Characterization of the Basic Membrane Properties of Neurons of the Rat Dorsal Motor Nucleus of the Vagus in Paraquat-Induced Models of Parkinsonism.

Most of Parkinson's disease (PD) patients experience gastrointestinal dysfunctions, including gastric hypomotility. The dorsal motor nucleus of the vagus (DMV) modulates the motility of the upper gastrointestinal (GI) tract. Paraquat (P) administration induces Parkinsonism in experimental models, and we have developed recently an environmental model of Parkinsonism in which rats are treated with subthreshold doses of P and lectins (P + L), in both models rats develop reduced gastric motility prodromal to the full extent of motor deficits. The aim of the present study was to examine whether the membrane properties of DMV neurons in these two experimental models of Parkinsonism were altered. Whole cell recordings in slices containing DMV neurons were conducted in male Sprague Dawley rats which received either injections of paraquat (10 mg/kg i.p.; 10P), or oral administration of paraquat (1 mg/kg) and lectin (0.05% w/v; P + L). Morphological reconstructions of DMV neurons were conducted at the end of the recordings. The repolarization kinetics of the afterhyperpolarization phase of the action potential was accelerated in 10P neurons vs control, while the phase plot revealed a slower depolarizing slope. At baseline, the amplitude of miniature excitatory postsynaptic currents was increased in P + L neurons. No differences in the morphology of DMV neurons were observed. These data indicate that the membrane and synaptic properties of DMV neurons are altered in rodent models of Parkinsonism, in which neurons of 10P and P + L rats demonstrate an increased excitatory transmission, perhaps in an attempt to counteract the paraquat-induced gastric hypomotility.

Ingestion of subthreshold doses of environmental toxins induces ascending Parkinsonism in the rat.

Increasing evidence suggests that environmental neurotoxicants or misfolded α-synuclein generated by such neurotoxicants are transported from the gastrointestinal tract to the central nervous system via the vagus nerve, triggering degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and causing Parkinson's disease (PD). We tested the hypothesis that gastric co-administration of subthreshold doses of lectins and paraquat can recreate the pathology and behavioral manifestations of PD in rats. A solution containing paraquat + lectin was administered daily for 7 days via gastric gavage, followed by testing for Parkinsonian behavior and gastric dysmotility. At the end of the experiment, brainstem and midbrain tissues were analyzed for the presence of misfolded α-synuclein and neuronal loss in the SNpc and in the dorsal motor nucleus of the vagus (DMV). Misfolded α-synuclein was found in DMV and SNpc neurons. A significant decrease in tyrosine hydroxylase positive dopaminergic neurons was noted in the SNpc, conversely there was no apparent loss of cholinergic neurons of the DMV. Nigrovagally-evoked gastric motility was impaired in treated rats prior to the onset of parkinsonism, the motor deficits of which were improved by l-dopa treatment. Vagotomy prevented the development of parkinsonian symptoms and constrained the appearance of misfolded α-synuclein to myenteric neurons. These data demonstrate that co-administration of subthreshold doses of paraquat and lectin induces progressive, l-dopa-responsive parkinsonism that is preceded by gastric dysmotility. This novel preclinical model of environmentally triggered PD provides functional support for Braak's staging hypothesis of idiopathic PD.

Perinatal high fat diet increases inhibition of dorsal motor nucleus of the vagus neurons regulating gastric functions.

BACKGROUND: Previous studies suggest an increased inhibition of dorsal motor nucleus of the vagus (DMV) neurons following exposure to a perinatal high fat diet (PNHFD); the underlying neural mechanisms, however, remain unknown. This study assessed the effects of PNHFD on inhibitory synaptic inputs to DMV neurons and the vagally dependent control of gastric tone and motility. METHODS: Whole-cell patch clamp recordings were made from DMV neurons in thin brainstem slices from Sprague-Dawley rats fed either a control diet or HFD (14 or 60% kcal from fat, respectively) from embryonic day 13 onwards; gastric tone and motility were recorded in in vivo anesthetized rats. KEY RESULTS: The non-selective GABAA antagonist, BIC (10 µmol L-1 ), induced comparable inward currents in PNHFD and control DMV neurons, but a larger current in PNHFD neurons at higher concentrations (50 µmol L-1 ). Differences were not apparent in neuronal responses to the phasic GABAA antagonist, gabazine (GBZ), the extrasynaptic GABAA agonist, THIP, the GABA transport blocker, nipecotic acid, or the gliotoxin, fluoroacetate, suggesting that PNHFD altered inhibitory transmission but not GABAA receptor density or function, GABA uptake or glial modulation of synaptic strength. Similarly, the increase in gastric motility and tone following brainstem microinjection of low doses of BIC (1-10 pmoles) and GBZ (0.01-0.1 pmoles) were unchanged in PNHFD rats while higher doses of BIC (25 pmoles) induced a significantly larger increase in gastric tone compared to control. CONCLUSIONS AND INFERENCES: These studies suggest that exposure to PNHFD increases the tonic inhibition of DMV neurons, possibly contributing to dysregulated vagal control of gastric functions.

Vagally mediated effects of brain stem dopamine on gastric tone and phasic contractions of the rat.

Dopamine (DA)-containing fibers and neurons are embedded within the brain stem dorsal vagal complex (DVC); we have shown previously that DA modulates the membrane properties of neurons of the dorsal motor nucleus of the vagus (DMV) via DA1 and DA2 receptors. The vagally dependent modulation of gastric tone and phasic contractions, i.e., motility, by DA, however, has not been characterized. With the use of microinjections of DA in the DVC while recording gastric tone and motility, the aims of the present study were 1) assess the gastric effects of brain stem DA application, 2) identify the DA receptor subtype, and, 3) identify the postganglionic pathway(s) activated. Dopamine microinjection in the DVC decreased gastric tone and motility in both corpus and antrum in 29 of 34 rats, and the effects were abolished by ipsilateral vagotomy and fourth ventricular treatment with the selective DA2 receptor antagonist L741,626 but not by application of the selective DA1 receptor antagonist SCH 23390. Systemic administration of the cholinergic antagonist atropine attenuated the inhibition of corpus and antrum tone in response to DA microinjection in the DVC. Conversely, systemic administration of the nitric oxide synthase inhibitor nitro-l-arginine methyl ester did not alter the DA-induced decrease in gastric tone and motility. Our data provide evidence of a dopaminergic modulation of a brain stem vagal neurocircuit that controls gastric tone and motility.NEW & NOTEWORTHY Dopamine administration in the brain stem decreases gastric tone and phasic contractions. The gastric effects of dopamine are mediated via dopamine 2 receptors on neurons of the dorsal motor nucleus of the vagus. The inhibitory effects of dopamine are mediated via inhibition of the postganglionic cholinergic pathway.

Downregulation of neuronal vasoactive intestinal polypeptide in Parkinson's disease and chronic constipation.

BACKGROUND: Chronic constipation (CC) is a common and severe gastrointestinal complaint in Parkinson's disease (PD), but its pathogenesis remains poorly understood. This study evaluated functionally distinct submucosal neurons in relation to colonic motility and anorectal function in PD patients with constipation (PD/CC) vs both CC and controls. METHODS: Twenty-nine PD/CC and 10 Rome III-defined CC patients were enrolled. Twenty asymptomatic age-sex matched subjects served as controls. Colonic transit time measurement and conventional anorectal manometry were evaluated in PD/CC and CC patients. Colonoscopy was performed in all three groups. Colonic submucosal whole mounts from PD/CC, CC, and controls were processed for immunohistochemistry with antibodies for vasoactive intestinal polypeptide (VIP) and peripheral choline acetyltransferase, markers for functionally distinct submucosal neurons. The mRNA expression of VIP and its receptors were also assessed. KEY RESULTS: Four subgroups of PD/CC patients were identified: delayed colonic transit plus altered anorectal manometry (65%); delayed colonic transit (13%); altered manometric pattern (13%); and no transit and manometric impairment (9%). There were no differences in the number of neurons/ganglion between PD/CC vs CC or vs controls. A reduced number of submucosal neurons containing VIP immunoreactivity was found in PD/CC vs controls (P<.05). VIP, VIPR1, and VIPR2 mRNA expression was significantly reduced in PD/CC vs CC and controls (P<.05). CONCLUSIONS AND INFERENCES: Colonic motor and rectal sensory functions are impaired in most PD/CC patients. These abnormalities are associated with a decreased VIP expression in submucosal neurons. Both sensory-motor abnormalities and neurally mediated motor and secretory mechanisms are likely to contribute to PD/CC pathophysiology.

Diminished vagal tone is a predictive biomarker of necrotizing enterocolitis-risk in preterm infants.

BACKGROUND: Necrotizing enterocolitis (NEC) is an acute neonatal inflammatory disease which may lead to intestinal necrosis, multisystem failure, and death. Currently, NEC is diagnosed by a combination of laboratory and radiographic tests conducted a posteriori i.e., when NEC is already clinically significant. Given the acute onset and rapid progression of NEC, a non-invasive biomarker that allows early detection of patients at risk is required as a matter of urgency. We evaluated whether the high frequency (HF) component of heart rate variability (HRV), a measure of vagal efferent tonic cholinergic activity may be used as a predictive biomarker for NEC-risk before the onset of clinical disease. METHODS: In this prospective study, stable preterm (gestational age 28-35 weeks) infants had HRV power spectra analyzed from surface electrocardiogram waveforms taken at rest on day 5-8 of life. We used regression modeling to determine the utility of HF-HRV in predicting NEC. KEY RESULTS: HF-HRV power was 21.5 ± 2.7 and 3.9 ± 0.81 ms(2) in infants that remained healthy and those that later developed stage 2+ NEC, respectively (p < 0.001). Nine of 70 enrolled infants developed NEC. The ROC discriminated a HF-HRV value of 4.68 ms(2) predictive for developing NEC with a sensitivity and specificity of 89% and 87%, and positive and negative predictive value of 50% and 98%, respectively. With predictive regression modeling, the risk (odds ratio) of developing NEC was 10 per every one SD decrease in HF-HRV. CONCLUSIONS & INFERENCES: Our preliminary data indicate that HF-HRV may serve as a potential, non-invasive predictive biomarker of NEC-risk in NICU infants.

Ghrelin increases vagally mediated gastric activity by central sites of action.

BACKGROUND: Vagally dependent gastric reflexes are mediated through vagal afferent fibers synapsing upon neurons of the nucleus tractus solitarius (NTS) which, in turn modulate the preganglionic parasympathetic dorsal motor nucleus of the vagus (DMV) neurons within the medullary dorsal vagal complex (DVC). The expression and transport of ghrelin receptors has been documented for the afferent vagus nerve, and functional studies have confirmed that vagal pathways are integral to ghrelin-induced stimulation of gastric motility. However, the central actions of ghrelin within the DVC have not been explored fully. METHODS: We assessed the responses to ghrelin in fasted rats using: (i) in vivo measurements of gastric tone and motility following IVth ventricle application or unilateral microinjection of ghrelin into the DVC and (ii) whole cell recordings from gastric-projecting neurons of the DMV. KEY RESULTS: (i) IVth ventricle application or unilateral microinjection of ghrelin into the DVC-elicited contractions of the gastric corpus via excitation of a vagal cholinergic efferent pathway and (ii) ghrelin facilitates excitatory, but not inhibitory, presynaptic transmission to DMV neurons. CONCLUSIONS & INFERENCES: Our data indicate that ghrelin acts centrally by activating excitatory synaptic inputs onto DMV neurons, resulting in increased cholinergic drive by way of vagal motor innervation to the stomach.

A critical re-evaluation of the specificity of action of perivagal capsaicin.

Perivagal application of capsaicin (1% solution) is considered to cause a selective degeneration of vagal afferent C fibres and has been used extensively to examine the site of action of many gastrointestinal (GI) neuropeptides. The actions of both capsaicin and GI neuropeptides may not be restricted to vagal afferent fibres, however, as other non-sensory neurones have displayed sensitivity to capsaicin and brainstem microinjections of these neuropeptides induce GI effects similar to those obtained upon systemic application. The aim of the present study was to test the hypothesis that perivagal capsaicin induces degeneration of vagal efferents controlling GI functions. Experiments were conducted 7-14 days after 30 min unilateral perivagal application of 0.1-1% capsaicin. Immunohistochemical analyses demonstrated that, as following vagotomy, capsaicin induced dendritic degeneration, decreased choline acetyltransferase but increased nitric oxide synthase immunoreactivity in rat dorsal motor nucleus of the vagus (DMV) neurones. Electrophysiological recordings showed a decreased DMV input resistance and excitability due, in part, to the expression of a large conductance calcium-dependent potassium current and the opening of a transient outward potassium window current at resting potential. Furthermore, the number of DMV neurones excited by thyrotrophin-releasing hormone and the gastric motility response to DMV microinjections of TRH were decreased significantly. Our data indicate that perivagal application of capsaicin induced DMV neuronal degeneration and decreased vagal motor responses. Treatment with perivagal capsaicin cannot therefore be considered selective for vagal afferent C fibres and, consequently, care is needed when using perivagal capsaicin to assess the mechanism of action of GI neuropeptides.

Experimental spinal cord injury in rats diminishes vagally-mediated gastric responses to cholecystokinin-8s.

BACKGROUND: We have shown recently that our model of experimental high-thoracic spinal cord injury (T3-SCI) mirrors the gastrointestinal clinical presentation of neurotrauma patients, whereby T3-SCI animals show diminished gastric emptying and dysmotility. In this study we used cholecystokinin as a model peptide to test the hypothesis that the T3-SCI induced gastroparesis is due, in part, to an impaired vagally-mediated response to gastrointestinal peptides. METHODS: We measured the responses to sulfated cholecystokinin (CCK-8s) in control and T3-SCI (3 or 21 days after injury) rats utilizing: (i) c-fos expression in the nucleus tractus solitarius (NTS) following peripherally administered CCK-8s; (ii) in vivo gastric tone and motility following unilateral microinjection of CCK-8s into the dorsal vagal complex (DVC); and (iii) whole cell recordings of glutamatergic synaptic inputs to NTS neurons. KEY RESULTS: Our results show that: (i) medullary c-fos expression in response to peripheral CCK-8s was significantly lower in T3-SCI rats 3 days after the injury, but recovered to control values at 3 weeks post-SCI, (ii) Unilateral microinjection of CCK-8s in the DVC induced a profound gastric relaxation in control animals, but did not induce any response in T3-SCI rats at both 3 and 21 days after SCI, (iii) Perfusion with CCK-8s increased glutamatergic currents in 55% of NTS neurons from control rats, but failed to induce any response in NTS neurons from T3-SCI rats. CONCLUSIONS & INFERENCES: Our data indicate alterations of vagal responses to CCK-8s in T3-SCI rats that may reflect a generalized impairment of gastric vagal neurocircuitry, leading to a reduction of gastric functions after SCI.

Plasticity of vagal brainstem circuits in the control of gastric function.

BACKGROUND: Sensory information from the viscera, including the gastrointestinal (GI) tract, is transmitted through the afferent vagus via a glutamatergic synapse to neurons of the nucleus tractus solitarius (NTS), which integrate this sensory information to regulate autonomic functions and homeostasis. The integrated response is conveyed to, amongst other nuclei, the preganglionic neurons of the dorsal motor nucleus of the vagus (DMV) using mainly GABA, glutamate and catecholamines as neurotransmitters. Despite being modulated by almost all the neurotransmitters tested so far, the glutamatergic synapse between NTS and DMV does not appear to be tonically active in the control of gastric motility and tone. Conversely, tonic inhibitory GABAergic neurotransmission from the NTS to the DMV appears critical in setting gastric tone and motility, yet, under basal conditions, this synapse appears resistant to modulation. PURPOSE: Here, we review the available evidence suggesting that vagal efferent output to the GI tract is regulated, perhaps even controlled, in an 'on-demand' and efficient manner in response to ever-changing homeostatic conditions. The focus of this review is on the plasticity induced by variations in the levels of second messengers in the brainstem neurons that form vago-vagal reflex circuits. Emphasis is placed upon the modulation of GABAergic transmission to DMV neurons and the modulation of afferent input from the GI tract by neurohormones/neurotransmitters and macronutrients. Derangement of this 'on-demand' organization of brainstem vagal circuits may be one of the factors underlying the pathophysiological changes observed in functional dyspepsia or hyperglycemic gastroparesis.

Modulation of inhibitory neurotransmission in brainstem vagal circuits by NPY and PYY is controlled by cAMP levels.

Pancreatic polypeptides such as neuropeptide Y (NPY) and peptide YY (PYY) exert profound, vagally mediated effects on gastrointestinal (GI) motility. Vagal efferent outflow to the GI tract is determined principally by tonic GABAergic synaptic inputs onto dorsal motor nucleus of the vagus (DMV) neurons, yet neither peptide modulates GABAergic transmission. We showed recently that opioid peptides appear similarly ineffective because of the low resting cAMP levels. Using whole cell recordings from identified DMV neurons, we aimed to correlate the influence of brainstem cAMP levels with the ability of pancreatic polypeptides to modulate GABAergic synaptic transmission. Neither NPY, PYY, nor the Y1 or Y2 receptor selective agonists [Leu,Pro]NPY or NPY(3-36) respectively, inhibited evoked inhibitory postsynaptic current (eIPSC) amplitude unless cAMP levels were elevated by forskolin or 8-bromo-cAMP, by exposure to adenylate cyclase-coupled modulators such as cholecystokinin octapeptide (sulfated) (CCK-8s) or thyrotropin releasing hormone (TRH), or by vagal deafferentation. The inhibition of eIPSC amplitude by [Leu,Pro]NPY or NPY(3-36) was stable for approximately 30 min following the initial increase in cAMP levels. Thereafter, the inhibition declined gradually until the agonists were again ineffective after 60 min. Analysis of spontaneous and miniature currents revealed that such inhibitory effects were due to actions at presynaptic Y1 and Y2 receptors. These results suggest that, similar to opioid peptides, the effects of pancreatic polypeptides on GABAergic transmission depend upon the levels of cAMP within gastric inhibitory vagal circuits.

Glucagon-like peptide-1 excites pancreas-projecting preganglionic vagal motoneurons.

Glucagon-like peptide-1 (GLP-1) increases pancreatic insulin secretion via a direct action on pancreatic beta-cells. A high density of GLP-1-containing neurons and receptors is also present in brain stem vagal circuits; therefore, the aims of the present study were to investigate 1) whether identified pancreas-projecting neurons of the dorsal motor nucleus of the vagus (DMV) respond to exogenously applied GLP-1, 2) the mechanism(s) of action of GLP-1, and 3) whether the GLP-1-responsive neurons (putative modulators of endocrine secretion) could be distinguished from DMV neurons responsive to peptides that modulate pancreatic exocrine secretion, specifically pancreatic polypeptide (PP). Whole cell recordings were made from identified pancreas-projecting DMV neurons. Perfusion with GLP-1 induced a concentration-dependent depolarization in approximately 50% of pancreas-projecting DMV neurons. The GLP-1 effects were mimicked by exendin-4 and antagonized by exendin-(9-39). In approximately 60% of the responsive neurons, the GLP-1-induced depolarization was reduced by tetrodotoxin (1 microM), suggesting both pre- and postsynaptic sites of action. Indeed, the GLP-1 effects were mediated by actions on potassium currents, GABA-induced currents, or both. Importantly, neurons excited by GLP-1 were unresponsive to PP and vice versa. These data indicate that 1) GLP-1 may act on DMV neurons to control pancreatic endocrine secretion, 2) the effects of GLP-1 on pancreas-projecting DMV neurons are mediated both via a direct excitation of their membrane as well as via an effect on local circuits, and 3) the GLP-1-responsive neurons (i.e., putative endocrine secretion-controlling neurons) could be distinguished from neurons responsive to PP (i.e., putative exocrine secretion-controlling neurons).

Effects of cholecystokinin-8s in the nucleus tractus solitarius of vagally deafferented rats.

We have shown recently that cholecystokinin octapeptide (CCK-8s) increases glutamate release from nerve terminals onto neurons of the nucleus tractus solitarius pars centralis (cNTS). The effects of CCK on gastrointestinal-related functions have, however, been attributed almost exclusively to its paracrine action on vagal afferent fibers. Because it has been reported that systemic or perivagal capsaicin pretreatment abolishes the effects of CCK, the aim of the present work was to investigate the response of cNTS neurons to CCK-8s in vagally deafferented rats. In surgically deafferented rats, intraperitoneal administration of 1 or 3 mug/kg CCK-8s increased c-Fos expression in cNTS neurons (139 and 251% of control, respectively), suggesting that CCK-8s' effects are partially independent of vagal afferent fibers. Using whole cell patch-clamp techniques in thin brain stem slices, we observed that CCK-8s increased the frequency of spontaneous and miniature excitatory postsynaptic currents in 43% of the cNTS neurons via a presynaptic mechanism. In slices from deafferented rats, the percentage of cNTS neurons receiving glutamatergic inputs responding to CCK-8s decreased by approximately 50%, further suggesting that central terminals of vagal afferent fibers are not the sole site for the action of CCK-8s in the brain stem. Taken together, our data suggest that the sites of action of CCK-8s include the brain stem, and in cNTS, the actions of CCK-8s are not restricted to vagal central terminals but that nonvagal synapses are also involved.

Melanin concentrating hormone innervation of caudal brainstem areas involved in gastrointestinal functions and energy balance.

Neural signaling by melanin-concentrating hormone and its receptor (SLC-1) has been implicated in the control of energy balance, but due to the wide distribution of melanin-concentrating hormone-containing fibers throughout the neuraxis, its critical sites of action for a particular effect have not been identified. The present study aimed to anatomically and functionally characterize melanin-concentrating hormone innervation of the rat caudal brainstem, as this brain area plays an important role in the neural control of ingestive behavior and autonomic outflow. Using retrograde tracing we demonstrate that a significant proportion (5-15%) of primarily perifornical and far-lateral hypothalamic melanin-concentrating hormone neurons projects to the dorsal vagal complex. In the caudal brainstem, melanin-concentrating hormone-ir axon profiles are distributed densely in most areas including the nucleus of the solitary tract, dorsal motor nucleus of the vagus, and sympathetic premotor areas in the ventral medulla. Close anatomical appositions can be demonstrated between melanin-concentrating hormone-ir axon profiles and tyrosine hydroxylase, GABA, GLP-1, NOS-expressing, and nucleus of the solitary tract neurons activated by gastric nutrient infusion. In medulla slice preparations, bath application of melanin-concentrating hormone inhibited in a concentration-dependent manner the amplitude of excitatory postsynaptic currents evoked by solitary tract stimulation via a pre-synaptic mechanism. Fourth ventricular administration of melanin-concentrating hormone (10 microg) in freely moving rats decreased core body temperature but did not change locomotor activity and food and water intake. We conclude that the rich hypothalamo-medullary melanin-concentrating hormone projections in the rat are mainly inhibitory to nucleus of the solitary tract neurons, but are not involved in the control of food intake. Projections to ventral medullary sites may play a role in the inhibitory effect of melanin-concentrating hormone on energy expenditure.

Cholecystokinin octapeptide increases spontaneous glutamatergic synaptic transmission to neurons of the nucleus tractus solitarius centralis.

Cholecystokinin (CCK) is released from enteroendocrine cells after ingestion of nutrients and induces multiple effects along the gastrointestinal tract, including gastric relaxation and short-term satiety. We used whole cell patch-clamp and immunohistochemical techniques in rat brain stem slices to characterize the effects of CCK. In 45% of the neurons of nucleus tractus solitarius subnucleus centralis (cNTS), perfusion with the sulfated form of CCK (CCK-8s) increased the frequency of spontaneous excitatory currents (sEPSCs) in a concentration-dependent manner (1-300 nM). The threshold for the CCK-8s excitatory effect was 1 nM, the EC(50) was 20 nM, and E(max) was 100 nM. The excitatory effects of CCK-8s were still present when the slices were preincubated with tetrodotoxin or bicuculline or when the recordings were conducted with Cs(+) electrodes. Pretreatment with the CCK-A receptor antagonist, lorglumide (1 microM), antagonized the effects of CCK-8s, whereas perfusion with the CCK-B preferring agonist CCK-8 nonsulfated (CCK-ns, 1 microM) did not affect the frequency of sEPSCs. Similarly, pretreatment with the CCK-B receptor antagonist, triglumide (1 microM), did not prevent the actions of CCK-8s. Although the majority (i.e., 76%) of CCK-8s unresponsive cNTS neurons had a bipolar somata shape and were TH-IR negative, no differences were found in either the morphological or the neurochemical phenotype of cNTS neurons responsive to CCK-8s. Our results suggest that the excitatory effects of CCK-8s on terminals impinging on a subpopulation of cNTS neurons are mediated by CCK-A receptors; these responsive neurons, however, do not have morphological or neurochemical characteristics that automatically distinguish them from nonresponsive neurons.

Characterization of neurons of the nucleus tractus solitarius pars centralis.

Esophageal sensory afferent inputs terminate principally in the central subnucleus of the tractus solitarius (cNTS). Neurons of the cNTS comprise two major neurochemical subpopulations. One contains neurons that are nitric oxide synthase (NOS) immunoreactive (-IR) while the other comprises neurons that are tyrosine hydroxylase (TH)-IR. We have shown recently that TH-IR neurons are involved in esophageal-distention induced gastric relaxation. We used whole cell patch clamp techniques in rat brainstem slices combined with immunohistochemical and morphological reconstructions to characterize cNTS neurons. Postrecording reconstruction of cNTS neurons revealed two morphological neuronal subtypes; one group of cells (41 out of 131 neurons, i.e., 31%) had a multipolar soma, while the other group (87 out of 131 neurons, i.e., 66%) had a bipolar soma. Of the 43 cells in which we conducted a neurochemical examination, 15 displayed TH-IR (9 with bipolar morphology, 6 with multipolar morphology) while the remaining 28 neurons did not display TH-IR (18 with bipolar morphology, 10 with multipolar morphology). Even though the range of electrophysiological properties varied significantly, morphological or neurochemical distinctions did not reveal characteristics peculiar to the subgroups. Spontaneous excitatory postsynaptic currents (sEPSC) recorded in cNTS neurons had a frequency of 1.5 +/- 0.15 events s(-1) and an amplitude of 27 +/- 1.2 pA (Vh = -50 mV) and were abolished by pretreatment with 30 muM AP-5 and 10 muM CNQX, indicating the involvement of both NMDA and non-NMDA receptors. Some cNTS neurons also received a GABAergic input that was abolished by perfusion with 30-50 muM bicuculline. In conclusion, our data show that despite the heterogeneity of morphological and neurochemical membrane properties, the electrophysiological characteristics of cNTS neurons are not a distinguishing feature.

Noradrenergic neurons in the rat solitary nucleus participate in the esophageal-gastric relaxation reflex.

Activation of esophageal mechanosensors excites neurons in and near the central nucleus of the solitary tract (NSTc). In turn, NSTc neurons coordinate the relaxation of the stomach [i.e., the receptive relaxation reflex (RRR)] by modulating the output of vagal efferent neurons of the dorsal motor nucleus of the vagus (DMN). The NSTc area contains neurons with diverse neurochemical phenotypes, including a large population of catecholaminergic and nitrergic neurons. The aim of the present study was to determine whether either one of these prominent neuronal phenotypes was involved in the RRR. Immunohistochemical techniques revealed that repetitive esophageal distension caused 53% of tyrosine hydroxylase-immunoreactive (TH-ir) neurons to colocalize c-Fos in the NSTc. No nitric oxide synthase (NOS)-ir neurons in the NSTc colocalized c-Fos in either distension or control conditions. Local brain stem application (2 ng) of alpha-adrenoreceptor antagonists (i.e., alpha1-prazosin or alpha2-yohimbine) significantly reduced the magnitude of the esophageal distension-induced gastric relaxation to approximately 55% of control conditions. The combination of yohimbine and prazosin reduced the magnitude of the reflex to approximately 27% of control. In contrast, pretreatment with either the NOS-inhibitor NG-nitro-l-arginine methyl ester or the beta-adrenoceptor antagonist propranolol did not interfere with esophageal distension-induced gastric relaxation. Unilateral microinjections of the agonist norepinephrine (0.3 ng) directed at the DMN were sufficient to mimic the transient esophageal-gastric reflex. Our data suggest that noradrenergic, but not nitrergic, neurons of the NSTc play a prominent role in the modulation of the RRR through action on alpha1- and alpha2-adrenoreceptors. The finding that esophageal afferent stimulation alone is not sufficient to activate NOS-positive neurons in the NSTc suggests that these neurons may be strongly gated by other central nervous system inputs, perhaps related to the coordination of swallowing or emesis with respiration.

Glucose effects on gastric motility and tone evoked from the rat dorsal vagal complex.

1. To examine the effects of glucose on the central components of the vago-vagal reflex control of gastric function, we performed both in vivo and in vitro experiments on neurones in the medial nucleus of the tractus solitarius (mNTS) and in the dorsal motor nucleus of the vagus (DMV). 2. In the in vivo anaesthetized rat preparation, unilateral microinjection of D-glucose (10 or 50 mM (60 nl)(-1)) in mNTS produced inhibition of gastric motility and an increase in intragastric pressure. D-glucose had no effect in the DMV. 3. In the in vitro rat brainstem slice preparation, whole-cell recordings of DMV neurones showed that increasing the glucose concentration of the perfusion solution from 5 mM to 15 or 30 mM produced outward currents of 35 +/- 5 pA (n = 7) and 51 +/- 10 pA (n = 11), respectively. These were blocked by tetrodotoxin and picrotoxin, indicating that glucose was acting indirectly to cause the release of GABA. Decreasing the glucose concentration of the perfusing solution by one-half produced an inward current of 36 +/- 5 pA (n = 7). 4. Stimulation of the NTS evoked inhibitory postsynaptic currents (IPSCs) in DMV neurones. The amplitude of the evoked IPSCs was positively correlated with glucose concentration. Perfusion with the ATP-sensitive K(+) (K(ATP)) channel opener diazoxide mimicked the effect of reduced glucose, while perfusion with the K(ATP) channel blocker glibenclamide mimicked the effects of increased glucose. 5. Our data indicate that glucose had no direct excitatory effect on DMV neurones, but DMV neurones appear to be affected by an action of glucose on cell bodies of mNTS neurones via effects on an ATP-sensitive potassium channel.

Receptors and transmission in the brain-gut axis: potential for novel therapies. V. Fast and slow extrinsic modulation of dorsal vagal complex circuits.

Vago-vagal reflex circuits in the medulla are responsible for the smooth coordination of the digestive processes carried out from the oral cavity to the transverse colon. In this themes article, we concentrate mostly on electrophysiological studies concerning the extrinsic modulation of these vago-vagal reflex circuits, with a particular emphasis on two types of modulation, i.e., by "fast" classic neurotransmitters and by "slow" neuromodulators. These examples review two of the most potent modulatory processes at work within the dorsal vagal complex, which have dramatic effects on gastrointestinal function. The reader should be mindful of the fact that many more different inputs from other central nervous system (CNS) loci or circulating humoral factors add to this complex mix of modulatory inputs. It is likely that similar long-term modulations of synaptic transmission occur with other neurotransmitters and may represent an important mechanism for the integration and regulation of neuronal behavior. Of course, this fact strongly militates against the success of any single drug or approach in the treatment of motility disorders having a CNS component.

Effects of substance P on identified neurons of the rat dorsal motor nucleus of the vagus.

Previous evidence suggests that substance P (SP) activates subpopulations of neurons within the dorsal motor nucleus of the vagus (DMV). In this study we aimed at identifying these subpopulations in relation to their gastrointestinal projection organs or vagal branches and characterizing pharmacologically the SP response. Using whole cell patch-clamp recordings from identified gastrointestinal-projecting vagal motoneurons, we found that SP induced an inward current in all neuronal groups except for cecum-projecting cells. The lowest percentage of SP-responding neurons was found in fundus-projecting cells, where SP also had a concentration-response curve that was shifted to the left (P < 0.05). Independently from the projections, the SP response was reduced by sendide and MEN 10,376 and mimicked by a combination of [Sar(9)-Met(O(2))(11)]SP and alpha-neurokinin. SP and alpha-neurokinin also increased the frequency, but not the amplitude, of postsynaptic currents. In conclusion, we demonstrated that SP induces both pre- and postsynaptic effects on DMV neurons via activation of neurokinin NK(1) and NK(2) receptors. The magnitude of the SP response was correlated to the peripheral target organ.