Morphological characteristics of astrocytes of the fastigial nucleus.

Astrocytes are a diverse and morphologically complex class of glial cells restricted to the central nervous system which have been implicated in the modulation of neuronal activity. The cerebellum is involved in planning movements and motor learning. Within the cerebellum three deep cerebellar nuclei (dentate, interposed and fastigial) provide the sole neuronal output. The fastigial nucleus participates in saccadic and vestibular function, and recent evidence disclosed neuronal projections to cognitive, affective, and motor areas. However, thus far there are no reliable descriptions of the distribution and morphological classifications of astrocytes in this nucleus. This work aims to describe the characteristics of astrocytes of the fastigial nucleus based on the expression of GFP in a transgenic mouse model.

hESC-derived striatal progenitors grafted into a Huntington's disease rat model support long-term functional motor recovery by differentiating, self-organizing and connecting into the lesioned striatum.

BACKGROUND: Huntington's disease (HD) is a motor and cognitive neurodegenerative disorder due to prominent loss of striatal medium spiny neurons (MSNs). Cell replacement using human embryonic stem cells (hESCs) derivatives may offer new therapeutic opportunities to replace degenerated neurons and repair damaged circuits. METHODS: With the aim to develop effective cell replacement for HD, we assessed the long-term therapeutic value of hESC-derived striatal progenitors by grafting the cells into the striatum of a preclinical model of HD [i.e., adult immunodeficient rats in which the striatum was lesioned by monolateral injection of quinolinic acid (QA)]. We examined the survival, maturation, self-organization and integration of the graft as well as its impact on lesion-dependent motor alterations up to 6 months post-graft. Moreover, we tested whether exposing a cohort of QA-lesioned animals to environmental enrichment (EE) could improve graft integration and function. RESULTS: Human striatal progenitors survived up to 6 months after transplantation and showed morphological and neurochemical features typical of human MSNs. Donor-derived interneurons were also detected. Grafts wired in both local and long-range striatal circuits, formed domains suggestive of distinct ganglionic eminence territories and displayed emerging striosome features. Moreover, over time grafts improved complex motor performances affected by QA. EE selectively increased cell differentiation into MSN phenotype and promoted host-to-graft connectivity. However, when combined to the graft, the EE paradigm used in this study was insufficient to produce an additive effect on task execution. CONCLUSIONS: The data support the long-term therapeutic potential of ESC-derived human striatal progenitor grafts for the replacement of degenerated striatal neurons in HD and suggest that EE can effectively accelerate the maturation and promote the integration of human striatal cells.

Analysis of a cell niche with proliferative potential at the roof of the aqueduct of Sylvius.

The aqueduct of Sylvius connects the third with the fourth ventricle and is surrounded by the Periaqueductal Grey. Here, we report a novel niche of cells in the dorsal section of the aqueduct, hereby named dorsal aqueduct niche or DAN, by applying a battery of selective markers and transgenic mouse lines. The somata of DAN cells are located toward the lumen of the ventricle forming multiple layers in close association with the cerebrospinal fluid (CSF). A single process emerges from the soma and run with the blood vessels. Cells of the DAN express radial glia/stem cell markers such as GFAP, vimentin and nestin, and the glutamate transporter GLAST or the oligodendrocyte precursor/pericyte marker NG2, thereby suggesting their potential for the generation of new cells. Morphologically, DAN cells resemble tanycytes of the third ventricle, which transfer biochemical signals from the CSF to the central nervous system and display proliferative capacity. The aqueduct ependymal lining can proliferate as observed by the integration of BrdU and expression of Ki67. Thus, the dorsal section of the aqueduct of Sylvius possesses cells that may act a niche of new glial cells in the adult mouse brain.

Origin of thyrotropin-releasing hormone neurons that innervate the tuberomammillary nuclei.

Hypophysiotropic thyrotropin-releasing hormone (TRH) neurons function as metabolic sensors that regulate the thyroid axis and energy homeostasis. Less is known about the role of other hypothalamic TRH neurons. As central administration of TRH decreases food intake and increases histamine in the tuberomammillary nuclei (TMN), and TMN histamine neurons are densely innervated by TRH fibers from an unknown origin, we mapped the location of TRH neurons that project to the TMN. The retrograde tracer, cholera toxin B subunit (CTB), was injected into the TMN E1-E2, E4-E5 subdivisions of adult Sprague-Dawley male rats. TMN projecting neurons were observed in the septum, preoptic area, bed nucleus of the stria terminalis (BNST), perifornical area, anterior paraventricular nucleus, peduncular and tuberal lateral hypothalamus (TuLH), suprachiasmatic nucleus and medial amygdala. However, CTB/pro-TRH178-199 double-labeled cells were only found in the TuLH. The specificity of the retrograde tract-tracing result was confirmed by administering the anterograde tracer, Phaseolus vulgaris leuco-agglutinin (PHAL) into the TuLH. Double-labeled PHAL-pro-TRH boutons were identified in all subdivisions of the TMN. TMN neurons double-labeled for histidine decarboxylase (Hdc)/PHAL, Hdc/Trh receptor (Trhr), and Hdc/Trh. Further confirmation of a TuLH-TRH neuronal projection to the TMN was established in a transgenic mouse that expresses Cre recombinase in TRH-producing cells following microinjection of a Cre recombinase-dependent AAV that expresses mCherry into the TuLH. We conclude that, in rodents, the TRH innervation of TMN originates in part from TRH neurons in the TuLH, and that this TRH population may contribute to regulate energy homeostasis through histamine Trhr-positive neurons of the TMN.

Organization of the ventricular zone of the cerebellum.

The roof of the fourth ventricle (4V) is located on the ventral part of the cerebellum, a region with abundant vascularization and cell heterogeneity that includes tanycyte-like cells that define a peculiar glial niche known as ventromedial cord. This cord is composed of a group of biciliated cells that run along the midline, contacting the ventricular lumen and the subventricular zone. Although the complex morphology of the glial cells composing the cord resembles to tanycytes, cells which are known for its proliferative capacity, scarce or non-proliferative activity has been evidenced in this area. The subventricular zone of the cerebellum includes astrocytes, oligodendrocytes, and neurons whose function has not been extensively studied. This review describes to some extent the phenotypic, morphological, and functional characteristics of the cells that integrate the roof of the 4V, primarily from rodent brains.

Calcium Signaling in the Cerebellar Radial Glia and Its Association with Morphological Changes during Zebrafish Development.

Radial glial cells are a distinct non-neuronal cell type that, during development, span the entire width of the brain walls of the ventricular system. They play a central role in the origin and placement of neurons, since their processes form structural scaffolds that guide and facilitate neuronal migration. Furthermore, glutamatergic signaling in the radial glia of the adult cerebellum (i.e., Bergmann glia), is crucial for precise motor coordination. Radial glial cells exhibit spontaneous calcium activity and functional coupling spread calcium waves. However, the origin of calcium activity in relation to the ontogeny of cerebellar radial glia has not been widely explored, and many questions remain unanswered regarding the role of radial glia in brain development in health and disease. In this study we used a combination of whole mount immunofluorescence and calcium imaging in transgenic (gfap-GCaMP6s) zebrafish to determine how development of calcium activity is related to morphological changes of the cerebellum. We found that the morphological changes in cerebellar radial glia are quite dynamic; the cells are remarkably larger and more elaborate in their soma size, process length and numbers after 7 days post fertilization. Spontaneous calcium events were scarce during the first 3 days of development and calcium waves appeared on day 5, which is associated with the onset of more complex morphologies of radial glia. Blockage of gap junction coupling inhibited the propagation of calcium waves, but not basal local calcium activity. This work establishes crucial clues in radial glia organization, morphology and calcium signaling during development and provides insight into its role in complex behavioral paradigms.

Inter-fastigial projections along the roof of the fourth ventricle.

The fastigial nucleus (FN) is a bilateral cerebellar integrative center for saccadic and vestibular control associated with non-motor functions such as feeding and cardiovascular regulation. In a previous study, we identified a tract of myelinated axons embedded in the subventricular zone (SVZ) that is located between the ependymal cells that form the dorsal wall of the ventricle and the glia limitans at the roof of the fourth ventricle González-González (Sci Rep 2017, 7:40768). Here, we show that this tract of axons, named subventricular axons or SVa, contains projection neurons that bilaterally interconnect both FNs. The approach consisted of the use of a battery of fluorescent neuronal tracers, transgenic mouse lines, and immunohistofluorescence. Our observations show that the SVa belong to a wide network of GABAergic projection neurons mainly located in the medial and caudal region of the FN. The SVa should be considered a part of a continuum of the cerebellar white matter that follows an alternative pathway through the SVZ, a region closely associated with the physiology of the fourth ventricle. This finding adds to our understanding of the complex organization of the FN; however, the function of the interconnection remains to be elucidated.

Identification of novel cellular clusters define a specialized area in the cerebellar periventricular zone.

The periventricular zone of cerebellum is a germinative niche during the embryonic development, nevertheless its structural organization and functional implications in adult have not been widely studied. Here we disclose the presence of two novel clusters of cells in that area. The first one was named the subventricular cellular cluster (SVCC) and is composed of cells that express glial and neuronal markers. The second was named the ventromedial cord (VMC) and appears as a streak of biciliated cells with microvillosities facing the ventricle, that includes GFAP+ and nestin+ cells organized along the periventricular vasculature. The dorsal limit of the SVCC is associated with myelinated axons of neurons of unknown origin. This paper describes the characteristics and organization of these groups of cells. They can be observed from late embryonic development in the transgenic mouse line GFAP-GFP. The SVCC and VMC expand during early postnatal development but are restricted to the central area of the ventricle in adulthood. We did not find evidence of cell proliferation, cell migration or the presence of fenestrated blood vessels. These findings provide new insights into the knowledge of the cellular composition and structural organization of the periventricular zone of cerebellum.

Fluoxetine Requires the Endfeet Protein Aquaporin-4 to Enhance Plasticity of Astrocyte Processes.

Morphological alterations in astrocytes are characteristic for post mortem brains of patients affected by major depressive disorder (MDD). Recently, a significant reduction in the coverage of blood vessels (BVs) by aquaporin-4 (AQP-4)-positive astrocyte endfeet has been shown in the prefrontal cortex (PFC) of MDD patients, suggesting that either alterations in the morphology of endfeet or in AQP-4 distribution might be responsible for the disease phenotype or constitute a consequence of its progress. Antidepressant drugs (ADs) regulate the expression of several proteins, including astrocyte-specific ones. Thus, they may target AQP-4 to induce morphological changes in astrocytes and restore their proper shape or relocate AQP-4 to endfeet. Using an animal model of depression, rats selectively bred for high anxiety-like behavior (HAB), we confirmed a reduced coverage of BVs in the adult PFC by AQP-4-immunoreactive (AQP-4-IR) astrocyte processes with respect to non-selected Wistar rats (NAB), thereby validating it for our study. A further evaluation of the morphology of astrocyte in brain slices (ex vivo) and in vitro using an antibody against the astrocyte-specific cytoskeletal protein glial fibrillary acidic protein (GFAP) revealed that HAB astrocytes extended less processes than NAB cells. Furthermore, short-term drug treatment in vitro with the AD fluoxetine (FLX) was sufficient to increase the plasticity of astrocyte processes, enhancing their number in NAB-derived cells and recovering their basal number in HAB-derived cells. This enhanced FLX-dependent plasticity occurred, however, only in the presence of intact AQP-4, as demonstrated by the lack of effect after the downregulation of AQP-4 with RNAi in both NAB and HAB cells. Nonetheless, a similar short-term treatment did neither modulate the coverage of BVs with AQP-4-positive astrocyte endfeet in NAB nor in HAB rats, although dosage and time of treatment were sufficient to fully recover GFAP expression in HAB brains. Thus, we suggest that longer treatment regimes may be needed to properly restore the coverage of BVs or to relocate AQP-4 to astrocyte endfeet. In conclusion, FLX requires AQP-4 to modulate the plasticity of astrocyte processes and this effect might be essential to re-establish a functional glia-vasculature interface necessary for a physiological communication between bloodstream and brain parenchyma.