Transplantation of fetal mesencephalic and medullary raphe tissues into the cerebellum of denervated adult rats--an immunohistochemical study.

Pieces of mesencephalic and medullary raphe tissues were transplanted into the cerebella of 5,6-dihydroxytryptamine-treated adult rats. The extent of axonal outgrowth of serotonergic and dopaminergic neurons in the grafts was immunohistochemically studied. At 3 months after transplantation, numerous dopaminergic neurons with many processes extending within the graft were detected in the mesencephalic raphe graft, but not in the medullary raphe graft. In contrast, both the mesencephalic and medullary raphe grafts contained numerous serotonergic neurons and a dense plexus of their fibers. The outgrowing serotonergic fibers from the mesencephalic raphe graft showed a hyperinnervation pattern in the cerebellar cortex adjacent to the graft. Furthermore, a glomerulus-like accumulation of serotonergic fibers was observed in the granular layer. In the cases of medullary raphe grafts, the relatively abundant outgrowing serotonergic fibers showed a laminar organization in the cerebellar cortex near the graft, which was similar to the normal distributional pattern. These results indicate that serotonergic and dopaminergic neurons located within the mesencephalic raphe graft clearly differed from each other in their ability to extend their processes into the host cerebellum, which provides further evidence for the existence of specific interactions between outgrowing serotonergic fibers and their terminal fields (targets).

Development of the nuclei and cell migration in the medulla oblongata. Application of the quail-chick chimera system.

We have tried to obtain new insight into the development of the medulla oblongata by using the quail-to-chick chimera system. Five types of isotopic and isochronic grafts were carried out, between quail and chick embryos, at the 10- to 12-somite stage: exchanges of (I) the entire myelencephalon, (II) the dorsal half of the myelencephalon, (III) the ventral half of the myelencephalon, (IV) the right half of the myelencephalon and (V) the dorsal quarter of the myelencephalon. Before analyzing the chimeric embryos, we studied the ontogeny of the various nuclei in the medulla oblongata of normal birds. The first appearance of nuclei in quail embryos preceded in many cases that of their chick counterpart by 12 to 24 h. The adult pattern of the nuclei was established by E8 in quail and E9 in chick. Similarly, during early development of chimeras, the migration of quail cells began earlier than that of chick cells. This shows that the species specific temporal sequence of proliferation and migration is not significantly altered by transplantation into the host. The possibility of grafting selectively the ventral or dorsal half of the neural tube allowed us to distinguish the fate of the cells belonging respectively to the alar and the basal plate. The nuclei with a total or partial motor function, such as the nucleus nervi abducentis, the nucleus nervi facialis, the nucleus nervi glossopharyngei and the nucleus motorius dorsalis nervi vagi, have either an exclusive or predominant origin from the basal plate. In contrast, the nuclei with essentially or exclusively sensory components (i.e., nucleus angularis, nucleus laminaris, nucleus magnocellularis) arise from the alar plate. The reticular formation such as the nucleus reticularis gigantocellularis and the nucleus reticularis subtrigeminalis was strikingly mixed, with both alar and basal plate origin of neurons. Active dorsoventral migrations of cells originating migrations from the dorsal neural tube, the "rhombic lip", contribute the ventral nuclei (i.e., nuclei pontis medialis, lateralis and olivaris inferior), whose functions are essentially associative. This study shows different types of cell migration. Dorsoventral and ventrodorsal movements are essentially active from E5 to E8. In the medulla oblongata, the dorsoventral stream is highly predominant. From E8 to E9, cells belonging to the marginal stream cross the midline laterally in both directions. Beyond E12, longitudinal migrations occur ventrally in both rostrocaudal and caudorostral directions. The immunohistochemical analyses carried out on chimeras generated in experiment V revealed the existence of fibers in marginal zones prior to the onset of the migration of cell bodies.(ABSTRACT TRUNCATED AT 400 WORDS)

[Estrogen effect on the development of neurons in an intraocular transplant of rat solitary tract nuclei]. / Vliianie éstrogenov na razvitie neironov intraokuliarnogo transplantata iader solitarnogo trakta krys.

At the light and electron-microscopic levels, using morphometric methods, we investigated the estradiol-17 beta effects on neural growth in the solitary tract nucleus grafts. Estradiol-17 beta increased the functional activity in some neurons of the grafts and caused their cell processes to grow. It also affected the synaptogenesis in the neuropil of the graft tissues.

[Transplantation of embryonic medulla oblongata into cerebella of adult rats].

Pieces of medulla oblongata anlagen were dissected free from embryonic 13-20 day (E 13 to E 20) rat brain, and these were transplanted into the cerebellar vermis of adult rats (Fischer 344). After grafting, host animals survived for 4-9 months. Cytoarchitectonic organization of the graft and the relationship between host and graft were analyzed light microscopically in 34 animals using the Nissl and silver impregnation methods. Fine structures of the graft were analyzed in 4 animals using electron microscope. Grafts from E 13-14 donor tissue showed the highest survival rate (90%), which decreased as the donor embryonic age increased (i.e., E 15-16: 33%, E 17-20: 15%). In the surviving grafts, small (5-10 microns diameter), medium-sized (10-20 microns) and large (20-30 microns) neurons, whose cytoplasmic organelles appeared normal, were observed. Bundles of myelinated fibers traversed in every direction and neurons were often clustered, indicating characteristic features of the medulla oblongata. Electron microscopically, various types of synaptic formations were also observed. Degenerative profiles of nerve-fiber endings, containing dense bodies and lysosomal figures, were also seen. The degeneration seemed to be caused by the failure of their establishing connections with their proper targets in the host. In both the host tissue and the graft-host interface, neuronal processes apparently derived from the graft were frequently observed. Some axonal processes contained large-cored vesicles, and some dendritic processes were enlarged at their stalks and tips. Aberrant axon terminals of unmyelinated fibers in the host medullary layer were considered to be the graft origin. These fibers were always accompanied by prominent glial proliferation. There was no indication of forming myelinated fiber bundles that entered the host cerebellum from the donor tissue, although the former was the target of the latter. Cell bodies of host granule cells and oligodendroglia in the graft-host interface were surrounded by processes of oligodendroglia, forming thin myelin lamellae. The present study showed that unmyelinated fibers from the graft could enter the host cerebellum non-specifically accompanied by glial proliferation, whereas myelinated fibers could hardly penetrate into the host crossing the glial scar.

Expression of phenylethanolamine N-methyltransferase (PNMT) and neuropeptide Y (NPY) in embryonic rat medulla oblongata grown in Oculo.

Expression and development of specific markers of the adrenergic phenotype were studied in central neurons grown in transplant system. Medulla oblongata from embryonic day 12.5 (E12.5) or E18 rat was grafted into the anterior chamber of the eye of adult rat hosts. After two months, grafts were examined for the presence of immunoreactivity (IR) and catalytic activity to the epinephrine-synthesizing enzyme, phenylethanolamine N-methyltransferase (PNMT, E.C. 2.1.1.28), a specific adrenergic marker. In addition, grafts were examined for immunoreactivity to neuropeptide Y (NPY) and tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine synthesis. In E12.5 grafts, PNMT was expressed de novo, enzyme activity developed to levels similar to those in adult rat brainstem and PNMT-IR neurons were observed. TH-IR and NPY-IR neurons were also observed. In contrast, PNMT-IR was not observed in E18 grafts even though these already contained PNMT-IR neurons at the time of grafting. This was not due to poor growth of E18 grafts, in general, since TH-IR neurons were present and the protein content of the grafts was similar to that of E12.5 grafts. These studies suggest that adrenergic neurons survive well in oculo if they are transplanted prior to the age when neuroblasts have initially expressed the adrenergic phenotype, migrated to their final positions and elaborated processes. In addition, these studies establish a transplant system in which factors required for the development of central adrenergic neurons can be more easily studied than in situ.

Transmitter expression and morphological development of embryonic medullary and mesencephalic raphé neurones after transplantation to the adult rat central nervous system. II. Grafts to the hippocampus.

Suspensions of cells taken from the mesencephalic or medullary raphé regions of the 13-14 day old embryonic rat brain were injected into the hippocampus of adult rats which had previously been denervated of its serotoninergic input by 5,7-dihydroxytryptamine. At periods of up to 14 months after implantation, the brains were taken for immunohistochemical analysis of 5-hydroxytryptamine (5HT)-, substance P (SP)- and thyrotropin releasing hormone (TRH)-like immunoreactivity. Surviving grafts were found in all animals. The implants derived from mesencephalic raphé contained neurones immunoreactive to 5HT, SP or both substances together. On average 19% of the potential number of mesencephalic 5HT neurones were found in the grafts. Outgrowth of 5HT-immunoreactive fibres was extensive, and displayed the typical pattern of 5HT innervation in the normal hippocampus-the densest plexuses were found in the dentate gyrus, with sparser networks in the CA1 and CA2 regions. SP-positive processes were principally found only in the graft itself. Transplants of medullary raphé cells contained 5HT-immunoreactive neurones, some of which also contained SP-and/or TRH-like immunoreactivity, thereby mirroring the situation found in the caudal raphé complex in situ. An average of 18% of the total available medullary serotoninergic neurones were found at each graft site. A rich outgrowth of 5HT-immunoreactive varicose processes was evident, with the same pattern as the 5HT innervation by the mesencephalic raphé grafts, and as in the normal hippocampus. SP- and TRH-positive fibres were essentially detectable only in the graft, but not in the host hippocampus. The present studies indicate that the milieu of the hippocampus does not preferentially attenuate the survival of the serotoninergic cells which do not normally project to it. Nor is the fibre outgrowth of these medullary raphé 5HT neurones significantly different from that of the mesencephalic raphé. However, the hippocampal environment may be responsible for the appearance of SP-LI in the otherwise apparently solely 5HT-containing mesencephalic raphé neurones, and for repressing the outgrowth of fibres containing TRH-, or SP-like immunoreactivity, regardless of their origin.

Transmitter expression and morphological development of embryonic medullary and mesencephalic raphé neurones after transplantation to the adult rat central nervous system. III. Grafts to the striatum.

Implants have been made of dissociated embryonic mesencephalic or medullary raphé cells into the adult rat striatum, previously depleted of its 5HT innervation. The transmitter complement and fibre outgrowth of the grafted neurones were analysed immunocytochemically. Serotonin-containing cells were found in both types of transplant, and the proportionate survival of the potential number of implanted 5HT cells was similar for each type of graft. However, these proportions were both greater than that described previously in transplants of mesencephalic raphé cells to the spinal cord. In addition, the proportionate survival of medullary substance P neurones grafted to the striatum was greater than that of medullary 5HT cells implanted in the same region. The transmitter complement of the medullary neurones was largely unaltered after transplantation. However, the mesencephalic grafts contained neurones storing 5HT- and/or substance P-, or TRH-like immunoreactivity. The 5HT/substance P and TRH neurones have so far not been encountered in the mesencephalon in situ using the present immunohistochemical methodology. Invasion of the host striatum by 5HT processes from the transplanted mesencephalic cells was extensive. Fibres from medullary raphé neurones, however, were restricted principally to within the graft itself. It is concluded that there may exist in the adult rat striatum a set of trophic factors for 5HT and substance P neurones different from those found in other regions of the central nervous system, such as spinal cord. Moreover, trophic agents in the host striatum appear to operate differentially on mesencephalic and medullary raphé 5HT neurones to regulate their axonal outgrowth. Lastly, the neurotransmitter phenotypic expression of the embryonic mesencephalic raphé cells may be susceptible to influences from the host environment.

Transmitter expression and morphological development of embryonic medullary and mesencephalic raphé neurones after transplantation to the adult rat central nervous system. I. Grafts to the spinal cord.

Suspensions of cells derived from the mesencephalic raphé or medullary raphé regions of the 13-14 day old embryonic rat brain were injected into the spinal cord of adult rats which had been previously denervated with 5,7-dihydroxytryptamine. At periods of up to 12 months after grafting, the spinal cords were taken for immunohistochemical analysis of 5-hydroxytryptamine (5HT), substance P (SP) and thyrotropin releasing hormone (TRH). In nearly all cases, surviving transplants were found. The grafts derived from mesencephalic raphé cells contained neurones which were immunoreactive to 5HT, or SP, but not both together. On average 4% of the total possible number of the available embryonic mesencephalic serotoninergic cells were found. A very dense outgrowth of 5HT positive fibres from the transplant was observed, extending up to 1.5 cm in both the caudal and rostral directions from the graft locus. Some SP immunoreactive fibres were also apparent near the implant. The grafts derived from the medullary transplant also contained 5HT-immunoreactive cells, comprising on average 25% of the total 5HT neurones available from the embryonic medullary primordium. In addition, neurones co-localizing 5HT together with SP and TRH were visible, closely reflecting the situation found in the medullary raphé in situ. Dense plexi of fibres containing 5HT-LI extended both caudally and rostrally up to 12-15 mm from the transplant. Outgrowth of SP and TRH varicose fibres was also demonstrable, although to a lesser degree than for 5HT. It was also possible to find many motoneurones surrounded by varicose fibres containing both 5HT and SP, in contrast to the situation with the mesencephalic grafts, where no such patterns of innervation were seen. The experiments indicate that the milieu of the spinal cord may compromise the survival of mesencephalic raphé 5HT neurones far more than of medullary serotonin cells. However, despite this effect on cell survival, the outgrowth of fibres from the remaining mesencephalic 5HT neurones was apparently unaffected by their ectopic position. Similarly, the transmitter content of both classes of raphé cells was largely unaltered, either by the transplantation process or by the environment into which they were placed. It is concluded that although the adult denervated spinal cord can selectively affect neuronal survival, it is incapable either of inducing in other serotoninergic cells placed within it the transmitter phenotype typical of medullary raphé neurones, or of causing those ectopically located 5HT cells to form connections appropriate to the descending serotonin fibres.

[Transfer of behavior patterns through transplantation of systems of neuroanatomic structures in amphibian larvae. II. Xenoplastic transplantation of hind brain systems between Xenopus laevis (Daud.) as well as Hymenochirus boettgeri (Torn.) and Triturus vulgaris]. / Ubertragung von Verhaltensweisen durich Transplantation von Anlagen neuroanatomischer Strukturen bei Amphibienlarven. II. Xenoplastische Transplantationen von Nachhirnanlagen zwischen Xenopus laevis (Daud.) sowie Hymenochirus boettgeri (Torn.) und Triturus vulgaris.

By replacing parts of the neural plate (prospective medulla oblongata) of Triturus vulgaris by neural ectoderm of Xenopus laevis larval chimaeras were obtained, which showed donorlike rhythmical movements of distinct parts of the visceral musculature according to the place of implantation. Especially the branchial region with its external gills showed rhythmical movements and is the special object of this investigation.