RESUMO
Human cortical and subcortical areas integrate emotion, memory, and cognition when interpreting various environmental stimuli for the elaboration of complex, evolved social behaviors. Pyramidal neurons occur in developed phylogenetic areas advancing along with the allocortex to represent 70-85% of the neocortical gray matter. Here, we illustrate and discuss morphological features of heterogeneous spiny pyramidal neurons emerging from specific amygdaloid nuclei, in CA3 and CA1 hippocampal regions, and in neocortical layers II/III and V of the anterolateral temporal lobe in humans. Three-dimensional images of Golgi-impregnated neurons were obtained using an algorithm for the visualization of the cell body, dendritic length, branching pattern, and pleomorphic dendritic spines, which are specialized plastic postsynaptic units for most excitatory inputs. We demonstrate the emergence and development of human pyramidal neurons in the cortical and basomedial (but not the medial, MeA) nuclei of the amygdala with cells showing a triangular cell body shape, basal branched dendrites, and a short apical shaft with proximal ramifications as "pyramidal-like" neurons. Basomedial neurons also have a long and distally ramified apical dendrite not oriented to the pial surface. These neurons are at the beginning of the allocortex and the limbic lobe. "Pyramidal-like" to "classic" pyramidal neurons with laminar organization advance from the CA3 to the CA1 hippocampal regions. These cells have basal and apical dendrites with specific receptive synaptic domains and several spines. Neocortical pyramidal neurons in layers II/III and V display heterogeneous dendritic branching patterns adapted to the space available and the afferent inputs of each brain area. Dendritic spines vary in their distribution, density, shapes, and sizes (classified as stubby/wide, thin, mushroom-like, ramified, transitional forms, "atypical" or complex forms, such as thorny excrescences in the MeA and CA3 hippocampal region). Spines were found isolated or intermingled, with evident particularities (e.g., an extraordinary density in long, deep CA1 pyramidal neurons), and some showing a spinule. We describe spiny pyramidal neurons considerably improving the connectional and processing complexity of the brain circuits. On the other hand, these cells have some vulnerabilities, as found in neurodegenerative Alzheimer's disease and in temporal lobe epilepsy.
RESUMO
The human cortical amygdaloid nucleus (CoA) receives exteroceptive sensory stimuli, modulates the functions coded by the intrinsic amygdaloid circuit, and constitutes the beginning of the limbic lobe continuum with direct and indirect connections toward subcortical, allocortical, and higher order neocortical areas. To provide basic data on the human CoA, we characterized and classified the neurons using the thionin and the "single-section" Golgi method adapted for postmortem brain tissue and light microscopy. We found 10 different types of neurons named according to the morphological features of the cell body, dendritic branches, and spine distribution. Most cells are multipolar spiny neurons with two or more primary dendrites, including pyramidal-like ones. Three-dimensional reconstructions evidenced the types and diversity of the dendritic spines in each neuron. The unlike density of spines along dendritic branches, from proximal to distal ones, indicate that the synaptic processing and plasticity can be different in each CoA neuron. Our study provides novel data on the neuronal composition of the human CoA indicating that the variety of cells in this region can have phylogenetic, ontogenetic, morphological, and likely functional implications for the integrated human brain function. This can reflect both a more complex subcortical synaptic processing of sensory and emotional information and an adaptation for species-specific social behavior display.
Assuntos
Complexo Nuclear Corticomedial/citologia , Neurônios/fisiologia , Adulto , Idoso , Dendritos/ultraestrutura , Espinhas Dendríticas/ultraestrutura , Humanos , Processamento de Imagem Assistida por Computador , Masculino , Pessoa de Meia-Idade , Plasticidade Neuronal/fisiologia , Células Piramidais/fisiologia , Células Piramidais/ultraestrutura , Olfato/fisiologia , Sinapses/fisiologiaRESUMO
BACKGROUND: Different approaches aim to unravel detailed morphological features of neural cells. Dendritic spines are multifunctional units that reflect cellular connectivity, synaptic strength and plasticity. NEW METHOD: A novel three-dimensional (3D) reconstruction procedure is introduced for visualization of dendritic spines from human postmortem brain tissue using brightfield microscopy. The segmentation model was based on thresholding the intensity values of the dendritic spine image along 'z' stacks. We used median filtering and removed false positives. Fine adjustments during image processing confirmed that the reconstructed image of the spines corresponded to the actual original data. RESULTS: Examples are shown for the cortical amygdaloid nucleus and the CA3 hippocampal area. Structure of spine heads and necks was evaluated at different angles. Our 3D reconstruction images display dendritic spines either isolated or in clusters, in a continuum of shapes and sizes, from simple to more elaborated forms, including the presence of spinule and complex 'thorny excrescences'. COMPARISON WITH EXISTING METHODS: The procedure has the advantages already described for the adapted "single-section" Golgi method, since it provides suitable results using human brains fixed in formalin for long time, is relatively easy, requires minimal equipment, and uses an algorithm for 3D reconstruction that provides high quality images and more precise morphological data. CONCLUSION: The procedure described here allows the reliable visualization and study of human dendritic spines with broad applications for normal controls and pathological studies.
Assuntos
Espinhas Dendríticas , Imageamento Tridimensional/métodos , Microscopia/métodos , Coloração pela Prata , Idoso , Humanos , Masculino , Software , Lobo Temporal/citologiaRESUMO
The posterodorsal medial amygdala (MePD) is responsive to androgens and participates in the integration of olfactory/vomeronasal stimuli for the display of sexual behavior in rats. Adult gonadectomy (GDX) affects the MePD structural integrity at the same time that impairs male mating behavior. At the cellular level, dendritic spines modulate excitatory synaptic transmission, strength, and plasticity. Here, we describe the effect of GDX on the number and shape of dendritic spines in the right and left MePD using confocal microscopy and 3D image reconstruction. Age-matched adult rats were intact (n = 6), submitted to a sham procedure (n = 4) or castrated and studied 90 days after GDX (n = 5). The MePD neurons have a density of 1.1 spines/dendritic µm composed of thin, mushroom-like, stubby/wide, and few ramified or atypical spines. Irrespective of brain hemisphere, GDX decreased the dendritic spine density in the MePD, but induced different effects on each spine type. That is, compared to control groups, GDX reduced (i) the number (up to 50%) of thin, mushroom-like, and ramified spines, and (ii) the size and the neck length of thin spines as well as the head diameter of ramified spines. Besides, GDX increased the number of stubby/wide and atypical spines (up to 140% and 400%, respectively). These data show that GDX promotes a cellular and synaptic reorganization in a spine-specific manner in the MePD. By altering the number and shape of these connectional elements, GDX can affect the neural transmission and hinder the function of integrated brain circuitries in the male brain.
Assuntos
Tonsila do Cerebelo/citologia , Castração , Espinhas Dendríticas/fisiologia , Tonsila do Cerebelo/metabolismo , Animais , Espinhas Dendríticas/ultraestrutura , Masculino , Ratos , Ratos WistarRESUMO
The medial nucleus of the amygdala (Me) is a component of the neural circuit for the interpretation of multimodal sensory stimuli and the elaboration of emotions and social behaviors in primates. We studied the presence, distribution, diverse shape, and connectivity of dendritic spines in the human Me of adult postmortem men. Data were obtained from the five types of multipolar neurons found in the Me using an adapted Golgi method and light microscopy, the carbocyanine DiI fluorescent dye and confocal microscopy, and transmission electron microscopy. Three-dimensional reconstruction of spines showed a continuum of shapes and sizes, with the spines either lying isolated or forming clusters. These dendritic spines were classified as stubby/wide, thin, mushroom-like, ramified or with an atypical morphology including intermediate shapes, double spines, and thorny excrescences. Pleomorphic spines were found from proximal to distal dendritic branches suggesting potential differences for synaptic processing, strength, and plasticity in the Me neurons. Furthermore, the human Me has large and thin spines with a gemmule appearance, spinules, and filopodium. The ultrastructural data showed dendritic spines forming monosynaptic or multisynaptic contacts at the spine head and neck, and with asymmetric or symmetric characteristics. Additional findings included en passant, reciprocal, and serial synapses in the Me. Complex long-necked thin spines were observed in this subcortical area. These new data reveal the diversity of the dendritic spines in the human Me likely involved with the integration and processing of local synaptic inputs and with functional implications in physiological and various neuropathological conditions.
Assuntos
Tonsila do Cerebelo/anatomia & histologia , Dendritos/ultraestrutura , Espinhas Dendríticas/ultraestrutura , Idoso , Axônios/ultraestrutura , Cadáver , Humanos , Masculino , Microscopia Confocal , Microscopia Eletrônica de Transmissão , Pessoa de Meia-IdadeRESUMO
The posterodorsal medial amygdala (MePD) is a sexually dimorphic area in the social behavior neural network, with high concentration of oxytocin (OT) receptors. Wild type (WT) and OT knockout (OTKO) females were studied in proestrus, and Golgi-impregnated spines in the MePD were classified. Results show that the OTKO group has increased density of thin, mushroom, and stubby/wide spines when compared to the WT (p<0.01 in all cases). These data indicate that OT is an important synaptic modulator in the MePD, a finding that is likely involved with the display of the female sexual behavior.
Assuntos
Tonsila do Cerebelo/citologia , Tonsila do Cerebelo/fisiologia , Espinhas Dendríticas/fisiologia , Ocitocina/fisiologia , Receptores de Ocitocina/fisiologia , Animais , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proestro , Receptores de Ocitocina/genéticaRESUMO
The posterodorsal medial amygdala (MePD) is a sex-steroid-sensitive area that modulates reproductive behavior in rats. The volume of the neuronal cell body, density of dendritic spines, and glial fibrillary acidic protein immunoreactivity are sexually dimorphic or affected by gonadal hormones in the MePD. Here we add new data to this panorama and describe the ultrastructure of the glial and axonal coverage of the perikaryal membrane and the somatic spines in the MePD of males and cycling females (in diestrus, early proestrus, late proestrus, and estrus). Transmission electron microscopy data (mean values from seven to 11 neurons per rat, five or six animals per group) showed that the rat MePD has most of the perikaryal membrane covered by glial processes and a relatively large amount (up to 40%) of axonal processes contacting the neuronal cell body. No statistically significant difference was found between groups for these somatic coverages (P > 0.5). However, the density of somatic spines along the length of the perikaryal membrane was higher in the late proestrus than in estrus (P < 0.05), and somatic spines in early and late proestrus showed variable shapes with stubby/wide, thin, mushroom-like, ramified, transitional or atypical aspects. These findings add to the rapid adjustable synaptic changes in the MePD and in the integrated neural circuits that control neuroendocrine secretion and the hormonally modulated timely display of social behaviors in rats.
Assuntos
Tonsila do Cerebelo/ultraestrutura , Axônios/ultraestrutura , Espinhas Dendríticas/ultraestrutura , Ciclo Estral/fisiologia , Neuroglia/ultraestrutura , Caracteres Sexuais , Tonsila do Cerebelo/fisiologia , Animais , Axônios/fisiologia , Espinhas Dendríticas/fisiologia , Feminino , Masculino , Microscopia Eletrônica de Transmissão , Neuroglia/fisiologia , Ratos WistarRESUMO
The medial nucleus (Me) is a superficial component of the amygdaloid complex. Here we assessed the density and morphology of the neurons and glial cells, the glial fibrillary acidic protein (GFAP) immunoreactivity, and the ultrastructure of the synaptic sites in the human Me. The optical fractionator method was applied. The Me presented an estimated mean neuronal density of 1.53 × 105 neurons/mm³ (greater in the left hemisphere), more glia (72% of all cells) than neurons, and a nonneuronal:neuronal ratio of 2.7. Golgi-impregnated neurons had round or ovoid, fusiform, angular, and polygonal cell bodies (10-30 µm in diameter). The length of the dendrites varied, and pleomorphic spines were found in sparsely spiny or densely spiny cells (1.5-5.2 spines/dendritic µm). The axons in the Me neuropil were fine or coarsely beaded, and fibers showed simple or notably complex collateral terminations. The protoplasmic astrocytes were either isolated or formed small clusters and showed GFAP-immunoreactive cell bodies and multiple branches. Furthermore, we identified both asymmetrical (with various small, clear, round, electron-lucent vesicles and, occasionally, large, dense-core vesicles) and symmetrical (with small, flattened vesicles) axodendritic contacts, also including multisynaptic spines. The astrocytes surround and may compose tripartite or tetrapartite synapses, the latter including the extracellular matrix between the pre- and the postsynaptic elements. Interestingly, the terminal axons exhibited a glomerular-like structure with various asymmetrical contacts. These new morphological data on the cellular population and synaptic complexity of the human Me can contribute to our knowledge of its role in health and pathological conditions.
Assuntos
Tonsila do Cerebelo/citologia , Astrócitos/ultraestrutura , Neurônios/ultraestrutura , Sinapses/ultraestrutura , Idoso , Idoso de 80 Anos ou mais , Astrócitos/metabolismo , Axônios/ultraestrutura , Contagem de Células , Forma Celular , Corantes , Dendritos/ultraestrutura , Espinhas Dendríticas/ultraestrutura , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Fenotiazinas , Coloração pela Prata/métodos , Vesículas Sinápticas/ultraestruturaRESUMO
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.
Assuntos
Tonsila do Cerebelo/ultraestrutura , Forma Celular/fisiologia , Espinhas Dendríticas/ultraestrutura , Neurônios/ultraestrutura , Sinapses/ultraestrutura , Tonsila do Cerebelo/citologia , Tonsila do Cerebelo/fisiologia , Animais , Carbocianinas/farmacocinética , Espinhas Dendríticas/metabolismo , Corantes Fluorescentes/farmacocinética , Masculino , Microscopia Confocal/métodos , Neurônios/citologia , Neurônios/fisiologia , Ratos , Ratos Wistar , Sinapses/metabolismoRESUMO
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.