Postweaning Enriched Environment Enhances Cognitive Function and Brain-Derived Neurotrophic Factor Signaling in the Hippocampus in Maternally Separated Rats.

Adverse environments during early life may lead to different neurophysiological and behavioral consequences, including depression and learning and memory deficits that persist into adulthood. Previously, we demonstrated that exposure to an enriched environment during adolescence mitigates the cognitive impairment observed after maternal separation in a task-specific manner. However, underlying neural mechanisms are still not fully understood. The current study examines the effects of neonatal maternal separation (MS) and postweaning environmental enrichment (EE) on spatial learning and memory performance in a short version of the Barnes Maze, active and passive behaviors in the forced swim test, and on TrkB/BDNF receptor expression in the hippocampus. Our results revealed that MS impaired acquisition learning and that enriched rats performed better than non-enriched rats in acquisition trials, regardless of early conditions. During the probe, enriched-housed rats demonstrated better performance than those reared in standard conditions. No significant differences between groups were found in the forced swim test. Both MS and EE increase full-length TrkB expression, and the combination of MS and EE treatment caused the highest levels of this protein expression. Similarly, truncated TrkB expression was higher in the MS/EE group. Animal facility rearing (AFR) non-enriched groups present the lowest activation of phosphorylated Erk, a canonical downstream kinase of TrkB signaling. Taken together, our results demonstrate the importance of enriched environment as an intervention to ameliorate the effects of maternal separation on spatial learning and memory. TrkB/BDNF signaling could mediate neuroplastic changes related to learning and memory during exposure to enriched environment.

Understanding new molecular and cell biology findings based on progressive scientific practices and interconnected activities in undergraduate students.

Nowadays Molecular Cell Biology (MCB) must be taught as science is practiced. Even though there are several approaches based on scientific practices, a key aspect is to define the purpose of each of these teaching strategies and, most importantly, their implementation. Our goal was to train students to acquire, understand, and communicate new scientific knowledge in the field. The main feature of our new teaching methodology was progressive training in scientific practices associated with a back-and-forward interplay between activities and assessments. The methodology was implemented over 4 years, in students attending the MCB course of the undergraduate degree in Biological Sciences. In the first two modules, the students were prepared to comprehend MCB concepts and techniques and to experience activities based on scientific practices. In the third module, the students analyzed a primary paper in-depth. They were assessed by midterm exams based on a primary paper, written laboratory reports, and the oral presentation of a scientific paper. Our teaching proposal was evaluated through the students' academic performance and by their opinion on the teaching methodology. Most students were satisfied since they improved their acquisition of concepts, their interpretation and integration of scientific knowledge, and developed skills to communicate scientific knowledge in writing and orally. The novelty of transversal interconnections and progressive training in scientific practices provides students with skills in acquiring and understanding new scientific information, even beyond the MCB course.

Loss of TrkB Signaling Due to Status Epilepticus Induces a proBDNF-Dependent Cell Death.

Neurotrophins (NTs) are secretory proteins that bind to target receptors and influence many cellular functions, such as cell survival and cell death in neurons. The mammalian NT brain-derived neurotrophic factor (matBDNF) is the C-terminal mature form released by cleavage from the proBDNF precursor. The binding of matBDNF to the tyrosine kinase receptor B (TrkB) activates different signaling cascades and leads to neuron survival and plasticity, while the interaction of proBDNF with the p75 NT receptor (p75NTR)/sortilin receptor complex has been highly involved in apoptosis. Many studies have demonstrated that prolonged seizures such as status epilepticus (SE) induce changes in the expression of NT, pro-NT, and their receptors. We have previously described that the blockage of both matBDNF and proBDNF signaling reduces neuronal death after SE in vivo (Unsain et al., 2008). We used an in vitro model as well as an in vivo model of SE to determine the specific role of TrkB and proBDNF signaling during neuronal cell death. We found that the matBDNF sequestering molecule TrkB-Fc induced an increase in neuronal death in both models of SE, and it also prevented a decrease in TrkB levels. Moreover, SE triggered the interaction between proBDNF and p75NTR, which was not altered by sequestering matBDNF. The intra-hippocampal administration of TrkB-Fc, combined with an antibody against proBDNF, prevented neuronal degeneration. In addition, we demonstrated that proBDNF binding to p75NTR exacerbates neuronal death when matBDNF signaling is impaired through TrkB. Our results indicated that both the mature and the precursor forms of BDNF may have opposite effects depending on the scenario in which they function and the signaling pathways they activate.

Cdk5 Is Essential for Amphetamine to Increase Dendritic Spine Density in Hippocampal Pyramidal Neurons.

Psychostimulant drugs of abuse increase dendritic spine density in reward centers of the brain. However, little is known about their effects in the hippocampus, where activity-dependent changes in the density of dendritic spine are associated with learning and memory. Recent reports suggest that Cdk5 plays an important role in drug addiction, but its role in psychostimulant's effects on dendritic spines in hippocampus remain unknown. We used in vivo and in vitro approaches to demonstrate that amphetamine increases dendritic spine density in pyramidal neurons of the hippocampus. Primary cultures and organotypic slice cultures were used for cellular, molecular, pharmacological and biochemical analyses of the role of Cdk5/p25 in amphetamine-induced dendritic spine formation. Amphetamine (two-injection protocol) increased dendritic spine density in hippocampal neurons of thy1-green fluorescent protein (GFP) mice, as well as in hippocampal cultured neurons and organotypic slice cultures. Either genetic or pharmacological inhibition of Cdk5 activity prevented the amphetamine-induced increase in dendritic spine density. Amphetamine also increased spine density in neurons overexpressing the strong Cdk5 activator p25. Finally, inhibition of calpain, the protease necessary for the conversion of p35 to p25, prevented amphetamine's effect on dendritic spine density. We demonstrate, for the first time, that amphetamine increases the density of dendritic spine in hippocampal pyramidal neurons in vivo and in vitro. Moreover, we show that the Cdk5/p25 signaling and calpain activity are both necessary for the effect of amphetamine on dendritic spine density. The identification of molecular mechanisms underlying psychostimulant effects provides novel and promising therapeutic approaches for the treatment of drug addiction.

An alkaloid extract obtained from Phlegmariurus Saururus induces neuroprotection after status epilepticus.

BACKGROUND: The brain is exposed to many excitotoxic insults that can lead to neuronal damage. Among these, Epilepsy is a neurological disease that affects a large percentage of world population and is commonly associated with cognitive disorders and excitotoxic neuronal death. Most experimental strategies are focused on preventing Status Epilepticus (SE), but once it has already occurred, the key question is whether it is possible to save neurons. PURPOSE: The aim of this study was to determine if a purified alkaloid extract (AE) obtained from Phlegmariurus saururus, a genus of Lycophyte plants (sometimes known as firmossesor fir club mosses) could induce neuroprotection following SE. METHODS: In vitro and in vivo techniques were applied for this purpose. Protein levels were measured by western blotting procedures. Neuronal death analysis was performed by calcein-ethidium staining and the presence of the NeuN protein as a marker for presence or absence of cells (in vitro experiments) and by Fluoro Jade B staining for the in vivo experiments. RESULTS: The effect of AE in the hippocampal neurons culture was the first determination, where we found an increase in neuronal survival and in the level of pErk and TrkB activation, 24 h after the addition of AE. In a well-established in vitro model of SE, we found that 24 h after being added to the hippocampal neuron-astrocyte co-culture, the AE induces a significant increase in neuronal survival. In addition to this, in the in vivo Li-pilocarpine model of SE, the AE induced a remarkable neuroprotection in areas such as the entorhinal cortex and hippocampal CA1 area. CONCLUSION: These results make the AE an excellent candidate for potential clinical use in neurological disorders where memory impairment and neuronal death occurs.

Calpain-dependent truncated form of TrkB-FL increases in neurodegenerative processes.

Recent findings indicate that the mechanisms that drive reshaping of the nervous system are aberrantly activated in epilepsy and several neurodegenerative diseases. The recurrent seizures in epilepsy, particularly in the condition called status epilepticus, can cause permanent neurological damage, resulting in cognitive dysfunction and other serious neurological conditions. In this study, we used an in vitro model of status epilepticus to examine the role of calpain in the degeneration of hippocampal neurons. We grew neurons on a culture system that allowed studying the dendritic and axonal domains separately from the cell bodies. We found that a recently characterized calpain substrate, the neurotrophin receptor TrkB, is cleaved in the dendritic and axonal domain of neurons committed to die, and this constitutes an early step in the neuronal degeneration process. While the full-length TrkB (TrkB-FL) levels decreased, the truncated form of TrkB (Tc TrkB-FL) concurrently increased, leading to a TrkB-FL/Tc TrkB-FL imbalance, which is thought to be causally linked to neurodegeneration. We further show that the treatment with N-acetyl-Leu-Leu-norleucinal, a specific calpain activity blocker, fully protects the neuronal processes from degeneration, prevents the TrkB-FL/Tc TrkB-FL imbalance, and provides full neuroprotection. Moreover, the use of the TrkB antagonist ANA 12 at the time when the levels of TrkB-FL were significantly decreased, totally blocked neuronal death, suggesting that Tc TrkB-FL may have a role in neuronal death. These results indicate that the imbalance of these neurotrophins receptors plays a key role in neurite degeneration induced by seizures.