Walace Gomes-Leal

Long-term accumulation of microglia with proneurogenic phenotype concomitant with persistent neurogenesis in adult subventricular zone after stroke.

Neural stem cells (NSCs) in the adult rat subventricular zone (SVZ) generate new striatal neurons during several months after ischemic stroke. Whether the microglial response associated with ischemic injury extends into SVZ and influences neuroblast production is unknown. Here, we demonstrate increased numbers of activated microglia in ipsilateral SVZ concomitant with neuroblast migration into the striatum at 2, 6, and 16 weeks, with maximum at 6 weeks, following 2 h middle cerebral artery occlusion in rats. In the peri‐infarct striatum, numbers of activated microglia peaked already at 2 weeks and declined thereafter. Microglia in SVZ were resident or originated from bone marrow, with maximum proliferation during the first 2 weeks postinsult. In SVZ, microglia exhibited ramified or intermediate morphology, signifying a downregulated inflammatory profile, whereas amoeboid or round phagocytic microglia were frequent in the peri‐infarct striatum. Numbers of microglia expressing markers of antigen‐presenting cells (MHC‐II, CD86) increased in SVZ but very few lymphocytes were detected. Using quantitative PCR, strong short‐ and long‐term increase (at 1 and 6 weeks postinfarct) of insulin‐like growth factor‐1 (IGF‐1) gene expression was detected in SVZ tissue. Elevated numbers of IGF‐1‐expressing microglia were found in SVZ at 2, 6, and 16 weeks after stroke. At 16 weeks, 5% of microglia but no other cells in SVZ expressed the IGF‐1 protein, which mitigates apoptosis and promotes proliferation and differentiation of NSCs. The long‐term accumulation of microglia with proneurogenic phenotype in the SVZ implies a supportive role of these cells for the continuous neurogenesis after stroke.

Astrocytosis, microglia activation, oligodendrocyte degeneration, and pyknosis following acute spinal cord injury

Glial activation and degeneration are important outcomes in the pathophysiology of acute brain and spinal cord injury (SCI). Our main goal was to investigate the pattern of glial activation and degeneration during secondary degeneration in both gray matter (GM) and white matter (WM) following SCI. Adult rats were deeply anesthetized and injected with 20 nmol of N-methyl-D-aspartate (NMDA) into the ventral horn of rat spinal cord (SC) on T7. Animals were perfused after survival times of 1, 3, and 7 days. Ten-micrometer sections were submitted to immunocytochemistry for activated macrophages/microglia, astrocytes, oligodendrocytes, and myelin. Astrocyte activation was more intense in the vacuolated white matter than in gray matter and was first noticed in this former region. Microglial activation was more intense in the gray matter and was clear by 24 h following NMDA injection. Both astrocytosis and microglial activation were more intense in the later survival times. Conspicuous WM vacuolation was present mainly at the 3-day survival time and decreased by 7 days after the primary damage. Quantitative analysis revealed an increase in the number of pyknotic bodies mainly at the 7-day survival time in both ventral and lateral white matter. These pyknotic bodies were frequently found inside white matter vacuoles like for degenerating oligodendrocytes. These results suggest a differential pattern of astrocytosis and microglia activation for white and gray matter following SCI. This phenomenon can be related to the different pathological outcomes for this two SC regions following acute injury.

Microglial physiopathology: how to explain the dual role of microglia after acute neural disorders?

Microglia are the resident macrophages of the central nervous system (CNS). In physiological conditions, resting microglia maintain tissue integrity by scanning the entire CNS parenchyma through stochastic and complex movements of their long processes to identify minor tissue alterations. In pathological conditions, over‐activated microglia contribute to neuronal damage by releasing harmful substances, including inflammatory cytokines, reactive oxygen species, and proteinases, but they can provide tissue repair by releasing anti‐inflammatory cytokines and neurotrophic factors. The reasons for this apparent paradox are unknown. In this paper, we first review the physiological role as well as both detrimental and beneficial actions of microglial during acute CNS disorders. Further, we discuss the possible reasons for this microglial dual role following CNS insults, considering that the final microglial phenotype is a direct consequence of both noxious and beneficial stimuli released into the extracellular space during the pathological insult. The nature of these micro‐glial ligands is unknown, but we hypothesize that harmful and beneficial stimuli may be preferentially located at specific anatomical niches along the pathological environment triggering both beneficial and deleterious actions of these glial cells. According to this notion, there are no natural populations of detrimental microglia, but is the pathological environment that determines the final microglial phenotype.

Modulation of microglial activation enhances neuroprotection and functional recovery derived from bone marrow mononuclear cell transplantation after cortical ischemia

Activated microglia may exacerbate damage in neural disorders; however, it is unknown how they affect stem cells transplanted after stroke. Focal ischemia was induced by microinjections of 40 pmol of endothelin-1 into the motor cortex of adult rats. Ischemic animals were treated with sterile saline (n = 5), bone marrow mononuclear cells (BMMCs, n = 8), minocycline (n = 5) or concomitantly with minocycline and BMMCs (n = 5). BMMC-treated animals received 5 × 10(6)BMMCs through the caudal vein 24h post-ischemia. Behavioral tests were performed to evaluate functional recovery. Morphometric and histological analyses were performed to assess infarct area, neuronal loss and microglia/macrophage activation up to 21 days post-ischemia. Treatments with minocycline, BMMCs or minocycline-BMMCs reduced infarct area, increased neuronal survival and decreased the number of caspase-3+ and ED-1+ cells, but these effects were more prominent in the minocycline-BMMC group. Behavioral analyses using the modified sticky-tape and open-field tests showed that ischemic rats concomitantly treated with BMMCs and minocycline showed better motor performance than rats treated with BMMCs or minocycline only. The results suggest that proper modulation of the inflammatory response through the blockage of microglia activation enhances neuroprotection and functional recovery induced by intravenous transplantation of BMMCs after motor cortex ischemia.

Systematic analysis of axonal damage and inflammatory response in different white matter tracts of acutely injured rat spinal cord

The mechanisms of white matter (WM) damage during secondary degeneration are a fundamental issue in the pathophysiology of central nervous system (CNS) diseases. Our main goal was to describe the pattern of an acute inflammatory response and secondary damage to axons in different WM tracts of acutely injured rat spinal cord. Adult rats were deeply anesthetized and injected with 20 nmol of NMDA into the spinal cord ventral horn on T7. Animals were perfused after survival times of 1 day, 3 days and 7 days. Ten micrometer sections were submitted to immunocytochemical analysis for activated macrophages/microglia, neutrophils and damaged axons. There were inflammatory response and progressive tissue destruction of ventral WM (VWM) with formation of microcysts in both VWM and lateral WM (LWM). In the VWM, the number of beta-amyloid precursor protein (beta-APP) end-bulbs increased from 1 day with a peak at 3 days, decreasing by 7 days following the injection. APP end-bulbs were present in the dorsal WM (DWM) at 3 days survival time but were not in the LWM. Electron microscopic analysis revealed different degrees of myelin disruption and axonal pathology in the vacuolated WM up to 14 mm along the rostrocaudal axis. Quantitative analysis revealed a significant loss of medium and large axons (P < 0.05), but not of small axons (P > 0.05). Our results suggest that bystander axonal damage and myelin vacuolation are important secondary component of the pathology of WM tracts following rat SCI. Further studies are needed to understand the mechanisms of these pathological events.

Inflammatory response and white matter damage after microinjections of endothelin-1 into the rat striatum

Following acute and chronic neurodegenerative disorders, a cascade of pathological events including inflammatory response, excitotoxicity and oxidative stress induces secondary tissue loss in both gray and white matter. Axonal damage and demyelination are important components of the white matter demise during these diseases. In spite of this, a few studies have addressed the patterns of inflammatory response, axonal damage and demyelination following focal ischemic damage to the central nervous system (CNS). In the present study, we describe the patterns of inflammatory response, axonal damage and myelin impairment following microinjections of 10 pmol of endothelin-1 into the rat striatum. Animals were perfused at 1 day, 3 days and 7 days after injection. 20 mum sections were stained by hematoxylin and immunolabeled for neutrophils (anti-MBS-1), activated macrophages/microglia (anti-ED1), damaged axons (anti-betaAPP) and myelin (anti-MBP). The evolution of acute inflammation was quantitatively assessed by cell counts in different survival times. There was recruitment of both neutrophils and macrophages to the damaged striatal parenchyma with maximum recruitment at 1 day and 7 days, respectively. Progressive myelin impairment in the striatal white matter tracts has been observed mainly at later survival times. beta-APP+ endbulbs were not present in all evaluated time points. These results suggest that progress myelin impairment in the absence of damage to axonal cylinder is a feature of white matter pathology following endothelin-1-induced focal striatal ischemia.

Minocycline treatment reduces white matter damage after excitotoxic striatal injury

We investigated the protective effects of minocycline following white matter damage (WMD) in the rat striatum. Excitotoxic lesions were induced by N-Methyl-d-Aspartate (NMDA) microinjections and caused striatal damage, concomitant with microglial/macrophage activation. The excitotoxic lesion both damaged oligodendrocytes (Tau-1(+) cells) and caused a decrease in tissue reactivity for myelin basic protein (MBP) after post-lesional day 3 (PLD). Treatment with the semi-synthetic tetracycline antibiotic minocycline, however, led to oligodendrocyte preservation and decreased myelin impairment. Taken together, these results suggest that white matter damage (WMD) is an important component of the physiopathology of acute striatal damage and that microglial/macrophage activation contributes to this pathological phenomenon.

Masticatory deficiency as a risk factor for cognitive dysfunction.

Several studies have demonstrated that chewing helps to maintain cognitive functions in brain regions including the hippocampus, a central nervous system (CNS) region vital for memory and learning. Epidemiological studies suggest that masticatory deficiency is associated with development of dementia, which is related to spatial memory deficits especially in older animals. The purpose of this paper is to review recent work on the effects of masticatory impairment on cognitive functions both in experimental animals and humans. We show that several mechanisms may be involved in the cognitive deficits associated with masticatory deficiency. The epidemiological data suggest a positive correlation between masticatory deficit and Alzheimers disease. It may be concluded that chewing has important implications for the mechanisms underlying certain cognitive abilities.

Chronic ethanol exposure during adolescence through early adulthood in female rats induces emotional and memory deficits associated with morphological and molecular alterations in hippocampus

There is increasing evidence that heavy ethanol exposure in early life may produce long-lasting neurobehavioral consequences, since brain structural maturation continues until adolescence. It is well established that females are more susceptible to alcohol-induced neurotoxicity and that ethanol consumption is increasing among women, especially during adolescence. In the present study, we investigated whether chronic ethanol exposure during adolescence through early adulthood in female rats may induce hippocampal histological damage and neurobehavioral impairments. Female rats were treated with distilled water or ethanol (6.5 g/kg/day, 22.5% w/v) by gavage from the 35th–90th day of life. Ethanol-exposed animals displayed reduced exploration of the central area and increased number of fecal boluses in the open field test indicative of anxiogenic responses. Moreover, chronic high ethanol exposure during adolescence induced marked impairments on short-term memory of female rats addressed on social recognition and step-down inhibitory avoidance tasks. These neurobehavioral deficits induced by ethanol exposure during adolescence through early adulthood were accompanied by the reduction of hippocampal formation volume as well as the loss of neurons, astrocytes and microglia cells in the hippocampus. These results indicate that chronic high ethanol exposure during adolescence through early adulthood in female rats induces long-lasting emotional and memory deficits associated with morphological and molecular alterations in the hippocampus.

Hippocampal neuronal loss, decreased GFAP immunoreactivity and cognitive impairment following experimental intoxication of rats with aluminum citrate

Aluminum (Al) is a neurotoxic agent with deleterious actions on cognitive processes. Nevertheless, few studies have investigated the neuropathological effects underlying the Al-induced cognitive impairment. We have explored the effects of acute Al citrate intoxication on both hippocampal morphology and mnemonic processes in rodents. Adult male Wistar rats were intoxicated with a daily dose of Al citrate (320 mg/kg) during 4 days by gavage. Animals were perfused at 8 (G2), 17 (G3) and 31 days (G4) after intoxication. Control animals were treated with sodium citrate (G1). Animals were submitted to behavioral tests of open field and elevated T-maze. Immunohistochemistry was performed to label neurons (anti-NeuN) and astrocytes (anti-GFAP) in both CA1 and CA3 hippocampal regions. There was an increase in the locomotor activity in open field test for G2 in comparison to control group and other groups (ANOVA-Bonferroni, P<0.05). The elevated T-maze avoidance latency (AL) was higher in all intoxicated groups compared to control (P<0.05) in avoidance 1. These values remained elevated in avoidance 2 (P<0.05), but abruptly decreased in G2 and G3, but not in G1 and G4 animals in avoidance 3 (P<0.05). There were no significant differences for 1 and 2 escape latencies. There were intense neuronal loss and a progressive decrease in GFAP immunoreactivity in the hippocampus of intoxicated animals. The results suggest that Al citrate treatment induces deficits on learning and memory concomitant with neuronal loss and astrocyte impairment in the hippocampus of intoxicated rats.