Alexandre Valotta da Silva

Differential effects of spontaneous versus forced exercise in rats on the staining of parvalbumin-positive neurons in the hippocampal formation

To investigate whether the type of physical activity, voluntary or forced, would promote different morphological changes in hippocampal formation we performed an immunocytochemical study using the parvalbumin (PV) distribution as a marker. Animals submitted to the voluntary exercise were placed in a voluntary wheel running and those submitted to the forced exercise were placed in a motor driven treadmill. Both exercise groups were submitted to 10 consecutive days of physical activity. It was observed a higher number of PV-positive cells in the hilus of dentate gyrus (DG) in the voluntary and forced exercise groups when compared to the control group. The voluntary exercise group also presented a stronger fiber staining in the DG hilus than the forced exercise and control groups. In contrast, no differences were found in the pattern of PV staining in CA1/CA3 region between exercise and control groups. These findings demonstrate that physical activity leads to prominent plastic changes in the hippocampal formation of rats that were more evident following voluntary activity.

Whole transcriptome analysis of the hippocampus: toward a molecular portrait of epileptogenesis

BackgroundUncovering the molecular mechanisms involved in epileptogenesis is critical to better understand the physiopathology of epilepsies and to help develop new therapeutic strategies for this prevalent and severe neurological condition that affects millions of people worldwide.ResultsChanges in the transcriptome of hippocampal cells from rats subjected to the pilocarpine model of epilepsy were evaluated by microarrays covering 34,000 transcripts representing all annotated rat genes to date. Using such genome-wide approach, differential expression of nearly 1,400 genes was detected during the course of epileptogenesis, from the early events post status epilepticus (SE) to the onset of recurrent spontaneous seizures. Most of these genes are novel and displayed an up-regulation after SE. Noteworthy, a group of 128 genes was found consistently hyper-expressed throughout epileptogenesis, indicating stable modulation of the p38MAPK, Jak-STAT, PI3K, and mTOR signaling pathways. In particular, up-regulation of genes from the TGF-beta and IGF-1 signaling pathways, with opposite effects on neurogenesis, correlate with the physiopathological changes reported in humans.ConclusionsA consistent regulation of genes functioning in intracellular signal transduction regulating neurogenesis have been identified during epileptogenesis, some of which with parallel expression patterns reported in patients with epilepsy, strengthening the link between these processes and development of epilepsy. These findings reveal dynamic molecular changes occurring in the hippocampus that may serve as a starting point for designing alternative therapeutic strategies to prevent the development of epilepsy after acquired brain insults.

Updated Neuronal Scaling Rules for the Brains of Glires (Rodents/Lagomorphs)

Brain size scales as different functions of its number of neurons across mammalian orders such as rodents, primates, and insectivores. In rodents, we have previously shown that, across a sample of 6 species, from mouse to capybara, the cerebral cortex, cerebellum and the remaining brain structures increase in size faster than they gain neurons, with an accompanying decrease in neuronal density in these structures [Herculano-Houzel et al.: Proc Natl Acad Sci USA 2006;103:12138–12143]. Important remaining questions are whether such neuronal scaling rules within an order apply equally to all pertaining species, and whether they extend to closely related taxa. Here, we examine whether 4 other species of Rodentia, as well as the closely related rabbit (Lagomorpha), conform to the scaling rules identified previously for rodents. We report the updated neuronal scaling rules obtained for the average values of each species in a way that is directly comparable to the scaling rules that apply to primates [Gabi et al.: Brain Behav Evol 2010;76:32–44], and examine whether the scaling relationships are affected when phylogenetic relatedness in the dataset is accounted for. We have found that the brains of the spiny rat, squirrel, prairie dog and rabbit conform to the neuronal scaling rules that apply to the previous sample of rodents. The conformity to the previous rules of the new set of species, which includes the rabbit, suggests that the cellular scaling rules we have identified apply to rodents in general, and probably to Glires as a whole (rodents/lagomorphs), with one notable exception: the naked mole-rat brain is apparently an outlier, with only about half of the neurons expected from its brain size in its cerebral cortex and cerebellum.

Damage, Reorganization, and Abnormal Neocortical Hyperexcitability in the Pilocarpine Model of Temporal Lobe Epilepsy

Summary:  Purpose: Clinical, neuropathological, and electrophysiological data have shown that limbic structures are involved in the pathogenesis of temporal lobe epilepsy (TLE). In most cases, limbic‐originated seizures frequently spread to extrahippocampal areas. It is unclear whether such distant circuitries, especially the neocortex, exhibit abnormal electrophysiology as consequences of a chronic epileptogenic process. The present research studied neuropathological abnormalities and in vitro electrophysiological properties of sensorimotor neocortex in pilocarpine‐treated epileptic rats.

Alterations of the neocortical GABAergic system in the pilocarpine model of temporal lobe epilepsy: neuronal damage and immunocytochemical changes in chronic epileptic rats.

A wealth of previous studies reported pathological alterations in extrahippocampal regions in mesial temporal lobe epilepsy. Previous experimental findings have also demonstrated that the entorhinal cortex and the neocortex are damaged in different animal models of acute limbic seizures. The present study was aimed at verifying possible alterations in neocortical areas, and, in particular, structural changes of GABAergic interneurons in the sensorimotor cortex, in pilocarpine-induced chronic epilepsy in the rat. Series of sections were Nissl stained and processed for immunocytochemistry using antibodies that recognize nonphosphorylated neurofilament (SMI311), glial fibrillary acidic protein (GFAP), the calcium-binding protein parvalbumin (PV) which is expressed by a subset of cortical GABAergic neurons, the GABA transporter (GAT1), and isoform 65 of glutamic acid decarboxylase (GAD65), the GABA synthetic enzyme. Epileptic rats showed decreased cortical thickness, and diffuse gliosis was observed with GFAP antibody. Neurofilament alterations were also detected in sections processed using SMI311 antiserum. In addition, a diffuse decrease of PV, GAD65, and GAT1 immunoreactivity was observed in the sensorimotor cortex. Altered expression of PV, GAD65, and GAT1 pointed out specific neocortical disturbances in GABAergic inhibition, which could play a crucial role in seizure generation and expression. Thus, the present findings indicate that damage of GABAergic interneurons could be strictly associated with neocortical hyperexcitability in temporal lobe epilepsy.

Neuroprotective effect of pyruvate and oxaloacetate during pilocarpine induced status epilepticus in rats

Recent research data have shown that systemic administration of pyruvate and oxaloacetate causes an increased brain-to-blood glutamate efflux. Since increased release of glutamate during epileptic seizures can lead to excitotoxicity and neuronal cell death, we tested the hypothesis that glutamate scavenging mediated by pyruvate and oxaloacetate systemic administration could have a neuroprotective effect in rats subjected to status epilepticus (SE). SE was induced by a single dose of pilocarpine (350mg/kgi.p.). Thirty minutes after SE onset, a single dose of pyruvate (250mg/kgi.p.), oxaloacetate (1.4mg/kgi.p.), or both substances was administrated. Acute neuronal loss in hippocampal regions CA1 and hilus was quantitatively determined five hours after SE onset, using the optical fractionator method for stereological cell counting. Apoptotic cascade in the hippocampus was also investigated seven days after SE using caspase-1 and -3 activity assays. SE-induced neuronal loss in CA1 was completely prevented in rats treated with pyruvate plus oxaloacetate. The SE-induced caspase-1 activation was significantly reduced when rats were treated with oxaloacetate or pyruvate plus oxaloacetate. The treatment with pyruvate and oxaloacetate caused a neuroprotective effect in rats subjected to pilocarpine-induced SE.

Delayed physical and neurobehavioral development and increased aggressive and depression-like behaviors in the rat offspring of dams fed a high-fat diet.

Early maternal exposure to a high‐fat diet (HFD) may influence the brain development of rat offspring and consequently affect physiology and behavior. Thus, in the present study, we investigated the somatic, physical, sensory‐motor and neurobehavioral development of the offspring of dams fed an HFD (52% calories from fat, mainly saturated) and the offspring of dams fed a control diet (CD – 14.7% fat) during lactation from the 1st to the 21st postnatal day (P). Maternal body weights were evaluated during lactation. In the progeny, somatic (body weight, head and lengths axes) and physical (ear unfolding, auditory conduit opening, eruption of the incisors and eye opening) development and the consolidation of reflex responses (palm grasp, righting, vibrissa placing, cliff avoidance, negative geotaxis, auditory startle response and free‐fall righting) were determined during suckling. Depressive and aggressive behaviors were tested with the forced swimming test (FST) and the “foot‐shock” test on days 60 and 110, respectively. The open field test was used to assess motor function. Compared to controls, the HFD‐pups exhibited decreases in body weight (P7–P21) and body length (P4–P18), but by days P71 and P95, these pups were overweight. All indicators of physical maturation and the consolidation of the following reflexes, vibrissa placing, auditory startle responses, free‐fall righting and negative geotaxis, were delayed in HFD‐progeny. In addition, the pups from HFD dam rats also exhibited reduced swimming and climbing times in the FST and increased aggressive behavior. No changes in locomotion were observed. These findings show developmental and neurobehavioral changes in the rat offspring of dams fed the HFD during lactation and suggest possible disruption of physical and sensory‐motor maturation and increased susceptibility to depressive and aggressive‐like behavior.

Disruption of cortical development as a consequence of repetitive pilocarpine-induced status epilepticus in rats.

Summary:  Purpose: The aim of the present study was to observe possible cortical abnormalities after repetitive pilocarpine‐induced status epilepticus (SE) in rats during development.

Neocortical and Hippocampal Changes after Multiple Pilocarpine‐induced Status Epilepticus in Rats

Summary:  Purpose: Multiple episodes of pilocarpine‐induced status epilepticus (SE) in developing rats (P7–P9) lead to progressive epileptiform activity and severe cognitive impairment in adulthood. The present work studied possible underlying abnormalities in the neocortex and hippocampus of pilocarpine‐treated animals.

Improved detection of incipient vascular changes by a biotechnological platform combining post mortem MRI in situ with neuropathology

The histopathological counterpart of white matter hyperintensities is a matter of debate. Methodological and ethical limitations have prevented this question to be elucidated. We want to introduce a protocol applying state-of-the-art methods in order to solve fundamental questions regarding the neuroimaging-neuropathological uncertainties comprising the most common white matter hyperintensities [WMHs] seen in aging. By this protocol, the correlation between signal features in in situ, post mortem MRI-derived methods, including DTI and MTR and quantitative and qualitative histopathology can be investigated. We are mainly interested in determining the precise neuroanatomical substrate of incipient WMHs. A major issue in this protocol is the exact co-registration of small lesion in a tridimensional coordinate system that compensates tissue deformations after histological processing. The protocol is based on four principles: post mortem MRI in situ performed in a short post mortem interval, minimal brain deformation during processing, thick serial histological sections and computer-assisted 3D reconstruction of the histological sections. This protocol will greatly facilitate a systematic study of the location, pathogenesis, clinical impact, prognosis and prevention of WMHs.