Activation of cannabinoid system in anterior cingulate cortex and orbitofrontal cortex modulates cost-benefit decision making.

Despite the evidence for altered decision making in cannabis abusers, the role of the cannabinoid system in decision-making circuits has not been studied. Here, we examined the effects of cannabinoid modulation during cost-benefit decision making in the anterior cingulate cortex (ACC) and orbitofrontal cortex (OFC), key brain areas involved in decision making. We trained different groups of rats in a delay-based and an effort-based form of cost-benefit T-maze decision-making task. During test days, the rats received local injections of either vehicle or ACEA, a cannabinoid type-1 receptor (CB1R) agonist in the ACC or OFC. We measured spontaneous locomotor activity following the same treatments and characterized CB1Rs localization on different neuronal populations within these regions using immunohistochemistry. We showed that CB1R activation in the ACC impaired decision making such that rats were less willing to invest physical effort to gain high reward. Similarly, CB1R activation in the OFC induced impulsive pattern of choice such that rats preferred small immediate rewards to large delayed rewards. Control tasks ensured that the effects were specific for differential cost-benefit tasks. Furthermore, we characterized widespread colocalizations of CB1Rs on GABAergic axonal ends but few colocalizations on glutamatergic, dopaminergic, and serotonergic neuronal ends. These results provide first direct evidence that the cannabinoid system plays a critical role in regulating cost-benefit decision making in the ACC and OFC and implicate cannabinoid modulation of synaptic ends of predominantly interneurons and to a lesser degree other neuronal populations in these two frontal regions.

Increased physical activity is not enough to recover astrocytic population from dark-rearing. Synergy with multisensory enrichment is required.

Elimination of sensory inputs (deprivation) modifies the properties of the sensory cortex and serves as a model for studying plasticity during postnatal development. Many studies on the effects of deprivation have been performed in the visual cortex using dark-rearing as a visual deprivation model. It induces changes in all cellular and molecular components, including astrocytes, which play an important role in the development, maintenance, and plasticity of the cortex, mediated by cytokines which have been termed angioglioneurins. When one sense is deprived, a compensatory mechanism called cross-modal plasticity increases performance in the remaining senses. Environmental enrichment is so far the best-known method to compensate sensorial deprivation. The aim of this work is to study the effects of exercise alone, and of an enriched environment combined with exercise, on astroglial population in order to observe the effects of exercise by itself, or the potential synergistic effect during the rat visual system development. Pregnant Sprague-Dawley rats were raised in one of the following rearing conditions: in total darkness and enriched environment conditions with physical exercise, and in total darkness with voluntary physical exercise. Astrocytic density was estimated by immunohistochemistry for S-100ß protein and quantifications were performed in layer IV. The somatosensorial cortex barrel field was also studied as control. Our main result shows that an enriched environment combined with voluntary physical exercise manages to reverse the negative effects induced by darkness over the astroglial population of both the visual and the somatosensory cortices. On the other hand, exercise alone only produces effects upon the astroglial population of the somatosensory cortex, and less so when combined with an enriched environment.

Angiogenic signalling pathways altered in gliomas: selection mechanisms for more aggressive neoplastic subpopulations with invasive phenotype.

The angiogenesis process is a key event for glioma survival, malignancy and growth. The start of angiogenesis is mediated by a cascade of intratumoural events: alteration of the microvasculature network; a hypoxic microenvironment; adaptation of neoplastic cells and synthesis of pro-angiogenic factors. Due to a chaotic blood flow, a consequence of an aberrant microvasculature, tissue hypoxia phenomena are induced. Hypoxia inducible factor 1 is a major regulator in glioma invasiveness and angiogenesis. Clones of neoplastic cells with stem cell characteristics are selected by HIF-1. These cells, called "glioma stem cells" induce the synthesis of vascular endothelial growth factor. This factor is a pivotal mediator of angiogenesis. To elucidate the role of these angiogenic mediators during glioma growth, we have used a rat endogenous glioma model. Gliomas induced by prenatal ENU administration allowed us to study angiogenic events from early to advanced tumour stages. Events such as microvascular aberrations, hypoxia, GSC selection and VEGF synthesis may be studied in depth. Our data showed that for the treatment of gliomas, developing anti-angiogenic therapies could be aimed at GSCs, HIF-1 or VEGF. The ENU-glioma model can be considered to be a useful option to check novel designs of these treatment strategies.

Enriched and deprived sensory experience induces structural changes and rewires connectivity during the postnatal development of the brain.

During postnatal development, sensory experience modulates cortical development, inducing numerous changes in all of the components of the cortex. Most of the cortical changes thus induced occur during the critical period, when the functional and structural properties of cortical neurons are particularly susceptible to alterations. Although the time course for experience-mediated sensory development is specific for each system, postnatal development acts as a whole, and if one cortical area is deprived of its normal sensory inputs during early stages, it will be reorganized by the nondeprived senses in a process of cross-modal plasticity that not only increases performance in the remaining senses when one is deprived, but also rewires the brain allowing the deprived cortex to process inputs from other senses and cortices, maintaining the modular configuration. This paper summarizes our current understanding of sensory systems development, focused specially in the visual system. It delineates sensory enhancement and sensory deprivation effects at both physiological and anatomical levels and describes the use of enriched environment as a tool to rewire loss of brain areas to enhance other active senses. Finally, strategies to apply restorative features in human-deprived senses are studied, discussing the beneficial and detrimental effects of cross-modal plasticity in prostheses and sensory substitution devices implantation.

Vascular endothelial growth factor and other angioglioneurins: key molecules in brain development and restoration.

Angioneurines are a family of molecules that include vascular growth factors such as VEGF, neurotrophins such as BDNF, IGF-I, and Erythropoietin, among others. They affect both neural and vascular processes. Due to the fact that all of them act over glia, we propose the term angioglioneurins to name them. They play a key role in the neurogliovascular unit that represents the functional core maintaining BBB. Although delivery to CNS is still an unsolved problem nowadays, exogenous angioglioneurin administration represents a promising therapeutic strategy for many neurological pathologies due to their neurotrophic and neurogenic role. In brains, VEGF is produced by neurons and astrocytes in different stages and situation, binding to tyrosine kinase receptors and also to neuropilin family. This fact reinforces its key role in the cross talk between neural and vascular development and activity. Angioglioneurins described in this report might become an important therapeutic resource in CNS restoration, especially in pathologies as stroke or traumatic brain injury.

Effect of intracortical vascular endothelial growth factor infusion and blockade during the critical period in the rat visual cortex.

VEGF is the major angiogenic and vascular permeability factor in health and disease. Vascular development depends on function, and in sensory areas is experience-dependent. Our aim was to investigate, qualitatively and quantitatively, the effects of intracortical infusion and neutralisation of VEGF during the first days of the critical visual period, when peak levels of endogenous VEGF secretion are reached. VEGF was intracortically delivered into middle cortical layers of P18 Long-Evans rats. Another cohort received anti-VEGF. Vehicle (PBS)-infused and non-operated animals were used as controls. Various immunopathological analyses were performed: Endothelial Barrier Antigen (EBA) for the BBB integrity and GFAP for astroglial response. Vascular density was measured by Butyryl Cholinesterase Histochemistry, neuronal density by NeuN immunohistochemistry and apoptosis by TUNEL staining. VEGF levels were measured by Western Blot. Decreased vascular permeability was evoked in VEGF-infused rats whilst EBA expression remained constant, suggesting a preserved BBB function. When VEGF was blocked, tissue showed a higher degree of extravasation and a decreased number of EBA-positive vessels surrounding the injury. Lesion induced by cannula implantation annulled the normal increase in vascular density and the decrease in neuronal density during this time. VEGF rescued in part the vascular increase, and also prevented physiological and pathological neuronal death. VEGF blockade induced a higher amount of neural loss and lower astrocytic reaction. Our results support the role of VEGF as extending beyond vascularization, preventing physiological and pathological neuronal death, not only in the injured hemisphere but also in the intact one suggesting a process of transhemispheric diaschisis.

Broad characterization of endogenous peptides in the tree shrew visual system.

Endogenous neuropeptides, acting as neurotransmitters or hormones in the brain, carry out important functions including neural plasticity, metabolism and angiogenesis. Previous neuropeptide studies have focused on peptide-rich brain regions such as the striatum or hypothalamus. Here we present an investigation of peptides in the visual system, composed of brain regions that are generally less rich in peptides, with the aim of providing the first broad overview of peptides involved in mammalian visual functions. We target three important parts of the visual system: the primary visual cortex (V1), lateral geniculate nucleus (LGN) and superior colliculus (SC). Our study is performed in the tree shrew, a close relative of primates. Using a combination of data dependent acquisition and targeted LC-MS/MS based neuropeptidomics; we identified a total of 52 peptides from the tree shrew visual system. A total of 26 peptides, for example GAV and neuropeptide K were identified in the visual system for the first time. Out of the total 52 peptides, 27 peptides with high signal-to-noise-ratio (>10) in extracted ion chromatograms (EIC) were subjected to label-free quantitation. We observed generally lower abundance of peptides in the LGN compared to V1 and SC. Consistently, a number of individual peptides showed high abundance in V1 (such as neuropeptide Y or somatostatin 28) and in SC (such as somatostatin 28 AA1-12). This study provides the first in-depth characterization of peptides in the mammalian visual system. These findings now permit the investigation of neuropeptide-regulated mechanisms of visual perception.

Vascular endothelial growth factor: adaptive changes in the neuroglialvascular unit.

Brain postnatal development is modulated by adaptation and experience. Experience-mediated changes increase neuronal activity leading to increased metabolic demands that involve adaptive changes including ones at the microvascular network. Therefore, vascular environment plays a key role in central nervous system (CNS) development and function in health and disease. Trophic factors are crucial in CNS development and cell survival in adults. They participate in protection and proliferation of neuronal, glial and endothelial cells. Among the most important molecules are: the proangiogenic vascular endothelial growth factor (VEGF), the neurotrophin brain derived neurotrophic factor (BDNF), insulin growth factor (IGF-I) and the glycoprotein erythropoietin (EPO). We propose the term angioglioneurins to define molecules acting on the three components of the neurogliovascular unit. We have previously reported the effects of environmental modifications on the three components of the neurogliovascular unit during the postnatal development. We have also described the main role played by VEGF in the experience-induced postnatal changes. Angioglioneurin administration, alone or in combination with other neuroprotective strategies such as environmental enrichment, has been proposed as a non-invasive therapeutic strategy against several CNS diseases.

[Glial stem cells and their relationship with tumour angiogenesis process]. / Células madre gliales y su relación en el proceso de angiogénesis tumoral.

INTRODUCTION: A subpopulation of neoplastic cells with characteristics of stem cells has been described on human multiform glioblastomas. These cells play a pivotal role in tumour angiogenesis and malignancy being involved in infiltration of adjacent normal parenchyma. The named glial stem cells could be responsible for recurrences after surgery. This is due to their survival capacity after quimio/radiotherapy treatments. DEVELOPMENT: In this work we review the role of glial stem cells in relationship with angiogenesis process. We also review some findings related to the appearance of these cells during angiogenesis in a rat endogenous experimental model of gliomas. These cells were characterized by antibodies against the antigens CD133, nestin and the vascular endothelial growth factor (VEGF). Nestin+ cells were found in every stage of tumour development, whereas CD133+ cells were only present since intermediates stages corresponding with VEGF overexpression. This moment is known as start of angiogenesis or 'angiogenic switch'. We also found that some nestin+ cells co-expressed CD133 antigen. Glial stem cells are distributed in the experimental glioma model as well as in human multiform glioblastomas, shaping niches into perivascular or intra-tumoral hypoxic areas. CONCLUSION: Many evidences corroborate the hypothesis that glial stem cells have a close relationship with angiogenic switch, intratumor hypoxia and neoplastic microvascular network.

Combination of intracortically administered VEGF and environmental enrichment enhances brain protection in developing rats.

Postnatal development of the visual cortex is modulated by experience, especially during the critical period. In rats, a stable neuronal population is only acquired after this relatively prolonged period. Vascular endothelial growth factor (VEGF) is the most important angiogenic factor and also has strong neuroprotective, neurotrophic and neurogenic properties. Similar effects have been described for rearing in enriched environments. Our aim is to investigate the vascular and neuronal effects of combining VEGF infusion and environmental enrichment on the visual cortex during the initial days of the critical period. Results showed that a small percentage of Cleaved Caspase-3 positive cells colocalized with neuronal markers. The lesion produced by the cannula implantation resulted in decreased vascular, neuronal and Caspase-3 positive cell densities. Rearing under enriched environment was unable to reverse these effects in any group, whereas VEGF infusion alone partially corrected those effects. A higher effectiveness was reached by combining both the procedures, the most effective combination being when enriched-environment rearing was introduced only after minipump implantation. In addition to the angiogenic effect of VEGF, applied strategies also had synergic neuroprotective effects, and the combination of the two strategies had more remarkable effects than those achieved by each strategy applied individually.

Physical exercise is required for environmental enrichment to offset the quantitative effects of dark-rearing on the S-100beta astrocytic density in the rat visual cortex.

After birth, exposure to visual inputs modulates cortical development, inducing numerous changes in all of the components of the visual cortex. Most of the cortical changes thus induced occur during what is called the critical period. Astrocytes play an important role in the development, maintenance and plasticity of the cortex as well as in the structure and function of the vascular network. Visual deprivation induces a decrease in the astroglial population, whereas enhanced experience increases it. Exposure to an enriched environment has been shown to prevent the effects of dark-rearing in the visual cortex. Our purpose was to study the effects of an enriched environment on the density of astrocytes per reference surface at the visual cortex of dark-reared rats, in order to determine if enhanced experience is able to compensate the quantitative effects of visual deprivation and the role of physical exercise on the enrichment paradigm. Pregnant Sprague-Dawley rats were raised in one of the following rearing conditions: control rats with standard housing (12-h light/dark cycle); in total darkness for the dark-rearing experiments; and dark-rearing in conditions of enriched environment without and with physical exercise. The astrocytic density was estimated by immunohistochemistry for S-100beta protein. Quantifications were performed in layer IV. The somatosensorial cortex barrel field was also studied as control. The volume of layer IV was stereologically calculated for each region, age and experimental condition. From the beginning of the critical period, astrocyte density was higher in control rats than in the enriched environment group without physical exercise, with densities of astrocytes around 20% higher at all of the different ages. In contrast, when the animals had access to voluntary exercise, densities were significantly higher than even the control rats. Our main result shows that strategies to apply environmental enrichment should always consider the incorporation of physical exercise, even for sensorial areas such as the visual area, where complex enriched experience by itself is not enough to compensate the effects of visual deprivation.

Effects of visual experience on vascular endothelial growth factor expression during the postnatal development of the rat visual cortex.

The development of the cortical vascular network depends on functional maturation. External inputs are an essential requirement in the modeling of the visual cortex, mainly during the critical period, when the functional and structural properties of visual cortical neurons are particularly susceptible to alterations. Vascular endothelial growth factor (VEGF) is the major angiogenic factor, a key signal in the induction of vessel growth. Our study focused on the role of visual stimuli on the development of the vascular pattern correlated with VEGF levels. Vascular density and the expression of VEGF were examined in the primary visual cortex of rats reared under different visual environments (dark rearing, dark-rearing in conditions of enriched environment, enriched environment, and laboratory standard conditions) during postnatal development (before, during, and after the critical period). Our results show a restricted VEGF cellular expression to astroglial cells. Quantitative differences appeared during the critical period: higher vascular density and VEGF protein levels were found in the enriched environment group; both dark-reared groups showed lower vascular density and VEGF levels, which means that enriched environment without the physical exercise component does not exert effects in dark-reared rats.

Spatio-temporal distribution of apoptosis and the modulators thereof following a cortical microinfarct in rat brain.

Two mechanisms of brain cell death coexist, necrosis and apoptosis. We investigated the correlation between the apoptotic index and the expression of apoptosis modulators and stress response in an ultraviolet-induced cortical microinfarct. Adult rat neocortex was exposed to an ultraviolet beam and brains removed at different intervals after injury were paraffin-embedded and processed for TUNEL assay and immunohistochemistry against apoptotic modulators Bax and Bcl-2, and stress protein HSP70. During the 12-72h postirradiation period, apoptotic nuclei decreased from 11% to 4% in the infarcted area whereas only 1.2% of such nuclei was seen in the perilesional area. While Bcl-2 was always negative in the lesion focus, Bax was positive at all survival times, mainly in glial cells. HSP70 was expressed over a broad area of the ipsilateral hemisphere from 3h after brain injury, firstly in neurons and progressively in glial cells and finally in endothelium. At longer survival times, positive cells could be also seen in the contralateral hemisphere. Apoptosis seems to play only a quantitatively modest role in the progression of brain damage in penumbra areas despite the wide expression of pro-apoptotic factors. On the other hand HSP70 appears to be one of the main protective responses to injury stress.

Lack of experience-mediated differences in the immunohistochemical expression of blood-brain barrier markers (EBA and GluT-1) during the postnatal development of the rat visual cortex.

The development of the cortical vascular tree depends on functional development. External inputs are an essential requirement in the modeling of the visual cortex, mainly during the critical period, when congruous blood supply is needed. The blood brain barrier (BBB) function regulates the passage of substances between the blood and the brain parenchyma, which is one of the main differential features of central nervous system (CNS) microvessels. The endothelial barrier antigen (EBA) has been reported as a specific marker for the BBB physiological function in rats. We studied the postnatal development of EBA expression in the visual cortex of rats reared under opposite paradigms of visual experience, e.g., standard laboratory conditions, dark rearing, and enriched environment at 14, 21, 28, 35, 42, 49, 56, and 63 days postnatal (dpn). Parallel sections were immunohistochemically processed for endothelial barrier antigen (EBA) and glucose transporter-1 (GluT-1). Total vasculature was quantified by Lycopersicon esculentum (LEA) lectin histochemistry. No differences in EBA expression were found between groups, although quantitative differences were recorded paralleling differences in vascular density. Paradoxically, there was no expression in certain cortical vessels which were GluT-1 immunopositive and positivity was consistent in non-barrier areas such as the pineal gland. These findings were completely independent of age or experimental conditions. Therefore, the role of the EBA antigen in the BBB remains unclear: it has been undeniably linked to vascular permeability, but its presence in non-barrier vessels suggests another vascular function. Although visual experience modifies vascular density in the visual cortex, it has not been shown to have an influence on the maturation of the BBB function.

Visual deprivation effects on the s100beta positive astrocytic population in the developing rat visual cortex: a quantitative study.

After birth, exposure to visual inputs modulates cortical development, inducing numerous changes of all components of the visual cortex. Most of the cortical changes thus induced occur during what is called the critical period. Astrocytes play an important role in the development, maintenance and plasticity of the cortex, as well as in the structure and function of the vascular network. Dark-reared Sprague-Dawley rats and age-matched controls sampled at 14, 21, 28, 35, 42, 49, 56 and 63 days postnatal (dpn) were studied in order to elucidate quantitative differences in the number of positive cells in the striate cortex. The astrocytic population was estimated by immunohistochemistry for S-100beta protein. The same quantification was also performed in a nonsensory area, the retrosplenial granular cortex. S-100beta positive cells had adult morphology in the visual cortex at 14 dpn and their numbers were not significantly different in light-exposed and nonexposed rats up to 35 dpn, and were even higher in dark-reared rats at 21 dpn. However, significant quantitative changes were recorded after the beginning of the critical period. The main finding of the present study was the significantly lower astroglial density estimated in the visual cortex of dark-reared rats over 35 dpn as well as the lack of difference at previous ages. Our results also showed that there were no differences when comparing the measurements from a nonsensory area between both groups. This led us to postulate that the astrocytic population in the visual cortex is downregulated by the lack of visual experience.