Dynamic Involvement of Striatal NG2-glia in L-DOPA Induced Dyskinesia in Parkinsonian Rats: Effects of Doxycycline.

SUMMARY STATEMENT: NG2-glia alters its dynamics in response to L-DOPA-induced dyskinesia. In these animals, striatal NG2-glia density was reduced with cells presenting activated phenotype while doxycycline antidyskinetic therapy promotes a return to NG2-glia cell density and protein to a not activated state.

Decoding astrocyte heterogeneity: New tools for clonal analysis.

The importance of astrocyte heterogeneity came out as a hot topic in neurosciences especially over the last decades, when the development of new methodologies allowed demonstrating the existence of big differences in morphological, neurochemical and physiological features between astrocytes. However, although the knowledge about the biology of astrocytes is increasing rapidly, an important characteristic that remained unexplored, until the last years, has been the relationship between astrocyte lineages and cell heterogeneity. To fill this gap, a new method called StarTrack was recently developed, a powerful genetic tool that allows tracking astrocyte lineages forming cell clones. Using StarTrack, a single astrocyte progenitor and its progeny can be specifically labeled from its generation, during embryonic development, to its final fate in the adult brain. Because of this specific labeling, astrocyte clones, exhibiting heterogeneous morphologies and features, can be easily analyzed in relation to their ontogenetic origin. This review summarizes how astrocyte heterogeneity can be decoded studying the embryonic development of astrocyte lineages and their clonal relationship. Finally, we discuss about some of the challenges and opportunities emerging in this exciting area of investigation.

Peripheral contributions to olfactory bulb cell populations (migrations towards the olfactory bulb).

The olfactory system represents one of the most suitable models to study interactions between the peripheral and central nervous systems. The developing olfactory epithelium (olfactory placode and pit) gives rise to several cell populations that migrate towards the telencephalic vesicle. One of these cell populations, called the Migratory Mass (MM), accompanies the first emerging olfactory axons from the olfactory placode, but the fate of these cells and their contribution to the Olfactory Bulb (OB) populations has not been properly addressed. To asses this issue we performed ultrasound-guided in utero retroviral injections at embryonic day (E) 11 revealing the MM as an early source of Olfactory Ensheathing Cells in later postnatal stages. Employing a wide number of antibodies to identify the nature of the infected cells we described that those cells generated within the MM at E11 belong to different cell populations both in the mesenchyma, where they envelop olfactory axons and express the most common glial markers, and in the olfactory bulb, where they are restricted to the Olfactory Nerve and Glomerular layers. Thus, the data reveal the existence of a novel progenitor class within the MM, potentially derived from the olfactory placode which gives rise to different neural cell population including some CNS neurons, glia and olfactory ensheathing cells.

[Protocortex versus protomap: a perspective from the olfactory bulb]. / Protocórtex frente a protomapa: una perspectiva desde el bulbo olfativo.

INTRODUCTION AND AIM: The olfactory sensory system is a unique model for the research of guidance and connectivity of growing axons. During development, the olfactory epithelium, the olfactory bulb and the olfactory cortex differentiate several cell types and extend projection axons. Because there is a close relationship between these three structures, we ask the question as to whether establishment of the olfactory bulb central projections can proceed independently of the arrival of the olfactory sensory afferents. This raises another more general question: is establishment of afferent connections necessary to awake a developmental program in target cells?. DEVELOPMENT: The initial establishment of the olfactory bulb central projections occurs independently of the arrival of the olfactory axons from the olfactory epithelium, which reinforces the idea that cortical regions are already patterned before migration of newborn neurons, at least for the olfactory bulb and maybe for the entire brain. This implies a strict intrinsic molecular control of the distinct olfactory structures, independent one of each other. CONCLUSIONS: How then, do axonal projections find their correct way within the brain? Contact-mediated mechanisms and chemotropic molecules cooperate to fix their position in the telencephalon, prevent bulbar axons from invading structures other than the olfactory cortex and, at the same time, stimulate axonal branching in an orchestra of both, attractive/promoting and repulsive/inhibiting signals. At later stages, the mature appearance of the olfactory bulb will be completed and refined.

Protocórtex frente a protomapa: una perspectiva desde el bulbo olfativo / Protocortex versus protomap: a perspective from the olfactory bulb

Introducción y objetivo. El sistema olfativo es un modelo extraordinario para la investigación de la guía y la conectividad del crecimiento axonal. Durante el desarrollo, el epitelio olfativo, el bulbo olfativo y la corteza olfativa diferencian varios tipos celulares, y extienden sus proyecciones axonales. Dado que hay una relación estrecha entre estas tres estructuras, nos hacemos la siguiente pregunta: ¿es necesaria la llegada de las aferencias sensoriales procedentes del epitelio olfativo para iniciarse la formación de las proyecciones centrales del bulbo olfativo? Esto nos lleva a otra pregunta más general: ¿es necesario el establecimiento de conexiones aferentes para inducirse un programa de desarrollo en las células diana? Desarrollo. El establecimiento inicial de las proyecciones centrales del bulbo olfativo ocurre independientemente de la llegada de los axones del epitelio olfativo, lo que refuerza la idea de que las diferentes regiones corticales se predeterminan ya antes de la migración de neuronas posmitóticas, por lo menos en el caso del bulbo olfativo. Esto implica un control molecular, intrínseco y estricto de las distintas estructuras del sistema olfativo. Conclusiones. Entonces, ¿cómo encuentran las proyecciones axonales su correcta localización dentro del cerebro? Mecanismos por contacto y moléculas quimiotrópicas cooperan para fijar su posición en el telencéfalo, y evitan que los axones del bulbo invadan otras estructuras diferentes a la corteza olfativa. Al mismo tiempo, estimulan la formación de colaterales axónicas, en una orquesta de señales atrayentes/permisivas y repulsivas/inhibidoras. En etapas posteriores del desarrollo, se completará la apariencia madura del bulbo olfativo (AU)

Introduction and aim. The olfactory sensory system is a unique model for the research of guidance and connectivity of growing axons. During development, the olfactory epithelium, the olfactory bulb and the olfactory cortex differentiate several cell types and extend projection axons. Because there is a close relationship between these three structures, we ask the question as to whether establishment of the olfactory bulb central projections can proceed independently of the arrival of the olfactory sensory afferents. This raises another more general question: is establishment of afferent connections necessary to awake a developmental program in target cells? Development. The initial establishment of the olfactory bulb central projections occurs independently of the arrival of the olfactory axons from the olfactory epithelium, which reinforces the idea that cortical regions are already patterned before migration of newborn neurons, at least for the olfactory bulb and maybe for the entire brain. This implies a strict intrinsic molecular control of the distinct olfactory structures, independent one of each other. Conclusions. How then, do axonal projections find their correct way within the brain? Contact-mediated mechanisms and chemotropic molecules cooperate to fix their position in the telencephalon, prevent bulbar axons from invading structures other than the olfactory cortex and, at the same time, stimulate axonal branching in an orchestra of both, attractive/promoting and repulsive/inhibiting signals. At later stages, the mature appearance of the olfactory bulb will be completed and refined (AU)

The olfactory bulb as an independent developmental domain.

The olfactory system is a good model to study the mechanisms underlying guidance of growing axons to their appropriate targets. The formation of the olfactory bulb involves differentiation of several populations of cells and the initiation of the central projections, all under the temporal and spatial patterns of gene expression. Moreover, the nature of interactions between the olfactory epithelium, olfactory bulb and olfactory cortex at early developmental stages is currently of great interest. To explore these questions more fully, the present review aims to correlate recent data from different developmental studies, to gain insight into the mechanisms involved in the specification and development of the olfactory system. From our studies in the pax6 mutant mice (Sey(Neu)/Sey(Neu)), it was concluded that the initial establishment of the olfactory bulb central projections is able to proceed independently of the olfactory sensory axons from the olfactory epithelium. The challenge that now remains is to consider the validity of the olfactory bulb as an independent development domain. In the course of evaluating these ideas, we will review the orchestra of molecular cues involved in the formation of the projection from the OB to the olfactory cortex.

Further studies on cortical tangential migration in wild type and Pax-6 mutant mice.

In this study we present new data concerning the tangential migration from the medial and lateral ganglionic eminences (MGE and LGE) to the cerebral cortex during development. We have used Calbindin as a useful marker to follow the itinerary of tangential migratory cells during early developmental stages in wild-type and Pax-6 homozygous mutant mice. In the wild-type mice, at early developmental stages, migrating cells advance through the intermediate zone (IZ) and preplate (PP). At more advanced stages, migrating cells were present in the subplate (SP) and cortical plate (CP) to reach the entire developing cerebral cortex. We found that, in the homozygous mutant mice (Pax-6(Sey-Neu)/Pax-6(Sey-Neu)), this tangential migration is severely affected at early developmental stages: migrating cells were absent in the IZ, which were only found some days later, suggesting that in the mutant mice, there is a temporal delay in tangential migration. We have also defined some possible mechanisms to explain certain migratory routes from the basal telencephalon to the cerebral cortex. We describe the existence of two factors, which we consider to be essential for the normal migration; the first one is the cell adhesion molecule PSA-NCAM, whose role in other migratory systems is well known. The second factor is Robo-2, whose expression delimits a channel for the passage of migratory cells from the basal telencephalon to the cerebral cortex.

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 Sey(Neu)/Sey(Neu) 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.

Development of the mammillothalamic tract in normal and Pax-6 mutant mice.

The mammillary bodies represent important relay stations for one of the major neuronal circuits in the brain: the limbic circuit. Mammillary projections traveling through the principal mammillary tract are established early during development, forming the mammillotegmental bundle, which appears fully developed by embryonic day 15 (E15). The mammillothalamic tract develops later, around E17-E18, forming a compact system of collateral fibers originating from the principal mammillary tract and reaching the thalamus by E20. The Pax-6 gene is expressed in various regions of the developing brain, among which the border separating the ventral thalamus from the dorsal thalamus, known as the zona limitans intrathalamica, is especially significant. In this report, the development of the efferent mammillary system of fibers was studied in wild type and Pax-6 mutant mice by using carbocyanine tracers and Golgi preparations. In mutant mice, the mammillotegmental bundle developed normally; however, the mammillothalamic tract was missing. By using anti-Pax-6 antibodies in wild type mice, the existence of an immunoreactive cell cluster is described surrounding the bifurcation point of the principal mammillary tract. The results of this study suggest that there is a correlation of these cells with a particular type of Golgi impregnated neuron.

Central olfactory structures in Pax-6 mutant mice.

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.

Protein synthesis inhibitors delay transneuronal death in the piriform cortex of young adult rats.

It has been demonstrated that apoptotic cell death is an active process that is dependent on RNA and protein synthesis. The question remains as to whether neuronal death in adult, mammalian brains can also be demonstrated in vivo to be dependent on protein synthesis. To address this question we have analysed transneuronal death in the piriform (olfactory) cortex. Following unilateral olfactory bulb ablation in young adult rats, layer IIa of the piriform cortex undergoes rapid degeneration, that commences 12 h after ablation and that is almost complete at 48 h. In order to block protein synthesis, three to six subcutaneous injections of the short acting protein synthesis inhibitor anisomycin, were given at 2 h intervals beginning just before the ablation of the olfactory bulb. In other cases a single injection of the long acting protein synthesis inhibitor emetine were made intracerebrally just before or after olfactory bulb ablation. The number of dying cells was then counted in sections through the rostrocaudal extent of the piriform cortex. Both anisomycin and emetine injections markedly reduced the number of pyknotic cells in layer IIa of the piriform cortex after olfactory bulb ablation. The effect of anisomycin was dose-dependent, near lethal doses leading to an almost complete absence of cell death (six injections of 100 mg/kg). As the doses of anisomycin were reduced, more dying cells were observed. Emetine was only effective at near lethal doses (10 mg/kg) and showed a greater capacity to reduce the levels of cell death when injected into structures near the piriform cortex (e.g., accumbens nucleus) than when injected into more distant structures. To further confirm that the cell death observed was due to apoptosis, we analysed sections by tunel staining to demonstrate DNA fragmentation. We found that tunel-positive cells were also always pyknotic, one of the landmarks of apoptosis. The appearance of pyknotic cells labelled by the tunel method demonstrated that the dying cells in the piriform cortex did indeed undergo apoptosis.

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 beta-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 (immature 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 the Escherichia coli lac-Z gene.

Dynamics of cell migration from the lateral ganglionic eminence in the rat.

From previous developmental studies, it has been proposed that the neurons of the ventrolateral cortex, including the primary olfactory cortex, differentiate from progenitor cells in the lateral ganglionic eminence. The objective of the present study was to test this hypothesis. The cells first generated in the forebrain of the rat migrate to the surface of the telencephalic vesicle by embryonic day (E) 12. Using [3H]thymidine, we found that most of these cells contributed to the formation of the deep layer III of the primary olfactory cortex. To study the migratory routes of these cells, we made localized injections of the carbocyanine fluorescent tracers Dil and DiA into various parts of the lateral ganglionic eminence in living embryos at E12-E14 and subsequently maintained the embryos in a culture device for 17-48 hr. After fixation, most migrating cells were located at the surface of the telencephalic vesicle, whereas others were seen coursing tangentially into the preplate. Injections made at E13 and in fixed tissue at E15 showed that migrating cells follow radial glial fibers extending from the ventricular zone of the lateral ganglionic eminence to the ventrolateral surface of the telencephalic vesicle. The spatial distribution of radial glial fibers was studied in Golgi preparations, and these observations provided further evidence of the existence of long glial fibers extending from the ventricular zone of the lateral ganglionic eminence to the ventrolateral cortex. We conclude that cells of the primary olfactory cortex derive from the lateral ganglionic eminence and that some early generated cells migrating from the lateral ganglionic eminence transgress the cortico-striatal boundary entering the preplate of the neocortical primordium.

Early onset of the rat olfactory bulb projections.

Using the fluorescent carbocyanine tracer DiI, we examined in detail the early development of the projections emanating from the rat olfactory bulb. The study commenced at embryonic day 13 when the first fibres can be detected and ended at embryonic day 20, when all major fibre systems have been established. The first axons arising from the prospective olfactory bulb area are seen at embryonic day 13. Labelled fibres are provided with elaborate axonal growth cones advancing through the ventrolateral part of the telencephalic vesicle. At embryonic day 14, while the main fibre tract has not developed much further, some isolated fibres are located quite distally from the prospective olfactory bulb. These early fibres apparently course within a narrow cell-free space that extends caudally along the ventrolateral part of the telencephalic vesicle. At embryonic day 15, a number of labelled fibres form a compact bundle, corresponding to the lateral olfactory tract, that ultimately reaches the prospective primary olfactory cortex. The fibres do not stop growing, but continue to extend caudally at embryonic day 17. The results of this study provide new information on the development of axonal tracts in the olfactory system. We show that the olfactory tract projection develops earlier than the morphological appearance of the olfactory bulbs. This suggests that the early development of olfactory projections might not depend on the arrival of the olfactory epithelium axons and thus, could be governed by factors intrinsic to the neurons and/or cues present in the target environment.

Time of origin and early fate of preplate cells in the cerebral cortex of the rat.

The time of origin, development, and morphology of neurons originating before the establishment of the cortical plate (preplate cells) were studied in the telencephalic vesicle of the rat from the embryonic day 10 (E10) to E15. The first postmitotic neurons settle superficial to the ventricular zone in the preplate at E12. We have observed mitotic figures in the preplate at E12 and E13, and regarded them as a possible source of cortical neurons outside the ventricular zone. It is suggested that these cells may correspond to Cajal-Retzius cells. The appearance of cells before the formation of the cortical plate was studied in animals injected with tritiated thymidine at E12 and E13 and correlated with observations made using an antibody to neuron specific class III beta-tubulin that becomes detectable immediately after the last mitotic division. Immunoreactive cells detected with this antibody concentrate at E12 immediately below the pial surface constituting the preplate. Radially oriented cells, most probably subplate neurons, were also present spanning the full thickness of the neuroepithelium, suggesting that the preplate contains different cell classes. We present a working hypothesis that explains the sequence of developmental events during the early phases of cortical histogenesis.

The telencephalic vesicles are innervated by olfactory placode-derived cells: a possible mechanism to induce neocortical development.

During early embryonic development, the olfactory placode is the source of different cell types migrating toward the telencephalic vesicle. Among these cell types are the ensheathing cells, the luteinizing hormone-releasing hormone-producing cells and the olfactory marker protein-immunoreactive cells. We have identified a novel group of olfactory placode-derived migratory cells using an antibody against beta-tubulin to label neurons and acetylcholinesterase histochemistry to label posmitotic cells. In this paper we describe the morphology, migration and fate of this novel group of cells. The first neurons detected in the rostral prosencephalon with acetylcholinesterase and anti-beta-tubulin antibody are localized in the olfactory placodes at embryonic day 11 in the rate. At embryonic day 12, anti-beta-tubulin antibody-positive cells were observed in the mesenchymal tissue between the olfactory pit and the rostral pole of the telencephalic vesicle. Anti-beta-tubulin antibody-positive cells were seen running superficially over the pial (dorsal) side of the telencephalic vesicle at embryonic day 13. The majority of these cells have a bipolar profile with short leading and trailing processes, suggesting that they are migratory elements. However, some of these cells showed elaborate processes extending for quite long distances, overlying the pial surface of the telencephalic vesicle. A mass of cells extending over the telencephalic vesicle from the developing olfactory epithelium were observed at embryonic day 13 using acetylcholinesterase histochemistry. Some of these acetylcholinesterase-positive cells were identified as neurons with the specific neuronal marker anti-beta-tubulin antibody. On embryonic day 12, neurons from the olfactory epithelium send axonal fibers toward the telencephalic vesicles. Most of these fibers spread over the anteroventral pole of the vesicles but others entered deep into the telencephalon, reaching the germinal ventricular zone. We also show that fibers run rostrocaudally over the surface of the telencephalic vesicles. We suggest that these cells and fibers, apparently originating in the olfactory placode and migrating through non-conventional routes, might play a significant role in the earliest stages of telencephalic vesicle development.

Persistence of early-generated neurons in the rodent subplate: assessment of cell death in neocortex during the early postnatal period.

In the rat, the deepest neocortical layer forms a conspicuous cell band known as layer Vlb. Cells in layer Vlb are among the first to differentiate, and it has been regarded as an homolog to the subplate of primates and carnivores. Cell death has been considered a universal feature of subplate cells. In order to assess the validity of this assertion, we examined the sequence of generation and the extent of cell death in layer Vlb. This was achieved using injections of 3H-thymidine and two methods for the direct visualization of apoptotic figures. Single injections of 3H-thymidine were performed between E12 and E15 (E0 is the day of insemination), and brains were examined at different postnatal ages between P1 and P63. The number of heavily labeled cells were counted in layer Vlb in six standard, equally spaced coronal sections in each brain. Single injections at E12 labels about 3% of the entire population of layer Vlb cells, 17% at E13, 30% at E14, and < 1% at E15. Our results indicate that the absolute number of heavily labeled cells in layer Vlb remains constant. The analysis of variance (one-way ANOVA) showed that the difference among the group means was not significant from P1 to P63 after injections at either E12, E13, or E14. In order to confirm these results, we evaluated the distribution of pyknotic (apoptotic) cell bodies in the neocortex. Apoptotic cells were visualized in Nissl preparations and by histochemical staining using an in situ apoptosis detection kit. The analysis was performed in rats from E18 to P15. Both methods gave comparable results. We found that the amount of cell death in layer Vlb is neither particularly prominent nor significantly different from that which occurs in the remaining neocortical layers, apart from layer II and in the white matter of the corpus callosum. We conclude that neuronal death does not play any significant role in the rodent subplate.

Action of a diffusible target-derived chemoattractant on cortical axon branch induction and directed growth.

Cortical axons innervate their brainstem target, the basilar pons, by the initiation and extension of collateral branches interstitially along their length. To address whether a diffusible pons-derived chemoattractant controls these events, we used cocultures in collagen matrices and time-lapse microscopy. Pontine explants enhanced by 5-fold the de novo initiation of transient branches along cortical axons; most branches were directed toward pons. Of the branches extended toward pons, 2%-3% were stabilized; those extended away were not. Pontine explants also enhanced the stable bifurcation of growth cones and prompted directional changes by growth cone turning and collateral extension. These effects were distance dependent and mimicked by pons-conditioned medium. This evidence indicates that the pons activity promotes branch initiation interstitially along cortical axons, a novel property for a chemoattractant, and provides a directional cue for their growth. These findings suggest that the pons chemoattractant serves as a diffusible target-recognition molecule.

Neuronal and synaptic composition of the mediodorsal thalamic nucleus in the rat: a light and electron microscopic Golgi study.

The distribution and dendritic domain of neurons in each segment of the mediodorsal thalamic nucleus (MD) have been studied in the rat with the Golgi technique. In addition, a combined Golgi method-electron microscopic (Golgi-EM) study was undertaken to determine the distribution of morphologically distinct synapse types along the dendrites of individual identified neurons in MD. All the subdivisions or "segments" of MD (medial, central, lateral) contained both stellate and fusiform cells. The dendritic domain of both types of cells was predominantly restricted to the same segment of MD that contained the cell body of the neuron. Typical stellate neurons were found near the center of each segment, with radiating dendrites that extended to but not across the boundaries of the segment. Fusiform cells were usually located close to the segmental or nuclear boundaries and tended to have dendrites oriented parallel to those borders; again, the dendrites tended not to extend across borders between segments or at the outer edge of MD. In the medial segment of MD many fusiform cells had especially bipolar dendritic configurations, generally with a dorsoventral orientation. Because no small neurons were identified that might correspond to thalamic interneurons, all the impregnated cells in MD are presumed to be thalamocortical projection neurons. These results indicate that cells and their major dendrites are confined to a single segment of MD, with little dendritic overlap across segmental or nuclear borders. The segments of MD may therefore be considered to be relatively independent subnuclei. The distribution of the four types of synapses previously identified in MD (Kuroda and Price, J. Comp. Neurol., 303:513-533, 1991) was determined along several identified dendrites studied with the Golgi-EM method. Primary dendrites were contacted mostly by large axon terminals, including both large, round vesicle (LR) terminals and large, pleomorphic vesicle (LP) terminals, as well as a few small to medium sized terminals with pleomorphic vesicles (SMP). No small terminals with round vesicles (SR terminals) were observed to make synapses with primary dendrites. Secondary and tertiary dendrites received synapses from all types of axon terminals. Higher order dendrites were contracted predominantly by SR boutons, but they also carried some LR and SMP terminals. In addition, SMP boutons were often found to form symmetric contacts with cell somata.

Neuroglial arrangements in the olfactory glomeruli of the hedgehog.

The olfactory glomeruli represent morphological and functional units in which olfactory information is processed in specialized synaptic arrangements established between the central processes of sensory neurons, whose cell bodies are located in the olfactory epithelium, and the terminal (intraglomerular) portions of the dendrites of periglomerular, tufted, and mitral cells. The olfactory glomeruli are surrounded by distinctive glial formations in which the peripheral glia interacts with the central glia. We have studied the morphology and organization of neuroglial cells in the layer of olfactory nerves and the glomerular layer of the olfactory bulb in the insectivorous hedgehog (Erinaceus europaeus) with the electron microscope, Golgi method, and immunohistochemistry by using antibodies to glial fibrillary acidic protein (GFAP) and "rip," a monoclonal antibody that stains oligodendrocytes and their processes in the rat (Friedman et al.: Glia 2:380-390, '89). The peripheral glia is represented by a special category of cells that are closely related to astrocytes and known as sheathing cells. They accompany olfactory axons to their entrance in the glomeruli where they interact with the central glia, represented by astrocytes and oligodendrocytes. The sheathing cells typically display indented nuclei and protoplasmic expansions forming laminar processes wrapping several axons together. Astrocytes surrounding the glomerular neuropil belong to the velate type. They display numerous sheet-like processes enveloping dendritic segments and periglomerular cell bodies. Oligodendrocytes were found surrounding the glomeruli and at the interstices separating different glomeruli. Myelinated dendritic segments and cell bodies were found surrounding the olfactory glomeruli. These myelin coverings probably derive from oligodendrocytes. Together with the astrocytic lamellar expansions, they provide a rigid structural support that contributes to the segregation of group of different cells while remaining relatively isolated from other influences at the periphery of the glomeruli.