Building momentum through networks: Bioimaging across the Americas.

In September 2023, the two largest bioimaging networks in the Americas, Latin America Bioimaging (LABI) and BioImaging North America (BINA), came together during a 1-week meeting in Mexico. This meeting provided opportunities for participants to interact closely with decision-makers from imaging core facilities across the Americas. The meeting was held in a hybrid format and attended in-person by imaging scientists from across the Americas, including Canada, the United States, Mexico, Colombia, Peru, Argentina, Chile, Brazil and Uruguay. The aims of the meeting were to discuss progress achieved over the past year, to foster networking and collaborative efforts among members of both communities, to bring together key members of the international imaging community to promote the exchange of experience and expertise, to engage with industry partners, and to establish future directions within each individual network, as well as common goals. This meeting report summarises the discussions exchanged, the achievements shared, and the goals set during the LABIxBINA2023: Bioimaging across the Americas meeting.

Neuroprotective Effects of Metformin Through AMPK Activation in a Neurotoxin-Based Model of Cerebellar Ataxia.

Cerebellar ataxia is a heterogeneous group of neural disorders clinically characterized by cerebellar dysfunction. The diagnosis of patients with progressive cerebellar ataxia is complex due to the direct correlation with other neuron diseases. Although there is still no cure for this pathological condition, some metabolic, hereditary, inflammatory, and immunological factors affecting cerebellar ataxia are being studied and may become therapeutic targets. Advances in studying the neuroanatomy, pathophysiology, and molecular biology of the cerebellum (CE) contribute to a better understanding of the mechanisms behind the development of this disorder. In this study, Wistar rats aged 30 to 35 days were injected intraperitoneally with 3-acetylpyridine (3-AP) and/or metformin (for AMP-activated protein kinase (AMPK) enzyme activation) and euthanized in 24 hours and 4 days after injection. We analyzed the neuromodulatory role of the AMPK on cerebellar ataxia induced by the neurotoxin 3-AP in the brain stem (BS) and CE, after pre-treatment for 7 and 15 days with metformin, a pharmacological indirect activator of AMPK. The results shown here suggest that AMPK activation in the BS and CE leads to a significant reduction in neuroinflammation in these regions. AMPK was able to restore the changes in fatty acid composition and pro-inflammatory cytokines caused by 3-AP, suggesting that the action of AMPK seems to result in a possible neuroprotection on the cerebellar ataxia model.

Adrenoreceptor phylogeny and novel functions of nitric oxide in ascidian immune cells.

Nitric oxide (NO) is a simple molecule involved in many biological processes and functions in the cardiovascular, neural, and immune systems. In recent years, NO has also been recognized as a crucial messenger in communication between the nervous and immune systems. Together with NO, catecholamines are the main group of neurotransmitters involved in cross-talk between the nervous and immune systems. Catecholamines such as noradrenaline, can act on immune cells through adrenoreceptors (ARs) present on the cell surface, and NO can cross the cell membrane and interact with secondary messengers, modulating catecholamine production. Here, we analyzed the mutual modulation by noradrenaline and NO in Phallusia nigra immune cells for specific subtypes of ARs. We also investigated the involvement of protein kinases A and C as secondary messengers to these specific subtypes of ARs in the adrenergic signaling pathway that culminates in NO modulation, and the phylogenetic distribution of ARs in deuterostome genomes. This analysis provided evidence for single-copy orthologs of α1, α2 and ß-AR in ascidian genomes, suggesting that NO and NA act on a less diverse set of ARs in urochordates. Pharmacological assays showed that high levels of NO can induce ascidian immune cells to produce catecholamines. We also observed that protein kinases A and C are the secondary messengers involved in downstream modulation of NO production through an ancestral ß-AR. Taken together, these results provide new information on NO as a modulator of immune cells, and reveal the molecules involved in the signaling pathway of ARs. The results also indicate that ARs may participate in NO modulation. Finally, our results suggest that the common ancestor of urochordates possessed a less complex system of ARs required for immune action and diverse pharmacological responses, since the α-ARs are phylogenetically more related to D1-receptors than are the ß-ARs.

Activation of cannabinoid type 1 receptor (CB1) modulates oligodendroglial process branching complexity in rat hippocampal cultures stimulated by olfactory ensheathing glia-conditioned medium.

The endocannabinoid system (ECS) refers to a complex cell-signaling system highly conserved among species formed by numerous receptors, lipid mediators (endocannabinoids) and synthetic and degradative enzymes. It is widely distributed throughout the body including the CNS, where it participates in synaptic signaling, plasticity and neurodevelopment. Besides, the olfactory ensheathing glia (OEG) present in the olfactory system is also known to play an important role in the promotion of axonal growth and/or myelination. Therefore, both OEG and the ECS promote neurogenesis and oligodendrogenesis in the CNS. Here, we investigated if the ECS is expressed in cultured OEG, by assessing the main markers of the ECS through immunofluorescence, western blotting and qRT-PCR and quantifying the content of endocannabinoids in the conditioned medium of these cells. After that, we investigated whether the production and release of endocannabinoids regulate the differentiation of oligodendrocytes co-cultured with hippocampal neurons, through Sholl analysis in oligodendrocytes expressing O4 and MBP markers. Additionally, we evaluated through western blotting the modulation of downstream pathways such as PI3K/Akt/mTOR and ERK/MAPK, being known to be involved in the proliferation and differentiation of oligodendrocytes and activated by CB1, which is the major endocannabinoid responsive receptor in the brain. Our data show that OEG expresses key genes of the ECS, including the CB1 receptor, FAAH and MAGL. Besides, we were able to identify AEA, 2-AG and AEA related mediators palmitoylethanolamide (PEA) and oleoylethanolamide (OEA), in the conditioned medium of OEG cultures. These cultures were also treated with URB597 10-9 M, a FAAH selective inhibitor, or JZL184 10-9 M, a MAGL selective inhibitor, which led to the increase in the concentrations of OEA and 2-AG in the conditioned medium. Moreover, we found that the addition of OEG conditioned medium (OEGCM) enhanced the complexity of oligodendrocyte process branching in hippocampal mixed cell cultures and that this effect was inhibited by AM251 10-6 M, a CB1 receptor antagonist. However, treatment with the conditioned medium enriched with OEA or 2-AG did not alter the process branching complexity of premyelinating oligodendrocytes, while decreased the branching complexity in mature oligodendrocytes. We also observed no change in the phosphorylation of Akt and ERK 44/42 in any of the conditions used. In conclusion, our data show that the ECS modulates the number and maturation of oligodendrocytes in hippocampal mixed cell cultures.

Use of invertebrates to model chemically induced parkinsonism-symptoms.

The prevalence of neurological diseases is currently growing due to the combination of several factor, including poor lifestyle and environmental imbalance which enhance the contribution of genetic factors. Parkinson's disease (PD), a chronic and progressive neurological condition, is one of the most prevalent neurodegenerative human diseases. Development of models may help to understand its pathophysiology. This review focuses on studies using invertebrate models to investigate certain chemicals that generate parkinsonian-like symptoms models. Additionally, we report some preliminary results of our own research on a crustacean (the crab Ucides cordatus) and a solitary ascidian (Styela plicata), used after induction of parkinsonism with 6-hydroxydopamine and the pesticide rotenone, respectively. We also discuss the advantages, limits, and drawbacks of using invertebrate models to study PD. We suggest prospects and directions for future investigations of PD, based on invertebrate models.

Central nervous system regeneration in ascidians: cell migration and differentiation.

Adult ascidians have the capacity to regenerate the central nervous system (CNS) and are therefore excellent models for studies on neuroregeneration. The possibility that undifferentiated blood cells are involved in adult neuroregeneration merits investigation. We analyzed the migration, circulation, and role of hemocytes of the ascidian Styela plicata in neuroregeneration. Hemocytes were removed and incubated with superparamagnetic iron oxide nanoparticles (SPION), and these SPION-labeled hemocytes were injected back into the animals (autologous transplant), followed by neurodegeneration with the neurotoxin 3-acetylpyridine (3AP). Magnetic resonance imaging showed that 1, 5, and 10 days after injury, hemocytes migrated to the intestinal region, siphons, and CNS. Immunohistochemistry revealed that the hemocytes that migrated to the CNS were putative stem cells (P-element-induced wimpy testis + or PIWI + cells). In the cortex of the neural ganglion, migrated hemocytes started to lose their PIWI labeling 5 days after injury, and 10 days later started to show ß-III tubulin labeling. In the neural gland, however, the hemocytes remained undifferentiated during the entire experimental period. Transmission electron microscopy revealed regions in the neural gland with characteristics of neurogenic niches, not previously reported in ascidians. These results showed that migration of hemocytes to the hematopoietic tissue and to the 3AP-neurodegenerated region is central to the complex mechanism of neuroregeneration.

Neuroprotective Gene Therapy by Overexpression of the Transcription Factor MAX in Rat Models of Glaucomatous Neurodegeneration.

Purpose: Based on our preview evidence that reduced nuclear content of the transcription factor Myc-associated protein X (MAX) is an early event associated with degeneration of retinal ganglion cells (RGCs), in the present study, our purpose was to test whether the overexpression of human MAX had a neuroprotective effect against RGC injury. Methods: Overexpression of either MAX or green fluorescent protein (GFP) in the retina was achieved by intravitreal injections of recombinant adenovirus-associated viruses (rAAVs). Lister Hooded rats were used in three models of RGC degeneration: (1) cultures of retinal explants for 30 hours ex vivo from the eyes of 14-day-old rats that had received intravitreal injections of rAAV2-MAX or the control vector rAAV2-GFP at birth; (2) an optic nerve crush model, in which 1-month-old rats received intravitreal injection of either rAAV2-MAX or rAAV2-GFP and, 4 weeks later, were operated on; and (3) an ocular hypertension (OHT) glaucoma model, in which 1-month-old rats received intravitreal injection of either rAAV2-MAX or rAAV2-GFP and, 4 weeks later, were subject to cauterization of the limbal plexus. Cell death was estimated by detection of pyknotic nuclei and TUNEL technique and correlated with MAX immunocontent in an ex vivo model of retinal explants. MAX expression was detected by quantitative RT-PCR. In the OHT model, survival of RGCs was quantified by retrograde labeling with DiI or immunostaining for BRN3a at 14 days after in vivo injury. Functional integrity of RGCs was analyzed through pattern electroretinography, and damage to the optic nerve was examined in semithin sections. Results: In all three models of RGC insult, gene therapy by overexpression of MAX prevented RGC death. Also, ON degeneration and electrophysiologic deficits were prevented in the OHT model. Conclusions: Our experiments offer proof of concept for a novel neuroprotective gene therapy for glaucomatous neurodegeneration based on overexpression of MAX.

Physical exercise influences astrocytes in the striatum of a Parkinson's disease male mouse model.

Physical exercise is considered an adjuvant treatment to Parkinson's disease (PD) patients, possibly reducing inflammatory responses in the brain. Studies have stated that physical exercise protects dopaminergic neurons in PD models produced by neurotoxins. However, few studies focused on immunohistochemically reacted astrocytes and morphometric analyses of these cells in a PD mouse model submitted to physical exercise. We investigated the effects of treadmill training on striatal astrocytes of a PD mouse model combining immunohistochemistry and western-blotting for glial fibrillary acidic protein (GFAP) with morphometric analyses. Male Swiss mice were divided into 4 groups: sedentary control (SEDCONT), exercise control (EXERCONT), sedentary Parkinson (SEDPD), and exercise Parkinson (EXERPD). Stereotaxic bilateral injections of 6-hydroxydopamine into the striatum were adopted for PD groups. Striatal astrocytes showed increased GFAP in EXERPD, and we observed a higher level of GFAP in EXERPD than SEDPD. The number of primary and secondary processes was similar in striatal astrocytes of control groups and EXERPD. The astrocyte primary processes of SEDPD were larger than those of EXERPD, EXERCONT and SEDCONT. Cell body diameters and areas showed no difference between groups. We concluded that physical exercise influences striatal astrocytes in exercised parkinsonian mice.

Physical exercise increases the production of tyrosine hydroxylase and CDNF in the spinal cord of a Parkinson's disease mouse model.

Previous research advocates that exercise is a non-pharmacological therapy for Parkinson's disease (PD). However, few studies have investigated the effects of exercise on central nervous system structures other than the nigrostriatal pathway by using PD animal models. This study investigated the effects of exercise on tyrosine hydroxylase (TH)- and cerebral dopamine neurotrophic factor (CDNF)-containing spinal-cord neurons. Male Swiss mice were divided into 4 groups: sedentary control (SEDCONT), exercise control (EXERCONT), sedentary Parkinson (SEDPD), and exercise Parkinson (EXERPD). The PD groups were submitted to a surgical procedure for stereotaxic bilateral injection of 6-hydroxydopamine into the striatum. TH- and CDNF-containing spinal-cord neurons were evaluated in all groups, using immunohistochemistry and western-blotting. TH content in the ventral horn differed notably between the SEDPD and EXERPD groups. CDNF content was highest in the EXERPD group. SEDPD and EXERPD groups differed the most, as shown by immunohistochemistry and western-blotting. The EXERPD group showed the most intense labeling in immunohistochemistry compared to the SEDCONT and EXERCONT groups. Therefore, we showed here that exercise increased the content of both TH and CDNF in the spinal-cord neurons of a bilateral PD mouse model. We may assume that the spinal cord is affected in a PD model, and therefore this central nervous system region deserves more attention from researchers dealing with PD.

Cell proliferation in the central nervous system of an adult semiterrestrial crab.

Neurogenesis occurs in adults of most organisms, both vertebrates and invertebrates. In semiterrestrial crabs of the infraorder Brachyura, the deutocerebrum, where neurogenesis occurs, processes the olfactory sensory information from the antennae. The deutocerebrum is composed of a pair of olfactory lobes associated with cell clusters 9 and 10 (Cl 9 and Cl 10), containing proliferating cells. Because the location of the neurogenic niche in brachyuran semiterrestrial crabs has not been defined, here we describe a neurogenic niche in the central olfactory system of the crab Ucides cordatus and report two types of glial cells in the deutocerebrum, based on different markers. Serotonin (5-hydroxytryptamine) labeling was used to reveal neuroanatomical aspects of the central olfactory system and the neurogenic niche. The results showed a zone of proliferating neural cells within Cl 10, which also contains III beta-tubulin (Tuj1)+ immature neurons, associated with a structure that has characteristics of the neurogenic niche. For the first time, using two glial markers, glial fibrillary acidic protein (GFAP) and glutamine synthetase (GS), we identified two types of astrocyte-like cells in different regions of the deutocerebrum. This study adds to the understanding of neurogenesis in a brachyuran semiterrestrial crustacean and encourages comparative studies between crustaceans and vertebrates, including mammals, based on shared aspects of both mechanisms of neurogenesis and regenerative potentials.

Lipid Rafts from Olfactory Ensheathing Cells: Molecular Composition and Possible Roles.

Olfactory ensheathing cells (OECs) are specialized glial cells of the olfactory system, believed to play a role in the continuous production of olfactory neurons and ensheathment of their axons. Although OECs are used in therapeutic applications, little is known about the cellular mechanisms underlying their migratory behavior. Recently, we showed that OEC migration is sensitive to ganglioside blockage through A2B5 and Jones antibody in OEC culture. Gangliosides are common components of lipid rafts, where they participate in several cellular mechanisms, including cell migration. Here, we characterized OEC lipid rafts, analyzing the presence of specific proteins and gangliosides that are commonly expressed in motile neural cells, such as young neurons, oligodendrocyte progenitors, and glioma cells. Our results showed that lipid rafts isolated from OECs were enriched in cholesterol, sphingolipids, phosphatidylcholine, caveolin-1, flotillin-1, gangliosides GM1 and 9-O-acetyl GD3, A2B5-recognized gangliosides, CNPase, α-actinin, and ß1-integrin. Analysis of the actin cytoskeleton of OECs revealed stress fibers, membrane spikes, ruffled membranes and lamellipodia during cell migration, as well as the distribution of α-actinin in membrane projections. This is the first description of α-actinin and flotillin-1 in lipid rafts isolated from OECs and suggests that, together with ß1-integrin and gangliosides, membrane lipid rafts play a role during OEC migration. This study provides new information on the molecular composition of OEC membrane microdomains that can impact on our understanding of the role of OEC lipid rafts under physiological and pathological conditions of the nervous system, including inflammation, hypoxia, aging, neurodegenerative diseases, head trauma, brain tumor, and infection.

Progressive muscle-strength protocol for the functionality of upper limbs and quality of life in individuals with Parkinson's disease: Pilot study.

The effects of physical-therapy intervention on the motor function of upper limbs and the quality of life in patients with Parkinson's disease (PD) are not fully understood. We evaluated the effects of a progressive muscle-strengthening protocol for upper limbs on the functionality and quality of life. Patients were divided into two groups: Intervention (n = 6) and Control (n = 7). Assessment tools used were: Unified Parkinson's Disease Rating Scale (UPDRS), Parkinson's Disease Questionnaire, Nine-Hole Peg Test (9HPT), Test d'Évaluation des Membres Supérieurs de Personnes Âgées (TEMPA), 10-Repetition Maximum (10-RM) and handgrip dynamometer, which were applied pre- and post-intervention, with follow-up for one month after the last training session. Only, the Intervention group (post-intervention) showed significant statistical differences, with the following outcomes: UPDRS III (p = 0.042); 9HPT, right (p = 0.028) and left side (p = 0.028); TEMPA for total right side (p = 0.028), left side (p = 0.028) and total bilateral tasks (p = 0.028); TEMPA task 2 - open a jar and take a spoonful of coffee (p = 0.028), task 3 - pick up a pitcher and pour water into a glass for right (p = 0.046) and left side (p = 0.028), task 5 - write on an envelope and stick on a stamp (p = 0.028), and task 6 - shuffle and deal playing cards (p = 0.028). We observed significant statistical differences between groups (post-intervention) for TEMPA task 6 (p = 0.032), total right side (p = 0.032), and total bilateral tasks (p = 0.032). An increase in the maximum load in the post-intervention stage, based on the 10-RM test, was observed on the right (p = 0.003) and left (p = 0.007) sides. Our results showed an improvement in upper-limb functionality in PD patients submitted to progressive muscle-strength training, although not in quality of life.

Measurement properties from the Brazilian Portuguese version of the QUIP-RS.

Parkinson's disease (PD) has numerous motor and non-motor symptoms. Among non-motor manifestations impulse control disorders (ICDs) stand out. ICDs include compulsions for gambling, shopping, eating, and sexual behavior, and "related disorders" such as hobbyism, simple motor activities, and dopamine dysregulation syndrome. There is no rating scale translated and adapted transculturally into Brazilian Portuguese language. Therefore, we cross-culturally adapted and investigated the measurement properties of the Brazilian version of the Questionnaire for Impulsive-Compulsive Disorders in Parkinson's Disease-Rating Scale (QUIP-RS). Fifty-three patients participated in the study. Inter-evaluator and test-retest (patient and health professional) reliabilities (intraclass correlation coefficient) were all excellent (0.93, 0.93, and 0.99). The internal consistency was high (α = 0.92). The Minimal detectable change (MDC) value was 5.8 (patient) and 2.3 (health professional) points. There was a floor, but no ceiling, effect. In summary, the Brazilian version of the QUIP-RS has high reliability and content validity.

A Balancing Act: The Immune System Supports Neurodegeneration and Neurogenesis.

Decapod crustaceans, like mammals, retain the ability to make new neurons throughout life. In mammals, immune cells are closely associated with stem cells that generate adult-born neurons. In crayfish, evidence suggests that immune cells (hemocytes) originating in the immune system travel to neurogenic regions and transform into neural progenitor cells. This nontraditional immune activity takes place continuously under normal physiological conditions, but little is known under pathological conditions (neurodegeneration). In this study, the immune system and its relationship with neurogenesis were investigated during neurodegeneration (unilateral antennular ablation) in adult crayfish. Our experiments show that after ablation (1) Proliferating cells decrease in neurogenic areas of the adult crayfish brain; (2) The immune response, but not neurogenesis, is ablation-side dependent; (3) Inducible nitric oxide synthase (iNOS) plays a crucial role in the neurogenic niche containing neural progenitors during the immune response; (4) Brain areas targeted by antennular projections respond acutely (15 min) to the lesion, increasing the number of local immune cells; (5) Immune cells are recruited to the area surrounding the ipsilateral neurogenic niche; and (6) The vasculature in the niche responds acutely by dilation and possibly also neovascularization. We conclude that immune cells are important in both neurodegeneration and neurogenesis by contributing in physiological conditions to the maintenance of the number of neural precursor cells in the neurogenic niche (neurogenesis), and in pathological conditions (neurodegeneration) by coordinating NO release and vascular responses associated with the neurogenic niche. Our data suggest that neural damage and recovery participate in a balance between these competing immune cell roles.

Oxidative stress: Noradrenaline as an integrator of responses in the neuroendocrine and immune systems of the ascidian Phallusia nigra.

Neurotransmitters play key roles in regulating the homeostasis of organisms in stressful environments. Noradrenaline (NA) is the main neurotransmitter known to modulate immunological parameters, and is important in the crosstalk between the neuroendocrine and immune systems. In this study, using the ascidian Phallusia nigra, we analyzed the level of catecholamines (CA) in the plasma after mechanical stress, and the effect of NA on the oxidative stress (OS) displayed by immune cells. We measured the concentration of reactive oxygen species (ROS), and analyzed whether α- and/or ß-adrenoreceptors (ARs) are involved in ROS modulation, lipid peroxidation (LPO), antioxidant capacity against peroxyl radicals (ACAP), and activity of the enzymes catalase (CAT) and glutathione S transferase (GST) in immune cells after incubation with different concentrations of NA, with or without zymosan (ZnA) challenge. The results showed that NA reduced ROS production, even in immune cells challenged with ZnA, and that this modulation occurred through α1-and ß1-ARs. ACAP levels showed different responses, depending on whether immune cells were challenged or not with ZnA, and also depending on the NA concentration: 1.0 µM NA increased ACAP levels, but 10.0 µM reduced ACAP levels. NA enhanced the activity of CAT and GST in ZnA-challenged and non-challenged immune cells, while 1.0 and 10.0 µM NA effectively reduced LPO. Taken together, these results show that NA can protect cells from ROS damage, decreasing ROS production and LPO, and enhancing ACAP as well as the activity of CAT and GST. The approach used here with this model contributes to understanding the relationship between the neuroendocrine and immune systems, revealing new effects of NA on OS regulation in ascidians.

Putative stem cells in the hemolymph and in the intestinal submucosa of the solitary ascidian Styela plicata.

BACKGROUND: In various ascidian species, circulating stem cells have been documented to be involved in asexual reproduction and whole-body regeneration. Studies of these cell population(s) are mainly restricted to colonial species. Here, we investigate the occurrence of circulating stem cells in the solitary Styela plicata, a member of the Styelidae, a family with at least two independent origins of coloniality. RESULTS: Using flow cytometry, we characterized a population of circulating putative stem cells (CPSCs) in S. plicata and determined two gates likely enriched with CPSCs based on morphology and aldehyde dehydrogenase (ALDH) activity. We found an ALDH + cell population with low granularity, suggesting a stem-like state. In an attempt to uncover putative CPSCs niches in S. plicata, we performed a histological survey for hemoblast-like cells, followed by immunohistochemistry with stem cell and proliferation markers. The intestinal submucosa (IS) showed high cellular proliferation levels and high frequency of undifferentiated cells and histological and ultrastructural analyses revealed the presence of hemoblast aggregations in the IS suggesting a possible niche. Finally, we document the first ontogenetic appearance of distinct metamorphic circulatory mesenchyme cells, which precedes the emergence of juvenile hemocytes. CONCLUSIONS: We find CPSCs in the hemolymph of the solitary ascidian Styela plicata, presumably involved in the regenerative capacity of this species. The presence of proliferating and undifferentiated mesenchymal cells suggests IS as a possible niche.

A subacute model of glaucoma based on limbal plexus cautery in pigmented rats.

Glaucoma is a neurodegenerative disorder characterized by the progressive functional impairment and degeneration of the retinal ganglion cells (RGCs) and their axons, and is the leading cause of irreversible blindness worldwide. Current management of glaucoma is based on reduction of high intraocular pressure (IOP), one of its most consistent risk factors, but the disease proceeds in almost half of the patients despite such treatments. Several experimental models of glaucoma have been developed in rodents, most of which present shortcomings such as high surgical invasiveness, slow learning curves, damage to the transparency of the optic media which prevents adequate functional assessment, and variable results. Here we describe a novel and simple method to induce ocular hypertension in pigmented rats, based on low-temperature cauterization of the whole circumference of the limbal vascular plexus, a major component of aqueous humor drainage and easily accessible for surgical procedures. This simple, low-cost and efficient method produced a reproducible subacute ocular hypertension with full clinical recovery, followed by a steady loss of retinal ganglion cells and optic axons, accompanied by functional changes detected both by electrophysiological and behavioral methods.

A role for gangliosides and ß1-integrin in the motility of olfactory ensheathing glia.

Olfactory ensheathing glia (OEG) are found in the olfactory mucosa, nerve and bulb, and provide in vivo ensheathment for the unmyelinated olfactory axons within the central and peripheral nervous system domains. OEG cells are able to migrate long distances within the neuropil of the central nervous system. Because gangliosides such as 9-O-acetyl GD3 have crucial regulatory roles in neuronal migration during development, we analyzed whether OEG in organotypical cultures are revealed by anti-9-O-acetyl GD3 and/or gangliosides are recognized by the A2B5 antibody (G-A2B5), and whether these gangliosides are involved in OEG migration. Our results showed that all OEG migrating out of a section of olfactory bulb onto a laminin substrate bound to the 9-O-acetyl GD3 and A2B5 antibodies, and that 2',3'-cyclic nucleotide phosphodiesterase (CNPase) colocalized with 9-O-acetyl GD3 and with G-A2B5. Additionally, we showed that the immune blockade of 9-O-acetyl GD3 or G-A2B5 reduced the migration of OEG on laminin, and that 9-O-acetyl GD3 and G-A2B5 colocalized with the ß1-integrin subunit. We also confirmed the phenotype of in-vitro-grown OEG cells derived from adult rats, showing that they express CNPase, and also α-smooth muscle actin, which is not expressed by Schwann cells. Our data showed that the gangliosides 9-O-acetyl GD3 and G-A2B5 participate in the migratory activity of OEG cells, and that the ß1-integrin subunit colocalizes with these gangliosides. These results suggest a new role for ß1-integrin and gangliosides in the polarized migration of OEG cells, and provide new information on the molecules controlling OEG motility and behavior.

The Neurotropic Parasite Toxoplasma gondii Induces Sustained Neuroinflammation with Microvascular Dysfunction in Infected Mice.

Toxoplasmosis is one of the leading parasitic diseases worldwide. Some data suggest that chronic acquired toxoplasmosis could be linked to behavioral alterations in humans. The parasite infects neurons, forming immunologically silent cysts. Cerebral microcirculation homeostasis is determinant to brain functions, and pathologic states can alter capillarity or blood perfusion, leading to neurodegeneration and cognitive deficits. Albino mice were infected with Toxoplasma gondii (ME49 strain) and analyzed after 10, 40, and 180 days. Infected mice presented decreased cerebral blood flow at 10 and 40 days post infection (dpi), which were restored at 180 dpi, as shown by laser speckle contrast imaging. Intravital microscopy demonstrated that infection led to significant capillary rarefaction, accompanied by neuroinflammation, with microglial activation and increased numbers of rolling and adherent leukocytes to the wall of cerebral capillaries. Acetylcholine-induced vasodilation was altered at all time points, and blood brain barrier permeability was evident in infected animals at 40 dpi. Infection reduced angiogenesis, with a decreased number of isolectin B4-stained blood vessels and a decrease in length and branching of laminin-stained capillaries. Sulfadiazine reduced parasite load and partially repaired microvascular damages. We conclude that T. gondii latent infection causes a harmful insult in the brain, promoting neuroinflammation and microcirculatory dysfunction in the brain, with decreased angiogenesis and can contribute to a neurodegenerative process.

Acute exposure to water-soluble fractions of marine diesel oil: Evaluation of apoptosis and oxidative stress in an ascidian.

To understand the mechanisms involved in organisms' responses to toxicity from oil pollution, we studied the effect of acute exposure (24 h) to the marine water-soluble fraction of diesel oil (WFDO) on the ascidian Styela plicata. We evaluated the mortality and behavior by means of the siphon reflex, and the response of blood cells (hemocytes) contained in the pharynx, by means of the production of nitric oxide (NO) and reactive oxygen species (ROS), in addition to the activity of the antioxidant enzyme catalase (CAT). We also correlated oxidative stress with the activation of apoptotic pathways. No mortality occurred 24 h after the ascidians were exposed to 5% and 10% marine WFDO; however, the siphon reflex, a behavioral test based on the time that the animals took to close their siphons, increased. We also observed an inflammatory response, as estimated by the increase in the number of hemocytes in the pharynx. NO and ROS production and CAT activity were reduced, whereas caspase-3, a signaling molecule involved in apoptosis, was activated. This suggests that in ascidians acutely exposed to oil, another mechanism can occur in addition to oxidative stress. Another possibility is that WFDO may directly interact with cellular macromolecules and activate caspase-3, independently of generating oxidative stress. The results showed that components of diesel oil affected a marine organism, which showed reduced ROS production in the pharynx cells, including hemocytes, and activation of apoptotic pathways.