Loss of zebrafish Ataxin-7, a SAGA subunit responsible for SCA7 retinopathy, causes ocular coloboma and malformation of photoreceptors.

Polyglutamine (polyQ) expansion in Ataxin-7 (ATXN7) results in spinocerebellar ataxia type 7 (SCA7) and causes visual impairment. SCA7 photoreceptors progressively lose their outer segments (OSs), a structure essential for their visual function. ATXN7 is a subunit of the transcriptional coactivator Spt-Ada-Gcn5 Acetyltransferase complex, implicated in the development of the visual system in flies. To determine the function of ATXN7 in the vertebrate eye, we have inactivated ATXN7 in zebrafish. While ATXN7 depletion in flies led to gross retinal degeneration, in zebrafish, it primarily results in ocular coloboma, a structural malformation responsible for pediatric visual impairment in humans. ATXN7 inactivation leads to elevated Hedgehog signaling in the forebrain, causing an alteration of proximo-distal patterning of the optic vesicle during early eye development and coloboma. At later developmental stages, malformations of photoreceptors due to incomplete formation of their OSs are observed and correlate with altered expression of crx, a key transcription factor involved in the formation of photoreceptor OS. Therefore, we propose that a primary toxic effect of polyQ expansion is the alteration of ATXN7 function in the daily renewal of OS in SCA7. Together, our data indicate that ATXN7 plays an essential role in vertebrate eye morphogenesis and photoreceptor differentiation, and its loss of function may contribute to the development of human coloboma.

Anatomy of the stemmata in the Photuris firefly larva.

Fireflies (Coleoptera: Lampyridae) have distinct visual systems at different stages of development. Larvae have stemmata and adults have compound eyes. Adults use compound eyes to mediate photic communication during courtship. Larvae do not manifest this behavior, yet they are bioluminescent. We investigated the structure of stemmata in Photuris firefly larvae to identify anatomical substrates (i.e., rhabdomeres) conferring visual function. Stemmata were located bilaterally on the antero-lateral surfaces of the head. Beneath the ~ 130 µm diameter lens, we identified a pigmented eye-cup. At its widest point, the eye-cup was ~ 150 µm in diameter. The optic nerve exited the eye-cup opposite the lens. Two distinct regions, asymmetric in size and devoid of pigmentation, were characterized in stemmata cross-sections. We refer to these regions as lobes. Each lobe contained a rhabdom of a radial network of rhabdomeres. Pairs of rhabdomeres formed interdigitating microvilli contributed from neighboring photoreceptor cell bodies. The optic nerve contained 88 axons separable into two populations based on size. The number of axons in the optic nerve together with distinct rhabdoms suggests these structures were formed from 'fusion stemmata.' This structural specialization provides an anatomical substrate for future studies of visually mediated behaviors in Photuris larvae.

Expression of the lymphatic marker podoplanin (D2-40) in human fetal eyes.

During human ocular development, expression of proteins varies in different maturation stages. This study aims to characterize structures in human fetal eyes stained by the lymphatic marker podoplanin (D2-40) with emphasis on the stage of maturation and the presence of intraocular lymphatic structures. Formalin-fixed paraffin-embedded eyes from 40 human fetuses between 10 and 38 weeks of gestation (WoG) were investigated. Immunohistochemical stains were performed for D2-40, LYVE-1 as a secondary lymphatic marker, and CD34 as a control for endothelial reactivity. A semiquantitative analysis of antigen expression in different segments of the eye was performed by light microscopy. The intensity of antigen expression was graded with a score ranging from 0 to 3. Podoplanin expression was found with a variable intensity in 97.5% of the eyes, in particular in lymphatic vessels of the conjunctiva (n = 26), conjunctival and corneal epithelium (n = 33), corneal endothelium (n = 4), trabecular meshwork (n = 28), and optic nerve sheaths (n = 23). A slight, equivocal staining reaction was noted in the choroid (n = 14). There was a correlation of antigen reactivity and the gestational age for corneal endothelial reactivity in earlier gestational stages (p = 0.003) and trabecular meshwork in older eyes (p = 0.031). D2-40 positive Müller cells were detected in two eyes ≥32 WoG. Thus, aside from conjunctival lymphatic vessels, podoplanin was expressed in several structures of the human fetal eye and the ocular adnexae at different gestational stages. Podoplanin positive structures were also found in the choroid and the chamber angle. However, lymphatic vessels or its progenitors could not be unequivocally identified in intraocular structures during 10-38 weeks of gestation. There is no evidence from our data that transient intraocular lymphactics develop in the fetal eye between 10 and 38 weeks of gestation.

Relaxation-expansion model for self-driven retinal morphogenesis: a hypothesis from the perspective of biosystems dynamics at the multi-cellular level.

The generation of complex organ structures such as the eye requires the intricate orchestration of multiple cellular interactions. In this paper, early retinal development is discussed with respect to the structure formation of the optic cup. Although recent studies have elucidated molecular mechanisms of retinal differentiation, little is known about how the unique shape of the optic cup is determined. A recent report has demonstrated that optic-cup morphogenesis spontaneously occurs in three-dimensional stem-cell culture without external forces, indicating a latent intrinsic order to generate the structure. Based on this self-organizing phenomenon, we introduce the "relaxation-expansion" model to mechanically interpret the tissue dynamics that enable the spontaneous invagination of the neural retina. This model involves three consecutive local rules (relaxation, apical constriction, and expansion), and its computer simulation recapitulates the optic-cup morphogenesis in silico.

Dynamics of optic canal and orbital cavity development revealed by microCT.

PURPOSE: Numerous studies have attempted to clarify the exact anatomy and variations of the optic canal with non-conclusive results due to its close proximity to many vulnerable structures. We sought to determine the dynamics of growth and development of these structures on fetal skulls, which will help us to better understand of gender and age-dependent variations, as well as fatal malformations. METHODS: Fifteen previously macerated fetal frontal and sphenoid bones were analyzed and the diameters of optic canal, and distance of orbit from frontomaxillary suture to frontozygomatic suture were measured using 3D reconstruction images obtained by micro-CT. RESULTS: Average diameter of the optic canal in 300 mm fetus was measured to be 1,546 ± 36 µm, in 400 mm fetus 2,470 ± 123 µm and in 500 mm fetus 3,757 ± 203 µm. This trend indicates a linear enlargement of optic canal during the fetal period. During the same time period, diameter of the orbit enlarges from 12,319 ± 559 µm in 300 mm fetus to 19,788 ± 736 µm in 500 mm fetus. Growth curve is significantly lower in comparison with the same curve in optic canal data. We also calculated the ratio of orbit diameter and optic canal diameter between those groups which decreased from a value of 7.9 ± 0.4 for 300 mm fetus to 5.3 ± 0.2 for 500 mm fetus. CONCLUSION: Dynamics of optic canal and orbital cavity development is different in early and late fetal period. Diameters of those structures are in better correlation with the fetal length.

Olfactomedin 1 interacts with the Nogo A receptor complex to regulate axon growth.

Olfm1, a secreted highly conserved glycoprotein, is detected in peripheral and central nervous tissues and participates in neural progenitor maintenance, cell death in brain, and optic nerve arborization. In this study, we identified Olfm1 as a molecule promoting axon growth through interaction with the Nogo A receptor (NgR1) complex. Olfm1 is coexpressed with NgR1 in dorsal root ganglia and retinal ganglion cells in embryonic and postnatal mice. Olfm1 specifically binds to NgR1, as judged by alkaline phosphatase assay and coimmunoprecipitation. The addition of Olfm1 inhibited the growth cone collapse of dorsal root ganglia neurons induced by myelin-associated inhibitors, indicating that Olfm1 attenuates the NgR1 receptor functions. Olfm1 caused the inhibition of NgR1 signaling by interfering with interaction between NgR1 and its coreceptors p75NTR or LINGO-1. In zebrafish, inhibition of optic nerve extension by olfm1 morpholino oligonucleotides was partially rescued by dominant negative ngr1 or lingo-1. These data introduce Olfm1 as a novel NgR1 ligand that may modulate the functions of the NgR1 complex in axonal growth.

Mutations in the homeobox gene HESX1/Hesx1 associated with septo-optic dysplasia in human and mouse.

During early mouse development the homeobox gene Hesx1 is expressed in prospective forebrain tissue, but later becomes restricted to Rathke's pouch, the primordium of the anterior pituitary gland. Mice lacking Hesx1 exhibit variable anterior CNS defects and pituitary dysplasia. Mutants have a reduced prosencephalon, anopthalmia or micropthalmia, defective olfactory development and bifurcations in Rathke's pouch. Neonates exhibit abnormalities in the corpus callosum, the anterior and hippocampal commissures, and the septum pellucidum. A comparable and equally variable phenotype in humans is septo-optic dysplasia (SOD). We have cloned human HESX1 and screened for mutations in affected individuals. Two siblings with SOD were homozygous for an Arg53Cys missense mutation within the HESX1 homeodomain which destroyed its ability to bind target DNA. These data suggest an important role for Hesx1/HESX1 in forebrain, midline and pituitary development in mouse and human.

Is the term "fasciculus opticus cerebralis" more justifiable than the term "optic nerve"?

The terminology of the optic nerve had already been changed three times, since 1895 until 1955 when the term "nervus opticus" was introduced in the "Terminologia Anatomica". Following our study we claim that, from the aspect of phylogenetic evolution of binocular vision development as well as optical embryogenesis where opticus is evidently presented as a product of diencephalic structures, the addition of the term "nervus" to opticus is not adequate and justified. From the clinical aspect the term "nervus opticus" is also inadequate, both as a "nerve" that has no functional regenerative properties, unlike other cranial nerves, as well as from a pedagogical and didactical aspect of educating future physicians. We suggest that the term "Fasciculus Opticus Cerebralis" should be used as it much better explains the origin as well as its affiliation to the central nervous system.

Megalin mediates the influence of sonic hedgehog on oligodendrocyte precursor cell migration and proliferation during development.

Oligodendrocyte precursor cells (OPCs) of the optic nerve are generated in the preoptic area, from where they migrate to colonize it entirely. Sonic hedgehog (Shh) induces the proliferation of these cells as well as influencing their migration, acting through its canonical receptor (Ptc-1). However, the multiligand receptor megalin (or LRP-2) is also involved in Shh-induced OPC proliferation and migration, and thus, we have evaluated the relevance of this interaction. During the stages at which Shh influences OPC development, we found megalin to be selectively expressed by optic nerve astrocytes, whereas Ptc-1 and Gli1 were found in OPCs. Indeed, this pattern of expression paralleled the rostral-caudal expression of the three Shh-related molecules during the time course of plp-dm20(+) -OPC colonization. The blockage of megalin partially abolished OPC chemoattraction and fully impaired Shh-induced proliferation. Using in vitro co-cultures of dissociated optic nerve cells, we demonstrated that Shh was internalized by astrocytes via megalin, and sufficient Shh was subsequently released to produce the biological effects on OPCs observed in the nerve. Together, these data indicate that at least part of the influence of Shh on OPCs is mediated by megalin during optic nerve development, and that astrocytes expressing megalin transiently capture Shh to present it to OPCs and/or to control the gradient of this molecule during development.

AP-2alpha knockout mice exhibit optic cup patterning defects and failure of optic stalk morphogenesis.

Appropriate development of the retina and optic nerve requires that the forebrain-derived optic neuroepithelium undergoes a precisely coordinated sequence of patterning and morphogenetic events, processes which are highly influenced by signals from adjacent tissues. Our previous work has suggested that transcription factor activating protein-2 alpha (AP-2alpha; Tcfap2a) has a non-cell autonomous role in optic cup (OC) development; however, it remained unclear how OC abnormalities in AP-2alpha knockout (KO) mice arise at the morphological and molecular level. In this study, we show that patterning and morphogenetic defects in the AP-2alpha KO optic neuroepithelium begin at the optic vesicle stage. During subsequent OC formation, ectopic neural retina and optic stalk-like tissue replaced regions of retinal pigment epithelium. AP-2alpha KO eyes also displayed coloboma in the ventral retina, and a rare phenotype in which the optic stalk completely failed to extend, causing the OCs to be drawn inward to the midline. We detected evidence of increased sonic hedgehog signaling in the AP-2alpha KO forebrain neuroepithelium, which likely contributed to multiple aspects of the ocular phenotype, including expansion of PAX2-positive optic stalk-like tissue into the OC. Our data suggest that loss of AP-2alpha in multiple tissues in the craniofacial region leads to severe OC and optic stalk abnormalities by disturbing the tissue-tissue interactions required for ocular development. In view of recent data showing that mutations in human TFAP2A result in similar eye defects, the current findings demonstrate that AP-2alpha KO mice provide a valuable model for human ocular disease.

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.

The dural sheath of the optic nerve: descriptive anatomy and surgical applications.

The aim of this work was to clarify the descriptive anatomy of the optic dural sheath using microanatomical dissections on cadavers. The orbit is the rostral part of the extradural neural axis compartment; the optic dural sheath forms the central portion of the orbit.In order to describe this specific anatomy, we carefully dissected 5 cadaveric heads (10 orbits) up to the meningeal structure of the orbit and its contents. 1 cadaveric head was reserved for electron microscopy to add to our knowledge of the collagen structure of the optic dural sheath.In this chapter, we describe the anatomy of the interperiostal-dural concept and the anatomy of the orbit. The optic dural sheath contains three portions: the intracranial, the intracanalicular and the intraorbital segment. Each one has specific anatomic relations which result in particular surgical considerations.

Spatiotemporal distribution of chondroitin sulfate proteoglycans in the developing mouse retina and optic nerve.

The aim of the present study was to determine the distribution of chondroitin sulfate proteoglycans in the mouse retina and optic nerve of the prenatal and postnatal mouse by immunohistochemistry. At embryonic day (E) 18, chondroitin-4-sulfate (C4S), chondroitin-6-sulfate (C6S) and biglycan were detected in the retina and optic nerve. However, aggrecan was seen in the retina but not in the optic nerve. At postnatal day (P) 7, aggrecan and biglycan were clearly observed in the optic nerve, inner nuclear layer and ganglion cell layer and diffuse in the outer retina. C4S diffusely distributed in the retina and optic nerve, but C6S was mainly confined to the photoreceptor layer and optic nerve sheath. At P42, biglycan showed diffuse distribution in the retina and optic nerve with intense staining in nerve-fiber rich layers. Aggrecan showed weak staining at the inner plexiform layer with higher density in the outer and inner nuclear layers, outer plexiform layer and ganglion cell layer. Both C4S and C6S were detected in the optic nerve and retina, but C6S showed strong immunostaining in the photoreceptor layer. The distributions of these proteoglycans with respect of time course during development of the retina and optic nerve suggest that they may have unique or overlapping roles in development and maintenance of the retina and optic nerve.

VEGF-C is a trophic factor for neural progenitors in the vertebrate embryonic brain.

Vascular endothelial growth factor C (VEGF-C) was first identified as a regulator of the vascular system, where it is required for the development of lymphatic vessels. Here we report actions of VEGF-C in the central nervous system. We detected the expression of the VEGF-C receptor VEGFR-3 in neural progenitor cells in Xenopus laevis and mouse embryos. In Xenopus tadpole VEGF-C knockdowns and in mice lacking Vegfc, the proliferation of neural progenitors expressing VEGFR-3 was severely reduced, in the absence of intracerebral blood vessel defects. In addition, Vegfc-deficient mouse embryos showed a selective loss of oligodendrocyte precursor cells (OPCs) in the embryonic optic nerve. In vitro, VEGF-C stimulated the proliferation of OPCs expressing VEGFR-3 and nestin-positive ventricular neural cells. VEGF-C thus has a new, evolutionary conserved function as a growth factor selectively required by neural progenitor cells expressing its receptor VEGFR-3.

Eph receptors are negatively controlled by protein tyrosine phosphatase receptor type O.

Eph receptors are activated by the autophosphorylation of tyrosine residues upon the binding of their ligands, the ephrins; however, the protein tyrosine phosphatases (PTPs) responsible for the negative regulation of Eph receptors have not been elucidated. Here, we identified protein tyrosine phosphatase receptor type O (Ptpro) as a specific PTP that efficiently dephosphorylates both EphA and EphB receptors as substrates. Biochemical analyses revealed that Ptpro dephosphorylates a phosphotyrosine residue conserved in the juxtamembrane region, which is required for the activation and signal transmission of Eph receptors. Ptpro thus seems to moderate the amount of maximal activation of Eph receptors. Using the chick retinotectal projection system, we show that Ptpro controls the sensitivity of retinal axons to ephrins and thereby has a crucial role in the establishment of topographic projections. Our findings explain the molecular mechanism that determines the threshold of the response of Eph receptors to ephrins in vivo.

Disruption of Nectin-like 1 cell adhesion molecule leads to delayed axonal myelination in the CNS.

Nectin-like 1 (Necl-1) is a neural-specific cell adhesion molecule that is expressed in both the CNS and PNS. Previous in vitro studies suggested that Necl-1 expression is essential for the axon-glial interaction and myelin sheath formation in the PNS. To investigate the in vivo role of Necl-1 in axonal myelination of the developing nervous system, we generated the Necl-1 mutant mice by replacing axons 2-5 with the LacZ reporter gene. Expression studies revealed that Necl-1 is exclusively expressed by neurons in the CNS. Disruption of Necl-1 resulted in developmental delay of axonal myelination in the optic nerve and spinal cord, suggesting that Necl-1 plays an important role in the initial axon-oligodendrocyte recognition and adhesion in CNS myelination.

Impairing retinoic acid signalling in the neural crest cells is sufficient to alter entire eye morphogenesis.

Retinoic acid (RA) is known to be required at various levels of eye patterning via Retinoic Acid Receptors (RAR); however the molecular and cellular mechanisms triggered by these nuclear receptors are still obscure. The genetic studies performed here enable us to present a new model to study RA action during eye development. By inactivating the three RARs, specifically in the periocular mesenchyme, we discriminate the individual contribution of each RAR during eye development and describe a new function for RARs during the formation of the optic nerve. We demonstrate that RARalpha is the only receptor that mediates RA signalling in the neurectoderm during ocular development. Surprisingly, and despite a sophisticated pattern of RA-activity in the developing retina, we observed that RA signalling is not autonomously required in this tissue for eye formation. We show that the action of RA during eye morphogenesis is occurring specifically in neural crest-derived periocular mesenchyme and is mediated by all three RARs. Furthermore, we point out that Pitx2, which encodes a homeodomain transcription factor, is a key RA-responsive gene in neural crest cells during eye development. Interestingly, we observed that RA is required in the neural crest cells for normal position of the extraocular muscle.

Topologically restricted appearance in the developing chick retinotectal system of Bravo, a neural surface protein: experimental modulation by environmental cues.

A novel neural surface protein, Bravo, shows a pattern of topological restriction in the embryonic chick retinotectal system. Bravo is present on the developing optic fibers in the retina; however, retinal axons in the tectum do not display Bravo. The appearance of Bravo in vitro is modulated by environmental cues. Axons growing out from retinal explants on retinal basal lamina, their natural substrate, express Bravo, whereas such axons growing on collagen do not. Retinal explants provide a valuable system to characterize the mechanism of Bravo restriction, as well as the cellular signals controlling it. Bravo was identified with monoclonal antibodies from a collection generated against exposed molecules isolated by using a selective cell surface biotinylation procedure. The NH2-terminal sequence of Bravo shows similarity with L1, a neural surface molecule which is a member of the immunoglobulin superfamily. This possible relationship to L1, together with its restricted appearance, suggests an involvement of Bravo in axonal growth and guidance.

Overproduction and elimination of retinal axons in the fetal rhesus monkey.

Quantitative electron microscopic analysis reveals 2.85 million retinal axons in fetal rhesus monkeys--a number that is more than twice the 1.2 million present in the adult. More than 1 million supernumerary optic axons are eliminated before birth, simultaneously with the segregation of inputs from the two eyes into separate layers of the lateral geniculate nucleus. Selective elimination of optic axons may not only play a role in the segregation of binocular visual connections but, secondarily, may establish the ratio of crossed and uncrossed retinogeniculate projections.