory profiles in adult male offspring of preeclamptic rats.

Previous studies showed that preeclampsia (PE) amplifies cardiovascular dysfunction induced by endotoxemia in adult male, but not female, offspring. Here, we asked if such aggravated endotoxic insult could be nullified by modulators of the renin-angiotensin system (RAS). PE was induced by gestational administration of Nω-nitro-L-arginine methyl ester(L-NAME, a nitric oxide synthase inhibitor). Adult male offspring of PE mothers treated gestationally with angiotensin 1-7 (Ang1-7, angiotensin II-derived vasodilator), losartan (AT1 receptor antagonist), pioglitazone (peroxisome proliferator-activated receptor gamma, PPARγ, agonist), or combined losartan/pioglitazone were instrumented with femoral indwelling catheters and challenged intravenously with a 5-mg/kg dose of lipopolysaccharides (LPS, 5mg/kg). LPS caused significant decreases in blood pressure (BP) and spectral index of overall heart rate variability and increases in heart rate and left ventricular contractility (dP/dtmax). These effects were mostly reduced to similar magnitudes by individual drug therapies. In offspring born to Ang1-7-treated dams, the spectral index of cardiac sympathovagal balance showed elevated sympathetic dominance in response to LPS. Immunohistochemistry revealed that Ang1-7, but not losartan/pioglitazone, abolished the exaggerated increases in toll-like receptor 4 (TLR-4) expression caused by PE/LPS in heart tissues and neuronal circuits of brainstem rostral ventrolateral medulla (RVLM). By contrast, the losartan/pioglitazone regimen, but not Ang1-7, decreased and increased angiotensin converting enzyme (ACE) and ACE2 expression, respectively. Together, gestational fetal reprogramming of Ang II (depression) and Ang1-7 (activation) arms of RAS effectively counterbalance worsened endotoxic cardiovascular and inflammatory profiles in adult male offspring of PE rats.

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.

MiR-193b-3p and miR-346 Exert Antihypertensive Effects in the Rostral Ventrolateral Medulla.

Rostral ventrolateral medulla (RVLM) neuron hyperactivity raises sympathetic outflow, causing hypertension. MicroRNAs (miRNAs) contribute to diverse biological processes, but their influence on RVLM neuronal excitability and blood pressure (BP) remains widely unexplored. The RVLM miRNA profiles in spontaneously hypertensive rats were unveiled using RNA sequencing. Potential effects of these miRNAs in reducing neuronal excitability and BP and underlying mechanisms were investigated through various experiments. Six hundred thirty-seven miRNAs were identified, and reduced levels of miR-193b-3p and miR-346 were observed in the RVLM of spontaneously hypertensive rats. Increased miR-193b-3p and miR-346 expression in RVLM lowered neuronal excitability, sympathetic outflow, and BP in spontaneously hypertensive rats. In contrast, suppressing miR-193b-3p and miR-346 expression in RVLM increased neuronal excitability, sympathetic outflow, and BP in Wistar Kyoto and Sprague-Dawley rats. Cdc42 guanine nucleotide exchange factor (Arhgef9) was recognized as a target of miR-193b-3p. Overexpressing miR-193b-3p caused an evident decrease in Arhgef9 expression, resulting in the inhibition of neuronal apoptosis. By contrast, its downregulation produced the opposite effects. Importantly, the decrease in neuronal excitability, sympathetic outflow, and BP observed in spontaneously hypertensive rats due to miR-193b-3p overexpression was greatly counteracted by Arhgef9 upregulation. miR-193b-3p and miR-346 are newly identified factors in RVLM that hinder hypertension progression, and the miR-193b-3p/Arhgef9/apoptosis pathway presents a potential mechanism, highlighting the potential of targeting miRNAs for hypertension prevention.

Effects of the Nucleus Raphe Magnus Stimulation on Nociceptive Neurons of the Rat Caudal Ventrolateral Medulla in Normal Conditions and after Intestinal Inflammation

Effects of the Nucleus Raphe Magnus Stimulation on Nociceptive Neurons of the Rat Caudal Ventrolateral Medulla in Normal Conditions and after Intestinal Inflammation

Acupuncture modulation of the ACE/Ang II/AT1R and ACE2/Ang(1-7)/MasR pathways in the rostral ventrolateral medulla reduces sympathetic output and prevents cardiac injury caused by SHR hypertension.

Acupuncture can reduce blood pressure, heart rate (HR), and ameliorate cardiac damage by modulating the excitability of the sympathetic nervous system, but the exact mechanism of this effect remains unclear. This study investigated the potential mechanisms of acupuncture in the treatment of cardiac damage in hypertension. Spontaneously hypertensive rats (SHR) were used as the hypertension model with Wistar-Kyoto rats as the control. Manual acupuncture, electroacupuncture, and metoprolol were used as interventions. Systolic and diastolic blood pressure (SBP, DBP) plus HR were monitored with cardiac structure determined using Masson staining. Angiotensin II (Ang II) and norepinephrine in myocardium were detected with ELISA as was Ang(1-7) and gamma aminobutyric acid (GABA) in the rostral ventrolateral medulla (RVLM). Expression of mRNA for collagen type I (Col-I), Col-III, actin α1 (ACTA1), and thrombospondin 4 (THBS4) in myocardium was detected using real-time PCR. Expression of angiotensin converting enzyme (ACE), Ang II, angiotensin II type 1 receptor (AT1R), ACE2, and Mas receptor (MasR) proteins in RVLM was monitored using western blot. After manual acupuncture and electroacupuncture treatment, SHRs showed decreased SBP, DBP and HR, reduced myocardial damage. There was decreased expression of the ACE/Ang II/AT1R axis, and increased expression of the ACE2/Ang(1-7)/MasR axis within the RVLM. GABA levels were increased within the RVLM and norepinephrine levels were decreased in myocardial tissue. Metoprolol was more effective than either manual acupuncture or electroacupuncture. Acupuncture directed against hypertensive cardiac damage may be associated with regulation of ACE/Ang II/AT1R and the ACE2/Ang(1-7)/MasR pathway within the RLVM to reduce cardiac sympathetic excitability.

Molecular Organization of Autonomic, Respiratory, and Spinally-Projecting Neurons in the Mouse Ventrolateral Medulla.

The ventrolateral medulla (VLM) is a crucial region in the brain for visceral and somatic control, serving as a significant source of synaptic input to the spinal cord. Experimental studies have shown that gene expression in individual VLM neurons is predictive of their function. However, the molecular and cellular organization of the VLM has remained uncertain. This study aimed to create a comprehensive dataset of VLM cells using single-cell RNA sequencing in male and female mice. The dataset was enriched with targeted sequencing of spinally-projecting and adrenergic/noradrenergic VLM neurons. Based on differentially expressed genes, the resulting dataset of 114,805 VLM cells identifies 23 subtypes of neurons, excluding those in the inferior olive, and five subtypes of astrocytes. Spinally-projecting neurons were found to be abundant in seven subtypes of neurons, which were validated through in situ hybridization. These subtypes included adrenergic/noradrenergic neurons, serotonergic neurons, and neurons expressing gene markers associated with premotor neurons in the ventromedial medulla. Further analysis of adrenergic/noradrenergic neurons and serotonergic neurons identified nine and six subtypes, respectively, within each class of monoaminergic neurons. Marker genes that identify the neural network responsible for breathing were concentrated in two subtypes of neurons, delineated from each other by markers for excitatory and inhibitory neurons. These datasets are available for public download and for analysis with a user-friendly interface. Collectively, this study provides a fine-scale molecular identification of cells in the VLM, forming the foundation for a better understanding of the VLM's role in vital functions and motor control.

The effect of microvascular decompression of the CN IX-X root entry/exit zone and the ventrolateral medulla in neurogenic hypertension involving the vertebral/basilar artery.

Neurogenic hypertension (HTN) is a type of HTN characterized by increased activity of the sympathetic nervous system. Vascular compression is one of the pathogenic mechanisms of neurogenic HTN. Despite Jannetta's solid anatomical and physiological arguments in favor of neurogenic HTN in the 1970's, the treatment for essential HTN by microvascular decompression (MVD) still lacks established selection criteria. Therefore, the subjects selected for our center were limited to patients with primary trigeminal neuralgia (TN) and primary hemifacial spasm (HFS) of the vertebral/basilar artery (VA/BA) responsible vessel type coexisting with neurogenic HTN who underwent MVD of the brainstem to further explore possible indications for MVD in the treatment of neurogenic HTN. A retrospective analysis of 63 patients who were diagnosed with neurogenic HTN had symptoms of HFS and TN cranial nerve disease. Patients were treated at our neurosurgery department from January 2018 to January 2023. A preoperative magnetic resonance examination of the patients revealed the presence of abnormally located vascular compression in the rostral ventrolateral medulla (RVLM) and the root entry zone (REZ) of the IX and X cranial nerves (CN IX- X). There was no significant difference between the two groups in terms of gender, age, course of HFS, course of TN, course of HTN, degree of HTN, or preoperative blood pressure. Based on the postoperative blood pressure levels, nine out of 63 patients were cured (14.28%), eight cases (12.70%) showed a marked effect, 16 cases (25.40%) were effective, and 30 cases were invalid (47.62%). The overall efficacy was 52.38%. However, 39 cases of combined cranial nerve disease were on the left side of the efficacy rate (66.67%) and 24 cases of combined cranial nerve disease were on the right side of the efficacy rate (29.16%). Over the last few decades, many scholars have made pioneering progress in the clinical retrospective study of MVD for neurogenic hypertension, and our study confirms the efficacy of MVD in treating vertebral/basilar artery-type neurogenic hypertension by relieving the vascular pressure of RVLM. In the future, with the development and deepening of pathological mechanisms and clinical observational studies, MVD may become an important treatment for neurogenic hypertension by strictly grasping the surgical indications. MVD is an effective treatment for neurogenic HTN. Indications may include the following: left-sided TN or HFS combined with neurogenic HTN; VA/BA compression in the left RVLM and REZ areas on MRI; and blood pressure in these patients cannot be effectively controlled by drugs.

Mapping of afferent and efferent connections of phenylethanolamine N-methyltransferase-expressing neurons in the nucleus tractus solitarii.

Phenylethanolamine N-methyltransferase (PNMT)-expressing neurons in the nucleus tractus solitarii (NTS) contribute to the regulation of autonomic functions. However, the neural circuits linking these neurons to other brain regions remain unclear. This study aims to investigate the connectivity mechanisms of the PNMT-expressing neurons in the NTS (NTSPNMT neurons). The methodologies employed in this study included a modified rabies virus-based retrograde neural tracing technique, conventional viral anterograde tracing, and immunohistochemical staining procedures. A total of 43 upstream nuclei projecting to NTSPNMT neurons were identified, spanning several key brain regions including the medulla oblongata, pons, midbrain, cerebellum, diencephalon, and telencephalon. Notably, dense projections to the NTSPNMT neurons were observed from the central amygdaloid nucleus, paraventricular nucleus of the hypothalamus, area postrema, and the gigantocellular reticular nucleus. In contrast, the ventrolateral medulla, lateral parabrachial nucleus, and lateral hypothalamic area were identified as the primary destinations for axon terminals originating from NTSPNMT neurons. Additionally, reciprocal projections were evident among 21 nuclei, primarily situated within the medulla oblongata. Our research findings demonstrate that NTSPNMT neurons form extensive connections with numerous nuclei, emphasizing their essential role in the homeostatic regulation of vital autonomic functions.

Esaxerenone: blood pressure reduction and cardiorenal protection without reflex sympathetic activation in salt-loaded stroke-prone spontaneously hypertensive rats.

Mineralocorticoid receptor (MR) is involved in the mechanisms of blood pressure elevation, organ fibrosis, and inflammation. MR antagonists have been used in patients with hypertension, heart failure, or chronic kidney disease. Esaxerenone, a recently approved MR blocker with a nonsteroidal structure, has demonstrated a strong blood pressure-lowering effect. However, blood pressure reduction may lead to sympathetic activation through the baroreflex. The effect of esaxerenone on the sympathetic nervous system remains unclear. We investigated the effect of esaxerenone on organ damage and the sympathetic nervous system in salt-loaded stroke-prone spontaneously hypertensive rats (SHRSP), a well-established model of essential hypertension with sympathoexcitation and organ damage. Three-week administration of esaxerenone or hydralazine successfully attenuated the blood pressure elevation. Both esaxerenone and hydralazine comparably suppressed left ventricular hypertrophy and urinary albumin excretion. However, renal fibrosis and glomerular sclerosis were suppressed by esaxerenone but not hydralazine. Furthermore, plasma norepinephrine level, a parameter of systemic sympathetic activity, was significantly increased by hydralazine but not by esaxerenone. Consistent with these findings, the activity of the control centers of sympathetic nervous system, the parvocellular region of the paraventricular nucleus in the hypothalamus and the rostral ventrolateral medulla, was enhanced by hydralazine but remained unaffected by esaxerenone. These results suggest that esaxerenone effectively lowers blood pressure without inducing reflex sympathetic nervous system activation. Moreover, the organ-protective effects of esaxerenone appear to be partially independent of its blood pressure-lowering effect. In conclusion, esaxerenone demonstrates a blood pressure-lowering effect without concurrent sympathetic activation and exerts organ-protective effects in salt-loaded SHRSP. Esaxerenone has antihypertensive and cardiorenal protective effects without reflex sympathetic activation in salt-loaded stroke-prone spontaneously hypertensive rats.

Ventrolateral medullary compression by vascular contact in primary hemifacial spasm: a radiological analysis.

The neurovascular conflict (NVC) causing hemifacial spasm (HFS) can also cause compression of ventrolateral medulla (VLM) which contains the central sympathetic neurons. VLM compression has been associated with hypertension. Whether the VLM compression in HFS patients is associated with hypertension is not clear. To determine the frequency, severity of VLM compression and its association with hypertension in HFS patients. A cross-sectional, hospital-based, case control study and recruited 120 study subjects (50 cases of primary HFS, 30 hypertensive and 40 normotensive age-, sex- matched controls). The VLM compression was assessed in magnetic resonance imaging Constructive Interference in Steady State (CISS) 3D sequences. Hypertension was present in 30 cases (60%). Six patients with HFS (20%) were detected to be hypertensive after the onset of HFS. VLM compression was seen in 24 cases (48%), 7 hypertensive controls (23.3%) and 5 normotensive controls (10%) (p = 0.03). Twenty-four patients with hypertension had VLM compression and remaining 6 patients with hypertension did not have VLM compression (80% vs 20%; p = 0.02). Normotensive patients did not have VLM compression. Vertebral artery was the most common artery causing VLM compression (22 patients; 7 hypertensive and 5 normotensive controls). VLM compression is more common in HFS patients as compared to hypertensive and normotensive controls. It is more common in hypertensive HFS patients in comparison with normotensive HFS patients. Microvascular decompression is an option in hypertensive HFS patients with VLM compression if the hypertension is medically refractory.

Molecular organization of adrenergic and noradrenergic neurons in the mouse ventrolateral medulla

Catecholaminergic neurons in the ventrolateral medulla (VLM) regulate many important aspects of physiology, including the sympathetic control of blood pressure, glucose homeostasis, immune function, and the neuroendocrine responses to inflammation and stress. Previous studies have parsed catecholaminergic VLM neurons into distinct sub-populations based on neurochemical content, connectivity, and distinct response to the activation of autonomic reflexes. However, there is no consensus for the molecular organization of catecholaminergic neurons. In this study, we used high-throughput single nuclei RNA sequencing combined with anatomically- and genetically-targeted cell-sorting of spinally-projecting and catecholaminergic neurons to systematically characterize C1 and A1 neurons in the mouse VLM. Our results, based on an analysis of 894 catecholaminergic neurons derived from a dataset of 114,805 single-nuclei transcriptomes, indicate that the VLM contains 7 molecularly distinct subpopulations of catecholaminergic neurons, including 5 adrenergic subtypes and 2 noradrenergic subtypes identified by the expression of enzymes involved in the production of catecholamine ( Th, Dbh) and the vesicular transporters for glutamate ( Slc6a17) and noradrenaline ( Slc6a2). Neurons with projections to the spinal cord were present in 4 of 5 adrenergic cell clusters, and 1 noradrenergic cell cluster. We histologically verified two novel markers of the adrenergic ( Hk2) and noradrenergic ( Eya1) neurons using RNA fluorescent in situ hybridization. Hk2 expression was found primarily in Dbh+ neurons in the rostral VLM, including both bulbospinal and non-bulbospinal neurons, as well as majority of Pnmt+ neurons in the rostral VLM and C2/C3 regions. Eya1 co-localized with Dbh and Slc6a2 consistently in the caudal VLM, aligning with the location of the A1 neurons, whereas Eya1 signal was diffuse in the rostral VLM. Collectively, this shows that Hk2 expression is enriched in adrenergic neurons as judged by conventional criteria irrespective of the cell's location in the medulla, whereas Eya1 is enriched in brainstem noradrenergic neurons and provides a marker for A1 neurons. These data show molecularly distinct adrenergic and noradrenergic neurons are spatially intermixed in the VLM, and provides novel gene markers for distinct subtypes of catecholaminergic neurons that can be used to dissect the function in future studies. This work was supported by National Institute of Health (NIH) R01 HL148004 to SBGA; and R01 HL153916, Pathway to Stop Diabetes Initiator Award 1-18-INI-14, and UVA 3Cavaliers grant to JNC. 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.

Decreased inhibition of ventral brainstem liver-related neurons in high-fat diet fed mice

Pre-sympathetic liver-related neurons in the ventrolateral and ventromedial medulla (VLM/VMM) participate in the regulation of hepatic carbohydrate and lipid metabolism. Increased sympathetic activity observed in diet-induced obese mice likely contributes to excessive hepatic glucose production, and thus to the development, and/or progression of type 2 diabetes mellitus. At the cellular level, change in neuronal activity and/or synaptic balance is one of the underlying mechanisms that can lead to increased sympathetic outflow. In this study, we tested the hypothesis that high-fat diet (HFD) diminishes glycinergic inhibition of liver-related neurons in the VLM/VMM. We crossed heterozygous glycine transporter 2 Cre (GlyT2Cre) mice with floxed channelrhodopsin 2 (ChR2-EYFP) mice to generate GlyT2-ChR2 expressing mice (GlyT2ChR2/EYFP). Liver-related neurons were identified in the VLM/VMM with a retrograde viral tracer and patch-clamp recordings were conducted. Whole-cell recordings from pre-sympathetic, liver-related neurons revealed that spontaneous inhibitory postsynaptic currents (sIPSCs) are mediated by GABA and glycine. Our data showed that increasing the activity of inhibitory inputs leads to increased glycine release. Next, light stimulation was used to trigger glycine release in GlyT2ChR2/EYFP mice. Light stimulation of ChR2-expressing glycinergic fibers triggered evoked IPSCs (eIPSCs) generated by co-release of GABA and glycine with identical time courses for the two neurotransmitters. Intriguingly, in HFD-fed mice we found that the amplitude of light evoked IPSCs was significantly decreased. These data suggest that liver-related neurons in the VLM/VMM are co-inhibited by GABA and glycine and in HFD-fed mice inhibition of liver-related neurons is reduced. This reduced inhibition of ventral brainstem neurons may contribute to increased sympathetic activity in diet-induced obesity. Supported by: NIDDK122842 and Tulane Brain Institute Marko Spark Innovation Research Fund. 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.

The roles of extrinsic neurons in modulating neuroimmune crosstalk during acute enteric infection with Citrobacter rodentium

Background: The intestinal tract is one of the most densely innervated organs in our body, containing extrinsic sensory and sympathetic neurons. With their cell bodies located outside of the gut, these neurons directly detect enteric pathogens and release immunomodulatory neurotransmitters or neuropeptides to communicate with immune cells in the gut. Specifically, sympathetic neurons secrete neurotransmitter norepinephrine, which activate β2 adrenergic receptors (β2ΑR) on immune cells to regulate immune functions. Recent studies from our lab, as well as others, have begun to highlight the neural regulation of immune responses during enteric infections. However, the precise pathways responsible for orchestrating these effects remain elusive. Hypothesis: We sought to determine whether extrinsic sensory and sympathetic neurons are implicated in host protective responses against enteric pathogen Citrobacter rodentium (C. rodentium). Methods: Mice were inoculated by oral gavage with C. rodentium (108 colony-forming units) or sterile luria broth. Bacterial colonization was quantified by homogenization of distal colon tissues and plating serial dilutions onto MacConkey agar. Relevant tissues were collected for qPCR, flow cytometry or histology. Genetically engineered Arc Targeted Recombination in Active Population (TRAP) mice were used to identify neurons that were activated during infection. These ArcTRAP mice received single intraperitoneal (i.p.) injection of 4-hydroxytamoxifen post-infection to “trap” active neurons, marking them tdTomato+. 6-hydroxydopamine (6OHDA) was used to chemically ablate sympathetic innervations to the colon via 3 consecutive daily i.p. injections. Results: At day 4 post-infection, there were significantly more active tdTomato+ neurons in the rostral ventrolateral medulla (RVLM) of the infected mice compared to the non-infected controls. This increase was more pronounced as the infection peaked at day 10, indicating that more neurons were being activated in the RVLM in response to C. rodentium in the gut. 6OHDA treatment significantly increased bacterial burden in distal colon at day 10 post-infection. This was associated with decreased tyrosine hydroxylase expression compared to the vehicle controls. Subsequently, qPCR revealed that there was increased pro-inflammatory cytokines expression in the colon of sympathectomized infected mice compared to the infected controls. Similarly, β2AR knockout significantly increased bacterial burden in distal colon at day 3 and 10 post-infection. Increased pro-inflammatory cytokines expression suggests an immunomodulatory role for β2AR during C. rodentium infection. Conclusions: These studies provided novel insights into the neuroimmune crosstalk that is crucial for acute enteric infections with C. rodentium. Specifically, these experiments may reveal how extrinsic neurons shape local immune responses in the gut. NIH NIAID R01AI150647; Graduate Student Support Program (UC Davis: School of Veterinary Medicine). 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.

Pre-sympathetic C1 RVLM neurons drives sleep disordered breathing in heart failure through activation of central chemoreceptors

Sleep-disordered breathing is closely linked to heart failure (HF) morbidity and mortality. Recently, we showed that photo-stimulation of pre-sympathetic motor neurons located within the rostro ventrolateral medulla (RVLM) drive sleep-disordered breathing in healthy conditions. However, the neural circuitry required for these effects and whether this pathway is relevant for HF pathophysiology is not known. Accordingly, we aimed to determine: i) if the generation of RVLM-induced altered breathing encompasses the activation of chemoreceptor neurons from the retrotrapezoid nucleus (RTN), and ii) if selective ablation of RVLM neurons improves breathing control and sleep architecture in HF rats. Sprague-Dawley rats received stereotaxic injections of a lenti-viral vector containing a light-sensitive cation channel (Channelrhodopsin2) in the RVLM and a Substance P-saporin toxin conjugate into the RTN. Then, 200 mm fiber optics were implanted for RVLM neurons photo-stimulation. A second group underwent volume overload to induced HF and ablation of RVLM neurons was performed using a DbH-saporin toxin (DbH-SAP). EEG and EMG leads were implanted, and rats allowed to recover before sleep/breathing studies. We found that photo-stimulation of RVLM neurons triggered breathing disorders only in the presence of a functional RTN neurons. Indeed, breathing irregularity score following RVLM photo-stimulation showed a two-fold increase (9±2 vs. 20±3, pre vs. post-stimulation) and this was absent in the presence of Substance P-saporin (10±3 vs. 12±3, pre vs. post-stimulation with Substance P-saporin). Compared to Sham rats, HF animals showed overt signs of disordered breathing and sleep fragmentation and ablation of RVLM neurons using DbH-SAP significantly improves both breathing and sleep function in HF. Our results showed that RVLM drives breathing disorders through activation of the RTN and that selective ablation of RVLM neurons offer salutary benefit for the control of sleep and irregular breathing patterns in the setting of HF. Fondecyt 1220950. 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.