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Several studies show direct connections between primary sensory cortices involved in multisensory integration. The purpose of this study is to understand the microcircuitry of the reciprocal connections between visual and somatosensory cortices. The laminar distribution of retrogradely labeled cell bodies in V1 and in the somatosensory cortex both in (S1BF) and outside (S1) the barrel field was studied to provide layer indices in order to determine whether the connections are of feedforward, feedback or lateral type. Single axons were reconstructed and the size of their swellings was stereologically sampled. The negative layer indices in S1 and S1BF and the layer index near zero in V1 indicate that the connection from S1BF to V1 is of feedback type while the opposite is of lateral type. The greater incidence of larger axonal swellings in the projection from V1 to S1BF strongly suggests that S1BF receives a stronger driver input from V1 and that S1BF inputs to V1 have a predominant modulatory influence.
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Neuronas/citología , Corteza Somatosensorial/fisiología , Vías Visuales , Animales , Ratones Endogámicos C57BL , Vías NerviosasRESUMEN
Background: Brain banks provide small tissue samples to researchers, while gross anatomy laboratories could provide larger samples, including complete brains to neuroscientists. However, they are preserved with solutions appropriate for gross-dissection, different from the classic neutral-buffered formalin (NBF) used in brain banks. Our previous work in mice showed that two gross-anatomy laboratory solutions, a saturated-salt-solution (SSS) and an alcohol-formaldehyde-solution (AFS), preserve antigenicity of the main cellular markers (neurons, astrocytes, microglia, and myelin). Our goal is now to compare the quality of histology and antigenicity preservation of human brains fixed with NBF by immersion (practice of brain banks) vs. those fixed with a SSS and an AFS by whole body perfusion, practice of gross-anatomy laboratories. Methods: We used a convenience sample of 42 brains (31 males, 11 females; 25-90 years old) fixed with NBF (N = 12), SSS (N = 13), and AFS (N = 17). One cm3 tissue blocks were cut, cryoprotected, frozen and sliced into 40 µm sections. The four cell populations were labeled using immunohistochemistry (Neurons = neuronal-nuclei = NeuN, astrocytes = glial-fibrillary-acidic-protein = GFAP, microglia = ionized-calcium-binding-adaptor-molecule1 = Iba1 and oligodendrocytes = myelin-proteolipid-protein = PLP). We qualitatively assessed antigenicity and cell distribution, and compared the ease of manipulation of the sections, the microscopic tissue quality, and the quality of common histochemical stains (e.g., Cresyl violet, Luxol fast blue, etc.) across solutions. Results: Sections of SSS-fixed brains were more difficult to manipulate and showed poorer tissue quality than those from brains fixed with the other solutions. The four antigens were preserved, and cell labeling was more often homogeneous in AFS-fixed specimens. NeuN and GFAP were not always present in NBF and SSS samples. Some antigens were heterogeneously distributed in some specimens, independently of the fixative, but an antigen retrieval protocol successfully recovered them. Finally, the histochemical stains were of sufficient quality regardless of the fixative, although neurons were more often paler in SSS-fixed specimens. Conclusion: Antigenicity was preserved in human brains fixed with solutions used in human gross-anatomy (albeit the poorer quality of SSS-fixed specimens). For some specific variables, histology quality was superior in AFS-fixed brains. Furthermore, we show the feasibility of frequently used histochemical stains. These results are promising for neuroscientists interested in using brain specimens from anatomy laboratories.
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Neuroscientific research that requires brain tissue depends on brain banks that provide very small tissue samples fixed by immersion in neutral-buffered formalin (NBF), while anatomy laboratories could provide full brain specimens. However, these brains are generally fixed by perfusion of the full body with solutions other than NBF generally used by brain banks, such as an alcohol-formaldehyde solution (AFS) that is typically used for dissection and teaching. Therefore, fixation quality of these brains needs to be assessed to determine their usefulness in post-mortem investigations through magnetic resonance imaging (MRI) and histology, two common neuroimaging modalities. Here, we report the characteristics of five brains fixed by full body perfusion of an AFS from our Anatomy Laboratory suspected of being poorly fixed, given the altered signal seen on T1w MRI scans in situ. We describe 1- the characteristics of the donors; 2- the fixation procedures applied for each case; 3- the tissue contrast characteristics of the T1w and T2w images; 4- the macroscopic tissue quality after extraction of the brains; 5- the macroscopic arterial characteristics and presence or absence of blood clots; and 6- four histological stains of the areas that we suspected were poorly fixed. We conclude that multiple factors can affect the fixation quality of the brain. Nevertheless, cases in which brain fixation is suboptimal, consequently altering the T1w signal, still have T2w of adequate gray-matter to white-matter contrast and may also be used for histology stains with sufficient quality.
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Encéfalo , Imagen por Resonancia Magnética , Fijación del Tejido , Humanos , Encéfalo/diagnóstico por imagen , Encéfalo/anatomía & histología , Fijación del Tejido/métodos , Imagen por Resonancia Magnética/métodos , Masculino , Femenino , Anciano , Fijadores , Persona de Mediana Edad , Formaldehído , Anciano de 80 o más Años , AdultoRESUMEN
The purpose of this study was to identify and compare the afferent projections to the primary visual cortex in intact and enucleated C57BL/6 mice and in ZRDCT/An anophthalmic mice. Early loss of sensory-driven activity in blind subjects can lead to activations of the primary visual cortex by haptic or auditory stimuli. This intermodal activation following the onset of blindness is believed to arise through either unmasking of already present cortical connections, sprouting of novel cortical connections or enhancement of intermodal cortical connections. Studies in humans have similarly demonstrated heteromodal activation of visual cortex following relatively short periods of blindfolding. This suggests that the primary visual cortex in normal sighted subjects receives afferents, either from multisensory association cortices or from primary sensory cortices dedicated to other modalities. Here cortical afferents to the primary visual cortex were investigated to determine whether the visual cortex receives sensory input from other modalities, and whether differences exist in the quantity and/or the structure of projections found in sighted, enucleated and anophthalmic mice. This study demonstrates extensive direct connections between the primary visual cortex and auditory and somatosensory areas, as well as with motor and association cortices in all three animal groups. This suggests that information from different sensory modalities can be integrated at early cortical stages and that visual cortex activations following visual deprivations can partly be explained by already present intermodal corticocortical connections.
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Corteza Visual/fisiología , Vías Visuales/fisiología , Animales , Anoftalmos/fisiopatología , Corteza Auditiva/fisiología , Ceguera/fisiopatología , Ceguera/cirugía , Enucleación del Ojo , Ratones , Ratones Endogámicos C57BL , Neuronas/citología , Neuronas/fisiología , Corteza Somatosensorial/fisiología , Corteza Visual/citología , Corteza Visual/fisiopatología , Vías Visuales/citología , Vías Visuales/fisiopatologíaRESUMEN
Background: Histology remains the gold-standard to assess human brain biology, so ex vivo studies using tissue from brain banks are standard practice in neuroscientific research. However, a larger number of specimens could be obtained from gross anatomy laboratories. These specimens are fixed with solutions appropriate for dissections, but whether they also preserve brain tissue antigenicity is unclear. Therefore, we perfused mice brains with solutions used for human body preservation to assess and compare the tissue quality and antigenicity of the main cell populations. Materials and methods: Twenty-eight C57BL/6J mice were perfused with 4% formaldehyde (FAS, N = 9), salt-saturated solution (SSS, N = 9), and alcohol solution (AS, N = 10). The brains were cut into 40 µm sections for antigenicity analysis and were assessed by immunohistochemistry of four antigens: neuronal nuclei (NeuN), glial fibrillary acidic protein (GFAP astrocytes), ionized calcium-binding adaptor molecule 1 (Iba1-microglia), and myelin proteolipid protein (PLP). We compared the fixatives according to multiple variables: perfusion quality, ease of manipulation, tissue quality, immunohistochemistry quality, and antigenicity preservation. Results: The perfusion quality was better using FAS and worse using AS. The manipulation was very poor in SSS brains. FAS- and AS-fixed brains showed higher tissue and immunohistochemistry quality than the SSS brains. All antigens were readily observed in every specimen, regardless of the fixative solution. Conclusion: Solutions designed to preserve specimens for human gross anatomy dissections also preserve tissue antigenicity in different brain cells. This offers opportunities for the use of human brains fixed in gross anatomy laboratories to assess normal or pathological conditions.
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The cortical processing of visual information is thought to follow a hierarchical framework. This framework of connections between visual areas is based on the laminar patterns of direct feedforward and feedback cortico-cortical projections. However, this view ignores the cortico-thalamo-cortical projections to the pulvinar nucleus in the thalamus, which provides an alternative transthalamic information transfer between cortical areas. It was proposed that corticothalamic (CT) pathways follow a similar hierarchical pattern as cortico-cortical connections. Two main types of CT projections have been recognized: drivers and modulators. Drivers originate mainly in Layer 5 whereas modulators are from Layer 6. Little is known about the laminar distribution of these projections to the pulvinar across visual cortical areas. Here, we analyzed the distribution of CT neurons projecting to the lateral posterior (LP) thalamus in two species: cats and mice. Injections of the retrograde tracer fragment B of cholera toxin (CTb) were performed in the LP. The morphology and cortical laminar distribution of CTb-labeled neurons was assessed. In cats, neurons were mostly found in Layer 6 except in Area 17, where they were mostly in Layer 5. In contrast, CT neurons in mice were mostly located in Layer 6 across all areas. Thus, our results demonstrate that CT projections in mice do not follow the same organization as cats suggesting that the transthalamic pathways play distinct roles in these species.
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Gatos/anatomía & histología , Corteza Cerebral/citología , Ratones/anatomía & histología , Pulvinar/citología , Vías Visuales/citología , Animales , Femenino , Masculino , Ratones Endogámicos C57BL , Especificidad de la EspecieRESUMEN
Signals from lower cortical visual areas travel to higher-order areas for further processing through cortico-cortical projections, organized in a hierarchical manner. These signals can also be transferred between cortical areas via alternative cortical transthalamic routes involving higher-order thalamic nuclei like the pulvinar. It is unknown whether the organization of transthalamic pathways may reflect the cortical hierarchy. Two axon terminal types have been identified in corticothalamic (CT) pathways: the types I (modulators) and II (drivers) characterized by thin axons with small terminals and by thick axons and large terminals, respectively. In cats, projections from V1 to the pulvinar complex comprise mainly type II terminals, whereas those from extrastriate areas include a combination of both terminals suggesting that the nature of CT terminals varies with the hierarchical order of visual areas. To test this hypothesis, distribution of CT terminals from area 21a was charted and compared with 3 other visual areas located at different hierarchical levels. Results demonstrate that the proportion of modulatory CT inputs increases along the hierarchical level of cortical areas. This organization of transthalamic pathways reflecting cortical hierarchy provides new and fundamental insights for the establishment of more accurate models of cortical signal processing along transthalamic cortical pathways.
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BACKGROUND: MRI-histology correlation studies of the ex vivo brain mostly employ fresh, extracted (ex situ) specimens, aldehyde fixed by immersion, which has several disadvantages for MRI scanning (e.g. deformation of the organ). A minority of studies are done ex vivo-in situ (unfixed brain), requiring an MRI scanner readily available within a few hours of the time of death. NEW METHOD: We propose a new technique, exploited by anatomists, for scanning the ex vivo brain: fixation by whole body perfusion, which implies fixation of the brain in situ. This allows scanning the brain surrounded by fluids, meninges, and skull, preserving the structural relationships of the brain in vivo. To evaluate the proposed method, five heads perfused-fixed with a saturated sodium chloride solution were employed. Three sequences were acquired on a 1.5â¯T MRI scanner: T1weighted, T2weighted-FLAIR, and Gradient-echo. Histology analysis included immunofluorescence for myelin basic protein and neuronal nuclei. RESULTS: All MRIs were successfully processed through a validated pipeline used with in vivo MRIs. All cases exhibited positive antigenicity for myelin and neuronal nuclei. COMPARISON WITH EXISTING METHODS: All scans registered to a standard neuroanatomical template in pseudo-Talairach space more accurately than an ex vivo-ex situ scan. The time interval to scan the ex vivo brain in situ was increased to at least 10 months. CONCLUSIONS: MRI and histology study of the ex vivo-in situ brain fixed by perfusion is an alternative approach that has important procedural and practical advantages over the two standard methods to study the ex vivo brain.
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Técnicas Histológicas , Imagen por Resonancia Magnética , Encéfalo/diagnóstico por imagen , HumanosRESUMEN
Anatomical and imaging studies show ample evidence for auditory activation of the visual cortex following early onset of blindness in both humans and animal models. Anatomical studies in animal models of early blindness clearly show intermodal pathways through which auditory information can reach the primary visual cortex. There is clear evidence for intermodal corticocortical pathways linking auditory and visual cortex and also novel connections between the inferior colliculus and the visual thalamus. A recent publication [L.K. Laemle, N.L. Strominger, D.O. Carpenter, Cross-modal innervation of primary visual cortex by auditory fibers in congenitally anophthalmic mice, Neurosci. Lett. 396 (2006) 108-112] suggested the presence of a direct reciprocal connection between the inferior colliculus and the primary visual cortex (V1) in congenitally anophthalmic ZRDCT/An mice. This implies that this mutant mouse would be the only known vertebrate having a direct tectal connection with a primary sensory cortex. The presence of this peculiar pathway was reinvestigated in the ZRDCT/An mouse with highly sensitive neuronal tracers. We found the connections normally described in the ZRDCT/An mouse between: (i) the inferior colliculus and the dorsal lateral geniculate nucleus, (ii) V1 and the superior colliculus, (iii) the lateral posterior nucleus and V1 and between (iv) the inferior colliculus and the medial geniculate nucleus. We also show unambiguously that the auditory subcortical structures do not connect the primary visual cortex in the anophthalmic mouse. In particular, we find no evidence of a direct projection from the auditory mesencephalon to the cortex in this animal model of blindness.
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Anoftalmos/patología , Vías Auditivas/patología , Mapeo Encefálico , Tálamo/patología , Corteza Visual/patología , Animales , Vías Auditivas/fisiopatología , Biotina/análogos & derivados , Biotina/metabolismo , Dextranos/metabolismo , Ratones , Ratones Mutantes NeurológicosRESUMEN
The development of the corpus callosum (CC) and the anterior commissure (CA) is well known in a wide variety of species. No study, however, has described the development of the commissure of the superior colliculus (CSC) from embryonic state to adulthood in mammals. In this study, by using the lipophylic tracer DiI, we investigated the ontogeny of this mesencephalic commissure in the hamster at various ages. The development of axonal terminals, growth cone morphologies, and axons branching were described for the superior colliculus (SC) contralateral to the tracer injection. The first CSC axons cross the midline at embryonic day 11 (E-11) and grow further into the intermediate layers of the contralateral SC between E-12 and E-14. There is little axon growth therein between E-14 and the day of birth (P-0). Growth cones at the tip of these axons adopt complex morphologies at E-12 and progressively simplify until P-0. Pioneer axons are clearly visible between E-14 and P-1. These are followed by other axons progressively more numerous between P-0 and P-5. Axons do not show any branching until P-2. Between P-3 and P-9, the axons progressively arborize in the intermediate layers. Some axons reach the superficial layers at P-5, and they become more numerous around P-11, and only a few axons remain therein by P-21. Myelinated axons appear at P11 and are very dense at P-21. Our results indicate that the CSC follows developmental schemes similar to those of the CC and the AC but that initial axon midline crossing occurs earlier.
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Mesocricetus , Colículos Superiores , Animales , Carbocianinas/metabolismo , Cricetinae , Embrión de Mamíferos/anatomía & histología , Colorantes Fluorescentes/metabolismo , Mesocricetus/anatomía & histología , Mesocricetus/crecimiento & desarrollo , Neuronas/citología , Neuronas/metabolismo , Colículos Superiores/anatomía & histología , Colículos Superiores/crecimiento & desarrolloRESUMEN
In blind individuals, visually deprived occipital areas are activated by non-visual stimuli. The extent of this cross-modal activation depends on the age at onset of blindness. Cross-modal inputs have access to several anatomical pathways to reactivate deprived visual areas. Ectopic cross-modal subcortical connections have been shown in anophthalmic animals but not in animals deprived of sight at a later age. Direct and indirect cross-modal cortical connections toward visual areas could also be involved, yet the number of neurons implicated is similar between blind mice and sighted controls. Changes at the axon terminal, dendritic spine or synaptic level are therefore expected upon loss of visual inputs. Here, the proteome of V1, V2M and V2L from P0-enucleated, anophthalmic and sighted mice, sharing a common genetic background (C57BL/6J x ZRDCT/An), was investigated by 2-D DIGE and Western analyses to identify molecular adaptations to enucleation and/or anophthalmia. Few proteins were differentially expressed in enucleated or anophthalmic mice in comparison to sighted mice. The loss of sight affected three pathways: metabolism, synaptic transmission and morphogenesis. Most changes were detected in V1, followed by V2M. Overall, cross-modal adaptations could be promoted in both models of early blindness but not through the exact same molecular strategy. A lower metabolic activity observed in visual areas of blind mice suggests that even if cross-modal inputs reactivate visual areas, they could remain suboptimally processed.
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Anoftalmos/genética , Anoftalmos/fisiopatología , Ceguera/fisiopatología , Corteza Visual/fisiopatología , Vías Visuales/fisiopatología , Animales , Ceguera/genética , Enucleación del Ojo , Expresión Génica/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteoma , Transmisión Sináptica , Corteza Visual/citologíaRESUMEN
The laminar distribution of several distinct populations of neurofilament protein containing neurons has been used as a criterion for the delineation of cortical areas in hamsters. SMI-32 is a monoclonal antibody that recognizes a non-phosphorylated epitope on the medium- and high-molecular weight subunits of neurofilament proteins. As in carnivores and primates, SMI-32 immunoreactivity in the hamster neocortex was present in cell bodies, proximal dendrites and axons of some medium and large pyramidal neurons located in cortical layers III, V and VI. A small population of labeled multipolar cells was also found in layer IV. Neurofilament protein immunoreactive neurons were found throughout isocortical areas. Very few labeled cells were encountered in supplemental motor area, insular cortex, medial portion of associative visual cortex and in parietal association cortex. Our data indicate that SMI-32 immunoreactive cells can be efficiently used to trace boundaries between neocortical areas in the hamster's brain. The regional distribution SMI-32 immunoreactivity in the hamster cortex corresponds quite closely with cortical areas as defined by their cytoarchitecture and myeloarchitecture. The primary sensory cortical areas contain the most intense of SMI-32 immunoreactivity and are also those with the highest density of myelinated axons. Very low SMI-32 immunoreactivity was found in orbital, insular, perirhinal, cingulate and infralimbic cortices, which are also poor in myelinated axons. This supports the association between SMI-32 immunoreactivity and myelin contents.
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Corteza Cerebral/metabolismo , Mesocricetus/metabolismo , Proteínas de Neurofilamentos/metabolismo , Neuronas/metabolismo , Animales , Especificidad de Anticuerpos/inmunología , Mapeo Encefálico , Corteza Cerebral/citología , Cricetinae , Dendritas/metabolismo , Inmunohistoquímica , Mesocricetus/anatomía & histología , Fibras Nerviosas Mielínicas/metabolismo , Proteínas de Neurofilamentos/inmunología , Neuronas/citología , Células Piramidales/citología , Células Piramidales/metabolismoRESUMEN
Brains have evolved to optimize sensory processing. In primates, complex cognitive tasks must be executed and evolution led to the development of large brains with many cortical areas. Rodents do not accomplish cognitive tasks of the same level of complexity as primates and remain with small brains both in relative and absolute terms. But is a small brain necessarily a simple brain? In this review, several aspects of the visual cortical networks have been compared between rodents and primates. The visual system has been used as a model to evaluate the level of complexity of the cortical circuits at the anatomical and functional levels. The evolutionary constraints are first presented in order to appreciate the rules for the development of the brain and its underlying circuits. The organization of sensory pathways, with their parallel and cross-modal circuits, is also examined. Other features of brain networks, often considered as imposing constraints on the development of underlying circuitry, are also discussed and their effect on the complexity of the mouse and primate brain are inspected. In this review, we discuss the common features of cortical circuits in mice and primates and see how these can be useful in understanding visual processing in these animals.
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Ratones/anatomía & histología , Primates/anatomía & histología , Corteza Visual/anatomía & histología , Animales , Evolución Biológica , Humanos , Vías Nerviosas/anatomía & histología , Tamaño de los ÓrganosRESUMEN
In the mouse, visual extrastriate areas are located within distinct acallosal zones. It has been proposed that the striate-extrastriate and callosal projections are interdependent. In visually deprived mice, the normal patterns of callosal and striate-extrastriate projections are disrupted. It remains unknown whether visual deprivation affects the topography of V1-extrastriate projections and their relationship with callosal projections. Two anterograde tracers were injected in V1 and multiple retrograde tracer injections were performed in the contralateral hemisphere of intact and enucleated C57BL/6 mice and in ZRDCT/An mice to determine the effects of prenatal and postnatal afferent sensory activity on the topography of V1-extrastriate and callosal projections. Greater topographic anomalies were found in striate-extrastriate projections of anophthalmic than enucleated mice. In enucleated mice, the relationship between striate-extrastriate projections and callosal zones was highly variable. In anophthalmic mice, there was also a greater overlap between these projections. These results suggest that the prenatal afferent sensory activity regulates some aspects of the distribution of V1-extrastriate and callosal projections, in addition to the development of a normal topographic representation in extrastriate areas.
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Cuerpo Calloso/citología , Cuerpo Calloso/fisiología , Corteza Visual/citología , Corteza Visual/fisiología , Animales , Anoftalmos , Enucleación del Ojo , Ratones , Ratones Endogámicos C57BL , Vías Visuales/citología , Vías Visuales/fisiologíaRESUMEN
Anophthalmia is a condition in which the eye does not develop from the early embryonic period. Early blindness induces cross-modal plastic modifications in the brain such as auditory and haptic activations of the visual cortex and also leads to a greater solicitation of the somatosensory and auditory cortices. The visual cortex is activated by auditory stimuli in anophthalmic mice and activity is known to alter the growth pattern of the cerebral cortex. The size of the primary visual, auditory and somatosensory cortices and of the corresponding specific sensory thalamic nuclei were measured in intact and enucleated C57Bl/6J mice and in ZRDCT anophthalmic mice (ZRDCT/An) to evaluate the contribution of cross-modal activity on the growth of the cerebral cortex. In addition, the size of these structures were compared in intact, enucleated and anophthalmic fourth generation backcrossed hybrid C57Bl/6J×ZRDCT/An mice to parse out the effects of mouse strains and of the different visual deprivations. The visual cortex was smaller in the anophthalmic ZRDCT/An than in the intact and enucleated C57Bl/6J mice. Also the auditory cortex was larger and the somatosensory cortex smaller in the ZRDCT/An than in the intact and enucleated C57Bl/6J mice. The size differences of sensory cortices between the enucleated and anophthalmic mice were no longer present in the hybrid mice, showing specific genetic differences between C57Bl/6J and ZRDCT mice. The post natal size increase of the visual cortex was less in the enucleated than in the anophthalmic and intact hybrid mice. This suggests differences in the activity of the visual cortex between enucleated and anophthalmic mice and that early in-utero spontaneous neural activity in the visual system contributes to the shaping of functional properties of cortical networks.
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Anoftalmos/patología , Corteza Cerebral/crecimiento & desarrollo , Corteza Cerebral/patología , Enucleación del Ojo/efectos adversos , Animales , Anoftalmos/fisiopatología , Corteza Cerebral/fisiopatología , Ratones Endogámicos C57BL , Tamaño de los Órganos , Especificidad de la Especie , Núcleos Talámicos/crecimiento & desarrollo , Núcleos Talámicos/patología , Núcleos Talámicos/fisiopatologíaRESUMEN
The visual system in humans is considered the gateway to the world and plays a principal role in the plethora of sensory, perceptual and cognitive processes. It is therefore not surprising that quality of vision is tied to quality of life . Despite widespread clinical and basic research surrounding the causes of visual disorders, many forms of visual impairments, such as retinitis pigmentosa and macular degeneration, lack effective treatments. Non-human primates have the closest general features of eye development to that of humans. Not only do they have a similar vascular anatomy, but amongst other mammals, primates have the unique characteristic of having a region in the temporal retina specialized for high visual acuity, the fovea(1). Here we describe a general technique for dissecting the primate retina to provide tissue for retinal histology, immunohistochemistry, laser capture microdissection, as well as light and electron microscopy. With the extended use of the non-human primate as a translational model, our hope is that improved understanding of the retina will provide insights into effective approaches towards attenuating or reversing the negative impact of visual disorders on the quality of life of affected individuals.
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Encéfalo/citología , Disección/métodos , Ojo/citología , Retina/citología , Animales , Encéfalo/anatomía & histología , Crioultramicrotomía/métodos , Ojo/anatomía & histología , Humanos , Inmunohistoquímica/métodos , Primates , Retina/anatomía & histologíaRESUMEN
The use of non-human primates provides an excellent translational model for our understanding of developmental and aging processes in humans(1-6). In addition, the use of non-human primates has recently afforded the opportunity to naturally model complex psychiatric disorders such as alcohol abuse(7). Here we describe a technique for blocking the brain in the coronal plane of the vervet monkey (Chlorocebus aethiops sabeus) in the intact skull in stereotaxic space. The method described here provides a standard plane of section between blocks and subjects and minimizes partial sections between blocks. Sectioning a block of tissue in the coronal plane also facilitates the delineation of an area of interest. This method provides manageable sized blocks since a single hemisphere of the vervet monkey yields more than 1200 sections when slicing at 50 microm. Furthermore by blocking the brain into 1cm blocks, it facilitates penetration of sucrose for cyroprotection and allows the block to be sliced on a standard cryostat.
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Encéfalo/anatomía & histología , Disección/métodos , Técnicas Estereotáxicas , Animales , Encéfalo/fisiología , Encéfalo/cirugía , Chlorocebus aethiops , PrimatesRESUMEN
During development, retinal ganglion cells (RGCs) extend their axons toward their thalamic and mesencephalic targets. Their navigation is largely directed by guidance cues present in their environment. Since cAMP is an important second messenger that mediates the neural response to guidance molecules and its intracellular levels seem to decrease significantly following birth, we tested whether modulation of the cAMP/protein kinase A (PKA) pathway would affect the normal development of RGC axons. At postnatal day 1, hamsters received a unilateral intraocular injection of either 0.9% saline solution, 12 mM of the membrane-permeable cAMP analogue (dibutyryl cAMP; db-cAMP), or 10 microM of the PKA inhibitor KT5720. Intraocular elevation of cAMP significantly accelerated RGC axonal growth while inhibition of PKA activity decreased it. Moreover, when highly purified RGC cultures were treated with forskolin (an activator of adenylate cyclase) or cAMP analogues (db-cAMP and Sp-cAMP), neurite length, growth cone (GC) surface area and GC filopodia number were significantly increased. This indicates that intraocular elevation of cAMP acts directly on RGCs. Since these effects were prevented by PKA inhibitors, it demonstrates that cAMP also exerts its action via the PKA pathway. Taken together, these results suggest that the cAMP/PKA cascade is essential for the normal development of retinothalamic projections.
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Proteínas Quinasas Dependientes de AMP Cíclico/biosíntesis , Células Ganglionares de la Retina/citología , Células Ganglionares de la Retina/enzimología , Animales , Animales Recién Nacidos , Carbazoles/farmacología , Células Cultivadas , Cricetinae , AMP Cíclico/biosíntesis , AMP Cíclico/genética , Proteínas Quinasas Dependientes de AMP Cíclico/antagonistas & inhibidores , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Mesocricetus , Ratones , Pirroles/farmacología , Células Ganglionares de la Retina/efectos de los fármacosRESUMEN
The occipital cortex, normally visual, can be activated by auditory or somatosensory tasks in the blind. This cross-modal compensation appears after early or late onset of blindness with differences in activation between early and late blind. This could support the hypothesis of a reorganization of sensory pathways in the early blind that does not occur in later onset blindness. Using immunohistochemistry of the c-Fos protein following a white noise stimulus and injections of the anterograde tracer dextran-biotin in the inferior colliculus, we studied how the occurrence of blindness influences cross-modal compensation in the mutant anophthalmic mouse strain and in C57BL/6 mice enucleated at birth. We observed, in mutant mice, immunolabeled nuclei in the visual thalamus - the dorsal lateral geniculate nucleus - in the primary visual area (V1) and a few labeled nuclei in the secondary visual area (V2). In enucleated mice, we observed auditory activity mainly in V2 but also sparsely in V1. No labeled cells could be found in the visual thalamus. Tracing studies confirmed the difference between anophthalmic and birth-enucleated mice: whereas the first group showed inferior colliculus projections entering both the dorsal lateral geniculate and the latero-posterior nuclei, in the second, auditory fibers were found only within the latero-posterior thalamic nucleus. None was found in controls with intact eyes. We suggest that the prenatal period of spontaneous retinal activity shapes the differences of the sensory reorganization in mice.
Asunto(s)
Anoftalmos/fisiopatología , Percepción Auditiva/fisiología , Ceguera/fisiopatología , Vías Visuales/fisiopatología , Estimulación Acústica/métodos , Análisis de Varianza , Animales , Animales Recién Nacidos , Corteza Auditiva/metabolismo , Corteza Auditiva/patología , Biotina/análogos & derivados , Biotina/metabolismo , Recuento de Células/métodos , Dextranos/metabolismo , Modelos Animales de Enfermedad , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Proteínas Proto-Oncogénicas c-fos/metabolismo , Vías Visuales/patologíaRESUMEN
The morphology and distribution of neurons immunoreactive (ir) to parvalbumin (PV), calretinin (CR) and calbindin (CB) were studied in the primary visual (V1) and auditory (A1) cortices of hamsters. Cortical cell populations were labelled immunohistochemically using a glucose oxidase-diaminobenzidine-nickel combined revelation method. Quantitative analysis revealed significant differences between V1 and A1 in the density and distribution of their neuronal population. CBir cells exhibited several typologies in both cortical regions. Most cells were multipolar even though many of them had bitufted or bipolar morphologies. These cells were distributed in layers II/III and in layer V of both A1 and V1, but were more numerous in layer V of V1. CRir cells were of the fusiform type with long bipolar dendritic arbours. These were similarly distributed in both cortices with a peak in superficial layers II/III. PVir cells were also found in both cortices and had round or oval-shaped somata with multipolar processes. They were mostly located in layer V for V1 and in layers III/IV for A1. Visual and auditory primary cortices can thus be differentiated on the basis of their immunoreactivity to specific calcium binding proteins.