non-NMDA Excitatory Amino Acid Receptors Research Articles

The ontogeny of glutamate receptors in rat barrel field cortex

The ontogeny of N-methyl- d-aspartate (NMDA) and non-NMDA excitatory amino acid receptors in rat barrel field cortex were characterized using receptor autoradiography. NMDA receptors showed a different pattern of development than that of non-NMDA receptors recognizing quisqualate (QUIS sites). During the first 14 days, high densities of QUIS sites were localized in barrel centers forming a sensory map of the rat whisker pad. After that time, the density of QUIS sites in barrel centers decreased so that the pattern was no longer apparent by postnatal day 21. In contrast to QUIS sites, NMDA sites did not exhibit a somatotopic pattern until postnatal day 21, when the lower density of sites in barrel septa formed an outline of barrel centers. At all ages examined, the density of NMDA sites did not differ significantly between barrel centers and surrounding cortex. Of the non-NMDA receptors examined in the postnatal day 10 old rat, both metabotropic sites and the NNKQ sites, which are [ 3H]glutamate binding sites that are not displaceable by NMDA, kainate or QUIS, showed a pattern of higher densities in barrel centers than surrounding tissue, whereas AMPA sites exhibited a complementary pattern. [ 3H]Glutamate binding to metabotropic sites was not significantly displaced by QUIS, whereas both NNKQ sites and metabotropic sites were potently blocked by the metabotropic agonist trans-ACPD. These results suggest that the NNKQ sites are low affinity QUIS metabotropic receptors, which, due to their high density in the immature barrel field, are in a position to influence barrel formation.

Localization and interaction of N-methyl-D-aspartate and non-N-methyl-D-aspartate receptors of lamprey spinal neurons

Small volumes of N-Methyl-D-Aspartate (NMDA) and non-NMDA excitatory amino acid receptor agonists were applied to localized regions of the dendritic trees of lamprey spinal neurons along their medial-lateral axis to obtain a spatial map of glutamate receptor distribution. Voltage clamp and frequency domain methods were used to obtain quantitative kinetic data of the voltage dependent ionic channels located both on the soma and on highly branched dendritic membranes. Pressure pulses of NMDA applied to the most peripheral regions of the dendritic tree elicited large somatic impedance increases, indicating that the most peripheral dendrites are well supplied with NMDA receptors. Experiments done with kainate did not elicit somatic responses to agonist applications on peripheral dendrites. The data obtained are consistent with the hypothesis that the activation of NMDA receptors by exogenous glutamate is significantly modified by the simultaneous activation of non-NMDA receptors, which shunts the NMDA response. The non-NMDA shunting hypothesis was tested by a combined application of kainate and NMDA to mimic the action of glutamate showing that the shunting effect of non-NMDA receptor activation virtually abolished the marked voltage dependency typical of NMDA receptor activation. These data were interpreted with a compartmental neuronal model having both NMDA and non-NMDA receptors.

A role for non-NMDA excitatory amino acid receptors in regulating the basal activity of rat globus pallidus neurons and their activation by the subthalamic nucleus

We have investigated the hypothesis that excitatory amino acid (EAA) receptors in the globus pallidus (GP) play a significant role in maintaining the firing rates of GP neurons under basal conditions and following activation of the subthalamic nucleus (STN). Drugs were infused directly into the GP and/or STN while the extracellular single unit activity of Type II GP neurons was recorded in ketamine-anesthetized rats. Local infusions of the EAA agonists NMDA (30–300 pmol/200 nl) or AMPA (0.1–1 pmol/200 nl) elicited increases in the firing rate of GP neurons in a dose-dependent fashion. Infusion of the GABA A receptor antagonist bicuculline methiodine (1–10 pmol/100 nl) into the STN also elicited dose-related increases in the firing rate of GP neurons. Intrapallidal infusion of the non-NMDA (AMPA/kainate) receptor antagonist NBQX (0.1–1.0 nmol) reduced the basal firing rate of GP neurons by 40%. In contrast, the NMDA antagonist MK-801 (0.01–0.1 nmol) produced no significant effect on basal firing rate. Intrapallidal infusion of the non-selective EAA receptor antagonist kynurenic acid or NBQX reversed or blocked the increase in firing rate of GP neurons following bicuculline-induced activation of the STN. Similar treatment with MK-801, however, had no significant effect on this response. These results indicate that tonic stimulation of non-NMDA receptors plays an important role in maintaining the basal activity of GP neurons and in mediating the effects of increased excitatory input from subthalamic afferent neurons.

Age-dependent appearance of synaptic currents in rat neocortical neurons in culture.

Rat neocortical neurons grown in dissociated cell culture were recorded with the whole-cell patch-clamp technique. Spontaneous inward currents were observed in cells that were held at a membrane potential of -80 mV in medium containing tetrodotoxin and Cd2+. These currents displayed amplitudes up to 140 pA and rise time of 1.8 +/- 0.2 ms (mean +/- SD, n = 15). They reversed near 0 mV and showed no voltage-dependent frequency of occurrence. Hence, they were presumably due to spontaneous release of transmitter. The inward currents appeared around day 10 in culture and were detected up to 4 weeks. When cells of different ages were compared, the maximal probability of recording these inward events occurred at around 3 weeks in culture. The inward currents were not reduced by application of bicuculline methiodide which is a competitive antagonist of the GABAA receptor, but were blocked by the broad-spectrum glutamate receptor antagonist kynurenic acid. Moreover, spontaneous inward events were not affected by DL-2-aminophosphono-valerate (NMDA receptor antagonist) but disappeared following application of the non-NMDA receptors antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Our observations indicate that the inward currents represent miniature synaptic events that are primarily mediated by non-NMDA excitatory amino acid receptor subtypes. Furthermore, our findings indicate that they develop over time and are not present in neurons that are grown in culture for less than 10 days.

Membrane properties and synaptic responses of interneurons located near the stratum lacunosum-moleculare/radiatum border of area CA1 in whole-cell recordings from rat hippocampal slices.

1. The membrane properties and synaptic inputs of interneurons, located at the stratum (s.) lacunosum-moleculare and radiatum border (L-M) of the CA1 region, were examined with the use of current-clamp whole-cell recordings in rat hippocampal slices. 2. Biocytin-labeled L-M interneurons had nonpyramidal somata and aspinous, often beaded, dendrites that arborized in s. lacunosum-moleculare and radiatum, sometimes as far as s. moleculare of the dentate gyrus. Their axon coursed and branched in s. lacunosum-moleculare and radiatum. Axon collaterals were also observed traversing the hippocampal fissure and arborizing in s. moleculare of the dentate gyrus and s. radiatum of the CA3 region. 3. Several membrane properties of interneurons were typically nonpyramidal: they had large input resistances, short-duration action potentials followed by prominent fast afterhyperpolarizations, and responded to hyperpolarizing current pulses with little membrane rectification. L-M interneurons showed significant anodal break responses, and their mean membrane time constant was 33 ms. After-depolarizations elicited by subthreshold depolarizing current pulses were larger in amplitude and decayed more slowly at depolarized than hyperpolarized membrane potentials. 4. The majority of L-M interneurons (35 of 49 cells) were silent at resting membrane potentials, whereas other displayed either spontaneous single action potentials (n = 12) or rhythmic bursts (n = 2). The rhythmic bursts were insensitive to the N-methyl-D-aspartate (NMDA) and non-NMDA excitatory amino acid receptor antagonists, 2-amino-5-phosphonopentanoic acid (AP-5; 50 microM) and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 20 microM), respectively. Both spontaneous single action potentials and burst firing were blocked by membrane hyperpolarization, suggesting that they were intrinsically rather than synaptically generated. 5. L-M interneurons responded with regular sustained firing to depolarizing current pulses at resting membrane potential. However, at more hyperpolarized membrane potentials (near -75 mV), depolarizing current pulses elicited action-potential firing with a delayed onset. This suggests that voltage-sensitive, transient outward currents may be activated in L-M interneurons from hyperpolarized membrane potentials. 6. Electrical stimulation of s. radiatum or lacunosum-moleculare elicited predominantly long-duration excitatory postsynaptic potentials (EPSPs; n = 20 cells), or both EPSPs and inhibitory postsynaptic potentials (IPSPs; n = 17 cells). In most L-M interneurons (35/37), with increasing intensities, up to two action potentials were elicited. Occasionally, larger bursts (3-5 action potentials) were observed (n = 2). 7. The multiphasic components of the synaptic responses became more evident when stimulations were repeated at different membrane potentials.(ABSTRACT TRUNCATED AT 400 WORDS)

The role of NMDA and non-NMDA excitatory amino acid receptors in the functional organization of primate retinal ganglion cells.

The role of excitatory amino acid (EAA) receptors in primate retinal ganglion cell function was analyzed in a superfused retina-eyecup preparation using single-unit, extracellular recording techniques. The effects of bath applied L-2-amino-4-phosphonobutyrate (APB), N-methyl-D-aspartate (NMDA), and non-NMDA EAA receptor agonists and antagonists were examined on the light-evoked responses and resting firing rates of ganglion cells. APB (30-100 microM) reduced or blocked the light-evoked responses and resting firing rates of all ON-center ganglion cells; higher doses of APB (100 microM) were required to block the light-evoked responses of ON-transient cells. In contrast, an increase in resting firing rates was observed when L-APB was applied to some OFF-center ganglion cells. The EAA agonists kainate (KA) (10-20 microM) and NMDA (200-350 microM) increased the firing rate of virtually all ganglion cells examined. Quisqualate (10-20 microM) increased firing in most cells, but occasionally (4/13 cases) produced inhibition. The NMDA antagonist D-amino-phosphono-heptanoic acid (D-AP7) (200-250 microM) reduced the light-evoked responses of ganglion cells by an average of 12% from control levels, while resting firing rates declined 37%. In the presence of D-AP7, the basic receptive-field characteristics of cells were not significantly altered. In contrast, two non-NMDA receptor antagonists, NBQX (2,3-Dihydroxy-6-nitro-7-sulfamoyl-benzo-(F)-quinoxalinedione) and DNQX (6,7-dinitro-quinoxaline-2,3-dione), produced substantial reductions in the light-evoked responses (82%) and resting firing rates (87%) of all ganglion cell classes. A striking observation in some neurons was the recovery of a persistent transient light-evoked response in the presence of NBQX. This NBQX-insensitive, light-evoked response was always blocked by adding D-AP7. Thus, neurotransmission from bipolar to ganglion cells in primates is mediated predominantly by non-NMDA EAA receptors, with NMDA receptors forming a minor component of the light-evoked response.

The pharmacology of the nicotinic antagonist, chlorisondamine, investigated in rat brain and autonomic ganglion.

1. A single administration of the ganglion blocker, chlorisondamine (10 mg kg-1, s.c.) is known to produce a quasi-irreversible blockade of the central actions of nicotine in the rat. The mechanism of this persistent action is not known. It is also unclear whether chlorisondamine can block neuronal responses to excitatory amino acids and whether chronic blockade of nicotinic responses also occurs in the periphery. 2. Acute administration of chlorisondamine (10 mg kg-1, s.c.) to rats resulted in a blockade of central nicotinic effects (ataxia and prostration) when tested 1 to 14 days later, but caused no detectable cell death in tissue sections sampled throughout the rostrocaudal extent of the brain which were stained in order to reveal neuronal degeneration. 3. Long-term blockade of central nicotinic effects by chlorisondamine was not associated with significant alterations in the density (Bmax) of high-affinity [3H]-nicotine binding to forebrain cryostat-cut sections. 4. In cultured dissociated mesencephalic cells of the foetal rat, chlorisondamine and mecamylamine inhibited [3H]-dopamine release evoked by N-methyl-D-aspartate (NMDA, 10(-4) M), but only at high concentrations (IC50 approx. 600 and 70 microM, respectively). A high concentration of chlorisondamine (10(-3) M) had no effect on responses to quisqualate (10(-5) M) and only slightly reduced responses to kainate (10(-4) M). Mecamylamine (10(-3) M) was ineffective against both agonists. 5. In adult rat hippocampal slices, chlorisondamine depressed NMDA receptor-mediated synaptically-evoked field potentials, but again only at high concentrations (10(-4)-10(-3) M). Synaptic responses that were mediated by non-NMDA excitatory amino acid receptors were less affected. 6. In rat isolated superior cervical ganglion, electrically-evoked synaptic transmission was reduced 1 h after acute in vivo administration of chlorisondamine (0.1 mg kg-1, s.c.). However, in vivo administration of a higher dose (10 mg kg-1, s.c.) did not significantly affect ganglionic transmission when tested two weeks later, despite the continued presence of central nicotinic blockade.7. These results indicate that the persistent CNS nicotinic blockade by chlorisondamine is not accompanied by changes in nicotinic [3H]-nicotine binding site density or by neuronal degeneration in the brain; that at doses sufficient to produce nicotinic receptor blockade, chlorisondamine acts in a pharmacologically selective manner; and that chronic central blockade is not accompanied by long-term peripheral ganglionic blockade.

Synthesis of a New Kainic Acid Based Selective Ligand as a Potential Photoaffinity Label of Non-NMDA Excitatory Amino Acid Receptors in Chicken Brain.

The synthesis of two analogues of kainic acid (KA) incorporating photo-activatible moieties attached either on the gamma-carboxy function (gamma-amide 1) or the isopropenyl side-chain (amide 2) is described. The synthesis of the former amide involves coupling of N-(tert-butoxycarbonyl)-protected alpha-diphenylmethyl kainate with 2-(4-azidobenzamido)ethylamine (5) followed by trifluoroacetic acid mediated complete deprotection. Amide 2 was synthesized by palladium-mediated allylic amination, with 4,4'-dimethoxybenzhydrylamine (DMBA), of N-(9-fluorenylmethoxycarbonyl)-protected dimethyl kainate, followed by splitting the DMB-group with formic acid, coupling with N-hydroxysuccinimidoyl 4-azidobenzoate and finally complete deprotection by saponification. Preliminary pharmacological studies in chicken brain membranes showed that amide 2 is a stronger inhibitor of [3H]KA binding on chicken cerebellar membranes than is amide 1 and that amide 2 has specificity only for the cerebellar, as opposed to the telencephalon, type of non-NMDA binding sites.

Activation of α-adrenoceptors indirectly facilitates sodium pumping in frog motoneurons

The effects of clonidine on Na + pumping in motoneurons of the isolated frog spinal cord was investigated using sucrose gap recordings from ventral roots. Na + pump activity, induced in motoneurons either by tetanizing the dorsal root or by rapidly exposing the cord to normal medium following 30 min in K +-free Ringer's solution (K +-activated hyperpolarizations), was increased by application of clonidine (100 μM). These actions of clonidine were blocked by the preferential α 2-adrenergic antagonist yohimbine, but not by α 1-adrenergic antagonist prazosin or the β-blocker propranolol. Clonidine's effects on Na + pumping appeared to be indirect (presumably via interneurons) because its effects on K +-activated hyperpolarizations were reduced by tetrodotoxin (TTX) or high concentrations of Mg 2+. This indirect mechanism involved activation of non-NMDA excitatory amino acid receptors. Thus, in the presence of clonidine, CNQX, but not APH, limited the ability of clonidine to enhance K +-activated hyperpolarizations. The AMPA receptor may play a role in the process. K +-activated hyperpolarizations were augmented by the presence of AMPA; NMDA had no effect. The present results are consistent with the idea that activation of α 2-adrenoceptors produces the following: the release of excitatory amino acids from interneurons; the activation of non-NMDA receptors on motoneurons; increased Na + influx and loading and increased Na + pump activity.

Evidence for Enhanced Synaptic Excitation in Transplanted Neostriatal Neurons

Fetal neostriatal tissue was transplanted into either the neostriatum or substantia nigra of adult rats. One to 6 months after transplantation, coronal brain slices were taken through the rostrocaudal extent of the transplants and neurons were characterized electrophysiologically using an in vitro slice preparation. When compared to control neurons taken from intact adult neostriata, transplanted neostriatal neurons (TSNs) had higher input resistances and longer time constants. All other passive and active membrane properties assessed were comparable between transplanted and control neostriatal neurons. Regardless of the transplantation site, local extracellular stimulation outside the graft elicited high-amplitude, long-duration depolarizing synaptic potentials that typically triggered bursts of action potentials. These synaptic potentials contrast with lower amplitude, shorter duration synaptic potentials consistently elicited in control neostriatal neurons. The depolarizing synaptic potentials evoked in the TSNs appeared to be mediated by a combined activation of N -methyl- d-aspartate (NMDA) and non-NMDA excitatory amino acid receptors. Both the broad-spectrum excitatory amino acid antagonist kynurenic acid and the specific non-NMDA receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione significantly reduced postsynaptic potentials elicited in TSNs. The specific NMDA antagonist 2-amino-5-phosphonovalerate had less effect on the amplitude but markedly reduced the duration of the synaptic potentials. The duration and amplitude of the bursts were augmented by the γ,-aminobutyric acid (GABA) A receptor antagonist bicuculline methiodide, indicating that inhibition occurred in TSNs. TSNs were also more sensitive than control neurons to direct application of glutamate or NMDA. These findings demonstrate that TSNs express altered electrophysiological properties. The pharmacological analysis indicates that depolarizing postsynaptic potentials were mediated by activation of excitatory amino acid receptors, suggesting either innervation of the graft by host fibers which contain excitatory amino acids or development of novel local excitatory interactions intrinsic to the graft. Furthermore, the occurrence of high-amplitude, long-duration depolarizing synaptic potentials in TSNs, regardless of the site of transplantation, suggests that grafted neostriatal neurons become hyperexcitable to synaptic input.

Kindling to the benzodiazepine receptor inverse agonist, FG 7142: Evidence for involvement of NMDA, but not non-NMDA, glutamatergic receptors

Repeated administration of the β-carboline FG 7142 to mice leads to the development of kindled convulsions. In order to investigate a role for glutamatergic mechanisms in the processes underlying FG 7142 kindling, the N- methyl- d-aspartate (NMDA) antagonist, 2-amino-7-phosphonoheptanoic acid (AP7; 25 nmol), was administered intracerebroventricularly (i.c.v.) daily before administration of FG 7142 (40 mg/kg, i.p.). Under these conditions, kindling to FG 7142 did not occur. Administration of two antagonists at non-NMDA excitatory amino acid receptors, 6-cyano-7-nitroquinoxaline-2, 3-dione (CNQX) and γ- d-glutamylaminomethylsulphonic acid (γ- d-GAMS; both 25 nmol) did not prevent the development of Seizures; these doses were, however, adequate and selective in protecting against seizures induced by respectively quisqualic and kainic acids given by i.c.v. The susceptibility of mice kindled with FG 7142 to Scizures induced by NMDA, or kainate or quisqualate was similar in mice which had shown 5 kindled Scizures to that seen in drug-naive mice; mice which had shown 10 kindled Scizures showed a decreased sensitivity to NMDA-induced convulsions (ED 50 was increased from 0.24 to 0.31 nmol). No changes were seen in the convulsant thresholds of either NMDA or non-NMDA agonists. These observations suggest that although NMDA receptors appear to be involved in the processes underlying FG 7142 kindling, such kindling is not necessarily associated with an increased sensitivity of glutamate receptors, and in animals which have convulsed, a decreased sensitivity to NMDA agonists occurs.

Epileptiform activity induced by 4-aminopyridine in guinea-pig and rat neocortices

Extracellular field recordings were performed in guinea-pig and rat neocortical slice preparations maintained in vitro. Bath application of the convulsant drug 4-aminopyridine (4-AP, 100 μM) induced spontaneous epileptiform potentials in 80% of the guinea-pig neocortical slices and only in 6% of the neocortical slices from rat. In both species spontaneous epileptiform activity consisted of a 4–16 s long ictal-like discharge that recurred with a frequency range of 0.01–0.02 Hz. In rat neocortical slices stimulus-induced responses resembled the spontaneous occurring epileptiform events. Ictal-like discharges in guinea-pig neocortical slices were blocked by the N- methyl- d-aspartate (NMDA) receptor antagonist 3-((±-2-carboxypiperazine-4-yl)propyl-1-phosphonic acid (5 μM), while those in the rat desappeared during perfusion with the non-NMDA excitatory amino acid receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (1–3 μM). These results indicate that the neocortex of guinea-pig has a higher propensity to generate 4-AP-induced spontaneous epileptiform activity than that of rat. Furthermore the epileptiform activity in these two species requires a different involvement of excitatory amino acid receptors.

Properties of [3H]AMPA Binding in Postmortem Human Brain from Psychotic Subjects and Controls: Increases in Caudate Nucleus Associated with Suicide

[3H]AMPA binding, a measure of the non-NMDA excitatory amino acid receptors, was measured in the frontal cortex, caudate nucleus, and nucleus accumbens of postmortem human brain tissue samples. In normal human frontal cortex, the binding data were best fit by a two-site model, with Kd values of 137 nM and 11.3 μM, and Bmax values of 2780 fmol/mg protein and 67.6 pmol/mg protein, respectively. Binding was linearly related to protein concentration and was strongly inhibited by glutamic acid and quisqualic acid. Binding was partially inhibited by kainic acid and glutamic acid diethyl ester and only slightly inhibited by N-methyl-d-aspartic acid. AMPA binding was not inhibited by neuroleptic drugs, in vitro. Freezing and storage did not result in a loss of AMPA binding, and there tended to be an increase in AMPA binding with extended freezer storage. When tissue frozen intact was compared to tissue frozen as a homogenate, the high-affinity binding parameters were unchanged, but there was an increase in the affinity and Bmax of the low-affinity site for the tissue frozen intact. Thus it appears that only the high-affinity site can be measured accurately in tissue frozen intact. AMPA binding was not significantly altered by premortem neuroleptic administration, age, postmortem delay, or by moderate durations of freezer storage. No differences in AMPA binding were found in psychotic subjects compared to normal controls. There was, however, a pronounced increase in total AMPA binding in the caudate nucleus in subjects that had committed suicide.

Pulmonary stretch receptor afferents activate excitatory amino acid receptors in the nucleus tractus solitarii in rats.

1. The goal of the present study was to identify potential neurotransmitter candidates in the Breuer-Hering (BH) reflex pathway, specifically at synapses between the primary afferents and probable second-order neurones (pump cells) within the nucleus tractus solitarii (NTS). We hypothesized that if activation of specific receptors in the NTS is required for production of the BH reflex, then (1) injection of the receptor agonist(s) would mimic the reflex response (apnoea), (2) injection of appropriate antagonists would impair the apnoea produced by either lung inflation or agonist injection, and (3) second-order neurones in the pathway would be excited by either lung inflation or agonists while antagonists would prevent the response to either. 2. Studies were carried out either in spontaneously breathing or in paralysed, thoracotomized and ventilated rats in which either diaphragm EMG or phrenic nerve activity, expired CO2 concentration and arterial pressure were continuously monitored. The BH reflex was physiologically activated by inflating the lungs. 3. Pressure injections (0.03-15 pmol) of selective excitatory amino acid (EAA) receptor agonists, quisqualic acid (Quis) and N-methyl-D-aspartic acid (NMDA) into an area of the NTS shown previously to contain neurones required for production of the BH reflex produced dose-dependent apnoeas that mimicked the response to lung inflation. Injection of substance P (0.03-4 pmol) did not alter baseline respiratory pattern. 4. Injections of the EAA antagonists, kynurenic acid (Kyn; 0.6-240 pmol), 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX) or 6,7-dinitroquinoxaline-2,3-dione (DNQX) into the BH region of the NTS reversibly impaired the apnoea produced by lung inflation. All three antagonists reduced or abolished the apnoeas resulting from injection of Quis or NMDA, and slowed baseline respiratory frequency. In contrast, injections of the highly selective NMDA receptor antagonist, D-2-amino-5-phosphonovaleric acids (AP5), in doses sufficient to block the apnoeic response to NMDA, neither altered the reflex apnoea evoked by lung inflation nor the baseline respiratory pattern. 5. Pump cells located within the BH region were excited by pressure injections of the broad spectrum EAA agonist, DL-homocysteic acid (DLH). Kyn reversibly blocked the excitation of pump cells in response to either lung inflation or DLH injection. 6. These findings suggest that EAAs mediate primary afferent excitation of second-order neurones in the Breuer-Hering reflex pathway, primarily through the activation of non-NMDA EAA receptor subtypes.

A1 neurons and excitatory amino acid receptors in rat caudal medulla mediate vagal excitation of supraoptic vasopressin cells

Extracellular recordings from the supraoptic nucleus of the rat established that vasopressinergic neurosecretory cells were excited by stimulation of cervical but not abdominal vagal afferents. This response was absent or significantly attenuated after microinjection of γ-aminobutyric acid into a region of the caudal medulla known to contain the A1 noradrenaline cell group. Consistent with the possible involvement of the A1 group, vagal stimulation approximately doubled the frequency of proto-oncogene expression in A1 noradrenaline neurons, as indicated by the occurrence of nuclear Fos-like immunoreactivity in tyrosine hydroxylase-positive neurons of the caudal ventrolateral medulla. Finally, A1 region microinjection of either the N- methyl- d-aspartic acid (NMDA) receptor antagonist dl-2-amino-5-phosphonovaleric acid (APV), or the non-NMDA antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), significantly reduced vasopressin cell responses to vagal stimulation. These findings suggest that: (i) the A1 group is an essential component in a pathway which relays facilitatory vagal input of cardiopulmonary origin to neurosecretory vasopressin cells, and (ii) the activation of A1 neurons in this pathway involves both NMDA and non-NMDA excitatory amino acid receptors, an observation consistent with an input to A1 cells which generates ‘mixed’ excitatory postsynaptic potentials.

Intrathecal administration of non-NMDA receptor agonists increases arterial pressure and heart rate in the rat

We have found that spinal NMDA receptors are involved in control of sympathetic output in pathways to the heart and vessels. The present study was done to determine whether spinal non-NMDA excitatory amino acid receptors participate in cardiovascular regulation. Experiments were done on urethane-anesthetized Sprague-Dawley rats, giving the non-NMDA receptor agonists, quisqualate and kainate, and the antagonist, kynurenate, intrathecally at the spinal T 9 level. Both quisqualate (30 nmol; n = 7; to activate AMPA receptors) and kainate (2 nmol; n = 6; to activate K receptors) increased arterial pressure and heart rate. The responses were characterized by a rapid onset, achieving, in most cases, > 80% of the maximum response within 1–4 min, and a persistence throughout the remaining 20–24 min of the experiment. I.v. injection of hexamethonium (10 mg/kg) prevented the effects of intrathecal administration of quisqualate ( n = 5) but not of kainate ( n = 7). To determine whether the hexamethonium-resistant effects of kainate were due to a peripheral action, kainate was given i.v. ( n = 6); it was found to be without effect on arterial pressure or heart rate. The increases in arterial pressure and heart rate produced by intrathecal administration of quisqualate (30 nmol; n = 6), kainate (2 nmol; n = 6), glutamate (1 μmol; n = 6) and NMDA (2 nmol; n = 6) but not carbachol (27.4 nmol; n = 6) were prevented by similar preadministration of kynurenate (125 nmol). Intrathecal administration of kynurenate (125 nmol; n = 6; 500 nmol; n = 7) decreased arterial pressure and/or heart rate. The greatest decreases in the group given the higher dose occurred at about 10 min (systolic pressures, 26.7 ± 1.5 mmHg; diastolic pressure, 13.6 ± 1.8 mmHg) and 4 min (heart rate, 13.3 ± 5.0 bpm). These responses persisted for 7–25 min. The results indicate: (1) spinal non-NMDA receptors are involved in activation of sympathetic output regulating cardiovascular function; (2) pathways activated by quisqualate and by kainate at the spinal level are mediated in sympathetic ganglia via nicotinic and non-nicotinic mechanisms, respectively; (3) there appears to be a tonic activation of non-NMDA receptors in pathways to the heart.

Induction of long-term potentiation in the basolateral amygdala does not depend on NMDA receptor activation.

Long-term potentiation (LTP) can be induced in the lateral and basolateral amygdala by stimulating synaptic afferents in the external capsule (EC). We examined the sensitivity of amygdaloid LTP to the NMDA receptor antagonist 2-amino-5-phosphonopentanoate (AP5), which is known to block LTP induction in the Schaffer collateral/CA1 synapses in the hippocampus. While relatively high concentrations (100 microM) of DL-AP5 were effective in preventing LTP induction in the lateral and basolateral amygdala in vitro, the same concentrations also significantly depressed synaptic responses to low-frequency stimulation. Furthermore, at 50 microM, a concentration sufficient to block both synaptic responses mediated by NMDA receptors and LTP induction in the hippocampus and neocortex, AP5 did not affect the probability of inducing LTP in the amygdala. Application of 10 microM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), which blocks non-NMDA excitatory amino acid receptors, reduced the monosynaptic response to EC stimulation by 85%. The remaining CNQX-insensitive response did not appear to be mediated by NMDA-type receptors, since it was not reduced by 50 or 100 microM AP5, and showed none of the voltage sensitivity characteristic of NMDA responses. These data suggest that while the induction of LTP in the amygdala produced by EC stimulation is blocked by high doses of AP5, plasticity at these synapses probably does not require activation of NMDA receptors.

The role of NMDA and non-NMDA excitatory amino acid receptors in the excitation of primate spinothalamic tract neurons by mechanical, chemical, thermal, and electrical stimuli

The role of excitatory amino acids (EAAs) in the excitation of monkey spinothalamic tract (STT) neurons following activation of cutaneous primary afferent fibers by noxious and non-noxious stimuli was investigated. The responses of STT neurons to either NMDA or non-NMDA EAA ligands were blocked by infusion of specific antagonists through a microdialysis fiber into the region surrounding the cells. Our results show that blockade of non-NMDA receptors results in a nearly complete elimination of the responses of STT neurons to all stimuli. Blockade of NMDA receptors results in an attenuation of the responses to noxious stimuli but, in addition, prevents the development of the sensitization of STT neurons that is often observed after intradermal injection of capsaicin. These observations further support a role of EAAs in the transmission of sensory information from primary afferent fibers to dorsal horn neurons and a role for NMDA receptors in the generation of hyperalgesia.

Activation of skeletal muscle nicotinic acetylcholine receptors.

Work over the past ten years has greatly increased our understanding of both the structure and function of the muscle nicotinic acetylcholine receptor. There is a strongly supported general picture of how the receptor functions: agonist binds rapidly to sites of low affinity and channel opening occurs at a rate comparable to the agonist dissociation rate. Channel closing is slow, so the channel has a high probability of being open if both agonist-binding sites are occupied by ACh. Results of expression studies have shown that each subunit can influence AChR activation and have given a structural basis for the major physiological change known for muscle AChR, the developmental change in AChR activation. These general statements notwithstanding, there are still major areas of uncertainty which limit our understanding. We have emphasized these areas of uncertainty in this review, to indicate what needs to be done. First, the quantitative estimates of rate constants are not as strongly supported as they should be. The major reasons are twofold--uncertainties about the interpretation of components in the kinetic data and difficulties of resolving brief events. As a result, any inferences about the functional consequences of structural alterations must remain tenuous. Second, the functional behavior of individual AChRs is not as well understood as it should be. The kinetic behavior of an individual receptor clearly can be complex (section II). In addition, there is evidence that superimposed on this complexity there may be stable and kinetically distinguishable populations of receptors (section III). Until the basis for the kinetically defined populations is clarified, kinetic parameters for receptors of defined structure cannot be unambiguously obtained. Finally, it is not surprising that the studies of AChR of altered structure have not given definitive results. Two reasons should be apparent from the preceding points: there is not a fully supported approach for kinetic analysis, and the "normal" population may not be clearly defined. An additional complication is also emerging, in that the available data support the idea that specific residues distributed over all subunits may influence AChR activation. This possibility renders the task of analysis that much more difficult. The muscle nicotinic AChR has served as a prototype for the family of transmitter-gated membrane channels, which includes the muscle and neuronal nicotinic receptors, the GABAA, the glycine and possibly the non-NMDA excitatory amino acid receptor (Stroud et al., 1990). It is interesting to note that the functional properties of the GABAA receptor, probably the best-studied of the other members of the family are rather similar.(ABSTRACT TRUNCATED AT 400 WORDS)

Abnormal action-potential bursts and synchronized, GABA-mediated inhibitory potentials in an in vitro model of focal epilepsy.

Focal, freeze-induced lesions were made in isolated hemispheres of turtle cerebral cortex in vitro, permitting the investigation of epileptiform discharges in a preparation with preserved intracortical circuitry. Freeze lesions resulted in interictal discharges and occasional ictal-like events. The interictal discharges were dependent upon activation of non-NMDA excitatory amino acid receptors and were affected by but did not require NMDA receptor activation. Voltage clamp and current clamp recordings revealed abnormal bursts of low-amplitude action potentials in 36% of recorded neurons, while large, repetitive inhibitory potentials, mediated by GABAA receptors, were recorded in 90% of the neurons. Thus, prominent findings in this model include abnormalities of both excitatory and inhibitory activity. Since these changes in neuronal excitability resulted from a localized physical injury, they may resemble the changes that occur in acute posttraumatic epilepsy.