Juan A. De Carlos

Time frame of mitral cell development in the mice olfactory bulb.

Along with tufted cells, mitral cells are the principal projection neurons in the olfactory bulb (OB). During the development of the OB, mitral cells migrate from the ventricular zone to the intermediate zone, where they begin to send axons along the lateral olfactory tract (LOT) to the cortical olfactory zones. Subsequently, they lose their tangential orientation, enabling them to make contact with the axons of the olfactory sensory neurons (OSN) that innervate the whole OB. Here, we investigated the distinct morphological features displayed by developing mitral cells and analyzed the relationship between the changes undertaken by these neurons and the arrival of the OSN axons. Immunostaining for specific markers of developing axons and dendrites, coupled with the use of fluorescent tracers, revealed the morphological changes, the continuous reorientation, and the final refinement that these cells undergo. We found that some of these changes are dependent on the arrival of the OSN axons. Indeed, we identified three main chronological events: 1) newly generated neurons become established in the intermediate zone and project to the LOT; 2) the cells reorient and spread their dendrites at the same time as OSN axons penetrate the OB (this is a sensitive period between embryonic day (E)15–16, in which the arrival of afferents establishes a spatial and temporal gradient that facilitates protoglomerulus and glomerulus formation); and 3) final refinement of the radially orientated cells to adopt a mature morphology. These results suggest that both afferent inputs and intrinsic factors participate to produce the well‐defined sensory system. J. Comp. Neurol. 495:529–543, 2006.

Origins and migratory routes of murine Cajal-Retzius cells.

The first layer that appears in the cortical neuroepithelium, the preplate, forms in the upper part of the cortex immediately below the pial surface. In mice, this layer exists between embryonic days (E) 10 and 13, and it hosts different cell populations. Here, we have studied the first cell population generated in the preplate, the Cajal‐Retzius cells. There is considerable confusion regarding these cells with respect to both their site of generation and the migratory routes that they follow. This perhaps is due largely to the different opinions that exist regarding their characterization. We have studied the site of origin of these cells, their migratory routes, and the molecular markers that may distinguish them by injecting tracers into early embryos, culturing them in toto for 24 hours, and then performing immunohistochemistry. We found that the Cajal‐Retzius cells are most likely generated in the cortical hem by comparing with other cortical or extracortical origins. These cells are generated mainly at E10 and E11, and they subsequently migrate tangentially to cover the whole cortical mantle in 24 hours. From their site of origin in the medial wall of the telencephalon, they spread in a caudorostral direction, following an oblique migratory path toward the lateral part of the neuroepithelium. Prior to the splitting of the preplate, a percentage of the Cajal‐Retzius cells that can be distinguished by the expression of reelin do not contain calretinin. Furthermore, there were no early‐migrating neurons that expressed calbindin. J. Comp. Neurol. 500:419–432, 2007.

Evidence for intrinsic development of olfactory structures in Pax-6 mutant mice.

It has been reported that the arrival of primary olfactory axons is required to induce the development of the olfactory bulb (OB). On the other hand, the SeyNeu/SeyNeu mutant mouse (Small eye) has been previously described as a model for the absence of olfactory bulbs, owing to the lack of olfactory epithelium (OE). In the present report, we take advantage of this mutant and study a neural structure in the rostral pole of the telencephalon that phenotypically resembles the prospective OB. We named this formation olfactory bulb‐like structure (OBLS). We also report the occurrence, in the mutants, of small epithelial vesicles in the malformed craneofacial pits, resembling an atrophic OE, although a mature olfactory nerve was not identified. Axonal tracing, birthdating, immunohistochemistry, and in situ hybridization using antibodies and probes expressed in the olfactory system, indicated that two distinct structures observed in the OBLS correspond to the main and accessory olfactory bulbs of the control mouse. We propose that the OBLS has developed independently of the external influences exerted by the olfactory nerve. The presence of a prospective OB in the mutants, without intervening olfactory fibers, suggests that intrinsic factors could define brain territories even in absence of the proper afferent innervation. The intrinsic mechanisms and environmental cues in the telencephalon could be sufficient to promote axonogenesis in the projection neurons of the OB and guide their axons in a lateral prospective tract, in the absence of olfactory axons. J. Comp. Neurol. 428:511–526, 2000.

A neuronal migratory pathway crossing from diencephalon to telencephalon populates amygdala nuclei

Neurons usually migrate and differentiate in one particular encephalic vesicle. We identified a murine population of diencephalic neurons that colonized the telencephalic amygdaloid complex, migrating along a tangential route that crosses a boundary between developing brain vesicles. The diencephalic transcription factor OTP was necessary for this migratory behavior.

A historical reflection of the contributions of Cajal and Golgi to the foundations of neuroscience

In 1906, the Spaniard Santiago Ramón y Cajal and the Italian Camillo Golgi shared the Nobel Prize in Physiology or Medicine, in recognition of their work on the structure of the nervous system. Although both were well-known scientists who had made a large number of important discoveries regarding the anatomy of the nervous system, each defended a different and conflicting position in relation to the intimate organization of the grey matter that makes up the brain. In this communication we will review the importance of Cajals studies using the method of impregnation discovered by Golgi, as well as the relevant studies carried out by Golgi, the concession of the Nobel Prize and the events that occurred during the Nobel conferences. In summary, we will précis the important contribution of both scientists to the founding of modern Neuroscience.

EARLY OLFACTORY FIBER PROJECTIONS AND CELL MIGRATION INTO THE RAT TELENCEPHALON

The formation and development of primary olfactory axons was studied in the rat embryo using acetylcholinesterase histochemistry, immunocytochemistry for neuron‐specific β‐tubulin (TuJ1) and growth associated protein 43 (GAP43), and a fluorescent tracer DiI. Olfactory axons extend from the olfactory receptor neurons localized in the olfactory epithelium. These fibers grow to reach and enter the olfactory bulbs, where they form the first relay and integrative synaptic station in the olfactory system: the olfactory glomerulus. In this communication we address the development of primary olfactory fibers: first from the olfactory placode and later from the olfactory epithelium. Olfactory fibers enter the olfactory bulbs apparently in a disordered manner but soon arrange themselves in hook shaped aggregates of fibers, with many boutons (inmature synaptic terminals), to form the glomeruli. We detected this kind of structure for the first time at embryonic day 16. The olfactory receptor cells are usually anchored in the basal lamina of the olfactory epithelium but some of them, after reaching their targets, lose their epithelial attachment, leave the olfactory epithelium and migrate to and enter the olfactory bulbs. The traffic of cells between the olfactory epithelium and the brain lasts late into embryonic development. We describe four types of migratory mechanism used by different populations of cells to reach their targets in the telencephalic vesicle and propose the existence of migrating cells that enter the telencephalon. These data were corroborated by injections into the olfactory epithelium a of murine retrovirus carrying theEscherichia coli lac‐Z gene.

Different astroglia permissivity controls the migration of olfactory bulb interneuron precursors

The rostral migratory stream (RMS) is a well defined migratory pathway for precursors of olfactory bulb (OB) interneurons. Throughout the RMS an intense astroglial matrix surrounds the migratory cells. However, it is not clear to what extent the astroglial matrix participates in migration. Here, we have analyzed the migratory behavior of neuroblasts cultured on monolayers of astrocytes isolated from areas that are permissive (RMS and OB) and nonpermissive (cortex and adjacent cortical areas) to migration. Our results demonstrate robust neuroblast migration when RMS‐explants are cultured on OB or RMS‐astrocytes, in contrast to their behavior on astroglia derived from nonpermissive areas. These differences, mediated by astrocyte‐derived nonsoluble factors, are related to the overexpression of extracellular matrix and cell adhesion molecules, as revealed by real‐time qRT‐PCR. Our results show that astroglia heterogeneity could play a significant role in migration within the RMS and in cell detachment in the OB.

Central Olfactory Structures in Pax-6 Mutant Micea

ABSTRACT: During the development of the olfactory system, cells located in the olfactory placode/olfactory pit send their axons toward the rostral part of the telencephalic vesicles (TVs). Some of these enter the TV inducing the formation of the olfactory bulbs (OBs), whereas, mitral and tufted cell axons form the lateral olfactory tract (LOT). Our recent studies have shown that the beginning of the central olfactory projections is independent of the arrival of olfactory receptor neuron (ORN) axons to the TV. Here we have used the mouse carrying a mutation in the Pax‐6 gene to study whether the nasal olfactory structures intervene in the formation of central olfactory structures. This mutant as well as lacking a nose and eyes, is reported to lack olfactory epithelium and OB. However, we have found an ovoid cellular structure localized in the rostral part of the brain, and some cells in this structure project axons toward the piriform cortex forming a presumptive LOT. We conclude that the referred structure is an OB, which fails to develop because the mutation in the Pax‐6 gene affects the formation of nasal structures. As such, fibers of the ORNs are necessary for the protrusion and layered formation of the OB, but these inputs are not necessary for the establishment of the central olfactory projections.

Evolution and development of the mammalian cerebral cortex.

Comparative developmental studies of the mammalian brain can identify key changes that can generate the diverse structures and functions of the brain. We have studied how the neocortex of early mammals became organized into functionally distinct areas, and how the current level of cortical cellular and laminar specialization arose from the simpler premammalian cortex. We demonstrate the neocortical organization in early mammals, which helps to elucidate how the large, complex human brain evolved from a long line of ancestors. The radial and tangential enlargement of the cortex was driven by changes in the patterns of cortical neurogenesis, including alterations in the proportions of distinct progenitor types. Some cortical cell populations travel to the cortex through tangential migration whereas others migrate radially. A number of recent studies have begun to characterize the chick, mouse and human and nonhuman primate cortical transcriptome to help us understand how gene expression relates to the development and anatomical and functional organization of the adult neocortex. Although all mammalian forms share the basic layout of cortical areas, the areal proportions and distributions are driven by distinct evolutionary pressures acting on sensory and motor experiences during the individual ontogenies.

Origin of the Cortical Layer I in Rodents

Using birthdating techniques, we have studied when cells that settle in the marginal zone (future layer 1) of the cortical neuroepithelium are generated in developing rat embryos. The majority of marginal zone cells are generated at embryonic day 12 (E12), E13 and E14, although some cells generated later can incorporate into this stratum after the cortical plate forms. The nature and the origin of the cell populations that colonize the preplate/marginal zone was studied by means of immunohistochemistry using cell markers for γ-amino butyric acid (GABA), reelin and the calcium binding proteins calretinin and calbindin. At early stages of development, the preplate is formed by Cajal-Retzius cells, subplate cells, subpial granular layer cells, some interneurons and some glial cells. With the arrival of the cortical plate cells, the subplate cells descend to occupy the stratum below. Layer 1 cells are of diverse origin as some of them are generated in the ventricular zone of the cortical neuroepithelium, whereas other cell populations come from extracortical regions such as the olfactory placode or the ganglionic eminences of the basal telencephalon. The predominant cell type in the marginal zone is the Cajal-Retzius cell, which expresses reelin and calretinin, and is probably generated in the cortical neuroepithelium. These cells can be readily distinguished from cells that come from the ganglionic eminences as these later populations mainly express GABA and calbindin. Finally, our results suggest that the cells of the subpial granular layer might be generated in the rostral pole of the lateral ganglionic eminences.