Interneuron Origins in the Embryonic Porcine Medial Ganglionic Eminence.

Interneurons contribute to the complexity of neural circuits and maintenance of normal brain function. Rodent interneurons originate in embryonic ganglionic eminences, but developmental origins in other species are less understood. Here, we show that transcription factor expression patterns in porcine embryonic subpallium are similar to rodents, delineating a distinct medial ganglionic eminence (MGE) progenitor domain. On the basis of Nkx2.1, Lhx6, and Dlx2 expression, in vitro differentiation into neurons expressing GABA, and robust migratory capacity in explant assays, we propose that cortical and hippocampal interneurons originate from a porcine MGE region. Following xenotransplantation into adult male and female rat hippocampus, we further demonstrate that porcine MGE progenitors, like those from rodents, migrate and differentiate into morphologically distinct interneurons expressing GABA. Our findings reveal that basic rules for interneuron development are conserved across species, and that porcine embryonic MGE progenitors could serve as a valuable source for interneuron-based xenotransplantation therapies.SIGNIFICANCE STATEMENT Here we demonstrate that porcine medial ganglionic eminence, like rodents, exhibit a distinct transcriptional and interneuron-specific antibody profile, in vitro migratory capacity and are amenable to xenotransplantation. This is the first comprehensive examination of embryonic interneuron origins in the pig; and because a rich neurodevelopmental literature on embryonic mouse medial ganglionic eminence exists (with some additional characterizations in other species, e.g., monkey and human), our work allows direct neurodevelopmental comparisons with this literature.

Electrophysiological properties of rat nodose ganglion neurons co-transplanted with carotid bodies into the chick chorioallantoic membrane

The electrophysiological properties of nodose ganglion neurons were evaluated immediately after removing nodose ganglia from young adult rats and 3 to 10 days after nodose ganglia implantation _either alone or co-implanted with carotid bodies_ onto the chick chorioallantoic membrane. Implanted and co-implanted nodose neurons were less excitable than acutely recorded nodose neurons. Co-implanted neurons also showed reduced amplitudes for both action potentials and spike after-hyperpolarizations relative to those found in acutely recorded nodose ganglion neurons and a smaller time constant (ô) than that found in implanted neurons. In addition, no spontaneous activity was recorded from nodose ganglion neurons co-implanted with carotid bodies during 3-9 days, which suggests that functional synapses between carotid glomus cells and nodose neurons were not yet established. Results indicate the feasibility of obtaining viable nodose neurons for up to 10 days grafted onto the chick chorioallantoic membrane, where they can conserve most of their passive and active membrane properties and also are susceptible to carotid bodies trophic influences. They also suggest that nodose neurons would need more time for the development of functional synapses when grafted with carotid body glomus cells.

Comparison of reactive processes in the rat brain elicited by xenotransplantation of nervous tissues of chicken or pulmonate snail.

It is known that a histocompatibility system is not developed to the same extent in lower invertebrates as in vertebrate animals. We assumed that the xenografts from the newborn invertebrate nervous system would not exert destructive effects on the brain of the vertebrate recipient even without immunosuppressive therapy. In search of brain xenografts (XG) capable to survive in the brain of a recipient without intensive immunosuppression, we transplanted ganglia of terrestrial snails into the rat brain. We compared effects of transplantation of the XG taken from anterior brain of the 18-day embryo chicken (XGC) and from ganglia of a newborn terrestrial pulmonate snail (Helix aspersa L., XGSn). Part of the XGSn were stained by vital fluorescent dyes Bisbenzimid or Fast Blue before grafting. The XGSn were implanted into the neocortex parenchyma in each hemisphere. Rat brains with the XGC were examined 5 days after, and brains with the XGSn - 5 and 28 days after the transplantation. Nonstained sections with the XGSn labeled with fluorescent dyes prior to transplantation were investigated in fluorescent microscope and stained later with tionin and cresyl-violet. Quantitative videoimage analysis of lymphocyte aggregations, reactive gliosis, morphology of the XG areas, and implantation trace was performed. It was found that the XGSn transplantation did not elicit in the rat brain an intensive immunological conflict 5 and 28 days after transplantation. In contrast, the XGC rapidly elicited a strong immune response resulting in massive obliterations in the rat brain and were rejected in 5 days. Labeled snail glia and vessels were observed in the stained XGSn 28 days after transplantation by fluorescence imaging. Putative snail vessels grew into the rat brain from the place of snail tissue transplantation serving the humoral integration of the XG and the host brain. Migration of molluscan glial cells was observed in the brain of recipients.

In utero fate mapping reveals distinct migratory pathways and fates of neurons born in the mammalian basal forebrain.

Recent studies suggest that neurons born in the developing basal forebrain migrate long distances perpendicularly to radial glia and that many of these cells reach the developing neocortex. This form of tangential migration, however, has not been demonstrated in vivo, and the sites of origin, pathways of migration and final destinations of these neurons in the postnatal brain are not fully understood. Using ultrasound-guided transplantation in utero, we have mapped the migratory pathways and fates of cells born in the lateral and medial ganglionic eminences (LGE and MGE) in 13.5-day-old mouse embryos. We demonstrate that LGE and MGE cells migrate along different routes to populate distinct regions in the developing brain. We show that LGE cells migrate ventrally and anteriorly, and give rise to the projecting medium spiny neurons in the striatum, nucleus accumbens and olfactory tubercle, and to granule and periglomerular cells in the olfactory bulb. By contrast, we show that the MGE is a major source of neurons migrating dorsally and invading the developing neocortex. MGE cells migrate into the neocortex via the neocortical subventricular zone and differentiate into the transient subpial granule neurons in the marginal zone and into a stable population of GABA-, parvalbumin- or somatostatin-expressing interneurons throughout the cortical plate.

Phenotypic development of the human embryonic striatal primordium: a study of cultured and grafted neurons from the lateral and medial ganglionic eminences.

Basic parameters which are crucial for the survival of human embryonic striatal grafts need to be investigated before initiating clinical trials in Huntington's disease. In order to define the dissection of human striatal-donor tissue which gives rise to the largest amount of striatal neurons after intrastriatal transplantation, we studied the lateral and medial ganglionic eminences of embryonic striatal primordia obtained from human embryos sized 17-30 mm in crown-to-rump length (corresponding to Carnegie stages 18-23). Anatomical landmarks that demarcated the lateral and medial ganglionic eminences from each other were present only in embryos with 20 mm crown-to-rump length or larger. In monolayer cultures, the lateral ganglionic eminence gave rise to a six-fold higher yield of dopamine- and cyclic AMP-regulated phosphoprotein 32-immunoreactive striatal neurons as compared to the medial ganglionic eminence. We also xenografted the lateral and medial ganglionic eminences from five embryos sized 21-30 mm in crown-to-rump length to the ibotenate lesioned striatum of immunosuppressed rats. The grafts were evaluated with respect to general morphology, survival and integration using (immuno-) histochemical stains for acetylcholinesterase/Cresyl Violet, nicotinamide adenine dinucleotide phosphate-diaphorase, dopamine- and cyclic AMP-regulated phosphoprotein-32, tyrosine hydroxylase and calbindin-D28KD. As assessed 9-25 weeks after implantation, 13 out of 16 and 8 out of 13 grafts, in the groups grafted with the medial and lateral ganglionic eminences, respectively, had survived. Previous studies with rat donor tissue have indicated that the functional efficacy of striatal grafts is related to the development of striatal-specific P-zone regions and that these are enriched in transplants derived from the lateral as opposed to the medial ganglionic eminence. Also in the human striatal xenografts of the present study, P-zones appeared more abundant when the donor tissue was derived from the lateral ganglionic eminence. However, the proportion of graft tissue that expressed P-zone properties was always very low (at most 30%) and never approached the 80-90% previously observed in transplants of rat lateral ganglionic eminence. We conclude that the relative yield of striatal neurons in grafts of the human embryonic striatal primordium has to be improved before neural transplantation should be applied in patients with Huntington's disease.

Maintenance of sympathetic innervation into the hippocampal formation requires a continuous local availability of nerve growth factor.

The sprouting of peripheral sympathetic fibers into the septally denervated hippocampal formation is a well-characterized model of lesion-induced plasticity. While various studies have demonstrated the importance of nerve growth factor for evoking sympathetic sprouting, little is known concerning whether nerve growth factor continues to be required for maintaining innervation once it has occurred. In the present study we have addressed this point by (i) investigating the consequences of withdrawing exogenous nerve growth factor support from rats in which sympathetic innervation was enhanced by a nerve growth factor infusion and (ii) using blocking antibodies to interfere with the actions of endogenous nerve growth factor. The results of this investigation clearly indicate that a continuous supply of nerve growth factor (either exogenous or endogenous) is required to maintain sympathetic innervation within the hippocampal formation. Evidence is also provided demonstrating that the nerve growth factor must be made available locally within a given region to evoke and maintain the sympathetic innervation within this location. Axonal rearrangement within the developing and adult brain is believed to be an important mechanism underlying learning and memory is crucial for lesion-related plasticity. In various experimental paradigms, nerve growth factor has been shown to be an important cue for initiating axonal remodeling. In the current study, we have demonstrated that once such rearrangements have taken place, nerve growth factor may also be required to maintain them.

Paths taken sensory nerve fibres in aneural chick wing buds.

What constrains growing nerves to follow the paths they take during the development of peripheral nerve patterns? This paper examines two, related, topics concerning the pathways taken by sensory nerve fibres in the embryo chick wing: the constraints imposed on the nerves by limb tissues; and the timing of axon outgrowth. Sensory ganglia from 7-day-old chick embryos were grafted into younger host embryo wing buds which had been previously denervated. The resultant nerve patterns revealed that, first, nerve fibres could grow almost anywhere within the wing bud, with the exceptions of cartilage and a region just beneath the growing tip. Secondly, the younger the host wing bud at the time of grafting, the more likely the neurites were to form a thick fascicle which followed the limb's normal nerve pathways. The wing apparently does not impose a rigid restraint on nerves to grow only along certain routes; however, if a nerve fibre reaches a normal nerve pathway, it prefers to follow it.

Absence of sympathetic sprouting in the rat olfactory bulb after cholinergic denervation.

Sympathetic axons have been shown to invade the rat hippocampal formation after lesions of the septohippocampal (cholinergic) projection. To test the generality of this type of transmitter-specific sprouting within the central nervous system, lesions were placed within the horizontal limb of the diagonal band in order to denervate the main olfactory bulb (MOB) of its cholinergic innervation. Some animals also received autologous transplants of the superior cervical ganglion to the region of the MOB. There was no evidence of sympathetic axon growth into the MOB under any circumstance as assessed with qualitative fluorescence histochemistry. In addition, retrograde fluorescent dye studies demonstrated that the basal forebrain neurons projecting to the neocortex, where sympathetic sprouting has also been reported, are a separate population from those which project to the MOB. Although it is difficult to unequivocally interpret negative results, these data indicate that cholinergic denervation does not induce sympathetic sprouting throughout the central nervous system. The availability of a brain region where sympathetic sprouting does not occur may provide clues to understanding the conditions which permit such neuronal plasticity in the hippocampal formation.

Neuronotrophic activity in brain wounds of the developing rat. Correlation with implant survival in the wound cavity.

Neuronotrophic activity accumulates in a wound cavity created in the entorhinal/occipital cortex of developing rats. These trophic factors support the survival of neurons in monolayer cultures of chick embryo spinal cord, ciliary ganglion, sympathetic ganglion and dorsal root ganglion, as well as of mouse dorsal root ganglion. Trophic activity was very low both in non-injured brain tissue and in the wound cavity 1 day post-lesion, but it increased 15- to 300-fold during the subsequent 2-5 days. Together with the trophic activity in the wound fluid were other substances which interfered with the survival of spinal cord neurons. The neuronotrophic factors appeared to be proteins immunologically distinct from mouse submaxillary nerve growth factor. Fragments of rat embryo corpus striatum placed in the cortical wound cavity immediately after its formation showed very poor subsequent survival and no innervation of the host hippocampus. However, if implantation was delayed by 3 or 6 days with respect to the time at which the receiving cavity was made, the survival was greatly improved and innervation of the host took place. The time course for the accumulation of the trophic factors in the cavity paralleled the delay leading to increased survival of brain grafts. It is suggested that the neuronotrophic activity accumulating in the wound cavity during the delay period may be responsible for the increased survival of the implants.

Stimulation of spermatogonial DNA synthesis in slug gonad by a factor released from cerebral ganglia under the influence of long days.

Incorporation of [3H]thymidine into gonadal DNA was shown to increase 1 week after implantation into an immature slug (Limax maximus) of a "brain" (circumesophageal ring of ganglia) from a male-phase donor. Light microscope autoradiography revealed that in stimulated gonads labeling was localized primarily in the nuclei of spermatagonia. Implant-stimulated spermatogonial DNA synthesis was found to depend upon implantation of supraesophageal (cerebral) ganglia. Neither subesophageal ganglia implants nor immature supraesophageal implants had an effect. Thymidine incorporation could also be stimulated by exposure of slugs to long-day lightcycles (LD 16:8) for 3 to 4 weeks. Similar duration of long-day treatment was also adequate to trigger male-phase development even after animals were returned to short days (LD 8:16). The results are consistent with the view that 3 to 4 weeks of long-day lightcycles are required to promote irreversibly the release from slug cerebral ganglia of a male-phase gonadotropic factor which directly or indirectly promotes spermatogonial proliferation.

[Transplantation of nerve tissue]. / Transplantatsiia nervnoi tkani.

The results of transplantation of various parts of the central and peripheral nervous system are considered. Transplantation of nerve trunks is used clinically, and heterogenous regeneration of the nerves results in reinnervation of tissues and organs. The spinal ganglion transplantation is successfully used in experiments with both embryonic and mature differentiated neurons. Transplantation of different parts of the cortex, some subcortical structures, hyppocampus, hypothalamus, cerebellum and the spinal cord is made using immature neurons. Some attempts have been made to transplant the nerve tissue grown in vitro into a host.

Adrenergic differentiation of cells of the cholinergic ciliary and Remak ganglia in avian embryo after in vivo transplantation.

We have previously shown that the neural crest is regionalized early into "adrenergic" and "cholinergic" areas from which arise, respectively, the sympathetic and parasympathetic ganglioblasts of the autonomic nervous system. This regionalization does not correspond, however, to an irreversible determination of the neural crest cells since, under certain experimental conditions, cholinergic cells can arise from the adrenergic region of the crest and vice versa. The phenotypic expression of the presumptive ganglion cells appears to be responsive to the environmental conditions they encounter during and/or after their migration. In the present study we show that the developmental behavior of parasympathetic ganglion cells which have stopped migrating and at least some of which have started to differentiate into cholinergic neurons can be profoundly modified if they are transplanted into a younger embryo at the trunk neural crest level. The crest level. The grafted ganglion cells start migrating and stop in the same sites as the host neural crest cells. Their further differentiation depends on their localization. When situated in the adrenergic ganglia and in the suprarenal gland they synthesize catehcolamines, whereas they differentiate into nonfluorescent, silver-staining ganglion cells if they migrate in the gut wall. Thus, the differentiation of autonomic neurons is dependent on tissue interactions even after the neural crest cells have grouped to form ganglionic structures in which biochemical differentiation is already in progress.

Target discrimination in regenerating insect sensory nerve.

The paired abdominal cerci of the cricket Acheta domesticus are mechanosensory appendages which regenerate readily when amputated during larval life. Their peripherally-located sense cells form axons which project centrally as a purely sensory nerve to the terminal abdominal ganglion. In an attempt to analyze some of the factors which guide a regenerating sensory nerve to correct central terminations, implants of homologous, supernumerary terminal ganglia were made in cricket larvae and the host cerci amputated. The possibility that implants with multiple nerve stumps might release an attracting substance was considered. Surgical procedures used were (1) implant in posterior abdomen; (2) implant in posterior abdomen, ipsilateral to chronic cercal deprivation; (3) implant in mesothoracic leg socket, adjacent to heterotopically-transplanted regenerated cercus; (4) implant in posterior abdomen, ipsilateral host cercal motor nerve sectioned; (5) implant in posterior abdomen, ipsilateral margin of host terminal ganglion wounded. Results were determined after the adult molt, by conventional histology or by cobalt chloride filling of regenerated cercal nerves. In all procedures except (3) and (4), the regenerated afferent nerve bypassed the implant and terminated in the host terminal ganglion. In (3), the regenerated fibers from cercal grafts bypassed the implant; terminations were not found. In (4), some regenerated cercal axons connected with the implant and the majority terminated in the host ganglion. It is suggested that regenerating cercal afferents may depend in a facultative way on the cercal motor nerve as a pathway guide but there is as yet no clear evidence for a trophic influence from the central nervous system.

Brain tissue transplanted to the anterior chamber of the eye: 3. Substitution of lacking central noradrenaline input by host iris sympathetic fibers in the isolated cerebral cortex developed in oculo.

Fetal parietal cerebral cortex was homologously transplanted to the anterior chambers of the eyes of adult rats. The transplants got vascularized, proliferated, as measured by in vivo stereoscopic inspections, and differentiated into brain tissue similar to cortex cerebri in situ and survived for long times, greater than 41/2 months. Fibers from the intact sympathetic adrenergic ground plexus of the iris were able to innervate the transplants in an organotypic way regarding fluorescence morphology, pattern of distribution of the nerve terminals and, to a certain extent, density of innervation, the only variable parameter being density of innervation. Thus, in unpretreated or MAO inhibited transplants only rather few to scattered terminals could be found, while after preincubation in 10(-5)M alpha-methyl-noradrenaline the number of visible terminals was normal or slightly less than normal, as compared to cortex cerebri in situ. When superior cervical ganglia (SCG) were transplanted together with fetal cortex tissue to sympathetically denervated eyes the ingrowth in the cortex tissue was similar to that after single cortex transplantation combined with 5 day old SCG, while a marked hyperinnervation was encountered when combined with adult SCG. It is concluded that the developing cortex cerebri, deprived of its normal CNS source of adrenergic nerves, is able to receive sympathetic adrenergic nerves from the iris in an organotypic way upon transplantation to the anterior chamber of the eye.