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Swallowing is induced by a central pattern generator in the nucleus tractus solitarius (NTS). We aimed to create a medullary slice preparation to elucidate the neural architecture of the central pattern generator of swallowing (Sw-CPG) and record its neural activities. Experiments were conducted on 2-day-old Sprague-Dawley rats (n = 46). The brainstem-spinal cord was transected at the pontomedullary and cervicothoracic junctions; the medulla was sliced transversely at thicknesses of 600, 700, or 800 μm. The rostral end of the slice was 100 μm rostral to the vagus nerve. We recorded hypoglossal nerve activity and electrically stimulated the vagus nerve or microinjected bicuculline methiodide (BIC) into the NTS. The 800-μm slices generated both rhythmic respiratory activity and electrically elicited neural activity. The 700-μm slices generated only respiratory activity, while the 600-μm slices did not generate any neural activity. BIC microinjection into the NTS in 800-μm slices resulted in the typical activity that closely resembled the swallowing activity reported in other experiments. This swallowing-like activity consistently lengthened the respiratory interval. Despite complete inhibition of respiratory activity, weak swallowing-like activity was observed under bath application of a non-NMDA receptor antagonist. Contrastingly, bath application of NMDA receptor antagonists resulted in a complete loss of swallowing-like activity and no change in respiratory activity. These results suggest that the 800-μm medullary slice preparation contains both afferent and efferent neural circuits and pattern generators of swallowing activity. Additionally, NMDA receptors may be necessary for generating swallowing activity. This medullary slice preparation can therefore elucidate Sw-CPG neural networks., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

The central leptin signaling system has been found to facilitate breathing and is linked to obesity-related hypoventilation. Activation of leptin signaling in the nucleus tractus solitarii (NTS) and retrotrapezoid nucleus (RTN) enhances respiratory drive. In this study, we investigated how medullary leptin signaling contributes to hypoventilation and whether respective deletion of SOCS3 in the NTS and RTN could mitigate hypoventilation in diet-induced obesity (DIO) male mice. Our findings revealed a decrease in the number of CO 2 -activated NTS neurons and downregulation of acid-sensing ion channels in DIO mice compared to lean control mice. Moreover, NTS leptin signaling was disrupted, as evidenced by the downregulation of phosphorylated STAT3 and the upregulation of SOCS3 in DIO mice. Importantly, deleting SOCS3 in the NTS and RTN significantly improved the diminished hypercapnic ventilatory response in DIO mice. In conclusion, our study suggests that disrupted medullary leptin signaling contributes to obesity-related hypoventilation, and inhibiting the upregulated SOCS3 in the NTS and RTN can alleviate this condition., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Nesfatin-1, an 82-amino acid peptide encoded by the secreted precursor nucleobinin-2 (NUCB2), exerts potent anorexigenic action independently of leptin signaling. This propensity has propelled this peptide and its analogues as potential anti-obesity drug candidates. However, a more extensive comprehension of its biological actions is needed prior to envisaging its potential use in the treatment of metabolic diseases. Swallowing is an essential motor component of ingestive behavior, which induces the propulsion of the alimentary bolus from the mouth to the esophagus. The dorsal swallowing group (DSG) which constitutes a part of the central pattern generator of swallowing (SwCPG) is located within the solitary tract nucleus (STN), a region reported to contain nesfatin-1/NUCB2 expressing neurons. In this context, we investigate here the possible effects of nesfatin-1 on swallowing discharge. Nesfatin-1 dose-dependently inhibited swallowing reflex and activated neurons located in the DSG region. In addition, we provide evidences that strongly suggest that this nesfatin-1 inhibitory effect involved an oxytocinergic relay. Indeed, oxytocin (OT) injection at the brainstem level inhibited swallowing reflex and OT receptor antagonist prevented nesfatin-1 inhibitory action. Altogether, these data constitute the first demonstration that nesfatin-1 modulates swallowing reflex by acting at the brainstem level via an oxytocinergic relay., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Our previous studies have shown that electroacupuncture (EA) at the Jianshi-Neiguan acupoints (P5-6, overlying the median nerve) attenuates sympathoexcitatory responses through its influence on neuronal activity in the rostral ventrolateral medulla (rVLM). The nucleus tractus solitarii (NTS) receives input from somatic nerve stimulation. Connections between the NTS and the rVLM during EA stimulation have not been investigated and thus were the focus of the present study. Seven to ten days after unilateral microinjection of a rhodamine-conjugated microsphere retrograde tracer (100 nl) into the rVLM, rats were subjected to EA or sham-EA without electrical stimulation. EA was performed for 30 min at the P5-6 acupoints bilaterally. Perikarya containing the microsphere tracer were found in the NTS of both groups. Compared to controls (needle placement without electrical stimulation, n = 7), c-Fos immunoreactivity and neurons double-labeled with c-Fos, an immediate early gene, and the tracer were significantly increased in the NTS of EA-treated rats (all P < 0.05; n = 8), particularly, in the medial and lateral subdivisions of NTS at subpostremal and obex levels. These results suggest that EA at the P5-6 acupoints activates NTS neurons. Furthermore, EA-activated NTS neurons directly project to the rVLM and likely influence the rVLM activity., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Aims: Previous studies have shown that brain-derived thyroid-stimulating hormone (TSH) and its receptor (TSHr) are present in hypothalamic extracts. No studies investigating both the anatomical location and functional significance of putative TSHr proteins in specific central nervous system (CNS) nuclei involved in feeding controls have yet been conducted. The aim was thus to determine whether TSHr are present in nuclei associated with feeding behavior, and if such receptors may be functional., Methods: Brain tissue from adult rats was analyzed for gene expression and receptor protein expression was investigated with immunohistochemistry and western blotting. To investigate whether putative TSHr may be functional, we evaluated food intake of rats given intraparenchymal nanoinjections of TSH into the nucleus of the solitary tract (NTS)., Results: RT-qPCR confirmed previous reports that TSHr mRNA is expressed in CNS tissues of the adult rat. Immunohistochemistry showed TSHr-immunoreactivity in the arcuate, the ventromedial, the dorsomedial, and the paraventricular hypothalamic nuclei. We also found TSHr-ir in the dorsal hindbrain to be localized to the area postrema, NTS, dorsal motor nucleus of the vagus, and the hypoglossal motor nucleus. Further protein analysis with western blotting showed 120kDa TSHr-ir proteins present in the hypothalamus and brainstem. Injections of TSH into the NTS reduced food intake similar to the positive control, urocortin., Conclusions: These data suggest that functional TSHr are present in the caudal brainstem and hypothalamic nuclei of relevance for feeding control as a possibly uncleaved holoreceptor, and highlights a hindbrain component to central TSH inhibition of food intake., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Serotonin (5-HT) is a potent neuromodulator with multiple receptor types within the cardiorespiratory system, including the nucleus tractus solitarii (nTS)--the central termination site of visceral afferent fibers. The 5-HT7 receptor facilitates cardiorespiratory reflexes through its action in the brainstem and likely in the nTS. However, the mechanism and site of action for these effects is not clear. In this study, we examined the expression and function of 5-HT7 receptors in the nTS of Sprague-Dawley rats. 5-HT7 receptor mRNA and protein were identified across the rostrocaudal extent of the nTS. To determine 5-HT7 receptor function, we examined nTS synaptic properties following 5-HT7 receptor activation in monosynaptic nTS neurons in the in vitro brainstem slice preparation. Application of 5-HT7 receptor agonists altered tractus solitarii evoked and spontaneous excitatory postsynaptic currents which were attenuated with a selective 5-HT7 receptor antagonist. 5-HT7 receptor-mediated changes in excitatory postsynaptic currents were also altered by block of 5-HT1A and GABAA receptors. Interestingly, 5-HT7 receptor activation also reduced the amplitude but not frequency of GABAA-mediated inhibitory currents. Together these results indicate a complex role for 5-HT7 receptors in the nTS that mediate its diverse effects on cardiorespiratory parameters., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Previous behavioral studies have demonstrated that presynaptic N-methyl-d-aspartate (NMDA) receptors expressed on vagal afferent terminals are involved in food intake and satiety. Therefore, using in vitro live cell calcium imaging of prelabeled rat hindbrain slices, we characterized which NMDA receptor GluN2 subunits may regulate vagal afferent activity. The nonselective NMDA receptor antagonist d,l-2-amino-5-phosphonopentanoic acid (d,l-AP5) significantly inhibited vagal terminal calcium influx, while the excitatory amino acid reuptake inhibitor d,l-threo-β-benzyloxyaspartic acid (TBOA), significantly increased terminal calcium levels following pharmacological stimulation with ATP. Subunit-specific NMDA receptor antagonists and potentiators were used to identify which GluN2 subunits mediate the NMDA receptor response on the vagal afferent terminals. The GluN2B-selective antagonist, ifenprodil, selectively reduced vagal calcium influx with stimulation compared to the time control. The GluN2A-selective antagonist, 3-chloro-4-fluoro-N-[4-[[2-(phenylcarbonyl)hydrazino]carbonyl] benzyl]benzenesulfonamide (TCN 201) produced smaller but not statistically significant effects. Furthermore, the GluN2A/B-selective potentiator (pregnenolone sulfate) and the GluN2C/D-selective potentiator [(3-chlorophenyl)(6,7-dimethoxy-1-((4-methoxyphenoxy)methyl)-3,4-dihydroisoquinolin-2(1H)-yl)methanone; (CIQ)] enhanced vagal afferent calcium influx during stimulation. These data suggest that presynaptic NMDA receptors with GluN2B, GluN2C, and GluN2D subunits may predominantly control vagal afferent excitability in the nucleus of the solitary tract., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Activity of second-order relay neurons in the nucleus tractus solitarius (NTS) is regulated by peripheral and intrinsic synaptic inputs, and modulation of those inputs by metabotropic glutamate receptors (mGluRs) has been proposed. This study investigated effects of mGluR activation on glutamatergic transmission in the NTS second-order neurons of guinea pigs. Whole-cell patch-clamp recordings from the brainstem slices revealed that activation of mGluRs exerted its effects on the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) but not on the amplitude. The sEPSC frequency was increased by an agonist of group I mGluRs, and it was decreased by an mGluR1 antagonist but not by an mGluR5 antagonist. The agonists of group II and III mGluRs decreased the sEPSC frequency, while their antagonists alone had no effect. Perfusion of cystine or TBOA, either of which elevates extracellular glutamate concentration, resulted in an increase in the sEPSC frequency, leaving the amplitude unchanged. The increased frequency of sEPSCs was returned to control by an mGluR1 antagonist. The tractus solitarius-evoked EPSCs were not altered by an agonist of group I mGluRs, whereas they were decreased along with an increase in paired-pulse ratio by agonists of group II and III mGluRs. These results suggest that mGluRs are present at the presynaptic sites in the NTS second-order neurons in guinea pigs. The mGluR1s function to facilitate the release of glutamate from axon terminals of intrinsic interneurons and the group II and III mGluRs play an inhibitory role in glutamatergic transmission., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Serotonin (5-Hydroxytryptamine, 5-HT) and the 5-HT2 receptor modulate cardiovascular and autonomic function in part through actions in the nTS, the primary termination and integration point for cardiorespiratory afferents in the brainstem. In other brain regions, 5-HT2 receptors (5-HT2R) modify synaptic transmission directly, as well as through 5-HT2AR-induced endocannabinoid release. This study examined the role of 5-HT2AR as well as their interaction with endocannabinoids on neurotransmission in the nucleus tractus solitarii (nTS). Excitatory postsynaptic currents (EPSCs) in monosynaptic nTS neurons were recorded in the horizontal brainstem slice during activation and blockade of 5-HT2ARs. 5-HT2AR activation augmented solitary tract (TS) evoked EPSC amplitude whereas 5-HT2AR blockade depressed TS-EPSC amplitude at low and high TS stimulation rates. The 5-HT2AR-induced increase in neurotransmission was reduced by endocannabinoid receptor block and increased endogenous endocannabinoids in the synaptic cleft during high frequency, but not low, TS stimulation. Endocannabinoids did not tonically modify EPSCs. These data suggest 5-HT acting through the 5-HT2AR is an excitatory neuromodulator in the nTS and its effects are modulated by the endocannabinoid system., (© 2013 Published by Elsevier B.V.)

The embryonic development of synapses in the rostral nucleus of the solitary tract (rNST) was investigated in rat to determine when synapses begin to function. Using a brain slice preparation we studied appearance of synaptic receptors on second order rNST neurons and investigated the development of postsynaptic responses elicited by afferent nerve stimulation. Prenatal excitatory and inhibitory synaptic responses were recorded as early as E14. Glutamatergic and GABAergic postsynaptic responses were detected as early as E16. Both NMDA and AMPA receptors contributed to glutamatergic postsynaptic responses. GABAergic postsynaptic responses resulted primarily from activation of GABA(A) receptors. However, functional GABA(C) receptors were also demonstrated. A glycinergic postsynaptic response was not found although functional glycine receptors were demonstrated at E16. Solitary tract (ST) stimulation-evoked EPSCs, first detected at E16, were eliminated by glutamate receptor antagonists. ST-evoked IPSPs, also detected at E16, were eliminated by GABA(A) receptor antagonist. Thus, considerable prenatal development of rNST synaptic connections occurs and this will ensure postnatal function of central taste processing circuits., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Propofol, an intravenous anesthetic, is broadly used for general anesthesia and diagnostic sedations due to its fast onset and recovery. Propofol depresses respiratory and cardiovascular reflex responses, however, their underlying mechanisms are not well known. Cardiorespiratory information from visceral afferent vagus nerves is integrated in the nucleus tractus solitarii (NTS). Cardiac and respiratory signals transducing vagal afferent neurons release the excitatory neurotransmitter glutamate onto NTS neurons in an activity dependent manner and trigger negative feedback reflex responses. In this experiment, the effects of propofol on glutamatergic synaptic responses at NTS neurons was tested using patch clamp methods. Glutamatergic excitatory postsynaptic currents (EPSC) were recorded at chloride reversal potential (-49mV) without γ-aminobutyric acid type A (GABA(A)) receptor antagonists. Propofol (≥3μM) facilitated frequency of the spontaneous EPSCs in a concentration dependent manner without altering amplitude and decay time. The GABA(A) receptor selective antagonist, gabazine (6μM), attenuated propofol effects on glutamate release. Propofol (10μM) evoked glutamate release was also blocked in the presence of the voltage dependent Na(+) and Ca(2+) channel blockers TTX (0.3μM) and Cd(2+) (0.2mM), respectively. In addition, the Na(+)-K(+)-Cl(-) cotransporter type 1 antagonist bumetanide (10μM) also inhibited propofol evoked increase in sEPSC frequency. These results suggest that propofol evoked glutamate release onto NTS neurons by GABA(A) receptor-mediated depolarization of the presynaptic excitatory terminals., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Fifty-eight male Sprague-Dawley rats used in the present study to investigate the role of baroreceptor sensitive neurons of the nucleus tractus solitarius (NTS) in the regulation of cardiovascular inhibition during acupuncture at the auricular point Heart, single unit recording was made in anesthetized Sprague-Dawley rats. A neuron was considered to be excited or inhibited by acupuncture stimulation if it displayed 15% more or less spikes s(-1), respectively. NTS neurons were classified into cardiac-related (CR) neurons and non-cardiac-related neurons based on whether their rhythmic discharges were synchronized with the R-waves and responding to sodium nitroprusside (NP; 20 μg/kg, i.v.) administration. Manual acupuncture was applied at the auricular point Heart and somatic acupuncture points ST36 and PC6. Acupuncture at auricular point Heart showed a more significant inhibitory effect on arterial pressure (-22.1±2.4mm Hg; P<0.001) and heart rate (-12.7±1.7 bpm; P<0.001) than that at ST36 and PC6. Acupuncture at auricular point Heart also increased the level of response of CR neurons in the NTS (93.8%±26.0% increase in discharge rate; P<0.01). Systemic or local administration of atropine attenuated the cardiovascular inhibition and activation of CR neurons evoked by auricular acupuncture, but had no effect on the same responses evoked by somatic acupuncture. Inactivation of the NTS with local anesthetics also decreased the cardiovascular inhibitory responses evoked by auricular acupuncture. Our results show that acupuncture at the auricular point Heart regulates cardiovascular function by activating baroreceptor sensitive neurons in the NTS in a similar manner as the baroreceptor reflex in cardiovascular inhibition., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Metabolic homeostasis reflects the complex output of endocrine, autonomic, and behavioral control circuits that extend throughout the central nervous system. Brain regions that control food intake and energy expenditure are privy to continuous visceral sensory feedback signals that presumably modulate appetite, satiety, digestion, and metabolism. Sensory signals from the gastrointestinal tract and associated digestive viscera are delivered to the brain primarily by vagal afferents that terminate centrally within the caudal nucleus of the solitary tract (NST), with signals subsequently relayed to higher brain regions by parallel noradrenergic and peptidergic projection pathways arising within the NST. This article begins with an overview of these ascending pathways identified in adult rats using a standard anterograde tracer microinjected into the caudal visceral sensory region of the NST, and also by immunocytochemical localization of glucagon-like peptide-1. NST projection targets identified by these two approaches are compared to the distribution of neurons that become infected after inoculating the ventral stomach wall with a neurotropic virus that transneuronally infects synaptically-linked chains of neurons in the anterograde (i.e., ascending sensory) direction. Although the focus of this article is the anatomical organization of axonal projections from the caudal visceral NST to the hypothalamus and limbic forebrain, discussion is included regarding the hypothesized role of these projections in modulating behavioral arousal and coordinating endocrine and behavioral (i.e., hypophagic) responses to stress., (2010 Elsevier B.V. All rights reserved.)

While the evidences emphasizing the role of astroglial cells in numerous aspects of information processing within the brain merges, the literature dealing with the involvement of this cell population in the signalization involved in feeding behavior and energetic homeostasis remains scarce. Nevertheless, some clues are now available indicating that glia could play a dynamic role in the regulation of energy balance, and that strengthening research effort in this field may further our understanding of the mechanisms controlling feeding behaviour. In the present review, we have summarized recent data indicating that the multifaceted glial compartment of the brainstem should be considered in future research aimed at identifying feeding-related processes operating at this level., (2010 Elsevier B.V. All rights reserved.)

Chorda tympani (CT) and glossopharyngeal (IXth) nerves relay taste information from anterior and posterior tongue to brainstem where they synapse with second order neurons in the rostral nucleus of solitary tract (rNST). rNST neurons monosynaptically connected to afferent gustatory input were identified both by anatomical labeling and synaptic latency measures. Anterograde tracing was used to label the CT and IXth terminal fields, and neurons surrounded by fluorescent neural profiles visualized with differential interference contrast (DIC) optics in horizontal brainstem slices. Anatomically identified neurons were patch-clamped and excitatory postsynaptic currents (EPSCs) evoked by electrically stimulating the solitary tract (ST) under GABA(A) receptor blockade. Monosynaptic connections were confirmed by measures of the standard deviation of synaptic latency (jitter). rNST neurons responded to ST stimulation with either all-or-none or graded amplitude EPSCs. Most (70%) of the rNST neurons with CT input and 30% with IX input responded with all-or-none EPSCs. The remainder of the neurons with CT and IX input responded with increasing EPSC amplitudes to greater intensity stimulus shocks. EPSCs evoked in rNST neurons by increasing shock frequency to both CT and IXth nerves resulted in reduced amplitude EPSCs characteristic of frequency-dependent synaptic depression. Our results suggest that the second order rNST neurons respond to afferent input with different patterns of EPSCs that potentially influence transmission of gustatory information. Frequency-dependent synaptic depression would act as a low pass filter important in the initial processing of gustatory derived sensory messages., (Copyright 2010 Elsevier B.V. All rights reserved.)

The present study examined whether vagal visceral inputs are involved in parasympathetic reflex vasodilatation in the masseter muscle in urethane-anesthetized and cervically sympathectomized rats. Electrical stimulation of the central cut end of the cervical vagus nerve (VN) including visceral afferent fibers, which consisted of cervical/thoracic branches (heart and lungs) and abdominal branches (entire gastrointestinal tract), elicited intensity- and frequency-dependent increases of blood flow in the masseter muscle (MBF). Activation of the abdominal VN inferior to the diaphragm failed to affect the MBF. MBF increases evoked by cervical VN stimulation were reduced significantly by hexamethonium. Pretreatment with atropine reduced the MBF increase evoked by VN stimulation significantly, whereas pretreatment with either propranolol or phentolamine had no effect on the response. MBF increases occurred with electrical stimulation of nucleus of the solitary tract (NTS), and these increases were significantly reduced by the administration of hexamethonium and atropine. MBF increases also occurred after microinjection of glutamate into the NTS in a dose-dependent manner. Microinjection of muscimol into the NTS caused a significant attenuation of the VN stimulation-induced MBF increases. Our results suggest that vagal visceral inputs passing to the NTS are involved in the parasympathetic reflex vasodilatation in the rat masseter muscle. The MBF increase evoked by the vagal-parasympathetic reflex mechanism occurred via visceral afferents running in the cervical VN, but not in the abdominal VN, suggesting that the vagal visceral afferents derived from cardiovascular and/or respiratory systems may play an important role in the regulation of the MBF., ((c) 2009 Elsevier B.V. All rights reserved.)

The HSD2 (11-beta-hydroxysteroid dehydrogenase-type 2 enzyme) containing neurons of the nucleus tractus solitarius (NTS) become activated during low-sodium and high-aldosterone states such as hypovolemia. This response may be due to hormonal and/or neural signals. Hormonal signals may activate neurons in the area postrema that innervate the HSD2 neurons. The vagus nerve projects directly to the HSD2 neurons and this could be another route whereby these neurons receive information about systemic sodium/aldosterone status. The peripheral sites of origin that contribute to this vagal projection remain unknown, and in the present study, we injected the transganglionic tracer, cholera toxin beta-subunit-horseradish peroxidase (CTb-HRP), into wall of various gastrointestinal organs (stomach, small and large intestine) or liver of rats. Confocal microscopy of brainstem sections stained by a double immunohistochemical procedure was used to analyze whether the HSD2 neurons received axonal contacts from specific gastrointestinal structures. The major source of afferents arose from the stomach, mainly from its pyloric antrum, but a weaker input originated from the fundus region. A trace amount originated from the duodenum. The terminal part of the small intestine and large intestine did not to contribute to this projection. Similarly, no afferent inputs from the liver or portal vein were found. In conclusion, HSD2 neurons receive an input mainly from the stomach and these results are considered as potential sites affecting sodium intake.

Caudal brainstem viscerosensory nuclei convey information about the body's internal state to forebrain regions implicated in feeding behavior and responses to immune challenge, and may modulate ingestive behavior following immune activation. Illness-induced appetite loss might be attributed to accentuated "satiety" pathways, activation of a distinct "danger channel" separate from satiety pathways, or both. To evaluate neural substrates that could mediate the effects of illness on ingestive behavior, we analyzed the pattern and phenotypes of medullary neurons responsive to consumption of a preferred food, sweetened milk, and to intraperitoneal lipopolysaccharide challenge that reduced sweetened milk intake. Brainstem sections were stained for c-Fos, dopamine beta-hydroxylase, phenylethanolamine-N-methyltransferase, and glucagon-like peptide-1 (GLP-1) immunoreactivity. Sweetened milk intake activated many neurons throughout the nucleus of the solitary tract (NTS), including A2 noradrenergic neurons in the caudal half of the NTS. LPS challenge activated a similar population of neurons in the NTS, in addition to rostral C2 adrenergic and mid-level A2 noradrenergic neurons in the NTS, many C1 and A1 neurons in the ventrolateral medulla, and in GLP-1 neurons in the dorsal medullary reticular nucleus. Increased numbers of activated GLP-1 neurons in the NTS were only associated with sweetened milk ingestion. Evidence for parallel processing was reflected in the parabrachial nucleus, where sweetened milk intake resulted in activation of the inner external lateral, ventrolateral and central medial portions, whereas LPS challenge induced c-Fos expression in the outer external lateral portions. Thus, signals generated in response to potentially dangerous physiological conditions seem to be propagated via specific populations of catecholaminergic neurons in the NTS and VLM, and likely include a pathway through the external lateral PBN. The data indicate that immune challenge engages multiple ascending neural pathways including both a distinct catecholaminergic "danger" pathway, and a possibly multimodal pathway derived from the NTS.

Taste and sensory information are closely associated and our electrophysiological studies showed a trigeminal modulation of gustatory neurons in the nucleus of the solitary tract (NST). Chorda tympani (CT) and lingual nerves (LN) converge centrally in the rostral subdivision of the NST in hamsters and rats. However, no study has yet revealed the details of this overlap on a same section. We therefore used a double-label strategy to visualize neurons in the NST that receive both trigeminal and gustatory inputs. An anterograde tracer (BDA, Biotinylated Dextran Amine) was applied unilaterally to the cut central end of the LN in male Sprague-Dawley rats. One week later, the ipsilateral CT was electrically stimulated, after which animals were perfused and brainstem sections double-labelled for Fos immunoreactivity of activated NST neurons and BDA labelling of LN afferents. Our results permitted to circumscribe the regional overlap of the trigeminal and CT afferents mainly in the rostral central (RC) subdivision of the gustatory NST. Fos-immunoreactive neurons were observed to be closely apposed by BDA-labelled fibres and terminal boutons. Such varicosities mainly "en passant" were especially present in the RC zone of the nucleus. These observations provide an anatomical substrate for trigemino-gustatory interactions.

Electrical stimulation of the chorda tympani nerve (CT; innervating taste buds on the rostral tongue) is known to initiate recurrent inhibition in cells in the nucleus of the solitary tract (NTS, the first central relay in the gustatory system). Here, we explored the relationship between inhibitory circuits and the breadth of tuning of taste-responsive NTS neurons. Initially, NTS cells with evoked responses to electrical stimulation of the CT (0.1 ms pulses; 1 Hz) were tested with each of four tastants (0.1 M NaCl, 0.01 M HCl, 0.01 M quinine and 0.5 M sucrose) in separate trials. Next, the CT was electrically stimulated using a paired-pulse (10-2000 ms interpulse interval; blocks of 100 trials) paradigm. Forty-five (30 taste-responsive) of 51 cells with CT-evoked responses (36 taste-responsive) were tested with paired pulses. The majority (34; 75.6%) showed paired-pulse attenuation, defined as fewer evoked spikes in response to the second (test) pulse compared with the first (conditioning) pulse. A bimodal distribution of the peak of paired-pulse attenuation was found with modes at 10 ms and 50 ms in separate groups of cells. Cells with early peak attenuation showed short CT-evoked response latencies and large responses to relatively few taste stimuli. Conversely, cells with late peak attenuation showed long CT-evoked response latencies and small taste responses with less selectivity. Results suggest that the breadth of tuning of an NTS cell may result from the combination of the sensitivities of peripheral nerve inputs and the recurrent influences generated by the circuitry of the NTS.

Eating is modulated by estradiol in females of many species and in women. To further investigate the estrogen receptor mechanism mediating this effect, ovariectomized rats and mice were treated with estradiol benzoate or the estrogen receptor-alpha (ER-alpha)-selective agonist PPT. PPT inhibited eating in rats much more rapidly than estradiol (approximately 2-6 h versus >24 h). In contrast, the latencies to vaginal estrus after PPT and estradiol were similar (>24 h). PPT also inhibited eating within a few hours in wild-type mice, but failed to inhibit eating in transgenic mice deficient in ER-alpha (ERalphaKO mice). PPT, but not estradiol, induced the expression of c-Fos in corticotrophin-releasing hormone (CRH)-expressing cells of the paraventricular nucleus (PVN) of the hypothalamus within 90-180 min in rats. Both PPT and estradiol reduced c-Fos expression in an ER-alpha-containing area of the nucleus of the solitary tract. The anomalously rapid eating-inhibitory effect of PPT suggests that PPT's neuropharmacological effect differs from estradiol's, perhaps because PPT differentially activates membrane versus nuclear ER-alpha or because PPT activates non-ER-alpha membrane estrogen receptors in addition to ER-alpha. The failure of PPT to inhibit eating in ERalphaKO mice, however, indicates that ER-alpha is necessary for PPT's eating-inhibitory action and that any PPT-induced activation of non-ER-alpha estrogen receptors is not sufficient to inhibit eating. Finally, the rapid induction of c-Fos in CRH-expressing cells in the PVN by PPT suggests that PPT elicits a neural response that is similar to that elicited by stress or aversive emotional stimuli.

Adiponectin is an adipocyte derived hormone which acts in the CNS to control autonomic function, energy and cardiovascular homeostasis. Two 7-transmembrane domain receptors, AdipoR1 and AdipoR2, expressed in the hypothalamus and brainstem mediate the actions of adiponectin. The medulla's nucleus of the solitary tract (NTS) is the primary viscerosensory integration site and an important nucleus in the regulation of cardiovascular function. Here we show the localization of both AdipoR1 and AdipoR2 mRNA in the NTS. We have investigated the consequences of receptor activation in response to exogenous application of adiponectin on cardiovascular (blood pressure and heart rate monitoring in vivo), and single neuron (whole cell current-clamp recordings in vitro) function. Microinjection of adiponectin in the medial NTS (mNTS) at the level of the area postrema resulted in a decrease in BP (mean AUC= -2055+/-648.1, n=5, mean maximum effect: -11.7+/-3.6 mm Hg) while similar commissural NTS (cNTS) microinjections were without effect. Patch clamp recordings from NTS neurons in a medullary slice preparation showed rapid (within 200 s of application) reversible (usually within 1000 s following washout) effects of adiponectin on the membrane potential of 62% of mNTS neurons tested (38/61). In 34% (n=21) of mNTS neurons adiponectin induced a depolarization of membrane potential (6.8+/-0.9 mV), while the remainder of mNTS cells influenced by adiponectin (n=17) hyperpolarized in response to this adipokine (-5.4+/-0.7 mV). Post-hoc single cell RT-PCR (ssRT-PCR) analysis of neurons showed that the majority of NPY mRNA positive mNTS neurons were depolarized by adiponectin (7/11), while 4 of these depolarized cells were also GAD67 positive. The results presented in this study suggest adiponectin acts in the NTS to control BP and suggest that such effects may occur as a direct result of the ability of this adipokine to modulate the excitability of discrete groups of neurons in the NTS. These studies identify the mNTS as a new CNS site which adiponectin may act to influence central autonomic processing.

This study was designed to examine the mechanism of heart rate (HR) responses elicited by the stimulation of hypothalamic paraventricular nucleus (PVN). Experiments were done in urethane-anesthetized, barodenervated, adult, male Wistar rats. Chemical stimulation of the PVN by unilateral microinjections of N-methyl-d-aspartic acid (NMDA) elicited increases in HR which were attenuated by bilateral vagotomy. PVN-induced tachycardia was also attenuated by the blockade of the spinal ionotropic glutamate receptors (iGLURs) which was accomplished by intrathecal injections at T9-T10 or direct application at T1-T4 of iGLUR antagonists. The blockade of spinal iGLURs combined with bilateral vagotomy completely blocked PVN-induced tachycardia. Blockade of GABA receptors in the medial nucleus tractus solitarius (mNTS) also attenuated the PVN-induced tachycardia. Complete blockade of PVN-induced tachycardia was also observed after the blockade of iGLURs in both the spinal cord and mNTS. Combination of the blockade of mNTS GABA receptors and spinal iGLURs also abolished PVN-induced tachycardia. PVN-induced tachycardia was not altered by the blockade of spinal vasopressin or oxytocin receptors at T1-T4. These results suggested that in barodenervated rats: 1) tachycardia elicited by the chemical stimulation of the PVN was mediated via both inhibition of vagal and activation of sympathetic outflows to the heart, 2) the vagal inhibition contributing to the PVN-induced tachycardia was mediated by the iGLURs and GABARs in the mNTS, 3) sympathetic activation contributing to the PVN-induced tachycardia was mediated via spinal iGLURs, and 4) spinal vasopressin and oxytocin receptors were not involved in the mediation of PVN-induced tachycardia.

Rostral forebrain structures like the gustatory cortex (GC), bed nucleus of the stria terminalis (BNST), central nucleus of the amygdala (CeA), and lateral hypothalamus (LH) send projections to the nucleus of solitary tract (NST) and the parabrachial nucleus (PBN) that modulate taste-elicited responses. However, the proportion of forebrain-induced excitatory and inhibitory effects often differs when taste cell recording changes from the NST to the PBN. The present study investigated whether this descending influence originates from a shared or distinct population of forebrain neurons. Under electrophysiological guidance, the retrograde tracers fast blue (FB) and fluorogold (FG) or green (GFB) and red (RFB) fluorescent latex microbeads were injected iontophoretically or by pressure pulses (10 ms at 20 psi) into the taste-responsive regions of the NST and the ipsilateral PBN in six rats. Seven days later, the animals were euthanized and tissue sections containing the LH, CeA, BNST, and GC were processed for co-localization of FB and FG or GFB and RFB. The results showed that the CeA is the major source of input to the NST (82.3+/-7.6 cells/section) and the PBN (76.7+/-11.5), compared to the BNST (31.8+/-4.5; 37.0+/-4.8), the LH (35.0+/-5.4; 33.6+/-5.7), and the GC (27.5+/-4.0; 29.0+/-4.6). Of the total number of retrogradely labeled cells, the incidence of tracer co-localization was 17+/-3% in the GC, 17+/-2% in the CeA, 15+/-3% in the BNST and 16+/-1% in the LH. Thus, irrespective of forebrain source the majority of descending input to the gustatory NST and PBN originates from distinct neuronal populations. This arrangement provides an anatomical substrate for differential modulation of taste processing in the first and second central relays of the ascending gustatory system.

Food intake is controlled by peripheral signals from the gastrointestinal tract and adipocytes, which are integrated within the central nervous system. There is evidence that signals from the GI tract are modulated by long term changes in diet, possibly leading to hyperphagia and increased body weight. We tested the hypothesis that diet-induced obese-prone (DIO-P) and obese-resistant (DIO-R) mice strains differ in the long term adaptive response of the gut-brain pathway to a high fat diet. Immunochemical detection of Fos protein was used as a measure of neuronal activation in the nucleus of the solitary tract (NTS) in response to intragastric administration of lipid in DIO-P (C57Bl6) and DIO-R (129sv) mouse strains maintained on chow or high fat, high energy diets (45% or 60% kcal from fat). Intragastric lipid administration activated neurons in the NTS in both DIO-P and DIO-R mice; the number of activated neurons was significantly greater in DIO-P than in DIO-R mice (P<0.001). However, lipid-induced activation of NTS neurons in DIO-P mice was attenuated by approximately 30% after maintenance on either 45% or 60% HF diet, for 4 or 8 weeks, compared to chow fed controls (P<0.05). In contrast, in DIO-R mice, maintenance on a HF diet (45% or 60%) had no effect on lipid-induced activation of NTS neurons. These results demonstrate that DIO-P and DIO-R mice strains differ in the adaptation of the pathway to long term ingestion of high fat diets, which may contribute to decrease satiation and increased food intake.

After ipsilateral injections of biotinylated dextran amine (BDA) into the ventrolateral subnucleus of the nucleus tractus solitarius (vlNTS) and Fluoro-gold (FG) into the rostral ventral respiratory group (rVRG) region or into the phrenic nucleus (PhN) region in the rat, an overlapping distribution of BDA-labeled axon terminals and FG-labeled neurons was found in the Kölliker-Fuse (KF) nucleus ipsilateral to the injection sites. Using retrograde tracing combined with immunohistochemistry for glutamic acid decarboxylase isoform 67 (GAD67), we indicated that as many as 40% of the vlNTS neurons projecting to the KF were immunoreactive for GAD67. Using a combination of anterograde and retrograde tracing techniques, and immunohistochemistry for GAD67, we further demonstrated that the vlNTS axon terminals with GAD67 immunoreactivity established close contact to the rVRG- or PhN-projecting KF neurons. The present results suggest that GABAergic vlNTS fibers may exert inhibitory influences on the rVRG- as well as PhN-projecting KF neurons and these circuits may be involved in the respiratory reflexes such as the Hering-Breuer reflex.

Taste neurons in the nucleus of the solitary tract (NST) not only send axons to the parabrachial nuclei (PbN), but also receive descending projections from gustatory nuclei in the forebrain in rodents. The parvicellular portion of the ventroposteromedial nucleus of the thalamus (VPMpc) receives projections from the bilateral PbN and transmits taste information to the gustatory cortex. Here, we examined the influence of bilateral stimulation of the VPMpc on taste-responsive neurons in the NST. Extracellular single unit activity was recorded from the urethane-anesthetized hamster. Taste responses were confirmed by delivery of four basic tastants to the anterior tongue. After identifying a taste neuron in the NST, the VPMpc was stimulated bilaterally. Thirty seven out of 83 neurons were orthodromically activated following VPMpc stimulation: 30 were excited and seven were inhibited. Among these cells, seven were excited and one was inhibited bilaterally. In addition, four NST neurons were antidromically invaded from the ipsilateral VPMpc. The effect of VPMpc activation on taste-driven responses was tested on 8 of 30 cells that were excited, and all seven cells that were inhibited by the VPMpc stimulation. The VPMpc stimulation enhanced responses to the effective taste stimuli or suppressed the taste-evoked activities in all eight and seven cells tested, respectively, parallel to the type of the inputs which they received from the VPMpc. These results suggest that a subset of taste neurons in the NST is under the influence from the bilateral VPMpc and that the VPMpc activation modulates taste responses of these cells.

The nucleus of the solitary tract (NTS) is the site of the first synapse of cardiovascular afferent fibers in the central nervous system. Important mechanisms for cardiovascular regulation are also present in the caudal pressor area (CPA) localized at the caudal end of the ventrolateral medulla. In the present study we sought to investigate the role of the commissural subnucleus of the NTS (commNTS) on pressor and tachycardic responses induced by l-glutamate injected into the CPA. Male Holtzman rats (n=8 rats/group) anesthetized with urethane (1.2 g/kg of body weight, iv) received injections of the GABA(A) receptor agonist muscimol into the commNTS. Unilateral injection of L-glutamate (10 nmol/100 nL) into the CPA increased mean arterial pressure (MAP, 31+/-4 mm Hg, vs. saline: 3+/-2 mm Hg) and heart rate (HR, 44+/-8 bpm, vs. saline: 10 +/-7 bpm). Inhibition of commNTS neurons with muscimol (120 pmol/60 nL) abolished the increase in MAP (9+/-4 mm Hg) and HR (17+/-7 bpm) produced by l-glutamate into the CPA. The present results suggest that the pressor and tachycardic responses to CPA activation are dependent on commNTS mechanisms.

The caudal ventrolateral medulla (CVLM) provides tonic inhibitory and also excitatory inputs to the rostral ventrolateral medulla (RVLM). These experiments evaluated the role of RVLM gamma-amino butyric acid (GABA) receptor subtypes and glycine receptors in mediating CVLM sympathoinhibition. In Inactin anesthetized female rats, the CVLM and RVLM were functionally defined by pressor and depressor responses to microinjected GABA (500 pmol, 50 nl). Although reduced, pressor and sympathoexcitatory responses due to inhibition of the CVLM with GABA persisted following ipsilateral RVLM GABA(A) receptor blockade (bicuculline, BIC, 400 pmol, 100 nl; n=12) in rats with contralateral nucleus tractus solitarius (NTS) lesion. In the presence of either ipsilateral (+contralateral NTS lesion; n=8) or bilateral (n=6) GABA(A) and GABA(B) receptor blockade of the RVLM (400 pmol BIC+400 pmol CGP35348, 100 nl), inhibition of the CVLM still increased MAP and renal sympathetic nerve activity (RSNA). Thus neither GABA(B) receptors nor a contralateral CVLM to RVLM GABAergic pathway explains residual responses to CVLM blockade. The addition of strychnine (300 pmol, 100 nl) to the RVLM eliminated responses to CVLM inhibition, suggesting that a GABA(A) and GABA(B) independent sympathoinhibitory influence from CVLM to RVLM is mediated by glycine receptors. Decreases in MAP and RSNA due to activation of the CVLM with glutamate (500 pmol, 50 nl) were reversed to increases in the presence of RVLM GABA(A) receptor blockade (n=7). Thus, a sympathoexcitatory pathway from the CVLM can be activated in the presence of RVLM GABA receptor blockade, but sympathoinhibitory influences from the CVLM predominate.

The nucleus of the solitary tract (NTS) is a critical structure involved in coordinating autonomic and visceral activities. Previous independent studies have demonstrated efferent projections from the NTS to the nucleus paragigantocellularis (PGi) and the central nucleus of the amygdala (CNA) in rat brain. To further characterize the neural circuitry originating from the NTS with postsynaptic targets in the amygdala and medullary autonomic targets, distinct green or red fluorescent latex microspheres were injected into the PGi and the CNA, respectively, of the same rat. Thirty-micron thick tissue sections through the lower brainstem and forebrain were collected. Every fourth section through the NTS region was processed for immunocytochemical detection of tyrosine hydroxylase (TH), a marker of catecholaminergic neurons. Retrogradely labeled neurons from the PGi or CNA were distributed throughout the rostro-caudal segments of the NTS. However, the majority of neurons containing both retrograde tracers were distributed within the caudal third of the NTS. Cell counts revealed that approximately 27% of neurons projecting to the CNA in the NTS sent collateralized projections to the PGi while approximately 16% of neurons projecting to the PGi sent collateralized projections to the CNA. Interestingly, more than half of the PGi and CNA-projecting neurons in the NTS expressed TH immunoreactivity. These data indicate that catecholaminergic neurons in the NTS are poised to simultaneously coordinate activities in limbic and medullary autonomic brain regions.

Changes in heart rate (HR), thoracic sympathetic nerve activity (tSNA) and frequency of phrenic nerve discharge (PND) in response to microinjection of L-glutamate before and after local microinjection of ionotropic or metabotropic glutamate receptors antagonists into the commissural subnucleus of the NTS (comNTS) were investigated. The experiments were performed in an in situ unanesthetized decerebrated working heart-brainstem preparation (WHBP), and the main findings were as follows: (a) microinjection of increasing concentrations of L-glutamate (5, 25, 50, 250 and 500 mM) into the comNTS produced bradycardia, increase in tSNA and reduction in the frequency of the PND in a concentration-dependent manner; (b) both bradycardia and increase in tSNA were almost abolished by kynurenic acid (KYN, 250 mM, a nonselective ionotropic glutamate receptor antagonist); (c) the reduction in the frequency of the PND was reversed to an increase in the frequency of the PND after KYN and this increase was blocked by the sequential microinjection of MCPG (100 mM, a nonselective metabotropic glutamate receptor antagonist); and (d) microinjection of increasing concentrations of trans-ACPD (0.5, 1.0, 2.5, 5.0 and 10 mM, a metabotropic glutamate receptor agonist), elicited bradycardia and increase in the frequency of the PND in a concentration-dependent manner, which were blocked by MCPG. Taken together, these data indicate that l-glutamate and its ionotropic receptors are involved in the sympathoexcitatory, bradycardic and reduction in the frequency of the PND responses whereas/although its metabotropic receptors are involved in the bradycardic and mainly in the increase in the frequency of the PND to microinjection of L-glutamate into the comNTS in the WHBP.

Neurons in the caudal nucleus of the tractus solitarius (cNTS) are quite heterogeneous in cell size (50 to 450 microm(2) in somal area) and other morphologic characteristics. For a more objective classification of cNTS neurons, their morphologic features were analyzed quantitatively based on reconstructed biocytin-filled cells after whole-cell patch-clamp recordings. According to the patterns of axonal branching behaviors, cNTS cells could be classified into two groups: smaller cells (94.1 microm(2) in mean somal area, range 62-120 microm(2), n = 22) and larger cells (245 microm(2) in mean somal area, range 142-411 microm(2), n = 23). Extensive axonal arborization with numerous possible synaptic boutons was specifically associated with smaller neurons, while larger cells possessed no or few axon collaterals, suggesting their distinct roles as local circuit neurons (or interneurons) and projection neurons, respectively. With regard to somatodendritic characteristics, the following correlations with cell size were found: smaller cells had larger form factors than larger cells (P < 0.05). Larger neurons had more extensive dendritic arborization, expressed by total dendritic length (P < 0.01) and number of dendritic branching points (P < 0.01), than smaller cells. It was suggested that small cNTS neurons contribute specifically to an integration of input information generated in the local circuits, while large neurons convey the integrated information to other autonomic brain regions.

After ipsilateral injections of cholera toxin B subunit (CTb) into the nucleus of the solitary tract (NST) and biotinylated dextran amine (BDA) into the insular cortex (IC) or into the central amygdaloid nucleus (ACe) in the rat, the prominent overlapping distribution of CTb-labeled neurons and BDA-labeled axon terminals was found in the posterolateral hypothalamus (PLH) region just medial to the subthalamic nucleus ipsilateral to the injection sites. At the electron microscopic level, the IC terminals formed asymmetrical synaptic contacts with dendrites and dendritic spines of the NST-projecting PLH neurons, whereas the ACe terminals formed symmetrical synaptic contacts with somata and dendrites of the NST-projecting PLH neurons. The present data suggest that output signals from the IC and ACe may exert excitatory and inhibitory influences, respectively, upon the PLH neurons that project to the NST for regulating cardiovascular functions.

The changes in mean arterial pressure (MAP) and respiratory frequency (RF) in response to microinjection of NMDA or AMPA into the commissural subnucleus of the NTS (comNTS) at the calamus scriptorius level of awake rats were evaluated. Under tribromoethanol anesthesia, the rats received guide-cannulae in direction of the NTS and a catheter was inserted into the femoral artery for measurement of arterial pressure. Changes in RF were evaluated with the rats inside a plethysmographic chamber. Randomly microinjections of 5 doses of NMDA (0.001, 0.01, 0.1, 1.0 and 2 nmol/50 nL; n = 10) or AMPA (1, 5, 10, 25 and 50 pmol/50 nL; n = 8) into the comNTS were performed at 15 min intervals and produced a dose-dependent increase in MAP [NMDA (3 +/- 2, 4 +/- 3, 25 +/- 4, 41 +/- 4 and 51 +/- 4 mm Hg) and AMPA (0 +/- 1, 14 +/- 4, 17 +/- 3, 27 +/- 5 and 34 +/- 3 mm Hg)]. Microinjection of NMDA (1 nmol/50 nL; n = 7) or AMPA (50 pmol/50 nL; n = 4) into the comNTS produced a long lasting apnea. The pressor responses to microinjection of NMDA or AMPA into the comNTS were blocked by prazosin, a alpha(1)-adrenoceptor antagonist, indicating that the increase in arterial pressure in both cases was sympathetically mediated. The data show that microinjection of NMDA and AMPA into the comNTS produced pressor response and apnea, indicating that both ionotropic l-glutamate receptors may play a role in the neurotransmission/neuromodulation of the autonomic and respiratory components of the cardiovascular reflexes at this level.

Cardiovascular changes occur during mental stress and in certain types of mood disorders. The neural basis for this phenomenon is unknown but it may be dependent on CNS neurons that provide branched projections to affective processing regions of the brain, such as the medial prefrontal cortex, and to the sympathetic outflow system. Because these putative neurons may be connected to these two target sites by chains of neurons, we performed double virus transneuronal tracing experiments and show here that a select subset of neurons in the medial preoptic nucleus (MPN), lateral hypothalamic area (LHA), and nucleus tractus solitarius (NTS) are co-linked to these two sites. Neurotensin MPN, orexin-containing LHA, and catecholamine NTS neurons were the major phenotypes involved in these projections. This novel class of neurons may coordinate cardiovascular changes seen in different emotional states.

The paratrigeminal nucleus (PTN) receives orofacial somatic and visceral afferent fibers and contains many calbindin-D28k neurons (CB-containing neurons) that project to nucleus of the solitary tract (NTS). In the present study, retrograde and transganglionic tracing methods combined with immunofluorescence histochemistry and confocal laser scanning microscopy were used. After Fluoro-gold (FG) injection into the unilateral NTS, 74.4% FG-labeled neurons of ipsilateral PTN were double-labeled with CB. Furthermore, 41.0% and 32.5% FG/CB double-labeled neurons co-existed with Fos induced by nociceptive stimulation of the lips and the upper alimentary tract, respectively. In the PTN unilateral to FG injection site, 26.6% CB-LI neurons were double-labeled with PAG, 61.5% and 79.0% CB/PAG double-labeled neurons were triple-labeled with FG and Fos, and 22.9% FG/CB double-labeled neurons were triple-labeled with PAG, 84.3% FG/PAG double-labeled neurons expressed Fos induced by the upper alimentary tract stimulation. In the intact animals, 62.8% CB-LI neurons and 88.3% PAG-LI neurons co-existed with GABA(B)R, respectively. In addition, some terminals from the inferior alveolar nerve (IAN) were closely apposed to CB/Fos double-labeled or CB single-labeled neurons. These results suggested that CB-containing neurons in the PTN receive the nociceptive information converge from the orofacial area and visceral organs, and comprising the glutamatergic excitatory transmission pathway from the PTN to the NTS. This pathway might be modulated by GABA via the GABA(B) receptor.

Microinjections of low doses (in the femtomolar or low picomolar range) of angiotensin II (Ang II) into the nucleus tractus solitarii (NTS) evoke depressor responses. In this study we have mapped in the rat the precise location of the subregion within the NTS at which Ang II evokes significant sympathoinhibitory and depressor responses. Microinjections of 1 pmol of Ang II evoked large decreases (>or=20% of baseline) in renal sympathetic nerve activity (RSNA), from a highly restricted region in the medial NTS, at or very close to the level 0.2 mm caudal to the obex. Microinjections of the same dose of Ang II into the commissural or lateral NTS at the same rostrocaudal level, or into the medial and lateral NTS at the level of the obex evoked significantly smaller sympathoinhibitory responses, while microinjections into more rostral or caudal levels of the NTS evoked significant sympathoinhibitory responses even less frequently. In most cases (71%), the sympathoinhibitory responses were accompanied by depressor responses, the magnitudes of which were also greater within the medial NTS at the level 0.2 mm caudal to obex, as compared to the surrounding subregions. The results demonstrate that the cardiovascular effects of Ang II in the NTS are highly site-specific. Taken together with previous studies, the results also indicate that the neurons in the NTS that mediate the Ang II-evoked sympathoinhibition are a discrete subgroup of the population of sympathoinhibitory neurons within the nucleus.

The excitatory amino acid L-glutamate injected into the nucleus of the solitary tract (NTS) in unanesthetized rats similar to peripheral chemoreceptor activation increases mean arterial pressure (MAP) and reduces heart rate. In this study, we investigated the effects of acute (1 day) and chronic (15 days) electrolytic lesions of the preoptic-periventricular tissue surrounding the anteroventral third ventricle (AV3V region) on the pressor and bradycardic responses induced by injections of L-glutamate into the NTS or peripheral chemoreceptor activation in unanesthetized rats. Male Holtzman rats with sham or electrolytic AV3V lesions and a stainless steel cannula implanted into the NTS were used. Differently from the pressor responses (28+/-3 mm Hg) produced by injections into the NTS of sham-lesioned rats, L-glutamate (5 nmol/100 nl) injected into the NTS reduced MAP (-26+/-8 mm Hg) or produced no effect (2+/-7 mm Hg) in acute and chronic AV3V-lesioned rats, respectively. The bradycardia to l-glutamate into the NTS and the cardiovascular responses to chemoreflex activation with intravenous potassium cyanide or to baroreflex activation with intravenous phenylephrine or sodium nitroprusside were not modified by AV3V lesions. The results show that the integrity of the AV3V region is essential for the pressor responses to L-glutamate into the NTS but not for the pressor responses to chemoreflex activation, suggesting dissociation between the central mechanisms involved in these responses.

It is well known that metabotropic glutamate receptors (mGluRs) have multiple actions on neuronal excitability mediated by G-protein-coupled receptors, although the exact mechanisms by which these actions occur are not understood. This study examines the effects of mGluRs agonists on voltage-dependent Ca2+ channels (VDCCs) currents (ICa) in the nucleus tractus solitarius (NTS) of rats using patch-clamp recording methods. An application of (RS)-3,5-dihydroxyphenylglycine (DHPG, Group I mGluR agonist) caused both facilitation and inhibition of L-type and N/P/Q-types ICa, respectively. Neither (2S, 2'R, 3'R)-2-(2', 3'-dicarboxycyclopropyl)glycine (DCG, Group II mGluRs agonist) nor L-(+)-2-amino-4-phosphonobutyric acid (AP-4, Group III mGluRs agonist) nor (RS)-2-chloro-5-hydroxyphenylglycine (CHPG, mGluR5 agonist) modulated ICa. Intracellular dialysis of the Gq/11-protein antibody and Gi-protein antibody attenuated the DHPG-induced facilitation and inhibition, respectively. The phospholipase C (PLC) inhibitor, as well as inhibition of either the protein kinase C (PKC) or inositol-1,4,5-trisphosphate (IP3) attenuated the DHPG-induced facilitation of ICa but not a DHPG-induced inhibition. Application of a strong depolarizing voltage prepulse attenuated the DHPG-induced inhibition of ICa. These results indicate that mGluR1 facilitates L-type VDCCs via Gq/11-protein involving PKC including IP3 formation. On the other hand, mGluR1 inhibits N- and P/Q-types VDCCs via Gi-protein betagamma subunits.

The nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus nerve (DMV) constitute sensory and motor nuclei of the dorsal vagal complex, respectively. We used whole-cell patch-clamp recordings from DMV neurons in rat brain slices and three methods of stimulation (electrical, glutamate microdrop, glutamate photostimulation) to test the hypothesis that convergent excitatory and inhibitory inputs to DMV neurons originate from intact neurons in multiple NTS areas. Electrical stimulation of the NTS resulted in evoked excitatory and inhibitory postsynaptic currents (eEPSCs and eIPSCs) in DMV neurons. Stimulation of the dorsal NTS with glutamate microdrops, which selectively stimulates the soma and dendrites of intact neurons, resulted in 31% of DMV neurons receiving eEPSCs, 44% receiving eIPSCs, and 6% receiving convergent excitatory and inhibitory inputs. Glutamate photostimulation allowed selective activation of intact neurons in multiple, discrete areas of the NTS and resulted in 36% of DMV neurons receiving eEPSCs, 65% receiving eIPSCs and 20% receiving both inputs. Data obtained by stimulation of multiple NTS areas support the hypothesis that there are anatomically convergent inputs to DMV neurons originating from intact neurons within the NTS. These data support the hypothesis that there is transfer of convergent information from the NTS to the DMV, implying that significant sensory-motor processing occurs within the brainstem.

The aim of this study was to identify the medullary vagal nuclei involved in the different phases of swallowing activated physiologically in a species with an esophagus similar to human. In decerebrate cats, the pharyngeal (0.5-1.0 ml water in pharynx (N=6)) or esophageal (1-3 ml air in esophagus (N=5)) phases of swallowing were stimulated separately once per minute for 3 h, and we compared the resulting c-fos immunoreactivity within neuronal cell nuclei of the dorsal motor nucleus (DMN), nucleus tractus solitarius (NTS) and nucleus ambiguus (NA) with a sham control group (N=5). We found that the pharyngeal phase was associated with an elevated number of c-fos positive neurons in the intermediate (NTSim), interstitial (NTSis), ventromedial (NTSvm) subnuclei of the NTS, caudal DMN, and dorsal NA; and the esophageal phase was associated with an elevated number of c-fos positive neurons in the central (NTSce), ventral, dorsolateral, ventrolateral subnuclei of the NTS, rostral DMN, and ventral NA. We concluded that the pharyngeal and esophageal phases of swallowing are associated with different sets of NTS subnuculei; and the DMN and NA may contain functionally different populations of motor neurons situated rostrocaudally and dorsoventrally associated with the different phases of swallowing. The central pattern generator (CPG) for swallowing probably receives significant peripheral feedback, and the NTSvm may participate in the transition of the pharyngeal to the esophageal phase of swallowing.

Insular cortex (IC) is recognized as a potential site for "central command" of cardiorespiratory responses during exercise. Muscular contraction (MC) decreased the discharge rate of most IC neurons. Activity of most contraction sensitive neurons was either not altered by elevating blood pressure or showed a response converse to that of MC. IC may thus have a role in central command but the area is clearly modulated by MC., (Copyright 2004 Elsevier B.V.)

Migraine pain arises in the trigeminovascular system and is often associated with nausea and sometimes with vomiting. In this study, an in vivo cat model of trigeminovascular stimulation was used to determine first whether there is a functional connection between the trigeminovascular system and the nucleus tractus solitarius (NTS), which is involved in regulating vomiting, and second whether anti-migraine drugs have any effect on such a connection. Chloralose-anaesthetised cats (n=16) were prepared for single neuron recording. The superior sagittal sinus (SSS) was isolated and stimulated electrically. The brainstem near the obex was exposed and a metal microelectrode equipped with six glass barrels for microiontophoresis was placed in the NTS. Recordings were made from 44 NTS neurons which responded to SSS stimulation with A-delta latencies. Iontophoretic ejection (50 nA) of eletriptan or naratriptan suppressed the response in 75% (15/20) and 78% (11/14) of cells and caused an average suppression of cell firing of 42+/-5% (n=20) and 54+/-8% (n=14), respectively. This suppression could be antagonized by the concurrent ejection (20-50 nA) of the 5-HT(1B/1D) receptor antagonist GR127935. We conclude that activation of the trigeminovascular system excites cells in the NTS that can be inhibited by eletriptan and naratriptan through activation of 5-HT(1B/1D) receptors. It is possible that in patients having a migraine attack trigeminovascular activation triggers nausea and vomiting, and that the alleviation of these symptoms by anti-migraine compounds may be via an action at 5-HT(1B/1D) receptors in the NTS.

These studies examined whether posttraining activation of alpha1-noradrenergic receptors in the nucleus tractus solitarius (NTS) influences neural processes that are involved in encoding information into memory. Different groups of male Sprague-Dawley rats were trained in two separate learning tasks. In experiment 1, rats were given either a control solution or the alpha1-noradrenergic agonist phenylephrine (0.5, 1.0, 5.0, or 10 microg/0.5 microl) directly into the NTS immediately after they were given a footshock (0.35 mA, 0.5 s) in the dark compartment of an inhibitory apparatus. In a retention test given 48 h later, groups that received either 5.0 or 10.0 microg of phenylephrine avoided the dark compartment for a significantly longer period of time than the PBS control group (P<0.05 and P<0.01, respectively). In experiment 2, identical doses of phenylephrine were infused in the NTS following footshock delivery in one alley of a Y-maze. Animals given either 1.0 or 5.0 microg of phenylephrine performed significantly better than PBS controls on several different measures that served as indices of retention. The results indicate that activation of alpha1-noradrenergic receptors in the NTS plays a critical role in the transmission of signals from the periphery to brain systems that process memory for emotionally significant experiences.

Immunoreactivity for nociceptin, an endogenous ligand for the ORL1 opioid receptors, has been reported in the nucleus tractus solitarius (nTS). A midline area in the commissural subnucleus (nCom) of nTS is the site of peripheral chemoreceptor projections. This investigation was carried out in urethane-anesthetized, artificially ventilated, adult male Wistar rats, to study the cardiovascular effects of the activation of ORL1 receptors in a midline area of the nCom. Microinjections (30 nl) of nociceptin (0.15-0.62 mM) into the nCom elicited depressor and bradycardic responses. Prior microinjections of [N-Phe(1)]-nociceptin-(1-13)-NH(2) (4.5 mM), a specific antagonist for ORL1 opioid receptors, into the nCom blocked the effects of nociceptin (0.31 mM, the maximally effective concentration), but not endomorphin-2 (0.6 mM; an endogenous ligand for micro -opioid receptors). On of other hand, naloxone (0.125 mM; an antagonist for classical opioid receptors) did not block the effects of nociceptin, while it did block the effects of endomorphin-2. The blockade of nociceptin effects by [N-Phe(1)]-nociceptin-(1-13)-NH(2) and endomorphin-2 by naloxone, was not due to some nonspecific effects because the responses to L-Glu (5 mM) remained unaltered after the microinjection of these antagonists. These results indicate that activation of ORL1 receptors in the nCom may play a role in the regulation of cardiovascular function.

The aim of this work was to investigate the modulation of the cardiovascular effects of neuropeptide Y (NPY) in the nucleus tractus solitarii (NTS) by angiotensin II (Ang II) and to determine the NPY receptor subtype involved in this modulation. Anesthesized Sprague-Dawley rats received microinjections in the NTS of Ang II (threshold and ED(50) doses) with NPY Y(1) agonist Leu(31)Pro(34)NPY and NPY Y(2) agonist NPY(13-36) (threshold and ED(50) doses). The changes in mean arterial pressure (MAP) and heart rate (HR) recorded in the femoral artery were analyzed during 60 min after the microinjections. The injection of threshold doses of Ang II, Y(1) agonist or Y(2) agonist alone did not produce any change in cardiovascular parameters. However, the co-injections into the NTS of threshold doses of both Ang II and the Y(1) agonist elicited significant increases of MAP and HR of about 12 and 10%, respectively. The co-administration of threshold doses of Ang II with the Y(2) agonist also induced a significant vasopressor response. The vasodepressor and bradycardiac effect of an ED(50) dose of the Y(1) agonist was significantly counteracted (P<0.01) by a threshold dose of Ang II. The vasopressor effect elicited by an ED(50) dose of the Y(2) agonist was significantly enhanced by a threshold dose of Ang II (P<0.01). No significant change of cardiovascular responses elicited by an ED(50) dose of Ang II was observed in the presence of threshold doses of the Y(1) agonist or of the Y(2) agonist. The present study gives functional evidences for a differential modulatory activity of Ang II on the cardiovascular responses mediated by Y(1) and Y(2) receptor subtypes, which may be of relevance for central cardiovascular regulation in the NTS.

The lateral hypothalamus (LH) regulates metabolic, behavioral and autonomic functions. The influence of the LH on gastrointestinal function and feeding behavior may be mediated by the dorsal vagal complex (DVC). In the present experiment, we used tract tracing and neurophysiologic techniques to evaluate the interrelationship between the LH and DVC. Using the tracer DiI, we demonstrated that the LH projects to both the nucleus of the solitary tract (NST) and the dorsal motor nucleus of the vagus (DMNV). We determined the effects of electrical stimulation of the LH and/or distention of the gastrointestinal tract on the firing rates of 107 DMNV neurons and 68 NST neurons. As previously reported, the majority of the DMNV neurons were inhibited and the majority of the NST neurons were excited by gastrointestinal distention. Electrical stimulation of the LH significantly changed the spontaneous activities of 71% of the DMNV neurons (46 excited and 30 inhibited). Of the 68 NST neurons characterized, 25 neurons were inhibited and 8 were excited by LH stimulation. In a separate experiment, we characterized the effects of both electrical and chemical stimulation of the LH on 36 DMNV and 14 NST neurons. Glutamate (0.8 nM) induced similar responses in the DVC neurons as electrical stimulation of the LH. The results indicate that the LH influences the electrical activity of DVC neurons. This effect may be the mechanism by which the LH modulates gastrointestinal function and feeding behavior.

Experiments were performed to investigate the effect of 17beta-estradiol (E; 30 pg/ml plasma) treatment (15-25 days) in the ovariectomized (OVX) female Wistar rat on the cardiovascular responses to hypocretin-1 (hcrt-1) in the nucleus tractus solitarius (NTS). In an initial series of experiments, the distribution of hcrt-1-like immunoreactivity within the region of the NTS was mapped in both OVX only and OVX+E animals. Hcrt-1 immunoreactivity was found throughout the NTS region in both groups of females, predominantly within the caudal interstitial, commissural, medial and lateral subnuclei of the NTS. The relative density of hcrt-1 immunoreactivity in all NTS subnuclei was similar in both female groups. Microinjections of hcrt-1 (0.5-10 pmol) into the caudal lateral and medial subnuclei of the NTS complex of the alpha-chloralose of the urethane-anaesthetized E-treated OVX rat elicited a dose-related decrease in heart rate (HR). On the other hand, although a dose-response effect on arterial pressure was evident, significant arterial pressure responses were observed only at the higher dose of hcrt-1 (>2.5 pmol). In the OVX only female rat, microinjection of hcrt-1 into similar NTS sites elicited a bradycardia and depressor response only at the highest dose of hcrt-1, and these responses were significantly smaller in magnitude than those elicited in the OVX+E animal. In addition, in the OVX only animals, a few sites within the caudal commissural subnucleus of the NTS complex were found at which hcrt-1 elicited tachycardia and pressor responses. Finally, it was found that the reflex bradycardia to the activation of arterial baroreceptors as a result of increasing systemic arterial pressure with phenylephrine (2-4 microg/kg) was significantly potentiated in the OVX+E animals only. These data suggest that hcrt-1 in the NTS of the female activates a neuronal circuit that controls the circulation and that the circulating level of E alters the sensitivity of these cardiovascular circuits to hcrt-1.

Electrophysiological methods were used to investigate the effects of trigeminal nerve stimulation or transection on responses of single gustatory neurons in the nucleus of the solitary tract (NTS) to tastants (NaCl, sucrose, citric acid, monosodium glutamate) in pentobarbital-anesthetized rats. Unilateral transection of the lingual nerve, or the mandibular branch of the trigeminal nerve, resulted in significant reductions (by 21 and 29%, respectively; P<0.01) in tastant-evoked responses, with no further effect following bilateral transection. Electrical stimulation of the central cut end of the mandibular nerve directly excited nine of 14 gustatory NTS units. For these units, central mandibular stimulation facilitated the tastant-evoked responses in six, depressed responses in three, and had no effect in five. Facilitation of tastant-evoked responses peaked 4 min after mandibular stimulation and recovered within 8 min. Electrical stimulation of the peripheral cut end of the mandibular nerve significantly reduced tastant-evoked responses in nine other NTS units, with a maximal reduction at 4 min post-stimulation followed by recovery. Stimulation of the superior cervical sympathetic ganglion did not affect NTS tastant-evoked responses. These results suggest the presence of complex central modulation of NTS neurons by trigeminal afferents, as well as a peripheral depressant effect on gustatory processing possibly mediated via neuropeptide release from trigeminal nerve endings in the tongue.

Gustatory processing within the medulla is modulated by a number of physiologic and experiential factors. Several neurotransmitters, including excitatory amino acids, GABA, and substance P, are involved in synaptic processing within the rostral portion of the nucleus of the solitary tract (NST). Endogenous opiates have been implicated in the regulation of feeding behavior and in taste palatability and gustatory responses in the parabrachial nuclei are reduced by systemic morphine. In the present experiments, extracellular recording of neuronal activity within the NST in response to taste input was combined with local microinjection of met-enkephalin (Met-ENK) and naltrexone (NLTX) to determine the effect of these agents on gustatory activity. The anterior tongue was stimulated with anodal current pulses to determine the time course of drug action (n=85 cells) and with prototypical taste stimuli (0.032 M sucrose, NaCl, and quinine hydrochloride, and 0.0032 M citric acid) to investigate the effects of these opioid compounds on taste-evoked responses (n=80 cells). Among these 165 taste-responsive neurons in the NST, the activity of 39 (23.6%) was suppressed by Met-ENK. These effects were dose-dependent and blockable by NLTX, which alone was without effect, suggesting that opiates do not maintain a tonic inhibitory influence. Immunohistochemical experiments demonstrated both micro - and delta-opioid receptors within the gustatory portion of the NST; previous studies had shown numerous fiber terminals containing Met-ENK. These data suggest that endogenous opiates play an inhibitory role in gustatory processing within the medulla.