Electroacupuncture Promotes the Survival of the Grafted Human MGE Neural Progenitors in Rats with Cerebral Ischemia by Promoting Angiogenesis and Inhibiting Inflammation.

Stem cells have the potential as a regenerative therapy for cerebral ischemia by improving functional outcomes. However, cell transplantation has some limitations, including a low rate of the grafted cell survival. There is still a major challenge of promoting the harmonious symbiosis between grafted cells and the host. Acupuncture can effectively improve the functional outcome after cerebral ischemia. The present study evaluated the therapeutic effects and explored the mechanism of combined medial ganglionic eminence (MGE) neural progenitors differentiated from human embryonic stem cells (hESCs) with electroacupuncture (EA) in a bilateral common carotid artery occlusion (2VO) rat model. The results showed that EA could promote the survival of the grafted MGE neural progenitors differentiated from hESCs and alleviate learning and memory impairment in rats with cerebral ischemia. This may have partially resulted from inhibited expression of TNF-α and IL-1ß and increased vascular endothelial growth factor (VEGF) expression and blood vessel density in the hippocampus. Our findings indicated that EA could promote the survival of the grafted MGE neural progenitors and enhance transplantation therapy's efficacy by promoting angiogenesis and inhibiting inflammation.

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.

Transplanted interneurons improve memory precision after traumatic brain injury.

Repair of the traumatically injured brain has been envisioned for decades, but regenerating new neurons at the site of brain injury has been challenging. We show GABAergic progenitors, derived from the embryonic medial ganglionic eminence, migrate long distances following transplantation into the hippocampus of adult mice with traumatic brain injury, functionally integrate as mature inhibitory interneurons and restore post-traumatic decreases in synaptic inhibition. Grafted animals had improvements in memory precision that were reversed by chemogenetic silencing of the transplanted neurons and a long-lasting reduction in spontaneous seizures. Our results reveal a striking ability of transplanted interneurons for incorporating into injured brain circuits, and this approach is a powerful therapeutic strategy for correcting post-traumatic memory and seizure disorders.

Transplantation of inhibitory precursor cells from medial ganglionic eminence produces distinct responses in two different models of acute seizure induction.

Medial ganglionic eminence (MGE) is one of the sources of inhibitory interneurons during development. Following transplantation in postnatal developing brain, MGE cells can increase local inhibition suggesting a possible protection to GABAergic dysfunction in brain disorders, such as epilepsy. Since it has been shown that MGE-derived cells harvested as neurospheres are able to suppress seizures, it might be important to investigate whether these protective effects would change in different seizure models. Here, we used pentylenetetrazole-(PTZ) and maximal electroshock (MES)-induced seizure models to test whether the transplantation of MGE cells would increase the threshold to trigger acute seizures. When transplanted into the neocortex (layers 3-4) of neonatal mice (postnatal days 3-4), MGE cells were able to survive and were mainly found in piriform cortex, fimbria, and ventricular wall regions. Additionally, the number of GFP+ cells found in the brains of mice induced with PTZ and MES differed significantly and suggests proliferation and larger survival rate of MGE-transplanted cells after PTZ, but not MES-induced seizures. Following transplantation, there was a reduction in the number of animals presenting mild and severe seizures induced by PTZ. Furthermore, MGE-cell transplantation was able to increase threshold to seizures induced by PTZ, but was not able to prevent seizure spread induced by MES.

Direct Comparison of Rat- and Human-Derived Ganglionic Eminence Tissue Grafts on Motor Function.

Huntington's disease (HD) is a debilitating, genetically inherited neurodegenerative disorder that results in early loss of medium spiny neurons from the striatum and subsequent degeneration of cortical and other subcortical brain regions. Behavioral changes manifest as a range of motor, cognitive, and neuropsychiatric impairments. It has been established that replacement of the degenerated medium spiny neurons with rat-derived fetal whole ganglionic eminence (rWGE) tissue can alleviate motor and cognitive deficits in preclinical rodent models of HD. However, clinical application of this cell replacement therapy requires the use of human-derived (hWGE), not rWGE, tissue. Despite this, little is currently known about the functional efficacy of hWGE. The aim of this study was to directly compare the ability of the gold standard rWGE grafts, against the clinically relevant hWGE grafts, on a range of behavioral tests of motor function. Lister hooded rats either remained as unoperated controls or received unilateral excitotoxic lesions of the lateral neostriatum. Subsets of lesioned rats then received transplants of either rWGE or hWGE primary fetal tissue into the lateral striatum. All rats were tested postlesion and postgraft on the following tests of motor function: staircase test, apomorphine-induced rotation, cylinder test, adjusting steps test, and vibrissae-evoked touch test. At 21 weeks postgraft, brain tissue was taken for histological analysis. The results revealed comparable improvements in apomorphine-induced rotational bias and the vibrissae test, despite larger graft volumes in the hWGE cohort. hWGE grafts, but not rWGE grafts, stabilized behavioral performance on the adjusting steps test. These results have implications for clinical application of cell replacement therapies, as well as providing a foundation for the development of stem cell-derived cell therapy products.

Viral-mediated Labeling and Transplantation of Medial Ganglionic Eminence (MGE) Cells for In Vivo Studies.

GABAergic cortical interneurons, derived from the embryonic medial and caudal ganglionic eminences (MGE and CGE), are functionally and morphologically diverse. Inroads have been made in understanding the roles of distinct cortical interneuron subgroups, however, there are still many mechanisms to be worked out that may contribute to the development and maturation of different types of GABAergic cells. Moreover, altered GABAergic signaling may contribute to phenotypes of autism, schizophrenia and epilepsy. Specific Cre-driver lines have begun to parcel out the functions of unique interneuron subgroups. Despite the advances in mouse models, it is often difficult to efficiently study GABAergic cortical interneuron progenitors with molecular approaches in vivo. One important technique used to study the cell autonomous programming of these cells is transplantation of MGE cells into host cortices. These transplanted cells migrate extensively, differentiate, and functionally integrate. In addition, MGE cells can be efficiently transduced with lentivirus immediately prior to transplantation, allowing for a multitude of molecular approaches. Here we detail a protocol to efficiently transduce MGE cells before transplantation for in vivo analysis, using available Cre-driver lines and Cre-dependent expression vectors. This approach is advantageous because it combines precise genetic manipulation with the ability of these cells to disperse after transplantation, permitting greater cell-type specific resolution in vivo.

Long-lasting anxiolytic effect of neural precursor cells freshly prepared but not neurosphere-derived cell transplantation in newborn rats.

BACKGROUND: The GABAergic system plays an important role in modulating levels of anxiety. When transplanted into the brain, precursor cells from the medial ganglionic eminence (MGE) have the ability to differentiate into GABAergic interneurons and modify the inhibitory tone in the host brain. Currently, two methods have been reported for obtaining MGE precursor cells for transplantation: fresh and neurosphere dissociated cells. Here, we investigated the effects generated by transplantation of the two types of cell preparations on anxiety behavior in rats. RESULTS: We transplanted freshly dissociated or neurosphere dissociated cells into the neonate brain of male rats on postnatal (PN) day 2-3. At early adulthood (PN 62-63), transplanted animals were tested in the Elevated Plus Maze (EPM). To verify the differentiation and migration pattern of the transplanted cells in vitro and in vivo, we performed immunohistochemistry for GFP and several interneuron-specific markers: neuropeptide Y (NPY), parvalbumin (PV) and calretinin (CR). Cells from both types of preparations expressed these interneuronal markers. However, an anxiolytic effect on behavior in the EPM was observed in animals that received the MGE-derived freshly dissociated cells but not in those that received the neurosphere dissociated cells. CONCLUSION: Our results suggest a long-lasting anxiolytic effect of transplanted freshly dissociated cells that reinforces the inhibitory function of the GABAergic neuronal circuitry in the hippocampus related to anxiety-like behavior in rats.

GABAergic interneuron migration and the evolution of the neocortex.

A neocortex is present in all mammals but is not present in other classes of vertebrates, and the neocortex is extremely elaborate in humans. Changes in excitatory projection neurons and their progenitors within the developing dorsal pallium in the most recent common ancestor of mammals are thought to have been involved in the evolution of the neocortex. Our recent findings suggest that changes in the migratory ability of inhibitory interneurons derived from outside the neocortex may also have been involved in the evolution of the neocortex. In this article we review the literature on the migratory profile of inhibitory interneurons in several different species and the literature on comparisons between the intrinsic migratory ability of interneurons derived from different species. Finally, we propose a hypothesis about the mammalian-specific evolution of the migratory ability of interneurons and its potential contribution to the establishment of a functional neocortex.

Antibodies obtained by xenotransplantation of organ-cultured median eminence specifically recognize hypothalamic tanycytes.

Tanycytes are specialized ependymal cells lining the infundibular recess of the third ventricle of the cerebrum. Early and recent investigations involve tanycytes in the mechanism of gonadotropin-releasing hormone (GnRH) release to the portal blood. The present investigation was performed to obtain a specific immunological marker of tanycytes and to identify the compound(s) responsible for this labeling. After 30 days of organ culture, explants of bovine median eminence formed spherical structures mostly constituted by tanycytes. These tanycyte spheres were xenotransplanted to rats, and the antibodies raised by the host animals against the transplanted living tanycytes were used for immunochemical studies of the bovine and rat median eminence. This antiserum immunoreacted with two compounds of 60 kDa and 85 kDa present in extracts of bovine and rat median eminence. The individual immunoblotting analysis of rat medial basal hypothalami showed a decrease in the amount of the 85-kDa compound in castrated rats as compared to control rats processed at oestrus and dioestrus. The antiserum, labeled as anti-P85, when used for immunostaining of sections throughout the rat central nervous system, immunoreacted specifically with the hypothalamic tanycytes. Within tanycytes, P-85 immunoreactivity was exclusively present in the basal processes. It is suggested that the 85-kDa and 60-kDa compounds correspond to two novel proteins selectively expressed by tanycytes. The possibility that they are secretory proteins involved in GnRH release is discussed. Anti-P85 appears to be the first specific marker of hypothalamic tanycytes.

Alleviation of estrogen-induced hyperprolactinemia through intracerebral transplantation of hypothalamic tissue containing dopaminergic neurons.

Prolactin-secreting pituitary tumors can be induced in young rats through prolonged estrogen treatment. Recent evidence suggests that such tumors are associated with a degeneration of tuberoinfundibular dopaminergic (TI-DA) neurons, which normally inhibit prolactin secretion by the anterior pituitary's lactrophs. For this study, chronic hyperprolactinemia was induced in young, ovariectomized Fisher 344 rats through Silastic capsule implants of 17 beta-estradiol, placed subcutaneously for 1 month prior to removal. Rats with such estrogen-induced hyperprolactinemia then received transplants of neonatal arcuate-median eminence (ME) tissue (containing TI-DA neurons) or amygdala (control) tissue, placed either within the third ventricle or bilaterally within the hypothalamus. Blood samples were obtained 1 month after transplantation and prolactin concentrations measured by radioimmunoassay. Two of 4 animals receiving ventricularly-placed arcuate-ME transplants and 4 of 7 animals receiving bilateral arcuate-ME transplants showed substantial reductions in plasma prolactin levels compared to mean values in control animals. Follow-up catecholamine (CA) histochemistry indicated a bright fluorescence intensity in the median eminence of animals remaining hyperprolactinemic with ineffective transplants. Furthermore, in sharp contrast to the very low, nonpulsatile LH levels found during a second bleeding in recipients bearing ineffective transplants, recipients with effective arcuate-ME transplants had the high, pulsatile levels of LH characteristic of normal, ovariectomized rats. These data suggest that developing TI-DA neurons, within effective arcuate-ME transplants, became functional to reinstate or accentuate DA inhibition of prolactin secretion and, in so doing, indirectly normalized LH secretion as well.

Ultrastructure of homografts of the rat median eminence into the anterior chamber of the eye.

Median eminences (MEs) were transplanted into the anterior chamber of the eye and collected at times varying between 10 and 72 days. At 10 days, all axons had degenerated leaving behind a well-vascularized epitheloid glial organ. With increasing age of the grafts, the characteristic lipid inclusions in the glial cells diminished and even disappeared, and the clear ultrastructural distinction between ependymal (tanycytes) and other glial cells became difficult or even impossible. In both the perikarya and perivascular terminals, an increased number of membrane-bounded, pleomorphic, electron-dense granulated vesicles occurred, the significance of which is unknown. Regrafting of these transplants into various hypothalamic and hypophysial sites is expected to yield information on the function of the glial cells of the ME.