Expression of a Barhl1a reporter in subsets of retinal ganglion cells and commissural neurons of the developing zebrafish brain.

Promoting the regeneration or survival of retinal ganglion cells (RGCs) is one focus of regenerative medicine. Homeobox Barhl transcription factors might be instrumental in these processes. In mammals, only barhl2 is expressed in the retina and is required for both subtype identity acquisition of amacrine cells and for the survival of RGCs downstream of Atoh7, a transcription factor necessary for RGC genesis. The underlying mechanisms of this dual role of Barhl2 in mammals have remained elusive. Whole genome duplication in the teleost lineage generated the barhl1a and barhl2 paralogues. In the Zebrafish retina, Barhl2 functions as a determinant of subsets of amacrine cells lineally related to RGCs independently of Atoh7. In contrast, barhl1a expression depends on Atoh7 but its expression dynamics and function have not been studied. Here we describe for the first time a Barhl1a reporter line in vivo showing that barhl1a turns on exclusively in subsets of RGCs and their post-mitotic precursors. We also show transient expression of barhl1a:GFP in diencephalic neurons extending their axonal projections as part of the post-optic commissure, at the time of optic chiasm formation. This work sets the ground for future studies on RGC subtype identity, axonal projections and genetic specification of Barhl1a-positive RGCs and commissural neurons.

Ipsilateral and Contralateral Retinal Ganglion Cells Express Distinct Genes during Decussation at the Optic Chiasm.

The increasing availability of transcriptomic technologies within the last decade has facilitated high-throughput identification of gene expression differences that define distinct cell types as well as the molecular pathways that drive their specification. The retinal projection neurons, retinal ganglion cells (RGCs), can be categorized into distinct morphological and functional subtypes and by the laterality of their projections. Here, we present a method for purifying the sparse population of ipsilaterally projecting RGCs in mouse retina from their contralaterally projecting counterparts during embryonic development through rapid retrograde labeling followed by fluorescence-activated cell sorting. Through microarray analysis, we uncovered the distinct molecular signatures that define and distinguish ipsilateral and contralateral RGCs during the critical period of axonal outgrowth and decussation, with more than 300 genes differentially expressed within these two cell populations. Among the differentially expressed genes confirmed through in vivo expression validation, several genes that mark "immaturity" are expressed within postmitotic ipsilateral RGCs. Moreover, at least one complementary pair, Igf1 and Igfbp5, is upregulated in contralateral or ipsilateral RGCs, respectively, and may represent signaling pathways that determine ipsilateral versus contralateral RGC identity. Importantly, the cell cycle regulator cyclin D2 is highly expressed in peripheral ventral retina with a dynamic expression pattern that peaks during the period of ipsilateral RGC production. Thus, the molecular signatures of ipsilateral and contralateral RGCs and the mechanisms that regulate their differentiation are more diverse than previously expected.

Six3 regulates optic nerve development via multiple mechanisms.

Malformations of the optic nerve lead to reduced vision or even blindness. During optic nerve development, retinal ganglion cell (RGC) axons navigate across the retina, exit the eye to the optic stalk (OS), and cross the diencephalon midline at the optic chiasm en route to their brain targets. Many signalling molecules have been implicated in guiding various steps of optic nerve pathfinding, however much less is known about transcription factors regulating this process. Here we show that in zebrafish, reduced function of transcription factor Six3 results in optic nerve hypoplasia and a wide repertoire of RGC axon pathfinding errors. These abnormalities are caused by multiple mechanisms, including abnormal eye and OS patterning and morphogenesis, abnormal expression of signalling molecules both in RGCs and in their environment and anatomical deficiency in the diencephalic preoptic area, where the optic chiasm normally forms. Our findings reveal new roles for Six3 in eye development and are consistent with known phenotypes of reduced SIX3 function in humans. Hence, the new zebrafish model for Six3 loss of function furthers our understanding of the mechanisms governing optic nerve development and Six3-mediated eye and forebrain malformations.

Visualization of Nerve Fiber Orientations in the Human Optic Chiasm Using Photomicrographic Image Analysis.

PURPOSE: Hemidecussation of fibers entering the optic chiasm from the optic nerves is well recognized. The reason why bitemporal hemianopia results from chiasmal compression has not been fully explained. There is still a paucity of data relating to the precise details of the routes that the nerve fibers take through the chiasm and, in particular, where and how nerve fibers cross each other. This information is important to understanding why crossing fibers are selectively damaged as a result of chiasmal compression. METHODS: An optic chiasm obtained at postmortem was fixed, stained, and sectioned to allow high-resolution photomicrographs to be taken. The photomicrographs were integrated to allow regions of interest across entire sections to be analyzed for fiber direction and crossing. RESULTS: The results confirmed that fibers from the temporal retina pass directly backward in the lateral chiasm to the optic tract, whereas fibers from the nasal retina cross to the contralateral optic tract. Crossings take place in the paracentral regions of the chiasm rather than in the center of the chiasm (where the nerve fibers are traveling mostly in parallel). The paracentral crossing regions are distributed in a largely postero-superior to antero-inferior arrangement. CONCLUSIONS: These findings clarify the precise locations and crossing angles of crossing nerve fibers in the chiasm. This information may help explain the clinical observation of junctional scotoma and will provide a much better basis for structural modeling of chiasmal compression which, in turn, will improve our understanding of how and why bitemporal hemianopia occurs.

Birthdate and outgrowth timing predict cellular mechanisms of axon target matching in the developing visual pathway.

How axons select their appropriate targets in the brain remains poorly understood. Here, we explore the cellular mechanisms of axon target matching in the developing visual system by comparing four transgenic mouse lines, each with a different population of genetically labeled retinal ganglion cells (RGCs) that connect to unique combinations of brain targets. We find that the time when an RGC axon arrives in the brain is correlated with its target selection strategy. Early-born, early-arriving RGC axons initially innervate multiple targets. Subsequently, most of those connections are removed. By contrast, later-born, later-arriving RGC axons are highly accurate in their initial target choices. These data reveal the diversity of cellular mechanisms that mammalian CNS axons use to pick their targets and highlight the key role of birthdate and outgrowth timing in influencing this precision. Timing-based mechanisms may underlie the assembly of the other sensory pathways and complex neural circuitry in the brain.

Localization of protein kinase C isoforms in the optic pathway of mouse embryos and their role in axon routing at the optic chiasm.

Protein kinase C (PKC) plays a key role in many receptor-mediated signaling pathways that regulate cell growth and development. However, its roles in guiding axon growth and guidance in developing neural pathways are largely unknown. To investigate possible functions of PKC in the growth and guidance of axons in the optic chiasm, we first determined the localization of major PKC isoforms in the retinofugal pathway of mouse embryos, at the stage when axons navigate through the midline. Results showed that PKC was expressed in isoform specific patterns in the pathway. PKC-α immunoreactivity was detected in the chiasm and the optic tract. PKC-ßΙΙ was strong in the optic stalk but was attenuated on axons in the diencephalon. Immunostaining for PKC-ε showed a colocalization in the chiasmatic neurons that express a surface antigen stage specific embryonic antigen-1 (SSEA-1). These chiasmatic neurons straddled the midline of the optic chiasm, and have been shown in earlier studies a role in regulation of axon growth and guidance. Expression levels of PKC-ßΙ, -δ and -γ were barely detectable in the pathway. Blocking of PKC signaling with Ro-32-0432, an inhibitor specific for PKC-α and -ß at nanomolar concentration, produced a dramatic reduction of ipsilateral axons from both nasal retina and temporal crescent. We conclude from these studies that PKC-α and -ßΙΙ are the predominant forms in the developing optic pathway, whereas PKC-ε is the major form in the chiasmatic neurons. Furthermore, PKC-α and -ßΙΙ are likely involved in signaling pathways triggered by inhibitory molecules at the midline that guide optic axons to the uncrossed pathway.

Pituicitoma hipofisario / No disponible

Los pituicitomas son una entidad poco frecuente incluida en la Clasificación de los Tumores del Sistema Nervioso de la Organización Mundial de la Salud (OMS) en el año 2007. Son lesiones originadas en la neurohipófisis y han sido confundidas durante años con otros tumores hipofisarios. Presentamos el caso de una mujer de 31 años diagnosticada de una lesión supraselar en el contexto de un estudio de infertilidad debida a un hipogonadismo-hipogonadotropo con prolactina ligeramente aumentada, a la que se realizó un abordaje pterional consiguiendo su exéresis completa. Tras la cirugía aparecieron hemianopsia bitemporal, diabetes insípida y panhipopituitarismo, y los 2 últimos se corrigieron en pocas semanas. Realizamos además una revisión de sus presentaciones clínicas y radiográficas más frecuentes, así como de los tratamientos planteados en los casos publicados (AU)

Optic chiasm presentation of Semaphorin6D in the context of Plexin-A1 and Nr-CAM promotes retinal axon midline crossing.

At the optic chiasm, retinal ganglion cells (RGCs) project ipsi- or contralaterally to establish the circuitry for binocular vision. Ipsilateral guidance programs have been characterized, but contralateral guidance programs are not well understood. Here, we identify a tripartite molecular system for contralateral RGC projections: Semaphorin6D (Sema6D) and Nr-CAM are expressed on midline radial glia and Plexin-A1 on chiasm neurons, and Plexin-A1 and Nr-CAM are also expressed on contralateral RGCs. Sema6D is repulsive to contralateral RGCs, but Sema6D in combination with Nr-CAM and Plexin-A1 converts repulsion to growth promotion. Nr-CAM functions as a receptor for Sema6D. Sema6D, Plexin-A1, and Nr-CAM are all required for efficient RGC decussation at the optic chiasm. These findings suggest a mechanism by which a complex of Sema6D, Nr-CAM, and Plexin-A1 at the chiasm midline alters the sign of Sema6D and signals Nr-CAM/Plexin-A1 receptors on RGCs to implement the contralateral RGC projection.

Organization and metamorphosis of glia in the Drosophila visual system.

The visual system of Drosophila is an excellent model for determining the interactions that direct the differentiation of the nervous system's many unique cell types. Glia are essential not only in the development of the nervous system, but also in the function of those neurons with which they become associated in the adult. Given their role in visual system development and adult function we need to both accurately and reliably identify the different subtypes of glia, and to relate the glial subtypes in the larval brain to those previously described for the adult. We viewed driver expression in subsets of larval eye disc glia through the earliest stages of pupal development to reveal the counterparts of these cells in the adult. Two populations of glia exist in the lamina, the first neuropil of the adult optic lobe: those that arise from precursors in the eye-disc/optic stalk and those that arise from precursors in the brain. In both cases, a single larval source gives rise to at least three different types of adult glia. Furthermore, analysis of glial cell types in the second neuropil, the medulla, has identified at least four types of astrocyte-like (reticular) glia. Our clarification of the lamina's adult glia and identification of their larval origins, particularly the respective eye disc and larval brain contributions, begin to define developmental interactions which establish the different subtypes of glia.

Development of astroglia heterogeneously expressing Pax2, vimentin and GFAP during the ontogeny of the optic pathway of the lizard (Gallotia galloti): an immunohistochemical and ultrastructural study.

The successful regrowth of retinal ganglion cell (RGC) axons after optic nerve (ON) axotomy in Gallotia galloti indicates a permissive role of the glial environment. We have characterised the astroglial lineage of the lizard optic pathway throughout its ontogeny (embryonic stage 30 [E30] to adults) by using electron microscopy and immunohistochemistry to detect the proliferation marker PCNA (proliferating cell nuclear antigen), the transcription factor Pax2 and the gliofilament proteins vimentin (Vim) and GFAP (glial fibrillary acidic protein). PCNA(+) cells were abundant until E39, with GFAP(+)/PCNA(+) astrocytes being observed between E37 and hatching. Proliferation diminished markedly afterwards, being undetectable in the adult optic pathway. Müller glia of the central retina expressed Pax2 from E37 and their endfeet accumulated Vim from E33 and GFAP from E37 onwards. Astrocytes were absent in the avascular lizard retina, whereas abundant Pax2(+) astrocytes were observed in the ON from E30. A major subpopulation of these astrocytes coexpressed Vim from E35 and also GFAP from E37 onwards; thus the majority of mature astrocytes coexpressed Pax2/Vim/GFAP. The astrocytes were ultrastructurally identified by their gliofilaments, microtubules, dense bodies, desmosomes and glycogen granules, which preferentially accumulated in cell processes. Astrocytes in the adult ON coexpressed both gliofilaments and presented desmosomes indicating a reinforcement of the ON structure; this is physiologically necessary for local adaptation to mechanical forces linked to eye movement. We suggest that astrocytes forming this structural scaffold facilitate the regrowth of RGCs after ON transection.

VEGF mediates commissural axon chemoattraction through its receptor Flk1.

Growing axons are guided to their targets by attractive and repulsive cues. In the developing spinal cord, Netrin-1 and Shh guide commissural axons toward the midline. However, the combined inhibition of their activity in commissural axon turning assays does not completely abrogate turning toward floor plate tissue, suggesting that additional guidance cues are present. Here we show that the prototypic angiogenic factor VEGF is secreted by the floor plate and is a chemoattractant for commissural axons in vitro and in vivo. Inactivation of Vegf in the floor plate or of its receptor Flk1 in commissural neurons causes axon guidance defects, whereas Flk1 blockade inhibits turning of axons to VEGF in vitro. Similar to Shh and Netrin-1, VEGF-mediated commissural axon guidance requires the activity of Src family kinases. Our results identify VEGF and Flk1 as a novel ligand/receptor pair controlling commissural axon guidance.

In vivo diffusion tensor imaging of the human optic chiasm at sub-millimeter resolution.

In this work we report findings from an in vivo diffusion tensor imaging (DTI) study of the human optic chiasm at sub-millimeter voxel resolution. Data were collected at 3 T using a diffusion-weighted radial-FSE sequence, which provides images free from typical magnetic susceptibility artifacts. The general DTI features observed in the optic chiasm region were consistent across subjects. They included a central area with high anisotropy and highest diffusivity in a predominately right/left direction corresponding to the decussation of nasal hemiretinae fibers, surrounded by a band of low anisotropy reflecting heterogeneous orientation of fibers within the voxel, and a lateral area with high anisotropy and highest diffusivity in a predominately anterior/posterior direction corresponding to temporal hemiretinae fibers that do not cross. Animal studies indicate that there is a significant dorsal-ventral reorganization of the retinotopic distribution of fibers along the optic pathways. We found that diffusion ellipsoids in the central portion of the optic chiasm show considerable planar anisotropy in the coronal plane indicating fiber crossings in the superior/inferior direction, rather than strictly right/left. This architectural feature of the chiasm suggests that dorso-ventral reorganization of fibers in the optic pathways also occurs in humans. We have shown that by collecting sub-millimeter resolution data, DTI can be used to investigate fine details of small and complex white matter structures, in vivo, with a clinical scanner. High spatial resolution, however, is necessary in the slice direction as well as in-plane to reduce the CSF contribution to the signal and to increase fiber coherence within voxels.

Pilocytic astrocytoma of the optic pathway: a tumour deriving from radial glia cells with a specific gene signature.

Pilocytic astrocytomas are WHO grade I gliomas that occur predominantly in childhood. They share features of both astroglial and oligodendroglial lineages. These tumours affect preferentially the cerebellum (benign clinical course) and the optic pathway, especially the hypothalamo-chiasmatic region (poor prognosis). Understanding the molecular basis responsible for the aggressive behaviour of hypothalamo-chiasmatic pilocytic astrocytomas is a prerequisite to setting up new molecular targeted therapies. We used the microarray technique to compare the transcriptional profiles of five hypothalamo-chiasmatic and six cerebellar pilocytic astrocytomas. Validation of the microarray results and comparison of the tumours with normal developing tissue was done by quantitative real-time PCR and immunohistochemistry. Results demonstrate that cerebellar and hypothalamo-chiasmatic pilocytic astrocytomas are two genetically distinct and topography-dependent entities. Numerous genes upregulated in hypothalamo-chiasmatic pilocytic astrocytomas also increased in the developing chiasm, suggesting that developmental genes mirror the cell of origin whereas migrative, adhesive and proliferative genes reflect infiltrative properties of these tumours. Of particular interest, NOTCH2, a gene expressed in radial glia and involved in gliomagenesis, was upregulated in hypothalamo-chiasmatic pilocytic astrocytomas. In order to find progenitor cells that could give rise to hypothalamo-chiasmatic pilocytic astrocytomas, we performed a morphological study of the hypothalamo-chiasmatic region and identified, in the floor of the third ventricle, a unique population of vimentin- and glial fibrillary acidic protein-positive cells highly suggestive of radial glia cells. Therefore, pilocytic astrocytomas of the hypothalamo-chiasmatic region should be considered as a distinct entity which probably originates from a unique population of cells with radial glia phenotype.

Autonomous and non-autonomous Shh signalling mediate the in vivo growth and guidance of mouse retinal ganglion cell axons.

In non-mammalian vertebrates, the relatively homogeneous population of retinal ganglion cells (RGCs) differentiates and projects entirely to the contralateral side of the brain under the influence of sonic hedgehog (Shh). In mammals, by contrast, there are two different RGC types: the Zic2-positive ipsilateral projecting and the Isl2-positive contralateral projecting. We asked whether the axons of these two populations respond to Shh and if their response differs. We have also analysed whether midline- and RGC-derived Shh contributes to the growth of the axons in the proximal visual pathway. We show that these two RGC types are characterised by a differential expression of Shh signalling components and that they respond differently to Shh when challenged in vitro. In vivo blockade of Shh activity, however, alters the path and distribution mostly of the contralateral projecting RGC axons at the chiasm, indicating that midline-derived Shh participates in funnelling contralateral visual fibres in this region. Furthermore, interference with Shh signalling in the RGCs themselves causes abnormal growth and navigation of contralateral projecting axons in the proximal portion of the pathway, highlighting a novel cell-autonomous mechanism by which Shh can influence growth cone behaviour.

The growth-inhibitory protein Nogo is involved in midline routing of axons in the mouse optic chiasm.

We have investigated the role of Nogo, a protein that inhibits regenerating axons in the adult central nervous system, on axon guidance in the developing optic chiasm of mouse embryos. Nogo protein is expressed by radial glia in the midline within the optic chiasm where uncrossed axons turn, and the Nogo receptor (NgR) is expressed on retinal neurites and growth cones. In vitro neurite outgrowth from both dorsonasal and ventrotemporal retina was inhibited by Nogo protein, and this inhibition was abolished by blocking NgR activity. In slice cultures of the optic pathway, blocking NgR with a peptide antagonist produced significant reduction in the uncrossed projection but had no effect on the crossing axons. This result was confirmed by treating cultures with an anti-Nogo functional blocking antibody. In vitro coculture assays of retina and optic chiasm showed that NgR was selectively reduced on neurites and growth cones from dorsonasal retina when they contacted chiasm cells, but not on those from ventrotemporal retina. These findings provide evidence that Nogo signaling is involved in directing the growth of axons in the mouse optic chiasm and that this process relies on a differential regulation of NgR on axons from the dorsonasal and ventrotemporal retina.

Zic2 promotes axonal divergence at the optic chiasm midline by EphB1-dependent and -independent mechanisms.

Axons of retinal ganglion cells (RGCs) make a divergent choice at the optic chiasm to cross or avoid the midline in order to project to ipsilateral and contralateral targets, thereby establishing the binocular visual pathway. The zinc-finger transcription factor Zic2 and a member of the Eph family of receptor tyrosine kinases, EphB1, are both essential for proper development of the ipsilateral projection at the mammalian optic chiasm midline. Here, we demonstrate in mouse by functional experiments in vivo that Zic2 is not only required but is also sufficient to change the trajectory of RGC axons from crossed to uncrossed. In addition, our results reveal that this transcription factor regulates the expression of EphB1 in RGCs and also suggest the existence of an additional EphB1-independent pathway controlled by Zic2 that contributes to retinal axon divergence at the midline.

Diversity in mammalian chiasmatic architecture: ipsilateral axons are deflected at glial arches in the prechiasmatic optic nerve of the eutherian Tupaia belangeri.

Permanent ipsilaterally projecting axons approach the chiasmatic midline in rodents but are confined to lateral parts of the optic chiasm in marsupials. Hence, principally different mechanisms were thought to underlie axon pathway choice in eutherian (placental) and marsupial mammals. First evidence of diversity in eutherian chiasmatic architecture came from studies in the newborn and adult tree shrew Tupaia belangeri (Jeffery et al. [1998] J. Comp. Neurol. 390:183-193). Here, as in marsupials, ipsilaterally projecting axons do not approach the midline. The present study aims to clarify how the developing tree shrew chiasm is organized, how glial cells are arranged therein, and the extent to which the tree shrew chiasm is similar to that of marsupials or other eutherians. By using routinely stained serial sections as well as immunohistochemistry with antibodies against glial fibrillary acidic protein, vimentin, and medium-molecular-weight neurofilament protein, we investigated chiasm formation from embryonic day 18 (E18) to birth (E43). From E22 onward, ipsilaterally projecting axons diverged from contralaterally projecting axons in prechiasmatic parts of the optic nerve. They made sharp turns when arriving at glial arches found at the transition from the optic nerve to the chiasm. Thus, during the ingrowth period of axons, Tupaia belangeri and marsupials have specialized glial arrays in common, which probably help to deflect ipsilaterally projecting axons to lateral parts of the chiasm. Our observations provide new evidence of diversity in eutherian chiasmatic architecture and identify Tupaia belangeri as an appropriate animal model for studies on the mechanisms underlying axon guidance in the developing chiasm of higher primates.

Role of protein kinase C in selective inhibition of mouse retinal neurites during contacts with chondroitin sulfates.

Chondroitin sulfate proteoglycans elicit a selective inhibition to neurite growth from ventrotemporal (VT) but not dorsonasal (DN) retina, potentiating the bilateral routing of axons in the mouse optic chiasm. We examined whether this selective response is mediated by a difference in protein kinase C (PKC) expression. Effects of suppressing PKC activity in explant preparations of embryonic day 14 retinae with inhibitor Gö6976 or Ro-32-0432 abolished the chondroitin sulfate inhibition to the VT neurites but had no effect to the DN neurites. Whether these responses rely on a difference in expression of PKC in the growth cones was examined using antibodies against six isozymes of PKC. Among these the alpha, betaI and epsilon isozymes were expressed prominently in the retinal growth cones; whilst the betaII, delta and gamma isozymes were barely detected. Moreover, while the alpha and epsilon isozymes were abundant in the filopodial and lamellipodial processes, the betaI isozyme was restricted largely in the core region of the growth cones. Despite these subtype specific localization, there was no significant difference in expression of any of these PKC isozymes between growth cones from VT and DN retina, indicating that the selective response to chondroitin sulfates is not likely generated by a regulation of PKC expression, but by expression of surface molecules that interact with chondroitin sulfate proteoglycans.

High-resolution diffusion tensor imaging and tractography of the human optic chiasm at 9.4 T.

The optic chiasm with its complex fiber micro-structure is a challenge for diffusion tensor models and tractography methods. Likewise, it is an ideal candidate for evaluation of diffusion tensor imaging tractography approaches in resolving inter-regional connectivity because the macroscopic connectivity of the optic chiasm is well known. Here, high-resolution (156 microm in-plane) diffusion tensor imaging of the human optic chiasm was performed ex vivo at ultra-high field (9.4 T). Estimated diffusion tensors at this high resolution were able to capture complex fiber configurations such as sharp curves, and convergence and divergence of tracts, but were unable to resolve directions at sites of crossing fibers. Despite the complex microstructure of the fiber paths through the optic chiasm, all known connections could be tracked by a line propagation algorithm. However, fibers crossing from the optic nerve to the contralateral tract were heavily underrepresented, whereas ipsilateral nerve-to-tract connections, as well as tract-to-tract connections, were overrepresented, and erroneous nerve-to-nerve connections were tracked. The effects of spatial resolution and the varying degrees of partial volume averaging of complex fiber architecture on the performance of these methods could be investigated. Errors made by the tractography algorithm at high resolution were shown to increase at lower resolutions closer to those used in vivo. This study shows that increases in resolution, made possible by higher field strengths, improve the accuracy of DTI-based tractography. More generally, post-mortem investigation of fixed tissue samples with diffusion imaging at high field strengths is important in the evaluation of MR-based diffusion models and tractography algorithms.

Enzymatic removal of hyaluronan affects routing of axons in the mouse optic chiasm.

Perturbations of interaction of hyaluronan (HA) with its receptor CD44 cause multiple errors in axon routing at the mouse optic chiasm. To investigate this interaction further on the chiasm routing, we studied the axon routing after enzymatic removal of HA from slice preparations of the optic pathway. Hyaluronidase treatment produced an obvious reduction in midline crossing of the first generated axons in E13 chiasms, but had no influence on routing ofthe uncrossed axons in E15 and E16 slices. These findings support a direct role of HA, acting probably through CD44, on axon decussation during early phase of chiasm development, but argue against a direct function of HA on the turning of uncrossed axons in the mouse optic chiasm.