Sympathetic Premotor Research Articles

Activation of orexin-A (hypocretin-1) receptors in the Raphe Pallidus at different ambient temperatures in the rat: effects on thermoregulation, cardiovascular control, sleep, and feeding behavior.

The Raphe Pallidus (RPa) is a brainstem nucleus containing sympathetic premotor neurons that control thermogenesis and modulate cardiovascular function. It receives inputs from various hypothalamic areas, including the Lateral Hypothalamus (LH), a heterogeneous region intricately involved in several autonomic and behavioral functions. A key subpopulation of neurons in the LH expresses orexin/hypocretin, a neuropeptide which is crucially involved in the regulation of the wake-sleep states and feeding behavior. The RPa receives orexinergic projections from the LH and orexinergic signalling in the RPa has been shown to enhance thermogenesis in the anaesthetized rat, but only in the presence of an already existing thermogenic drive, without significantly affecting cardiovascular function. The present work was aimed at exploring the effects on thermoregulation and autonomic function and the possible role in the modulation of the wake-sleep states and feeding behavior of orexin injection in the RPa in the free-behaving rat. In order to assess the influence of an already present thermogenic drive on orexinergic signalling in the RPa, animals were studied at three different ambient temperatures (Ta, 10°C, 24°C, and 32°C). We found that orexin injection into the RPa variably affected the wake-sleep states, autonomic functions, motor activity, and feeding behavior, at the different Tas. In particular, in the first post-injection hour, we observed an increase in wakefulness, which was large at Ta 24°C and Ta 10°C and rather mild at Ta 32°C. Deep brain temperature was increased by orexin injection at Ta 10°C, but not at either Ta 24°C or Ta 32°C. Moreover, an increase in mean arterial blood pressure occurred at Ta 24°C, which was probably masked by the high baseline levels at Ta 10°C and was completely absent at Ta 32°C. Finally, an enhancement in feeding behavior was observed at Ta 24°C and 10°C only. In accordance with what observed in anaesthetized rats, orexinergic signalling in the RPa seems to be ineffective in the absence of any thermogenic drive. Moreover, the effects observed on the wake-sleep states and feeding behavior introduce the RPa as a novel player in the central neural network promoting wakefulness and feeding.

Mediobasal hypothalamic neurons contribute to the control of brown adipose tissue sympathetic nerve activity and cutaneous vasoconstriction

The mediobasal hypothalamus (MBH) contains heterogeneous neuronal populations that regulate food intake and energy expenditure. However, the role of MBH neurons in the neural control of thermoeffector activity for thermoregulation is not known. This study sought to determine the effects of modulating the activity of MBH neurons on the sympathetic outflow to brown adipose tissue (BAT), BAT thermogenesis, and cutaneous vasomotion. Pharmacological inhibition of MBH neurons by local administration of muscimol, a GABAA receptor agonist, reduced skin cooling-evoked BAT thermogenesis, expired CO2, body temperature, heart rate, and mean arterial pressure, while blockade of GABAA receptors by nanoinjection of bicuculline in the MBH induced large increases in BAT sympathetic nerve activity (SNA), BAT temperature, body temperature, expired CO2, heart rate, and cutaneous vasoconstriction. Neurons in the MBH send projections to neurons in the dorsal hypothalamic area and dorsomedial hypothalamus (DMH), which excite sympathetic premotor neurons in the rostral raphe pallidus area (rRPa) that control sympathetic outflow to BAT. The increases in BAT SNA, BAT temperature, and expired CO2 elicited by blockade of GABAA receptors in the MBH were reversed by blocking excitatory amino acid receptors in the DMH or in the rRPa. Together, our data show that MBH neurons provide a modest contribution to BAT thermogenesis for cold defense, while GABAergic disinhibition of these neurons produces large increases in the sympathetic outflow to BAT, and cutaneous vasoconstriction. Activation of glutamate receptors on BAT thermogenesis-promoting neurons of the DMH and rRPa is necessary for the increased sympathetic outflow to BAT evoked by disinhibition of MBH neurons. These data demonstrate neural mechanisms that contribute to the control of thermoeffector activity, and may have important implications for regulating body temperature and energy expenditure.

Acute deep brain stimulation of the paraventricular nucleus of the hypothalamus increases brown adipose tissue thermogenesis in rats

Brown adipose tissue (BAT) activity is controlled by the sympathetic nervous system. Activation of BAT has shown significant promise in preclinical studies to elicit weight loss. Since the hypothalamic paraventricular nucleus (PVN) contributes to the regulation of BAT thermogenic activity, we sought to determine the effects of electrical stimulation of the PVN as a model of deep brain stimulation (DBS) for increasing BAT sympathetic nerve activity (SNA). The rostral raphe pallidus area (rRPa) was also chosen as a target for DBS since it contains the sympathetic premotor neurons for BAT. Electrical stimulation (100 µA, 100 µs, 100 Hz, for 5 min at a 50 % duty cycle) of the PVN increased BAT SNA and BAT thermogenesis. These effects were prevented by a local nanoinjection of bicuculline, a GABAA receptor antagonist. We suggest that electrical stimulation of the PVN elicited local release of GABA, which inhibited BAT sympathoinhibitory neurons in PVN, thereby releasing a restraint on BAT SNA. Electrical stimulation of the rRPa inhibited BAT thermogenesis and this was prevented by a local nanoinjection of bicuculline, suggesting that local release of GABA suppressed BAT SNA. Electrical stimulation of the PVN activates BAT metabolism via a mechanism that may include activation of local GABAA receptors. These findings contribute to our understanding of the mechanisms underlying the effects of DBS in the regulation of fat metabolism and provide a foundation for further DBS studies targeting hypothalamic circuits regulating BAT thermogenesis as a therapy for obesity.

Descending pathways from the superior colliculus mediating autonomic and respiratory effects associated with orienting behaviour.

The ability to discriminate competing external stimuli and initiate contextually appropriate behaviours is a key brain function. Neurons in the deep superior colliculus (dSC) integrate multisensory inputs and activate descending projections to premotor pathways responsible for orienting, attention and defence, behaviours which involve adjustments to respiratory and cardiovascular parameters. However, the neural pathways that subserve the physiological components of orienting are poorly understood. We report that orienting responses to optogenetic dSC stimulation are accompanied by short-latency autonomic, respiratory and electroencephalographic effects in awake rats, closely mimicking those evoked by naturalistic alerting stimuli. Physiological responses were not accompanied by detectable aversion or fear, and persisted under urethane anaesthesia, indicating independence from emotional stress. Anterograde and trans-synaptic viral tracing identified a monosynaptic pathway that links the dSC to spinally projecting neurons in the medullary gigantocellular reticular nucleus (GiA), a key hub for the coordination of orienting and locomotor behaviours. In urethane-anaesthetized animals, sympathoexcitatory and cardiovascular, but not respiratory, responses to dSC stimulation were replicated by optogenetic stimulation of the dSC-GiA terminals, suggesting a likely role for this pathway in mediating the autonomic components of dSC-mediated responses. Similarly, extracellular recordings from putative GiA sympathetic premotor neurons confirmed short-latency excitatory inputs from the dSC. This pathway represents a likely substrate for autonomic components of orienting responses that are mediated by dSC neurons and suggests a mechanism through which physiological and motor components of orienting behaviours may be integrated without the involvement of higher centres that mediate affective components of defensive responses. KEY POINTS: Neurons in the deep superior colliculus (dSC) integrate multimodal sensory signals to elicit context-dependent innate behaviours that are accompanied by stereotypical cardiovascular and respiratory activities. The pathways responsible for mediating the physiological components of colliculus-mediated orienting behaviours are unknown. We show that optogenetic dSC stimulation evokes transient orienting, respiratory and autonomic effects in awake rats which persist under urethane anaesthesia. Anterograde tracing from the dSC identified projections to spinally projecting neurons in the medullary gigantocellular reticular nucleus (GiA). Stimulation of this pathway recapitulated autonomic effects evoked by stimulation of dSC neurons. Electrophysiological recordings from putative GiA sympathetic premotor neurons confirmed short latency excitatory input from dSC neurons. This disynaptic dSC-GiA-spinal sympathoexcitatory pathway may underlie autonomic adjustments to salient environmental cues independent of input from higher centres.

Disparate Roles of Oxidative Stress in Rostral Ventrolateral Medulla in Age-Dependent Susceptibility to Hypertension Induced by Systemic l-NAME Treatment in Rats.

This study aims to investigate whether tissue oxidative stress in the rostral ventrolateral medulla (RVLM), where sympathetic premotor neurons reside, plays an active role in age-dependent susceptibility to hypertension in response to nitric oxide (NO) deficiency induced by systemic l-NAME treatment, and to decipher the underlying molecular mechanisms. Systolic blood pressure (SBP) and heart rate (HR) in conscious rats were recorded, along with measurements of plasma and RVLM level of NO and reactive oxygen species (ROS), and expression of mRNA and protein involved in ROS production and clearance, in both young and adult rats subjected to intraperitoneal (i.p.) infusion of l-NAME. Pharmacological treatments were administered by oral gavage or intracisternal infusion. Gene silencing of target mRNA was made by bilateral microinjection into RVLM of lentivirus that encodes a short hairpin RNA (shRNA) to knock down gene expression of NADPH oxidase activator 1 (Noxa1). We found that i.p. infusion of l-NAME resulted in increases in SBP, sympathetic neurogenic vasomotor activity, and plasma norepinephrine levels in an age-dependent manner. Systemic l-NAME also evoked oxidative stress in RVLM of adult, but not young rats, accompanied by augmented enzyme activity of NADPH oxidase and reduced mitochondrial electron transport enzyme activities. Treatment with L-arginine via oral gavage or infusion into the cistern magna (i.c.), but not i.c. tempol or mitoQ10, significantly offset the l-NAME-induced hypertension in young rats. On the other hand, all treatments appreciably reduced l-NAME-induced hypertension in adult rats. The mRNA microarray analysis revealed that four genes involved in ROS production and clearance were differentially expressed in RVLM in an age-related manner. Of them, Noxa1, and GPx2 were upregulated and Duox2 and Ucp3 were downregulated. Systemic l-NAME treatment caused greater upregulation of Noxa1, but not Ucp3, mRNA expression in RVLM of adult rats. Gene silencing of Noxa1 in RVLM effectively alleviated oxidative stress and protected adult rats against l-NAME-induced hypertension. These data together suggest that hypertension induced by systemic l-NAME treatment in young rats is mediated primarily by NO deficiency that occurs both in vascular smooth muscle cells and RVLM. On the other hand, enhanced augmentation of oxidative stress in RVLM may contribute to the heightened susceptibility of adult rats to hypertension induced by systemic l-NAME treatment.

An oxytocinergic neural pathway that stimulates thermogenic and cardiac sympathetic outflow.

Oxytocin alters autonomic functions besides social behaviors. However, the central neuronal links between hypothalamic oxytocinergic neurons and the autonomic nervous system remain unclear. Here we show that oxytocinergic neurons in the rat paraventricular hypothalamic nucleus (PVH), a pivotal site for energy homeostasis, innervate sympathetic premotor neurons in the rostral medullary raphe region (rMR) to stimulate brown adipose tissue (BAT) thermogenesis and cardiovascular functions. Oxytocin receptor stimulation in the rMR evokes BAT thermogenesis and tachycardia. Invivo optogenetic stimulation of the PVH→rMR long-range oxytocinergic pathway, using a virus-mediated system for amplified gene expression in oxytocinergic neurons, not only elicits BAT thermogenic and cardiac responses but also potentiates sympathetic responses evoked by glutamatergic transmission in the rMR. The PVH→rMR oxytocinergic pathway connects the hypothalamic circuit for energy homeostasis to thermogenic and cardiac sympathetic outflow, and, therefore, its defects may cause obesity and impaired thermoregulation, as seen in Prader-Willi syndrome.

Hypoglycemia: Control of counter‐regulatory hormone secretion by the brain

Hypoglycaemia is a common side‐effect of the treatment of type 1 diabetes with insulin. The Diabetes Control and Complications Trial indicated that, while intensive insulin therapy reduced the incidence and severity of diabetic complications such as retinopathy, neuropathy and nephropathy, the incidence of severe hypoglycaemia was increased 2‐3 fold. In non‐diabetic individuals, hypoglycaemia triggers the glucose counter‐regulatory response. This response consists of inhibition of insulin secretion and activation of glucagon secretion. Prolonged hypoglycaemia also evokes adrenaline secretion and less importantly, growth hormone and cortisol secretion. However, in individuals that have type 1 diabetes, the glucagon response to hypoglycaemia is lost. In these patients, the pancreatic α‐cells continue to synthesise glucagon since glucagon secretagogues such as arginine produce a robust glucagon secretory response. Our research has examined the hypothalamic and medullary neurocircuits that mediate adrenaline and glucagon secretion under conditions of neuroglucoprivation or hypoglycaemia. These studies have shown that the response to neuroglucoprivation involves specialised sympathetic premotor neurons in the rostral ventrolateral medulla (RVLM) that mediate adrenaline secretion. These neurons are readily distinguishable from sympathetic vasomotorneurons also located in the RVLM. In addition, we have described the properties of perifornical hypothalamic (PeH) orexinergic (Ox) neurons that are activated by insulin‐induced hypoglycaemia (IHH) or neuroglucoprivation. PeH neurons project to many different targets including the RVLM and the dorsal motor nucleus of the vagus (DMV). Ox2‐R in the RVLM mediate the adrenaline secretory response. PeH neurons also mediate the glucagon response to IHH since the release of glucagon in response to IHH is diminished in Ataxin3 rats which have an age‐related loss of PeH orexin neurons.

Neural circuits mediating circulating interleukin-1β-evoked fever in the absence of prostaglandin E2 production

Infectious diseases and inflammatory conditions recruit the immune system to mount an appropriate acute response that includes the production of cytokines. Cytokines evoke neurally-mediated responses to fight pathogens, such as the recruitment of thermoeffectors, thereby increasing body temperature and leading to fever. Studies suggest that the cytokine interleukin-1β (IL-1β) depends upon cyclooxygenase (COX)-mediated prostaglandin E2 production for the induction of neural mechanisms to elicit fever. However, COX inhibitors do not eliminate IL-1β-induced fever, thus suggesting that COX-dependent and COX-independent mechanisms are recruited for increasing body temperature after peripheral administration of IL-1β. In the present study, we aimed to build a foundation for the neural circuit(s) controlling COX-independent, inflammatory fever by determining the involvement of brain areas that are critical for controlling the sympathetic outflow to brown adipose tissue (BAT) and the cutaneous vasculature. In anesthetized rats, pretreatment with indomethacin, a non-selective COX inhibitor, did not prevent BAT thermogenesis or cutaneous vasoconstriction (CVC) induced by intravenous IL-1β (2 µg/kg). BAT and cutaneous vasculature sympathetic premotor neurons in the rostral raphe pallidus area (rRPa) are required for IL-1β-evoked BAT thermogenesis and CVC, with or without pretreatment with indomethacin. Additionally, activation of glutamate receptors in the dorsomedial hypothalamus (DMH) is required for COX-independent, IL-1β-induced BAT thermogenesis. Therefore, our data suggests that COX-independent mechanisms elicit activation of neurons within the DMH and rRPa, which is sufficient to trigger and mount inflammatory fever. These data provide a foundation for elucidating the brain circuits responsible for COX-independent, IL-1β-elicited fevers.

Central sympathetic network for thermoregulatory responses to psychological stress

In mammals, many types of psychological stressors elicit a variety of sympathoexcitatory responses paralleling the classic fight-or-flight response to a threat to survival, including increased body temperature via brown adipose tissue thermogenesis and cutaneous vasoconstriction, and increased skeletal muscle blood flow via tachycardia and visceral vasoconstriction. Although these responses are usually supportive for stress coping, aberrant sympathetic responses to stress can lead to clinical issues in psychosomatic medicine. Sympathetic stress responses are mediated mostly by sympathetic premotor drives from the rostral medullary raphe region (rMR) and partly by those from the rostral ventrolateral medulla (RVLM). Hypothalamomedullary descending pathways from the dorsomedial hypothalamus (DMH) to the rMR and RVLM mediate important, stress-driven sympathoexcitatory transmission to the premotor neurons to drive the thermal and cardiovascular responses. The DMH also likely sends an excitatory input to the paraventricular hypothalamic nucleus to stimulate stress hormone release. Neurons in the DMH receive a stress-related excitation from the dorsal peduncular cortex and dorsal tenia tecta (DP/DTT) in the ventromedial prefrontal cortex. By connecting the corticolimbic emotion circuit to the central sympathetic and somatic motor systems, the DP/DTT → DMH pathway plays as the primary mediator of the psychosomatic signaling that drives a variety of sympathetic and behavioral stress responses. These brain regions together with other stress-related regions constitute a central neural network for physiological stress responses. This network model is relevant to understanding the central mechanisms by which stress and emotions affect autonomic regulations of homeostasis and to developing new therapeutic strategies for various stress-related disorders.

Dopaminergic input from the posterior hypothalamus to the raphe pallidus area inhibits brown adipose tissue thermogenesis.

Systemic administration of dopamine (DA) receptor agonists leads to falls in body temperature. However, the central thermoregulatory pathways modulated by DA have not been fully elucidated. Here we identified a source and site of action contributing to DA's hypothermic action by inhibition of brown adipose tissue (BAT) thermogenesis. Nanoinjection of the type 2 and type 3 DA receptor (D2R/D3R) agonist, 7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT), in the rostral raphe pallidus area (rRPa) inhibits the sympathetic activation of BAT evoked by cold exposure or by direct activation of N-methyl-d-aspartate (NMDA) receptors in the rRPa. Blockade of D2R/D3R in the rRPa with nanoinjection of SB-277011A increases BAT thermogenesis, consistent with a tonic release of DA in the rRPa contributing to inhibition of BAT thermogenesis. Accordingly, D2Rs are expressed in cold-activated and serotonergic neurons in the rRPa, and anatomical tracing studies revealed that neurons in the posterior hypothalamus (PH) are a source of dopaminergic input to the rRPa. Disinhibitory activation of PH neurons with nanoinjection of gabazine inhibits BAT thermogenesis, which is reduced by pretreatment of the rRPa with SB-277011A. In conclusion, the rRPa, the site of sympathetic premotor neurons for BAT, receives a tonically active, dopaminergic input from the PH that suppresses BAT thermogenesis.

Melanin-concentrating hormone neurons affect adipose tissues and modulate weight gain.

Melanin-concentrating hormone neurons affect adipose tissues and modulate weight gain.

Melanin-concentrating hormone-producing neurons in the hypothalamus regulate brown adipose tissue and thus contribute to energy expenditure.

Melanin-concentrating hormone (MCH) neuron-ablated mice exhibit increased energy expenditure and reduced fat weight. Increased brown adipose tissue (BAT) activity and locomotor activity-independent energy expenditure contributed to body weight reduction in MCH neuron-ablated mice. MCH neurons send inhibitory input to the medullary raphe nucleus to modulate BAT activity. Hypothalamic melanin-concentrating hormone (MCH) peptide robustly affects energy homeostasis. However, it is unclear whether and how MCH-producing neurons, which contain and release a variety of neuropeptides/transmitters, regulate energy expenditure in the central nervous system and peripheral tissues. We thus examined the regulation of energy expenditure by MCH neurons, focusing on interscapular brown adipose tissue (BAT) activity. MCH neuron-ablated mice exhibited reduced body weight, increased oxygen consumption, and increased BAT activity, which improved locomotor activity-independent energy expenditure. Trans-neuronal retrograde tracing with the recombinant pseudorabies virus revealed that MCH neurons innervate BAT via the sympathetic premotor region in the medullary raphe nucleus (MRN). MRN neurons were activated by MCH neuron ablation. Therefore, endogenous MCH neuron activity negatively modulates energy expenditure via BAT inhibition. MRN neurons might receive inhibitory input from MCH neurons to suppress BAT activity.

Characterization of Tyrosine Hydroxylase‐Positive Neurons Projections from the Paraventricular Nucleus of Hypothalamus to Hindbrain Autonomic Regulatory Centers

Hypothalamus is a key brain region for cardiovascular, behavioral, and metabolic regulation that contains complex type of excitatory or inhibitory neurons. Tyrosine hydroxylase (TH)-positive neurons in the hypothalamus play an important role in cardiometabolic regulation; however, the anatomical projections of TH neurons in the paraventricular nucleus hypothalamus (PVN) to key parasympathetic and sympathetic premotor nuclei in preautonomic regions of the brain and their connections to the liver remains undefined. This study aims to characterize the projections from THPVN neurons to ventral preautonomic regulating centers and their connection to liver neuronal projections. To achieve our goal, Cre recombinase-dependent TIT2L-eGFP reporter mice were microinjected with adeno-associated virus encoding the Cre recombinase driven by a rat TH promoter (AAV2-TH-Cre, 6.8 x 108 vg/site) bilaterally into the PVN using a stereotaxic apparatus. A retrograde pseudorabies virus with red fluorescence reporter (PRV-614-RFP) was injected into the liver (3 sites, 3ul/site) to identify liver-related neurons in the brainstem preautonotmic centers. One month after injection, mice were transcardially perfused and brain tissues were processed for confocal imaging. The AAV2-TH-Cre induced a robust expression of GFP mostly in the PVN neuronal cell bodies but not in other brain regions. In the brainstem preautonomic centers, we found dense projections in the nucleus tractus solitarius (NTS) and some projections to the dorsal motor nucleus of the vagus (DMV) and rostral ventrolateral medulla (RVLM). Interestingly, there was also dense projects into the median eminence (ME) that regulates neural hormones release. Importantly, the PRV-RFP retrograded from liver was found throughout the DMV and ventrolateral NTS . Particularly in the NTS, there was dense synaptic contact of the THPVN neuronal projections with the liver related neurons (PRV-RFP+ cell). In addition, we also observed that some PRV-RFP+ cells colocalized with TH-positive neurons in the PVN. In sum, the THPVN neurons project anatomically to the autonomic and neurosecretory regions of brain and may modulate the cardiometabolic activity and liver metabolism through these projections.

Relevance of differential control of sympathorespiratory response magnitudes in clinical assessments.

Relevance of differential control of sympathorespiratory response magnitudes in clinical assessments.

Ventromedial medullary pathway mediating cardiac responses evoked from periaqueductal gray.

Periaqueductal gray (PAG) is a midbrain region that projects to areas controlling behavioral and autonomic outputs and is involved in the behavioral and physiological components of defense reactions. Since Raphe Pallidus (RPa) is a medial medullary region comprising sympathetic premotor neurons governing heart function, it is worth considering the PAG-RPa path. We assessed: i) whether PAG projects to RPa; ii) the amplitude of cardiac responses evoked from PAG; iii) whether cardiovascular responses evoked from PAG rely on RPa. Experiments conducted in Wistar rats (±300g) were approved by Ethics Committee CEUA-UFG (092/18). Firstly, (n=3), monosynaptic retrograde tracer Retrobeads was injected into RPa; PAG slices were analyzed. Other two groups (n=6 each) were anesthetized with urethane (1.4g/kg) and chloralose (120mg/kg) and underwent craniotomy, tracheostomy, catheterization of femoral artery and vein and of cardiac left ventricle. In one group, we injected the GABAA receptor antagonist, bicuculline methiodide (BMI - 40pmol/100nL) into lateral/dorsolateral PAG. Another group was injected (100nL) with the GABAA receptor agonist muscimol (20mM) into RPa, 20min before BMI into PAG. The results were: i) retrogradely labelled neurons were found in PAG; ii) PAG activation by BMI caused positive chronotropism and inotropism, which were accompanied by afterload increases; iii) RPa inhibition with Muscimol reduced heart rate, arterial and ventricular pressures; iv) the subsequent PAG activation still increased arterial pressure, cardiac chronotropy and inotropy, but these responses were significantly attenuated. In conclusion, PAG activation increases cardiac chronotropy and inotropy, and these responses seem to rely on a direct pathway reaching ventromedial medullary RPa neurons.

Microbiota modulate sympathetic neurons via a gut–brain circuit

Gut-brain connections monitor the intestinal tissue and its microbial and dietary content1, regulating both intestinal physiological functions such as nutrient absorption and motility2,3, and brain–wired feeding behaviour2. It is therefore plausible that circuits exist to detect gut microbes and relay this information to central nervous system (CNS) areas that, in turn, regulate gut physiology4. We characterized the influence of the microbiota on enteric–associated neurons (EAN) by combining gnotobiotic mouse models with transcriptomics, circuit–tracing methods, and functional manipulations. We found that the gut microbiome modulates gut–extrinsic sympathetic neurons; while microbiota depletion led to increased cFos expression, colonization of germ-free mice with short-chain fatty acid–producing bacteria suppressed cFos expression in the gut sympathetic ganglia. Chemogenetic manipulations, translational profiling, and anterograde tracing identified a subset of distal intestine-projecting vagal neurons positioned to play an afferent role in microbiota–mediated modulation of gut sympathetic neurons. Retrograde polysynaptic neuronal tracing from the intestinal wall identified brainstem sensory nuclei activated during microbial depletion, as well as efferent sympathetic premotor glutamatergic neurons that regulate gastrointestinal transit. These results reveal microbiota–dependent control of gut extrinsic sympathetic activation through a gut-brain circuit.

Anomalous AMPK-regulated angiotensin AT1R expression and SIRT1-mediated mitochondrial biogenesis at RVLM in hypertension programming of offspring to maternal high fructose exposure

BackgroundTissue oxidative stress, sympathetic activation and nutrient sensing signals are closely related to adult hypertension of fetal origin, although their interactions in hypertension programming remain unclear. Based on a maternal high-fructose diet (HFD) model of programmed hypertension, we tested the hypothesis that dysfunction of AMP-activated protein kinase (AMPK)-regulated angiotensin type 1 receptor (AT1R) expression and sirtuin1 (SIRT1)-dependent mitochondrial biogenesis contribute to tissue oxidative stress and sympathoexcitation in programmed hypertension of young offspring.MethodsPregnant female rats were randomly assigned to receive normal diet (ND) or HFD (60% fructose) chow during pregnancy and lactation. Both ND and HFD offspring returned to ND chow after weaning, and blood pressure (BP) was monitored from age 6 to 12 weeks. At age of 8 weeks, ND and HFD offspring received oral administration of simvastatin or metformin; or brain microinfusion of losartan. BP was monitored under conscious condition by the tail-cuff method. Nutrient sensing molecules, AT1R, subunits of NADPH oxidase, mitochondrial biogenesis markers in rostral ventrolateral medulla (RVLM) were measured by Western blot analyses. RVLM oxidative stress was measured by fluorescent probe dihydroethidium and lipid peroxidation by malondialdehyde assay. Mitochondrial DNA copy number was determined by quantitative real-time polymerase chain reaction.ResultsIncreased systolic BP, plasma norepinephrine level and sympathetic vasomotor activity were exhibited by young HFD offspring. Reactive oxygen species (ROS) level was also elevated in RVLM where sympathetic premotor neurons reside, alongside augmented protein expressions of AT1R and pg91phox subunit of NADPH oxidase, decrease in superoxide dismutase 2; and suppression of transcription factors for mitochondrial biogenesis, peroxisome proliferator-activated receptor γ co-activator α (PGC-1α) and mitochondrial transcription factor A (TFAM). Maternal HFD also attenuated AMPK phosphorylation and protein expression of SIRT1 in RVLM of young offspring. Oral administration of a HMG-CoA reductase inhibitor, simvastatin, or an AMPK activator, metformin, to young HFD offspring reversed maternal HFD-programmed increase in AT1R and decreases in SIRT1, PGC-1α and TFAM; alleviated ROS production in RVLM, and attenuated sympathoexcitation and hypertension.ConclusionDysfunction of AMPK-regulated AT1R expression and SIRT1-mediated mitochondrial biogenesis may contribute to tissue oxidative stress in RVLM, which in turn primes increases of sympathetic vasomotor activity and BP in young offspring programmed by excessive maternal fructose consumption.

Anomalous AMPK‐Regulated Angiotensin AT1R Expression and SIRT1‐Mediated Mitochondrial Biogenesis at RVLM in Hypertension Programming of Offspring to Maternal High Fructose Exposure

Tissue oxidative stress, sympathetic activation and nutrient sensing signals are closely related to adult hypertension of fetal origin, although their interactions in hypertension programming remain unclear. Based on a maternal high‐fructose diet (HFD) model of programmed hypertension, we tested the hypothesis that dysfunction of AMP‐activated protein kinase (AMPK)‐regulated angiotensin type 1 receptor (AT1R) expression and sirtuin1 (SIRT1)‐dependent mitochondrial biogenesis contribute to tissue oxidative stress and sympathoexcitation in programmed hypertension of young offspring. Increased systolic and diastolic blood pressure, plasma norepinephrine level and sympathetic vasomotor activity were exhibited by young HFD offspring. Reactive oxygen species (ROS) level was also elevated in rostral ventrolateral medulla (RVLM) where sympathetic premotor neurons reside, alongside augmented protein expressions of AT1R and pg91phox subunit of NADPH oxidase, decrease in superoxide dismutase 2; and suppression of transcription factors for mitochondrial biogenesis, peroxisome proliferator‐activated receptor g co‐activator a (PGC‐1a) and mitochondrial transcription factor A (TFAM). Maternal HFD also attenuated AMPK phosphorylation and protein expression of SIRT1 in RVLM of young offspring. Oral administration of a HMG‐CoA reductase inhibitor, simvastatin, or an AMPK activator, metformin, to young HFD offspring reversed maternal HFD‐programmed increase in AT1R and decreases in SIRT1, PGC‐1a and TFAM; alleviated ROS production in RVLM, and attenuated sympathoexcitation and hypertension. Together these results suggest that dysfunction of AMPK‐regulated AT1R expression and SIRT1‐mediated mitochondrial biogenesis may contribute to tissue oxidative stress in RVLM, which in turn primes increases of sympathetic vasomotor activity and blood pressure in young offspring programmed by excessive maternal fructose consumption.Support or Funding InformationThis work was supported by research grants from the Chang Gung Medical Foundation to J.Y.H. Chan (CMRPG8C0053‐33 and OMRPG8G0011). Y.M. Chao was supported by a Postdoctoral Fellowship (MOST107‐2811‐B‐182A‐501) from the Ministry of Science and Technology, Taiwan.

Neurons in the rat superior colliculus translate visual threat into autonomic responses via a relay in the rostral ventromedial medulla

The superior colliculus (SC) is a sensory integration hub in the dorsal brainstem where multimodal information is combined and, depending on the saliency of the competing sensory inputs, appropriate motor commands and supportive autonomic changes initiated. In rodents, the SC is indispensable for initiating behavioral responses to stereotypical visual stimuli that resemble approaching objects, such as looming (an expanding overhead black circle). Here we report that presentation of overhead looming or naturalistic stimuli drove acute surges in blood pressure in telemetered conscious rats, an effect that was replicated by optogenetic stimulation of the deep SC (dSC). dSC stimulation also evoked increases in respiratory rate and tail vasoconstriction in the absence of detectable anxiety‐like behaviors and continued to exert excitatory effects on heart rate, respiratory rate, and sympathetic nerve activity under urethane anesthesia. The objective of the current study was to identify the central pathways responsible for mediating these physiological effects.Anterograde labeling of dSC neurons revealed a previously uncharacterized axonal projections to brainstem cell groups associated with arousal and autonomic control, including the locus coeruleus, A5 group and, most extensively, neurons within a region that spanned the medullary gigantocellular and raphe cell groups, collectively called the rostral ventromedial medulla (RVMM). Optogenetic stimulation of dSC terminals within the RVMM recapitulated some of the sympathetic and respiratory effects evoked by dSC stimulation, and optogenetic dSC activation evoked powerful excitatory effects on extracellular recordings of putative RVMM sympathetic premotor neurons, suggesting that elements of the physiological response dSC stimulation are mediated by direct activation of medullary autonomic neurons.To investigate the contribution of environmental stimuli to the excitability of this pathway we conducted single‐unit recordings of SC neuronal responses to visual and acoustic stimuli using high‐density silicon probes. In addition to responding to stereotypical audio‐visual looming stimuli, we report the presence of SC neurons with higher‐order visual capabilities relating to object detection that differ by subregion and are several orders of magnitude more complex than previously recognized. These tuning properties were also found in subpopulations of opto‐tagged SC neurons that project to the RVMM. Our data suggest that the SC is not only capable of nuanced object recognition, but can translate naturalistic visual cues into fast‐acting autonomic changes via direct medullary projectionsSupport or Funding InformationResearch was supported by the NHMRC and Hillcrest Foundation

Melatonin attenuates renal sympathetic overactivity and reactive oxygen species in the brain in neurogenic hypertension.

Sympathetic overactivation contributes to the pathogenesis of both experimental and human hypertension. We have previously reported that oxidative stress in sympathetic premotor neurons leads to arterial baroreflex dysfunction and increased sympathetic drive to the kidneys in an experimental model of neurogenic hypertension. In this study, we hypothesized that melatonin, a potent antioxidant, may be protective in the brainstem regions involved in the tonic and reflex control of blood pressure (BP) in renovascular hypertensive rats. Neurogenic hypertension was induced by placing a silver clip (gap of 0.2 mm) around the left renal artery, and after 5 weeks of renal clip placement, the rats were treated orally with melatonin (30 mg/kg/day) by gavage for 15 days. At the end of melatonin treatment, we evaluated baseline mean arterial pressure (MAP), renal sympathetic nerve activity (rSNA), and the baroreflex control of heart rate (HR) and rSNA. Reactive oxygen species (ROS) were detected within the brainstem regions by dihydroethidium staining. Melatonin treatment effectively reduced baseline MAP and sympathoexcitation to the ischemic kidney in renovascular hypertensive rats. The baroreflex control of HR and rSNA were improved after melatonin treatment in the hypertensive group. Moreover, there was a preferential decrease in ROS within the rostral ventrolateral medulla (RVLM) and the nucleus of the solitary tract (NTS). Therefore, our study indicates that melatonin is effective in reducing renal sympathetic overactivity associated with decreased ROS in brainstem regions that regulate BP in an experimental model of neurogenic hypertension.