The response of optic tract glia during regeneration of the goldfish visual system. I. Biosynthetic activity within different glial populations after transection of retinal ganglion cell axons

We monitored biosynthetic activity of optic tract glia during regeneration of retinal ganglion cell axons in the goldfish and found that the greatest level of incorporated [3H]thymidine and [3H]leucine occurred in glia by 10-15 days after axotomy. During this period there was a marked increase in the number of oligodendroglia and multipotential glia near the site of injury with no change occurring in the astroglial population. Electron microscopic autoradiography showed that oligodendroglia and multipotential cells incorporated 5-7-fold more thymidine than did cells of intact control preparations. Though all glial cell types incorporated more [3H]leucine during axonal regeneration, oligodendroglia and multipotential cells together accounted for more than 90% of measured radioactivity. In order to characterize glial-stimulating events specific to axonal regeneration, we produced axonal degeneration in the optic tract by removal of the retina. Optic tract glia during axonal degeneration incorporated less amino acid when compared to glia associated with regenerating axons. The degenerating optic tract also had less 2',3'-cyclic nucleotide 3'-phosphohydrolase, an enzyme produced by oligodendroglia, than that found in the regenerating visual system. Our results suggest that in response to ganglion cell axotomy oligodendroglia and multipotential glia of the goldfish optic tract proliferate. Moreover, regenerating axons provide one type of stimulant for glial protein biosynthesis.     

Time course and pattern of optic fiber regeneration following tectal lobe removal in the goldfish

Following single tectal lobe removal in the adult goldfish, Carassius auratus, the pattern of regeneration of the optic fibers which had previously projected to that tectum was examined at 1, 2, 4, 6, 8, 10, and 12 weeks postoperative using ³H-proline radioautography. We found that regenerating optic fibers grew across the midline through the transverse, minor, horizontal, and posterior commissures to innervate the remaining tectum. At early postoperative times innervation of the tectum was continuous, while later, the regenerating fibers segregated into discrete patches in the superficial layers of the tectum. In addition, regenerating fibers also grew into non-optic centers/pathways such as the habenula, the fasciculus retroflexus, the forebrain, the torus semicircularis, the valvula and corpus cerebelli, the hypothalamus, and the medulla. While optic fibers were no longer apparent in the habenula and the fasciculus retroflexus after 2 weeks postoperative, all other structures were still occupied by the fibers at 12 weeks postoperative. Since most of the innervated pathways were either tectal efferent pathways, which should contain degenerating debris and proliferating glial cells after the tectal removal, or pathways closely associated with traumatized areas, we suggest that degenerating axonal debris and proliferating glia may play an important role in guiding regenerating fibers in this system.     

Denervation of non-optic brain areas along the course of the optic tract does not affect the success of optic nerve regeneration in frogs

Unsuccessful axonal regeneration in frog or goldfish spinal cord is thought to be due in part to inappropriate denervated synaptic sites attracting regenerating axons as they pass by the lesion zone (Bernstein and Bernstein, '69; Bernstein and Gelderd, '73). The reported success of optic nerve regeneration in these same animals may result because there are no inappropriate synaptic sites available to the regenerating axons. To test for this we denervated areas within thalamus and mesencephalon which lie along the path of regenerating optic axons to determine whether or not abnormal connections would form in these areas and affect the success of optic nerve regeneration. After transection of the brainstem through the isthmal region, terminal degeneration was found in several zones adjacent to optic nerve targets or the optic tract (i.e., nucleus rotundus, corpus geniculatum laterale, and torus semicircularis). In adult frogs receiving left isthmal transection, we also crushed the right optic nerve and then examined the regenerated optic projection at intervals up to six months after nerve crush using anterograde transport of ³H-proline. In no instance did the regenerating optic axons alter their distribution within visual neuropil zones or invade those areas deafferented by the isthmal lesion. Histological study showed that the axons cut by the isthmal lesion did not regenerate back to their sites to prevent the invasion of optic axons into these zones. We then attempted to force optic axons into foreign territory by removing a major projection zone, the optic tectum. With tectal ablation and isthmal transection, regenerating optic axons were offered synaptic space made available by both lesions. However, we found abnormal optic projections only in the middle and posterior thalamic neuropil and in the remaining tectal hemisphere. Optic axons did not expand into any of the areas deafferented by the isthmal transection, even though some of them were further denervated by the tectal ablation. We conclude that optic axons will not invade non-optic areas deafferented by an isthmal lesion even if a large number of optic axons have no normal target to innervate.     

Distribution of [3H]RNA in goldfish optic tectum following intraocular or intracranial injection of [3H]uridine. Evidence of axonal migration of RNA in regenerating optic fibers.

The distribution of [³H]RNA in the goldfish optic tectum following eitherintra-ocular orintracranial injection of [³H]uridine during optic fiber regeneration has been studied by light (LMA) and electron (EMA) microscopic autoradiography.In one group of 4 fish both optic nerves were crushed, and 18 days later [³H]uridine was injected into the right eye. A second group of 5 fish, in which only one optic nerve had been crushed, received intracranial injections of [³H]uridine 18 or 22 days after the crush. All fish were sacrificed 24 days after crushing the optic nerves, a time when regenerating optic fibers have entered the tectum and are establishing functional reconnections. Tecta were fixed in situ with glutaraldehyde, dissected out, and samples were processed for LMA and EMA. Controls were carried out to ensure that [³H]RNA was the only radioactive component present in the tissue after fixation.The distribution of silver grains related to [³H]RNA in intraocularly injected goldfish was different from that following intracranial injection. Following intraocular injection virtually all the [³H]RNA was located in the layers of the left optic tectum (contralateral to the side of intraocular injection) where the regenerating optic fibers course and terminate, whereas virtually no radioactivity was present in the right optic tectum. EMA quantitative analysis of the labeled layers of the left optic tectum revealed that perikarya of cells, most of which are glial cells, had a density of grains related to [³H]RNA of 20–28 g/100 sq.μm; axonal growth cones had a density of 14–24 g/100 sq.μm. Grain densities over non-axonal cell structures were markedly lower, ranging between 3 and 6 g/sq.μm. Grains located over axons and growth cones accounted for 50–60% of all counted grains.Inintracranially injected goldfish, either 2 or 6 days after injection, silver grains were clustered over leptomeninges as well as vessels and parenchymal cells of the tectal strata containing the regenerating optic fibers. In the stratum opticum a high grain density was seen over glial cells, whereas virtually no grains were present over the fascicles of regenerating axons. EMA quantitative analysis revealed a grain density over glial and other parenchymal cells of the stratum opticum of 67 g/100 sq.μm, whereas densities over growth cones and regenerating axons were 1.3 g/100 sq.μm and 1.8 g/100 sq.μm respectively. Grains located over axons and growth cones accounted for 3.3% of all counted grains.On the basis of the present and previous findings it is suggested that followingintraocular injection of [³H]uridine the [³H]RNA present inside regenerating optic axons is transported from the ganglion cells of the retina; on the other hand, the [³H]RNA present in surrounding glial cells is the result of local utilization of [³H]RNA precursors which also migrate from the retina along with the [³H]RNA.It is also concluded that 2 and 6 days followingintracranial injection of [³H]uridine no substantial tranfer of [³H]RNA from glial cells to regenerating optic fibers occurs in the goldfish optic tectum.     

Anatomical evidence for the influence of degenerating pathways on regenerating optic fibers following surgical manipulations in the visual system of the goldfish

We have used [³H]proline radioautography to trace regenerating optic fibers in the goldfish following: (1) the removal of the right tectal lobe and the right eye, and (2) the removal of both tectal lobes. Our results indicate that following the removal of the right tectal lobe and the right eye, both the denervated tectal efferent pathways, and the denervated visual pathways and terminal zones of the enucleated eye were penetrated by the regenerating optic fibers. In addition, following bilateral lobectomy, the denervated tectal efferent pathways were bilaterally penetrated by the regenerating fibers. Since, in both types of operations, these denervated pathways and terminal zones should undergo degeneration, our results support the suggestion that the presence of degenerating axonal debris and proliferating glia may play an important role in guiding regenerating optic fibers in the visual system of the goldfish.     

Gliosis during optic fiber regeneration in the goldfish: an immunohistochemical study.

Antisera directed against the 48 kDa and 50 kDa cytoskeletal antigens were used to examine changes in the astroglial fabric of the goldfish visual pathways following optic nerve crush. Several major observations are described. First, an optic nerve crush lesion in these animals appears to be devoid of glial cells for at least the first month after surgery. As a corollary, regenerating axons that grow across the lesion may do so over an aglial substrate. Once the axons cross the lesion, their growth is confined to the astroglial domains of the proximal nerve stump. In the optic nerve, gliosis comprises hypertrophy of astrocytic processes such that the open framework characterizing the normal nerve is obscured. In addition, during regeneration, optic nerve glia express large amounts of the 50 kDa cytoskeletal protein, which they ordinarily express at only minimal levels. In the optic tract, gliosis is reflected in a markedly increased expression of the 50 kDa protein as well as an apparent increase in the number and complexity of glial processes. In addition, optic tract glia begin to express the 48 kDa antigen during regeneration. This protein is ordinarily confined for the most part to the optic nerve and is not seen in the tract glia. Finally, no obvious changes were seen in the glia of the optic tectum. These results demonstrate many points of similarity between gliosis in the goldfish and in mammals. However, in some particulars the two responses differ, and it is possible that these differences are related to the differing ability of central axons to regenerate in the two groups of organisms.     

Retinal projections throughout optic nerve regeneration in the ornate dragon lizard, Ctenophorus ornatus

In goldfish and frog, optic nerve regeneration is successful, with restoration of retinotopic projections in visual brain centres and the return of functional vision within 1-2 months. By contrast, at 1 year after unilateral optic nerve crush in the ornate dragon lizard (Ctenophorus ornatus), the regenerated retinotectal projections lack topographic order, presumably explaining why the lizards are blind via the experimental eye (Beazley et al. [1997] J. Comp. Neurol. 377:105-120). To determine whether other abnormalities are associated with the inability to restore topographic projections in the lizard, we charted anatomically the time course, accuracy, and stability of optic nerve regeneration by examining visual projections with the lipophillic dye 1,1'-dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine perchlorate (DiI) applied to the optic disk at intervals up to 1 year after optic nerve crush; in addition, DiI tracing of small groups of axons was used to examine the topicity of axons projecting to the tectum. Axons re-innervated visual centres from between 1 and 2 months, a time frame comparable with that in goldfish and frog. However, the projections in lizard were found to differ from those in goldfish and frog in three major ways. First, there was considerable variability within the projection patterns both between individual lizards at any one stage and with time. Second, the projections were inaccurate. As in normal lizards, the major projection was to the contralateral optic tectum, although it lacked detectable retinotopic axon order throughout. Furthermore, misrouting occurred such that regenerating axons formed a persistent projection to the ipsilateral side of the brain that was considerably stronger and more widespread than normal. Minor visual centres also became re-innervated but, in addition, regenerating axons formed persistent projections into the opposite optic nerve and to non-retino-recipient regions such as the nucleus rotundus, hypothalamus, and olfactory nerve, as well as the posterior and tectal commissures. Third, the projections appeared unstable. Projections to both tecta were strongest between 3 and 5 months, but they diminished thereafter. The results suggest that, compared with goldfish and frog, in lizards both pathway and target cues are degraded and/or cannot be read adequately; as a consequence, regenerating axons are unable to navigate exclusively to visual centres and cannot re-form stable connections.     

Aberrant growth of regenerating retinotectal axons subsequent to optic tract ablation in goldfish

This study examined the effect of optic tract ablation on retinotectal fiber regeneration in goldfish. Approximately two-thirds of the left optic tract was removed, and, at various times post lesion (10–75 days), the course of regenerating retinotectal fibers was traced using horseradish peroxidase. In all experimental animals, axons were observed regenerating through the visual pathway but at the brachia most of the fibers were channeled through the ventral brachium. We present evidence that fibers in the ventral brachium originated from ganglion cells in all regions of retina and that these fibers grew almost exclusively into ventral half tectum even though some of these fibers would normally synapse in dorsal half tectum. These observations suggest that optic tract ablation does not prevent retinal fiber regeneration but results in aberrant growth through the brachia and significant inhibition of exploratory fiber growth within the tectum.     

Changes in the topographically organized connections between the nucleus isthmi and the optic tectum after partial tectal ablation in adult goldfish

The projection of the nucleus isthmi to the ipsilateral optic tectum was examined in normal goldfish. This was compared to the projection in animals in which the entire visual field had been induced to compress onto a rostral half tectum by caudal tectal ablation. The isthmo-tectal projection was examined by making localized injections of horseradish peroxidase into the optic tecta and observing the patterns of labeled cells within the nucleus isthmi. The teleost nucleus isthmi consists of a cell sparse medulla covered by a cellular cortex, which is thick on the rostral, medial, and dorsal surfaces of the nucleus. Almost all isthmic cells projecting to the tectum were located in the area of thick cortex. In normal fish, rostral tectal injections labeled cells in the rostroventral portion of the thick cortex; injections midway in the rostrocaudal tectal axis labeled more caudodorsally located cells, and caudal tectal injections labeled cells a little further caudally in extreme dorsal cortex. The rostroventral to caudodorsal isthmic axis was therefore seen to project rostrocaudally along the tectum. This topography contrasts somewhat with the situation seen in amphibia where the rostrocaudal tectal axis receives projections from the rostrocaudal isthmic axis. In fish with half-tectal ablations, injections near the caudal edge of the half tectum (at a site that had originally been midtectal) labeled cells that had previously projected to caudal tectum. Rostral tectal injections in fish with compression of the visual field gave a normal pattern of labeled isthmic cells. The results indicate that a topographically ordered isthmo-tectal projection exists in goldfish that may be induced to compress onto a half tectum.     

The optic tectum regulates the transport of specific proteins in regenerating optic fibers of goldfish

The pattern of rapidly-transported proteins in regenerating optic fibers of the adult goldfish is regulated by interactions between these fibers and their main target, the optic tectum. When the optic fibers are allowed to interact with the tectum, the transport of proteins with molecular weights in the range of 110-145 kilodaltons (kDa) increases, whereas the transport of proteins in the 24-27 kDa range declines from the previously high level which has been induced by axotomy. If the optic fibers are prevented from interacting with the tectum, the transport of the 24-27 kDa proteins remains elevated for months. Amounts of other rapidly-transported retinal proteins (e.g. the acidic 43-49 kDa proteins that increase in regenerating optic fibers after axotomy) are relatively unaffected by tectal ablation.     

Effect of target removal on goldfish optic nerve regeneration: analysis of fast axonally transported proteins.

How is axonal transport in regenerating neurons affected by contact with their synaptic target? We investigated whether removing the target (homotopic) lobe of the goldfish optic tectum altered the incorporation of 3H-proline into fast axonally transported proteins in the regenerating optic nerve. Regeneration was induced either by an optic tract lesion (to reveal the changes in the original axon segment that remained connected to the cell body) or by an optic nerve lesion (to reveal the changes in the newly formed axon segment). Of 26 proteins analyzed by 2-dimensional gel electrophoresis and fluorography, all but one showed increased labeling as a result of tectal lobe ablation. By 2 d after the lesion, significantly increased labeling of some proteins was seen with a 6-hr labeling interval, but not with a 24-hr labeling interval. This is probably indicative of an increased velocity of transport, which may have been a nonspecific consequence of the surgery. Otherwise, tectal lobe removal had relatively little effect until 3 weeks, when there was a transitory increase in labeling of transported proteins in the new axon segments of the tectum-ablated animals. Beginning at 5 weeks, tectal lobe ablation caused considerably higher labeling of many of the proteins in the original axon segments. Because this was seen with both 6-hr and 24-hr labeling intervals, it is probably indicative of increased protein synthesis. The increased synthesis lasted until at least 12 weeks, though some proteins were beginning to show a diminished effect at this time. In the late stages of regeneration (8-12 weeks), there was also increased labeling of proteins in the new axon segments as a result of the absence of the target tectal lobe. This included a disproportionately large increase in the relative contribution of cytoskeletal proteins and of protein 4, which is the goldfish equivalent of the growth-associated protein GAP-43 (neuromodulin). We conclude that, after the regenerating axons begin to innervate the tectum, the expression of most of the proteins in fast axonal transport is down-regulated by interaction between the axons and their target. However, the changes in expression may be preceded by a modulation of the turnover and/or deposition of proteins in the newly formed axon segment.     

Goldfish retina and tectum influence each other's growth activity during regrowth of the retinotectal projection

Variations in retinal and tectal growth activity, during regrowth of the goldfish retinotectal projection, were monitored by measuring the rates of incorporation of [14C]leucine into soluble protein and tubulin-enriched fractions at different times after crushing the optic nerves. Other experiments tested for growth-modulating interactions between tectum and retina. Here we studied how the absence of one of these structures (i.e. tectal ablation or eye removal) affected the profile of biosynthetic activity in the other. Experiments were also conducted on groups of fish in which the tectum was reinnervated by a half-retina (either half-nasal (1/2 N) or half-temporal (1/2 T) retina). This was done to ascertain if growth interactions between retina and tectum display any position-dependent differences that may be relevant to retinotopic ordering during regeneration. Our studies have revealed that: the retina and tectum of 1/2 T and 1/2 N groups differ in their growth responses during regeneration of the visual pathway: the tectum may exert a stimulatory and at other times an inhibitory influence on retinal protein synthesis; and retina and tectum display a bimodal profile of biosynthetic activity during regeneration that coincides with two stages of increased cell division (primarily glia) which other workers have found occurs in the tectum and tract during regeneration of the retinotectal projection. Indeed it seems there may be a link between this glial proliferation and the neurotrophic and guiding influences which tectum and retina exert upon one another during regeneration.     

Visual recovery in goldfish following unilateral optic tectum ablation: Evidence of competition between optic axons for tectal targets

Anatomical studies suggest that regenerating optic axons which invade the ipsilateral lobe of the optic tectum following ablation of the contralateral lobe compete with resident optic axons for synaptic sites on tectal neurons. Invader optic axons are initially uniformly distributed over the entire tectal lobe. With time, the invader and resident optic axons progressively segregate so that the invaders are localized in bands or islands separated by areas that are innervated mainly by the residents. When the resident optic axons are destroyed by ablating the eye opposite to the experimental eye, the invader axons remain continuously distributed and the segregation process apparently does not occur. We investigated the relationship between the segregation process and the recovery of visual function by the invader axons. Visual recovery was measured with a behavioral method in which the index of vision was the occurrence of a branchial suppression response to a moving spot of red light that was classically conditioned to an electric shock stimulus. The minimum time to reappearance of vision following ablation of the contralateral lobe of the tectum in two-eye fish was similar to the reported time of onset of the segregation process. Visual recovery occurred sooner when the opposite eye was removed. The restored vision in both groups disappeared following subsequent ablation of the remaining lobe of the tectum. These results suggest that the goldfish optic tectum normally contains no free synaptic sites for anomalous optic afferents and that the invader axons must compete for targets with the resident optic afferents. The invader axons can apparently remain unconnected or non-functional for several weeks following their arrival in the ipsilateral tectal lobe.     

Axonal transport and transcellular transfer of nucleosides and polyamines in intact and regenerating optic nerves of goldfish: speculation on the axonal regulation of periaxonal cell metabolism

The axonal transport, metabolism, and transcellular transfer of uridine, adenosine, putrescine, and spermidine have been examined in intact and regenerating optic nerves of goldfish. Following intraocular injection of labeled nucleosides, axonal transport was determined by comparing left-right differences in tectal radioactivity, and transcellular transfer was indicated by light autoradiographic analysis. The results demonstrated axonal transport, transcellular transfer, and periaxonal cell utilization of both nucleosides in intact axons and severalfold increases of all of these processes in regenerating axons. Experiments in which the metabolism of the nucleosides was studied resulted in data which suggested that uridine and adenosine, when delivered to the tectum by axonal transport, are protected from degradation and thus are relatively more available for periaxonal cell utilization than nucleosides reaching these cells via the blood. In intact axons, the majority of the nonmetabolized radioactivity was present as UMP, UDP, and UTP following [3H]uridine injections, whereas the majority of the radioactivity following [3H]adenosine injections was present as adenosine, with the phosphorylated derivatives constituting a smaller proportion. During nerve regeneration, the relative proportion of nucleosides to nucleotides was reversed, with uridine being the principal labeled compound in the first case, and AMP, ADP, and ATP being the major labeled compounds in the latter case. The nucleosides also were found to be different from each other in that adenosine, but not uridine, can be taken up by optic axons and transported retrogradely from the tectum to retinal ganglion cell bodies in the eye. Following intraocular injection of [3H]spermidine, radioactivity was transported to the optic tectum and transferred to tectal cells in the vicinity of the regenerating axons. Following [3H]putrescine injections, silver grains were found over periaxonal glia, but preliminary findings suggest that they are not present over tectal neurons nor over radial glial cells in the periependymal layers. Analysis of tectal radioactivity showed in each case that it was composed primarily of the injected compounds. These studies indicate that, following axonal transport, the polyamines do not remain within regenerating axons but are transferred to cells surrounding the axon. On the basis of these and previous findings, we speculate that the axonal transport and transcellular transfer of uridine, adenosine, polyamines, and perhaps other small molecules are means of communication between axons and periaxonal cells; that the axon can affect RNA and protein synthesis in periaxonal cells by regulating the availability of these small molecules; and that, during nerve regeneration, the increased metabolic needs of periaxonal cells are met by an increased axonal supply of precursors (adenosine and uridine) and other molecules (polyamines) critical for protein synthesis.     

In vitro proliferation of radial glia within isolated tectal tissue explanted from adult goldfish

Radial glia located in tectal tissue isolated from adult goldfish retain the ability to incorporate exogenous thymidine into DNA up to 5 days in culture. The rate of their proliferation is maximally enhanced during reinnervation of the tectum by optic fibers about 5 weeks after unilateral optic nerve crush.     

Inhibition of non-neuronal cell proliferation in the goldfish visual pathway affects the regenerative capacity of the retina

Proliferating cells associated with the visual pathway were found in the present study to affect the regenerative capacity of the goldfish retina following optic nerve injury. The contribution of these cells to the process of regeneration was investigated in the goldfish visual system by reducing their proliferation in the optic tract and tecta, using X-irradiation. The regenerative ability of the retina was then evaluated by the following parameters: sprouting from retinal explants, protein synthesis in the retina and accumulation of radiolabeled transported components in the tectum. X-irridiation of the visual system at an early stage of the regeneration process had a promoting effect whereas irradiation at a later stage resulted in a reduced capacity to regenerate. The results are discussed with respect to the possibility that proliferating cells, possibly glia, exert two contradictory contributions: an inhibitory effect at the site of injury, whereas distal to it, a supportive, perhaps trophic effect.     

Axonal pathfinding during the regeneration of the goldfish optic pathway

Retinal ganglion cells in fish and amphibians regenerate their axons after transection of the optic nerve. Fiber tracing studies during the third month of regeneration show that the axons have reestablished a basically normal fiber order in the two brachia of the optic tract; axons originating in the ventral hemiretina are concentrated in the dorsal brachium, axons from the dorsal hemiretina in the ventral brachium. Attardi and Sperry (Exp. Neurol. 7:46-64, 1963) first suggested that the reestablishment of the fiber order reflects path-finding by the regenerating axons. Recently, however, Becker and Cook (Development 101:323-337, 1987) have claimed that the fiber order observed at later stages of regeneration is due to secondary axonal rearrangements and that the initial brachial choice is random. In order to evaluate whether regenerating axons are capable of navigating in the optic tract and brachia and on the tectum, the present study examined the pathway choices and the morphology of regenerating axons en route to their tectal targets in goldfish. Subsets of axons were labeled at various time intervals (2 to 30 days) following an optic nerve crush, by intraretinal application of the lipophilic fluorescent tracer 1,1-dioctadecyl-3-3-3'-3'-tetramethylcarbocyanine (DiI). After a survival time of 18 to 72 hours (to allow for diffusion of DiI along the axons), the experimental animals were perfused with fixative and their right and left optic pathways (nerve, tract, and tectum) were dissected free and separated at the chiasm. Fluorescently labeled axons were traced in whole-mounted pathways. Pathway choices were examined at the brachial bifurcation where axons from ventral and dorsal hemiretinae normally segregate. DiI was found to label axons reliably up to their growth cones, even at the earliest stages of regrowth. The pathway choices of the axons were nonrandom. The majority of the ventral axons reached the appropriate, dorsal hemitectum through the appropriate dorsal brachium of the tract. Dorsal axons reached the ventral hemitectum mainly through the ventral brachium. This suggests the presence of specific guidance cues, accessible to the regenerating axons. Differences in the complexity of the growth cones of the regenerating axons (simple in the nerve and tectal fiber layer, complex in the tract and the synaptic layer of the tectum) provide further evidence for specific interactions between the regenerating axons and their substrates along the pathway. These results argue that regenerating retinal axons in fish are capable of axonal path-finding.     

Spatiotemporal distribution of glia in and around the developing mouse optic tract

In the developing mouse optic tract, retinal ganglion cell (RGC) axon position is organized by topography and laterality (i.e., eye-specific or ipsi- and contralateral segregation). Our lab previously showed that ipsilaterally projecting RGCs are segregated to the lateral aspect of the developing optic tract and found that ipsilateral axons self-fasciculate to a greater extent than contralaterally projecting RGC axons in vitro. However, the full complement of axon-intrinsic and -extrinsic factors mediating eye-specific segregation in the tract remain poorly understood. Glia, which are known to express several guidance cues in the visual system and regulate the navigation of ipsilateral and contralateral RGC axons at the optic chiasm, are natural candidates for contributing to eye-specific pre-target axon organization. Here, we investigate the spatiotemporal expression patterns of both putative astrocytes (Aldh1l1+ cells) and microglia (Iba1+ cells) in the embryonic and neonatal optic tract. We quantified the localization of ipsilateral RGC axons to the lateral two-thirds of the optic tract and analyzed glia position and distribution relative to eye-specific axon organization. While our results indicate that glial segregation patterns do not strictly align with eye-specific RGC axon segregation in the tract, we identify distinct spatiotemporal organization of both Aldh1l1+ cells and microglia in and around the developing optic tract. These findings inform future research into molecular mechanisms of glial involvement in RGC axon growth and organization in the developing retinogeniculate pathway.