Glutamatergic and purinergic mechanisms on respiratory modulation in the caudal NTS of awake rats

In the present study we evaluated the role of ionotropic glutamate receptors and purinergic P2 receptors in the caudal commissural NTS (cNTS) on the modulation of the baseline respiratory frequency (fR), and on the tachypneic response to chemoreflex activation in awake rats. The selective antagonism of ionotropic glutamate receptors with kynurenic acid (2 nmol/50 nl) in the cNTS produced a significant increase in the baseline fR but no changes in the tachypneic response to chemoreflex activation. The selective antagonism of purinergic P2 receptors by PPADS (0.25 nmol/50 nl) in the cNTS produced no changes in the baseline fR or in the tachypneic response to chemoreflex activation. The data indicate that glutamate acting on ionotropic receptors in the cNTS plays a inhibitory role on the modulation of the baseline fR but had no effect on the tachypneic response to chemoreflex activation, while ATP acting on P2 receptors in the cNTS plays no major role in the modulation of the baseline fR or in the tachypneic response to chemoreflex activation. We suggest that neurotransmitters other than L-glutamate and ATP are involved in the processing of the tachypneic response of the chemoreflex at the cNTS level. (C) 2008 Elsevier B.V. All rights reserved.

Involvement of hypothalamic paraventricular nucleus non-N-methyl-d-aspartate receptors in the pressor response to noradrenaline microinjected into the bed nucleus of the stria terminalis of unanesthetized rats

Microinjection of noradrenaline into the bed nucleus of the stria terminalis (BST) has been reported to cause a pressor response in unanesthetized rats, which was shown to be mediated by acute vasopressin release into the systemic circulation. In the present study we verified the involvement of magnocellular neurons of the hypothalamic paraventricular (PVN) or supraoptic (SON) nuclei and the local neurotransmitter involved in the pressor response to noradrenaline microinjection into the BST. The PVN pretreatment with the non-selective neurotransmission blocker CoCl(2) (1 nmol/100 nL) inhibited the noradrenaline-evoked pressor response. However, responses were not affected by SON treatment with CoCl(2). Further experiments were carried out to test if glutamatergic neurotransmission in the PVN mediates the pressor response evoked by noradrenaline microinjection into the BST. Pretreatment of the PVN with the selective N-methyl-d-aspartate (NMDA) receptor antagonist LY235959 (2 nmol/100 nL) did not affect the noradrenaline-evoked pressor response. However, PVN pretreatment with the selective non-NMDA receptor antagonist NBQX (2 nmol/100 nL) significantly reduced the pressor response to noradrenaline microinjection into the BST. In conclusion, our results suggest that pressor responses to noradrenaline microinjection into the BST are mediated by PVN magnocellular neurons without involvement of SON neurons. They also suggest that a glutamatergic neurotransmission through non-NMDA glutamate receptors in the PVN mediates the response.

Ionotropic glutamatergic receptors in the rostral medullary raphe modulate hypoxia and hypercapnia-induced hyperpnea

It has been suggested that the medullary raphe (MR) plays a key role in the physiological responses to hypoxia and hypercapnia. We assessed the role of ionotropic glutamate receptors in the rostral MR (rMR) in the respiratory responses to hypoxia and hypercapnia by measuring pulmonary ventilation (V(E)) and body temperature (Tb) of male Wistar rats before and after microinjecting Kynurenic acid (KY, an ionotropic glutamate receptors antagonist, 0.1 mM) into the rMR followed by 60 min of hypoxia (7% O(2)) or hypercapnia exposure (7% CO(2)). Compared to the control group, the ventilatory response to hypoxia was attenuated in animals treated with KY intra-rMR, however the ventilatory response to hypercapnia increased significantly. No differences in Tb among groups were observed during hypoxia or hypercapnia. These data suggest that the glutamate acting on ionotropic receptors in the rMR exerts an excitatory modulation on hyperventilation induced by hypoxia but an inhibitory modulation on the hypercapnia-induced hyperpnea. (C) 2010 Elsevier B.V. All rights reserved.

Ethanol and gene expression in brain

This article represents the proceedings of a symposium at the 2000 ISBRA Meeting in Yokohama, Japan. The chairs were Izuru Matusmoto and Peter A. Wilce. The presentations were (1) GABA receptor subunit expression in the human alcoholic brain, by Tracey Buckley and Peter Dodd; (2) NMDAR gene expression during ethanol addiction, by Jorg Puzke, Rainer Spanagel, Walther Zieglgansberger, and Gerald Wolf; (3) Differentially expressed gene in the nucleus accumbens from ethanol-administered rat, by Shuangying Leng; (4) Expression of a novel gene in the alcoholic brain, by Peter A. Wilce; and (5) Investigations of haplotypes of the dopamine Da-receptor gene in alcoholics, by Hans Rommelspacher, Ulrich Finckh, and Lutz G. Schmidt.

Ethanol-related adaptive changes and physical dependence in rats after exposure to ethanol

Adaptive changes that occur after chronic exposure to ethanol are an important component in the development of physical dependence. We have focused our research on ethanol-induced changes in the expression of several genes that may be important in adaptation. In this article, we describe adaptive changes at the level of the N-methyl-D-aspartate receptor, in the protein expression and activity of the Egr transcription factors, and in the expression of a novel gene of unknown function. (C) 2001 Elsevier Science Inc. All rights reserved.

Identification of neuronal alterations induced by Shank3 mutations using iPS cells from Phelan-McDermid Patients' Fibroblasts

A família de proteínas Shank é o principal conjunto de proteinas de suporte e está localizada na densidade pós-sináptica das sinapses excitatórias. Existem 3 genes na família Shank, Shank1, Shank2 e Shank3 e são caracterizados por múltiplos domínios repetidos de anquirina próximo ao N-terminal seguido pelos domínios Src homologo 3 e PDZ, uma região longa rica em prolina e um domínio de motivo α estéril próximo ao C-terminal. Shank proteínas conectam duas subunidades de receptors glutamatérgicos, recetores NMDA e recetores metabotrópicos de glutamato do tipo-I (mGluRs). O domínio PDZ da Shank conecta-se ao C-terminal do GKAP e este, liga-se, ao complexo recetor PSD-95-NMDA. Por outro lado, a proteína Homer interage com o domínio rico em prolina para confirmar a associação entre a proteína Shank com o mGluR tipo-I. A proteína específica em estudo, Shank3, é haploinsuficiente em pacientes com sindrome Phelan-McDermid devido à deleções no braço comprido do cromossoma 22 levando à danos intelectuais, ausência ou atraso no discurso, comportamentos semelhantes ao autismo, hipotonia e características dismórficas. Neste trabalho, investigamos o papel da Shank3 na função sináptica para compreender a relação entre alterações nesta proteína e as características neurológicas presente em Pacientes com síndrome Phelan-McDermid. Foram utilizados dois modelos diferentes, ratinhos knockout Shank3 e hiPSC de pacientes com PMS. Ratinhos geneticamente modificados são ferramentas uteis no estudo de genes e na compreensão dos mecanismos que experiências in vitro não são capazes de reproduzir, mas de maneira a compreender melhor as patologias humanas, decidimos trabalhar também com células humanas. Os fibroblastos dos pacientes com síndrome Phelan-McDermid fora reprogramados em hiPS cells, diferenciados em neurónios e comparados com os neurónios obtidos a partir de doadores saudavéis e da mesma idade. A reprogramação em iPSC foi realizada por infecção de lentivirus com quatro genes de reprogramação OCT4, c-MYC, SOX2 e KFL4 para posteriormente serem diferenciados em neurónios, com cada passo sendo positivamente confirmado através de marcadores neuronais. Através dos neurónios diferenciados, analisamos a expressão de proteínas sinápticas. Pacientes com haploinsuficiencia na proteína Shank3 apresentam níveis elevados de proteína mGluR5 e decrescidos de proteína Homer sugerindo que a haploinsuficiencia leva a desregulação do complexo mGluR5-Homer-Shank3 conduzindo também, a defeitos na maturação sináptica. Assim, a expressão da proteína mGluR5 está alterada nos pacientes com PMS podendo estar relacionada com defeitos encontrados na diferenciação neuronal e maturação sináptica observados nos neurónios de pacientes. Conclusivamente, iPS cells representam um modelo fundamental no estudo da proteína Shank3 e a sua influência no sindrome de Phelan-McDermid.

Hipersonolência Diurna. Narcolepsia, Qual a Etiologia?

Introdução: A narcolepsia é uma doença do sono REM com desregulação do ciclo de sono-vigília, consequente sonolência diurna e eventual associação a alucinações hipnagógicas, paralisia do sono e cataplexia. A sua prevalência é de 0,05 a 0,02% no adulto mas desconhecida na idade pediátrica. Caso clínico: Criança de seis anos, previamente saudável com sonolência excessiva até 18 horas/dia e discinésia oromandibular, desequilíbrio na marcha e movimentos coreiformes dos membros superiores. Duas semanas antes realizara vacinação para a gripe pandémica. Registou-se ainda hiperfagia diurna e nocturna durante cinco dias com resolução espontânea, episódios de cataplexia perante riso e alterações emocionais e tremor da cabeça e dos membros superiores com melhoria clínica progressiva após oito dias. Realizou RMN-CE e EEG sem alterações. O exame líquido céfalo-raquidiano e PCR para painel de vírus herpes, Mycoplasma pneumoniae e enterovírus negativas. Nesta fase realizou polissonografia com teste de latências múltiplas do sono (TLMS) sem alterações. Exame cultural do exsudado faríngeo, TASO e anticorpo AntiDnase B negativos. Da exaustiva investigação que realizou apresentava serologias ELISA e WB compatíveis com infecção por Borrelia burdorferi, pelo que cumpriu ceftriaxone 14 dias. Serologias para influenza A mostraram IgM 39 UA/mL com IgG 194 UA/mL com segunda amostra com IgM 43 UA/mL e IgG 162 UA/mL (VR IgM<20;IgG<20). O estudo da autoimunidade revelou ANA 1/320, anticorpos anticardiolipina e antinucleares extraíveis negativos. Restantes autoanticorpos e doseamento de complemento normal. Anticorpos Anti-NMDA e VKCG negativos. Doseamento de hipocretina muito diminuído com HLA DR2 e DQB1*0602 presentes. A polissonografia com TLMS, sete meses após a primeira, confirmou sonolência excessiva com quatro inícios do sono REM sugestivos de narcolepsia. Faz terapêutica com metilfenidato, a sonolência diurna diminuiu e cumpre o seu horário escolar sem limitações. Comentários: O diagnóstico de narcolepsia foi sugerido pela clínica e confirmado pelo teste de latências múltiplas. O valor de hipocretina diminuído pode sugerir uma etiologia autoimune. Uma infecção como a borreliose ou a vacinação prévia para H1N1, responsabilizada por outros casos de narcolepsia podem ter sido desencadeantes de uma alteração imunitária responsável pela doença, nesta criança com a susceptibilidade HLA DR2 e DBQ1*0602.

Eficácia da memantina na doença de Alzheimer em seus estágios moderado a grave

INTRODUÇÃO: O objetivo desta revisão de literatura é avaliar a eficácia, a segurança e a tolerabilidade da memantina, um antagonista não-competitivo do receptor N-metil-D-aspartato (NMDA), no tratamento da doença de Alzheimer em seus estágios moderado a grave. MÉTODOS: Realizou-se uma busca no banco de dados MEDLINE com as palavras-chave memantine e Alzheimer's disease, sendo inseridos apenas ensaios clínicos randomizados, duplo-cegos e controlados com placebo. RESULTADOS: Foram incluídos quatro estudos que preenchiam os critérios de inclusão supracitados, realizados com pacientes portadores de doença de Alzheimer com grau moderado a grave. Todos os estudos indicaram um efeito benéfico da memantina com relação ao placebo para os seguintes parâmetros: melhora da capacidade funcional e maior participação nas atividades diárias. Dois estudos evidenciaram melhora cognitiva. A duração média dos estudos foi de 28 semanas e as doses mais eficazes variaram de 10 a 20mg/dia. Os efeitos adversos, em todos os estudos, foram maiores no grupo placebo. DISCUSSÃO: Apesar de ser uma droga nova e ainda de custo elevado, a memantina parece reduzir os custos totais e o tempo gasto do cuidador, além de produzir melhora global do paciente, gerando melhor qualidade de vida tanto para o paciente quanto para o cuidador. Estudos ainda não publicados, contudo, sugerem que o impacto dessa droga nos estágios mais avançados da demência de Alzheimer seja marginal.

La plasticidad sináptica en hipocampo como posible sustrato neurobiológico del desarrollo de tolerancia rápida a benzodiazepinas

La administración crónica de bezodiazepinas induce tolerancia a sus efectos sedativos, como así también los que se observan sobre las alteraciones motoras y a su efecto como relajante muscular. Es de destacar que el desarrollo de tolerancia a las distintas acciones mencionadas, se desarrolla de manera rápida. Recientemente distintos autores han descripto que la administración previa de antagonistas de los receptores al glutámico de tipo N-metil-D-aspartato (NMDA) es capaz de revertir el desarrollo de tolerancia al efecto hipolocomotor de diazepam (DZ). Resultados recientes de este laboratorio han demostrado que durante el desarrollo de tolerancia al efecto hipolocomotor de DZ se produce un incremento de la plasticidad sináptica en el hipocampo de estos animales. Ha sido sugerido que un fenómeno asociativo o de aprendizaje, subyace al desarrollo de tolerancia a benzodiazepina como así también al desarrollo de tolerancia a otras drogas psicoactivas. Los receptores de glutamato, involucrados en la prevención del desarrollo a la tolerancia a DZ, como así también a otras drogas psicoactivas, es un tipo de receptores al glutámico, llamado NMDA. Estos receptores han sido relacionados con la plasticidad neural y conductual. Más aún, se ha propuesto que proponen que la tolerancia y la dependencia a opiáceos depende de la activación de receptores NMDA. Objetivos del proyecto: * Demostrar que en el desarrollo rápido de tolerancia al efecto hipolocomotor de DZ participan elementos que demuestran la existencia de un proceso de aprendizaje. * Determinar si el incremento de la plasticidad sináptica observada en el hipocampo de animales en los cuales se desarrolló tolerancia al efecto hipolocomotor de DZ, puede ser el sustrato neurobiológico del proceso de aprendizaje postulado.

Efectos de las situaciones estresantes sobre la ansiogenésis y el consumo de sustancias: interacciones con las estrategias cognitivo-comportamentales y las drogas antidepresivas

El presente proyecto desarrolla algunas hipótesis acerca de una de las emociones más primitivas: la ansiedad. Su influencia en los trastornos mentales está ampliamente reconocida, aunque sus causas y mecanismos están aún bajo intentos científicos de comprensión. Por otra parte intenta establecer relaciones entre la ansiedad y el consumo oral de drogas ansiolíticas. El proyecto está diseñado para estudiar la ansiedad en un modelo animal. Plantea por un lado, un procedimiento para inducir ansiedad y por otro un test de ansiedad independiente de la situación de inducción de la ansiedad, cuyo objetivo es hacer más válido el modelo, ya que en los seres humanos la ansiedad se mantiene independientemente de las situaciones. El diseño experimental en primera instancia permite la evaluación de variables tanto psicológicas como neurobiológicas y también la medición del consumo oral de benzodiacepinas. Se prevé también la utilización de drogas antidepresivas para estudiar su efecto per se y la interacción de las variables psicológicas. Objetivos * Evaluar los niveles diferenciales de ansiedad en sujetos expuestos a situaciones estresantes con distintos grados de predectibilidad y controlabilidad. * Evaluar la posible influencia de las estrategias cognitivo-comportamentales sobre la ansiedad. * Determinar la relación entre ansiedad y consumo de ansiolíticos. * Estudiar comparativamente los efectos de drogas antidepresivas sobre los cambios en la ansiedad y el consumo de ansiolíticos inducidos por estrés. * Utilizar herramientas farmacológicas como los antagonistas de los receptores NMDA, los cuales afectan el establecimiento de las estrategias cognitivo-comportamentales, para confirmar la posible influencia de las mencionadas estrategias sobre la ansiedad y el consumo de ansiolíticos.

Bases neurobiológicas del efecto de grelina sobre ansiedad, memoria e ingesta. Neurobiological mechanisms of ghrelin effect on anxiety, memory and food intake.

El objetivo de este proyecto es investigar el sustrato neurobiológico que subyace a los efectos centrales de grelina (Gr) en estructuras extrahipotalámicas tales como núcleo dorsal del rafe (NDR), hipocampo(Hi) y amígdala(Am) donde hemos demostrado que el péptido incrementa la memoria e ingesta, Los mecanismos neurales, neurotrasmisores(nt), segundos mensajeros, etc., involucrados en estos procesos fisiológicos, inducidos por el péptido, necesitan aun ser esclarecidos. Hemos demostrado que grelina incrementa la retención de la memoria cuando es inyectado en Hi, NDR y Am. También incrementa la ingesta al ser administrado en Hi y NDR pero no así en Am. En Hi los efectos de Gr sobre la memoria se correlacionan con incremento en los niveles tisulares de óxido nítrico (NO) y con la disponibilidad del nt 5-HT. En lo que a electrofisiología se refiere hemos demostrado que Gr disminuye el umbral para generar potenciación a largo plazo (LTP). A fin de aportar nuevas evidencias que contribuyan a esclarecer los efectos del péptido sobre memoria e ingesta utilizaremos estudios conductuales, determinaciones bioquímicas y determinaciones electrofisiológicas. En lo que a ingesta se refiere intentaremos esclarecer el papel de los núcleos central y basolateral de la Am en aspectos hedónicos de la ingesta inducida por Gr En esta etapa, más específicamente nos proponemos:1) Determinar si los efectos de grelina sobre ingesta y memoria demostrados en hipocampo y NDR después de la su administración se correlacionan con modificaciones en la liberación de serotonina utilizando cortes de hipocampo precargados con 5HT tritiada en presencia y ausencia del péptido.2) Evaluar si el incremento de óxido nítrico inducido por grelina en hipocampo se correlaciona con cambios en la expresión de nNOS, utilizando Western-blot y la importancia de NOS/NO en la acción de grelina repitiendo los experimentos previo tratamiento de las ratas con inhibidores de NOS. 3)Estudiar la participación del nt glutamato en los efectos hipocampales de Gr sobre la memoria. Analizando a) si grelina modifica la liberación del nt a partir de sinaptosomas aislados de hipocampo de ratas pretratadas con Gr.b) la participación de los receptores NMDA y GABAa en los efectos de grelina previo bloqueo farmacológico del mencionado receptor y el test step down.c) la participación de los receptores NMDA y GABAa en los efectos de grelina utilizando electrofisiología y Western Blot. 4) Estudiar el efecto de la administración de Gr en Amigdala Central y Basolateral sobre aspectos hedónicos de la ingesta utilizando diferentes paradigmas conductuales en animales. Estudiaremos:a) si Gr modifica el consumo de alimento de diferente palatabilidad en animales y si afecta el componente motivacional de la conducta de ingesta paradigma de "runway".

Separate neuronal and glial Na+,K+-ATPase isoforms regulate glucose utilization in response to membrane depolarization and elevated extracellular potassium.

The role of cell type-specific Na+,K+-ATPase isozymes in function-related glucose metabolism was studied using differentiated rat brain cell aggregate cultures. In mixed neuron-glia cultures, glucose utilization, determined by measuring the rate of radiolabeled 2-deoxyglucose accumulation, was markedly stimulated by the voltage-dependent sodium channel agonist veratridine (0.75 micromol/L), as well as by glutamate (100 micromol/L) and the ionotropic glutamate receptor agonist N-methyl-D-aspartate (NMDA) (10 micromol/L). Significant stimulation also was elicited by elevated extracellular potassium (12 mmol/L KCl), which was even more pronounced at 30 mmol/L KCl. In neuron-enriched cultures, a similar stimulation of glucose utilization was obtained with veratridine, specific ionotropic glutamate receptor agonists, and 30 mmol/L but not 12 mmol/L KCl. The effects of veratridine, glutamate, and NMDA were blocked by specific antagonists (tetrodotoxin, CNQX, or MK801, respectively). Low concentrations of ouabain (10(-6) mol/L) prevented stimulation by the depolarizing agents but reduced only partially the response to 12 mmol/L KCl. Together with previous data showing cell type-specific expression of Na+,K+-ATPase subunit isoforms in these cultures, the current results support the view that distinct isoforms of Na+,K+-ATPase regulate glucose utilization in neurons in response to membrane depolarization, and in glial cells in response to elevated extracellular potassium.

Developmental critical period in gentic redox dysregulation: animal and human studies in schizophrenia

Converging evidence favors an abnormal susceptibility to oxidative stress in schizophrenia. Decreased levels of glutathione (GSH), the major cellular antioxidant and redox regulator, was observed in cerebrospinal-fluid and prefrontal cortex of patients. Importantly, abnormal GSH synthesis of genetic origin was observed: Two case-control studies showed an association with a GAG trinucleotide repeat (TNR) polymorphism in the GSH key synthesizing enzyme glutamate-cysteine-ligase (GCL) catalytic subunit (GCLC) gene. The most common TNR genotype 7/7 was more frequent in controls, whereas the rarest TNR genotype 8/8 was three times more frequent in patients. The disease associated genotypes (35% of patients) correlated with decreased GCLC protein, GCL activity and GSH content. Similar GSH system anomalies were observed in early psychosis patients. Such redox dysregulation combined with environmental stressors at specific developmental stages could underlie structural and functional connectivity anomalies. In pharmacological and knock-out (KO) models, GSH deficit induces anomalies analogous to those reported in patients. (a) morphology: spine density and GABA-parvalbumine immunoreactivity (PV-I) were decreased in anterior cingulate cortex. KO mice showed delayed cortical PV-I at PD10. This effect is exacerbated in mice with increased DA from PD5-10. KO mice exhibit cortical impairment in myelin and perineuronal net known to modulate PV connectivity. (b) physiology: In cultured neurons, NMDA response are depressed by D2 activation. In hippocampus, NMDA-dependent synaptic plasticity is impaired and kainate induced g-oscillations are reduced in parallel to PV-I. (c) cognition: low GSH models show increased sensitivity to stress, hyperactivity, abnormal object recognition, olfactory integration and social behavior. In a clinical study, GSH precursor N-acetyl cysteine (NAC) as add on therapy, improves the negative symptoms and decreases the side effects of antipsychotics. In an auditory oddball paradigm, NAC improves the mismatched negativity, an evoked potential related to pre-attention and to NMDA receptors function. In summary, clinical and experimental evidence converge to demonstrate that a genetically induced dysregulation of GSH synthesis combined with environmental insults in early development represent a major risk factor contributing to the development of schizophrenia Conclusion Based on these data, we proposed a model for PSIP1 promoter activity involving a complex interplay between yet undefined regulatory elements to modulate gene expression.

Rôle de l'activation de c-Jun N-terminal kinase (JNK) dans un modèle de glaucome chez le rat et neuroprotection par inhibition de la voie de JNK

RESUME : Dans ce travail effectué chez le rat adulte, l'excitotoxicité rétinienne est élicitée par injection intravitréenne de NMDA. Les lésions en résultant sont localisées dans la rétine interne. Elles prennent la forme de pycnoses dans la couche des cellules ganglionnaires (corps cellulaires des cellules ganglionnaires et amacrines déplacées) et dans la partie interne de la couche nucléaire interne (cellules amacrines). Cette localisation est liée à la présence de récepteurs au glutamate de type NMDA sur ces cellules. L'activation de ces récepteurs entraîne un influx calcique et l'activation de diverses enzymes (phospholipase A, calpaïnes, calmoduline, synthase d'oxyde nitrique). La signalisation se poursuit en aval en partie par les voies des Mitogen Activated Protein Kinase (MAPK) : ERK, p38, ]NK. Dans les expériences présentées, toutes trois sont activées après l'injection de NMDA. Dans les cascades de signalisation de JNK, trois kinases s'ancrent sur une protéine scaffold. Les MAPKKK phosphorylent MKK4 et MKK7, qui phosphorylent JNK. JNK a de nombreuses cibles nucléaires (dont le facteur de transcription c-Jun) et cytoplasmiques. La voie de JNK est bloquée par l'inhibiteur peptidique D-JNKI-1 en empêchant l'interaction de la kinase avec son substrat. L'inhibiteur est formé de 20 acides aminés du domaine de liaison JBD et de 10 acides aminés de la partie TAT du virus HIV. L'injection intravitréenne de D-JNKI-1 permet une diminution des taux de JNK et c-Jun phosphorylés dans les lysats de rétine. L'effet prépondérant est la restriction importante des altérations histologiques des couches internes de la rétine. L'évaluation par électrorétinogramme met en sus en évidence une sauvegarde de la fonction cellulaire. Ce travail a ainsi permis d'établir la protection morphologique et fonctionnelle des cellules de la rétine interne par inhibition spécifique de la voie de JNK lors d'excitotoxicité. SUMMARY Excitotoxicity in the retina associates with several pathologies like retinal ischemia, traumatic optic neuropathy and glaucoma. In this study, excitotoxicity is elicited by intravitreal NMDA injection in adult rats. Lesions localise in the inner retina. They present as pyknotic cells in the ganglion cell layer (ganglion cells and displaced amacrines) and the inner nuclear layer (amacrine cells). These cells express NMDA glutamate receptors. The receptor activation leads to a calcium flow into the cell and hence enzyme activation (phospholipase, calpains, calmodulin, nitric oxide synthase). The subsequent signaling pathways can involve the Mitogen Activated Protein Kinases (MAPK): ERK, p38 end JNK. These were all activated in our experiments. The signaling cascade organises around several scaffold proteins. The various MAPKKK phosphorylate MKK4 and MKK7, which phosphorylate JNK. JNK targets are of nuclear (c-Jun transcription factor) or cytoplasmic localisation. The peptidic inhibitor D-JNKI-1, 20 amino acids from the JNK binding domain JBD coupled to 10 amino acids of the TAT transporter, disrupts the binding of JNK with its substrate. Intravitreal injection of the inhibitor lowers phosphorylated forms of JNK and c-Jun in retinal extracts. It protects strongly against histological lesions in the inner retina and allows functional rescue.

Drug refractory epilepsy in brain damage: effect of dextromethorphan on EEG in four patients.

High doses of dextromethorphan (20-42 mg/kg/day) were given to four critically ill children with seizures and frequent epileptiform abnormalities in the EEG that were refractory to antiepileptic drugs. Their acute diseases (hypoxia, head trauma and hypoxia, neurodegenerative disease, hypoglycaemia) were thought to be due in part to N-methyl-D-aspartate (NMDA) receptor mediated processes. Treatment with dextromethorphan, an NMDA receptor antagonist, was started between 48 hours and 14 days after the critical incident. In three patients the EEG improved considerably within 48 hours and seizures ceased within 72 hours. In the patient with neurodegenerative disease the effect on the EEG was impressive, but the seizures were not controlled. Despite the improvement of the EEG the clinical outcome was poor in all children: three died in the critical period or due to the progressing disease; the patient with hypoglycaemia survived with severe neurological sequelae. Plasma concentrations of dextromethorphan varied between 74-1730 ng/ml and its metabolite dextrorphan varied between 349-3790 ng/ml. In one patient corresponding concentrations in CSF were lower than those in plasma. The suppression of epileptic discharges by the doses of dextromethorphan given suggests that such doses are sufficient to block NMDA receptors.