Immunohistochemical localization of neurocan and L1 in the formation of thalamocortical pathway of developing rats

We used immunohistochemistry to examine possible molecular interactions between the subplate and growing thalamocortical axons in rat fetuses. In the cortical anlage of embryonic day 16 (E16), the subplate first appeared below the cortical plate. Among chondroitin sulfate proteoglycans, phosphacan was uniformly distributed throughout the cortical wall, whereas neurocan was localized only in the subplate at E16. Neural cell adhesion molecules, NCAM-H, TAG-1, and L1, were detected in the cortical anlage. Both cortical neurons and growing axons were diffusely immunopositive for NCAM-H, and TAG-1 immunoreactivity was found on immature neurons and cortical efferent axons but not on thalamocortical axons. L1 immunoreactivity was specifically localized on the growing thalamocortical axons. When the locations of neurocan and L1 were compared in the developing cortex, L1-bearing axons were found to extend to neurocan-immunopositive regions; neurocan immunoreactivity was intense in the subplate at E16, when small numbers of L1-immunoreactive thalamocortical axons began to invade the cortex. At E17, many L1-positive axons were observed in the subplate that expressed neurocan specifically. Double immunostaining showed that L1-positive axons and neurocan immunoreactivity overlapped in the subplate at E17. After E18, neurocan expression gradually extended to the lower part of the cortical plate; it extended to the entire cortex by E21, 1 day before birth. By E21, L1-bearing axons had invaded the lower part of the cortical plate. The present study demonstrated that the neurocan expression precedes growth of L1-bearing thalamocortical afferent fibers. Because neurocan can bind to L1 molecule in vitro, these results suggest that neurocan and L1 play some important roles in pathfinding of the thalamocortical afferent fibers during rat corticogenesis. J. Comp. Neurol. 382:141-152, 1997. © 1997 Wiley-Liss, Inc.     

The first thalamocortical synapses are made in the cortical plate in the developing visual cortex of the wallaby (Macropus eugenii)

The time course of development and laminar distribution of thalamocortical synapses in the visual cortex of the marsupial mammal the wallaby (Macropus eugenii) has been studied by electron microscopy from the time of afferent ingrowth to the appearance of layer 4, the main target for thalamic axons. Axons were labeled from the thalamus by a fluorescent carbocyanine dye in fixed tissue or by transneuronal transport of horseradish peroxidase conjugated to wheat germ agglutinin from the eye. Thalamic axons first reached the cortex 2 weeks after birth and grew into the developing cortical plate without a waiting period in the subplate. The first thalamocortical synapses were detected 2 weeks later solely throughout the loosely packed zone of the cortical plate, where layer 6 cells previously have been shown to reside. As the thickness of the cortex increased with age, thalamocortical synapses were increasingly prevalent in the loosely packed zone of the cortical plate. With the appearance of layer 4, thalamocortical synapses were found there as well as in the marginal zone and layer 6. There was no evidence for an early population of thalamocortical synapses in the subplate. The first synapses made by thalamic axons were in a region containing layer 6 cells, one of their normal targets in the mature cortex.     

Monoaminergic afferents to the neocortex: a developmental histofluorescence study in normal and Reeler mouse embryos

The patterns of distribution of monoaminergic (MA) afferents during early histogenesis of the neocortex of normal and Reeler mice are studied by histofluorescence microscopy. Fluorescing fibers appear rostrally in the cortex of both genotypes on the 14th embryonic day (E14), which is within 24 h of the development of the cortical plate. They are distributed to all regions of the cortex by the time of birth. Although the patterns of intracortical deployment differ in the two genotypes, the fibers appear to have homologous target structures. These are: (1) the polymorphic cells of the subplate in the depths of the normal and in the superplate near the surface of the mutant cortex; and (2) the zones of consolidation of apical dendrites of pyramidal cells: the external plexiform zone of normal and a series of intracortical plexiform planes in the mutant cortex. By contrast, the axons of this system do not branch significantly among the compactly ordered somata of pyramidal cells within the cortical plate of either genotype.     

Ultrastructural Evidence for Synaptic Interactions between Thalamocortical Axons and Subplate Neurons

Thalamic axons are known to accumulate in the subplate for a protracted period prior to invading the cortical plate and contacting their ultimate targets, the neurons of layer 4. We have examined the synaptic contacts made by visual and somatosensory thalamic axons during the transition period in which axons begin to leave the subplate and invade the cortical plate in the ferret. We first determined when geniculocortical axons leave the subplate and begin to grow into layer 4 of the visual cortex by injecting 1,1′-dioctadecyl-3, 3, 3′, 3′-tetramethyl indocarbocyanine (Dil) into the lateral geniculate nucleus (LGN). By birth most LGN axons are still confined to the subplate. Over the next 10 days LGN axons grow into layer 4, but many axons retain axonal branches within the subplate. To establish whether thalamic axons make synaptic contacts within the subplate, the anterograde tracer PHA-L was injected into thalamic nuclei of neonatal ferrets between postnatal day 3 and 12 to label thalamic axons at the electron microscope level. The analysis of the PHA-L injections confirmed the Dil data regarding the timing of ingrowth of thalamic axons into the cortical plate. At the electron microscope level, PHA-L-labelled axons were found to form synaptic contacts in the subplate. The thalamic axon terminals were presynaptic primarily to dendritic shafts and dendritic spines. Between postnatal days 12 and 20 labelled synapses were also observed within layer 4 of the cortex. The ultrastructural appearance of the synapses did not differ significantly in the subplate and cortical plate, with regard to type of postsynaptic profiles, length of postsynaptic density or presynaptic terminal size. These observations provide direct evidence that thalamocortical axons make synaptic contacts with subplate neurons, the only cell type within the subplate possessing mature dendrites and dendritic spines; they also suggest that functional interactions between thalamic axons and subplate neurons could play a role in the establishment of appropriate thalamocortical connections.     

Growth and targeting of subplate axons and establishment of major cortical pathways.

In the developing mammalian neocortex, the first postmitotic neurons form the "preplate" superficial to the neuroepithelium. The preplate is later split into a marginal zone (layer 1) and subplate by cortical plate neurons that form layers 2-6. Cortical efferent axons from layers 5 and 6 and cortical afferent axons from thalamus pass between cortex and subcortical structures through the internal capsule. Here, we identify in rats the axonal populations that establish the internal capsule, and characterize the potential role of subplate axons in the development of cortical efferent and afferent projections. The early growth of cortical efferent and afferent axons was studied using 1-1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (Dil) as an anterograde and retrograde tracer in aldehyde-fixed brains of embryonic rats. Cortical axons first enter the nascent internal capsule on embryonic day (E) 14 and originate from lateral and anterior cortex; axons from posterior cortex extend rostrally but do not yet exit cortex. The labeled axons, tipped by growth cones with complex morphologies, take a pathway deep to the preplate. Preplate neurons extend these early cortical efferents, based on the developmental stage of the cortex, and on their location and morphology. Most of these cells later occupy the subplate. Cortical plate neurons extend axons into the internal capsule by E16. En route to the internal capsule, cortical plate axons take the same path as the earlier-growing preplate axons, through the intermediate zone deep to subplate. Subplate axons reach thalamus by E16; the first cortical plate axons enter thalamus about a day later. Thalamic axons enter cortex by E16, prior to other cortical afferents. On E15, both preplate and thalamic axons reach the midpoint of the internal capsule. To determine the subcortical distribution of subplate axons, we used Dil as a retrograde tracer in aldehyde-fixed brains and fast blue and rhodamine-B-isothiocyanate as in vivo retrograde markers in neonatal rats. Tracers were injected into the superior colliculus, the principal midbrain target of layer 5 neurons, at times before, during, and after the arrival of cortical axons, or into the subcortical pathway of primary layer 5 axons at two points, the cerebral peduncle caudal to the internal capsule, and the pyramidal decussation at the junction of the hindbrain and spinal cord, at times shortly after the passing of cortical axons. In every case, the labeled neurons are confined to layer 5; subplate neurons are not labeled.(ABSTRACT TRUNCATED AT 400 WORDS)     

Axon strata of the cerebral wall in embryonic mice

The stratification of principal fiber systems affiliated with the developing neocortex has been analyzed by means of HRP tracing methods, monoamine histofluorescence and silver impregnations in mouse embryos ranging from the 15th to 16th embryonic day (E15/16) to the end of gestation (E19 = the day of birth). As early as E15/16 a fiber stratum divides the subplate and marks the inferior boundary of the developing cortex. Axons coursing in this fiber plane, termed the external sagittal stratum (ESS), include components of at least 5 identifiable systems: thalamocortical, corticothalamic, ipsilateral corticocortical, callosal and monoaminergic. The neocortical afferents of extrinsic origin, i.e., the thalamocortical, callosal and monoaminergic systems, cross the intermediate zone from their separate directions and converge upon the ESS. After a variable course through this stratum, single fibers ascend from their parent fascicles to ramify densely in the cortical subplate (CSB). Fibers of each of the extrinsic afferent systems mingle with each other and with locally arising axons within the CSB. Axons of the monoaminergic projection as well as fibers of the thalamic projection cross the cortical plate to ramify in the marginal zone. Other axons apparently of local intracortical origin course tangentially through the cortical plate. Otherwise, the cortical plate is devoid of proliferating axons at this early developmental stage. The set of observations illustrates the existence of sharply defined boundaries between axon-rich and axon-poor strata of the developing neocortex. These boundaries also compartmentalize postmigratory neurons with respect to their state of differentiation.     

Thalamocortical projections in the reeler mutant mouse

The normal radial distribution of the different neocortical cell classes is inverted in the reeler mutant mouse. The organization of the thalamocortical projection in adult reelers has been investigated by using anterograde degeneration techniques. Thalarnocortical mons follow anomalous trajectories to their target cytoarchitectonic fields in the mutant. After leaving the internal capsule, the axons ascend in sigmoid-shaped fascicles to enter a fiber stratum near the cortical surface. The axons course through this superficial stratum until they reach their target fields and then descend to terminate in deeper cortical planes. In the normal animal, by contrast, the mons course tangentially at the interface of the cortex with the subcortical white matter; upon reaching their target fields, they ascend to terminate more superficially in the cortex. Thus, in both genotypes the tangential portions of the axon trajectories pass through the polymorphic cell population. Both with respect to degree of divergence and the radial distribution of terminals within the different cytoarchitectonic fields, the thalamocortical projection in reeler, like that in the normal animal, appears to be composed of two distinct axon classes. The “class I” axons, the less-divergent system which terminates densely in two or three tiers within the cortex, are the subject of the present analysis. The “class 1” projections form an orderly cortical representation of the thalamus: Despite distortions in the topography of the reeler thalamus and cortex, both the nucleus to field relationships and the detailed topologic organization of the projection appear to be normal. The terminals of class I projections are distributed in radially segregated tiers that resemble the tiered pattern of termination as in normal mice, and there are field-specific variations in the tiers that, like those of normal mice, are systematically related to variations of cortical cytoarchitecture. Thus, similar mechanisms, which may depend in part on the interaction of ingrowing axons with specific postsynaptic cell surfaces, must determine the restricted radial distribution of thalamocortical projections in both genotypes. However, there is an abnormal mix of somata and dendrites at all radial levels of the mutant cortex, suggesting that the spatial domain of termination of thalamocortical axons may be governed to some extent by factors other than the distribution of specific prospective postsynaptic cell surfaces. The reeler mutation alters the radial but not the tangential structure of the neocortex, suggesting that these two aspects of cortical organization develop under the control of independent mechanisms. The present results suggest that despite the anomalies of radial position, normal relationships between thalamocortical afferents and distinct classes of postsynaptic elements with characteristic radial distributions are largely conserved in reeler. Certain types of aberrant connections are highly probable, however.     

Afferent and efferent connections of the striate and extrastriate visual cortex of the normal and reeler mouse

In order to analyze the role of lamination in establishing the precisely ordered connectional pattern of the neocortex, we compared the afferent and efferent connections of the visual cortical areas in normal mice with those of the mutant mouse reeler (rl). The reeler mutation causes disruption of the laminar organization of the neocortex; all classes of neurons are present but are abnormally located. The corticocortical and thalamocortical connections of visual cortical areas 17, 18a, and 18b were determined in normal and reeler mice with injections of horseradish peroxidase (HRP) or HRP conjugated with wheat germ agglutinin (HRP-WGA). The diffusion of HRP-WGA is highly restricted due to the surface binding properties of the lectin; it was particularly effective in demonstrating retinotopically ordered connections. We found that the patterns of connections made by the reeler mutant are indistinguishable from normal. Cortical loci in area 17 are reciprocally connected to homotopic locations in areas 18a and 18b. Area 17 is also reciprocally connected with the dorsal lateral geniculate nucleus of the thalamus and projects to the superior colliculus. Areas 18a and 18b are reciprocally connected with each other and with the lateral posterior and lateral nuclei of the thalamus, respectively. In addition, we found evidence of reciprocal connections between the lateral posterior nucleus and area 17, and between the lateral nucleus and areas 17 and 18a. The results indicate that neurons in visual cortical areas of the reeler mutant mouse are capable of forming retinotopically organized corticocortical and thalamocortical connections in a pattern similar to that found in normal animals. Thus the genetic anomaly producing incorrect neuronal positioning during development of the reeler cortex does not seriously impede the pathway and target recognition mechanisms responsible for formation of functionally appropriate cortical connections.     

Multiple defects in the formation of rat cortical axonal pathways following prenatal X-ray irradiation

Prenatal X-ray irradiation is known to result in severe defects of neuronal migration and laminar formation in the cerebral cortex. We examined the formation of cortical afferent and efferent pathways in rats that had been exposed to X-ray irradiation (1.0 Gy) at embryonic day 14 (E14), by birthdating with bromodeoxyuridine (BrdU) and axonal labeling with 1-1'-dioctodecyl-3,3,3',3'- tetramethyl-indocarbocyanine perchlorate (DiI), in addition to immunohistochemical staining for various axonal markers including neurofilament, and cell adhesion molecules L1 and TAG-1. The results obtained were as follows. (i) The neuroepithelium formed germinal rosettes and concavities in the cortical anlage from 2 days after irradiation. Neurons generated in the neuroepithelium accumulated to form subcortical heterotopia and obstructed pathway formation in the intermediate zone, resulting in an aberrant trajectory of TAG-1-immunoreactive cortical efferent axons. (ii) In rats exposed to X-ray irradiation at E14, cystic cavities were formed in the cortex-striatum boundary region between E15 and E17, probably because of delayed cell death of neurons generated at E14. These cavities transiently interrupted both cortical afferent (L1-positive) and efferent (TAG-1-positive) axons. (iii) X-ray irradiation at E14 partially destroyed subplate neurons (transient targets of thalamic afferent axons) and disturbed the arrangement of the subplate layer. This resulted in a misrouting of neurofilament- and L1-immunoreactive thalamocortical axons that obliquely traversed the cortical plate to run up to the superficial layer. The present study demonstrates for the first time that X-ray irradiation during initial cortical development causes multiple defects in the formation of cortical afferent and efferent pathways.     

Pathfinding and target selection by developing geniculocortical axons.

During development of the mammalian cerebral cortex, thalamic axons must grow into the telencephalon and select appropriate cortical targets. In order to begin to understand the cellular interactions that are important in cortical target selection by thalamic axons, we have examined the morphology of axons from the lateral geniculate nucleus (LGN) as they navigate their way to the primary visual cortex. The morphology of geniculocortical axons was revealed by placing the lipophilic tracer Dil into the LGN of paraformaldehyde-fixed brains from fetal and neonatal cats between embryonic day 26 (E26; gestation is 65 d) and postnatal day 7 (P7). This morphological approach has led to three major observations. (1) As LGN axons grow within the intermediate zone of the telencephalon toward future visual cortex (E30-40), many give off distinct interstitial axon collaterals that penetrate the subplate of nonvisual cortical areas. These collaterals are transient and are not seen postnatally. (2) There is a prolonged period during which LGN axons are restricted to the visual subplate prior to their ingrowth into the cortical plate; the first LGN axons arrive within visual subplate by E36 but are not detected in layer 6 of visual cortex until about E50. (3) Within the visual subplate, LGN axons extend widespread terminal branches. This represents a marked change in their morphology from the simple growth cones present earlier as LGN axons navigate en route to visual cortex. The presence of interstitial collaterals suggests that there may be ongoing interactions between LGN axons and subplate neurons along the entire intracortical route traversed by the axons. From the extensive branching of LGN axons within the visual subplate during the waiting period, it appears that they are not simply "waiting." Rather, LGN axons may participate in dynamic cellular interactions within the subplate long before they contact their ultimate target neurons in layer 4. These observations confirm the existence of a prolonged waiting period in the development of thalamocortical connections and provide important morphological evidence in support of the previous suggestion that interactions between thalamic axons and subplate neurons are necessary for cortical target selection.     

Immunohistological localization of cell adhesion molecules L1, J1, N-CAM and their common carbohydrate L2 in the embryonic cortex of normal and reeler mice

The expression of the cell adhesion molecules L1, J1 and N-CAM and their shared carbohydrate L2 was studied in the embryonic cerebral cortex of normal and reeler mutant mice using light and electron microscopic immunocytochemistry. Apart from a general delay in their appearance in the reeler cortex, the 4 antigens were present with a cellular distribution in both genotypes reflecting the anatomical characteristics of normal and mutant phenotypes. The cell surface glycoprotein L1 was exclusively expressed by neurons, particularly axons, but was never detected at sites of neuron-glia contact. L1 was accumulated in the marginal zone and subplate of the normal cortex and in the homologous layers of the reeler cortex. The secreted glycoprotein J1 was found on glia and neurons. Although initially present in regions of fiber outgrowth, J1 became characteristically excluded from the large fiber tracts at later stages. J1 mapped in the marginal zone and subcortical plate of the normal cortex and in the corresponding layers of the mutant cortex. N-CAM had a more ubiquitous distribution and was present in ventricular zones, particularly at early stages, as well as on glia and neurons and large fiber tracts at later developmental stages. The distribution of the L2 epitope was quite similar to that of the J1 molecule but remained present on large fiber tracts, like N-CAM and L1, also at later developmental stages. These comparative observations in normal and reeler mutant mice lend support to previous suggestions that L1, together with N-CAM, may play a role in the aggregation of neuronal cell bodies after migration and in the fasciculation of developing fiber bundles. They also point to a possible function of the extracellular matrix component J1 in the guidance or support of fiber outgrowth in large fiber tracts.     

Development of functional thalamocortical synapses studied with current source-density analysis in whole forebrain slices in the rat

We analysed the laminar distribution of transmembrane currents from embryonic (E) day 17 until adulthood after selective thalamic stimulation in slices of rat forebrain to study the development of functional thalamocortical and cortico-cortical connections. At E18 to birth a short-latency current sink was observed in the subplate and layer 6, which was decreased, but not fully abolished in a cobalt containing solution or after the application of glutamate receptor blockers (APV and DNQX). This indicated that embryonic thalamic axons were capable of conducting action potentials to the cortex and some of them had already formed functional synapses there. Between birth and P3, when thalamic axons were completing their upward growth, a sink gradually appeared more superficially in the dense cortical plate and synchronously, a current source aroused in layer 5. Both sinks and sources completely disappeared after blocking synaptic transmission. The adult-like distribution of CSDs became apparent after P7. The component in layer 6 cannot be blocked completely after this age suggesting antidromic activation. This study demonstrated that cells of the lowest layers of the cortex received functional thalamic input before birth and that thalamocortical axons formed synapses with more superficial cells as they grew into the cortical plate.     

Development of connections in the human visual system during fetal mid-gestation: a DiI-tracing study

Animal studies have shown that connections between the retina, lateral geniculate nucleus (LGN), and visual cortex begin to develop prenatally. To study the development of these connections in humans, regions of fixed brain from fetuses of 20-22 gestational weeks (GW) were injected with the fluorescent tracer DiI. Placement of DiI in the optic nerve or tract labeled retinogeniculate projections. In the LGN, these projections were already segregated into eye-specific layers by 20 GW. Retinogeniculate segregation thus preceded cellular lamination of the LGN, which did not commence until 22 GW. Thalamocortical axons, labeled from DiI injections into the optic radiations, densely innervated the subplate, but did not significantly innervate the cortical plate. This pattern was consistent with observations of a "waiting period" in animals, when thalamocortical axons synapse in the subplate for days or weeks before entering the cortical plate. Cortical efferent neurons (labeled retrogradely from the optic radiations) were located in the subplate and deep layers of the cortical plate. In summary, human visual connections are partially formed by mid-gestation, and undergo further refinement during and after this period. The program for prenatal development of visual pathways appears remarkably similar between humans and other primates.     

Transient microcircuits formed by subplate neurons and their role in functional development of thalamocortical connections

Subplate neurons are a transient population of neurons in the brain forming one of the first functional cortical circuits. Past experiments have demonstrated their importance in growth of thalamocortical afferents into the cortical plate and later segregation of thalamocortical afferents. Recently, subplate neurons have been shown to be required for the functional maturation of both thalamocortical connections and mature visual responses in visual cortex. These findings suggest that thalamocortical afferents might not segregate properly in the absence of subplate neurons because the thalamocortical synapse does not mature. Subplate neurons are unique in that they form a circuit that appears to promote synaptic scaling and maturation. Although the precise contribution of subplate neurons within the context of cortical development is unknown, they might play an early role in providing thalamic input to cortex that then interacts with learning rules governing synaptic strengthening at the thalamocortical synapse. Because they appear to play multiple key roles at different stages of development, subplate neurons might also play a role in the pathology of developmental disorders, such as epilepsy and schizophrenia.     

Callosal axon guidance defects in p35(-/-) mice.

Mice lacking p35, an activator of cdk5 in the central nervous system (CNS), exhibit defects in a variety of CNS structures, most prominently characterized by a disruption in the laminar structure of the neocortex (Chae et al., 1997). In addition, alterations of certain axonal fiber tracts are found in the cortex of p35 mutant mice. Notably, the corpus callosum appears bundled at the midline, but dispersed lateral to the midline. Tracer injection experiments in adult p35 mutant mice reveal that projecting cortical axons fail to assimilate into the corpus callosum, and take oblique paths to the midline. After crossing the midline, cortical axons defasciculate prematurely from the corpus callosum and take similarly oblique paths through the cortex. This callosal phenotype is not detected in reeler mice, which also exhibit defects in cortical lamination, suggesting that the lack of fasciculation of callosal axons is not an inherent manifestation of a disruption of cortical lamination. The embryonic callosal axon tract is defasciculated before crossing the midline, suggesting that axon guidance may be affected during embryonic development of the corpus callosum. In addition, embryonic thalamocortical afferents also exhibit a defasciculated phenotype. These results suggest that defective axonal fasciculation and guidance may be primary responses to the loss of p35 in the cortex. Furthermore, this study postulates a role for the p35/cdk5 kinase in molecular signaling pathways necessary for proper guidance of selective axons during embryonic development.     

Role of semaphorin III in the developing rodent trigeminal system.

Semaphorins are a large family of secreted and transmembrane glycoproteins. Sema III, a member of the Class III semaphorins is a potent chemorepulsive signal for subsets of sensory axons and steers them away from tissue regions with high levels of expression. Previous studies in mutant mice lacking sema III gene showed various neural and nonneural abnormalities. In this study, we focused on the developing trigeminal pathway of sema III knockout mice. We show that the peripheral and central trigeminal projections are impaired during initial pathway formation when they develop into distinct nerves or tracts. These axons defasciculate and compromise the normal bundling of nerves and restricted alignment of the central tract. In contrast to trigeminal projections, thalamocortical projections to the barrel cortex appear normal. Furthermore, sema III receptor, neuropilin, is expressed during a short period of development when the tract is laid down, but not in the developing thalamocortical pathway. Peripherally, trigeminal axons express neuropilin for longer duration than their central counterparts. In spite of projection errors, whisker follicle innervation appears normal and whisker-related patterns form in the trigeminal nuclei and upstream thalamic and cortical centers. Our observations suggest that sema III plays a limited role during restriction of developing trigeminal axons to proper pathways and tracts. Other molecular and cellular mechanisms must act in concert with semaphorins in ensuring target recognition, topographic order of projections, and patterning of neural connections.     

Thalamic connectivity of the primary motor cortex of normal and reeler mutant mice

Reeler, an autosomal recessive mutation in mice, causes cytoarchitectonic abnormalities of the cerebral cortex, which are characterized by malposition of neurons. Retrograde and anterograde transport of horseradish peroxidase (HRP) was employed to examine the reciprocal connectivity between the hindlimb area of the primary motor cortex (MI) and thalamus of normal and reeler mutant mice. In the normal mouse, most of the cells labelled after HRP injection into the hindlimb area of MI were located in the ventrolateral nucleus, the lateral division of the ventrobasal nucleus, the central lateral, paracentral and central intralaminar nuclei, and the medial division of the posterior complex. HRP reaction product anterogradely transported was also observed in the same nuclei and in the thalamic reticular nucleus. In the reeler mutant mouse, retrogradely labelled neurons and anterogradely labelled terminals were again found in the nuclei referred to above, and the distribution pattern and morphology of HRP-filled neurons were also similar to those of normal controls. The present results suggest therefore that the normal reciprocal connectivity between MI (hindlimb representation) and thalamus is preserved in the reeler mouse. That is to say, dislocated cortical neurons appropriately project to their target nuclei of the thalamus, and conversely, thalamic neurons send their axons precisely to their target cortical areas of the radially disorganized cortex.