Estudio de citoquinas y proteínas de choque térmico en pacientes polifracturados / Study of cytokines and heat shock proteins in multiple fracture patients

Objetivo: Valorar la repercusión y comportamiento de los niveles séricos de factores proinflamatorios con respecto a la aparición de complicaciones médico–quirúrgicas en pacientes politraumatizados. Material y métodos: se incluyeron 18 pacientes politraumatizados, 10 hombres y 8 mujeres, con 2 o más fracturas óseas y un ISS>16, con edad media de 42 años y cuyo mecanismo lesional más frecuente fue el accidente de tráfico (44%). El valor medio del ISS fue de 26,83 y de 33,72 para el NISS. Se recogieron datos demográficos, lesiones ocasionadas, intervenciones quirúrgicas realizadas, datos de evolución, complicaciones y secuelas. Se analizó en sangre, leucocitos, fibrinógeno, proteína C reactiva, TNFα, interleucina 1β, interleucina 6, proteína de choque térmico HSP70i y anticuerpos antiHSP70i. Resultados: Los valores de TNFα, tienen una curva ascendente, con un aumento de la pendiente a partir de las 48 horas del traumatismo. La IL-1‚ mostró el pico máximo en la primera medición inmediatamente después del traumatismo, para disminuir de manera progresiva. La IL-6 presentó cifras por encima de 500 pg/ml. Los niveles séricos elevados de HSP70i máximos en el momento inicial para disminuir en las siguientes 48 horas. Conclusiones: Las curvas de reacción de factores proinflamatorios establecidas servirán de base para futuros estudios que los afiancen como biomarcadores de politraumatismo (AU)

Objective: To evaluate the repercussion and behavior of the serum levels of proinflammatory factors in relation to the appearance of clinical-surgical complications in polytraumatized patients. Material and methods: The study comprised 18 polytraumatized patients, 10 males and 8 females, with two or more bone fractures and an injury severity score (ISS) >16, and with a mean age of 42 years, in which traffic accidents were the main cause of injury (44%). The mean ISS was 26.83, with a new injury severity score (NISS) of 33.72. Demographic data were collected, together with information on the injuries produced, the surgical interventions, outcome, complications and sequelae. Blood tests were performed to record leukocyte count, fibrinogen, C-reactive protein, TNF·, interleukin 1‚, interleukin 6, heat shock protein HSP70i and antiHSP70i antibodies. Results: The TNF· values showed an ascending tendency, with an increase in slope starting 48 hours after trauma. IL-1 in turn showed a maximum value on occasion of the first measurement immediately after injury, followed by a gradual decrease. IL-6 showed values above 500 pg/ml. Peak serum HSP70i elevation were recorded at first determination, followed by a decrease over the following 48 hours. Conclusions: The established proinflammatory factor response curves will serve as a basis for future studies to consolidate them as biomarkers applicable to polytraumatized patients (AU)

Cavum septum pellucidum in schizophrenia: a case report.

Different structural alterations of the central nervous system (CNS), such as an increase in ventricular size, decrease in hippocampus and amygdala volume, and other abnormalities that are probably secondary to a loss of neuronal mass and neuropili (axons and dendrite ramifications) linked to a precocious alteration in neurodevelopment are described in patients with schizophrenia in greater proportion than in the general population. The cavum septum pellucidum is among the alterations whose role in the disease is more unknown, since the cavity between the two septum laminae draws attention in very few cases. According to several studies, this alteration is more frequent in patients with schizophrenia than in healthy subjects and currently its association to the disease is being studied. In the following, we present the case of a 37 year old patient with no outstanding background, who was admitted to our psychiatry department due to psychotic symptoms.

Estudio prospectivo de los reactantes fase aguda tras una artroplastia total de rodilla / Prospective study of acute phase response factors after total knee artrhoplasty

Introducción: La artroplastia total de rodilla actualmentees el tratamiento de elección para las artrosis evolucionadasde dicha articulación. Al igual que otras lesiones mayores, lasintervenciones quirúrgicas o los traumatismos, originan respuestasmetabólicas, hormonales y hemodinámicas. Nos proponemosvalorar los reactantes de fase aguda tanto en prótesistotales de rodilla normales como en infectadas.Material y método: De forma consecutiva, a partir de marzodel 2003, se reclutaron 36 pacientes a los que se les implantóuna prótesis total de rodilla (grupo I). Además se estudiaron10 pacientes (grupo II) a los que se les habíaimplantado una prótesis de rodilla y presentaban criterios deinfección.Resultado: En el grupo I, los leucocitos y la PCR presentanun pico máximo a las 24 horas de la intervención, observándoseniveles preoperatorios en la extracción obtenida al mes;El fibrinógeno y la VSG muestran un ascenso hasta la extraccióntomada al mes de la cirugía, y continúan con niveles superioresa los basales incluso después de tres meses.Lo pacientes del grupo II mostraron niveles, estadísticamentesignificativos, más altos de fibrinógeno, VSG y PCR.Conclusiones: Los reactantes de fase aguda ofrecen unamoderada eficacia diagnóstica por lo que deben apoyar aldiagnóstico basado en los criterios clínicos de infección

Introduction: Nowadays total knee arthroplasty (TKA) isthe choice treatment in evolved knee artrosis. Operations ortraumas cause metabolism and hormone responses.Patients and Methods: Consecutively, from 2003, we includedprospectively in this research, 36 patients undergoingelective TKA (group I). Also we studied 10 patients (group II)that were diagnosed as infected TKA.Results: Acute phase response evolution in group I showsmaximum leukocytes and C-reactive protein (CRP) levels duringthe first 24 hours after the surgery, returning to presurgicallevels after one month. Erythrocyte sedimentation (ESR)rate and fibrinogen are increased even 3 months after the surgery.The patients of the goup II showed hihger levels, statisticallysignificant, of fibrinogen, CRP and ESR.Discussion: Acute phase response factors offer modest diagnosticefficacy, so we always have to use the clinical criterionto achive a god diagnosis

Chronic lithium treatment antagonizes glutamate-induced decrease of phosphorylated CREB in neurons via reducing protein phosphatase 1 and increasing MEK activities.

The cyclic AMP response element binding protein (CREB) has major roles in mediating adaptive responses at glutamatergic synapses and in the neuroprotective effects of neurotrophins. CREB has been implicated as a potential mediator of antidepressant actions. In vitro, chronic lithium treatment has been shown to promote neuronal cell survival. In the present study, we have used cultures of cerebellar granule neurons to analyze the effects of acute and chronic lithium treatment on the response to toxic concentrations of glutamate. Such concentrations of glutamate decrease the phosphorylation of CREB at serine(133) in an N-methyl-D-aspartate (NMDA) receptor-dependent manner. Chronic, but not acute, lithium treatment suppresses glutamate-induced decreases in phosphorylated CREB, and transfection studies indicate that chronic lithium, in the presence of a glutamate stimulus, markedly increases CRE-driven gene expression. Experiments with selected pharmacological reagents indicate that the glutamate-induced decreases in phosphorylated CREB are regulated primarily by protein phosphatase 1. Chronic lithium treatment not only decreases protein phosphatase 1 activity under these circumstances, but also augments glutamate-induced increases in MEK activity. PD 98059, a MEK inhibitor, prevents chronic lithium treatment from increasing phosphorylated CREB levels in glutamate-treated neurons. We conclude from these results that chronic lithium treatment is permissive for maintaining higher phosphorylated CREB levels in the presence of glutamate in part by decreasing protein phosphatase 1 activity and in part by increasing MEK activity. Higher levels of phosphorylated CREB and CRE-responsive genes such as bcl-2 may be responsible for lithium's reported effects on neuronal survival.

Specific interference with gene expression and gene function mediated by long dsRNA in neural cells.

Double-stranded (ds) RNA-induced sequence-specific interference with gene expression, RNA interference (RNAi), has been extensively used in invertebrates, allowing for efficient and high-throughput gene silencing and gene function analysis. In vertebrates, however, use of RNAi to study gene function has been limited due to non-specific effects induced by double-stranded RNA (dsRNA)-dependent protein kinase and interferon activation. dsRNA-induced specific inhibition of vertebrate gene expression has only been shown in embryonic and non-differentiated mammalian cells. In this report, we demonstrate dsRNA-induced specific interference of gene expression and gene function in partially as well as fully differentiated mouse neuroblastoma cells. Specific silencing was observed in the expression of an integrated transgene coding for green fluorescent protein and a variety of endogenous genes. Moreover, we show that RNAi-mediated inhibition of poly (ADP-ribose) polymerase (PARP) expression induced cellular resistance to oxygen-glucose deprivation, consistent with the role of PARP in ischemia-induced brain damage. Our results indicate that RNAi can be used as a powerful tool to study gene function in neural cells.

[Interdisciplinarity of Spanish cardiovascular research teams]. / La interdisciplinariedad en los grupos españoles de investigación en el área cardiovascular.

OBJECTIVES: The incidence of interdisciplinarity (ID) in Spanish cardiovascular research teams was analyzed and scientists' opinions about interdisciplinary relationships were examined. METHODS: The data analyzed were obtained in a survey sent in 1999 to a sample of 310 researchers. They were selected using bibliometric techniques and/or for being research project leaders in recent years. RESULTS: The response rate was 61%. Data were obtained from 130 research teams located mainly in Madrid and Catalonia. Teams doing clinical research (81%) and those working in hospitals (64%) predominated. Different facets of the interdisciplinary nature of the teams were analyzed: scientist training, team composition, behavior patterns, collaboration, and publication and reading habits. A high ID was observed in the area: more than 70% of the teams are interdisciplinary according to the training of scientists, around 80% make use of knowledge or techniques from other disciplines, and around 90% read and publish in journals outside their own disciplines. Basic research teams had a higher ID than clinical ones. A total of 37 highly interdisciplinary teams were identified. These teams had a greater tendency towards collaboration. CONCLUSIONS: Interdisciplinary reading and publishing habits were the norm among Spanish researchers, even in single-discipline groups. The scattered nature of teams, their high rate of external collaborations, and the multidisciplinary context of centers enhance interdisciplinary relationships. Administrative barriers do not seem to be a major obstacle to establishing interdisciplinary contacts.

La interdisciplinariedad en los grupos españoles de investigación en el área cardiovascular / Interdisciplinarity of spanish cardiovascular research teams

Objetivos. Se analiza la presencia de interdisciplinariedad (ID) en los grupos españoles de investigación en el área cardiovascular, así como las opiniones de los investigadores sobre las relaciones interdisciplinarias. Métodos. Los datos analizados proceden de una encuesta enviada en 1999 a una muestra de 310 investigadores seleccionados por métodos bibliométricos o por haber dirigido proyectos de investigación dentro del área en los últimos años. Resultados. La tasa de respuesta fue del 61 por ciento. Se obtienen datos de 130 grupos de investigación localizados sobre todo en Madrid y Cataluña. Predominan los grupos que realizan investigación clínica (81 por ciento) y los que están ubicados en hospitales (64 por ciento). El carácter interdisciplinario de los grupos se analiza en distintas facetas: formación de los investigadores, composición de los grupos, pautas de comportamiento, prácticas de colaboración y hábitos de publicación y lectura. Se observa una alta ID en el área: más del 70 por ciento de los grupos es interdisciplinario por la formación de sus investigadores, el 80 por ciento admite aplicar conocimientos o técnicas de otras disciplinas y cerca del 90 por ciento utiliza revistas de disciplinas distintas de la propia para la lectura y publicación de sus investigaciones. Los grupos básicos tienden a mostrar mayor ID que los clínicos. Se detectan 37 grupos muy interdisciplinarios, que muestran mayor tendencia a colaborar que los equipos restantes. Conclusiones. Los hábitos de lectura y publicación interdisciplinarios son la norma entre los investigadores españoles, incluso en los grupos de composición unidisciplinaria. El carácter distribuido de los grupos, su alta tasa de colaboración externa y el ambiente multidisciplinario de los centros favorecen las relaciones entre disciplinas. Las delimitaciones administrativas no parecen ser un importante obstáculo para el desarrollo de contactos interdisciplinarios (AU)

Reduction of functional N-methyl-D-aspartate receptors in neurons by RNase P-mediated cleavage of the NR1 mRNA.

One approach to studying the functional role of individual NMDA receptor subunits involves the reduction in the abundance of the protein subunit in neurons. We have pursued a strategy to achieve this goal that involves the use of a small guide RNA which can lead to the destruction of the mRNA for a specific receptor subunit. We designed a small RNA molecule, termed 'external guide sequence' (EGS), which binds to the NR1 mRNA and directs the endonuclease RNase P to cleave the target message. This EGS has exquisite specificity and directed the RNase P-dependent cleavage at the targeted location within the NR1 mRNA. To improve the efficiency of this EGS, an in vitro evolution strategy was employed which led to a second generation EGS that was 10 times more potent than the parent molecule. We constructed an expression cassette by flanking the EGS with self-cleaving ribozymes and this permitted generation of the specified EGS RNA sequence from any promoter. Using a recombinant Herpes simplex virus (HSV), we expressed the EGS in neurons and showed the potency of the EGS to reduce NR1 protein within neurons. In an excitotoxicity assay, we showed that expression of the EGS in cortical neurons is neuroprotective. Our results demonstrate the utility of EGSs to reduce the expression of any gene (and potentially any splice variant) in neurons.

Histochemical analysis of nitric oxide synthase by NADPH diaphorase staining.

The presence of NOS coincides with that of NADPH diaphorase activity; therefore, NADPH diaphorase staining is widely used to detect NOS-containing cells in neural and non-neural tissues. With the appropriate fixation procedure, this method is capable of detecting cells containing any of the NOS isoforms.

Dynamic regulation of neuronal NO synthase transcription by calcium influx through a CREB family transcription factor-dependent mechanism.

Neuronal nitric oxide (NO) synthase (nNOS) is dynamically regulated in response to a variety of physiologic and pathologic stimuli. Although the dynamic regulation of nNOS is well established, the molecular mechanisms by which such diverse stimuli regulate nNOS expression have not yet been identified. We describe experiments demonstrating that Ca(2+) entry through voltage-sensitive Ca(2+) channels regulates nNOS expression through alternate promoter usage in cortical neurons and that nNOS exon 2 contains the regulatory sequences that respond to Ca(2+). Deletion and mutational analysis of the nNOS exon 2 promoter reveals two critical cAMP/Ca(2+) response elements (CREs) that are immediately upstream of the transcription start site. CREB binds to the CREs within the nNOS gene. Mutation of the nNOS CREs as well as blockade of CREB function results in a dramatic loss of nNOS transcription. These findings suggest that nNOS is a Ca(2+)-regulated gene through the interactions of CREB on the CREs within the nNOS exon 2 promoter and that these interactions are likely to be centrally involved in the regulation of nNOS in response to neuronal injury and activity-dependent plasticity.

Requirement for nitric oxide activation of p21(ras)/extracellular regulated kinase in neuronal ischemic preconditioning.

The mechanisms underlying neuronal ischemic preconditioning, a phenomenon in which brief episodes of ischemia protect against the lethal effects of subsequent periods of prolonged ischemia, are poorly understood. Ischemia can be modeled in vitro by oxygen-glucose deprivation (OGD). We report here that OGD preconditioning induces p21(ras) (Ras) activation in an N-methyl-D-aspartate receptor- and NO-dependent, but cGMP-independent, manner. We demonstrate that Ras activity is necessary and sufficient for OGD tolerance in neurons. Pharmacological inhibition of Ras, as well as a dominant negative mutant Ras, block OGD preconditioning whereas a constitutively active form of Ras promotes neuroprotection against lethal OGD insults. In contrast, the activity of phosphatidyl inositol 3-kinase is not required for OGD preconditioning because inhibition of phosphatidyl inositol 3-kinase with a chemical inhibitor or with a dominant negative mutant does not have any effect on the development of OGD tolerance. Furthermore, using recombinant adenoviruses and pharmacological inhibitors, we show that downstream of Ras the extracellular regulated kinase cascade is required for OGD preconditioning. Our observations indicate that activation of the Ras/extracellular regulated kinase cascade by NO is a critical mechanism for the development of OGD tolerance in cortical neurons, which may also play an important role in ischemic preconditioning in vivo.

Reduced editing of low-affinity kainate receptor subunits in optic nerve glial cells.

We have analyzed the degree of editing of adult optic nerve mRNAs encoding the low-affinity kainate receptor subunits, GluR5 and GluR6, the two major constituents of native receptors in this family. To this end, we used reverse transcription (RT) followed by polymerase chain reaction (PCR), and subsequent cloning and sequencing of the amplified fragments. Our results revealed that the GluR5 subunit is unedited at the Q/R site of cismembrane domain 2 (M2), whereas the GluR6 subunit is edited to a low extent at this site. These findings are in contrast to those reported by others using mRNAs from the adult brain in which GluR5 and GluR6 are edited at the Q/R site of M2 to a larger extent. In addition, we found that the adenosine deaminases, DRADA, RED1 and RED2, which edit ionotropic glutamate receptors in the brain, are expressed in the adult optic nerve and in oligodendrocytes, the major cell type in this structure. It thus appears that editing of kainate receptors in optic nerve cells is not limited by the availability of editing enzymes but rather by other, as yet unidentified factors. Overall, reduced editing of kainate receptor subunits in glial cells may result in different functional responses of the native receptors present in these cells with respect to those in neurons.

[Spanish scientific production in the cardiovascular area through Science Citation Index (1990-1996)]. / La producción científica española en el área cardiovascular a través del Science Citation Index (1990-1996).

INTRODUCTION AND OBJECTIVES: A bibliometric analysis of the Spanish scientific production in the Cardiovascular research area constitutes a useful approach to the study of the scientific activity conducted in the field. The most relevant aspects of the research and their evolution over time are analysed. MATERIAL AND METHODS: Publications covered by the Science Citation Index database under the heading "cardiovascular system" during the years 1990-1996 and signed by at least one Spanish address were studied. The distribution of the scientific production by geographic areas and institutional sectors was analysed and the most productive centers were identified. RESULTS: During these years, the scientific production in Cardiovascular research showed a growing rate of 83%, slightly lower than that of Biomedicine in Spain. A total of 1,434 documents were analysed (3% of the biomedical production), with 24% of meeting-abstracts. Although the production was concentrated in Madrid (33%) and Cataluña (30%), Cantabria and Navarra emerged when the values were normalised according to either R&D expenditures or population data. The most productive institutional sectors were Hospitals (86%) and Universities (22%). The most productive centres were identified, as well as their growth rate, their activity index and the average impact factor of the journals used. More than 50% of the documents were performed in collaboration among different centres. International collaboration showed an upward trend (from 16% to 21%). CONCLUSIONS: Spanish Cardiovascular research showed an increasing international visibility over the studied period, as shown by the growing number of publications in the Science Citation Index database, the up-rising trend towards high-impact-factor journals, and the ever-growing collaboration of Spanish authors with foreign partners.

[Evaluation of the scientific activity through bibliometric indices]. / Evaluación de la actividad científica a través de indicadores bibliométricos.

The scope of bibliometric studies is the treatment and quantitative analysis of scientific publications. They belong to the so-called "social studies of science", and science policy constitutes one of its main applied fields. These studies efficiently complement the opinions and judgements of experts, thus providing objective and useful tools for evaluating the results of scientific activity. Nevertheless, given the impact that these evaluations have on the assignment of funding for research and even on the professional career of investigators, it becomes essential to know in detail the characteristics of bibliometric indicators and the limitations of their use. The Science Citation Index database is one of the most employed. In the case of biomedical research it is useful to analyze the most internationally visible scientific production, since it satisfactorily covers biomedical journals; however, clinical research with local interest published in Spanish journals is not included in that database. Widely employed bibliometric indicators are those measuring scientific activity through the number of publications, those based on the citations received by published studies and, in between them, the impact of journals. The impact factor is an indicator very used in bibliometric studies; though occasionally a high impact factor is assumed to reflect high quality, this indicator specifically measures visibility and diffusion of the works published by these journals rather than their scientific quality.

On how altered glutamate homeostasis may contribute to demyelinating diseases of the CNS.

Glial cells communicate reciprocally with neurons in multiple ways, both in synaptic and non-synaptic regions of the central nervous system. In the latter, neuron to glial and glial to glial signals can be mediated by neurotransmitters. Here, we review the presence and some of the functional properties of glutamate transporters and receptors in oligodendrocytes. In addition, we present data illustrating that alterations in glutamate homeostasis can be excitotoxic to oligodendroglia and that the tissue lesions caused by overactivation of glutamate receptors resemble those observed in demyelinating diseases such as multiple sclerosis. Overall, this information indicates that aberrant glutamate signaling may contribute to the development of some white matter pathologies.

[Scientific activity of the most productive Spanish research teams in pharmacology and pharmacy during the period 1986-1993 as covered by the Science Citation Index (SCI)]. / Actividad científica de los grupos españoles más productivos en farmacología y farmacia durante el período 1986-1993 a través del Science Citation Index (SCI).

BACKGROUND: Bibliometric analyses at the research team level in the pharmacology and pharmacy (F&F) subfield allow in-depth analysis of the findings obtained at higher aggregation levels. MATERIAL AND METHODS: Publications of Spanish authors in pharmacological journals covered by the Science Citation Index (SCI) are studied and the most productive Spanish teams during 1986-1989 and 1990-1993 are identified through in-house programmes based on co-authorship analysis. Team composition is defined and team size, scientific output, productivity, expected impact factor and collaboration pattern of the most productive teams are analysed as well as its evolution over time. RESULTS: The observed annual increasing rate in the number of publications and authors in the field (11%) was similar to that of the number of researchers in the Spanish scientific system (8%). The number of highly productive teams doubled from the first to the second period (28 and 61 teams, respectively). The average team size was 8 researchers, showing an average scientific output of 3 documents/year and an average, productivity of 1.7 publications per author and team in each period. Over the years, an increase in the expected impact factor (EIF) of the publication journals and significant changes in the team composition were observed. A total of 13 stable teams were identified, with a greater size and output than the rest of the groups, and showing an upward trend in their expected impact factor and international collaboration rate. Stable teams did not show the negative correlation between team size and productivity observed for the total of the highly productive teams. CONCLUSIONS: The F&F subfield is in a very dynamic stage, with a great increase in the number of researchers, starting of new teams and consolidation of others. A trend was observed towards the concentration of activity in the most productive teams, that are the most visible internationally both in number of publications in the SCI and in the impact factor of the publication journals.

Nitric oxide mediates N-methyl-D-aspartate receptor-induced activation of p21ras.

N-methyl-D-aspartate (NMDA) glutamate receptor-mediated increases in intracellular calcium are thought to play a critical role in synaptic plasticity. The mechanisms by which changes in cytoplasmic calcium transmit the glutamate signal to the nucleus, which is ultimately important for long-lasting neuronal responses, are poorly understood. We show that NMDA receptor stimulation leads to activation of p21(ras) (Ras) through generation of nitric oxide (NO) via neuronal NO synthase. The competitive NO synthase inhibitor, L-nitroarginine methyl ester, prevents Ras activation elicited by NMDA and this effect is competitively reversed by the NO synthase substrate, L-arginine. NMDA receptor stimulation fails to activate Ras in neuronal cultures from mice lacking neuronal NO synthase. NMDA-induced Ras activation occurs through a cGMP-independent pathway as 1H-[1,2,4]oxadiazolo[4,3-alpha]quinoxalin-1-one (ODQ), a potent and selective inhibitor of guanylyl cyclase, has no effect on NMDA receptor-induced activation of Ras, and the cell-permeable cGMP analog, 8Br-cGMP, does not activate Ras. Furthermore, NO directly activates immunoprecipitated Ras from neurons. NMDA also elicits tyrosine phosphorylation of extracellular signal-regulated kinases, a downstream effector pathway of Ras, through a NO/non-cGMP dependent mechanism, thus supporting the physiologic relevance of endogenous NO regulation of Ras. These results suggest that Ras is a physiologic target of endogenously produced NO and indicates a signaling pathway for NMDA receptor activation that may be important for long-lasting neuronal responses.

Manganese superoxide dismutase protects nNOS neurons from NMDA and nitric oxide-mediated neurotoxicity.

Neuronal nitric oxide synthase (nNOS) neurons kill adjacent neurons through the action of NMDA-glutamate receptor activation, although they remain relatively resistant to the toxic effects of NMDA and NO. The molecular basis of the resistance of nNOS neurons to toxic insults is unknown. To begin to understand the molecular mechanisms of the resistance of nNOS neurons, we developed a pheochromacytoma-derived cell line (PC12) that is resistant to the toxic effects of NO. We found through serial analysis of gene expression (SAGE) that manganese superoxide dismutase (MnSOD) is enriched in the NO-resistant PC12 cell-derived line (PC12-R). Antisense MnSOD renders PC12-R cells sensitive to NO toxicity and increases the sensitivity to NO in the parental, NO-sensitive PC12 line (PC12-S). Adenoviral transfer of MnSOD protects PC12-S cells against NO toxicity. We extended these studies to cortical cultures and showed that MnSOD is enriched in nNOS neurons and that antisense MnSOD renders nNOS neurons susceptible to NMDA neurotoxicity, although it has little effect on the overall susceptibility of cortical neurons to NMDA toxicity. Overexpression of MnSOD provides dramatic protection against NMDA and NO toxicity in cortical cultures, but not against kainate or AMPA neurotoxicity. Furthermore, nNOS neurons from MnSOD -/- mice are markedly sensitive to NMDA toxicity. Adenoviral transfer of MnSOD to MnSOD-/- cultures restores resistance of nNOS neurons to NMDA toxicity. Thus, MnSOD is a major protective protein that appears to be essential for the resistance of nNOS neurons in cortical cultures to NMDA mediated neurotoxicity.

Regulation of neuronal nitric oxide synthase and identification of novel nitric oxide signaling pathways.

Neuronal nitric oxide synthase (nNOS) participate in a variety of physiologic and pathologic processes in the nervous system. nNOS was originally felt to be a constitutively expressed enzyme, but recent observations suggest that its levels are dynamically controlled in response to neuronal development, plasticity and injury. nNOS expression is regulated through alternative promoter usage through alternative mRNA splicing and it is likely that this plays an important role in the inducibility of gene expression in response to extracellular stimuli. Emerging data also suggests that NO may be the key mediator linking activity to gene expression and long-lasting neuronal responses through NO activating p21Ras through redox-sensitive modulation.