Embryonic Tissue Induces Growth of Adult Axons from Myelinated Fiber Tracts

Suspensions of late embryonic hippocampal tissue were microinjected so as to be completely enclosed within the myelinated fiber bundles of the adult rat fimbria. Previous studies have shown that the axons from such transplanted neurons readily cross the graft/host interface and extend rapidly through the host fiber tract. The present study shows that the adult axons from the host fiber tract can also cross this interface in the opposite direction and enter the transplants. Biotin dextran tracing shows that the adult host fimbrial axons traverse the embryonic grafts and also form terminal arborizations within the transplants. Electron microscopy of orthograde electron-dense degeneration confirms that these host axons form synaptic terminals accounting for at least 6.6% of the synapses in the neuropil of the transplant. Thus, contact with embryonic nervous tissue can induce elongative growth by the adult fibers in a myelinated central tract.     

Morphology and migration of cultured schwann cells transplanted into the fimbria and hippocampus in adult rats

Schwann cells cultured from neonatal rat peripheral nerve were injected into the fimbria and hippocampus of syngeneic adult rats by a microtransplantation technique which causes minimal disturbance to the host brain structure at the site of implantation, and thus allows the grafted cells to come into immediate contact with intact host tissue. Numerous Schwann cells could be identified for up to 6 weeks (and with decreasing frequency for up to 3 months) by intense immunoreactivity for low affinity nerve growth factor receptor. The transplanted cells adopted a distinctive elongated form, with a central, ovoid nucleus flanked by processes which were up to 300 μm long, and which ranged from swollen segments with a diameter as large as 12 μm down to thread-like fibres of 1 μm or less. This morphology is different from that of any of the host cells. The transplanted Schwann cells migrated freely into the host tissue along blood vessels and according to the position of the grafts, they either entered the hippocampal neuropil, or migrated (for distances of up to 2 mm) along the longitudinal axis of the fimbria, where they were interspersed in parallel with the interfascicular glial rows and axons. The host astrocytes did not appear to impede the migration of the donor Schwann cells. Although the host astrocytic processes became hypertrophic, with increased glial fibrillary acidic protein and vimentin expression, the predominant longitudinal orientation of the astrocytic tract processes was maintained. The transplanted Schwann cells did not form peripheral myelin (as detected by P_o immunoreactivity), and it is not clear whether they survive beyond the period at which we detect them. © 1993 Wiley-Liss, Inc.     

Myelination in the human hippocampal formation from midgestation to adulthood

Myelination, one of the last steps of neuronal development, was examined in the human fetal and postnatal hippocampal formation using immunohistochemistry to detect a protein component of the myelin sheath, the myelin basic protein synthesized by oligodendroglial cells. Myelin basic protein-immunoreactive oligodendroglial cells were first seen at the 20th gestational week in the fimbria fornicis and in the alveus. Between the 21st and 35th weeks, myelinated axons also appeared in the fimbria fornicis. At the age of 39 gestational weeks, short and thin myelinated fibers were present in the fimbria, in the alveus, and less so in the stratum oriens of the hippocampus, while the first oligodendroglial cells appeared in the stratum lacunosum-moleculare and in the hilus. By the 2nd postnatal week myelinated fibers appeared in the stratum lacunosum-moleculare of Ammon's horn. At the 3rd month, myelination was strong in the alveus, moderate in the strata oriens, lacunosum-moleculare and radiatum of Ammon's horn, while only a small number of myelinated fibers were detected in the hilus. By the 5th month, the first oligodendroglial cells were detected in the molecular layer of the dentate gyrus. Myelination continued in the following years, particularly in the dentate gyrus, where even at the age of 11 years the density of myelinated fibers did not reach the adult level. It appears that the first myelinated axons belong to the long-projecting large hippocampal pyramidal cells and/or to their subcortical and cortical afferents. The sequence of myelination follows the known developmental pattern of hippocampal afferent and efferent pathways, and the prolonged myelination might be a factor in the prolonged functional maturation of hippocampal circuitry.     

Sprouting of sympathetic axons in the hippocampal formation: Conditions necessary to elicit ingrowth

Sympathetic, noradrenaline (NA)-containing axons, which do not normally innervate the parenchyma of the hippocampal formation, grow into this structure following lesions of the fimbria. This anomalous innervation apparently arises by collateral sprouting from the normal sympathetic plexus on arteries within the subarachnoid space along the medial surface of the hemisphere. These NA axons travel orthogonal to the septotemporal axis within the dentate hilus and stratum oriens of CA3, and parallel to the septotemporal axis within the stratum lucidum of CA3. Regardless of the type of lesion (electrolytic, aspiration, knife cut) the septohippocampal input must be transected to elicit the ingrowth. No ingrowth follows selective lesions of the commissural or entorhinal cortical inputs or of the vasculature and cortex overlying the septal pole of the hippocampal formation, including central NA collaterals in the fasciculus cinguli. Discrete lesions of portions of the fimbria permit localized sympathetic ingrowth into the septal (medial fimbria) or temporal (lateral fimbria) ends of the structure. The pattern of anomalous innervation is neither extended by additional commissural or entorhinal cortical deafferentation, nor apparent in all areas deafferented by the septal lesion. Thus, the ingrowth is not regulated solely by the presence of degenerating terminal fields, and although damage to the septal afferent appears necessary, it is not sufficient to permit sympathetic axon and terminal proliferation within the entire hippocampal formation.     

Organotypic slice co‐cultures reveal that early postnatal hippocampal axons lose the ability to grow along the fimbria,while retaining the ability to invade and arborise in septal neuropil

The failure of cut axons to grow along fibre tracts in the adult CNS contrasts with their ability to do so in development. Organotypic slices culture of a number of areas enables the time of failure to be pinpointed to around the second week of postnatal life in the rat. ‘Heterochronic’ co‐culture of slices above and below this age shows that the failure is due to the inability of the older axons to grow into either the same age or younger targets. Using hippocampo‐septal slices the present experiments show that this failure is due to an inability to recognise the glial pathway of the fimbria, even when this is of a younger age. However, the older hippocampal neurons retain the ability to grow axons into septal target tissue when they are placed in direct contact with it. This exactly mirrors the inability of cut central axons to regenerate along their previous fibre pathways while they retain their ability to reinnervate neuropil.     

The topographic organization of hypothalamic and brain stem projections to the hippocampus

Direct projections primarily ipsilateral to hippocampus from medial septal, diagonal band, supramammillary, submammillothalamic, locus coeruleus, and dorsal and medianus raphe nuclei were demonstrated. The locus coeruleus projects primarily through the cingulum and fornix superior to the dorsal posterior hippocampus, with its terminal fields in the stratum lacunosum moleculare of the subiculum and areas CA 1-CA 2 of the dorsal posterior hippocampus. LC projections to the granular layer of the dentate hilus were not found. Raphe nuclei project through the cingulum, fornix superior, and primarily the fimbria, to the dorsal and ventral posterior hippocampus, with their terminal fields in the stratum lacunosum moleculare of the dorsal posterior subicular region, stratum radiatum of CA 1-CA 3 in the dorsal hippocampus, and the stratum polymorph of the dentate gyrus, primarily in its superficial part. Raphe projections to the anterior hippocampal rudiment were found. However, no projection was found to the subiculum of the ventral posterior hippocampus, nor to stratum oriens. Hypothalamic nuclei project through the fornix superior and the fimbria, mainly to the dorsal posterior hippocampus with abundant terminal fibers in the depth of the dentate hilus. Smaller cells in these hypothalamic nuclei appear projecting to the ventral hippocampus. The number of neurons in the entorhinal area, the diagonal band, and the hypothalamic nuclei projecting to the hippocampus suggests these groups as the main sources of the extrinsic hippocampal afferents. In addition, they may also serve as relay stations for inputs from more caudal nuclei, and the topographic organization of their terminal fields as described herein may have important functional implications.     

Axonal plasticity elicits long‐term changes in oligodendroglia and myelinated fibers

Axons are linked to induction of myelination during development and to the maintenance of myelin and myelinated tracts in the adult CNS. Currently, it is unknown whether and how axonal plasticity in adult CNS impacts the myelinating cells and their precursors. In this article, we report that newly formed axonal sprouts are able to induce a protracted myelination response in adult CNS. We show that newly formed axonal sprouts, induced by lesion of the entorhino–hippocampal perforant pathway, have the ability to induce a myelination response in stratum radiatum and lucidum CA3. The lesion resulted in significant recruitment of newly formed myelinating cells, documented by incorporation of the proliferation marker bromodeoxyuridine into chondroitin sulphate NG2 expressing cells in stratum radiatum and lucidum CA3 early after lesion, and the occurrence of a 28% increase in the number of oligodendrocytes, of which some had incorporated bromodeoxyuridine, 9 weeks post‐lesion. Additionally, a marked increase (41%) in myelinated fibres was detected in silver stained sections. Interestingly, these apparently new fibres achieved the same axon diameter as unlesioned mice but myelin thickness remained thinner than normal, suggesting that the sprouting axons in stratum radiatum and lucidum CA3 were not fully myelinated 9 weeks after lesion. Our combined results show that sprouting axons provide a strong stimulus to oligodendrocyte lineage cells to engage actively in the myelination processes in the adult CNS. © 2009 Wiley‐Liss, Inc.     

Transplantation of cultured type 1 astrocyte cell suspensions into young, adult and aged rat cortex: Cell migration and survival

The present study examined the fate and migration of transplanted astrocytes in different host ages. Additionally, the effect of donor cell age was examined in relation to cell migration. Cultured astrocytes from 5,12 and 30 days in vitro were transplanted into young (postnatal day 5 and 21), adult (4.5 month), and aged (21 month) animals. The transplanted cells were labeled with Fast Blue, Fluorogold or DiI. The results confirmed previous studies demonstrating that transplanted cells were able to migrate successfully through host central nervous system and extended those findings to show that the age of the host significantly influenced donor cell migration distance. Migration was most extensive in young animals, as conditions supporting cell migration appeared to be lacking in older animals. Donor cells preferentially migrated on myelinated fiber tracts, rather than on unmyelinated fiber tracts or gray matter. The donor cells were not glial fibrillary acidic protein positive, indicating that either the cultured type 1 astrocytes did not survive transplantation or underwent significant remodeling of the intermediate filament network. It is also possible that a subpopulation of cells, possibly immature astrocytes which are present in the transplanted cell suspensions, flourished and subsequently migrated in the host brains.     

Axonal composition of the marginal and basal optic tracts of the grass frog]

An electron-microscopy study of the marginal optic tracts (the medial and lateral divisions) and basal optic tract was performed for the frog Rana temporaria. The resemblance between the ultrastructure and composition of the marginal tracts is revealed. The basal tract is characterized by a relatively loose structure and considerable content of the myelinated large diameter fibres. The corresponding tract contains 3900 (4.7%), 4700, (4.9%), 700 (28%) of myelinated fibres and 79 800 (95.3%), 91500 (95.1%) and 1800 (72%) of unmyelinated ones. The spectra of the myelinated fibre diameters of the marginal tract are from 0.4 to 2.6 mum with the main peak at 1.0 mum and additional peak at 1.6 mum; the spectrum of the myelinated fibre diameters of the basal tract is from 0.4 to 4.0 mum with the main peak at 1.8 mum and the small additional peak at 3.2 mum. The spectra of the unmyelinated fibre diameters of all tracts are from 0.1 to 0.5 mum; 60% of them have a diameter of about 0.2 mum. 85 days after enucleation undegenerated myelinated fibres were not found in the basal tract of the frog kept at a temperature 18-20 degrees C. The medial and lateral marginal tracts contained a significant number of the undegenerated myelinated fibres, 33% and 13%, the diameter spectra being 0.6-1.4 mum and 0.6-1.0 mum, respectively. It is possible that these fibres participate in the organization of the ipsilateral visual projection on the tectum. Unmyelinated fibres remain unchanged in all the studied tracts.     

Temporal and spatial regulation of chondroitin sulfate, radial glial cells, growing commissural axons, and other hippocampal efferents in developing hamsters

We investigated the time and space relationship between growth of hippocampal efferents, particularly those forming the hippocampal commissure, and expression of extracellular matrix components related to radial glial cells. Developing hamster brains from embryonic day (E) 13 to postnatal day (P) 7 had 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) crystals implanted into the hippocampus or were processed for fluorescent immunohistochemistry against chondroitin sulfate (CS) glycosaminoglycans and glial fibrillary acidic protein (GFAP). The first, pioneer fibers from the hippocampus were seen crossing the midline at E15 and arriving at the contralateral hippocampus 24-48 hours later (P1), followed closely by a thick front of growing fibers. Before E15, CS expression was preceded by septal fusion and was concomitant with formation of the commissural tract. On E15, CS expression formed a U-shaped border below the fimbria. From E15 to P3, CS became expressed between the hippocampal commissure and the third ventricle and at the caudal borders of the fornix columns. As the hippocampal commissure expanded, CS expression became gradually lighter to virtually disappear by P7. On E15 and P1, GFAP-positive radial glial cells were present caudal (but not rostral) to the commissure at the midline, partially overlapping CS expression. Similar cells were present dorsal to the fimbria, extending their processes perpendicularly over the growing axons. The data reveal that CS and radial glial cells form a tunnel surrounding the developing fimbria and a border at the midline caudal to the hippocampal commissure. It is suggested that these cellular and molecular borders play a role in guidance of hippocampal efferents.     

Regeneration of the septohippocampal pathways in adult rats is promoted by utilizing embryonic hippocampal implants as bridges

The ability of embryonic hippocampal tissue to promote regeneration of cholinergic axons in the septohippocampal system has been studied in adult rats. Strips of embryonic hippocampus, taken from 7–40 mm rat fetuses, were implanted into a 2–3 mm wide cavity which completely transected the septal cholinergic axons innervating the intrinsic hippocampus. The ingrowth of cholinergic fibres into the denervated host hippocampal formation was monitored by measuring the activity of the enzyme, choline acetyltransferase (ChAT), and by acetylcholine esterase (AChE) histochemistry. The results demonstrated a gradual, partial return of both ChAT enzyme activity and AChE-positive fibres in the initially denervated hippocampal formation of the adult recipient. Time-course studies indicated that this ingrowth progressed from the implant into the rostral tip of the host hippocampus, and continued caudally to cover the entire dorsal hippocampus by 3–6 months post-operative Although the regenerating AChE-positive fibres reached the hippocampal target in the recipient along abnormal routes, they reinnervated selectively the appropriate terminal areas within the host hippocampus and dentate gyrus, suggesting the presence of quite specific mechanisms to guide the regenerating axons back to their original targets. Lesions of the medial septum-diagonal band area of the host and horseradish peroxidase (HRP) injections into the host hippocampus, caudal to the implant, indicated that the origin of the regenerating axons was predominately from the ipsilateral ventral medial septum and diagonal band area of the host. The results provide evidence that axonal regeneration and reinnervation of a denervated target zone can be promoted by utilizing implants of embryonic CNS tissue to bridge a tissue defect between the target and the lesioned axonal stumps.     

Cellular, histochemical and connective organization of the hippocampus and fascia dentata transplanted to different regions of immature and adult rat brains

The aims of the present study were to examine the survival and the cellular and connective differentiation of intracerebral transplants of fascia dentata and hippocampus. Pieces of immature dentate and hippocampal tissue were taken from late embryonic (E18) and early postnatal (1-9 days old) rats and transplanted into the brains of 1- to 13-day-old and adult rats. After survival times from 4 days to 2 years the cellular and connective organization of the transplants was monitored in parallel series of sections stained with thionin (cell bodies), Timm's sulphide silver method (terminal fields). Nauta and Fink-Heimer methods (normal and degenerating fibers) and a method for AChE activity (cholinergic afferents). The transplants survived well in all combinations of donor and recipient ages used, and they survived and differentiated in all parts of the recipient brains, although relations to pial and ventricular surfaces appeared to be optimal. Cell differentiation continued after transplantation, and a characteristic laminar organization was retained, although least in embryonic donor tissues. The distribution of intrinsic connections was determined by the types of subfields present in the transplants and interaction with ingrown host afferents. All aberrant intrinsic connections observed corresponded to aberrant connections formed in the hippocampus and fascia dentata denervated in situ and included supragranular mossy fibers in the fascia dentata, aberrant infrapyramidal mossy fibers in CA3, spread of CA4-associated afferents beyond the normal commissural-associational zone in the dentate molecular layer together with ingrowth of CA3-associated and CA1-subiculum-associated afferents. Most transplants received a cholinergic input of host origin irrespective of the localization in the host brain, but also non-cholinergic host pathways innervated the transplants, in particular when the transplants were in close contact with host fiber tracts, and when the recipients were immature. At various transplant locations the non-cholinergic host afferents belonged to the commissural hippocampo-dentate system, the commissural hippocampal system and the callosal system. Other cases suggested innervation of dentate transplant by host entorhinal afferents. The formation and distribution of intrinsic transplant connections and connections between transplant and host appeared to be regulated by the same factors that regulate the development and reorganization of fiber connections in the normal and the in situ denervated hippocampus and fascia dentata. As a special variety of this, the distribution of cholinergic afferents adjusted to the distribution of the major intrinsic and extrinsic non-cholinergic pathways.     

Fluorescent tracer in pilocarpine-treated rats shows widespread aberrant hippocampal neuronal connectivity

Neuronal fibres of the hippocampal formation of normal and chronic epileptic rats were investigated by fluorescent tracing methods using the pilocarpine model of limbic epilepsy. Two months after onset of spontaneous limbic seizures, hippocampal slices were prepared and maintained in vitro for 10 h. Small crystals of fluorescent dye [fluorescein (fluoro-emerald) and tetramethylrhodamine (fluoro-ruby)] were applied to different hippocampal regions. The main findings were: (i) in control rats there was no supragranular labelling when the mossy fibre tract was stained in stratum radiatum of area CA3. However, in epileptic rats a fibre network in the inner molecular layer of the dentate gyrus was retrogradely labelled; (ii) a retrograde innervation of area CA3 by CA1 pyramidal cells was disclosed by labelling remote CA1 neurons after dye injection into the stratum radiatum of area CA3 in chronic epileptic rats; (iii) labelling of CA1 neurons apart from the injection site within area CA1 was observed in epileptic rats but not in control animals; and (iv), a subicular-hippocampal projection was present in pilocarpine-treated rats when the tracer was injected just below the stratum pyramidale of area CA1. The findings show that fibre rearrangement in distinct regions of the epileptic hippocampal formation can occur as an aftermath of pilocarpine-induced status epilepticus.     

Region-specific differentiation of embryonic stem cell-derived neural progenitor transplants into the adult mouse hippocampus following seizures

Embryonic stem (ES) cells can generate neural progenitors and neurons in vitro and incorporate into the adult central nervous system (CNS) following transplantation, suggesting their therapeutic potential for treating neurological disorders. However, our understanding of the conditions that direct ES-derived neural progenitor (ESNP) migration and differentiation within different regions of the adult CNS is incomplete. Rodents treated with the chemoconvulsant kainic acid (KA) experience seizures and display hippocampal sclerosis, as well as enhanced hippocampal neurogenesis, similar to pathological findings in patients with temporal lobe epilepsy (TLE). To examine the potential for ESNPs to incorporate into the adult hippocampus and differentiate into hippocampal neurons or glia following seizure-induced damage, we compared the fates of ESNPs after they were transplanted into the CA3 region or fimbria 1 week following KA-induced seizures. After 4-8 weeks, ESNPs grafted into the CA3 region had migrated to the dentate gyrus (DG), where a small subset adopted neural stem cell fates and continued to proliferate, based on bromodeoxyuridine uptake. Others differentiated into neuroblasts or dentate granule neurons. In contrast, most ESNPs transplanted into the fimbria migrated extensively along existing fiber tracts and differentiated into oligodendrocytes or astrocytes. Hippocampal grafts in mice not subjected to seizures displayed a marked tendency to form tumors, and this effect was more pronounced in the DG than in the fimbria. Taken together, these data suggest that seizures induce molecular changes in the CA3 region and DG that promote region-specific neural differentiation and suppress tumor formation.     

Schwann cells transplanted into the CNS

A small volume of purified Schwann cells, cultured from early postnatal rat sciatic nerve, was injected into the hippocampus or fimbria of syngeneic adult hosts. The procedure caused minimal structural disturbance at the transplantation site, with close graft-host contact and maximal opportunity for integration.The donor Schwann cells were identified by a combination of light and electron microscopic features (which include characteristic deep and complex infoldings of a well marked nuclear envelope), antigenic profile (especially low affinity nerve growth factor receptor immunoreactivity), uptake of fluorescent latex microspheres and autoradiography of [³H]thymidine-labelled dividing cells.The donor Schwann cells adopted a distinctive elongated form, with a central, ovoid nucleus flanked by processes which were up to 300 μm long, and which ranged from swollen segments with a diameter as large as 12 μm down to thread-like fibres of 1 μm or less with growth cone-like expansions. Transplanted cells migrated from the graft, particularly along blood vessels, and could permeate all cytoarchitectonic regions of the adjacent host hippocampal neuropil. Donor Schwann cells also migrated along the longitudinal axis of the fimbria, where they were interspersed in parallel with the interfascicular glial rows and axons.The grafted cells induced a transient but marked host astrocytic hypertrophy, which did not appear to impede the migration of the donor Schwann cells.The transplanted Schwann cells did not form peripheral myelin (as detected by P_o immunoreactivity), and it is not clear whether they survive beyond the period at which we detect them.     

A quantitative study of morphological changes accompanying the initiation and progress of myelin production in the dorsal funiculus of the rat spinal cord

The dorsal funiculus in cervical spinal cords of rats from 3 to 120 days postnatal was studied in order to document and quantitate glial cell development and axonal growth as related to the initiation and progress of central myelination. Within the dorsal funiculus are three major and distinct tracts, each having distinct developmental trends and adult characteristics in terms of fiber sizes and amount of myelin. These tracts are the cuneate and gracile fasciculi and the cortico-spinal tracts. Glial cell counts and cross-sectional surface area determinations of each tract at increasing ages show that the initial rate of glial population increase is similar. However, each tract is unique in terms of the age at which a maximum population density is reached and the rate at which the expected population dilution takes place. An electron-microscopic examination indicates that oligodendrocytes constitute over 85% of the total glial population throughout the development period surveyed. As such, these cells are primarily responsible for the population density changes. The diameters of unmyelixgnated fibers, promyelin fibers and some myelinated fibers in these tracts were measured at 5, 10, 15, 20 and 120 days postnatal. This was done both for the purpose of relating glial population density changes with the initiation and decline of active myelination, and for determining whether or not a critical diameter for myelination exists in the CNS as was found in peripheral nerves (Matthews, '68). For each tract there is a characteristic sequence of events involving not only myelination, but also changes in diameter distribution just prior to the appearance of myelin and during the period of active myelin formation. These events coincide with the concentration and dilution of the glial population, but it is also evident that there is no critical and constant diameter in the CNS above which all axons are myelinated and below which all are unmyelinated. Myelin appears first on larger axons, but as the animal matures, it is found on progressively smaller axons until between 20 and 120 days, axons 0.2–0.4 μ in diameter acquire myelin. Thus, myelination begins with axons destined to be large and then extends down to those which enlarge very little prior to acquiring myelin and remain very small even in adult animals. Finally, from the determination, in adult rats, of the number of axons and oligodendrocytes in a defined volume of each tract and an estimation of internode length, the ratio of internodes to oligodendrocytes was calculated. The specific values obtained could vary by as much as ±50% and are only meant to serve as indicators of a trend. However, it is suggested that the number of internodes per oligodendrocyte may be inversely proportional to the length of the internode.     

Transplantation of olfactory ensheathing cells or Schwann cells restores rapid and secure conduction across the transected spinal cord

Olfactory ensheathing cells (OECs) or Schwann cells were transplanted into the transected dorsal columns of the rat spinal cord to induce axonal regeneration. Electrophysiological recordings were obtained in an isolated spinal cord preparation. Without transplantation of cells, no impulse conduction was observed across the transection site; but following cell transplantation, impulse conduction was observed for over a centimeter beyond the lesion. Cell labelling indicated that the regenerated axons were derived from the appropriate neuronal source, and that donor cells migrated into the denervated host tract. As reported in previous studies, the number of regenerated axons was limited. Conduction velocity measurements and morphology indicated that the regenerated axons were myelinated, but conducted faster and had larger axon areas than normal axons. These results indicate that the regenerated spinal cord axons induced by cell transplantation provide a quantitatively limited but rapidly conducting new pathway across the transection site.     

Metastasis-associated mts1 (S100A4) protein in the developing and adult central nervous system

We have found recently that white matter astrocytes in the spinal cord constitutively express immunoreactivity for Mts1 (S100A4) protein and that this expression is up-regulated ipsilaterally after sciatic nerve or dorsal root injury. Here, we have studied the expression pattern of Mts1 throughout the rat central nervous system (CNS). We found Mts1 immunoreactivity in myelinated tracts such as the olfactory tract, optic nerve, corpus callosum, internal capsule, fimbria, and spinal cord funiculi but not in cerebellar white matter. Mts1-immunoreactive (IR) cells were consistently astrocytic (glial fibrillary acidic protein positive). In addition to myelinated tracts, Mts1 immunoreactivity was also present in a few nonmyelinated or poorly myelinated areas, such as pituitary gland, olfactory bulb, and around the lateral ventricle. Based on location, three Mts1-IR astrocyte groups were distinguished: 1) astrocytes at the surfaces of the CNS, i.e., adjacent to the cerebrospinal fluid, organized perpendicularly to the bundles of axonal tracts; 2) astrocytes located in parallel to, and inserted between, axonal bundles; and 3) clusters of astrocytes around the lateral ventricle and in the olfactory bulb. We further analyzed the relationship between Mts1 immunoreactivity and the development of CNS fiber tracts by combining staining for Mts1 and myelin basic protein (MBP). Mts1 immunoreactivity appeared postnatally in recently myelinated areas. During the development of corpus callosum and the optic tract, Mts1 immunoreactivity was concentrated at the frontier of myelination. The developmental expression pattern suggests a role of Mts1-IR astrocytes in the maturation of myelinated fiber tracts. The preferential localization of Mts1 to the subpial region in the mature CNS suggests that Mts1 participates in astrocyte-mediated CNS-cerebrospinal fluid exchange.     

Monoclonal Antibodies to Late-differentiating Epitopes Identify Mossy Fibre Terminals Innervating Normal and Transplanted Hippocampal CA3 Pyramidal Cells

We have derived two monoclonal antibodies, MF-1 and MF-2, which both recognize the same 58-kD antigen. Light and electron microscopic immunocytochemistry showed that this antigen is highly expressed in the large mossy fibre terminals innervating the proximal portion of the apical dendrites of pyramidal neurons in hippocampal field CA3. Staining was seen in the adult hippocampus in rats and mice, and in a post mortem human sample. Comparison with the Timm stain showed that the antibodies recognize mossy fibres from all parts of the adult dentate gyrus except for the tip of the infrapyramidal blade (the latest part of the dentate gyrus to develop). The MF antigen is expressed by mature terminals, and is not detected immunohistochemically in developing hippocampal mossy terminals until the end of the first postnatal week (i.e. later than the Timm-positive material). It was also found in host mossy fibre terminals innervating embryonic CA3 pyramids transplanted into adult hosts, but not in areas of the graft containing transplanted CA1 pyramids. These results indicate that this previously undescribed, late-developing antigen provides a useful specific marker for the mossy fibre projection in both the normal hippocampus and in situations of experimentally manipulated connectivity.