Alfonso Fairen

Instituto de Neurociencias · Developmental Neurobiology Unit

In rodents, the medial nucleus of the amygdala receives direct inputs from the accessory olfactory bulbs and is mainly implicated in pheromone-mediated reproductive and defensive behaviors. The principal neurons of the medial amygdala are GABAergic neurons generated principally in the caudo-ventral medial ganglionic eminence and preoptic area. Besi...

A unique population of cells, called “lot cells,” circumscribes the path of the lateral olfactory tract (LOT) in the rodent brain and acts to restrict its position at the lateral margin of the telencephalon. Lot cells were believed to originate in the dorsal pallium (DP). We show that Lhx2 null mice that lack a DP show a significant increase in the...

Human mutations in ZIC2 have been identified in patients with holoprosencephaly and schizophrenia. Similarly, Zic2 mutant mice exhibit holoprosencephaly in homozygosis and behavioral and morphological schizophrenic phenotypes associated with forebrain defects in heterozygosis. Despite the devastating effects of mutations in Zic2, the cellular and m...

A systematic classification and accepted nomenclature of neuron types is much needed but is currently lacking. This article describes a possible taxonomical solution for classifying GABAergic interneurons of the cerebral cortex based on a novel, web-based interactive system that allows experts to classify neurons with pre-determined criteria. Using...

A definitive consequence of the aging process is the progressive deterioration of higher cognitive functions. Defects in DNA repair mechanisms mostly result in accelerated aging and reduced brain function. DNA polymerase µ is a novel accessory partner for the non-homologous end-joining DNA repair pathway for double-strand breaks, and its deficiency...

Polµ−/− mice display reduced exploratory activity and enhanced sensorimotor coordination during aging. (A–B) Motor activity (defined as number of beam interruptions per 10 min) of 3-, 8-, and 18-month-old wild-type (black bars) or Polµ−/− (white bars) mice. (C–D) Maximum time of permanency on the rota-rod (C) for 3-, 8-, and 18-month-old wild-type...

Quantitative analysis of the relationships between the percentage of CRs and fEPSP slopes for the different experimental groups across habituation, conditioning, and extinction sessions. Data collected from 3-month-old wild-type (A) and Polµ−/− (B) mice and from 18-month-old wild-type (C) and Polµ−/− (D) mice are illustrated. Each point represents...

Characteristics of eyeblink responses evoked in young and aged Polµ−/− and wild-type mice. (A) A diagram indicating the location of stimulating (St.) electrodes implanted on the supraorbital nerve and electromyographic (EMG) recording electrodes implanted in the orbicularis oculi (O.O.) muscle. (B) Three superimposed records of the O.O. EMG respons...

Paired-pulse facilitation of field excitatory postsynaptic potentials (fEPSP) recordings in the CA1 area following stimulation of the ipsilateral Schaffer collateral-commissural pathway. Data were collected from extracellular fEPSP paired traces collected from 18-month-old wild-type and Polµ−/− mice at different inter-pulse intervals. The data show...

Evaluation of mitochondrial activity in brain. Crude extract or mitochondrial fractions, prepared as previously described (Birch-Machin and Turnbull, 2001) from wild-type or Polµ−/− brains, were evaluated for their mitochondria number (A), defined as citrate synthetase activity (CS IU) per mg of protein, or the mtDNA content (B), expressed as the r...

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Mechanistic working model for in vivo Polµ function in NHEJ reactions. (A) NHEJ (error-prone) and HR (error-free) are the fundamental pathways for double-strand break (DSB) repair in mammals, and both compete for the same substrates. The contribution of either mechanism is dictated by the avidity of the heterodimer ku70/ku80 and the activity of the...

Molecular characterization of Polμ−/− mice associated with aging. (A) Paraffin brain sections from wild-type and Polµ−/− mice (18–20 months old) were processed and stained for 53BP1 (M). (B) qRT-PCR expression analysis of a selection of genes (Atg5l, Atg7l, Atg12l, Maplc3b, and Atg9a) critical for execution or regulation of autophagy. Black bars co...

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Comparative evaluation of DNA repair activity in brain extracts. (A). Evaluation of gap filling activity in brain (wild-type vs. Polµ−/−) extracts, using the indicated labeled-template primer (upper part). After incubation (30′–1 h, 30°C) with the clarified extracts (10 µg), with the addition of the indicated concentration of ddCTP, products were r...

qRT-PCR expression analysis of a panel of aging-associated functions in brain of aged Polµ−/− mice. (A). Differential transcript levels of selected aging-related (Zahn et al., 2006) functions, related to inflammation (C1qa, C5a, C3aR, C5aR, Cox2), mitochondrial activity (Timm17a, Nfa10, Mrps12, Mrpl28, Atp5a), and apoptosis or p53 targets (Acin1, E...

To study the potential role of neural cell adhesion molecule (NCAM) in the development of thalamocortical (TC) axon topography, wild type, and NCAM null mutant mice were analyzed for NCAM expression, projection, and targeting of TC afferents within the somatosensory area of the neocortex. Here we report that NCAM and its α-2,8-linked polysialic aci...

The articles in this Special Topic cover a range of issues concerning long-distance projecting cortical GABAergic neurons, in the context of interneuron diversity. As several authors report, these neurons are attracting renewed attention spurred by new techniques and markers which show great potential for deciphering their role in cortical organiza...

The preplate of the cerebral cortex contains projection neurons that connect the cortical primordium with the subpallium. These are collectively named pioneer neurons. After preplate partition, most of these pioneer neurons become subplate neurons. Certain preplate neurons, however, never associate with the subplate but rather with the marginal zon...

Neuroscience produces a vast amount of data from an enormous diversity of neurons. A neuronal classification system is essential to organize such data and the knowledge that is derived from them. Classification depends on the unequivocal identification of the features that distinguish one type of neuron from another. The problems inherent in this a...

Metabotropic glutamate receptor 1 (mGluR1) has been related to processes underlying learning in hippocampal circuits, but demonstrating its involvement in synaptic plasticity when measured directly on the relevant circuit of a learning animal has proved to be technically difficult. We have recorded the functional changes taking place at the hippoca...

This essay explores the contributions to the organization of neuronal microcircuits in the cerebral cortex by Rafael Lorente de Nó, a renowned disciple of Santiago Ramón y Cajal. Lorente de Nó was impressed by the advances in functional parcellation of the cerebral cortex, and wished to find an anatomical correlate, not in cytoarchitectonic charts...

We have studied the role of rostral medial prefrontal cortex (mPFC) on reflexively evoked blinks and on classically conditioned eyelid responses in alert-behaving rabbits. The rostral mPFC was identified by its afferent projections from the medial half of the thalamic mediodorsal nuclear complex. Classical conditioning consisted of a delay paradigm...

A long-held assumption states that each dendritic spine in the cerebral cortex forms a synapse, although this issue has not been systematically investigated. We performed complete ultrastructural reconstructions of a large (n=144) population of identified spines in adult mouse neocortex finding that only 3.6% of the spines clearly lacked synapses....

Perlecan is a proteoglycan expressed in the basal lamina of the neuroepithelium during development. Perlecan absence does not impair basal lamina assembly, although in the 55% of the mutants early disruptions of this lamina conducts to exencephaly, impairing brain development. The rest of perlecan-null brains complete its prenatal development, main...

We describe a novel spontaneous autosomal recessive mutation, cervelet-4 (crv4), which arose in a BALB/c strain. Mice homozygous for the mutation exhibit principally a reduced body size, a congenital neurological phenotype characterized by ataxic gait and intention tremor, with no gross anomalies observed in brain or cerebellum, and skeletal anomal...

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. Th...

Metabotropic gamma-aminobutyric acid receptors (GABAB) play modulatory roles in central synaptic transmission and are involved in controlling neuronal migration during development. We used immunohistochemical methods to elucidate the expression pattern as well as the cellular and the precise subcellular localization of the GABA(B1a/b) and GABAB2 su...

Pyramidal neurons of the mammalian cerebral cortex are generated in the ventricular zone of the pallium whereas the subpallium provides the cortex with inhibitory interneurons. The marginal zone contains a subpial stream of migratory interneurons and two different classes of transient neurons, the pioneer neurons provided with corticofugal axons, a...

We investigated the substrates supporting neuronal migration, and its routes, during early thalamic development in the rat. Neurons and axonal and glial fibres were identified in embryos with single and double immunohistochemistry; dynamic data were obtained with cell tracers in short-term organotypic cultured slices. The earliest thalamic neurons,...

The reelin signaling pathway regulates laminar positioning of radially migrating neurons during cortical development. It has been suggested that reelin secreted by Cajal-Retzius cells in the marginal zone could provide either a stop or an attractant signal for migratory neurons expressing reelin receptors, but the proposed models fail to explain re...

The marginal zone of the developing cerebral cortex is formed by different types of neurons, some of which were described more than one century ago. It is the case of Cajal-Retzius cells, which are known to synthetize and secrete Reelin, an extracellular matrix glycoprotein critically involved in the radial migration and early cortical cytoarchitec...

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 GABAB...

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 pre...

The marginal zone of the developing cerebral cortex is formed by different types of neurons, some of which were described more than one century ago. It is the case of Cajal-Retzius cells, which are known to synthesize and secrete Reelin, an extracellular matrix glycoprotein critically involved in the radial migration and early cortical cytoarchitec...

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...

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 thes...

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 relation...

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 proj...

ty (LI) was widely distributed in the rat brain and matched the distribution of the #4-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. T...

The subpial granular layer (SGL) is a transient layer of small, undifferentiated cells located immediately below the pial surface of the cerebral cortex. It was first described by Ranke (1910) in the marginal zone (MZ), the prospective cortical layer I, of human fetuses around midgestation. While Ranke believed the SGL to be composed mainly of glia...

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...

The distribution of the α4-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 nACh...

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...

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...

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 cons...

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 f...

It is widely believed that inhibitory synapses are not present or present in only small numbers in the rodent cerebral cortex during the early postnatal period when the cortex is being innervated by thalamocortical fibers. Quantitative electron microscopy was carried out on the posteromedial barrel subfield of mouse somatosensory cortex from postna...

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-inte...

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...

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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...

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 sy...

It is not a futile exercise to analyze the scientific accomplishments of our predecessors; it may serve to better understand our own intellectual attitudes towards science.

Immunocytochemical techniques were used to analyze the distribution of the calcium-binding proteins calbindin and parvalbumin during the pre- and postnatal development of the rat somatosensory cortex. Calbindin occurs in most early differentiated neurons that form the primordial plexiform layer at embryonic day 14. This expression in transient; dur...

Species-specific antisera against rat or mouse immunoglobulins, such as the commercial ones currently used as secondary antibodies in immunohistochemistry, revealed an intense, developmentally regulated immunostaining of the cerebral cortex in the homospecific rodent species. The immunostaining showed evident anatomical restrictions, favouring the...

We have searched for the possible correlation of naturally occurring cell death with spontaneously enhanced c-fos expression in the developing cerebral cortex of normal Wistar albino rats. During the late prenatal and early postnatal period, cells with irregular contours and intracytoplasmic electron-dense granules (granule-containing cells) were a...

The distribution of calbindin immunoreactivity was studied in the developing rat dorsal thalamus at embryonic days 14, 16, 18 and 20. At early stages (days 14-16), calbindin is expressed throughout the dorsal thalamic cell mass. Most intense labeling occurs in cells adjacent to the ventricular surface, in a spatial gradient reflecting the well-know...

The present study has explored with immunocytochemical methods the expression of the proto-oncogene c-fos during the pre- and postnatal development of the cerebral cortex of the rat. The immunostaining of the Fos protein follows a strikingly precise spatiotemporal pattern: it occurs uniquely within layer VIb of the developing cerebral cortex, and i...

The maturation of the calcium binding proteins calbindin-D28k (CB) and parvalbumin (PV) during the first 3 postnatal weeks was studied in the rat thalamus using immunohistochemistry. These two proteins display a non-homogeneous distribution in the adult thalamus. In the rat, CB is mainly localized in the neurons and neuropil of the thalamic midline...

The distribution of the GABAA receptor/benzodiazepine receptor/chloride channel complex was investigated in the thalamus of the rat by means of immunohistochemistry in adulthood, as well as during embryonic and postnatal development, using a monoclonal antibody. In adults, the immunoreactivity for the GABAA receptor complex was intensely expressed...

The ontogenesis of cells showing GABA-like immunoreactivity, and the distribution of the immunoreactivity for the GABAA receptor were studied immunocytochemically in the prenatal rat brain. By embryonic day 14, a few GABA-like immunoreactive (GABA-positive) cells scattered at the subpial limit of the marginal zone (primordial plexiform layer) in th...

A population of cortical neurons contains the opioid peptide dynorphin; the laminar distribution of these neurons in the adult cerebral cortex and their patterns of development are not well known. We have utilized in situ hybridization techniques to localize prodynorphin mRNA-containing neurons. Rats aged from embryonic day (E) 15 through postnatal...

The neurogenetic gradients of neurons showing glutamic acid decarboxylase (GAD) immunoreactivity were determined in the regio superior and in the regio inferior of the mouse hippocampus. Pregnant C57Bl mice received pulse injections of (3H)thymidine from E11 through E17 (E0 being the day of mating). Distributions of (3H)thymidine-labeled, GAD-posit...

The temporal patterns of neurogenesis of cells showing glutamic acid decarboxylase (GAD) immunoreactivity were determined in the area dentata of the mouse. Pregnant C57Bl mice received pulse injections of (3H)thymidine from E11 through E17 (E0 being the day of mating). The distribution of (3H)thymidine-labeled, GAD-positive neurons in the hilus and...

During the early postnatal development of the rat, γ-aminobutryic acid (GABA)-like immunoreactive axons were contained within the subcortical white matter. Some of the immunoreactive axons crossed the midline, while others followed a vertical or longitudinal trajectory within the lateral part of the corpus callosum. Growth cones were occasionally o...

In the barrel cortex of the mouse, GAD-immunoreactive cells are positioned following an 'inside-out' gradient. Each cortical layer is populated by a variety of classes of GABAergic neurons. The period of neuronal generation for each cortical layer lasts for several days, but it is not known whether the different classes of GABAergic neurons are gen...

A single, isolated interneuron with axonal arcades in the cat visual cortex was analysed in detail by both light and electron microscopy. The neuron was impregnated by the Golgi-Kopsch method, gold-toned, and processed for electron microscopy using the ethanolic phosphotungstic acid (PTA) staining method of Bloom & Aghajanian (1968). These methods,...

Injections of the fluorescent tracer Fluoro-Gold were made in transplanted and normal cerebral cortex of rats in order to investigate and compare the local connectivities of both. In the normal somatosensory cortex, small injections in superficial layers (I to III) produced retrograde cell labeling below the injection site in two bands: in layer V...

We describe the morphological types of glutamic acid decarboxylase (GAD) immunoreactive cells in the barrel cortex of the mouse. A method is introduced and applied, which combines qualitative and quantitative criteria to classify these cells through the use of multivariate statistics. With the aid of an interactive computer microscope, 2010 GAD-pos...

The time course of the neurogenesis of cells showing glutamic acid decarboxylase (GAD) immunoreactivity has been analyzed in the dorsal hippocampus and area dentata of the mouse. The quantitative data indicate that γ-aminobutyric acid (GABA)ergic cells are generated prenatally, and that their peaks of neurogenesis occur before the peaks of neurogen...

The birth dates of neurons showing glutamic acid decarboxylase (GAD) immunoreactivity have been determined in mouse somatosensory cortex. Pregnant C57B1 mice received pulse injections of (3H)thymidine from E10 through E17 (EO being the day of mating). The distributions of thymidinelabeled, GAD-positive and nonimmunoreactive (non-GAD) cells as a fun...

The use of cyanocrylic glue to fix pieces of Golgi-stained nervous tissue on a paraffin blank is proposed for obtaining thick sections of unembedded tissue with a sliding microtome. This procedure makes correct orientation of the tissue easy during sectioning and makes it possible to obtain tissue sections quickly. The sections are flat-mounted usi...

The axonal arborizations of the basket cells in the cerebral neocortex have long been considered as the source of the presynaptic terminals contacting the cell bodies of pyramidal cells. Given that the concept of the cortical basket cell is based upon indirect evidence only, it was deemed worthwhile to re-investigate this problem using the Golgi-EM...

The axonal arborization of chandelier cells is characterized by its conspicuous, vertically oriented, bouton aggregates. The efferent synaptic relationships established by these terminal formations were investigated by electron microscopy of Golgi preparations after gold toning and deimpregnation. In all cases examined from layers II and III of cat...

This chapter addresses two questions related to the circuitry of the visual cortex on the basis of Golgi observations. The first is: What types of neurons with intracortical axons can be contacted by geniculo-cortical fibers? The second is: How are these fibers three-dimensionally arranged in layer IV of the primary visual cortex? The Golgi method...