Adenosine release in nucleus tractus solitarii does not appear to mediate hypoxia-induced respiratory depression in rats

The time course of adenosine release in the nucleus tractus solitarii (NTS) and ventrolateral medulla (VLM) during acute systemic hypoxia was investigated in the anaesthetised rat by means of amperometric enzymatic sensors. It was found that acute hypoxia induced a significant delayed increase in adenosine level (reaching levels as high as 5 μM) in the NTS and that hypoxia-induced release of adenosine was similar at various regions of the NTS along its rostro-caudal axis. Significantly smaller or no increases in adenosine levels at all in response to hypoxia were observed in the VLM. The increase in adenosine level in the NTS occurred during reoxygenation after the termination of the hypoxic challenge and was accompanied by a smaller increase in inosine concentration. At the dorsal surface of the brainstem, only release of inosine was detected following acute hypoxia. Addition of the ecto-5'-nucleotidase inhibitor α,β-methylene ADP (200 μM) to the dorsal surface of the brainstem completely abolished the signal evoked by hypoxia, suggesting that the inosine arose from adenosine that was produced in the extracellular space by the prior release of ATP. This study indicates that following systemic hypoxia, adenosine levels in the NTS increase to a significantly greater extent than in the VLM. However, the increase in adenosine concentration in the NTS occurs too late to be responsible for the hypoxia-induced depression of the respiratory activity.     

Angiotensin AT1 receptor antagonist losartan and the defence reaction in the anaesthetised rat. Effect on the carotid chemoreflex

Modulation at the level of the nucleus tractus solitarii (NTS) appears to be an effective way of controlling cardiovascular reflexes. Angiotensin II acting on angiotensin AT1 receptors at the central nervous system appears to have an important role in these modulatory processes. The hypothalamic defence area (HDA) is a potential source of descending fibres containing angiotensin II that innervate the NTS. We investigated the effect of AT1 receptor blockade in the NTS on the response to stimulation of HDA in anaesthetised rats treated with the neuromuscular blocking agent pancuronium bromide. The characteristic increase in heart rate, blood pressure and phrenic nerve activity evoked by electrical stimulation of HDA is decreased by the microinjection of the AT1 receptor antagonist losartan into the NTS and the cardiovascular response to carotid body chemical stimulation is also reduced. These results support the hypothesis that AT1 receptors in the NTS play a role in the modulation of cardiovascular reflexes, and modify the influence exerted on the processing of these reflexes by other areas of the central nervous system.     

Release of adenosine-like material from isolated perfused dog adipose tissue following sympathetic nerve stimulation and its inhibition by adrenergic alpha-receptor blockade.

Following the intraarterial infusion of 3H-adenine to isolated perfused canine subcutaneous adipose tissue, its adenine nucleotides are labelled. A continuous release of radioactivity, comprised of non-nucleotide material, was observed. The rate of this release was markedly enhanced by sympathetic nerve stimulation. The major components of the enhanced release appeared to be inosine and adenosine. Adrenergic alpha-receptor blockade (phentolamine or Hydergin) abolished the enhanced nucleoside release, while glycerol release was enhanced. The release of radioactivity was decreased during mechanical blood flow reduction and enhanced afterwards. However, the magnitude of this enhancement of release after clamp was much less than following nerve stimulation. The results suggest that adenosine or a closely related compound is released from canine subcutaneous adipose tissue by sympathetic nerve stimulation and that the release is related to adrenergic alpha-receptor stimulation. Since adenosine is a potent inhibitor of catecholamine induced lipolysis in this tissue the possibility of a regulatory role must be considered.     

Release of Adenosine-like Material from Isolated Perfused Dog Adipose Tissue Following Sympathetic Nerve Stimulation and its Inhibition by Adrenergic α-Receptor Blockade

Following the intraarterial infusion of ^sH-adenine to isolated perfused canine subcutaneous adipose tissue, its adenine nucleotides are labelled. A continuous release of radioactivity, comprised of non-nucleotide material, was observed. The rate of this release was markedly enhanced by sympathetic nerve stimulation. The major components of the enhanced release appeared to be inosine and adenosine. Adrenergic α-receptor blockade (phentolamine or Hydergin®) abolished the enhanced nucleoside release, while glycerol release was enhanced. The release of radioactivity was decreased during mechanical blood flow reduction and enhanced afterwards. However, the magnitude of this enhancement of release after clamp was much less than following nerve stimulation. The results suggest that adenosine or a closely related compound is released from canine subcutaneous adipose tissue by sympathetic nerve stimulation and that the release is related to adrenergic α-receptor stimulation. Since adenosine is a potent inhibitor of catecholamine induced lipolysis in this tissue the possibility of a regulatory role must be considered.     

Attenuated hypothalamo-pituitary-adrenal axis responses to immune challenge during pregnancy: the neurosteroid–opioid connection

In late pregnancy maternal hypothalamo-pituitary-adrenal (HPA) axis responses to emotional and physical stressors are attenuated. This is expected to minimize the detrimental programming effects of glucocorticoid exposure on the fetuses. We have utilized a model of immune challenge, systemic administration of interleukin-1β (IL-1β), to investigate the underlying mechanisms. Intravenous IL-1β activates corticotropin-releasing hormone (CRH) neurones in the parvocellular division of the paraventricular nucleus (pPVN) via noradrenergic (A2 cell group) neurones in the nucleus tractus solitarii (NTS). Despite comparable activation of these brainstem neurones by IL-1β in virgin and in late pregnant rats, pPVN CRH neurones are activated only in virgin rats. As a consequence IL-1β fails to evoke ACTH and corticosterone secretion in late pregnant rats, in contrast to virgin rats. Suppressed responsiveness of the CRH neurones, and hence the HPA axis, following IL-1β in late pregnancy is explained by presynaptic inhibition of noradrenaline release in the pPVN, due to increased endogenous enkephalin and μ-opioid receptor production in brainstem NTS neurones. The factor that signals to the brain the pregnancy status of the animal and stimulates opioid production in the brainstem is allopregnanolone, a neurosteroid metabolite of progesterone. The supporting evidence for these mechanisms is discussed.     

Modification of the reflex response to stimulation of carotid sinus baroreceptors during and following stimulation of the hypothalamic defence area in the cat

1. The effects of stimulation of the hypothalamic defence area on carotid sinus baroreceptor reflexes have been investigated by examining the cardiovascular responses to a 15 sec period of increased pressure within the vascularly isolated carotid sinus before, during, and immediately following a 25 sec period of hypothalamic stimulation.2. Identification of the hypothalamic defence area was based on the occurence of atropine-sensitive muscle vasodilatation. Electrode positions were confirmed by histological examination.3. During hypothalamic stimulation the reflex fall in blood pressure resulting from a rise in sinus pressure was found to be undiminished whether sinus pressure was raised at the onset or at the 10th sec of hypothalamic stimulation.4. By contrast, in at least half the cats in which a reflex bradycardia could be evoked from the sinus, this bradycardia was largely if not completely suppressed during hypothalamic stimulation. This suppression of reflex bradycardia occurred when sinus pressure was raised at the onset as well as at the 10th sec of stimulation.5. During the first 5 sec after hypothalamic stimulation the hypotensive response to an increase in carotid sinus pressure was much reduced; on the other hand the reduction in heart rate was exaggerated, sometimes to a very marked degree.6. The results suggest that stimulation of the hypothalamic defence area can modify baroreceptor reflexes and that this modification can include selective alterations in the various components of the reflex response.     

Effects of hypothalamic stimulation on activity of dorsomedial medulla neurons that respond to subdiaphragmatic vagal stimulation

1. Effects of hypothalamic stimulation on activity of dorsomedial medulla neurons that responded to subdiaphragmatic vagal stimulation were investigated in urethan-anesthetized rats. 2. Extracellular recordings were made from 231 neurons in the nucleus of the tractus solitarius (NTS) that fired repetitively in response to single-pulse subdiaphragmatic vagal stimulation and from 320 neurons in the dorsal motor nucleus of the vagal nerve (DMV) that responded antidromically to subdiaphragmatic vagal stimulation. The mean latencies of responses to subdiaphragmatic vagal stimulation were 90.3 +/- 17.1 ms (mean +/- SD) for NTS neurons, and 90.8 +/- 11.2 ms for DMV neurons. This indicated that both afferent and efferent subdiaphragmatic vagal fibers were thin and unmyelinated and had a conduction velocity of approximately 1 m/s. 3. In extracellular recordings from 320 DMV neurons, marked inhibition preceded the antidromic response and subdiaphragmatic vagal stimulation evoked orthodromic spikes in only a few neurons. 4. Intracellular recordings from 66 DMV neurons revealed inhibitory postsynaptic potentials (IPSPs) before the antidromic responses. These IPSPs suppressed spontaneous firing and prevented excitatory postsynaptic potentials (EPSPs) from generating action potentials. 5. Stimulation in all hypothalamic loci studied, the ventromedial hypothalamic nucleus (VMH), the lateral hypothalamic area (LHA), and the paraventricular nucleus (PVN), induced responses with similar characteristics of excitation alone or excitation followed by inhibition in most NTS and DMV neurons. 6. No reciprocal effect of VMH and LHA stimulation was observed on NTS and DMV neurons. 7. Intracellular recordings from DMV neurons revealed monosynaptic EPSPs in response to stimulation of the VMH, the LHA, and the PVN. 8. PVN stimulation evoked significantly more responses in NTS and DMV neurons than VMH stimulation and more responses in DMV neurons than LHA stimulation. This suggests a difference in the number of connections between each hypothalamic site and the dorsomedial medulla. 9. The same dorsomedial medulla neurons were tested with VMH and LHA stimulation. The respective mean latencies of the antidromic and the orthodromic NTS neuron responses were 37.3 +/- 3.2 and 39.6 +/- 12.9 ms for VMH stimulation and 29.8 +/- 5.3 and 31.8 +/- 8.7 ms for LHA stimulation. The mean latencies of the orthodromic DMV neuron responses were 39.4 +/- 8.3 ms for VMH stimulation and 31.1 +/- 5.2 ms for LHA stimulation. The estimated conduction velocity from the VMH to the dorsomedial medulla was approximately 0.25 m/s and from the LHA it was approximately 0.33 m/s, which was significantly faster.(ABSTRACT TRUNCATED AT 400 WORDS)     

迷走神经刺激术对脑干孤束核部位GABA含量的影响

目的:探讨迷走神经刺激术(VNS)抗癫痫作用的机制是否为VNS引起孤束核部位GABA含量变化。方法:通过Waters高效液相色谱系统分析32只接受不同持续时间迷走神经刺激的大鼠脑干孤束核部位主要兴奋及抑制性氨基酸含量的变化。结果:1小时持续刺激组孤束核部位GABA含量较对照组升高且差异有显著意义。结论:迷走神经刺激术可能通过升高孤束核部位的GABA含量起到抗癫痫作用。     

Sensory Afferent Selective Role of P2 Receptors in the Nucleus Tractus Solitarii for Mediating the Cardiac Component of the Peripheral Chemoreceptor Reflex in Rats

We have assessed the functional role of type 2 purinergic (P2) receptors within the caudal aspect of the commissural nucleus tractus solitarii (NTS) in mediating the peripheral chemoreceptor reflex cardiorespiratory response in the arterially perfused in situ working heart-brainstem preparation of rats. Microinjection in NTS of either suramin (100 pmol) or pyrinoxalphosphate-6-azophenyl-2',4'-disulphonic acid tetrasodium salt (PPADS; 10 pmol) depressed the reflex bradycardia (by ≈50 %), but not the tachypnoea, following peripheral chemoreceptor stimulation. In contrast, the reflex bradycardia produced by stimulation of pharyngo-oesophageal receptors was unaffected. Furthermore, microinjections in NTS of the P2X receptor agonist α,β-methyleneadenosine 5'-triphosphate (10 pmol) evoked a bradycardia which was antagonized by suramin (100 pmol). This P2X agonist reversibly potentiated the peripheral chemoreceptor-evoked bradycardia. The effect of suramin was selective to purinergic receptors because the bradycardia evoked by microinjection of α,β-methyleneadenosine 5'-triphosphate was blocked while the bradycardic responses to microinjections of NMDA or non-NMDA receptor agonists were not affected. From whole-cell recordings, some NTS neurones received convergent excitatory synaptic inputs from both peripheral chemoreceptors and receptors at the pharyngo-oesophageal junction. The excitatory postsynaptic response evoked by chemoreceptor stimulation was depressed by suramin, but convergent excitatory inputs from pharyngo-oesophageal receptors were unperturbed. Our findings support the hypothesis that caudal commissural NTS P2 purinergic receptors play a role in the neurotransmission of the parasympathetic (bradycardic) component of the chemoreceptor reflex. This effect is highly selective in that the chemoreceptor afferent-evoked tachypnoea, as well as other visceral receptor-mediated reflex bradycardia, remain unaffected.     

An L-dopaergic relay from the posterior hypothalamic nucleus to the rostral ventrolateral medulla and its cardiovascular function in anesthetized rats.

We have proposed that L-3,4-dihydroxyphenylalanine (L-DOPA) is a neurotransmitter in the central nervous system [Misu Y. et al. (1996) Prog. Neurobiol. 49, 415-454]. Herein, we attempt to clarify whether lesions in the posterior hypothalamic nucleus decrease the tissue content of L-DOPA in the rostral ventrolateral medulla. We also attempt to clarify whether or not endogenous L-DOPA is evoked by electrical stimulation of the posterior hypothalamic nucleus. It is possible that evoked L-DOPA functions as a transmitter candidate to activate pressor sites of the rostral ventrolateral medulla in anesthetized rats. Electrolytic lesions were made in the bilateral posterior hypothalamic nucleus by a monopolar direct current of 2 mA for 10 s, 10 days before measurements. The effect of the lesions was to selectively decrease the tissue content of L-DOPA by one-half in the right rostral ventrolateral medulla. Decreases in the amounts of dopamine, noradrenaline or adrenaline were not observed. Decreases were also not evident in the right caudal ventrolateral medulla. During microdialysis of the right rostral ventrolateral medulla, extracellular basal levels of L-DOPA and three types of catecholamine were consistently detectable by high-performance liquid chromatography with electrochemical detection. Tetrodotoxin (1 microM) perfused into the right rostral ventrolateral medulla gradually decreased basal levels of L-DOPA by 25%; it decreased basal levels of noradrenaline and adrenaline by 25-30% and dopamine levels by 40%. Intensive electrical stimulation of the ipsilateral posterior hypothalamic nucleus (50 Hz, 0.3 mA, 0.1 ms duration, twice for 5 min at an interval of 5 min) selectively caused the release of L-DOPA in a repetitive and constant manner. The stimulation was accompanied by hypertension and tachycardia. However, catecholamines were not released. Tetrodotoxin suppressed the release of L-DOPA, but partially inhibited hypertension with only a slight inhibition of tachycardia evoked by stimulation of the posterior hypothalamic nucleus. L-DOPA methyl ester, a competitive L-DOPA antagonist, was bilaterally microinjected into pressor sites of the rostral ventrolateral medulla at 1.5 microg x 2 and 3 microg x 2. The antagonist dose-dependently and consistently antagonized pressor and tachycardiac responses to mild transient stimulation of the unilateral posterior hypothalamic nucleus (33 Hz, 0.2 mA, 0.1 ms duration, for 10 s). In addition, the antagonist alone (3 microg x 2) elicited hypotension and bradycardia. These results show that an L-DOPAergic relay may project from the posterior hypothalamic nucleus directly to pressor sites of the rostral ventrolateral medulla and/or indirectly to certain neurons near pressor sites in microcircuits of the same region. When released, L-DOPA appears to function tonically to activate pressor sites; it also appears to be involved in the maintenance and regulation of blood pressure and heart rate.     

Release of 3H-nucleosides from 3H-adenine labelled hypothalamic synaptosomes

[³H]adenine was taken up by a crude hypothalamic synaptosomal fraction and incorporated into mainly nucleotides. The synaptosomes were superfiTsed and after the initial washout a steady fractional release rate of 0.5-1 % of the content/min was found. Electrical pulses (2 ms, 50 Hz, 10–20 mA, 4 min) and veratridine (10 μM, 4 min) induced a Ca⁺⁺-dependent increase in purine release rate. K⁺ (30 mM, 4 min) caused a largely Ca⁺⁺independent increase. Most of the released material co-chromatographed with adenosine, inosine and hypoxanthine, while little or no nucleotide material was detected. Release of endogenous adenosine, inosine and hypoxanthine was detected by high performance liquid chromatography. However, following hypo-osmotic shock most of the released material was in nucleotides. The removal of glucose from the medium increased the fractional release rate 2–3 fold. Histamine, acetylcholine and glutamate were without effect. High amounts of noradrenaline caused an EGTA-inhibited release of purines, which was un-af-fected by propranolol or phentolamine. It is suggested that purines are released from neuronal structures and that the release reflects increased energy consumption and/or decreased energy production.     

Localization and action of adenosine A2a receptors in regions of the brainstem important in cardiovascular control

In vitro autoradiography and central microinjections of a P1 adenosine A2a receptor antagonist have been employed to investigate a possible role for centrally located adenosine A2a receptors in modulation of the baroreceptor reflex. In vitro autoradiography using [125I]4-(2-[7-amino-2-[2-furyl][3,2,4]triazolol[2,3-a][1,3,5]tr iazin-5-yl-amino]ethyl)phenol ([125I]ZM241385), the high-affinity adenosine A2a receptor antagonist, revealed a heterogeneous distribution of adenosine A2a binding sites within the lower brainstem of the rat. Image analysis showed high levels of binding in rostral regions of both the nucleus tractus solitarius and the ventrolateral medulla. Intermediate levels of binding were observed in the commissural nucleus tractus solitarius and the dorsal vagal motor nucleus, with low levels of binding in caudal regions of the nucleus tractus solitarius and the ventrolateral medulla, and the hypoglossal nucleus. Unilateral microinjections of unlabelled ZM241385 into the nucleus tractus solitarius had no effect on baseline levels of arterial pressure, heart rate and phrenic nerve activity recorded in anaesthetized, artificially ventilated rats. However, microinjections of ZM241385 reduced the bradycardia evoked by stimulation of the ipsilateral aortic nerve. In contrast, ZM241385 had no effect on the depressor response or the reduction in phrenic nerve activity evoked by aortic nerve stimulation. Our results indicate that adenosine A2a binding sites are located in a number of brainstem regions involved in autonomic function, consistent with the idea that adenosine acts as a neuromodulator of a variety of cardiorespiratory reflexes. Specifically, the data support the hypothesis that adenosine A2a receptors located within the nucleus tractus solitarius are activated during baroreceptor stimulation and have an important modulatory role in the pattern of cardiovascular changes associated with this reflex.     

Central nesfatin-1-expressing neurons are sensitive to peripheral inflammatory stimulus

Recently, a novel factor with anorexigenic properties was identified and called nesfatin-1. This protein (82 aac) is not only expressed in peripheral organs but it is also found in neurons located in specific structures including the hypothalamus and the brainstem, two sites strongly involved in food intake regulation. Here, we studied whether some of the neurons that become activated following an injection of an anorectic dose of lipopolysaccharides (LPS) exhibit a nesfatin-1 phenotype. To this end, we used double immunohistochemistry to target the expression of the immediate-early gene c-fos and of nesfatin-1 on coronal frozen sections of the rat brain. The number of c-Fos+/nesfatin-1+ neurons was evaluated in the immunosensitive structures reported to contain nesfatin-1 neurons; i.e. paraventricular hypothalamic nucleus (PVN), supraoptic nucleus (SON), arcuate nucleus (ARC) and nucleus of the solitary tract (NTS). LPS strongly increased the number of c-Fos+/nesfatin-1+ neurons in the PVN, SON and NTS, and to a lesser extent in the ARC. Triple labeling showed that a portion of the nesfatin-1 neurons activated in response to LPS within the NTS are catecholaminergic since they co-express tyrosine hydroxylase (TH). Our data therefore indicate that a portion of nesfatin-1 neurons of both the hypothalamus and brainstem are sensitive to peripheral inflammatory signals, and provide the first clues suggesting that centrally released nesfatin-1 may contribute to the neural mechanisms leading to endotoxaemic anorexia.     

Brainstem mediation of prefrontal stimulus-produced hypotension

Summary Electrical stimulation of the prefrontal cortex (PFC) in rats produces transient hypotension. It has been suggested that this stimulus-produced hypotension (SPH) may be mediated by direct PFC projections to either the posterolateral hypothalamus or the nucleus tractus solitarius (NTS). To initially test these hypotheses, microinjections (0.5 l) of 4% lidocaine were made into various brainstem sites, including the posterolateral hypothalamus, the PFC-NTS pathway and the NTS itself. Most injections made into the posterolateral hypothalamus, rostral portion of the PFC-NTS pathway or NTS were successful in blocking prefrontal SPH. In comparison, the majority of injections made into numerous other brainstem sites including the caudal portion of the PFC-NTS pathway did not block prefrontal SPH. These findings support the concept that prefrontal SPH maybe mediated via both the posterolateral hypothalamus and NTS. However, these findings do not support the hypothesis that prefrontal SPH is mediated by direct projections from the PFC to the NTS. To eliminate the possibility that the effects of the lidocaine injections made into the hypothalamus were due to the incapacitation of fibers alone, a series of experiments was conducted in which microinjections (0.5 l) of ibotenic acid, a neurotoxin that destroys perikarya but spares axons, were made into the posterolateral hypothalamus. Each of these injections resulted in the blockade of prefrontal SPH. These findings further support the role of hypothalamic involvement in prefrontal SPH.     

Cholinergic input to the supraoptic nucleus increases Fos expression and body temperature in rats

To examine the role played by cholinergic input and processes in the supraoptic nucleus (SON) in the control of body temperature and water intake in rats, we used microdialysis to stimulate and analyze SON without disturbing the behavior of unanesthetized rats. After microdialysis, we also investigated immunoreactivity for c-Fos protein in the brain as an index of neuronal activation. Stimulation with neostigmine, an acetylcholine esterase inhibitor, through the microdialysis probe increased the extracellular concentration of acetylcholine in the SON. This cholinergic stimulation dose-dependently increased body temperature but did not significantly change the water intake. The stimulation markedly increased c-Fos-like immunoreactivity (Fos-IR) in the SON and certain hypothalamic areas, including the paraventricular nucleus (PVN) and median preoptic nucleus (MnPO). Fos-IR was also evident in certain regions of the pons and brainstem, including the locus ceruleus (LC), area postrema (AP), and nucleus of the solitary tract (NTS). Addition of atropine, a muscarinic receptor antagonist, to the dialysis medium containing neostigmine attenuated the increase of Fos-IR and suppressed the neostigmine-induced responses in body temperature. These results suggest that cholinergic input and activation of the muscarinic cholinoceptive neurons in the SON contribute to the regulation of body temperature. Activation of noradrenergic pathways in the brainstem including LC and NTS may be involved in the thermoregulation mechanism.     

大白鼠前脑、脑干和小脑向孤束核的纤维投射——HRP法和ARG法研究

本实验用HRP法、WGA—HRP法和ARG法进一步研究了大白鼠前脑、脑干和小脑向孤束核的纤维投射及其局部定位关系。作者发现皮质32及8区、三叉神经脊束核尾侧亚核投射到孤束核头段;皮质24及8a区、三叉神经脊束核尾侧亚核投射到孤束核中段;皮质23及13区,中脑导水管周围灰质和三叉神经脊束核尾侧亚核投射到孤束核尾段。此外,结果还证明室旁核、下丘脑外侧区、兰斑核、K—F氏核、小脑顶核、中缝大核、网状大细胞核、外侧网状核、及延髓网状结构向孤束核的纤维投射,均存在着一定的局部定位关系。本文结合前脑、脑干向孤束核的纤维投射讨论了内脏活动调节及针刺镇痛效应的中枢机理并对大白鼠孤束核的分段(头、中及尾段)标准提出了新建议。     

Activation within dorsal medullary nuclei following stimulation in the hypothalamic paraventricular nucleus in rats

The influence of the hypothalamic paraventricular nucleus (PVN) on neurones in the dorsal medulla has been examined in 71 urethane/sagatal-anaesthetised rats. Of 536 neurones localised and tested for responses to electrical stimulation of both the vagus and/or the PVN, 378 were synaptically or antidromically activated following vagal stimulation 72 of which were synaptically activated by stimulation within PVN. The majority of those were located at the border between NTS and dorsal motor nucleus of the vagus in caudal NTS. None showed cardiac or ventilatory rhythm. Neurones showing such rhythms were not affected from PVN. Of 89 neurones in dorsal motor nucleus of the vagus, ten were synaptically activated and two synaptically depressed from PVN. PVN activated neurones in NTS tested for responses to stimulation of arterial baroreceptors and carotid body chemoreceptors were either unaffected or inhibited, but gastric inflation excited them. The results suggest a powerful PVN influence on the dorsal medulla, which is largely confined to the ventral and caudal NTS. There is little evidence for an effect on neurones with a cardiovascular function, but the abdominal vagal influence suggests a link with feeding.     

Localization and function of the Kv3.1b subunit in the rat medulla oblongata: focus on the nucleus tractus solitarii

The voltage-gated potassium channel subunit Kv3.1 confers fast firing characteristics to neurones. Kv3.1b subunit immunoreactivity (Kv3.1b-IR) was widespread throughout the medulla oblongata, with labelled neurones in the gracile, cuneate and spinal trigeminal nuclei. In the nucleus of the solitary tract (NTS), Kv3.1b-IR neurones were predominantly located close to the tractus solitarius (TS) and could be GABAergic or glutamatergic. Ultrastructurally, Kv3.1b-IR was detected in NTS terminals, some of which were vagal afferents. Whole-cell current-clamp recordings from neurones near the TS revealed electrophysiological characteristics consistent with the presence of Kv3.1b subunits: short duration action potentials (4.2 ± 1.4 ms) and high firing frequencies (68.9 ± 5.3 Hz), both sensitive to application of TEA (0.5 m m ) and 4-aminopyridine (4-AP; 30 μ m ). Intracellular dialysis of an anti-Kv3.1b antibody mimicked and occluded the effects of TEA and 4-AP in NTS and dorsal column nuclei neurones, but not in dorsal vagal nucleus or cerebellar Purkinje cells (which express other Kv3 subunits, but not Kv3.1b). Voltage-clamp recordings from outside-out patches from NTS neurones revealed an outward K⁺ current with the basic characteristics of that carried by Kv3 channels. In NTS neurones, electrical stimulation of the TS evoked EPSPs and IPSPs, and TEA and 4-AP increased the average amplitude and decreased the paired pulse ratio, consistent with a presynaptic site of action. Synaptic inputs evoked by stimulation of a region lacking Kv3.1b-IR neurones were not affected, correlating the presence of Kv3.1b in the TS with the pharmacological effects.     

Vagal afferent control of opioidergic effects in rat brainstem circuits

We demonstrated recently that increasing the levels of cAMP allows opioids to modulate GABAergic synaptic transmission between the nucleus of the tractus solitarius (NTS) and dorsal motor nucleus of the vagus (DMV). Using a combination of electrophysiological, immunohistochemical and biochemical approaches, we provide evidence that vagal afferent fibres dampen cAMP levels within the vagal brainstem circuits via tonic activation of group II metabotropic glutamate receptors (mGluRs). Whole-cell patch-clamp recordings were made from identified neurons of the rat DMV. Following chronic vagal deafferentation, the opioid agonist methionine-enkephalin (ME) inhibited the amplitude of evoked IPSC (eIPSC) in 32 of 33 neurons, without exogenous enhancement of cAMP levels. The ME-induced inhibition was prevented by the group II mGluR-selective agonist APDC. Following perfusion with the group II mGluR-selective antagonist EGLU, ME inhibited eIPSC amplitude in brainstem slices of control rats. Immunohistochemical experiments revealed that, following vagal deafferentation, μ-opioid receptors were colocalized on GABAergic profiles apposing DMV neurons; the number of colocalized profiles was significantly decreased by pretreatment with APDC. Radioimmunoassay and Western blot analysis showed that cAMP and phosphorylated cyclic AMP response element binding protein (pCREB) levels in the dorsal vagal complex were increased following vagal deafferentation. Our data show that by tonically dampening the levels of cAMP within the GABAergic synaptic contacts, activated group II mGluRs prevent the modulation of this synapse by endogenous opioids. These data suggest that the plasticity, hence the response, of central circuits controlling the vagal motor outflow to visceral organs is modulated and finely tuned by vagal afferent fibres.     

Substance P presynaptically depresses the transmission of sensory input to bronchopulmonary neurons in the guinea pig nucleus tractus solitarii

Substance P modulates the reflex regulation of respiratory function by its actions both peripherally and in the CNS, particularly in the nucleus tractus solitarii (NTS), the first central site for synaptic contact of the lung and airway afferent fibres. There is considerable evidence that the actions of substance P in the NTS augment respiratory reflex output, but the precise effects on synaptic transmission have not yet been determined. Therefore, we determined the effects of substance P on synaptic transmission at the first central synapses by using whole-cell voltage clamping in an NTS slice preparation. Studies were performed on second-order neurons in the slice anatomically identified as receiving monosynaptic input from sensory nerves in the lungs and airways. This was done by the fluorescent labelling of terminal boutons after 1,1'-dioctadecyl-3,3,3',3'-tetra-methylindocarbo-cyanine perchlorate (DiI) was applied via tracheal instillation. Substance P (1.0, 0.3 and 0.1 μM) significantly decreased the amplitude of excitatory postsynaptic currents (eEPSCs) evoked by stimulation of the tractus solitarius, in a concentration-dependent manner. The decrease was accompanied by an increase in the paired-pulse ratio of two consecutive eEPSCs, and a decrease in the frequency, but not the amplitude, of spontaneous EPSCs and miniature EPSCs, findings consistent with a presynaptic site of action. The effects were consistently and significantly attenuated by a neurokinin-1 (NK1) receptor antagonist (SR140333, 3 μM). The data suggest a new site of action for substance P in the NTS (NK1 receptors on the central terminals of sensory fibres) and a new mechanism (depression of synaptic transmission) for regulating respiratory reflex function.