Del laboratorio a la clínica en el ictus isquémico agudo. Modelos experimentales in vitro e in vivo / From the laboratory to the clinic in acute ischaemic stroke. In vitro and in vivo experimental models

Introducción: La enfermedad cerebrovascular es una de las principales causas de muerte, discapacidad y demencia en el mundo. La forma más frecuente de la enfermedad, el ictus isquémico, sólo tiene un fármaco disponible, el activador tisular del plasminógeno, y pocos pacientes pueden beneficiarse de esta terapia por los estrictos criterios de inclusión establecidos para su uso. Esta circunstancia hace crucial la búsqueda de nuevas formas de tratamiento para combatir las secuelas de la enfermedad, y para ello es necesario el desarrollo de nuevos modelos biomiméticos que permitan conocer mejor su evolución. Desarrollo: En esta revisión, actualizamos las plataformas y modelos más utilizados en los últimos años para estudiar la fisiopatología del ictus isquémico. Por un lado, repasamos las plataformas bi- y tridimensionales sobre las que se llevan a cabo los ensayos in vitro y, por otro lado, describimos los modelos experimentales in vivo más utilizados en la actualidad, así como las técnicas para evaluar el daño isquémico. Conclusiones: El desarrollo de buenos modelos experimentales tiene como fin último encontrar nuevas formas de tratamiento y, de esta manera, mejorar el pronóstico y la calidad de vida de los pacientes; por ello, es importante generar nuevos dispositivos in vitro y refinar más aún los modelos in vivo para hacer posible una buena traslación a la clínica.(AU)

Introduction: Cerebrovascular disease is one of the leading causes of death, disability and dementia around the world. For the most common form of the disease, ischaemic stroke, there is only one drug available, tissue plasminogen activator, and few patients can benefit from this therapy because of the strict inclusion criteria established for its use. This circumstance makes it crucial to search for new forms of treatment to combat the sequelae of the disease, and this requires the development of new biomimetic models that allow for a better understanding of its evolution. Development: In this review, we update the platforms and models most widely used in recent years to study the pathophysiology of ischaemic stroke. On the one hand, we review the two- and three-dimensional platforms on which in vitro assays are carried out and, on the other, we describe the most commonly used in vivo experimental models and techniques for assessing ischaemic damage. Conclusions: The ultimate aim of developing good experimental models is to find new forms of treatment and thus improve patients’ prognosis and quality of life. It is therefore important to generate new in vitro devices and to further refine in vivo models to enable a good clinical translation.(AU)

[From the laboratory to the clinic in acute ischaemic stroke. In vitro and in vivo experimental models]. / Del laboratorio a la clínica en el ictus isquémico agudo. Modelos experimentales in vitro e in vivo.

INTRODUCTION: Cerebrovascular disease is one of the leading causes of death, disability and dementia around the world. For the most common form of the disease, ischaemic stroke, there is only one drug available, tissue plasminogen activator, and few patients can benefit from this therapy because of the strict inclusion criteria established for its use. This circumstance makes it crucial to search for new forms of treatment to combat the sequelae of the disease, and this requires the development of new biomimetic models that allow for a better understanding of its evolution. DEVELOPMENT: In this review, we update the platforms and models most widely used in recent years to study the pathophysiology of ischaemic stroke. On the one hand, we review the two- and three-dimensional platforms on which in vitro assays are carried out and, on the other, we describe the most commonly used in vivo experimental models and techniques for assessing ischaemic damage. CONCLUSIONS: The ultimate aim of developing good experimental models is to find new forms of treatment and thus improve patients' prognosis and quality of life. It is therefore important to generate new in vitro devices and to further refine in vivo models to enable a good clinical translation.

TITLE: Del laboratorio a la clínica en el ictus isquémico agudo. Modelos experimentales in vitro e in vivo.Introducción. La enfermedad cerebrovascular es una de las principales causas de muerte, discapacidad y demencia en el mundo. La forma más frecuente de la enfermedad, el ictus isquémico, sólo tiene un fármaco disponible, el activador tisular del plasminógeno, y pocos pacientes pueden beneficiarse de esta terapia por los estrictos criterios de inclusión establecidos para su uso. Esta circunstancia hace crucial la búsqueda de nuevas formas de tratamiento para combatir las secuelas de la enfermedad, y para ello es necesario el desarrollo de nuevos modelos biomiméticos que permitan conocer mejor su evolución. Desarrollo. En esta revisión, actualizamos las plataformas y modelos más utilizados en los últimos años para estudiar la fisiopatología del ictus isquémico. Por un lado, repasamos las plataformas bi- y tridimensionales sobre las que se llevan a cabo los ensayos in vitro y, por otro lado, describimos los modelos experimentales in vivo más utilizados en la actualidad, así como las técnicas para evaluar el daño isquémico. Conclusiones. El desarrollo de buenos modelos experimentales tiene como fin último encontrar nuevas formas de tratamiento y, de esta manera, mejorar el pronóstico y la calidad de vida de los pacientes; por ello, es importante generar nuevos dispositivos in vitro y refinar más aún los modelos in vivo para hacer posible una buena traslación a la clínica.

Pathophysiological and pharmacological relevance of TLR4 in peripheral immune cells after stroke.

Stroke is a very common disease being the leading cause of death and disability worldwide. The immune response subsequent to an ischemic stroke is a crucial factor in its physiopathology and outcome. This response is not limited to the injury site. In fact, the immune response to the ischemic process mobilizes mainly circulating cells which upon activation will be recruited to the injury site. When a stroke occurs, molecules that are usually retained inside the cell bodies are released into the extracellular space by uncontrolled cell death. These molecules can bind to the Toll-like receptor 4 (TLR4) in circulating immune cells which are then activated, eliciting, although not exclusively, the inflammatory response to the stroke. In this review, we present an up-to-date summary of the role of the different peripheral immune cells in stroke as well as the role of TLR4 in the function of each cell type in ischemia. Also, we summarize the different antagonists developed against TLR4 and their potential as a pharmacological tool for stroke treatment.

Specific Features of SVZ Neurogenesis After Cortical Ischemia: a Longitudinal Study.

Stroke is a devastating disease with an increasing prevalence. Part of the current development in stroke therapy is focused in the chronic phase, where neurorepair mechanisms such as neurogenesis, are involved. In the adult brain, one of the regions where neurogenesis takes place is the subventricular zone (SVZ) of the lateral ventricles. Given the possibility to develop pharmacological therapies to stimulate this process, we have performed a longitudinal analysis of neurogenesis in a model of cortical ischemia in mice. Our results show an initial decrease of SVZ proliferation at 24 h, followed by a recovery leading to an increase at 14d and a second decrease 28d after stroke. Coinciding with the 24 h proliferation decrease, an increase in the eutopic neuroblast migration towards the olfactory bulb was observed. The analysis of the neuroblast ectopic migration from the SVZ toward the lesion showed an increase in this process from day 14 after the insult. Finally, our data revealed an increased number of new cortical neurons in the peri-infarct cortex 65d after the insult. In summary, we report here critical check-points about post-stroke neurogenesis after cortical infarcts, important for the pharmacological modulation of this process in stroke patients.

Pallister-Killian syndrome in a preterm newborn who died soon after precipitous delivery: cytogenetic analysis.

The authors report a preterm neonate with dysmorphic traits and cleft palate who was born preterm because of precipitous delivery and died soon after birth notwithstanding neonatal intensive care unit (NICU) support. The cytogenetic analysis on fibroblasts from post-mortem skin biopsy demonstrated a Pallister-Killian syndrome (PKS). PKS is a cytogenetically syndrome characterized by a tissue limited mosaic distribution of one isochromosome 12p (tetrasomy 12p). Clinical manifestations of PKS are variable, and some symptoms may overlap with other malformative syndromes, thus the correct diagnosis mainly depends on the demonstration of the specific cytogenetic abnormality.

The high-mobility group I-Y transcription factor is involved in cerebral ischemia and modulates the expression of angiogenic proteins.

The present study aims to identify transcription factors (TFs) contributing to angiogenesis, a mechanism involved in giving plasticity to the brain, as potential therapeutic targets after cerebral ischemia. The promoter sequences from candidate genes involved in angiogenesis were submitted to a comparative analysis by bioinformatics software. High-mobility group I-Y protein (HMGIY) TF characterization in a rat permanent focal cerebral ischemia model was performed by quantitative real time polymerase chain reaction and Western blot for the TF expression profile study. The TF functional study was carried out using a TF-TF interaction array and gene silencing by siRNA in rat brain microvascular endothelial cells. The results showed that the promoters shared a common TF binding site for HMGIY. The expression profile analysis in ischemic rat brain showed an increase in HMGIY mRNA in the acute phase and a progressive overexpression of protein over time post-ischemia. The interaction array analysis revealed that ischemia promotes the interaction of HMGIY with TFs involved in different cerebral plasticity processes. In vitro knockdown studies showed that angiopoietin 1 and vascular endothelial growth factor expression is controlled by HMGIY and that this TF is involved in cell survival in brain endothelial cells. These findings suggest that HMGIY is a potential therapeutic target that could promote brain repair functions after stroke.

Characterization of Gcf2/Lrrfip1 in experimental cerebral ischemia and its role as a modulator of Akt, mTOR and ß-catenin signaling pathways.

Leucine-rich repeat in Flightless-1 interaction protein 1 (Lrrfip1) is an up-regulated protein after cerebral ischemia whose precise role in the brain both in healthy and ischemic conditions is unclear. Different Lrrfip1 isoforms with distinct roles have been reported in human and mouse species. The present study aimed to analyze the Lrrfip1 transcriptional variants expressed in rat cortex, to characterize their expression patterns and subcellular location after ischemia, and to define their putative role in the brain. Five transcripts were identified and three of them (Lrrfip1, CRA_g and CRA_a' (Fli-I leucine-rich repeat associated protein 1 - Flap-1)) were analyzed by quantitative real-time polymerase chain reaction (qPCR). All the transcripts were up-regulated and showed differential expression patterns after in vivo and in vitro ischemia models. The main isoform, Lrrfip1, was found to be up-regulated from the acute to the late phases of ischemia in the cytoplasm of neurons and astrocytes of the peri-infarct area. This study demonstrates that Lrrfip1 activates ß-catenin, Akt, and mammalian target of rapamycin (mTOR) proteins in astrocytes and positively regulates the expression of the excitatory amino acid transporter subtype 2 (GLT-1). Our findings point to Lrrfip1 as a key brain protein that regulates pro-survival pathways and proteins and encourages further studies to elucidate its role in cerebral ischemia as a potential target to prevent brain damage and promote functional recovery after stroke.

Unbalanced 1q whole-arm translocation resulting in der(14)t(1;14)(q11-12;p11) in myelodysplastic syndrome.

Unbalanced whole-arm translocations (WATs) of the long arm of chromosome 1, resulting in complete trisomy 1q, are chromosomal abnormalities detectable in both solid tumors and hematologic neoplasms. Among the WATs of 1q to acrocentric chromosomes, a few patients with der(1;15) described as a dicentric chromosome have been reported so far, whereas cases of der(1;14) are much rarer. We report on a case of der(1;14) detected as single anomaly in a patient with myelodysplastic syndrome. The aim of our work was to investigate the breakpoints of the (1;14) translocation leading to the der(1;14). Fluorescence in situ hybridization (FISH) experiments have been performed on chromosome preparations from bone marrow aspirate, using specific centromeric probes of both chromosomes, as well as a probe mapping to 1q11 band. FISH results showed that in our patient the derivative chromosome was monocentric with a unique centromere derived from chromosome 14. The breakpoints of the translocation were located in the short arm of chromosome 14 and in the long arm of chromosome 1, between the alphoid D1Z5 and the satellite II domains. The 1q breakpoint was within the pericentromeric region of chromosome 1, which is notoriously an unstable chromosomal region, involved in different chromosomal rearrangements.

Daidzein has neuroprotective effects through ligand-binding-independent PPARγ activation.

Phytoestrogens are a group of plant-derived compounds that include mainly isoflavones like daidzein. Phytoestrogens prevent neuronal damage and improve outcome in experimental stroke; however, the mechanisms of this neuroprotective action have not been fully elucidated. In this context, it has been postulated that phytoestrogens might activate the peroxisome proliferator-activated receptor-γ (PPARγ), which exerts neuroprotective effects in several settings. The aim of this study was to determine whether the phytoestrogen daidzein elicits beneficial actions in neuronal cells by mechanisms involving activation of PPARγ. Our results show that daidzein (0.05-5 µM) decreases cell death induced by exposure to oxygen-glucose deprivation (OGD) from rat cortical neurons and that improves synaptic function, in terms of increased synaptic vesicle recycling at nerve terminals, being both effects inhibited by the PPARγ antagonist T0070907 (1 µM). In addition, this phytoestrogen activated PPARγ in neuronal cultures, as shown by an increase in PPARγ transcriptional activity. Interestingly, these effects were not due to binding to the receptor ligand site, as shown by a TR-FRET PPARγ competitive binding assay. Conversely, daidzein increased PPARγ nuclear protein levels and decreased cytosolic ones, suggesting nuclear translocation. We have used the receptor antagonist (RE) fulvestrant to study the neuroprotective participation of daidzein via estrogen receptor and at least in our model, we have discarded this pathway. These results demonstrate that the phytoestrogen daidzein has cytoprotective properties in neurons, which are due to an increase in PPARγ activity not mediated by direct binding to the receptor ligand-binding domain but likely due to post-translational modifications affecting its subcellular location and not depending to the RE and it is not additive with the agonist rosiglitazone.

Reduced infarct size and accumulation of microglia in rats treated with WIN 55,212-2 after neonatal stroke.

Cannabinoids have emerged as brain protective agents under neurodegenerative conditions. Many neuroprotective actions of cannabinoids depend on the activation of specific receptors, cannabinoid receptor type 1 (CB1R) and type 2 (CB2R). The aim of the present study was to determine whether the CB2R and CB1R agonist WIN 55,212-2 (WIN) protects neonatal brain against focal cerebral ischemia-reperfusion and whether anti-inflammatory mechanisms play a role in protection. Seven-day-old rats were subjected to 90-min middle cerebral artery occlusion (MCAO), and injured rats were identified by diffusion-weighted MRI during the occlusion. After reperfusion, rats were subcutaneously administered 1 mg/kg of WIN or vehicle twice daily until sacrifice. MCAO led to increased mRNA expression of CB2R (but not CB1R), chemokine receptors (CCR2 and CX3CR1), and cytokines (IL-1ß and TNFα), as well as increased protein expression of chemokines MCP-1 and MIP-1α and microglial activation 24 h after MCAO. WIN administration significantly reduced microglial activation at this point and attenuated infarct volume and microglial accumulation and proliferation in the injured cortex 72 h after MCAO. Cumulatively, our results show that the cannabinoid agonist WIN protects against neonatal focal stroke in part due to inhibitory effects on microglia.

Longitudinal studies of ischemic penumbra by using 18F-FDG PET and MRI techniques in permanent and transient focal cerebral ischemia in rats.

At present, the goal of stroke research is the identification of a potential recoverable tissue surrounding the ischemic core, suggested as ischemic penumbra, with the aim of applying a treatment that attenuates the growth of this area. Our purpose was to determine whether a combination of imaging techniques, including (18)F-FDG PET and MRI could identify the penumbra area. Longitudinal studies of (18)F-FDG PET and MRI were performed in rats 3 h, 24 h and 48 h after the onset of ischemia. A transient and a permanent model of focal cerebral ischemia were performed. Regions of interest were located, covering the ischemic core, the border that progresses to infarction (recruited tissue), and the border that recovers (recoverable tissue) with early reperfusion. Analyses show that permanent ischemia produces severe damage, whereas the transient ischemia model does not produce clear damage in ADC maps at the earliest time studied. The only significant differences between values for recoverable tissue, (18)F-FDG (84±2%), ADC (108±5%) and PWI (70±8%), and recruited tissue, (18)F-FDG (77±3%), ADC (109±4%) and PWI (77±4%), are shown in (18)F-FDG ratios. We also show that recoverable tissue values are different from those in non-infarcted tissue. The combination of (18)F-FDG PET, ADC and PWI MRI is useful for identification of ischemic penumbra, with (18)F-FDG PET being the most sensitive approach to its study at early times after stroke, when a clear DWI deficit is not observed.

Lack of adrenomedullin, but not complement factor H, results in larger infarct size and more extensive brain damage in a focal ischemia model.

Adrenomedullin (AM) and its binding protein, complement factor H (FH), are expressed throughout the brain. In this study we used a brain-specific conditional knockout for AM and a complete knockout for FH to investigate the effect of these molecules on the pathophysiology of stroke. Following 48 h of middle cerebral artery permanent occlusion, there was a statistically significant infarct size increase in animals lacking AM when compared to their wild type littermates. In contrast, lack of FH did not affect infarct volume. To investigate some of the mechanisms by which lack of AM may augment brain damage, markers of nitrosative stress, apoptosis, and autophagy were studied at the mRNA and protein levels. There was a significant increase of inducible nitric oxide synthase (iNOS), matrix metalloproteinase-9 (MMP9), fractin, and Beclin-1 in the peri-infarct area of AM-deficient mice when compared to their wild type counterparts and to contralateral and sham-operated controls. These data suggest that AM exerts a neuroprotective action in the brain and that this protection may be mediated by regulation of iNOS, matrix metalloproteases, and inflammatory mediators. In the future, substances that increase AM actions in the central nervous system may be used as potential neuroprotective agents in stroke.

Plasma levels of 15d-PGJ are not altered in multiple sclerosis.

BACKGROUND: The 15-deoxi delta prostaglandin J(2) (15d-PGJ(2)) is a peroxisome proliferator-activated receptor-gamma agonist with potent anti-inflammatory properties. It has been suggested that 15d-PGJ(2) may modulate multiple sclerosis (MS). METHODS: Here, we investigated the plasma levels of 15d-PGJ(2) by enzyme-linked immunoassay in 28 healthy controls and 140 MS patients [30 patients with primary-progressive MS, 28 patients with secondary-progressive MS, and 82 patients with relapsing-remitting MS (28 patients during clinical remission, 25 patients during relapse, and 29 treated with interferon-beta - IFN-beta)]. RESULTS: Levels of 15d-PGJ(2) were similar between healthy controls and untreated MS patients with different clinical courses of the disease. Treatment with IFN-beta had no effect on levels of 15d-PGJ(2). CONCLUSIONS: Although these findings suggest that 15d-PGJ(2) is not involved in the acute or chronic phases of the disease, further studies measuring 15d-PGJ(2) in cerebrospinal fluid samples are needed before excluding a role of 15d-PGJ(2) in MS.

Delayed post-ischemic administration of CDP-choline increases EAAT2 association to lipid rafts and affords neuroprotection in experimental stroke.

Glutamate transport is the only mechanism for maintaining extracellular glutamate concentrations below excitotoxic levels. Among glutamate transporters, EAAT2 is responsible for up to 90% of all glutamate transport and has been reported to be associated to lipid rafts. In this context, we have recently shown that CDP-choline induces EAAT2 translocation to the membrane. Since CDP-choline preserves membrane stability by recovering levels of sphingomyelin, a glycosphingolipid present in lipid rafts, we have decided to investigate whether CDP-choline increases association of EAAT2 transporter to lipid rafts. Flotillin-1 was used as a marker of lipid rafts due to its known association to these microdomains. After gradient centrifugation, we have found that flotillin-1 appears mainly in fractions 2 and 3 and that EAAT2 protein is predominantly found colocalised with flotillin-1 in fraction 2. We have also demonstrated that CDP-choline increased EAAT2 levels in fraction 2 at both times examined (3 and 6 h after 1 g/kg CDP-choline administration). In agreement with this, [(3)H] glutamate uptake was also increased in flotillin-associated vesicles obtained from brain homogenates of animals treated with CDP-choline. Exposure to middle cerebral artery occlusion also increased EAAT2 levels in lipid rafts, an effect which was further enhanced in those animals receiving 2 g/kg CDP-choline 4 h after the occlusion. Infarct volume measured at 48 h after ischemia showed a reduction in the group treated with CDP-choline 4 h after occlusion. In summary, we have demonstrated that CDP-choline redistributes EAAT2 to lipid raft microdomains and improves glutamate uptake. This effect is also found after experimental stroke, when CDP-choline is administered 4 h after the ischemic occlusion. Since we have also shown that this delayed post-ischemic administration of CDP-choline induces a potent neuroprotection, our data provides a novel target for neuroprotection in stroke.

Velázquez: farmacología básica y clínica

Prefacio, los directores. Prólogo a la primera edición (1930), Teófilo Hernando. Farmacología básica. Sistema nervioso periférico. Sistema nervioso central. Aparato cardiovascular. Autacoides, inflamación y respuesta inmunológica. Aparato digestivo. Sistema nervioso endocrino. Aparato respiratorio. Sangre. Quimioterapia antiinfecciosa y antitumoral. Miscelánea. Farmacología clínica. Variabilidad de la repsuesta farmacológica. Efectos no deseados de los medicamentos. Evaluación de los efectos de los medicamentos. Evaluación y mejora del uso de medicamentos. Nuevas perspectivas

Velázquez: farmacología básica y clínica

Prefacio, los directores. Prólogo a la primera edición (1930), Teófilo Hernando. Farmacología básica. Sistema nervioso periférico. Sistema nervioso central. Aparato cardiovascular. Autacoides, inflamación y respuesta inmunológica. Aparato digestivo. Sistema nervioso endocrino. Aparato respiratorio. Sangre. Quimioterapia antiinfecciosa y antitumoral. Miscelánea. Farmacología clínica. Variabilidad de la repsuesta farmacológica. Efectos no deseados de los medicamentos. Evaluación de los efectos de los medicamentos. Evaluación y mejora del uso de medicamentos. Nuevas perspectivas

Statin treatment withdrawal in ischemic stroke: a controlled randomized study.

BACKGROUND: Pretreatment with statins has been shown to reduce brain injury in cerebral ischemia. In this controlled randomized study, we investigated the influence of statin pretreatment and its withdrawal on the outcome of acute ischemic stroke patients. METHODS: From 215 patients admitted within 24 hours of a hemispheric ischemic stroke, 89 patients on chronic statin treatment were randomly assigned either to statin withdrawal for the first 3 days after admission (n = 46) or to immediately receive atorvastatin 20 mg/day (n = 43). The primary outcome event was death or dependency (modified Rankin Scale [mRS] score > 2) at 3 months. Early neurologic deterioration (END) and infarct volume at days 4 to 7 were secondary outcome variables. In a secondary analysis, outcome variables were compared with the nonrandomized patients without previous statin therapy (n = 126). RESULTS: Patients with statin withdrawal showed a higher frequency of mRS score > 2 at the end of follow-up (60.0% vs 39.0%; p = 0.043), END (65.2% vs 20.9%; p < 0.0001), and greater infarct volume (74 [45, 126] vs 26 [12, 70] mL; p = 0.002) compared with the non-statin-withdrawal group. Statin withdrawal was associated with a 4.66 (1.46 to 14.91)-fold increase in the risk of death or dependency, a 8.67 (3.05 to 24.63)-fold increase in the risk of END, and an increase in mean infarct volume of 37.63 mL (SE 10.01; p < 0.001) after adjusting for age and baseline stroke severity. Compared with patients without previous treatment with statins, statin withdrawal was associated with a 19.01 (1.96 to 184.09)-fold increase in the risk of END and an increase in mean infarct volume of 43.51 mL (SE 21.91; p = 0.048). CONCLUSION: Statin withdrawal is associated with increased risk of death or dependency at 90 days. Hence, this treatment should be continued in the acute phase of ischemic stroke.

A chronic treatment with CDP-choline improves functional recovery and increases neuronal plasticity after experimental stroke.

Chronic impairment of forelimb and digit movement is a common problem after stroke that is resistant to therapy. Although in the last years some studies have been performed to increase the efficacy of rehabilitative experience and training, the pharmacological approaches in this context remain poorly developed. We decided to study the effect of a chronic treatment with CDP-choline, a safe and well-tolerated drug that is known to stabilize membranes, on functional outcome and neuromorphological changes after stroke. To assess the functional recovery we have performed the staircase reaching test and the elevated body swing test (EBST), for studying sensorimotor integration and asymmetrical motor function respectively. The treatment with CDP-choline, initiated 24 h after the middle cerebral artery occlusion (MCAO) and maintained during 28 days, improved the functional outcome in both the staircase test (MCAO+CDP=87.0+/-6.6% pellets eaten vs. MCAO+SAL=40.0+/-4.5%; p<0.05) and the EBST (MCAO+CDP=70.0+/-6.8% vs. MCAO+SAL=88.0+/-5.4%; contralateral swing p<0.05). In addition, to study potential neuronal substrates of the improved function, we examined the dendritic morphology of layer V pyramidal cells in the undamaged motor cortex using a Golgi-Cox procedure. The animals treated with CDP-choline showed enhanced dendritic complexity and spine density compared with saline group. Our results suggest that a chronic treatment with CDP-choline initiated 24 h after the insult is able to increase the neuronal plasticity within noninjured and functionally connected brain regions as well as to promote functional recovery.