Distribution of glycine/GABA neurons in the ventromedial medulla with descending spinal projections and evidence for an ascending glycine/GABA projection

The ventromedial medulla (VM), subdivided in a rostral (RVM) and a caudal (CVM) part, has a powerful influence on the spinal cord. In this study, we have identified the distribution of glycine and GABA containing neurons in the VM with projections to the cervical spinal cord, the lumbar dorsal horn, and the lumbar ventral horn. For this purpose, we have combined retrograde tracing using fluorescent microspheres with fluorescent in situ hybridization (FISH) for glycine transporter 2 (GlyT2) and GAD67 mRNAs to identify glycinergic and/or GABAergic (Gly/GABA) neurons. Since the results obtained with FISH for GlyT2, GAD67, or GlyT2 + GAD67 mRNAs were not significantly different, we concluded that glycine and GABA coexisted in the various projection neurons. After injections in the cervical cord, we found that 29% ± 1 (SEM) of the retrogradely labeled neurons in the VM were Gly/GABA (RVM: 43%; CVM: 21%). After lumbar dorsal horn injections 31% ± 3 of the VM neurons were Gly/GABA (RVM: 45%; CVM: 12%), and after lumbar ventral horn injections 25% ± 2 were Gly/GABA (RVM: 35%; CVM: 17%). In addition, we have identified a novel ascending Gly/GABA pathway originating from neurons in the area around the central canal (CC) throughout the spinal cord and projecting to the RVM, emphasizing the interaction between the ventromedial medulla and the spinal cord. The present study has now firmly established that GABA and glycine are present in many VM neurons that project to the spinal cord. These neurons strongly influence spinal processing, most notably the inhibition of nociceptive transmission.     

Disruption of KCC2 reveals an essential role of K-Cl cotransport already in early synaptic inhibition

Synaptic inhibition by GABA(A) and glycine receptors, which are ligand-gated anion channels, depends on the electrochemical potential for chloride. Several potassium-chloride cotransporters can lower the intracellular chloride concentration [Cl(-)](i), including the neuronal isoform KCC2. We show that KCC2 knockout mice died immediately after birth due to severe motor deficits that also abolished respiration. Sciatic nerve recordings revealed abnormal spontaneous electrical activity and altered spinal cord responses to peripheral electrical stimuli. In the spinal cord of wild-type animals, the KCC2 protein was found at inhibitory synapses. Patch-clamp measurements of embryonic day 18.5 spinal cord motoneurons demonstrated an excitatory GABA and glycine action in the absence, but not in the presence, of KCC2, revealing a crucial role of KCC2 for synaptic inhibition.     

Functional modifications of acid-sensing ion channels by ligand-gated chloride channels

Together, acid-sensing ion channels (ASICs) and epithelial sodium channels (ENaC) constitute the majority of voltage-independent sodium channels in mammals. ENaC is regulated by a chloride channel, the cystic fibrosis transmembrane conductance regulator (CFTR). Here we show that ASICs were reversibly inhibited by activation of GABA(A) receptors in murine hippocampal neurons. This inhibition of ASICs required opening of the chloride channels but occurred with both outward and inward GABA(A) receptor-mediated currents. Moreover, activation of the GABA(A) receptors modified the pharmacological features and kinetic properties of the ASIC currents, including the time course of activation, desensitization and deactivation. Modification of ASICs by open GABA(A) receptors was also observed in both nucleated patches and outside-out patches excised from hippocampal neurons. Interestingly, ASICs and GABA(A) receptors interacted to regulate synaptic plasticity in CA1 hippocampal slices. The activation of glycine receptors, which are similar to GABA(A) receptors, also modified ASICs in spinal neurons. We conclude that GABA(A) receptors and glycine receptors modify ASICs in neurons through mechanisms that require the opening of chloride channels.     

Strychnine antagonizes vaginal stimulation-produced analgesia at the spinal cord

Vaginal-cervical mechanostimulation (VS) suppresses vocalization and withdrawal responses to noxious stimulation. To determine whether the inhibitory neurotransmitter, glycine, contributes to the action of VS, strychnine, a specific glycine receptor antagonist was administered perispinally via intrathecal catheter in dosages of 1,5,25 and 100 micrograms. Prior to strychnine administration, VS (400 g force) elevated thresholds to elicit vocalization in response to graded intensities of tail shock, and blocked vocalization elicited by stimulation of a skin area, previously sensitized by intradermal injection of a 20% yeast solution. After strychnine administration the analgesic effects of VS were significantly attenuated. These findings suggest that the analgesic action of VS is partially mediated by glycine at the spinal level.     

Glycine is taken up through GLYT1 and GLYT2 transporters into mouse spinal cord axon terminals and causes vesicular and carrier-mediated release of its proposed co-transmitter GABA

Glycine and GABA are likely co-transmitters in the spinal cord. Their possible interactions in presynaptic terminals have, however, not been investigated. We studied the effects of glycine on GABA release using superfused mouse spinal cord synaptosomes. Glycine concentration dependently elicited [(3)H]GABA release which was insensitive to strychnine or 5,7-dichlorokynurenic acid, but was Na(+) dependent and sensitive to the glycine uptake blocker glycyldodecylamide. The glycine effect was external Ca(2+) independent, but was reduced when intraterminal Ca(2+) was chelated with 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetracetic acid or depleted with thapsigargin, or when vesicular storage was impaired with bafilomycin. Glycine-induced [(3)H]GABA release was prevented, in part, by blocking GABA transport. The glycine effect was halved by sarcosine, a GLYT1 substrate/inhibitor, or by amoxapine, a GLYT2 blocker, and abolished by a mixture of the two. The sensitivity to sarcosine, used as a transporter inhibitor or substrate, persisted in synaptosomes prelabelled with [(3)H]GABA in the presence of beta-alanine, excluding major gliasome involvement. To conclude, in mice spinal cord, transporters for glycine (both GLYT1 and GLYT2) and for GABA coexist on the same axon terminals. Activation of the glycine transporters elicits GABA release, partly by internal Ca(2+)-dependent exocytosis and partly by transporter reversal.     

Pain and itch: insights into the neural circuits of aversive somatosensation in health and disease

Although pain and itch are distinct sensations, most noxious chemicals are not very specific to one sensation over the other, and recent discoveries are revealing that Trp channels function as transducers for both. A key difference between these sensations is that itch is initiated by irritation of the skin, whereas pain can be elicited from almost anywhere in the body; thus, itch may be encoded by the selective activation of specific subsets of neurons that are tuned to detect harmful stimuli at the surface and have specialized central connectivity that is specific to itch. Within the spinal cord, cross-modal inhibition between pain and itch may help sharpen the distinction between these sensations. Moreover, this idea that somatosensory modalities inhibit one another may be generalizable to other somatosensory subtypes, such as cold and hot. Importantly, just as there are inhibitory circuits in the dorsal horn that mediate cross-inhibition between modalities, it appears that there are also excitatory connections that can be unmasked upon injury or in disease, leading to abnormally elevated pain states such as allodynia. We are now beginning to understand some of this dorsal horn circuitry, and these discoveries are proving to be relevant for pathological conditions of chronic pain and itch.     

Antisera to gamma-aminobutyric acid. II. Immunocytochemical application to the central nervous system

An antiserum to gamma-aminobutyric acid (GABA) was tested for the localization of GABAergic neurons in the central nervous system using the unlabeled antibody enzyme method under pre- and postembedding conditions. GABA immunostaining was compared with glutamate decarboxylase (GAD) immunoreactivity in the cerebellar cortex and in normal and colchicine-injected neocortex and hippocampus of cat. The types, distribution, and proportion of neurons and nerve terminals stained with either sera showed good agreement in all areas. Colchicine treatment had little effect on the density of GABA-immunoreactive cells but increased the number of GAD-positive cells to the level of GABA-positive neurons in normal tissue. GABA immunoreactivity was abolished by solid phase adsorption to GABA and it was attenuated by adsorption to beta-alanine or gamma-amino-beta-hydroxybutyric acid, but without selective loss of immunostaining. Reactivity was not affected by adsorption to glutamate, aspartate, taurine, glycine, cholecystokinin, or bovine serum albumin. The concentration (0.05-2.5%) of glutaraldehyde in the fixative was not critical. The antiserum allows the demonstration of immunoreactive GABA in neurons containing other neuroactive substances; cholecystokinin and GABA immunoreactivities have been shown in the same neurons of the hippocampus. In conclusion, antisera to GABA are good markers for the localization of GABAergic neuronal circuits.     

GABAB receptors in the NTS mediate the inhibitory effect of trigeminal nociceptive inputs on parasympathetic reflex vasodilation in the rat masseter muscle

The present study was designed to examine whether trigeminal nociceptive inputs are involved in the modulation of parasympathetic reflex vasodilation in the jaw muscles. This was accomplished by investigating the effects of noxious stimulation to the orofacial area with capsaicin, and by microinjecting GABA(A) and GABA(B) receptor agonists or antagonists into the nucleus of the solitary tract (NTS), on masseter hemodynamics in urethane-anesthetized rats. Electrical stimulation of the central cut end of the cervical vagus nerve (cVN) in sympathectomized animals bilaterally increased blood flow in the masseter muscle (MBF). Increases in MBF evoked by cVN stimulation were markedly reduced following injection of capsaicin into the anterior tongue in the distribution of the lingual nerve or lower lip, but not when injected into the skin of the dorsum of the foot. Intravenous administration of either phentolamine or propranolol had no effect on the inhibitory effects of capsaicin injection on the increases of MBF evoked by cVN stimulation, which were largely abolished by microinjecting the GABA(B) receptor agonist baclofen into the NTS. Microinjection of the GABA(B) receptor antagonist CGP-35348 into the NTS markedly attenuated the capsaicin-induced inhibition of MBF increase evoked by cVN stimulation, while microinjection of the GABA(A) receptor antagonist bicuculline did not. Our results indicate that trigeminal nociceptive inputs inhibit vagal-parasympathetic reflex vasodilation in the masseter muscle and suggest that the activation of GABA(B) rather than GABA(A) receptors underlies the observed inhibition in the NTS.     

Spinal GABA(A) receptors do not mediate the sympathetic baroreceptor reflex in the rat

Activation of baroreceptors causes efferent sympathetic nerve activity (SNA) to fall. Two mechanisms could account for this sympathoinhibition: disfacilitation of sympathetic preganglionic neurons (SPN) and/or direct inhibition of SPN. The roles that spinal GABA and glycine receptors play in the baroreceptor reflex were examined in anesthetized, paralyzed, and artificially ventilated rats. Spinal GABA(A) receptors were blocked by an intrathecal injection of bicuculline methiodide, whereas glycine receptors were blocked with strychnine. Baroreceptors were activated by stimulation of the aortic depressor nerve (ADN), and a somatosympathetic reflex was used as control. After an intrathecal injection of vehicle, there was no effect on any measured variable or evoked reflex. In contrast, bicuculline caused a dose-dependent increase in arterial pressure, SNA, phrenic nerve discharge, and it significantly facilitated the somatosympathetic reflex. However, bicuculline did not attenuate either the depressor response or sympathoinhibition evoked after ADN stimulation. Similarly, strychnine did not affect the baroreceptor-induced depressor response. Thus GABA(A) and glycine receptors in the spinal cord have no significant role in baroreceptor-mediated sympathoinhibition.     

Evidence that glycine and GABA mediate postsynaptic inhibition of bulbar respiratory neurons in the cat.

Experiments were carried out on decerebrate cats to identify transsynaptic mediators of spontaneous postsynaptic inhibition of bulbar inspiratory and postinspiratory neurons. Somatic membrane potentials were recorded through the central micropipette of a coaxial multibarreled electrode. Blockers of type A gamma-aminobutyric acid (GABA-A) and glycine receptors were iontophoresed extracellularly from peripheral micropipettes surrounding the central pipette. Effective antagonism was demonstrated by iontophoresis of agonists with antagonists; application of strychnine antagonized the action of glycine but not GABA, and application of bicuculline antagonized the action of GABA but not glycine. In both types of neurons, iontophoresis of either antagonist depolarized the somatic membrane and increased input resistance throughout the respiratory cycle. Bicuculline preferentially depolarized the somatic membrane in both types of neurons during inactive phases. Strychnine increased the firing rate of inspiratory neurons during inspiration despite maintenance of somatic membrane potential at preiontophoresis levels. Tetrodotoxin reduced the effects of iontophoresed bicuculline and strychnine, suggesting that the action of the antagonists required presynaptic axonal conduction. The present results suggest that presynaptic release of both GABA and glycine contributes to tonic postsynaptic inhibition of bulbar respiratory neurons. GABA-A receptors appear to contribute to inhibition during inactive phases in inspiratory and postinspiratory neurons, whereas glycinergic mechanisms appear to contribute to inspiratory inhibition in inspiratory neurons.     

Asymmetric cross-inhibition between GABAA and glycine receptors in rat spinal dorsal horn neurons

Presynaptic nerve terminals of inhibitory synapses in the dorsal horn of the spinal cord and brain stem can release both GABA and glycine, leading to coactivation of postsynaptic GABAA and glycine receptors. In the present study we have analyzed functional interactions between GABAA and glycine receptors in acutely dissociated neurons from rat sacral dorsal commissural nucleus. Although the application of GABA and glycine activates pharmacologically distinct receptors, the current induced by a simultaneous application of these two transmitters was less than the sum of currents induced by applying two transmitters separately. Sequential application of glycine and GABA revealed that the GABA-evoked current is more affected by glycine than glycine-evoked responses by GABA. Activation of glycine receptors decreased the amplitude and accelerated the rate of desensitization of GABA-induced currents. This asymmetric cross-inhibition is reversible, dependent on the agonist concentration applied, but independent of both membrane potential and intracellular calcium concentration or changes in the chloride equilibrium potential. During sequential applications, the asymmetric cross-inhibition was prevented by selective GABAA or glycine receptor antagonists, suggesting that occupation of binding sites did not suffice to induce glycine and GABAA receptors functional interaction, and receptor channel activation is required. Furthermore, inhibition of phosphatase 2B, but not phosphatase 1 or 2A, prevented GABAA receptor inhibition by glycine receptor activation, whereas inhibition of phosphorylation pathways rendered cross-talk irreversible. Taken together, our results demonstrated that there is an asymmetric cross-inhibition between glycine and GABAA receptors and that a selective modulation of the state of phosphorylation of GABAA receptor and/or mediator proteins underlies the asymmetry in the cross-inhibition.     

Plasticity of pre- and postsynaptic GABAB receptor function in the paraventricular nucleus in spontaneously hypertensive rats

GABA(B) receptor function is upregulated in the paraventricular nucleus (PVN) of the hypothalamus in spontaneously hypertensive rats (SHR), but it is unclear whether this upregulation occurs pre- or postsynaptically. We therefore determined pre- and postsynaptic GABA(B) receptor function in retrogradely labeled spinally projecting PVN neurons using whole cell patch-clamp recording in brain slices in SHR and Wistar-Kyoto (WKY) rats. Bath application of the GABA(B) receptor agonist baclofen significantly decreased the spontaneous firing activity of labeled PVN neurons in both SHR and WKY rats. However, the magnitude of reduction in the firing rate was significantly greater in SHR than in WKY rats. Furthermore, baclofen produced larger membrane hyperpolarization and outward currents in labeled PVN neurons in SHR than in WKY rats. The baclofen-induced current was abolished by either including G protein inhibitor GDPbetaS in the pipette solution or bath application of the GABA(B) receptor antagonist in both SHR and WKY rats. Blocking N-methyl-d-aspartic acid receptors had no significant effect on baclofen-elicited outward currents in SHR. In addition, baclofen caused significantly greater inhibition of glutamatergic excitatory postsynaptic currents (EPSCs) in labeled PVN neurons in brain slices from SHR than WKY rats. By contrast, baclofen produced significantly less inhibition of GABAergic inhibitory postsynaptic currents (IPSCs) in labeled PVN neurons in SHR than in WKY rats. Although microinjection of the GABA(B) antagonist into the PVN increases sympathetic vasomotor tone in SHR, the GABA(B) antagonist did not affect EPSCs and IPSCs of the PVN neurons in vitro. These findings suggest that postsynaptic GABA(B) receptor function is upregulated in PVN presympathetic neurons in SHR. Whereas presynaptic GABA(B) receptor control of glutamatergic synaptic inputs is enhanced, presynaptic GABA(B) receptor control of GABAergic inputs in the PVN is attenuated in SHR. Changes in both pre- and postsynaptic GABA(B) receptors in the PVN may contribute to the control of sympathetic outflow in hypertension.     

The interaction between glycine and GABA postsynaptic effects in the spinal cord neurons of frog Rana temporaria

Patch-clamp study in the whole multipolar cell (presumably motoneuron) was performed, the cells having been mechanically isolated from the spinal cord of the frog Rana ridibunda. It was shown that GABA and glycine, when applied simultaneously, produced a transmembrane current. Its amplitude was lower than the summed amplitude of currents produced in the same neuron by separate applications of GABA and glycine. The investigation of this occlusion showed that the superfusion of the neurons with solution containing 0.2 mM of glycine totally blocked the responses to GABA (5 mM) application, and vice versa. The crossinhibition can lie in the basis of this phenomenon. It could be due to either the existence of a common receptor complex sensitive to both GABA and glycine or to interaction between GABA and glycine receptors.     

Behavioral and Electrophysiological Evidence for the Differential Functions of TRPV1 at Early and Late Stages of Chronic Inflammatory Nociception in Rats

We previously reported that vanilloid receptor type 1 (VR1, or TRPV1) was up-regulated in dorsal root ganglion (DRG) and the spinal dorsal horn after chronic inflammatory pain produced by complete Freund’s adjuvant (CFA) injection into the plantar of rat hind paw. In the present study, we found that subcutaneous or intrathecal application of capsazepine (CPZ), a TRPV1 competitive antagonist, could inhibit thermal hyperalgesia on day 1 and on day 14 but not on day 28 after CFA injection. With extracellular electrophysiological recording, the effect of CPZ on noxious electrical or heat stimulation evoked responses of wide dynamic range (WDR) neurons in the deep layers of the spinal dorsal horn was evaluated. Under noxious electrical stimulation to sciatic nerve, CPZ applied to the spinal cord produced an inhibition on Aδ- and C-fiber evoked responses of WDR neurons on day 1 and 14, but not on day 28. Under radiant heat stimulation to the receptive field skin, subcutaneous application of CPZ significantly inhibited the background activity and extended the response latency of WDR neurons on day 14. These results provide new evidence for the functional significance of TRPV1 at the early stage, but not the late stage, in the rat model of CFA-induced inflammatory pain. Special issue article in honor of Dr. Ji-Sheng Han. Hao Luo, Isabella Shi Xu, Yi Chen are Co-first authors.     

Mechanisms of interactions between glycine- and GABA-mediated responses in spinal cord neurons

The interactions of GABA- and glycine-mediated responses have been analyzed, the whole cell patch-clamp method being used. The response induced by co-application of glycine and GABA was a lesser one than the sum of responses induced by applying two transmitters separately. The molecular mechanisms underlying this effect have been determined. Due to applications of high concentrations of neurotransmitters it was revealed that GABA could activate glycine receptors in frog spinal motoneurons with relatively high efficiency (EC50 = 1.2 mM). The sequential application of neurotransmitters showed that even a single application of glycine could significantly boost the "run-down" of the GABA-mediated current, suggesting that there was a strong phosphorylation-dependent mechanism of GABAa-receptors inhibition. These mechanisms are likely to take place in frog spinal motoneurons when GABA and glycine are co-released from the same synaptic terminal.     

GABA transporters mediate glycine release from cerebellum nerve endings: roles of Ca(2+)channels, mitochondrial Na(+)/Ca(2+) exchangers, vesicular GABA/glycine transporters and anion channels

GABA transporters accumulate GABA to inactivate or reutilize it. Transporter-mediated GABA release can also occur. Recent findings indicate that GABA transporters can perform additional functions. We investigated how activation of GABA transporters can mediate release of glycine. Nerve endings purified from mouse cerebellum were prelabeled with [(3)H]glycine in presence of the glycine GlyT1 transporter inhibitor NFPS to label selectively GlyT2-bearing terminals. GABA was added under superfusion conditions and the mechanisms of the GABA-evoked [(3)H]glycine release were characterized. GABA stimulated [(3)H]glycine release in a concentration-dependent manner (EC(50) = 8.26 μM). The GABA-evoked release was insensitive to GABA(A) and GABA(B) receptor antagonists, but it was abolished by GABA transporter inhibitors. About 25% of the evoked release was dependent on external Ca(2+) entering the nerve terminals through VSCCs sensitive to ω-conotoxins. The external Ca(2+)-independent release involved mitochondrial Ca(2+), as it was prevented by the Na(+)/Ca(2+) exchanger inhibitor CGP37157. The GABA uptake-mediated increases in cytosolic Ca(2+) did not trigger exocytotic release because the [(3)H]glycine efflux was insensitive to clostridial toxins. Bafilomycin inhibited the evoked release likely because it reduced vesicular storage of [(3)H]glycine so that less [(3)H]glycine can become cytosolic when GABA taken up exchanges with [(3)H]glycine at the vesicular inhibitory amino acid transporters shared by the two amino acids. The GABA-evoked [(3)H]glycine efflux could be prevented by niflumic acid or NPPB indicating that the evoked release occurred essentially by permeation through anion channels. In conclusion, GABA uptake into GlyT2-bearing cerebellar nerve endings triggered glycine release which occurred essentially by permeation through Ca(2+)-dependent anion channels. Glial GABA release mediated by anion channels was proposed to underlie tonic inhibition in the cerebellum; the present results suggest that glycine release by neuronal anion channels also might contribute to tonic inhibition.     

Microiontophoretically applied THIP effects upon nociceptive responses of neurons in medial thalamus

Sprague-Dawley rats anesthetized with urethane were used to study the single cell responses of medial thalamic neurons following noxious input and their interactions with gamma-aminobutyric acid (GABA) agonist THIP (4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol) and morphine sulfate applied microintophoretically . The majority of the medial thalamic neurons responded to noxious stimulation by an increase in their firing rate. Local application of both THIP and morphine attenuated the spontaneous and the noxious evoked responses of these neurons. The possibility of a role for GABA in mediating nonopiate pain suppression is discussed.     

Evidence for GABA-mediated control of putative nociceptive modulating neurons in the rostral ventromedial medulla: iontophoresis of bicuculline eliminates the off-cell pause.

This study was undertaken to test the hypothesis that gamma-aminobutyric acid (GABA) is an endogeneous neurotransmitter regulating the activity of a class of putative nociceptive modulatory neurons (termed "off-cells") in the rostral ventromedial medulla (RVM) of the barbiturate-anesthetized rat. Off-cells, which are believed to correspond to the RVM output neuron that inhibits nociceptive processing at the level of the spinal cord, exhibit an abrupt pause in firing that begins immediately prior to the occurrence of the tail flick response (TF), a nocifensive reflex evoked by application of noxious heat to the tail. Single-unit recording and iontophoretic techniques were used to examine the ability of the GABAA receptor antagonist bicuculline methiodide (BIC) to antagonize selectively the characteristic off-cell pause. Iontophoretic application of BIC (5-30 nA) blocked the TF-related pause in each of the off-cells tested. This effect of BIC was generally slow in onset, and outlasted the period of application by several minutes. BIC iontophoresis also eliminated the cyclic alternation between active and silent periods that is often displayed by off-cells in lightly anesthetized rats. BIC application did not have a consistent effect on the firing of two other classes of RVM neurons ("on-cells" and "neutral cells"). Iontophoretically applied BIC antagonized the inhibitory effect of iontophoretically applied GABA, but not that produced by glycine. The glycine receptor antagonist strychnine did not mimic the action of BIC on off-cell activity. These data demonstrate antagonism of a synaptically evoked response using iontophoretic application of BIC, and provide strong evidence that the inhibitory neurotransmitter GABA mediates the TF-related off-cell pause. Taken together with behavioral experiments demonstrating that a GABA-mediated inhibitory process within RVM is crucial in permitting execution of the TF response, the present observations point to the significant functional relevance of GABA transmission within RVM in modulation of nociception.     

Integration of cornea and cardiorespiratory afferents in the nucleus of the solitary tract of the rat

We determined the activity of neurons within the nucleus of the solitary tract (NTS) after stimulation of the cornea and assessed whether this input affected the processing of baroreceptor and peripheral chemoreceptor inputs. In an in situ, unanesthetized decerebrate working heart-brain stem preparation of the rat, noxious mechanical or electrical stimulation was applied to the cornea, and extracellular single unit recordings were made from NTS neurons. Cornea nociceptor stimulation evoked bradycardia and an increase in the cycle length of the phrenic nerve discharge. Of 90 NTS neurons with ongoing activity, corneal stimulation excited 51 and depressed 39. There was a high degree of convergence to these NTS neurons from either baroreceptors or chemoreceptors. The excitatory synaptic response in 12 of 19 baroreceptive and 10 of 15 chemoreceptive neurons was attenuated significantly during concomitant electrical stimulation of the cornea. This inhibition was GABA(A) receptor mediated, being blocked by pressure ejection of bicuculline. Thus the NTS integrates information from corneal receptors, some of which converges onto neurons mediating reflexes from baroreceptors and chemoreceptors to inhibit these inputs.     

Development of sensitivity to glycine, GABA and beta-alanine in cultured spinal neurons

Whole cell patch-clamp recording of cultured chick spinal neurons, presumed to be motoneurons, revealed currents elicited in these cells by GABA, glycine and beta-alanine, corresponding to the opening of chloride channels. During maturation, sensitivity to all three transmitters were first detected on the day 6 of culture, and appeared in most neurons by day 8. However, at all stages of development, a fraction of the cells were sensitive only to GABA; this observation supports the notion that the GABA and the glycine receptors are distinct. On the other hand, separate activation by glycine and beta-alanine was never observed, in agreement with the postulate that these amino acids bind to the same receptor.