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A Fairén

Publications and source records attributed to A Fairén.

At least 19 recordsLinked to original sources
PubMed2005

Pioneering a golden age of cerebral microcircuits: the births of the combined Golgi-electron microscope methods.

Theodor W. Blackstad devised methods by which the synaptic connectivity of neuron somata and their dendritic and axonal processes in the CNS could be analyzed by the combined use of light and electron microscope techniques. His first publication on that subject dates from 1965 and was contemporary to the independent research by William K. Stell. The Golgi method was an obvious neuronal marker at those times, and Blackstad and Stell showed that the Golgi precipitate is electron-dense and intracellular and, therefore, it could help identify in the electron microscope, with great accuracy, profiles of neurons initially visualized in light microscopy. Besides this convergent research, Blackstad demonstrated for the first time that anterograde axonal degeneration could be combined with the Golgi-electron microscope method, allowing the identification of the neurons whose dendritic or somatic profiles were postsynaptic to the severed axonal afferent projections. Last, but not least, Blackstad pioneered de-impregnation techniques for electron microscopy of Golgi preparations. This had a great impact in the study of synaptic circuitry. The present account is a remembrance of the events that linked these early attempts with the development of a de-impregnation method based on gold toning by Alan Peters and the present author.

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PubMed2002

Expression and distribution of metabotropic GABA receptor subtypes GABABR1 and GABABR2 during rat neocortical development.

To understand the possible contribution of metabotropic gamma-aminobutyric acid receptors (GABABR) in cortical development, we investigated the expression pattern and the cellular and subcellular localization of the GABABR1 and GABABR2 subtypes in the rat neocortex from embryonic day 14 (E14) to adulthood. At the light microscopic level, both GABABR1 and GABABR2 were detected as early as E14. During prenatal development, both subtypes were expressed highly in the cortical plate. Using double immunofluorescence, GABABR1 colocalized with GABABR2 in neurons of the marginal zone and subplate, indicating that these proteins are coexpressed and could be forming functional GABABRs during prenatal development in vivo. In contrast, only GABABR1 but not GABABR2 was detected in the tangentially migratory cells in the lower intermediate zone. During postnatal development, immunoreactivity for GABABR1 and GABABR2 was distributed mainly in pyramidal cells. Discrete GABABR1-immunopositive cell bodies of interneurons were present throughout the neocortex. In addition, GABABR1 but not GABABR2 was found in identified Cajal-Retzius cells in layer I. At the electron microscopic level, immunoreactivity for GABABR1 and GABABR2 was found in dendritic spines and dendritic shafts at extrasynaptic and perisynaptic sites throughout postnatal development. We further demonstrated the presynaptic localization of GABABR1 and GABABR2, as well as the association of the receptors with asymmetrical synaptic junctions. These results indicate potentially important roles for the GABABRs in the regulation of migratory processes during corticogenesis and in the modulation of synaptic transmission during early development of cortical circuitry.

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PubMed2002

Differential distribution of group I metabotropic glutamate receptors during rat cortical development.

Neurons in the rat cerebral cortex are enriched in group I metabotropic glutamate receptor (mGluR) subtypes and respond to their activation during development. To understand better the mechanisms by which mGluR1 and mGluR5 mediate these effects, the goal of this study was to elucidate the expression pattern and to determine the cellular and the precise subcellular localization of these two receptor subtypes in the rat neocortex and hippocampus during late prenatal and postnatal development. At the light microscopic level, mGluR1alpha and mGluR5 were first detected in the cerebral cortex with different expression levels at embryonic day E18. Thus, mGluR5 had a moderate expression, whereas mGluR1alpha was detected as a diffuse and weak labeling. mGluR5 was localized in some Cajal- Retzius cells as well as in other cell types, such as pioneer neurons of the marginal zone. During postnatal development, the distribution of the receptors dramatically changed. From P0 to around P10, mGluR1alpha was localized in identified, transient Cajal-Retzius cells of neocortex and hippocampus, until these cells disappear. In addition, a population of interneurons localized the receptor from the second/third postnatal week. In contrast, mGluR5 was localized mainly in pyramidal cells and in some interneurons, with a neuropilar staining throughout the cerebral cortex. At the electron microscopic level, the immunoreactivity for both group I mGluR subtypes was expressed postsynaptically. Using immunogold methods, mGluR1alpha and mGluR5 immunoreactivities were found throughout postnatal development at the edge of postsynaptic specialization of asymmetrical synapses. These results show that the two group I mGluRs have a differential expression pattern in neocortex and hippocampus that may suggest roles for the receptors in the early processing of cortical information and in the control of cortical developmental events.

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PubMed2001

Whole-mount confocal immunofluorescence of mammalian CNS.

Bright-field wholemount labeling techniques applied to the mammalian central nervous system (CNS) offer advantages over conventional methods based on sections since an immediate and three-dimensional view of the stained components is provided. It thereby becomes possible to survey and count large number of cells and fibers in their natural relationships. The ability of confocal laser scanning microscopy to visualize in one focal plane the fluorescence associated with multiple markers could be most valuable by the availability of reliable wholemount fluorescent techniques. Accordingly, based in our previously published bright-field wholemount protocols [Brain Res. Prot. 2 (1998) 165-173], we have devised an effective immmunofluorescence wholemount procedure. We show that reliable wholemount fluorescent staining can be obtained using isolated complete CNS aged up to rat embryonic day 17, with antibodies penetration in the millimeter range. Examples are shown of preparations in which colocalization can be observed in nerve cells of cytoskeletal and calcium-binding proteins.

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PubMed2001

Cajal-Retzius cells in early postnatal mouse cortex selectively express functional metabotropic glutamate receptors.

Glutamate receptors have been linked to the regulation of several developmental events in the CNS. By using cortical slices of early postnatal mice, we show that in layer I cells, glutamate produces intracellular calcium ([Ca(2+)](i)) elevations mediated by ionotropic and metabotropic glutamate receptors (mGluRs). The contribution of mGluRs to these responses was demonstrated by application of tACPD, an agonist to groups I and II mGluRs, which evoked [Ca(2+)](i) increases that could be reversibly blocked by MCPG, an antagonist to groups I and II mGluRs. In the absence of extracellular Ca(2+), repetitive applications of tACPD or quisqualate, an agonist to group I mGluRs, elicited decreasing [Ca(2+)](i) responses that were restored by refilling a thapsigargin-sensitive Ca(2+) store. The use of specific group I mGluR agonists CHPG and DHPG indicated that the functional mGluR in layer I was of the mGluR1 subtype. Subtype specific antibodies confirmed the presence of mGlur1 alpha, but not mGluR5, in Cajal-Retzius (Reelin-immunoreactive) neurons.

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PubMed2000

Cellular mosaics in the rat marginal zone define an early neocortical territorialization.

We describe a novel neuronal group of the neocortical primordium that is formed by the pioneer neurons, unlike the Cajal-Retzius cells and the subplate component of the preplate. These pioneer neurons of the preplate and the marginal zone send transient axonal projections into the nascent internal capsule, preceding the formation of the axonal projection from the subplate neurons. In wholemount preparations, the pioneer neurons of the preplate and the marginal zone cover the prospective neocortical territory from embryonic day (E) 12 to E17. Two subpopulations of pioneer neurons (defined by differential expression of calcium-binding proteins) group into well-defined cell clusters, separated by spaces containing a lower packing density of cells immunoreactive to the corresponding calcium-binding protein. In both cases, cell clustering was concomitant with fasciculation of their axons. Although both subpopulations cohabit in the same areas of the marginal zone, their clustering occurs in specific, well-delineated territories, giving a mosaic appearance to the surface of the neocortical primordium before the arrival of thalamocortical axons. We hypothesize that the fascicles of descending axons arising from defined territories of the marginal zone may intervene in the initial guidance of the subcortical projection from the subplate.

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PubMed1999

Ultrastructural localization of the alpha4-subunit of the neuronal acetylcholine nicotinic receptor in the rat substantia nigra.

The distribution of the alpha4-subunit of the neuronal nicotinic acetylcholine receptor (nAChR) in the rat brain was examined at light and electron microscopy levels using immunohistochemical staining. In the present study we demonstrate the specificity, in both tissue homogenates and brain sections, of a polyclonal antibody raised against the rat nAChR alpha4-subunit. The characterization of this antibody involved: (1) Western blot analysis of rat brain homogenates and membrane extracts from cells previously transfected with diverse combinations of neuronal nAChR subunits, and (2) immunohistochemistry using transfected cells and rat brain tissue. At the light microscope level, the alpha4-subunit-like-immunoreactivity (LI) was widely distributed in the rat brain and matched the distribution of the alpha4-subunit transcripts observed previously by in situ hybridization. Strong immunohistochemical labeling was detected in the mesencephalic dopaminergic nuclei. The nAChRs in this region are thought to be responsible for the modulation of dopaminergic transmission. The neurotransmitter identity of alpha4-immunolabeled neurons in the substantia nigra pars compacta (SNpc) and the ventral tegmental area was thus assessed by investigating the possible colocalization of the nAChR alpha4-subunit with tyrosine hydroxylase using confocal microscopy. The double labeling experiments unambiguously indicated that the alpha4-subunit-LI is present in dopaminergic neurons. At the electron microscope level, the neurons in the SNpc exhibited alpha4-subunit-LI in association with a minority of postsynaptic densities, suggesting that the alpha4-subunit may be a component of functional nAChRs mediating synaptic transmission between midbrain cholinergic neurons and mesencephalic dopaminergic neurons.

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PubMed1999

Functional NMDA and GABAA receptors in pioneer neurons of the cortical marginal zone.

Transient pioneer neurons in the neocortical marginal zone generate an early corticofugal axonal projection at E12-E16 (Meyer et al. 1998). We have analysed the functional activity of glutamate and GABA receptors in such cells by measuring changes in intracellular calcium concentrations ([Ca2+]i). The activation of GABAA receptors with muscimol, as well as bath application of glutamate, lead to increases in [Ca2+]i in pioneer neurons. The stimulatory action of glutamate is mostly produced through the NMDA-type of ionotropic receptors. Metabotropic glutamate receptor activation has no effect on [Ca2+]i. Consistent with such results, immunocytochemical studies showed a prominent expression of GABAA and NMDA receptors in pioneer neurons. The activation of such receptors may be implicated in the remodelling of pioneer neurons during development.

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PubMed1998

Different origins and developmental histories of transient neurons in the marginal zone of the fetal and neonatal rat cortex.

Two major classes of early-born neurons are distinguished during early corticogenesis in the rat. The first class is formed by the cortical pioneer neurons, which are born in the ventricular neuroepithelium all over the cortical primordium. They appear at embryonic day (E) 11.5 in the lateral aspect of the telencephalic vesicle and cover its whole surface on E12. These cells, which show intense immunoreactivity for calbindin and calretinin, are characterized by their large size and axonal projection. They remain in the marginal zone after the formation of the cortical plate; they project first into the ventricular zone, and then into the subplate and the internal capsule. Therefore, these cells are the origin of the earliest efferent pathway of the developing cortex. Pioneer neurons are only present in prenatal brains. The second class is formed by subpial granule neurons, which form the subpial granular layer (SGL), previously considered to be found exclusively in the human cortex. SGL neurons are smaller than pioneer neurons. They are generated in a transient compartment of the retrobulbar ventricle between E12 and E14, and we propose the hypothesis that they invade the marginal zone, through tangential subpial migration, at different moments of fetal life. SGL neurons contain calbindin, calretinin, and gamma-aminobutyric acid (GABA), but the GABA-immunoreactive group becomes inconspicuous before birth. The extracellular matrix-like glycoprotein reelin, a molecule crucial for cortical lamination, is prenatally expressed by SGL neurons; postnatally, it is present in both Cajal-Retzius cells and subpial pyriform cells, both derivatives of SGL cells. In the rat, Cajal-Retzius cells are horizontal neurons that remain only until the end of the first postnatal week. They are located in layer I at a critical distance of approximately 20 microm from the pial surface and express reelin and, only occasionally, calretinin. Subpial pyriform cells coexpress reelin and calretinin and remain in layer I longer than Cajal-Retzius cells. Both pioneer neurons and subpial granule neurons are specific to the cortex. They mark the limit between the rudimentary cerebral cortex and olfactory bulb in the rat during early corticogenesis.

Aging↗
PubMed1998

GABA-immunoreactive cells of the cortical primordium contribute to distinctly fated neuronal populations.

The roles of GABA during development, as either a putative neurotransmitter or a nonsynaptic trophic factor, are being discussed intensely in recent literature. We offer an anatomical framework to better understand these possible roles in the developing cerebral cortex. During the early development of the cerebral cortex, GABA-containing cells constitute an outstanding cell population in the primordial plexiform layer, but they later distribute into at least four compartments. These include (1) Cajal-Retzius cells in layer I and (2) the subplate cells. Certain of these GABA-containing cell groups may disappear either by ceasing their expression of GABA, dilution in a growing brain volume, or cell death, possibilities that are reviewed here. The chemical tags that characterize Cajal-Retzius cells, both in the forming isocortex and Ammon's horn, are discussed. Another cell population that also belongs to the primordial plexiform layer is formed by (3) the tangentially migrating cells of the deep intermediate layer. These migrate away from the isocortical primordium to invade, and contribute cells to, the forming stratum oriens of the Ammon's horn. Since these cells cross cortical area boundaries, their tangential migration is relevant to the issue of cortical area specification during development. Finally, GABA-immunoreactive cells in the developing cortical plate are considered to be (4) the future GABAergic interneurons. A hypothetical mechanism is presented here to explain their acquisition of laminar positions, which is known to take place simultaneously, and with an identical "inside-out gradient," to the pyramidal cells generated contemporarily.

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PubMed1996

Neurogranin in the development of the rat telencephalon.

We have used a novel antibody to map the distribution of the protein kinase C substrate protein RC3/neurogranin during the development of the rat telencephalon. Neurogranin appearance in the rat brain is biphasic: it shows an early stage of anatomically restricted, low-intensity expression, and a juvenile stage of anatomically widespread, high-intensity expression. Most of the structures that express neurogranin during development conserve it in the adult stage. Neurogranin expression starts on embryonic day 18 in two different sites-the amygdalar primordium and in the piriform cortex-and is confined to these structures until the first postnatal day (P1). On P1, neurogranin expression increases dramatically in intensity, and appears in the olfactory cortex, isocortex, subiculum and hippocampus. In the striatum, expression starts on P1 and extends to the caudoputamen and parts of the globus pallidus and septum. Particularly complex patterns of labelling can be seen in the amygdala and cerebral cortex. Cortical layers showing early expression are the presumptive layers 4 and 5 in the somatosensory cortex, and layers 2 and 5 in the anterior cingulate and agranular insular cortices. Immunoreactivity is found mostly in cell bodies during the early and juvenile stages, but by the end of the first postnatal week it starts being more apparent in the neuropil. This phenomenon probably reflects the intracellular translocation of neurogranin to distal parts of the dendrites and dendritic spines. This process culminates by the end of the second postnatal week, when the adult pattern is reached. According to the timing and anatomy of its distribution, expression of neurogranin seems to be independently regulated in each telencephalic region by specific signalling mechanisms. It is proposed, on this basis, that neurogranin could be implicated in neuronal differentiation and synaptogenesis during telencephalic development.

Aging↗
PubMed1996

Parvovirus minute virus of mice strain i multiplication and pathogenesis in the newborn mouse brain are restricted to proliferative areas and to migratory cerebellar young neurons.

Newborn BALB/c mice intranasally inoculated at birth with a lethal dose of the immunosuppressive strain of the parvovirus minute virus of mice (MVMi) developed motor disabilities and intention tremors with a high incidence by the day 6 postinfection (dpi). These neurological syndromes paralleled the synthesis of virus intermediate DNA replicative forms and yield of infectious particles in the brain, with kinetics that peaked by this time. The preferred virus replicative sites in the brain were established early in the infection (2 dpi) and at the onset of clinical symptoms (6 dpi) and were compared with major regions of cellular proliferative activity found after intraperitoneal injection of bromodeoxyuridine 24 h before encephalons were subjected to immunohistochemistry detection. At 2 dpi, viral capsid antigen was located in the laterodorsal thalamic and the pontine nuclei but not in the extensive proliferative regions of the mouse brain at this postnatal day. At 6 dpi, however, the neurotropism of the MVMi was highlighted by its ability to target the subventricular zone of the ventricles, the subependymal zone of the olfactory bulb, and the dentate gyrus of the hippocampus, which are the three main germinal centers of the cerebrum in mouse postbirth neurogenesis. Unexpectedly, in the cerebellum, the MVMi capsid antigen was confined exclusively to cells that have undergone mitosis and have migrated to the internal granular layer (IGL) and not to the proliferative external granular layer (EGL), which was stained with antiproliferative cell nuclear antigen antibody and is the main target in other parvovirus infections. This result implies temporal or differentiation coupling between MVMi cycle and neuroblast morphogenesis, since proliferative granules of the EGL should primarily be infected but must migrate in a virus carrier state into the IGL in order to express the capsid proteins. During migration, many cells undergo destruction, accounting for the marked hypocellularity specifically found in the IGL and the irregular alignment of Purkinje cell bodies, both consistent histopathological hallmarks of animals developing cerebellar symptoms. We conclude that MVMi impairs postmitotic neuronal migration occurring in the first postnatal week, when, through the natural respiratory route of infection, the virus titer peaks in the encephalon. The results illustrate the intimate connection between MVMi neuropathogenesis and mouse brain morphogenetic stage, underscoring the potential of parvoviruses as markers of host developmental programs.

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PubMed1994

Cortical cells that migrate beyond area boundaries: characterization of an early neuronal population in the lower intermediate zone of prenatal rats.

Studies of the early development of the mammalian cerebral cortex have revealed that the earliest generated neurons that form the primordial plexiform layer (also called preplate or marginal zone) distribute among layer I and layer VII (subplate). By means of bromodeoxyuridine labelling of cells becoming postmitotic, we have found evidence that, in the rat, an additional group of neurons of the primordial plexiform layer remains in the close vicinity of the ventricular zone. This finding, in line with the proposal by Marín-Padilla (Z. Anat. Entwicklungsgesch., 134, 117-145, 1971), implies that the primordial plexiform layer suffers a tripartition after the formation of the cortical plate and of the intermediate zone (the latter soon becomes the embryonic white matter). Thus, primordial plexiform layer derivatives are in layer I, layer VII (subplate) and in the lower part of the embryonic white matter. This early generated neuronal population is also revealed with an antibody that recognizes the larger (67 kDa) isoform of glutamic acid decarboxylase (Kaufman et al., Science, 232, 1138-1140, 1986). This is in accord with the earlier finding of a GABA-containing cell population showing a similar spatiotemporal distribution. The early generated neurons of the embryonic white matter migrate tangentially and, in early postnatal animals, are found as interstitial cells in the medial regions of the subcortical white matter and at the midline in the corpus callosum. At caudal levels, similar cells invade the subpyramidal strata of the developing hippocampus. This tangential migration might explain the tangential dispersion of neural cell clones described in recent studies of cell lineage in the cerebral cortex.

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PubMed1993

A simple and reliable method for correlative light and electron microscopic studies.

We describe in detail a simple method for flat-embedding that can be subsequently used in correlative light and electron microscopic studies. The method can be applied to any material suitable for electron microscopy and is especially useful for study of the synaptology and ultrastructural characteristics of immunocytochemically or morphologically identified neurons or their processes. We present here an example to show how accurately one can delineate the fine details of a complex axonal arborization impregnated with the Golgi method in the mouse cerebral cortex. Golgi-impregnated sections to be studied at the electron microscopic level are osmicated, dehydrated, infiltrated with Araldite resin, flat-embedded, and identified cells or processes photographed. Serial semi-thin sections (1-2 microns thick) are then cut with an ultramicrotome, examined with the light microscope, and the elements rephotographed. Selected semi-thin sections are then resectioned on the ultramicrotome at 60-70 nm and examined electron microscopically. This method allows the systematic and accurate localization of stained cells and processes throughout the successive steps of the procedure.

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PubMed1992

Electron microscopy of Golgi-impregnated interneurons: notes on the intrinsic connectivity of the cerebral cortex.

The Golgi-electron microscope technique has opened new avenues to explore the synaptic organization of the brain. In this article, we shall discuss basic methodological principles necessary to analyze axonal arborizations with this combined technique. To illustrate the applications of the method, we shall review the forms and distribution of the synapses in which the axonal arborizations of local cortical interneurons engage.

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