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Context: Understanding of central nervous system mechanisms related to age-related infertility remains limited. Fibril α-synuclein, distinct from its monomer form, is implicated in age-related diseases and propagates among neurons akin to prions. Aims: We compared α-synuclein expression in gonadotropin-releasing hormone-expressing neurons (GnRH neurons) in the pre-optic area, arcuate nucleus, and median eminence of healthy heifers and aged cows to determine its role in age-related infertility. Methods: We analysed mRNA and protein expression, along with fluorescent immunohistochemistry for GnRH and α-synuclein, followed by Congo red staining to detect amyloid deposits, and confocal microscopy. Key results: Both mRNA and protein expressions of α-synuclein were confirmed by reverse transcription-polymerase chain reaction (RT-PCR) and western blots in bovine cortex, hippocampus, and anterior and posterior hypothalamus tissues. Significant differences in α-synuclein mRNA expression were observed in the cortex and hippocampus between young and old cows. Western blots showed five bands of α-synuclein, probably reflecting monomer, dimer, and oligomers, in the cortex, hippocampus, hypothalamus tissues, and there were significant differences in some bands between young and old cows. Bright-field and polarised light microscopy did not detect obvious amyloid deposition in aged hypothalami; however, higher-sensitive confocal microscopy unveiled strong positive signal of Congo red and α-synuclein in GnRH neurons in aged hypothalami. Additionally, α-synuclein expression was detected in immortalised GnRH neurons, GT1-7 cells. Conclusion: Alpha-synuclein was expressed in GnRH neurons, and some differences were observed between young and old hypothalami. Implications: Alpha-synuclein may play an important role in aging-related infertility. In our investigation into age-related infertility, we examined the presence of α-synuclein, a protein associated with brain aging, particularly in gonadotropin-releasing hormone (GnRH) neurons in hypothalami. Contrasting samples from young and old cows, we observed significant differences in α-synuclein levels across the specific hypothalamus regions linked to reproductive function. These findings suggest that α-synuclein may play a pivotal role in the mechanisms underlying age-related infertility, paving the way for further inquiries into potential therapeutic avenues in reproductive health. Image by Hiroya Kadokawa. [ABSTRACT FROM AUTHOR]

Background: Glucagon-like peptide 1 (GLP-1) is involved in the regulation of energy and glucose homeostasis. As GLP-1 has similar effects on the energy homeostasis as the hypophysiotropic thyrotropin-releasing hormone (TRH) neurons that regulate the hypothalamic–pituitary–thyroid (HPT) axis, we raised the possibility that the TRH neurons are involved in the mediation of the effects of GLP-1. Therefore, the relationship and interaction of the GLP-1 system and the TRH neurons of the hypothalamic paraventricular nucleus (PVN) were studied. Methods: To examine the anatomical and functional relationship of TRH neurons and the GLP-1 system in the PVN, immunocytochemistry, in situ hybridization, in vitro patch-clamp electrophysiology, metabolic phenotyping, and explant experiments were performed. Results: Our data demonstrate that the TRH neurons of the PVN are innervated by GLP-1 producing neurons and express the GLP-1 receptor (GLP-1R). However, not only do the GLP-1-innervated TRH neurons express GLP-1R but the receptor is also present in the axons of the hypophysiotropic TRH neurons in the blood–brain barrier free median eminence (ME) suggesting that peripherally derived GLP-1 may also influence the TRH neurons. In vitro, GLP-1 increased the firing rate of TRH neurons and depolarized them. In addition, GLP-1 directly stimulated the GABAergic input of a population of TRH neurons. Furthermore, GLP-1 inhibited the release of TRH from the hypophysiotropic axons in the ME. In vivo, peripheral GLP-1R agonist administration markedly inhibited the food intake and the energy expenditure, but had no effect on the TRH expression in the PVN and resulted in lower circulating free T4 levels. Conclusions: Our results indicate that GLP-1R activation has a direct stimulatory effect on TRH neurons in the PVN, but the activation of GLP-1R may also inhibit TRH neurons by facilitating their inhibitory inputs or by inhibiting the axon terminals of these cells in the ME. The innervation of TRH neurons by GLP-1 neurons suggests that TRH neurons might be influenced by both circulating GLP-1 and by GLP-1 neurons of the nucleus tractus solitarii. The lack of GLP-1R agonist-induced regulation of TRH neurons in vivo suggests that the HPT axis does not mediate the GLP-1R agonist-induced weight loss. [ABSTRACT FROM AUTHOR]

The human cortex contains inhibitory interneurons derived from the medial ganglionic eminence (MGE), a germinal zone in the embryonic ventral forebrain. How this germinal zone generates sufficient interneurons for the human brain remains unclear. We found that the human MGE (hMGE) contains nests of proliferative neuroblasts with ultrastructural and transcriptomic features that distinguish them from other progenitors in the hMGE. When dissociated hMGE cells are transplanted into the neonatal mouse brain, they reform into nests containing proliferating neuroblasts that generate young neurons that migrate extensively into the mouse forebrain and mature into different subtypes of functional interneurons. Together, these results indicate that the nest organization and sustained proliferation of neuroblasts in the hMGE provide a mechanism for the extended production of interneurons for the human forebrain.

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.

Neurofibromatosis 1 (NF1) is caused by mutations in the NF1 gene, which encodes the protein, neurofibromin, an inhibitor of Ras activity. Cortical GABAergic interneurons (CINs) are implicated in NF1 pathology, but the cellular and molecular changes to CINs are unknown. We deleted mouse Nf1 from the medial ganglionic eminence, which gives rise to both oligodendrocytes and CINs that express somatostatin and parvalbumin. Nf1 loss led to a persistence of immature oligodendrocytes that prevented later-generated oligodendrocytes from occupying the cortex. Moreover, molecular and cellular properties of parvalbumin (PV)-positive CINs were altered by the loss of Nf1, without changes in somatostatin (SST)-positive CINs. We discovered that loss of Nf1 results in a dose-dependent decrease in Lhx6 expression, the transcription factor necessary to establish SST+ and PV+ CINs, which was rescued by the MEK inhibitor SL327, revealing a mechanism whereby a neurofibromin/Ras/MEK pathway regulates a critical CIN developmental milestone.

Manipulations that enhance GABAergic inhibition have been associated with improved behavioral phenotypes in autism models, suggesting that autism may be treated by correcting underlying deficits of inhibition. Interneuron transplantation is a method for increasing recipient synaptic inhibition, and it has been considered a prospective therapy for conditions marked by deficient inhibition, including neuropsychiatric disorders. It is unknown, however, whether interneuron transplantation may be therapeutically effective only for conditions marked by reduced inhibition, and it is also unclear whether transplantation improves behavioral phenotypes solely by normalizing underlying circuit defects. To address these questions, we studied the effects of interneuron transplantation in male and female mice lacking the autism-associated gene, Pten, in GABAergic interneurons. Pten mutant mice exhibit social behavior deficits, elevated synaptic inhibition in prefrontal cortex, abnormal baseline and social interaction-evoked electroencephalogram (EEG) signals, and an altered composition of cortical interneuron subtypes. Transplantation of wild-type embryonic interneurons from the medial ganglionic eminence into the prefrontal cortex of neonatal Pten mutants rescued social behavior despite exacerbating excessive levels of synaptic inhibition. Furthermore, transplantation did not normalize recipient EEG signals measured during baseline states. Interneuron transplantation can thus correct behavioral deficits even when those deficits are associated with elevated synaptic inhibition. Moreover, transplantation does not exert therapeutic effects solely by restoring wild-type circuit states. Our findings indicate that interneuron transplantation could offer a novel cell-based approach to autism treatment while challenging assumptions that effective therapies must reverse underlying circuit defects.SIGNIFICANCE STATEMENT Imbalances between neural excitation and inhibition are hypothesized to contribute to the pathophysiology of autism. Interneuron transplantation is a method for altering recipient inhibition, and it has been considered a prospective therapy for neuropsychiatric disorders, including autism. Here we examined the behavioral and physiological effects of interneuron transplantation in a mouse genetic model of autism. They demonstrate that transplantation rescues recipient social interaction deficits without correcting a common measure of recipient inhibition, or circuit-level physiological measures. These findings demonstrate that interneuron transplantation can exert therapeutic behavioral effects without necessarily restoring wild-type circuit states, while highlighting the potential of interneuron transplantation as an autism therapy.

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.

Immature neurons generated by the subpallial MGE tangentially migrate to the cortex where they become parvalbumin-expressing (PV+) and somatostatin (SST+) interneurons. Here, we show that the Sp9 transcription factor controls the development of MGE-derived cortical interneurons. SP9 is expressed in the MGE subventricular zone and in MGE-derived migrating interneurons. Sp9 null and conditional mutant mice have approximately 50% reduction of MGE-derived cortical interneurons, an ectopic aggregation of MGE-derived neurons in the embryonic ventral telencephalon, and an increased ratio of SST+/PV+ cortical interneurons. RNA-Seq and SP9 ChIP-Seq reveal that SP9 regulates MGE-derived cortical interneuron development through controlling the expression of key transcription factors Arx, Lhx6, Lhx8, Nkx2-1, and Zeb2 involved in interneuron development, as well as genes implicated in regulating interneuron migration Ackr3, Epha3, and St18. Thus, Sp9 has a central transcriptional role in MGE-derived cortical interneuron development.

The CCCTC-binding factor (CTCF) is a central regulator of chromatin topology recently linked to neurodevelopmental disorders such as intellectual disability, autism, and schizophrenia. The aim of this study was to identify novel roles of CTCF in the developing mouse brain. We provide evidence that CTCF is required for the expression of the LIM homeodomain factor LHX6 involved in fate determination of cortical interneurons (CINs) that originate in the medial ganglionic eminence (MGE). Conditional Ctcf ablation in the MGE of mice of either sex leads to delayed tangential migration, abnormal distribution of CIN in the neocortex, a marked reduction of CINs expressing parvalbumin and somatostatin (Sst), and an increased number of MGE-derived cells expressing Lhx8 and other markers of basal forebrain projection neurons. Likewise, Ctcf-null MGE cells transplanted into the cortex of wild-type hosts generate fewer Sst-expressing CINs and exhibit lamination defects that are efficiently rescued upon reexpression of LHX6. Collectively, these data indicate that CTCF regulates the dichotomy between Lhx6 and Lhx8 to achieve correct specification and migration of MGE-derived CINs.SIGNIFICANCE STATEMENT This work provides evidence that CCCTC-binding factor (CTCF) controls an early fate decision point in the generation of cortical interneurons mediated at least in part by Lhx6. Importantly, the abnormalities described could reflect early molecular and cellular events that contribute to human neurological disorders previously linked to CTCF, including schizophrenia, autism, and intellectual disability.

Mafb and c-Maf transcription factor (TF) expression is enriched in medial ganglionic eminence (MGE) lineages, beginning in late-secondary progenitors and continuing into mature parvalbumin (PV+) and somatostatin (SST+) interneurons. However, the functions of Maf TFs in MGE development remain to be elucidated. Herein, Mafb and c-Maf were conditionally deleted, alone and together, in the MGE and its lineages. Analyses of Maf mutant mice revealed redundant functions of Mafb and c-Maf in secondary MGE progenitors, where they repress the generation of SST+ cortical and hippocampal interneurons. By contrast, Mafb and c-Maf have distinct roles in postnatal cortical interneuron (CIN) morphological maturation, synaptogenesis, and cortical circuit integration. Thus, Mafb and c-Maf have redundant and opposing functions at different steps in CIN development.

Objective: Oligodendrocyte progenitor cell differentiation is regulated by nutritional signals in the adult median eminence (ME), but the consequences on local myelination are unknown. The aim of this study was to characterize myelin plasticity in the ME of adult mice in health or in response to chronic nutritional challenge and determine its relevance to the regulation of energy balance. Methods: We assessed new oligodendrocyte (OL) and myelin generation and stability in the ME of healthy adult male mice using bromodeoxyuridine labelling and genetic fate mapping tools. We evaluated the contribution of microglia to ME myelin plasticity in PLX5622-treated C57BL/6J mice and in Pdgfra-Cre/ERT2;R26R-eYFP;Myrffl/fl mice, where adult oligodendrogenesis is blunted. Next, we investigated how high-fat feeding or caloric restriction impact ME OL lineage progression and myelination. Finally, we characterized the functional relevance of adult oligodendrogenesis on energy balance regulation. Results: We show that myelinating OLs are continuously and rapidly generated in the adult ME. Paradoxically, OL number and myelin amounts remain remarkably stable in the adult ME. In fact, the high rate of new OL and myelin generation in the ME is offset by continuous turnover of both. We show that microglia are required for continuous OL and myelin production, and that ME myelin plasticity regulates the recruitment of local immune cells. Finally, we provide evidence that ME myelination is regulated by the body’s energetic status and demonstrate that ME OL and myelin plasticity are required for the regulation of energy balance and hypothalamic leptin sensitivity. Conclusions: This study identifies a new mechanism modulating leptin sensitivity and the central control of energy balance and uncovers a previously unappreciated form of structural plasticity in the ME.

Abstract Aims Fatty acid–binding protein (FABP) regulates polyunsaturated fatty acid (PUFA) intracellular trafficking and signal transduction. Our previous studies demonstrated that the alteration of PUFA in the hypothalamus is involved in pain process. However, how FABP subtypes change during pain remain unclear. Here, we examined the expression changes and localization in the hypothalamic FABP subtype in postoperative pain model mice. Methods Paw incision–induced postoperative methods were adopted as a pain model in male ddY mice. Mechanical allodynia was examined using the von Frey test. The analysis of several FABPs mRNA was measured by real‐time PCR, and cellular localization of its protein level was measured by immunofluorescent study. Results Postoperative pain mouse elicited mechanical allodynia on Day 2 after paw incision, and mRNA expression of FABP3 increased significantly in the hypothalamus in the postoperative pain mouse model compared to that in control mice. FABP3 protein expressed in the median eminence and the arcuate nucleus, and colocalized with Iba‐1, which is a microglial cell marker. Its protein level significantly increased in the median eminence on Day 2 after incision and returned to the control level on Day 4 after incision. Conclusions Our findings indicate that FABP3 in the median eminence may change in pain stimuli and may represent a molecular link controlling pain.

Transplanting embryonic precursors of GABAergic neurons from the medial ganglionic eminence (MGE) into adult mouse spinal cord ameliorates mechanical and thermal hypersensitivity in peripheral nerve injury models of neuropathic pain. Although Fos and transneuronal tracing studies strongly suggest that integration of MGE-derived neurons into host spinal cord circuits underlies recovery of function, the extent to which there is synaptic integration of the transplanted cells has not been established. Here, we used electron microscopic immunocytochemistry to assess directly integration of GFP-expressing MGE-derived neuronal precursors into dorsal horn circuitry in intact, adult mice with short- (5-6 weeks) or long-term (4-6 months) transplants. We detected GFP with pre-embedding avidin-biotin-peroxidase and GABA with post-embedding immunogold labeling. At short and long times post-transplant, we found host-derived synapses on GFP-immunoreactive MGE cells bodies and dendrites. The proportion of dendrites with synaptic input increased from 50% to 80% by 6 months. In all mice, MGE-derived terminals formed synapses with GFP-negative (host) cell bodies and dendrites and, unexpectedly, with some GFP-positive (i.e., MGE-derived) dendrites, possibly reflecting autoapses or cross talk among transplanted neurons. We also observed axoaxonic appositions between MGE and host terminals. Immunogold labeling for GABA confirmed that the transplanted cells were GABAergic and that some transplanted cells received an inhibitory GABAergic input. We conclude that transplanted MGE neurons retain their GABAergic phenotype and integrate dynamically into host-transplant synaptic circuits. Taken together with our previous electrophysiological analyses, we conclude that MGE cells are not GABA pumps, but alleviate pain and itch through synaptic release of GABA.

We demonstrate that cortical interneurons derived from ventral eminences, including the caudal ganglionic eminence, undergo programmed cell death. Moreover, with the exception of VIP interneurons, this occurs in a manner that is activity-dependent. In addition, we demonstrate that, within interneurons, Calcineurin, a calcium-dependent protein phosphatase, plays a critical role in sequentially linking activity to maturation (E15-P5) and survival (P5-P20). Specifically, embryonic inactivation of Calcineurin results in a failure of interneurons to morphologically mature and prevents them from undergoing apoptosis. By contrast, early postnatal inactivation of Calcineurin increases apoptosis. We conclude that Calcineurin serves a dual role of promoting first the differentiation of interneurons and, subsequently, their survival.

The excitatory action of gamma-aminobutyric-acid (GABA) in the median-eminence (ME) led to the steady-state release of corticotropinreleasing hormone (CRH) from CRH axon terminals, which modulates the hypothalamic-pituitary-adrenal (HPA) axis. However, in ME, the source of excitatory GABAergic input is unknown. We examined agouti-related peptide (AgRP) expressing neurons in the arcuate nucleus as a possible source for excitatory GABAergic input. Here, we show that a subpopulation of activated AgRP neurons directly project to the CRH axon terminals in ME elevates serum corticosterone levels in 60% food-restricted mice. This increase in serum corticosterone is not dependent on activation of CRH neuronal soma in the paraventricular nucleus. Furthermore, conditional deletion of Na+-K+-2Cl- cotransporter-1 (NKCC1), which promotes depolarizing GABA action, from the CRH axon terminals results in significantly lower corticosterone levels in response to food restriction. These findings highlight the important role of a subset of AgRP neurons in HPA axis modulation via NKCC1-dependent GABAergic excitation in ME. [ABSTRACT FROM AUTHOR]

ObjectiveA significant proportion of the more than 50 million people worldwide currently suffering with epilepsy are resistant to antiepileptic drugs (AEDs). As an alternative to AEDs, novel therapies based on cell transplantation offer an opportunity for long-lasting modification of epileptic circuits. To develop such a treatment requires careful preclinical studies in a chronic epilepsy model featuring unprovoked seizures, hippocampal histopathology, and behavioral comorbidities.MethodsTransplantation of progenitor cells from embryonic medial or caudal ganglionic eminence (MGE, CGE) were made in a well-characterized mouse model of status epilepticus-induced epilepsy (systemic pilocarpine). Behavioral testing (handling and open field), continuous video-electroencephalographic (vEEG) monitoring, and slice electrophysiology outcomes were obtained up to 270 days after transplantation (DAT). Post-hoc immunohistochemistry was used to confirm cell identity.ResultsMGE progenitors transplanted into the hippocampus of epileptic mice rescued handling and open field deficits starting at 60 DAT. In these same mice, an 84% to 88% reduction in seizure activity was observed between 180 and 210 DAT. Inhibitory postsynaptic current frequency, measured on pyramidal neurons in acute hippocampal slices at 270 DAT, was reduced in epileptic mice but restored to naïve levels in epileptic mice receiving MGE transplants. No reduction in seizure activity was observed in epileptic mice receiving intrahippocampal CGE progenitors.InterpretationOur findings demonstrate that transplanted MGE progenitors enhance functional GABA-mediated inhibition, reduce spontaneous seizure frequency, and rescue behavioral deficits in a chronic epileptic animal model more than 6 months after treatment. Ann Neurol 2017;82:530-542.

BackgroundDespite recent insights into the pathophysiology of acute and chronic itch, chronic itch remains an often intractable condition. Among major contributors to chronic itch is dysfunction of spinal cord gamma aminobutyric acidergic (GABAergic) inhibitory controls.ObjectivesWe sought to test the hypothesis that selective GABA agonists as well as cell transplant-derived GABA are antipruritic against acute itch and in a transgenic mouse model of atopic dermatitis produced by overexpression of the TH2 cell-associated cytokine, IL-31 (IL-31Tg mice).MethodsWe injected wild-type and IL-31Tg mice with combinations of GABA-A (muscimol) or GABA-B (baclofen) receptor agonists 15 to 20 minutes prior to injection of various pruritogens (histamine, chloroquine, or endothelin-1) and recorded spontaneous scratching before and after drug administration. We also tested the antipruritic properties of intraspinal transplantation of precursors of GABAergic interneurons in the IL-31Tg mice.ResultsSystemic muscimol or baclofen are antipruritic against both histamine-dependent and -independent pruritogens, but the therapeutic window using either ligand alone was very small. In contrast, combined subthreshold doses of baclofen and muscimol produced a significant synergistic antipruritic effect, with no sedation. Finally, transplant-mediated long-term enhancement of GABAergic signaling not only reduced spontaneous scratching in the IL-31Tg mice but also dramatically resolved the associated skin lesions.ConclusionsAlthough additional research is clearly needed, existing approved GABA agonists should be considered in the management of chronic itch, notably atopic dermatitis.

Many subtypes of cortical interneurons (CINs) are found in adult mouse cortices, but the mechanism generating their diversity remains elusive. We performed single-cell RNA sequencing on the mouse embryonic medial ganglionic eminence (MGE), the major birthplace for CINs, and on MGE-like cells differentiated from embryonic stem cells. Two distinct cell types were identified as proliferating neural progenitors and immature neurons, both of which comprised sub-populations. Although lineage development of MGE progenitors was reconstructed and immature neurons were characterized as GABAergic, cells that might correspond to precursors of different CINs were not identified. A few non-neuronal cell types were detected, including microglia. In vitro MGE-like cells resembled bona fide MGE cells but expressed lower levels of Foxg1 and Epha4. Together, our data provide detailed understanding of the embryonic MGE developmental program and suggest how CINs are specified.

Medial ganglionic eminence (MGE) transplantation rescues disease phenotypes in various preclinical models with interneuron deficiency or dysfunction, including epilepsy. While underlying mechanism(s) remains unclear to date, a simple explanation is that appropriate synaptic integration of MGE-derived interneurons elevates GABA-mediated inhibition and modifies the firing activity of excitatory neurons in the host brain. However, given the complexity of interneurons and potential for transplant-derived interneurons to integrate or alter the host network in unexpected ways, it remains unexplored whether synaptic connections formed by transplant-derived interneurons safely mirror those associated with endogenous interneurons. Here, we combined optogenetics, interneuron-specific Cre driver mouse lines, and electrophysiology to study synaptic integration of MGE progenitors. We demonstrated that MGE-derived interneurons, when transplanted into the hippocampus of neonatal mice, migrate in the host brain, differentiate to mature inhibitory interneurons, and form appropriate synaptic connections with native pyramidal neurons. Endogenous and transplant-derived MGE progenitors preferentially formed inhibitory synaptic connections onto pyramidal neurons but not endogenous interneurons. These findings demonstrate that transplanted MGE progenitors functionally integrate into the postnatal hippocampal network.

Cell transplantation offers an attractive alternative to pharmacotherapy for the management of a host of clinical conditions. Most importantly, the transplanted cells provide a continuous, local delivery of therapeutic compounds, which avoids many of the adverse side effects associated with systemically administered drugs. Here, we describe the broad therapeutic utility of transplanting precursors of cortical inhibitory interneurons derived from the embryonic medial ganglionic eminence (MGE), in a variety of chronic pain and itch models in the mouse. Despite the cortical environment in which the MGE cells normally develop, these cells survive transplantation and will even integrate into the circuitry of an adult host spinal cord. When transplanted into the spinal cord, the cells significantly reduce the hyperexcitability that characterizes both chronic neuropathic pain and itch conditions. This MGE cell-based strategy differs considerably from traditional pharmacological treatments as the approach is potentially disease modifying (i.e., the therapy targets the underlying etiology of the pain and itch pathophysiology).

Cell transplantation offers an attractive alternative to pharmacotherapy for the management of a host of clinical conditions. Most importantly, the transplanted cells provide a continuous, local delivery of therapeutic compounds, which avoids many of the adverse side effects associated with systemically administered drugs. Here, we describe the broad therapeutic utility of transplanting precursors of cortical inhibitory interneurons derived from the embryonic medial ganglionic eminence (MGE), in a variety of chronic pain and itch models in the mouse. Despite the cortical environment in which the MGE cells normally develop, these cells survive transplantation and will even integrate into the circuitry of an adult host spinal cord. When transplanted into the spinal cord, the cells significantly reduce the hyperexcitability that characterizes both chronic neuropathic pain and itch conditions. This MGE cell-based strategy differs considerably from traditional pharmacological treatments as the approach is potentially disease modifying (i.e., the therapy targets the underlying etiology of the pain and itch pathophysiology).

Aims: Fatty acid–binding protein (FABP) regulates polyunsaturated fatty acid (PUFA) intracellular trafficking and signal transduction. Our previous studies demonstrated that the alteration of PUFA in the hypothalamus is involved in pain process. However, how FABP subtypes change during pain remain unclear. Here, we examined the expression changes and localization in the hypothalamic FABP subtype in postoperative pain model mice. Methods: Paw incision–induced postoperative methods were adopted as a pain model in male ddY mice. Mechanical allodynia was examined using the von Frey test. The analysis of several FABPs mRNA was measured by real‐time PCR, and cellular localization of its protein level was measured by immunofluorescent study. Results: Postoperative pain mouse elicited mechanical allodynia on Day 2 after paw incision, and mRNA expression of FABP3 increased significantly in the hypothalamus in the postoperative pain mouse model compared to that in control mice. FABP3 protein expressed in the median eminence and the arcuate nucleus, and colocalized with Iba‐1, which is a microglial cell marker. Its protein level significantly increased in the median eminence on Day 2 after incision and returned to the control level on Day 4 after incision. Conclusions: Our findings indicate that FABP3 in the median eminence may change in pain stimuli and may represent a molecular link controlling pain. [ABSTRACT FROM AUTHOR]

The maturation of inhibitory GABAergic cortical circuits regulates experience-dependent plasticity. We recently showed that the heterochronic transplantation of parvalbumin (PV) or somatostatin (SST) interneurons from the medial ganglionic eminence (MGE) reactivates ocular dominance plasticity (ODP) in the postnatal mouse visual cortex. Might other types of interneurons similarly induce cortical plasticity? Here, we establish that caudal ganglionic eminence (CGE)-derived interneurons, when transplanted into the visual cortex of neonatal mice, migrate extensively in the host brain and acquire laminar distribution, marker expression, electrophysiological properties, and visual response properties like those of host CGE interneurons. Although transplants from the anatomical CGE do induce ODP, we found that this plasticity reactivation is mediated by a small fraction of MGE-derived cells contained in the transplant. These findings demonstrate that transplanted CGE cells can successfully engraft into the postnatal mouse brain and confirm the unique role of MGE lineage neurons in the induction of ODP.

The median eminence (ME) of the hypothalamus comprises the hypothalamic nerve terminals, glia (especially tanycytes) and the portal capillary vasculature that transports hypothalamic neurohormones to the anterior pituitary gland. The ultrastructure of the ME is dynamically regulated by hormones and undergoes organizational changes during development and reproductive cycles in adult females, but relatively little is known about the ME during aging, especially in nonhuman primates. Therefore, we used a novel transmission scanning electron microscopy technique to examine the cytoarchitecture of the ME of young and aged female rhesus macaques in a preclinical monkey model of menopausal hormone treatments. Rhesus macaques were ovariectomized and treated for 2 years with vehicle, estradiol (E2), or estradiol + progesterone (E2 + P4). While the overall cytoarchitecture of the ME underwent relatively few changes with age and hormones, changes to some features of neural and glial components near the portal capillaries were observed. Specifically, large neuroterminal size was greater in aged compared to young adult animals, an effect that was mitigated or reversed by E2 alone but not by E2 + P4 treatment. Overall glial size and the density and tissue fraction of the largest subset of glia were greater in aged monkeys, and in some cases reversed by E2 treatment. Mitochondrial size was decreased by E2, but not E2 + P4, only in aged macaques. These results contrast substantially with work in rodents, suggesting that the ME of aging macaques is less vulnerable to age-related disorganization, and that the effects of E2 on monkeys' ME are age specific.

The arcuate nucleus of the hypothalamus is central in the regulation of body weight homeostasis through its ability to sense peripheral metabolic signals and relay them, through neural circuits, to other brain areas, ultimately affecting physiological and behavioural changes. The early postnatal development of these neural circuits is critical for normal body weight homeostasis, such that perturbations during this critical period can lead to obesity. The role for peripheral regulators of body weight homeostasis, including leptin, insulin and ghrelin, in this postnatal development is well described, yet some of the fundamental processes underpinning axonal and dendritic growth remain unclear. Here, we hypothesised that molecules known to regulate axonal and dendritic growth processes in other areas of the developing brain would be expressed in the postnatal arcuate nucleus and/or target nuclei where they would function to mediate the development of this circuitry. Using state‐of‐the‐art RNAscope® technology, we have revealed the expression patterns of genes encoding Dcc/Netrin‐1, Robo1/Slit1 and Fzd5/Wnt5a receptor/ligand pairs in the early postnatal mouse hypothalamus. We found that individual genes had unique expression patterns across developmental time in the arcuate nucleus, paraventricular nucleus of the hypothalamus, ventromedial nucleus of the hypothalamus, dorsomedial nucleus of the hypothalamus, median eminence and, somewhat unexpectedly, the third ventricle epithelium. These observations indicate a number of new molecular players in the development of neural circuits regulating body weight homeostasis, as well as novel molecular markers of tanycyte heterogeneity. [ABSTRACT FROM AUTHOR]

Mutations in the phosphatase PTEN are strongly implicated in autism spectrum disorder (ASD). Here, we investigate the function of Pten in cortical GABAergic neurons using conditional mutagenesis in mice. Loss of Pten results in a preferential loss of SST(+) interneurons, which increases the ratio of parvalbumin/somatostatin (PV/SST) interneurons, ectopic PV(+) projections in layer I, and inhibition onto glutamatergic cortical neurons. Pten mutant mice exhibit deficits in social behavior and changes in electroencephalogram (EEG) power. Using medial ganglionic eminence (MGE) transplantation, we test for cell-autonomous functional differences between human PTEN wild-type (WT) and ASD alleles. The PTEN ASD alleles are hypomorphic in regulating cell size and the PV/SST ratio in comparison to WT PTEN. This MGE transplantation/complementation assay is efficient and is generally applicable for functional testing of ASD alleles in vivo.

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.

GABAergic inhibition has been shown to play an important role in the opening of critical periods of brain plasticity. We recently have shown that transplantation of GABAergic precursors from the embryonic medial ganglionic eminence (MGE), the source of neocortical parvalbumin- (PV(+)) and somatostatin-expressing (SST(+)) interneurons, can induce a new period of ocular dominance plasticity (ODP) after the endogenous period has closed. Among the diverse subtypes of GABAergic interneurons PV(+) cells have been thought to play the crucial role in ODP. Here we have used MGE transplantation carrying a conditional allele of diphtheria toxin alpha subunit and cell-specific expression of Cre recombinase to deplete PV(+) or SST(+) interneurons selectively and to investigate the contributions of each of these types of interneurons to ODP. As expected, robust plasticity was observed in transplants containing PV(+) cells but in which the majority of SST(+) interneurons were depleted. Surprisingly, transplants in which the majority of PV(+) cells were depleted induced plasticity as effectively as those containing PV(+) cells. In contrast, depleting both cell types blocked induction of plasticity. These findings reveal that PV(+) cells do not play an exclusive role in ODP; SST(+) interneurons also can drive cortical plasticity and contribute to the reshaping of neural networks. The ability of both PV(+) and SST(+) interneurons to induce de novo cortical plasticity could help develop new therapeutic approaches for brain repair.

Summary: The mediobasal hypothalamus (MBH; arcuate nucleus of the hypothalamus [ARH] and median eminence [ME]) is a key nutrient sensing site for the production of the complex homeostatic feedback responses required for the maintenance of energy balance. Here, we show that refeeding after an overnight fast rapidly triggers proliferation and differentiation of oligodendrocyte progenitors, leading to the production of new oligodendrocytes in the ME specifically. During this nutritional paradigm, ME perineuronal nets (PNNs), emerging regulators of ARH metabolic functions, are rapidly remodeled, and this process requires myelin regulatory factor (Myrf) in oligodendrocyte progenitors. In genetically obese ob/ob mice, nutritional regulations of ME oligodendrocyte differentiation and PNN remodeling are blunted, and enzymatic digestion of local PNN increases food intake and weight gain. We conclude that MBH PNNs are required for the maintenance of energy balance in lean mice and are remodeled in the adult ME by the nutritional control of oligodendrocyte differentiation.

Seizure disorders debilitate more than 65,000,000 people worldwide, with temporal lobe epilepsy (TLE) being the most common form. Previous studies have shown that transplantation of GABA-releasing cells results in suppression of seizures in epileptic mice. Derivation of interneurons from human pluripotent stem cells (hPSCs) has been reported, pointing to clinical translation of quality-controlled human cell sources that can enhance inhibitory drive and restore host circuitry. In this study, we demonstrate that hPSC-derived maturing GABAergic interneurons (mGINs) migrate extensively and integrate into dysfunctional circuitry of the epileptic mouse brain. Using optogenetic approaches, we find that grafted mGINs generate inhibitory postsynaptic responses in host hippocampal neurons. Importantly, even before acquiring full electrophysiological maturation, grafted neurons were capable of suppressing seizures and ameliorating behavioral abnormalities such as cognitive deficits, aggressiveness, and hyperactivity. These results provide support for the potential of hPSC-derived mGIN for restorative cell therapy for epilepsy.

The transmission of pruritoceptive (itch) messages involves specific neural circuits within the spinal cord that are distinct from those that transmit pain messages. These itch-specific circuits are tonically regulated by inhibitory interneurons in the dorsal horn. Consistent with these findings, it has previously been reported that loss of GABAergic interneurons in mice harboring a deletion of the transcription factor Bhlhb5 generates a severe, nonremitting condition of chronic itch. Here, we tested the hypothesis that the neuropathic itch in BHLHB5-deficient animals can be treated by restoring inhibitory controls through spinal cord transplantation and integration of precursors of cortical inhibitory interneurons derived from the embryonic medial ganglionic eminence. We specifically targeted the transplants to segments of the spinal cord innervated by areas of the body that were most severely affected. BHLHB5-deficient mice that received transplants demonstrated a substantial reduction of excessive scratching and dramatic resolution of skin lesions. In contrast, the scratching persisted and skin lesions worsened over time in sham-treated mice. Together, these results indicate that cell-mediated restoration of inhibitory controls has potential as a powerful, cell-based therapy for neuropathic itch that not only ameliorates symptoms of chronic itch, but also may modify disease.

Most cortical interneurons originate in a region of the embryonic subpallium called the medial ganglionic eminence (MGE). When MGE cells are transplanted into cerebral cortex, these progenitors migrate extensively and differentiate into functional inhibitory neurons. Although MGE progenitors have therapeutic potential following transplantation, it is unknown precisely how these cells distribute within neocortical lamina of the recipient brain. Here we transplanted mouse embryonic day 12.5 MGE progenitors into postnatal neocortex and evaluated laminar distribution of interneuron subtypes using double- and triple-label immunohistochemistry. Studies were performed using wild type (WT) or donor mice lacking a metabotropic GABA(B) receptor subunit (GABA(B1)R KO). MGE-derived neurons from WT and GABA(B1)R KO mice preferentially and densely distributed in neocortical layers 2/3, 5 and 6. As expected, MGE-derived neurons differentiated into parvalbumin+ and somatostatin+ interneurons within these neocortical lamina. Our findings provide insights into the anatomical integration of MGE-derived interneurons following transplantation.

Most cortical interneurons originate in a region of the embryonic subpallium called the medial ganglionic eminence (MGE). When MGE cells are transplanted into cerebral cortex, these progenitors migrate extensively and differentiate into functional inhibitory neurons. Although MGE progenitors have therapeutic potential following transplantation, it is unknown precisely how these cells distribute within neocortical lamina of the recipient brain. Here we transplanted mouse embryonic day 12.5 MGE progenitors into postnatal neocortex and evaluated laminar distribution of interneuron subtypes using double- and triple-label immunohistochemistry. Studies were performed using wild type (WT) or donor mice lacking a metabotropic GABA(B) receptor subunit (GABA(B1)R KO). MGE-derived neurons from WT and GABA(B1)R KO mice preferentially and densely distributed in neocortical layers 2/3, 5 and 6. As expected, MGE-derived neurons differentiated into parvalbumin+ and somatostatin+ interneurons within these neocortical lamina. Our findings provide insights into the anatomical integration of MGE-derived interneurons following transplantation.

Llamas are induced non-reflex ovulators, which ovulate in response to the hormonal stimulus of the male protein beta-nerve growth factor (β-NGF) that is present in the seminal plasma; this response is dependent on the preovulatory gonadotrophin-releasing hormone (GnRH) release from the hypothalamus. GnRH neurones are vital for reproduction, as these provide the input that controls the release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) from the pituitary gland. However, in spontaneous ovulators, the activity of GnRH cells is regulated by kisspeptin neurones that relay the oestrogen signal arising from the periphery. Here, we investigated the organisation of GnRH and kisspeptin systems in the hypothalamus of receptive adult female llamas. We found that GnRH cells exhibiting different shapes were distributed throughout the ventral forebrain and some of these were located in proximity to blood vessels; sections of the mediobasal hypothalamus (MBH) displayed the highest number of cells. GnRH fibres were observed in both the organum vasculosum laminae terminalis (OVLT) and median eminence (ME). We also detected abundant kisspeptin fibres in the MBH and ME; kisspeptin cells were found in the arcuate nucleus (ARC), but not in rostral areas of the hypothalamus. Quantitative analysis of GnRH and kisspeptin fibres in the ME revealed a higher innervation density of kisspeptin than of GnRH fibres. The physiological significance of the anatomical findings reported here for the ovulatory mechanism in llamas is still to be determined.

Here we have provided evidence that nitric oxide-cyclic GMP (NO-cGMP) signaling regulates neurite length and migration of immature neurons derived from the medial ganglionic eminence (MGE). Dlx1/2(-/-) and Lhx6(-/-) mouse mutants, which exhibit MGE interneuron migration defects, have reduced expression of the gene encoding the α subunit of a soluble guanylate cyclase (Gucy1A3). Furthermore, Dlx1/2(-/-) mouse mutants have reduced expression of NO synthase 1 (NOS1). Gucy1A3(-/-) mice have a transient reduction in cortical interneuron number. Pharmacological inhibition of soluble guanylate cyclase and NOS activity rapidly induces neurite retraction of MGE cells in vitro and in slice culture and robustly inhibits cell migration from the MGE and caudal ganglionic eminence. We provide evidence that these cellular phenotypes are mediated by activation of the Rho signaling pathway and inhibition of myosin light chain phosphatase activity.

Directed differentiation from human pluripotent stem cells (hPSCs) has seen significant progress in recent years. However, most differentiated populations exhibit immature properties of an early embryonic stage, raising concerns about their ability to model and treat disease. Here, we report the directed differentiation of hPSCs into medial ganglionic eminence (MGE)-like progenitors and their maturation into forebrain type interneurons. We find that early-stage progenitors progress via a radial glial-like stem cell enriched in the human fetal brain. Both in vitro and posttransplantation into the rodent cortex, the MGE-like cells develop into GABAergic interneuron subtypes with mature physiological properties along a prolonged intrinsic timeline of up to 7 months, mimicking endogenous human neural development. MGE-derived cortical interneuron deficiencies are implicated in a broad range of neurodevelopmental and degenerative disorders, highlighting the importance of these results for modeling human neural development and disease.

In humans and experimental animals, the administration of ciliary neurotrophic factor (CNTF) reduces food intake and body weight. To gain further insights into the mechanism(s) underlying its satiety effect, we: (i) evaluated the CNTF-dependent activation of the Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) pathway in mouse models where neuropeptide Y (NPY) and pro-opiomelanocortin (POMC) neurons can be identified by green fluorescent protein (GFP); and (ii) assessed whether CNTF promotes leptin signaling in hypothalamic feeding centers. Immunohistochemical experiments enabled us to establish that intraperitoneal injection of mouse recombinant CNTF activated the JAK2-STAT3 pathway in a substantial proportion of arcuate nucleus (ARC) NPY neurons (18.68% ± 0.60 in 24-h fasted mice and 25.50% ± 1.17 in fed mice) but exerted a limited effect on POMC neurons (4.15% ± 0.33 in 24-h fasted mice and 2.84% ± 0.45 in fed mice). CNTF-responsive NPY neurons resided in the ventromedial ARC, facing the median eminence (ME), and were surrounded by albumin immunoreactivity, suggesting that they are located outside the blood-brain barrier (BBB). In both normally fed and high-fat diet (HFD) obese animals, CNTF activated extracellular signal-regulated kinase signaling in ME β1- and β2-tanycytes, an effect that has been linked to the promotion of leptin entry into the brain. Accordingly, compared to the animals treated with leptin, mice treated with leptin/CNTF showed: (i) a significantly greater leptin content in hypothalamic protein extracts; (ii) a significant increase in phospho-STAT3 (P-STAT3)-positive neurons in the ARC and the ventromedial hypothalamic nucleus of normally fed mice; and (iii) a significantly increased number of P-STAT3-positive neurons in the ARC and dorsomedial hypothalamic nucleus of HFD obese mice. Collectively, these data suggest that exogenously administered CNTF reduces food intake by exerting a leptin-like action on distinctive NPY ARC neurons and by promoting leptin signaling in hypothalamic feeding centers.

Arginine vasopressin (AVP), when released into portal capillaries with corticotrophin‐releasing factor (CRF) from terminals of parvocellular neurones of the hypothalamic paraventricular nucleus (PVH), facilitates the secretion of adrenocorticotrophic hormone (ACTH) in stressed rodents. The AVP gene encodes a propeptide precursor containing AVP, AVP‐associated neurophysin II (NPII), and a glycopeptide copeptin, although it is currently unclear whether copeptin is always cleaved from the neurophysin and whether the NPII and/or copeptin have any functional role in the pituitary. Furthermore, for primates, it is unknown whether CRF, AVP, NPII and copeptin are all colocalised in neurosecretory vesicles in the terminal region of the paraventricular CRF neurone axons. Therefore, we investigated, by fluorescence and immunogold immunocytochemistry, the cellular and subcellular relationships of these peptides in the CRF‐ and AVP‐producing cells in unstressed Japanese macaque monkeys (Macaca fuscata). Reverse transcription‐polymerase chain reaction analysis showed the expression of both CRF and AVP mRNAs in the monkey PVH. As expected, in the magnocellular neurones of the PVH and supraoptic nucleus, essentially no CRF immunoreactivity could be detected in NPII‐immunoreactive (AVP‐producing) neurones. Immunofluorescence showed that, in the parvocellular part of the PVH, NPII was detectable in a subpopulation (approximately 39%) of the numerous CRF‐immunoreactive neuronal perikarya, whereas, in the outer median eminence, NPII was more prominent (approximately 52%) in the CRF varicosities. Triple immunoelectron microscopy in the median eminence demonstrated the presence of both NPII and copeptin immunoreactivity in dense‐cored vesicles of CRF‐containing axons. The results are consistent with an idea that the AVP propeptide is processed and NPII and copeptin are colocalised in hypothalamic‐pituitary CRF axons in the median eminence of a primate. The CRF, AVP and copeptin are all co‐packaged in neurosecretory vesicles in monkeys and are thus likely to be co‐released into the portal capillary blood to amplify ACTH release from the primate anterior pituitary. [ABSTRACT FROM AUTHOR]

Tanycytes are peculiar ependymoglial cells lining the bottom and the lateral wall of the third ventricle. For a decade, the utilization of molecular genetic approaches allowed us to make important discoveries about their diverse physiological functions. Here, I review the current methods used to target tanycytes, focusing on their specificity, their efficiency, their limitations, as well as their potential future improvements. [ABSTRACT FROM AUTHOR]

In humans and experimental animals, the administration of ciliary neurotrophic factor (CNTF) reduces food intake and body weight. To gain further insights into the mechanism(s) underlying its satiety effect, we: (i) evaluated the CNTF-dependent activation of the Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) pathway in mouse models where neuropeptide Y (NPY) and pro-opiomelanocortin (POMC) neurons can be identified by green fluorescent protein (GFP); and (ii) assessed whether CNTF promotes leptin signaling in hypothalamic feeding centers. Immunohistochemical experiments enabled us to establish that intraperitoneal injection of mouse recombinant CNTF activated the JAK2-STAT3 pathway in a substantial proportion of arcuate nucleus (ARC) NPY neurons (18.68% ± 0.60 in 24-h fasted mice and 25.50% ± 1.17 in fed mice) but exerted a limited effect on POMC neurons (4.15% ± 0.33 in 24-h fasted mice and 2.84% ± 0.45 in fed mice). CNTF-responsive NPY neurons resided in the ventromedial ARC, facing the median eminence (ME), and were surrounded by albumin immunoreactivity, suggesting that they are located outside the blood-brain barrier (BBB). In both normally fed and high-fat diet (HFD) obese animals, CNTF activated extracellular signal-regulated kinase signaling in ME β1- and β2-tanycytes, an effect that has been linked to the promotion of leptin entry into the brain. Accordingly, compared to the animals treated with leptin, mice treated with leptin/CNTF showed: (i) a significantly greater leptin content in hypothalamic protein extracts; (ii) a significant increase in phospho-STAT3 (P-STAT3)-positive neurons in the ARC and the ventromedial hypothalamic nucleus of normally fed mice; and (iii) a significantly increased number of P-STAT3-positive neurons in the ARC and dorsomedial hypothalamic nucleus of HFD obese mice. Collectively, these data suggest that exogenously administered CNTF reduces food intake by exerting a leptin-like action on distinctive NPY ARC neurons and by promoting leptin signaling in hypothalamic feeding centers. [ABSTRACT FROM AUTHOR]