Interactions between mitochondria and the transcription factor myocyte enhancer factor 2 (MEF2) regulate neuronal structural and functional plasticity and metaplasticity.

The classical view of mitochondria as housekeeping organelles acting in the background to simply maintain cellular energy demands has been challenged by mounting evidence of their direct and active participation in synaptic plasticity in neurons. Time-lapse imaging has revealed that mitochondria are motile in dendrites, with their localization and fusion and fission events regulated by synaptic activity. The positioning of mitochondria directly influences function of nearby synapses through multiple pathways including control over local concentrations of ATP, Ca(2+) and reactive oxygen species. Recent studies have also shown that mitochondrial protein cascades, classically associated with apoptosis, are involved in neural plasticity in healthy cells. These findings link mitochondria to the plasticity- and metaplasticity-associated activity-dependent transcription factor myocyte enhancer factor 2 (MEF2), further repositioning mitochondria as potential command centres for regulation of synaptic plasticity. Intriguingly, MEF2 and mitochondrial functions appear to be intricately intertwined, as MEF2 is a target of mitochondrial apoptotic caspases and, in turn, MEF2 regulates mitochondrial genome transcription essential for production of superoxidase and hydrogen peroxidase. Here, we review evidence supporting mitochondria as central organelles controlling the spatiotemporal expression of neuronal plasticity, and attempt to disentangle the MEF2-mitochondria relationship mediating these functions.

Lipofuscin Granules in the Epileptic Human Temporal Neocortex with Age.

Lipofuscin granules (LGs), the "age pigments", are autofluorescent cell products from lysosomes that diverge in number and size among brain regions. Human temporal cortex from 20- to 55-year-old epileptic subjects were studied with the fat soluble dye Sudan Black, under confocal and electron microscopy. Ultrastructural analysis showed that with age LGs increase in area, but not in number. Proportionally to the LGs area, the electron lucid portion increases and the electron dense reduces over time. The robust increase in lipid components is possibly due to modifications in the neuronal metabolism with age in physiological and pathological conditions.

Selectively RNA interaction by a hnRNPA/B-like protein at presynaptic terminal of squid neuron.

In previous works, we identified a RNA-binding protein in presynaptic terminal of squid neurons, which is likely involved in local mRNA processing. Evidences indicate this strongly basic protein, called p65, is an SDS-stable dimer protein composed of ~ 37 kDa hnRNPA/B-like subunits. The function of p65 in presynaptic regions is not well understood. In this work, we showed p65 and its subunit p37 are concentrated in RNA-enriched regions in synaptosomes. We performed in vitro binding studies with a recombinant protein and showed its propensity to selectively bind actin mRNA at the squid presynaptic terminal. Biochemical analysis using lysed synaptosomes suggested RNA integrity may affect p65 and p37 functions. Mass spectrometry analysis of oligo(dT) pull down indicated squid hnRNPA1, hnRNPA1-like 2, hnRNPA3 and ELAV-like proteins as candidates to interact with p65 and p37 forming a ribonucleoprotein complex, suggesting a role of squid hnRNPA/B-like proteins in site-specific RNA processing.

Plasma hormonal profiles and dendritic spine density and morphology in the hippocampal CA1 stratum radiatum, evidenced by light microscopy, of virgin and postpartum female rats.

Successful reproduction requires that changes in plasma follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), oxytocin (OT), estrogen (E(2)) and progesterone (P(4)) occur together with the display of maternal behaviors. Ovarian steroids and environmental stimuli can affect the dendritic spines in the rat hippocampus. Here, studying Wistar rats, it is described: (a) the sequential and concomitant changes in the hormonal profile of females at postpartum days (PP) 4, 8, 12, 16, 20 and 24, comparing to estrous cycle referential values; (b) the dendritic spine density in the stratum radiatum of CA1 (CA1-SR) Golgi-impregnated neurons in virgin females across the estrous cycle and in multiparous age-matched ones; and (c) the proportion of different types of spines in the CA1-SR of virgin and postpartum females, both in diestrus. Plasma levels of gonadotrophins and ovarian hormones remained low along PP while LH increased and PRL decreased near the end of the lactating period. The lowest dendritic spine density was found in virgin females in estrus when compared to diestrus and proestrus phases or to postpartum females in diestrus (p<0.03). Other comparisons among groups were not statistically significant (p>0.4). There were no differences in the proportions of the different spine types in nulliparous and postpartum females (p>0.2). Results suggest that medium layer CA1-SR spines undergo rapid modifications in Wistar females across the estrous cycle (not quite comparable to Sprague-Dawley data or to hormonal substitutive therapy following ovariectomy), but persistent effects of motherhood on dendritic spine density and morphology were not found in this area.

Sex Differences and Estrous Cycle Changes in Synaptic Plasticity-related microRNA in the Rat Medial Amygdala.

The posterodorsal medial amygdala (MePD) is a sex steroid-sensitive and sexually dimorphic subcortical area that dynamically modulates social behaviors in rats. As different microRNA (miRNA) can act as post-transcriptional regulators of synaptic processing, we addressed changes that occur in miRNA expression in the MePD of males and females along the estrous cycle. The expression of miR25-3p, miR132-3p, miR138-5p, miR181a-5p, miR195-5p, and miR199a-5p, involved in neuronal cytoskeleton remodeling and synaptic plasticity, were evaluated by RT-qPCR. We found that the expression of miR138-5p was higher in males than in females along the different phases of the estrous cycle. Males also showed higher levels of miR-181a when compared to females in diestrus and estrus. On the other hand, when compared to females in proestrus, males presented lower levels of miR132-3p and miR199a-5p. The expression of miR25-3p was higher in diestrus females than in proestrus females. In addition, diestrus females showed higher values of miR25-3p, miR181a-5p, and miR195-5p when compared to estrus females. These miRNA expression profiles indicate a variable and fine-tuned protein regulation in the adult MePD. It is likely that these miRNA can be involved in structural and functional synaptic features and plasticity characteristic of males and cycling females and for the MePD regulation of mammalian reproduction.

Distribution of glutamate receptors in the posterodorsal medial amygdala of adult male rats.

The rat posterodorsal medial amygdala (MePD) has a remarkable neuronal plasticity and responds to olfactory/pheromonal stimuli to modulate emotional and reproductive behaviors. Glutamate is locally released by incoming sensorial pathways to establish and enforce synaptic inputs. Here, we combined DiI dye and immunolabeling procedure under confocal microscopy to describe the presence and distribution of glutamate receptors on neurons of the MePD of adult male rats. Western blot analysis interrogated binding specificity. Both AMPA (GluA1-4 subunits) and NMDA (GluN1 subunit) receptors were immunolabeled on cell bodies and along proximal and distal dendritic shafts. AMPA receptors were mainly observed on mushroom and stubby/wide spines, whereas NMDA receptors were found on thin spines. Colocalization of AMPA and NMDA receptors occurred in some spines. Filopodium did not show immunolabeled puncta on it. Our results are different from the distribution of glutamate receptors in the amygdaloid lateral nucleus, an upstream area involved with emotional processing, and suggest a region-specific excitatory transmission at proximal and distal dendritic branches. Altogether, these data provide new information for synaptic processing in the MePD likely related to the modulation of social behavior in rats.

Inhibitory and multisynaptic spines, and hemispherical synaptic specialization in the posterodorsal medial amygdala of male and female rats.

The density of dendritic spines is sexually dimorphic and variable throughout the female estrous cycle in the rat posterodorsal medial amygdala (MePD), a relevant area for the modulation of reproductive behavior in rats. The local synaptic activity differs between hemispheres in prepubertal animals. Here we used serial section transmission electron microscopy to produce 3D reconstructions of dendritic shafts and spines to characterize synaptic contacts on MePD neurons of both hemispheres in adult males and in females along the estrous cycle. Pleomorphic spines and nonsynaptic filopodia occur in the MePD. On average, 8.6% of dendritic spines received inputs from symmetric gamma-aminobutyric acid (GABA)-immunoreactive terminals, whereas 3.6% received two synaptic contacts on the spine head, neck, or base. Presynaptic terminals in female right MePD had a higher density of synaptic vesicles and docked vesicles than the left MePD, suggesting a higher rate of synaptic vesicle release in the right MePD of female rats. In contrast, males did not show laterality in any of those parameters. The proportion of putative inhibitory synapses on dendritic shafts in the right MePD of females in proestrus was higher than in the left MePD, and higher than in the right MePD in males, or in females in diestrus or estrus. This work shows synaptic laterality depending on sex and estrous cycle phase in mature MePD neurons. Most likely, sexual hormone effects are lateralized in this brain region, leading to higher synaptic activity in the right than in the left hemisphere of females, mediating timely neuroendocrine and social/reproductive behavior.

Dendritic spines of the medial amygdala: plasticity, density, shape, and subcellular modulation by sex steroids.

The medial nucleus of the amygdala (MeA) is a complex component of the "extended amygdala" in rats. Its posterodorsal subnucleus (MePD) has a remarkable expression of gonadal hormone receptors, is sexually dimorphic or affected by sex steroids, and modulates various social behaviors. Dendritic spines show remarkable changes relevant for synaptic strength and plasticity. Adult males have more spines than females, the density of dendritic spines changes in the course of hours to a few days and is lower in proestrous and estrous phases of the ovarian cycle, or is affected by both sex steroid withdrawal and hormonal replacement therapy in the MePD. Males also have more thin spines than mushroom-like or stubby/wide ones. The presence of dendritic fillopodia and axonal protrusions in the MePD neuropil of adult animals reinforces the evidence for local plasticity. Estrogen affects synaptic and cellular growth and neuroprotection in the MeA by regulating the activity of the cyclic AMP response element-binding protein (CREB)-related gene products, brain-derived neurotrophic factor (BDNF), the anti-apoptotic protein B-cell lymphoma-2 (Bcl-2) and the activity-regulated cytoskeleton-related protein (Arc). These effects on signal transduction cascades can also lead to local protein synthesis and/or rearrangement of the cytoskeleton and subsequent numerical/morphological alterations in dendritic spines. Various working hypotheses are raised from these experimental data and reveal the MePD as a relevant region to study the effects of sex steroids in the rat brain.

Blocking Effects of Human Tau on Squid Giant Synapse Transmission and Its Prevention by T-817 MA.

Filamentous tau inclusions are hallmarks of Alzheimer's disease and related neurodegenerative tauopathies, but the molecular mechanisms involved in tau-mediated changes in neuronal function and their possible effects on synaptic transmission are unknown. We have evaluated the effects of human tau protein injected directly into the presynaptic terminal axon of the squid giant synapse, which affords functional, structural, and biochemical analysis of its action on the synaptic release process. Indeed, we have found that at physiological concentration recombinant human tau (h-tau42) becomes phosphorylated, produces a rapid synaptic transmission block, and induces the formation of clusters of aggregated synaptic vesicles in the vicinity of the active zone. Presynaptic voltage clamp recordings demonstrate that h-tau42 does not modify the presynaptic calcium current amplitude or kinetics. Analysis of synaptic noise at the post-synaptic axon following presynaptic h-tau42 microinjection revealed an initial phase of increase spontaneous transmitter release followed by a marked reduction in noise. Finally, systemic administration of T-817MA, a proposed neuro-protective agent, rescued tau-induced synaptic abnormalities. Our results show novel mechanisms of h-tau42 mediated synaptic transmission failure and identify a potential therapeutic agent to treat tau-related neurotoxicity.

Descriptive findings on the morphology of dendritic spines in the rat medial amygdala.

The rat posterodorsal medial amygdala (MePD) is a brain area in which gonadal hormones induce notable plastic effects in the density of dendritic spines. Dendritic spines are post-synaptic specializations whose shape and spacing change neuronal excitability. Our aim was to obtain new data on the dendritic spines morphology and density from MePD neurons using the carbocyanine dye DiI under confocal microscopy. In adult male rats, the dendritic spine density of the medial branches of the left MePD (mean+/-SD) was 1.15+/-0.67spines/dendritic microm. From the total sampled, approximately 53% of the spines were classified as thin, 22.5% as "mushroom-like", and 21.5% as stubby/wide. Other spine shapes (3%) included those ramified, with a filopodium-like or a gemule appearance, and others with a protruding spinule. Additional experiment joining DiI and synaptophysin (a pre-synaptic protein) labeling suggested synaptic sites on dendritic shafts and spines. Dendritic spines showed synaptophysin puncta close to their head and neck, although some spines had no evident labeled puncta on them or, conversely, multiple puncta appeared upon one spine. These results advance previous light microscopy results by revealing features and complexities of the dendritic spines at the same time that give new insight on the possible synaptic organization of the adult rat MePD.

The "single-section" Golgi method adapted for formalin-fixed human brain and light microscopy.

The Golgi method has been used for over a century to describe the general morphology of neurons in the nervous system of different species. The "single-section" Golgi method of Gabbott and Somogyi (1984) and the modifications made by Izzo et al. (1987) are able to produce consistent results. Here, we describe procedures to show cortical and subcortical neurons of human brains immersed in formalin for months or even years. The tissue was sliced with a vibratome, post-fixed in a combination of paraformaldehyde and picric acid in phosphate buffer, followed by osmium tetroxide and potassium dicromate, "sandwiched" between cover slips, and immersed in silver nitrate. The whole procedure takes between 5 and 11 days to achieve good results. The Golgi method has its characteristic pitfalls but, with this procedure, neurons and glia appear well-impregnated, allowing qualitative and quantitative studies under light microscopy. This contribution adds to the basic techniques for the study of human nervous tissue with the same advantages described for the "single-section" Golgi method in other species; it is easy and fast, requires minimal equipment, and provides consistent results.

O ciclo da vesícula sináptica, espinhos dendríticos e a transdução de sinal / Synaptic vesicle cycle, dendritic spines and signal transduction

No sistema nervoso, a sinapse é a estrutura que permite a um neurônio passar um sinal elétrico ou químico a outro neurônio ou outra célula (muscular ou glandular). A palavra sinapse vem de "synaptein", palavra que Sir Charles Scott Sherrington e seus colegas acunharam do grego "syn" (junto) e "haptein"(afivelar). As sinapses podem ser separadas entre elétricas e químicas, porém a maior parte da transmissão sináptica é realizada através das sinapses químicas. Apesar das sinapses químicas terem uma resposta mais lenta que as elétricas, elas possuem a vantagem da amplificação do sinal gerada através de uma cascata de segundos mensageiros. As sinapses químicas podem ser excitatórias ou inibitórias e são caracterizadas por um terminal pré-sináptico (onde estão presentes as vesículas que contêm os neurotransmissores) em contato com um terminal pós-sináptico (onde estão presentes os receptores ionotrópicos e metabotrópicos para esses neurotransmissores) separados pela fenda sináptica. As sinapses típicas acontecem sobre axônios (axo-axônicas), sobre dendritos (axo-dendríticas), sobre o soma de outro neurônio (axo-somáticas) e sobre os espinhos dendríticos...

In the nervous system, the synapse is the structure that allows a neuron pass an electrical or chemical signal to another neuron or another cell (muscle or glandular). The word synapse comes from "synaptein" that Sir Charles Scott Sherrington and his colleagues minted from the Greek "syn" (together) and "haptein"(buckling). Most part of the synaptic transmission is performed through chemical synapses. Chemical synapses have a slower response than the electric ones; they have the advantage of amplifying the signal generated through a cascade of second messengers. Chemical synapses can be excitatory or inhibitory and are characterized by a presynaptic terminal (where there are vesicles that contain the neurotransmitters) in contact with a postsynaptic terminal (where there are the ionotropic and metabotropic receptors) separated by the synaptic cleft. Synapses can occur on axons (axo-axonal), on dendrites (axodendritic), on soma (axo-somatic) and on dendritic spines...

Neurotransmissão glutamatérgica e plasticidade sináptica: aspectos moleculares, clínicos e filogenéticos / Glutamatergic neurotransmission and synaptic plasticity: molecular, clinical, and phylogenetic aspects

A comunicação entre neurônios é passível de constantes modificações, até mesmo no encéfalo adulto. Esta capacidade de circuitos neuronais fortalecerem ou enfraquecerem suas interações sinápticas específicas (fenômeno conhecido como plasticidade sináptica) pode ocorrer de acordo com as diferentes demandas ambientais, o que favorece a noção de que alterações dinâmicas na comunicação entre neurônios estão na base da flexibilidade comportamental (i.e., processos de aprendizagem e memória). Nas últimas décadas, o avanço das neurociências tem permitido uma melhor compreensão a respeito da plasticidade sináptica, especialmente a plasticidade de sinapses glutamatérgicas, cujos processos moleculares de modificação sináptica parecem estar entre os mais comuns de todo o sistema desse progresso na ciência básica tem contribuído para uma melhor compreensão acerca dos processos patológicos envolvendo as sinapses glutamatérgicas, como a doença de Alzheimer. Além disso, a crescente compreensão sobre o funcionamento da comunicação glutamatérgica tem ajudado a esclarecer como as sinapses, em geral, teriam se originado e evoluído na escala filogenética do reino animal (Metazoa)...

Communication between neurons is subject to constant changes, even in the adult brain. This ability of neural circuits to strengthen or weaken their specific synaptic interactions (a phenomenon known assynaptic plasticity) may occur according to different environmental demands, which favors the idea that dynamic changes in the communication between neurons underlie behavioral flexibility (i.e., learning and memory processes). In recent decades, advances in neuroscience has allowed a better understanding of synaptic plasticity, specially the plasticity of glutamatergic synapses, whose molecular processes of synaptic change appear to be among the most common throughout the central nervous system.Much of this progress in basic science has contributed to a better understanding of pathological processes involving the glutamatergic synapses, such as Alzheimer's disease. Furthermore, the growing understanding about the physiology of glutamatergic communication has helped explain how synapses, in general, would have originated and evolved in the phylogenetic scale of the Metazoa...