2-3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline and Epilepsy

Epilepsy may arise following acute brain insults, but no treatments exist that prevent epilepsy in patients at risk. Here we examined whether a combination of two glutamate receptor antagonists, NBQX and ifenprodil, acting at different receptor subtypes, exerts antiepileptogenic effects in the intrahippocampal kainate mouse model of epilepsy. These drugs were administered over 5 days following kainate. Spontaneous seizures were recorded by video/EEG at different intervals up to 3 months. Initial trials showed that drug treatment during the latent period led to higher mortality than treatment after onset of epilepsy, and further, that combined therapy with both drugs caused higher mortality at doses that appear safe when used singly. We therefore refined the combined-drug protocol, using lower doses. Two weeks after kainate, significantly less mice of the NBQX/ifenprodil group exhibited electroclinical seizures compared to vehicle controls, but this effect was lost at subsequent weeks. The disease modifying effect of the treatment was associated with a transient prevention of granule cell dispersion and less neuronal degeneration in the dentate hilus. These data substantiate the involvement of altered glutamatergic transmission in the early phase of epileptogenesis. Longer treatment with NBQX and ifenprodil may shed further light on the apparent temporal relationship between dentate gyrus reorganization and development of spontaneous seizures.

Topics: Animals; Anticonvulsants; Dentate Gyrus; Disease Models, Animal; Drug Administration Schedule; Drug Therapy, Combination; Electroencephalography; Epilepsy; Humans; Kainic Acid; Male; Mice; Neurons; Piperidines; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Time Factors; Treatment Outcome

Hypersynchronous neuronal excitation manifests clinically as seizure (ictogenesis), and may recur spontaneously and repetitively after a variable latency period (epileptogenesis). Despite tremendous research efforts to describe molecular pathways and signatures of epileptogenesis, molecular pathomechanisms leading to chronic epilepsy remain to be clarified. We hypothesized that epigenetic modifications may form the basis for a cellular memory of epileptogenesis, and used a primary neuronal cell culture model of the rat hippocampus to study the translation of massive neuronal excitation into persisting changes of epigenetic signatures and pro-epileptogenic target gene expression. Increased spontaneous activation of cultured neurons was detected 3 and 7 days after stimulation with 10 μM glutamate when compared to sham-treated time-matched controls using calcium-imaging in vitro. Chromatin-immunoprecipitation experiments revealed short-term (3 h, 7 h, and 24 h) and long-term (3 d and 2 weeks) changes in histone modifications, which were directly linked to decreased expression of two selected epilepsy target genes, e.g. excitatory glutamate receptor genes Gria2 and Grin2a. Increased promoter methylation observed 4 weeks after glutamate stimulation at respective genes suggested long-term repression of Gria2 and Grin2a genes. Inhibition of glutamatergic activation or blocking the propagation of action potentials in cultured neurons rescued altered gene expression and regulatory epigenetic modifications. Our data support the concept of a cellular memory of epileptogenesis and persisting epigenetic modifications of epilepsy target genes, which are able to turn normal into pro-epileptic neurons and circuits.

Topics: Action Potentials; Animals; Animals, Newborn; Cells, Cultured; DNA Methylation; Epigenesis, Genetic; Epilepsy; Excitatory Amino Acid Agents; Gene Expression; Glutamic Acid; Hippocampus; Microtubule-Associated Proteins; Models, Biological; Neurons; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Sodium Channel Blockers; Tetrodotoxin; Valine

The neural precursor cell expressed developmentally down-regulated gene 4-2, Nedd4-2, is an epilepsy-associated gene with at least three missense mutations identified in epileptic patients. Nedd4-2 encodes a ubiquitin E3 ligase that has high affinity toward binding and ubiquitinating membrane proteins. It is currently unknown how Nedd4-2 mediates neuronal circuit activity and how its dysfunction leads to seizures or epilepsies. In this study, we provide evidence to show that Nedd4-2 mediates neuronal activity and seizure susceptibility through ubiquitination of GluA1 subunit of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor, (AMPAR). Using a mouse model, termed Nedd4-2andi, in which one of the major forms of Nedd4-2 in the brain is selectively deficient, we found that the spontaneous neuronal activity in Nedd4-2andi cortical neuron cultures, measured by a multiunit extracellular electrophysiology system, was basally elevated, less responsive to AMPAR activation, and much more sensitive to AMPAR blockade when compared with wild-type cultures. When performing kainic acid-induced seizures in vivo, we showed that elevated seizure susceptibility in Nedd4-2andi mice was normalized when GluA1 is genetically reduced. Furthermore, when studying epilepsy-associated missense mutations of Nedd4-2, we found that all three mutations disrupt the ubiquitination of GluA1 and fail to reduce surface GluA1 and spontaneous neuronal activity when compared with wild-type Nedd4-2. Collectively, our data suggest that impaired GluA1 ubiquitination contributes to Nedd4-2-dependent neuronal hyperactivity and seizures. Our findings provide critical information to the future development of therapeutic strategies for patients who carry mutations of Nedd4-2.

Topics: 14-3-3 Proteins; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amino Acid Sequence; Animals; Animals, Newborn; Blotting, Western; Cells, Cultured; Endosomal Sorting Complexes Required for Transport; Epilepsy; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Genetic Predisposition to Disease; HEK293 Cells; Humans; Lysine; Male; Mice, Knockout; Microscopy, Confocal; Mutation, Missense; Nedd4 Ubiquitin Protein Ligases; Neurons; Quinoxalines; Receptors, AMPA; Seizures; Ubiquitin-Protein Ligases

Epilepsy is a serious brain disorder with diverse seizure types and epileptic syndromes. AMPA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzoquinoxaline-2,3-dione (NBQX) attenuates spontaneous recurrent seizures in rats. However, the anti-epileptic effect of NBQX in chronic epilepsy model is poorly understood. Perineuronal nets (PNNs), specialized extracellular matrix structures, surround parvalbumin-positive inhibitory interneurons, and play a critical role in neuronal cell development and synaptic plasticity. Here, we focused on the potential involvement of PNNs in the treatment of epilepsy by NBQX. Rats were intraperitoneally (i.p.) injected with pentylenetetrazole (PTZ, 50 mg/kg) for 28 consecutive days to establish chronic epilepsy models. Subsequently, NBQX (20 mg/kg, i.p.) was injected for 3 days for the observation of behavioral measurements of epilepsy. The Wisteria floribundi agglutinin (WFA)-labeled PNNs were measured by immunohistochemical staining to evaluate the PNNs. The levels of three components of PNNs such as tenascin-R, aggrecan and neurocan were assayed by Western blot assay. The results showed that there are reduction of PNNs and decrease of tenascin-R, aggrecan and neurocan in the medial prefrontal cortex (mPFC) in the rats injected with PTZ. However, NBQX treatment normalized PNNs, tenascin-R, aggrecan and neurocan levels. NBQX was sufficient to decrease seizures through increasing the latency to seizures, decrease the duration of seizure onset, and reduce the scores for the severity of seizures. Furthermore, the degradation of mPFC PNNs by chondroitinase ABC (ChABC) exacerbated seizures in PTZ-treated rats. Finally, the anti-epileptic effect of NBQX was reversed by pretreatment with ChABC into mPFC. These findings revealed that PNNs degradation in mPFC is involved in the pathophysiology of epilepsy and enhancement of PNNs may be effective for the treatment of epilepsy.

Topics: Aggrecans; Animals; Anticonvulsants; Behavior, Animal; Chondroitin ABC Lyase; Disease Models, Animal; Down-Regulation; Epilepsy; Immunohistochemistry; Male; Neurocan; Pentylenetetrazole; Peripheral Nerves; Prefrontal Cortex; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Tenascin

Hyaluronan (HA), a large anionic polysaccharide (glycosaminoglycan), is a major constituent of the extracellular matrix of the adult brain. To address its function, we examined the neurophysiology of knock-out mice deficient in hyaluronan synthase (Has) genes. Here we report that these Has mutant mice are prone to epileptic seizures, and that in Has3(-/-) mice, this phenotype is likely derived from a reduction in the size of the brain extracellular space (ECS). Among the three Has knock-out models, namely Has3(-/-), Has1(-/-), and Has2(CKO), the seizures were most prevalent in Has3(-/-) mice, which also showed the greatest HA reduction in the hippocampus. Electrophysiology in Has3(-/-) brain slices demonstrated spontaneous epileptiform activity in CA1 pyramidal neurons, while histological analysis revealed an increase in cell packing in the CA1 stratum pyramidale. Imaging of the diffusion of a fluorescent marker revealed that the transit of molecules through the ECS of this layer was reduced. Quantitative analysis of ECS by the real-time iontophoretic method demonstrated that ECS volume was selectively reduced in the stratum pyramidale by ∼ 40% in Has3(-/-) mice. Finally, osmotic manipulation experiments in brain slices from Has3(-/-) and wild-type mice provided evidence for a causal link between ECS volume and epileptiform activity. Our results provide the first direct evidence for the physiological role of HA in the regulation of ECS volume, and suggest that HA-based preservation of ECS volume may offer a novel avenue for development of antiepileptogenic treatments.

Topics: Action Potentials; Animals; Brain; Electric Stimulation; Electroencephalography; Epilepsy; Excitatory Amino Acid Antagonists; Extracellular Space; Glucuronosyltransferase; Hyaluronan Synthases; Hyaluronic Acid; In Vitro Techniques; Mice; Mice, Knockout; Models, Neurological; Mutation; Nerve Net; Neurons; Phosphopyruvate Hydratase; Quinoxalines

EPAC (Exchange Proteins Activated by cAMP) regulates glutamate transmitter release in the central neurons, but a role underlying this regulation has yet to be identified. Here we show that EPAC binds directly to the intracellular loop of an ATP-sensitive potassium (KATP) channel type-1 sulfonylurea receptor (SUR1) receptor consisting of amino acids 859-881 (SUR1(859-881)). Ablation of EPAC or expression of SUR1(859-881), which intercepts EPAC-SUR1 binding, increases the open probability of KATP channels consisting of the Kir6.1 subunit and SUR1. Opening of KATP channels inhibits glutamate release and reduces seizure vulnerability in adult mice. Therefore, EPAC interaction with SUR1 controls seizure susceptibility and possibly acts via regulation of glutamate release.

Topics: Animals; Bicuculline; Biophysics; Disease Models, Animal; Electric Stimulation; Epilepsy; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; GABA-A Receptor Antagonists; Glutamic Acid; Guanine Nucleotide Exchange Factors; Hippocampus; Humans; Immunoprecipitation; In Vitro Techniques; Kainic Acid; KATP Channels; Male; Membrane Potentials; Mice; Mice, Knockout; Patch-Clamp Techniques; Probability; Protein Binding; Quinoxalines; Synaptosomes; Transduction, Genetic

We describe a form of very fast oscillation (VFO) in patient electrocorticography (ECoG) recordings, that can occur prior to ictal events, in which the frequency increases steadily from ≈ 30-40 to >120 Hz, over a period of seconds. We dub these events "glissandi" and describe a possible model for them.. Four patients with epilepsy had presurgical evaluations (with ECoG obtained in two of them), and excised tissue was studied in vitro, from three of the patients. Glissandi were seen spontaneously in vitro, associated with ictal events-using acute slices of rat neocortex-and they were simulated using a network model of 15,000 detailed layer V pyramidal neurons, coupled by gap junctions.. Glissandi were observed to arise from human temporal neocortex. In vitro, they lasted 0.2-4.1 s, prior to ictal onset. Similar events were observed in the rat in vitro in layer V of frontal neocortex when alkaline solution was pressure-ejected; glissandi persisted when γ-aminobutyric acid A (GABA(A)), GABA(B), and N-methyl-d-aspartate (NMDA), and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors were blocked. Nonalkaline conditions prevented glissando generation. In network simulations it was found that steadily increasing gap junction conductance would lead to the observed steady increase in VFO field frequency. This occurred because increasing gap junction conductance shortened the time required for an action potential to cross from cell to cell.. The in vitro and modeling data are consistent with the hypothesis that glissandi arise when pyramidal cell gap junction conductances rise over time, possibly as a result of an alkaline fluctuation in brain pH.

Topics: 2-Amino-5-phosphonovalerate; Action Potentials; Adult; Animals; Axons; Computer Simulation; Cortical Synchronization; Electric Stimulation; Electroencephalography; Epilepsy; Excitatory Amino Acid Antagonists; Female; GABA Antagonists; Gap Junctions; Humans; In Vitro Techniques; Male; Middle Aged; Models, Neurological; Neocortex; Nerve Net; Neurons; Phosphinic Acids; Propanolamines; Quinoxalines; Rats; Rats, Wistar; Reaction Time

Epileptiform discharges recorded in the 4-aminopyridine (4-AP) in vitro epilepsy model are mediated by glutamatergic and GABAergic signaling. Using a 60-channel perforated multi-electrode array (pMEA) on corticohippocampal slices from 2 to 3 week old mice we recorded interictal- and ictal-like events. When glutamatergic transmission was blocked, interictal-like events no longer initiated in the hilus or CA3/CA1 pyramidal layers but originated from the dentate gyrus granule and molecular layers. Furthermore, frequencies of interictal-like events were reduced and durations were increased in these regions while cortical discharges were completely blocked. Following GABA(A) receptor blockade interictal-like events no longer propagated to the dentate gyrus while their frequency in CA3 increased; in addition, ictal-like cortical events became shorter while increasing in frequency. Lastly, drugs that affect tonic and synaptic GABAergic conductance modulated the frequency, duration, initiation and propagation of interictal-like events. These findings confirm and expand on previous studies indicating that multiple synaptic mechanisms contribute to synchronize neuronal network activity in forebrain structures. This article is part of a Special Issue entitled 'Trends in neuropharmacology: in memory of Erminio Costa'.

Topics: 4-Aminopyridine; Animals; Anticonvulsants; Bicuculline; CA3 Region, Hippocampal; Disease Models, Animal; Electrodes; Epilepsy; GABA-A Receptor Antagonists; gamma-Aminobutyric Acid; Hippocampus; In Vitro Techniques; Isoxazoles; Mice; Mice, Inbred C57BL; Microarray Analysis; Motion Pictures; Piperazines; Potassium Channel Blockers; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Software; Somatosensory Cortex

Dark neurons have plagued the interpretation of brain tissue sections, experimentally and clinically. Seen only when perturbed but living tissue is fixed in aldehydes, their mechanism of production is unknown. Since dark neurons are seen in cortical biopsies, experimental ischemia, hypoglycemia, and epilepsy, we surmised that glutamate release and neuronal transmembrane ion fluxes could be the perturbation leading to dark neuron formation while the fixation process is underway. Accordingly, we excised biopsies of rat cortex to simulate neurosurgical production of dark neurons. To ascertain the role of glutamate, blockade of N-methyl-D-aspartate (NMDA) and non-NMDA receptors was done prior to formaldehyde fixation. To assess the role of transmembrane sodium ion (and implicitly, water) fluxes, tetraethylammonium (TEA) was used. Blockade of NMDA receptors with MK-801 and non-NMDA receptors with the quinoxalinediones (CNQX and NBQX) abolished dark neuron formation. More delayed exposure of the tissue to the antagonist, CNQX, by admixing it with the fixative directly, allowed for some production of dark neurons. Aminophosphonoheptanoate (APH), perhaps due to its polarity, and TEA, did not prevent dark neurons, which were abundant in control formaldehyde fixed material unexposed to either receptor or ion channel antagonists. The results demonstrate a role for the pharmacologic subtypes of glutamate receptors in the pathogenetic mechanism of dark neuron formation. Our results are consistent with the appearance of dark neurons in biopsy where the cerebral cortex has been undercut, and rendered locally ischemic and hypoglycemic, as well as in epilepsy, hypoglycemia, and ischemia, all of which lead to glutamate release. Rather than a pressure-derived mechanical origin, we suggest that depolarization, glutamate release or receptor activation are more likely mechanisms of dark neuron production.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Artifacts; Biopsy; Brain Ischemia; Cerebral Cortex; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Antagonists; Glutamic Acid; Hypoglycemia; Male; Neurons; Potassium Channel Blockers; Quinoxalines; Rats; Rats, Wistar; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Tetraethylammonium

The highest incidence of seizures during lifetime is found in the neonatal period and neonatal seizures lead to a propensity for epilepsy and long-term cognitive deficits. Here, we identify potential mechanisms that elucidate a critical role for AMPA receptors (AMPARs) in epileptogenesis during this critical period in the developing brain. In a rodent model of neonatal seizures, we have shown previously that administration of antagonists of the AMPARs during the 48 h after seizures prevents long-term increases in seizure susceptibility and seizure-induced neuronal injury. Hypoxia-induced seizures in postnatal day 10 rats induce rapid and reversible alterations in AMPAR signaling resembling changes implicated previously in models of synaptic potentiation in vitro. Hippocampal slices removed after hypoxic seizures exhibited potentiation of AMPAR-mediated synaptic currents, including an increase in the amplitude and frequency of spontaneous and miniature EPSCs as well as increased synaptic potency. This increased excitability was temporally associated with a rapid increase in phosphorylation at GluR1 S845/S831 and GluR2 S880 sites and increased activity of the protein kinases CaMKII (calcium/calmodulin-dependent protein kinase II), PKA, and PKC, which mediate the phosphorylation of these AMPAR subunits. Postseizure administration of AMPAR antagonists NBQX (2,3-dihydroxy-6-nitro-7-sulfonyl-benzo[f]quinoxaline), topiramate, or GYKI-53773 [(1)-1-(4-aminophenyl)-3-acetyl-4-methyl-7,8-methylenedioxy-3,4-dihydro-5H-2,3-benzodiazepine] attenuated the AMPAR potentiation, phosphorylation, and kinase activation and prevented the concurrent increase in in vivo seizure susceptibility. Thus, the potentiation of AMPAR-containing synapses is a reversible, early step in epileptogenesis that offers a novel therapeutic target in the highly seizure-prone developing brain.

Topics: Animals; Animals, Newborn; Anticonvulsants; Benzodiazepines; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cyclic AMP-Dependent Protein Kinases; Disease Susceptibility; Enzyme Activation; Epilepsy; Excitatory Postsynaptic Potentials; Fructose; Hypoxia; Male; Phosphorylation; Protein Kinase C; Quinoxalines; Rats; Rats, Long-Evans; Receptors, AMPA; Synapses; Topiramate

Excitatory amino acids play an important role in generation of epileptic seizures. To study the participation of different types of their receptors in cortical epileptic afterdischarges, a noncompetitive NMDA receptor antagonist dizocilpine and a competitive AMPA receptor antagonist NBQX were used. Adult rats with implanted epidural stimulation and registration electrodes were pretreated either with NBQX (30 or 60 mg/kg i.p.) or with dizocilpine (0.1 or 0.5 mg/kg i.p.) and low-frequency stimulation of sensorimotor cortical area was repeatedly applied with stepwise increased current intensities. Lower dose of NBQX unexpectedly decreased thresholds for elicitation of spike-and-wave afterdischarges (ADs), clonic seizures accompanying this type of ADs and for transition into the second, limbic type of ADs. Lower dose of dizocilpine increased these three thresholds. Higher doses of either drug did not significantly change threshold intensities. Duration of ADs was also influenced by the two antagonists in opposite directions: higher dose of NBQX resulted in prolongation of ADs mainly due to an increased duration of the spike-and-wave part of ADs whereas dizocilpine shortened ADs in a dose-dependent manner affecting both types of ADs. In addition, NBQX did not influence interhemispheric responses meanwhile dizocilpine moderately suppressed these evoked potentials. According to our results, NMDA receptors are important for generation of cortical epileptic afterdischarges meanwhile the role of AMPA receptors is not clear and has to be analyzed.

Topics: Animals; Cerebral Cortex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Electric Stimulation; Electroencephalography; Epilepsy; Evoked Potentials; Excitatory Amino Acid Antagonists; Quinoxalines; Rats; Rats, Wistar

Glutamatergic mossy fibers of the hippocampus sprout in temporal lobe epilepsy and establish aberrant synapses on granule cells from which they originate. There is currently no evidence for the activation of kainate receptors (KARs) at recurrent mossy fiber synapses in epileptic animals, despite their important role at control mossy fiber synapses. We report that KARs are involved in ongoing glutamatergic transmission in granule cells from chronic epileptic but not control animals. KARs provide a substantial component of glutamatergic activity, because they support half of the non-NMDA receptor-mediated excitatory drive in these cells. KAR-mediated EPSC(KA)s are selectively generated by recurrent mossy fiber inputs and have a slower kinetics than EPSC(AMPA). Therefore, in addition to axonal rewiring, sprouting of mossy fibers induces a shift in the nature of glutamatergic transmission in granule cells that may contribute to the physiopathology of the dentate gyrus in epileptic animals.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Disease Models, Animal; Electric Stimulation; Epilepsy; Excitatory Postsynaptic Potentials; Hippocampus; In Vitro Techniques; Male; Mossy Fibers, Hippocampal; Patch-Clamp Techniques; Quinoxalines; Rats; Rats, Wistar; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Synapses

Hypoxia of the brain may alter further seizure susceptibility in a different way. In this study, we tried to answer the question how episode of convulsion induced by hypoxia (HS) changes further seizure susceptibility, and how N-methyl-D-aspartic acid (NMDA) and AMPA/KA receptor antagonists influence this process. Adult Albino Swiss mice exposed to hypoxia (5% O(2)) developed clonic/tonic convulsions after about 340 s. Mice which underwent 10 s but not 5 s seizures episode subsequently exhibited significantly increased seizure susceptibility to low doses (equal ED(16)) of bicuculline (BCC) and NMDA during a 3-week observation period. No morphological signs of brain tissue damage were seen in light microscope on the third day after a hypoxia-induced seizure (HS). Learning abilities assessed in passive avoidance test as well as spontaneous alternation were not disturbed after an HS episode. Pretreatment with AMPA/KA receptor antagonist NBQX effectively prolonged latency to HS and given immediately after seizure episode also attenuated subsequent convulsive susceptibility rise, however, NMDA receptor antagonist, MK-801, appeared to be ineffective. These results suggest that a seizure episode induced by hypoxia, depending on its duration, may play an epileptogenic role. The AMPA/KA receptor antagonist prolongs the latency to HS, and given after this episode, prevents the long-term epileptogenic effect.

Topics: Animals; Avoidance Learning; Bicuculline; Brain Chemistry; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; GABA Antagonists; gamma-Aminobutyric Acid; Hypoxia; Mice; N-Methylaspartate; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Seizures

The intrahippocampal administration of 4-aminopyridine (4-AP) induces epileptic seizures and neurodegeneration, due probably to stimulation of glutamate release from synaptic terminals. We have studied the time course of the neurodegenerative changes produced by 4-AP, perfused through microdialysis cannulas in rat hippocampus, and correlated them with the expression of the inducible heat shock protein 70 (HSP70), detected immunocytochemically. Electroencephalographic seizure activity appeared immediately after the beginning of 4-AP perfusion. The first signs of histological neuronal damage were observed in CA1 and CA3 subfields of the perfused hippocampus 3 h after treatment and progressed until reaching a maximal neuronal loss at 24 h. In 4-AP-treated rats HSP70 was expressed mainly in neurons of the contralateral hippocampus, with a time course and cellular distribution very similar to the neurodegeneration observed in the perfused hippocampus, but no neuronal damage was observed. The N-methyl-D-aspartate (NMDA) receptor antagonists MK-801 and (3-phosphonopropyl)-piperazine-2-carboxylic acid prevented the seizures, the neurodegeneration and the expression of HSP70. These data demonstrate that the 4-AP-induced release of endogenous glutamate overactivates NMDA receptors in the perfused hippocampus and that the resulting neuronal hyperexcitability propagates to the contralateral hippocampus, generating a glutamate-mediated neuronal stress sufficient to induce the expression of HSP70 but not to produce neurodegeneration. These findings provide a useful model for investigating the relationships between neuronal hyperexcitation, neurodegeneration and the role of HSP expression.

Topics: 4-Aminopyridine; Animals; Anticonvulsants; Cell Count; Chromatography, High Pressure Liquid; Dizocilpine Maleate; Drug Interactions; Electroencephalography; Epilepsy; Extracellular Space; Functional Laterality; Gene Expression; Glutamic Acid; Hippocampus; HSP70 Heat-Shock Proteins; Immunohistochemistry; Male; Microdialysis; Nerve Degeneration; Neuroprotective Agents; Piperazines; Potassium Channel Blockers; Quinoxalines; Rats; Rats, Wistar; Somatosensory Cortex; Stress, Physiological; Time Factors

Recent evidence suggests that excessive GABA(A) receptor-mediated transmission can lead to neuronal hyperexcitability and hypersynchrony. We show now that exposure of a rat hippocampal slice to GABA(B) receptor antagonists (CGP 55845A and CGP 35348) in the absence of ionotropic glutamatergic transmission leads to a progressive synchronization of spontaneous interneuronal activity. In about 30% of over 200 slices examined, the GABA(A)-mediated spontaneous activity produced field responses in the CA1 soma region with a positive-going phase of up to 5 mV, followed by a long-lasting negative deflection with a simultaneous extracellular K(+) transient. These bicarbonate-dependent GABAergic ictal-like events (GIEs) were associated with biphasic (hyperpolarizing/depolarizing) intracellular responses and with synchronous bursting of the pyramidal neurons. The GIEs could not be reversed by wash-out of the GABA(B) receptor antagonists or by baclofen, but they were inhibited by agonists acting on presynaptic mu-opioid and cannabinoid (CB1) receptors pointing to a down-regulation of presynaptic GABA(B) receptors. GIEs were dependent on intracellular carbonic anhydrase, and potentiated by maneuvers that increase intracellular pH. They were blocked by the Cx36-specific gap-junction (gj) blocker, quinine/quinidine, as well as by the broad-spectrum gj blocker, octanol. These data suggest that enhanced GABAergic activity with functional interneuronal connectivity via gjs is sufficient to trigger epileptiform activity in the absence of ionotropic glutamatergic transmission.

Topics: 2-Amino-5-phosphonovalerate; Animals; Carbonic Anhydrase Inhibitors; Electric Stimulation; Electrophysiology; Epilepsy; Female; GABA Antagonists; Gap Junctions; Hippocampus; Hydrogen-Ion Concentration; In Vitro Techniques; Interneurons; Male; Membrane Potentials; Organophosphorus Compounds; Patch-Clamp Techniques; Phosphinic Acids; Propanolamines; Quinoxalines; Rats; Rats, Wistar; Receptors, GABA-A; Receptors, GABA-B

Glial cells limit local K(+)-accumulation by K(+)-uptake through different mechanisms, sensitive to Ba(2+), ouabaine, furosemide, or DIDS. Since the relative contribution of these mechanisms has not yet been determined, we studied the effects of bath-applied barium (2 mM), ouabaine (9 microM), furosemide (2 mM), and DIDS (1 mM) on ionophoretically-induced rises in [K(+)](o) in the pyramidal layer of area CA1 from normal rat slices, in the presence of glutamate receptor (Glu-R) antagonists. We also investigated the effect of barium on ionophoretically-induced tetrapropylammonium (TPA(+))-signals in order to test for barium-induced changes of the extracellular space. Finally, we repeated the barium experiment on slices from human non-sclerotic and sclerotic hippocampal specimens to assess a reduced glial capability for barium-sensitive K(+)-uptake in sclerotic tissue from epilepsy patients. In normal rat slices barium augmented ionophoretically-induced rises in [K(+)](o) by approximately 120%, also in the presence of tetrodotoxin (TTX) (by approximately 150%), but did not significantly affect the TPA(+)-signal. Ouabaine also augmented the K(+)-signal, but only by 27%. Furosemide and DIDS had negligible effects. In slices from sclerotic human hippocampus an augmentation of the K(+)-signal by barium was absent. Thus barium augments ionophoretically-induced K(+)-signals to a similar extent as previously shown for stimulus-induced signals. We suggest that glial barium-sensitive K(+)-buffer mechanisms reduce fast local rises of [K(+)](o) by at least 50%. This capability of glial cells is extremely reduced in area CA1 of slices from human sclerotic hippocampal specimens.

Topics: 2-Amino-5-phosphonovalerate; 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Anesthetics, Local; Animals; Barium; Buffers; Diuretics; Enzyme Inhibitors; Epilepsy; Excitatory Amino Acid Antagonists; Extracellular Space; Furosemide; Hippocampus; Humans; In Vitro Techniques; Iontophoresis; Ouabain; Potassium; Quaternary Ammonium Compounds; Quinoxalines; Rats; Rats, Wistar; Sclerosis; Tetrodotoxin

The granule cells of the dentate gyrus (DG) send a strong glutamatergic projection, the mossy fibre tract, toward the hippocampal CA3 field, where it excites pyramidal cells and neighbouring inhibitory interneurons. Despite their excitatory nature, granule cells contain small amounts of GAD (glutamate decarboxylase), the main synthetic enzyme for the inhibitory transmitter GABA. Chronic temporal lobe epilepsy results in transient upregulation of GAD and GABA in granule cells, giving rise to the speculation that following overexcitation, mossy fibres exert an inhibitory effect by release of GABA. We therefore stimulated the DG and recorded synaptic potentials from CA3 pyramidal cells in brain slices from kindled and control rats. In both preparations, DG stimulation caused excitatory postsynaptic potential (EPSP)/inhibitory postsynaptic potential (IPSP) sequences. These potentials could be completely blocked by glutamate receptor antagonists in control rats, while in the kindled rats, a bicuculline-sensitive fast IPSP remained, with an onset latency similar to that of the control EPSP. Interestingly, this IPSP disappeared 1 month after the last seizure. When synaptic responses were evoked by high-frequency stimulation, EPSPs in normal rats readily summate to evoke action potentials. In slices from kindled rats, a summation of IPSPs overrides that of the EPSPs and reduces the probability of evoking action potentials. Our data show for the first time that kindling induces functionally relevant activity-dependent expression of fast inhibition onto pyramidal cells, coming from the DG, that can limit CA3 excitation in a frequency-dependent manner.

Topics: 2-Amino-5-phosphonovalerate; Animals; Epilepsy; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Kindling, Neurologic; Mossy Fibers, Hippocampal; Neural Inhibition; Neural Pathways; Neuronal Plasticity; Pyramidal Cells; Quinoxalines; Rats; Rats, Wistar; Reaction Time

Bilateral intracerebroventricular infusion of dl-homocysteic acid (DL-HCA) (600 nmol on each side) to immature 12-day-old rats induced generalized clonic-tonic seizures, recurring frequently for at least 90 min, with a high rate of survival. Electrographic recordings from sensorimotor cortex, hippocampus, and striatum demonstrated isolated spikes in the hippocampus and/or striatum as the first sign of dl-HCA action. Generalization of epileptic activity occurred during generalized clonic-tonic seizures, but electroclinical correlation was very low; dissociation between EEG pattern and motor phenomena was common. Seizures were accompanied by large decreases of cortical glucose and glycogen and by approximately 7- to 10-fold accumulation of lactate. ATP and phosphocreatine (PCr) levels remained unchanged even during longlasting (3 h) convulsions. Metabolite levels became normalized during the recovery period (24 h). The examination of the effect of selected antagonists of NMDA [AP7 (18.5 and 37 mg/kg, respectively), MK-801 (0.5 mg/kg)] and non-NMDA [NBQX (10, 15 and 30 mg/kg, respectively)] receptors revealed that seizures could be attenuated or prevented (depending on the dose employed) by antagonists of both NMDA and non-NMDA receptors, as evaluated not only according to the suppression of behavioral manifestations of seizures, but also in terms of the protection of metabolite changes accompanying seizures. All antagonists employed, when given alone in the same doses as those used for seizure protection, did not influence metabolite levels, with the exception of increased glucose concentrations. Furthermore, the pronounced anticonvulsant effect could be achieved by the combined treatment with low subthreshold doses of NMDA (AP7) and non-NMDA (NBQX) receptor antagonists, which may be of potential significance for a new approach to the treatment of epilepsy.

Topics: 2-Amino-5-phosphonovalerate; Age Factors; Animals; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Electroencephalography; Energy Metabolism; Epilepsy; Excitatory Amino Acid Antagonists; Glucose; Homocysteine; Injections, Intraventricular; Male; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Seizures

The involvement of alpha-amino-3-hydroxy-5-methylizoxazole-4-propionic acid (AMPA) receptors in induction of long-term potentiation (LTP) was examined in rat hippocampal slice preparation. Using conventional extracellular recording, excitatory postsynaptic potentials (EPSPs) and population action potentials (PSs), evoked by low-frequency stimulation of the Schaffer collateral-commissural fibres, were recorded in the CA1 region. The effects of a competitive AMPA receptor antagonist, 6-nitro-7-sulfamoylbenzo(f)quinoxaline-2, 3-dione (NBQX), and that of a non-competitive blocker, 1-(4-aminophenyl)-4-methyl-7,8-methylendioxy-5H-2,3-benzodiazepine (GYKI 52466) have been examined. 0.25-0.5 microM of NBQX and 20-40 microM of GYKI 52466 did not suppress the induction of LTP. LTP was attenuated only at the highest concentrations tested (1 microM NBQX or 80 microM GYKI 52466). These in vitro concentrations, however, exceed the brain levels needed for in vivo anticonvulsant action. Furthermore, even at the highest concentrations both compounds suppressed only the expression but not the induction of LTP. Namely after their washout LTP reappeared. Thus, at pharmacologically relevant concentrations these AMPA receptor antagonists apparently do not suppress LTP, a cellular mechanism underlying memory formation. These experiments suggest that in clinical practice AMPA receptor blockade may have some advantage over N-methyl-D-aspartate receptor antagonism, which is accompanied by severe memory impairment.

Topics: Animals; Anti-Anxiety Agents; Benzodiazepines; Dose-Response Relationship, Drug; Epilepsy; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Hippocampus; Long-Term Potentiation; Male; Memory; Neurodegenerative Diseases; Neurons; Organ Culture Techniques; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA

Paired intracellular recordings were used to investigate recurrent excitatory transmission in layers II, III and V of the rat entorhinal cortex in vitro. There was a relatively high probability of finding a recurrent connection between pairs of pyramidal neurons in both layer V (around 12%) and layer III (around 9%). In complete contrast, we have failed to find any recurrent synaptic connections between principal neurons in layer II, and this may be an important factor in the relative resistance of this layer in generating synchronized epileptiform activity. In general, recurrent excitatory postsynaptic potentials in layers III and V of the entorhinal cortex had similar properties to those recorded in other cortical areas, although the probabilities of connection are among the highest reported. Recurrent excitatory postsynaptic potentials recorded in layer V were smaller with faster rise times than those recorded in layer III. In both layers, the recurrent potentials were mediated by glutamate primarily acting at alpha-amino-3-hydroxy-5-methyl-4-isoxazole receptors, although there appeared to be a slow component mediated by N-methyl-D-aspartate receptors. In layer III, recurrent transmission failed on about 30% of presynaptic action potentials evoked at 0.2Hz. This failure rate increased markedly with increasing (2, 3Hz) frequency of activation. In layer V the failure rate at low frequency was less (19%), and although it increased at higher frequencies this effect was less pronounced than in layer III. Finally, in layer III, there was evidence for a relatively high probability of electrical coupling between pyramidal neurons. We have previously suggested that layers IV/V of the entorhinal cortex readily generate synchronized epileptiform discharges, whereas layer II is relatively resistant to seizure generation. The present demonstration that recurrent excitatory connections are widespread in layer V but not layer II could support this proposal. The relatively high degree of recurrent connections and electrical coupling between layer III cells may be a factor in it's susceptibility to neurodegeneration during chronic epileptic conditions.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Action Potentials; Animals; Entorhinal Cortex; Epilepsy; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; In Vitro Techniques; Lysine; Male; Neural Pathways; Pyramidal Cells; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission

The entorhinal cortex (EC) is a major gateway for sensory information into the hippocampus and receives a cholinergic input from the forebrain. Therefore, we studied muscarinic effects on excitability and intracellular Ca2+ signalling in layer II stellate and layer III pyramidal projection neurons of the EC. In both classes of neurons, local pressure-pulse application of carbachol (1 mM) caused small, atropine-sensitive membrane depolarizations that were not accompanied by any detectable changes in [Ca2+]i. At a higher concentration (10 mM), carbachol induced a larger membrane depolarization associated with synaptic oscillations and epileptiform activity in both classes of neurons. In contrast to the intrinsic theta rhythm in stellate cells with one dominant peak frequency at approximately 7 Hz, the synaptically mediated oscillation induced by carbachol showed three characteristic peaks in the theta and gamma frequency range at approximately 11, 23 and 40 Hz. Although carbachol-induced epileptiform activity was associated with increases in intracellular free Ca2+ in both layer II and III cells, the observed [Ca2+]i accumulation was significantly larger in layer III than in layer II cells. Responses to intracellular current injections showed differences in Ca2+ accumulation in layer II and III cells at the same membrane potentials, suggesting a dominant expression of low- and high-voltage-activated Ca2+ channels in these layer II and III cells, respectively. In conclusion, we present evidence for significant differences in the [Ca2+]i regulation between layer II stellate and layer III pyramidal cells of the medial EC.

Topics: 2-Amino-5-phosphonovalerate; Action Potentials; Afferent Pathways; Animals; Calcium; Calcium Signaling; Carbachol; Cholinergic Agonists; Electrophysiology; Entorhinal Cortex; Epilepsy; Excitatory Amino Acid Antagonists; Organ Culture Techniques; Periodicity; Pyramidal Cells; Quinoxalines; Rats; Rats, Wistar; Synaptic Transmission; Tetrodotoxin

The anticonvulsant effects of some novel 2,3-benzodiazepines acting as alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid/kainate (AMPA/KA) antagonists were evaluated in genetically epilepsy prone rats. The ED50 values against clonic and tonic seizures (in micromol/kg) revealed that the rank order of anticonvulsant activity was: GYKI 52466 > 2,3BZ-2 > 2,3 MBZ-2 > NBQX. Maximal anticonvulsant protection was observed 15-45 min after the i.p. administration of NBQX and GYKI 52466, 30-90 min after the i.p. administration of 2,3BZ-2, and 45-120 min after the i.p. administration of 2,3MBZ-2. The time course of plasma levels of rats treated with GYKI 52466 showed that peak plasma concentration was observed 15 min after i.p. administration, 2,3BZ-2 revealed that peak plasma concentration was achieved 45 min after i.p. administration, whereas following 2,3MBZ-2 administered i.p., two curves were detected; one is referred to the parent compound and the other to its demethylate metabolite that corresponds to 2,3BZ-2. The therapeutic index (ratio of TD50 values for impaired rotarod performance and ED50 values for anticonvulsant activity) revealed that NBQX and GYKI 52466 were slighly more toxic than 2,3BZ-2 and 2,3MBZ-2. The present data suggest that 2,3-benzodiazepines acting at AMPA/kainate receptors play an important role in the generation and/or propagation of the audiogenic seizures in genetically epilepsy-prone rats.

Topics: Animals; Anti-Anxiety Agents; Anticonvulsants; Benzodiazepines; Epilepsy; Motor Activity; Motor Skills Disorders; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA

The molecular basis for developing symptomatic epilepsy (epileptogenesis) remains ill defined. We show here in a well characterized hippocampal culture model of epilepsy that the induction of epileptogenesis is Ca2+-dependent. The concentration of intracellular free Ca2+ ([Ca2+]i) was monitored during the induction of epileptogenesis by prolonged electrographic seizure activity induced through low-Mg2+ treatment by confocal laser-scanning fluorescent microscopy to directly correlate changes in [Ca2+]i with alterations in membrane excitability measured by intracellular recording using whole-cell current-clamp techniques. The induction of long-lasting spontaneous recurrent epileptiform discharges, but not the Mg2+-induced spike discharges, was prevented in low-Ca2+ solutions and was dependent on activation of the N-methyl-D-aspartate (NMDA) receptor. The results provide direct evidence that prolonged activation of the NMDA-Ca2+ transduction pathway causes a long-lasting plasticity change in hippocampal neurons causing increased excitability leading to the occurrence of spontaneous, recurrent epileptiform discharges.

Topics: 2-Amino-5-phosphonovalerate; Animals; Animals, Newborn; Benzoates; Calcium; Cells, Cultured; Dizocilpine Maleate; Egtazic Acid; Epilepsy; Excitatory Amino Acid Antagonists; Glycine; Hippocampus; Magnesium; Membrane Potentials; Microscopy, Confocal; Neurons; Nifedipine; Patch-Clamp Techniques; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction

The aim of this study was to assess whether a drug which combines an antagonistic action at both NMDA and non-NMDA receptors offers advantages for treatment of epileptic seizures compared to drugs which antagonize only one of these ionotropic glutamate receptors. The novel glutamate receptor antagonist LU 73068 (4,5-dihydro-1-methyl-4-oxo-7-trifluoromethylimidazo[1,2a]quinoxal ine-2-carbonic acid) binds with high affinity to both the glycine site of the NMDA receptor (Ki 185 nM) and to the AMPA receptor (Ki 158 nM). Furthermore, binding experiments with recombinant kainate receptor subunits showed that LU 73068 binds to several of these subunits, particularly to rGluR7 (Ki 104 nM) and rGluR5 (Ki 271 nM). In comparison, the prototype non-NMDA receptor antagonist NBQX (2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo[f]quinoxaline) binds with high affinity to AMPA receptors only. Both NBQX and LU 73068 were about equieffective after i.p. injection in mice to block lethal convulsions induced by AMPA or NMDA. In the rat amygdala kindling model of temporal lobe epilepsy, LU 73068 dose-dependently increased the focal seizure threshold (afterdischarge threshold, ADT). When rats were stimulated with a current 20% above the individual control ADT, LU 73068 completely blocked seizures with an ED50 of 4.9 mg kg(-1). Up to 20 mg kg(-1), only moderate adverse effects, e.g. slight ataxia, were observed. NBQX, 10 mg kg(-1), and the glycine/NMDA site antagonist L-701,324 (7-chloro-4-hydroxy-3-(3-phenoxy)phenyl-quinoline-2(1H)one), 2.5 or 5 mg kg(-1), exerted no anticonvulsant effects in kindled rats when administered alone, but combined treatment with both drugs resulted in a significant ADT increase. The data indicate that combination of glycine/NMDA and non-NMDA receptor antagonism in a single drug is an effective means of developing a potent and effective anticonvulsant agent.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anticonvulsants; Disease Models, Animal; Dizocilpine Maleate; Drug Synergism; Epilepsy; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Imidazoles; Kindling, Neurologic; Male; Mice; Mice, Inbred Strains; N-Methylaspartate; Quinolones; Quinoxalines; Receptors, AMPA; Receptors, Glycine; Receptors, N-Methyl-D-Aspartate; Tritium

Intrahippocampal perfusion of bicuculline (50 microM) in Mg2+-free medium caused elevation of extracellular cGMP and epileptic-like behaviour. Both effects were partially prevented by blocking NMDA receptors with MK-801 or Mg2+ ions. Similarly, the GABA(B) receptor antagonists CGP52432 (0.1-30 microM) and CGP35348 (0.3-1 mM) evoked increases of extracellular cGMP. CGP52432 also elicited behavioural responses ranging from wet dog shakes to convulsions. MK-801 or Mg2+ ions reduced the effects of CGP52432. Local application of muscimol (100-300 microM) or (-)baclofen (300 microM) caused inhibition of extracellular cGMP. Administration of the AMPA/kainate receptor antagonist NBQX (100 microM) caused cGMP elevation which was almost abolished by co-perfusion of muscimol and (-)baclofen. In the presence of physiological Mg2+, perfusion of AMPA (30 microM) failed to affect cGMP levels, although rats displayed wet dog shakes episodes. When AMPA was co-perfused with low concentrations of bicuculline or CGP52432, cGMP elevations were observed in 60% of the rats. Addition of both antagonists to AMPA resulted in 85% of rats displaying a cGMP response. To conclude: (a) extracellular hippocampal cGMP is controlled by inhibitory GABA(A) and GABA(B) receptors tonically activated through GABAergic interneurons receiving AMPA/kainate-mediated glutamatergic inputs; (b) the GABAergic receptors are not endogenously saturated and can be further stimulated by exogenous agonists; (c) blockade of the GABA-mediated inhibition causes increase of cGMP and epileptic-like behaviour, due largely to endogenous activation of NMDA receptors; (d) reproducible cGMP responses to AMPA can be observed when the inhibitory GABAergic inputs to the NO/guanylyl cyclase system are blocked, confirming the previously proposed existence of AMPA/kainate receptors able to increase the nucleotide synthesis.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Behavior, Animal; Cyclic GMP; Epilepsy; Excitatory Amino Acid Agonists; gamma-Aminobutyric Acid; Hippocampus; Microdialysis; Nitric Oxide; Quinoxalines; Rats; Receptors, AMPA; Receptors, GABA-A; Receptors, GABA-B; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate

Clinically, and in experimental models, perinatal hypoxic encephalopathy is commonly associated with seizures. We previously described a rat model in which hypoxia induces seizures and permanently increases in seizure susceptibility in immature rats [postnatal day (P) 10-12] but not in older rats. In the present study, we compared the effect of pretreatment with the excitatory amino acid antagonists MK-801 and NBQX versus lorazepam in our rat model of perinatal hypoxia. Animals exposed to hypoxia at P10 without treatment have frequent seizures during hypoxia and subsequently exhibit increased seizure susceptibility to flurothyl. Treatment with 6-nitro-7-sulfamoylbenzo(f)quinoxaline-2,3-dione (NBQX 20 mg/kg) effectively suppressed hypoxia-induced seizures in immature rats and also protected against permanent changes in flurothyl threshold in adulthood, whereas treatment with MK-801 (1 mg/kg) or lorazepam (LZP 1 mg/kg) did not prevent these hypoxia-related epileptogenic effects. These results suggest that activation of alpha-amino-3-hydroxy-5-methyl-4-isoxazol propionic acid (AMPA) receptors may partly mediate the age-dependent epileptogenic effect of hypoxia in the perinatal period.

Topics: Age Factors; Animals; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Antagonists; Flurothyl; Hypoxia, Brain; Lorazepam; Male; Quinoxalines; Rats; Receptors, AMPA

To investigate the role of non-NMDA receptors in epileptic seizures, we examined the antiepileptogenic and anticonvulsant effects of NBQX (2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)-quinoxaline), a potent and selective AMPA receptor antagonist, in the rat kindling model. Systemic administration of 10-40 mg/kg NBQX significantly and dose dependently suppressed previously kindled seizures from the amygdala (AM), assessed in terms of the motor seizure stage and afterdischarge (AD) duration. The maximal effects were observed at 0.5-1 h after drug injection. When the intensity of electrical stimulation was increased to twice the generalized seizure-triggering threshold (GST), the anticonvulsant effects of NBQX on AM-kindled seizures were not reversed, suggesting that the effects were not due to non-specific elevation of the GST. In contrast to AM-kindled seizures, 20-40 mg/kg NBQX significantly suppressed only the motor seizure stage without reducing the AD duration of previously hippocampal-kindled seizures. Daily administration of 15 or 30 mg/kg NBQX prior to each electrical stimulation of the AM markedly and significantly suppressed the development of kindling. During drug sessions, the growth of the AD duration was blocked almost completely, while the waveform of ADs became more complex. These results indicate that NBQX has potent antiepileptogenic and anticonvulsant actions on kindling, at least from the AM and that non-NMDA receptors have an important role in seizure propagation.

Topics: Amygdala; Animals; Anticonvulsants; Dose-Response Relationship, Drug; Electric Stimulation; Electroencephalography; Epilepsy; Hippocampus; Kindling, Neurologic; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Time Factors

Sound-induced seizures in genetically epilepsy-prone rats were used to compare the anticonvulsant effect of phenytoin and diazepam with compounds which decrease glutamatergic neurotransmission including excitatory amino acid antagonists acting at N-methyl-D-aspartate (NMDA) receptors: D(-)CPPene, CGP 37849 and MK 801 or at the glycine/NMDA site: ACPC (1-aminocyclopropane-dicarboxylic acid) (partial agonist) or non-NMDA receptors: NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo[f]-quinoxaline.Li) and GYKI 52466 (1-(aminophenyl)-4-methyl-7,8-methylene-dioxy-5H-2,3-benzodiazepin e.HCl) or acting at sodium channels to decrease glutamate release: lamotrigine and BW 1003C87 (5(2,3,5-trichlorophenyl)-2,4-diaminopyrimidine ethane sulphonate). ED50 values against clonic seizures (in mumol/kg at the time of peak anticonvulsant effect) were: phenytoin 30.5 (2 h), diazepam 0.5 (0.5 h), MK 801 0.01 (4 h), D(-)CPPene 1.9 (4 h), CGP 37849 2 (1 h), GYKI 52466 24 (0.25 h), NBQX 40 (0.5 h), ACPC 1053 (0.5 h), BW 1003C87 2.2 (1 h), lamotrigine 4.8 (4 h). BW 1003C87, lamotrigine, MK 801, phenytoin, diazepam and CGP 37849 had the most favourable therapeutic indices (rotarod locomotor deficit ED50/anticonvulsant ED50).

Topics: 2-Amino-5-phosphonovalerate; Acoustic Stimulation; Amino Acids; Amino Acids, Cyclic; Analysis of Variance; Animals; Anticonvulsants; Behavior, Animal; Diazepam; Disease Models, Animal; Dizocilpine Maleate; Epilepsy; Female; Lamotrigine; Male; Motor Activity; Phenytoin; Pyrimidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Time Factors; Triazines

Using a grease-seal technique on cortical slices, methohexitone (10-316 microM) dose dependently and reversibly reduced depolarising responses to kainate more than those to alpha-amino-3-hydroxy-4-isoxazolepropionate (AMPA) and N-methyl-D-aspartate (NMDA). The respective pA2 values were 4.9 +/- 0.07, 3.6 +/- 0.03 and 4.0 +/- 0.05 whereas, for 6-nitro,7-sulphamoylbenz[F]quinoxalinedione (NBQX), they were 5.8 +/- 0.06, 6.7 +/- 0.05 and < 4.0. Methohexitone was also more effective than NBQX in reducing the spontaneous epileptiform activity occurring in these cortical slices. Thus 10 and 20 microM of this short-acting barbiturate reduced afterpotentials and burst frequencies respectively by about 50% whereas NBQX 10 microM only reduced burst frequency by some 15%. The results are discussed with respect to a putative methohexitone- and kainate-sensitive autoreceptor which facilitates presynaptic glutamate release.

Topics: Action Potentials; Animals; Cerebral Cortex; Dose-Response Relationship, Drug; Epilepsy; gamma-Aminobutyric Acid; In Vitro Techniques; Kainic Acid; Methohexital; Quinoxalines; Rats; Receptors, Glutamate; Receptors, Kainic Acid

Two potent glutamate antagonists, NBQX and GYKI 52466, that act selectively on non-NMDA receptors, have been tested for anticonvulsant activity in 3 models of reflex epilepsy (sound-induced seizures in DBA/2 mice and in genetically epilepsy-prone rats and photically-induced myoclonus in Papio papio) and in amygdala kindled rats. Both compounds potently but transiently suppress reflexly-induced epileptic responses. GYKI 52466 also reduces behavioral seizures and afterdischarge duration in amygdala kindled rats, but with a lower potency than it suppresses reflex epilepsy. These data are similar to earlier results with antagonists acting selectively on NMDA receptors; they do not support a specific involvement of enhanced AMPA receptor sensitivity as a major factor in the expression of kindled seizures.

Topics: Acoustic Stimulation; Amygdala; Animals; Anti-Anxiety Agents; Anticonvulsants; Benzodiazepines; Cerebral Cortex; Electroencephalography; Epilepsies, Myoclonic; Epilepsy; Evoked Potentials; Excitatory Amino Acid Antagonists; Hippocampus; Kindling, Neurologic; Mice; Mice, Inbred DBA; Neural Pathways; Papio; Photic Stimulation; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, AMPA; Receptors, Glutamate; Reflex; Synaptic Transmission

The effect of i.p. or i.v. administration of the non-N-methyl-D-aspartate antagonists, GYKI 52466 (1-(4-aminophenyl)-4-methyl-7,8-methylendioxy-5H-2,3-benzodiazepin e.HCl, molecular weight 330) and NBQX (2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)-quinoxaline, molecular weight 342) on sound-induced seizures in rats and photically induced myoclonus in baboons was studied. In both species an anticonvulsant effect occurred 15-60 min after administration of GYKI 52466 or NBQX. The ED50 value for clonic seizure suppression for GYKI 52466 at 30 min was 39 (rats, i.p.) and at 15 min was 13 (Papio papio, i.v.) mumol kg-1 and for NBQX at 30 min was 40 (rats, i.p.) and at 15 min approximately 10 (Papio papio, i.v.) mumol kg-1. Side effects were not observed in rats; apparent side effects in baboons probably arose from drug formulation. The anticonvulsant actions of GYKI 52466 and NBQX suggest a possible role for non-NMDA antagonists in the therapy of epilepsy.

Topics: Acoustic Stimulation; Animals; Anti-Anxiety Agents; Anticonvulsants; Behavior, Animal; Benzodiazepines; Dose-Response Relationship, Drug; Epilepsies, Myoclonic; Epilepsy; Epilepsy, Tonic-Clonic; Female; Male; Papio; Photic Stimulation; Quinoxalines; Rats; Rats, Inbred Strains; Reflex

Kindling, induced by repeated subconvulsive electrical or chemical stimulations leads to progressive and permanent amplification of seizure activity, culminating in generalized seizures. We report that kindling induced by electrical stimulation in the ventral hippocampus leads to a marked and transient increase in mRNA for NGF and BDNF in the dentate gyrus, the parietal cortex, and the piriform cortex. BDNF mRNA increased also in the pyramidal layer of hippocampus and in the amygdaloid complex. No change was seen in the level of HDNF/NT-3 mRNA. The increased expression of NGF and BDNF mRNAs was not influenced by pretreatment with the NMDA receptor antagonist MK801, but was partially blocked by the quisqualate, AMPA receptor antagonist NBQX. The presumed subsequent increase of the trophic factors themselves may be important for kindling-associated plasticity in specific neuronal systems in the hippocampus, which could promote hyperexcitability and contribute to the development of epileptic syndromes.

Topics: Animals; Brain; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Epilepsy; Kindling, Neurologic; Male; Nerve Growth Factors; Nerve Tissue Proteins; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; RNA, Messenger