Selective stabilization of retinotectal synapses by an activity-dependent mechanism

How does each ingrowing retinal fiber select the right spot in the overall retinotopic projection? Chemospecific surface interactions appear to be sufficient only to organize a crude retinotopic map on the tectum during regeneration of the optic nerve of goldfish. Precise retinotopic ordering is achieved via an activity-dependent stabilization of appropriate synapses, based on the correlated activity of neighboring ganglion cells of the same receptive field type in the retina. Four treatments have been found to block the sharpening process: 1) blocking activity of the ganglion cells with intraocular tetrodotoxin (TTX); 2) rearing in total darkness; 3) correlated activation of all ganglion cells via stroboscopic illumination in a featureless environment; 4) block of retinotectal synaptic transmission with alpha-bungarotoxin. These experiments support a role for normal visually driven activity in sharpening the diffuse projection, and demonstrate that the correlated activity of the optic fibers interacts within the postsynaptic cells, probably through the summation of excitatory postsynaptic potentials. Intraocular TTX experiments suggest that a similar mechanism may drive both the formation of ocular dominance patches in fish tectum and kitten visual cortex and the segregation of different receptive field types in the lateral geniculate nucleus. Thus, it may be a general mechanism whereby the diffuse projections of early development are brought to a mature level of organization.     

Formation of retinotopic connections: Selective stabilization by an activity-dependent mechanism

During regeneration of the optic nerve in goldfish, the ingrowing retinal fibers successfully seek out their correct places in the overall retinotopic projection on the tectum. Chemospecific cell-surface interactions appear to be sufficient to organize only a crude retinotopic map on the tectum during regeneration. Precise retinotopic ordering appears to be achieved via an activity-dependent stabilization of appropriate synapses and is based upon the correlated activity of neighboring ganglion cells of the same receptive-field type in the retina. Four treatments have been found to block the sharpening process: (a) blocking the activity of the ganglion cells with intraocular tetrodotoxin (TTX), (b) rearing in total darkness, (c) correlating the activation of all ganglion cells via stroboscopic illumination and (d) blocking retinotectal synaptic transmission with alpha-bungarotoxin (alphaBTX). These experiments support a role for correlated visually driven activity in sharpening the diffuse projection and suggest that this correlated activity interacts within the postsynaptic cells, probably through the summation of excitatory postsynaptic potentials (EPSPs). Other experiments support the concept that effective synapses are stabilized: a local postsynaptic block of transmission causes a local disruption in the retinotectal map. The changes that occur during this disruption suggest that each arbor can move to maximize its synaptic efficacy. In development, initial retinotectal projections are often diffuse and may undergo a similar activity-dependent sharpening. Indirect retinotectal maps, as well as auditory maps, appear to be brought into register with the direct retinotopic projections by promoting the convergence of contacts with correlated activity. A similar mechanism may drive both the formation of ocular dominance patches in fish tectum and kitten visual cortex and the segregation of different receptive-field types in the lateral geniculate nucleus. Activity-dependent synaptic stabilization may therefore be a general mechanism whereby the diffuse projections of early development are brought to the precise, mature level of organization.     

金鱼再生的视神经在顶盖投射精确化的DiI顺行标记研究

本文用微量显微注射法,在金鱼视网膜的背侧用亲脂类荧光染料DiI标记少量神经节细胞,通过顺行标记研究了视神经再生过程中视网膜顶盖投射的精确化过程。在损伤视神经后的不同时期观察了再生视神经纤维在顶盖整装片上的分布。在再生早期它们以超出正常的途径由背腹两侧进入顶盖,广泛分布。但其中大部分仍分布于顶盖腹侧的靶区。在再生晚期通过精确化,重建如正常鱼一样精确的视网膜顶盖投射。这个精确化过程表现在以下三方面:(1)再生于顶盖错误区域的再生视神经纤维的消失;(2)再生早期视神经纤维主干上生长的侧部分支的消失;(3)到达靶区的再生视神经纤维形成重迭的终末分支。由以上结果推测,顶盖中可能存在两类不同的因子:一类是普通诱向因子,存在于整个顶盖中,它在再生早期引导再生的视神经纤维长入顶盖。另一类是神经营养因子,它具区域特异性,在再生晚期引导视神经纤维到达顶盖靶区,形成精确的视网膜顶盖投射。     

Up-regulation of protein kinase C in regenerating optic nerve fibers of goldfish: Immunohistochemistry and kinase activity assay

Protein kinase C (PKC) activation has been associated with synaptic plasticity in many projections, and manipulating PKC in the retinotectal projection strongly affects the activity-driven sharpening of the retinotopic map. This study examined levels of PKC in the regenerating retinotectal projection via immunostaining and assay of activity. A polyclonal antibody to the conserved C2 (Ca²⁺ binding) domain of classical PKC isozymes (anti-panPKC) recognized a single band at 79–80 kD on Western blots of goldfish brain. It stained one class of retinal bipolar cells and the ganglion cells in normal retina, as shown previously. Strong staining was not present in the optic fiber layer of retina or in optic nerve, optic tract, or terminal zone in tectum, with the exception of a single fascicle of optic nerve fibers that by their location and by L1 (E587) staining were identified as those arising from newly added ganglion cells at the retinal margin. Normal tectal sections showed dark staining of a subclass of type XIV neuron with somas at the top of the periventricular layer and an apical dendrite ascending to stratum opticum. In regenerating retina, swollen ganglion cells stained darkly and stained axons were seen in the optic fiber layer. In regenerating optic nerve (2–11 weeks postcrush), all fascicles of optic fibers stained darkly for both PKC and L1(E587). At 5 weeks postcrush, PKC staining could also be seen in the medial and lateral optic tracts and stratum opticum at the front half of the tectum and very lightly over the terminal zones. PKC activity was measured in homogenized tissues dissected from a series of fish with unilateral nerve crush from 1 to 5 weeks previously. Activity levels stimulated by phorbols and Ca²⁺ were measured by phosphorylation of a specific peptide and referred to levels measured in the opposite control side. Regeneration did not increase overall PKC activity in retina or tectum, but in optic nerve there was an 80% rise after the first week. The increased activity verifies that the increased staining in nerve represented an up-regulation of functional PKC during nerve regeneration. © 1998 John Wiley & Sons, Inc. J Neurobiol 36: 315–324, 1998     

Activity sharpens the regenerating retinotectal projection in goldfish: sensitive period for strobe illumination and lack of effect on synaptogenesis and on ganglion cell receptive field properties

The regenerating optic nerve of goldfish first reestablishes a rough retinotopic map on the contralateral tectum and then sharpens it. Disruption of visual activity, either by blocking activity with intraocular tetrodotoxin (TTX; Schmidt and Edwards, 1983) or by synchronizing activity with strobe illumination (Schmidt and Eisele, 1985), disrupts the sharpening process: the map is correctly oriented but the multiunit receptive fields at each point average 25-40 degrees in diameter. In order to test whether strobe and TTX interfere with the same mechanism, we have tested whether their sensitive periods are the same, and whether strobe, like TTX treatment, does not affect either ganglion cell receptive field properties or synaptogenesis. In parallel studies, we exposed fish to 2 weeks of either strobe illumination or intraocular TTX beginning at various times after crush and determined via electrophysiological recordings that the periods of sensitivity were nearly identical. There was no effect of either treatment during the first 2 weeks (before the fibers arrive at the tectum), maximal disruption of sharpening between 14 and 50 days (the period of rapid synaptogenesis), decreasing disruption between 50 and 125 days, and no effect beyond that point or in the normal projection. In addition, long strobe exposures of up to 142 days produced no greater disruptions than shorter 2-3-week exposures, indicating no cumulative effect. The reestablishment of synaptic transmission in tectum, assayed by recording field potentials elicited by optic nerve shock, was not affected by stroboscopic illumination. Finally, individual ganglion cells, recorded intraretinally following long-term strobe exposure, had receptive fields that were normal both in size and in their characteristic responses to light-on, to light-off, or to both on and off. These findings support the hypothesis that strobe-like TTX prevents retinotopic refinement by preventing the correction of errors initially made by the ingrowing optic axons (Schmidt et al., 1988).     

Competitive and positional cues in the patterning of nerve connections

The visual system of lower vertebrates has served as an important testing ground for the mechanisms that generate topographic neuronal connections. During both the outgrowth and the regeneration of the optic nerve, a smoothly ordered map of the retina is formed on its major target, the optic tectum (the retinotectal projection). Experiments performed on this projection have offered support for a variety of mechanisms, including the matching of positional cues in the retina and tectum, the guidance of nerve fibers by interactions between fibers, competition for synaptic space, and the refinement of connections based on neuronal activity. Unfortunately, individual experiments that support any one of these mechanisms have been taken at times as evidence against the involvement of any other mechanism; for example, experiments demonstrating the importance of positional cues have been thought mistakenly to indicate that activity-based interactions are unimportant. Computer simulations, in which multiple, somewhat opposed, mechanisms are allowed to operate in concert demonstrate that such a hybrid model is able to generate a full range of experimental results. More importantly, the elimination of any one of the mechanisms renders the model unable to fit entire classes of findings. Thus, the patterning of the retinotectal projection is best viewed as a process in which the optic nerve terminals attempt to satisfy multiple constraints in selecting their target sites.     

Ipsilateral retinal projections into the tectum during regeneration of the optic nerve in the cichlid fish Haplochromis burtoni: a Dil study in fixed tissue.

Retinal projections were experimentally manipulated in a bony fish to reveal conditions under which considerably enlarged ipsilateral projections developed and persisted. Three experimental groups were studied: animals after unilateral enucleation, after unilateral nerve crush, and after enucleation and crush of the remaining optic nerve. At 29 days after unilateral enucleation alone, no enhanced ipsilateral projection had developed. After nerve crush, however, large numbers of retinal fibers regenerated into the ipsilateral tectum. Retrogradely filled, ipsilaterally projecting ganglion cells were distributed throughout the entire retina. After 15 days regenerating retinal fibers covered the entire ipsilateral tectum. At later stages the ipsilateral projection showed progressive reduction in coverage of the tectum. Combining enucleation with nerve crush led to an ipsilateral projection that covered the tectum at 28 days and later. In this experimental situation the development of an ipsilateral projection appears to be a two-step process: (1) Fibers are rerouted to the ipsilateral side at the diencephalon, and (2) ipsilateral fibers persist in the tectum only in the absence of a contralateral projection while they appear to be eliminated in the other cases.     

Ipsilateral retinal projections into the tectum during regeneration of the optic nerve in the cichlid fish Haplochromis burtoni: A dil study in fixed tissue

Retinal projections were experimentally manipulated in a bony fish to reveal conditions under which considerably enlarged ipsilateral projections developed and persisted. Three experimental groups were studied: animals after unilateral enucleation, after unilateral nerve crush, and after enucleation and crush of the remaining optic nerve. At 29 days after unilateral enucleation alone, no enhanced ipsilateral projection had developed. After nerve crush, however, large numbers of retinal fibers regenerated into the ipsilateral tectum. Retrogradely filled, ipsilaterally projecting ganglion cells were distributed throughout the entire retina. After 15 days regenerating retinal fibers covered the entire ipsilateral tectum. At later stages the ipsilateral projection showed progressive reduction in coverage of the tectum. Combining enucleation with nerve crush led to an ipsilateral projection that covered the tectum at 28 days and later. In this experimental situation the development of an ipsilateral projection appears to be a two-step process: (1) Fibers are rerouted to the ipsilateral side at the diencephalon, and (2) ipsilateral fibers persist in the tectum only in the absence of a contralateral projection while they appear to be eliminated in the other cases. © 1992 John Wiley & Sons, Inc.     

Persistence of Disorganized Pathways and Tortuous Trajectories of Regenerating Retinal Fibers in the Adult Newt Cynops pyrrhogaster

The anatomical pathways and trajectories of regenerating retinal fibers within the optic tract and tectum of adult newts were examined, 7 months after transection of the optic nerve. In spite of restoration of retinotopic ordered central connection within the tectum, the pathways of individual regenerating retinal fibers within the optic tract were greatly disorganized; the misrouted retinal fibers exhibited tortuous trajectories on the tectal surface in approach to their sites of normal innervation. These results suggest that the regenerating retinal fibers with abnormal pathways and tortuous trajectories can be maintained provided they are in contact with appropriate targets.     

Evidence for a driving role of ingrowing axons for the shifting of older retinal terminals in the tectum of fish.

In amphibians and teleosts, retina and tectum grow incongruently. In order to maintain the retinotopy of the retinotectal projection, Gaze, Keating, and Chung (1974) postulated a shifting of terminals throughout growth. In order to test the possibility that ingrowing retinal fibers are the driving force for this shifting, we induced a permanent retinal projection into the ipsilateral tectum in juveniles of the cichlid fish Haplochromis burtoni. The surface of the tectum had increased (11-18 months later) 2.5-5.8 times, and the surface of the retina 8.6-14 times. Filling of ganglion cells with horseradish peroxidase (HRP) retrogradely from the tectum showed ipsilaterally regenerating ganglion cells only in the center of the retina. The position of ganglion cells indicated that the ipsilateral projection derived only from axotomized and regenerating retinal ganglion cells but not from those newly born. Ipsilaterally projecting retinal fibers showed terminals only in the rostral half of the tectum. Comparison of area of terminations of ipsilaterally projecting ganglion cells at various times after the crush provided no evidence for expansion or a shift into caudal tectal areas throughout the period of growth. These findings are compatible with the idea that newly ingrowing fibers induce older terminals to move caudally.     

Alterations in the Xenopus retinotectal projection by antibodies to Xenopus N-CAM

The patterned neural projection from the eye to the optic tectum of lower vertebrates (the retinotectal projection) has been proposed to be ordered by interactions between the optic nerve fibers and their surrounding tissues. To investigate the role of one such defined cell interaction, agarose implants containing antibodies to the neural cell adhesion molecule, N-CAM, were inserted into the tectum of the African clawed frog, Xenopus laevis. Both monoclonal and polyclonal antibodies against N-CAM reversibly and specifically distorted the pattern of the retinotectal projection, decreasing the precision of the projection as determined by electrophysiological techniques as well as decreasing the density of retinal innervation of the tectum and the branching of single axons as determined by horseradish peroxidase tracing. The anatomical effects became maximal at 4 to 6 days after implantation and returned to undetectable levels by 2 weeks, whereas the physiological effects became maximal by 8 to 10 days and a normal physiological map was reestablished within 4 weeks. The results are consistent with the hypothesis that anti-N-CAM antibodies perturb the ongoing growth and retraction of the terminal arbors of the optic nerve fibers, such that a region of the tectum becomes largely denuded of fibers. The physiological defects may then be a consequence both of the initial retraction of optic nerve terminals and of the rapid ingrowth of the perturbed and neighboring optic nerve fibers into the denuded region after the antibodies were cleared from the tectum. These results support the concept of a major role for N-CAM-mediated adhesion during map regeneration and maintenance.     

Dynamic aspects of retinotectal map formation revealed by a vital-dye fiber-tracing technique

In the visual system of Xenopus laevis, the axons from the retinal ganglion cells of the eye form a topographic projection onto the optic tectum. Many studies have focused on revealing the mechanisms responsible for this precise and regular projection pattern. In contrast to the static view of the system that one might expect from examining the regularity of the projection, recent work on its regeneration and its changes during larval development indicate that part of the patterning process involves the dynamic behavior of optic fibers. Typically, anatomical and electrophysiological techniques have been used to obtain static views of the developing retinotectal projection which then must be complied to provide a glimpse of any dynamic behavior. Here we report on experiments using a newly developed fiber tracing technique to directly follow the emergence of topography in the developing retinotectal projection. Defined halves of the developing eyebud were labeled with a vital fluorescent dye which fills the growing axons, and the projection of the labeled cells was followed for up to 2 weeks in individual animals. The experiments confirm that dorsal and ventral optic nerve fibers sort out into an ordered projection early in development. In contrast, nasal and temporal fibers initially overlap, and the same sets of prelabeled fibers then sort out into the adult topography over a period of days.     

Evidence for a driving role of ingrowing axons for the shifting of older retinal terminals in the tectum of fish

In amphibians and teleosts, retina and tectum grow incongruently. In order to maintain the retinotopy of the retinotectal projection, Gaze, Keating, and Chung (1974) postulated a shifting of terminals throughout growth. In order to test the possibility that ingrowing retinal fibers are the driving force for this shifting, we induced a permanent retinal projection into the ipsilateral tectum in juveniles of the cichlid fish Haplochromis burtoni. The surface of the tectum had increased (11–18 months later) 2.5–5.8 times, and the surface of the retina 8.6–14 times. Filling of ganglion cells with horseradish peroxidase (HRP) retrogradely from the tectum showed ipsilaterally regenerating ganglion cells only in the center of the retina. The position of ganglion cells indicated that the ipsilateral projection derived only from axotomized and regenerating retinal ganglion cells but not from those newly born. Ipsilaterally projecting retinal fibers showed terminals only in the rostral half of the tectum. Comparison of area of terminations of ipsilaterally projecting ganglion cells at various times after the crush provided no evidence for expansion or a shift into caudal tectal areas throughout the period of growth. These findings are compatible with the idea that newly ingrowing fibers induce older terminals to move caudally.     

Rapid activity-dependent sprouting of optic fibers into a local area denervated by application of beta-bungarotoxin in goldfish tectum

The retinotectal projection is known to be capable of extensive long-term expansion of connections, but it is not known how fast such changes can occur or what triggers sprouting of terminals. We studied sprouting of optic fibers into an area denervated by local microinjection of beta-bungarotoxin (β-BTX), a specific presynaptic neurotoxin with phospholipase A₂ activity that destroys nerve terminals at the neuromuscular junction. After injection of 0.1 pmol of β-BTX, the optic terminals fired spontaneously with decreasing amplitude and became silent within 1 to 2 h. Outside the injection zone, the retinotectal map was normal, so the silent zone was associated with a scotoma in the visual field. Horseradish peroxidase (HRP) staining of the entire optic nerve showed a denervated region at the injection site with beaded, degenerating fibers at its edge. Between 3 and 9 days later, optic units were recorded within the injection zone whose receptive fields lay just outside the scotoma in the visual field, indicating that intact surrounding terminals had sprouted into the area. These sprouts made functional connections, as indicated by field potential recordings and current source-density analysis. At this time, HRP staining also demonstrated retinal innervation within the injection zone. By 12 days, normal maps with no scotoma were recorded and HRP staining was normal at the injection site, indicating that the β-BTX-damaged fibers had regenerated to reclaim their tectal sites. The results show that the retinotectal projection of goldfish is very dynamic, since intact optic fibers can sprout into adjacent vacant postsynaptic territory within 2 to 3 days, much faster than previously reported. In a final experiment, we showed that this sprouting is activity-dependent, since it could be prevented by blocking retinal activity with intraocular tetrodotoxin (TTX) during the first 2 days postinjection, even though TTX block of activity does not block regeneration in this system. One possible mechanism for this rapidly triggered sprouting is that arachidonic acid liberated by β-BTX acts as a sprouting factor to attract surrounding healthy fibers into the denervated region but requires activity at the terminals to be effective. © 1996 John Wiley & Sons, Inc.     

Competitive interactions between retinal ganglion axons for tectal targets explain plasticity of retinotectal projection in the servomechanism model of retinotectal mapping

The mechanism of topographic mapping of retinal ganglion cells to the midbrain was previously elucidated by the servomechanism model, which is based on the fact that cells expressing Eph-receptors respond specifically to surface expressing membrane-bound ephrin-ligands at a critical level. The retina has increased nasal-to-temporal gradient of Eph receptor-density, and the optic tectum/superior colliculus has increased rostral-to-caudal gradient of membrane-bound ephrin-ligand. An axon from the retina has an identification tag of a certain level of Eph-receptor density depending on its retinal position, and adheres to the site on the tectum/superior colliculus expressing ephrin-ligands at a critical ligand-density level. The servomechanism model rigidly defines positions of axon terminals on the midbrain. However, optic nerve regeneration experiments combined with halved retina or tectum show a plastic or flexible mapping (expansion, compression and transposition of tectal projections). To reconcile the discrepancy between the rigid model and the plastic behavior, competition between retinal axon terminals for a target site was introduced to the servomechanism. The servomechanism/competition model succeeded in computer simulations of the plastic mapping of retinal axons on the tectum. Recent experiments of upregulated ligand-density on the tectum during nerve regeneration and the role of axonal competition are discussed.     

Spontaneous bursting and long-lived local correlation in normal and denervated tectum of goldfish

The formation of fine retinotopic order by growing optic fibers in the goldfish is thought to be mediated by the correlated firing of optic fibers from neighboring retinal ganglion cells. Although the activity of the tectal cells must also be important for this activity-dependent refinement, few studies have analyzed the pattern and local correlation of the intrinsic activity of tectal neurons and the effect of denervation on this activity. To address this issue, spontaneous (nonoptic driven) activity was analyzed and cross-correlograms were computed between individual tectal neurons using single and double electrode extracellular recordings. Recordings were made in normally innervated tectum in which the contribution of optic activity was eliminated by short-term intraocular blockade with tetrodotoxin and in denervated tecta in which the optic nerve had been severed several weeks prior. Several observations were relevant to activitydependent refinement: First, coupling between neighboring tectal cells is weak. Second, the time duration for local correlation is relatively long, as long as 200 ms. Third, tectal neurons exhibit spontaneous bursting. Fourth, denervation increased the level of spontaneous activity in the tectum. The increased spontaneous activity and bursting following denervation implies that tectal neurons are more excitable when optic fibers are beginning to reinnervate the tectum. This could make it possible for optic fibers to drive tectal neurons at a time when their input to individual neurons is severely weakened by a lack of spatial convergence. The weak coupling between tectal cells and the consequent long-time constant for correlated activity implies a constraint on the duration of correlated retinal activity that is used for activitydependent refinement. Since optic fibers likely need to detect the postsynaptic activity of a local group of tectal neurons, rather than that of a single neuron, the long tectal time constant means that retinal activity need not be correlated with precision much better than 200 ms because the postsynaptic circuitry cannot generate shorter correlations. © 1995 John Wiley & Sons, Inc.     

The development of ipsilateral retinal projections into the tectum in the cichlid fish Haplochromis burtoni: a Dil study in fixed tissue.

The normal development of the retinal projection was studied in a bony fish with Dil. Between 5.5 and 10 days postfertilization the contralateral retinal projection grows from the rostral pole of the tectum across its center. A maximum of 15 retinal fibers reaches the ipsilateral tectum. In 33-day-old juvenile animals, less than 15 ipsilateral fibers terminate in the entire tectum. Ipsilaterally projecting ganglion cells (maximal number = 20 cells) are scattered throughout the entire retina, and the location of ganglion cells in the retina and axonal terminations in the tectum display a large interindividual variability. This suggests that the small adult contingent of ipsilateral fibers in this bony fish develops without an initial exuberant ipsilateral retinal projection that is later pruned back.     

The development of ipsilateral retinal projections into the tectum in the cichlid fish Haplochromis burtoni: A dil study in fixed tissue

The normal development of the retinal projection was studied in a bony fish with Dil. Between 5.5 and 10 days postfertilization the contralateral retinal projection grows from the rostral pole of the tectum across its center. A maximum of 15 retinal fibers reaches the ipsilateral tectum. In 33-day-old juvenile animals, less than 15 ipsilateral fibers terminate in the entire tectum. Ipsilaterally projecting ganglion cells (maximal number = 20 cells) are scattered throughout the entire retina, and the location of ganglion cells in the retina and axonal terminations in the tectum display a large interindividual variability. This suggests that the small adult contingent of ipsilateral fibers in this bony fish develops without an initial exuberant ipsilateral retinal projection that is later pruned back. © 1992 John Wiley & Sons, Inc.     

Goldfish retinal axons respond to position-specific properties of tectal cell membranes in vitro

Using a special in vitro assay, we tested whether retinal ganglion cell axons in an adult vertebrate, the goldfish (which can regenerate a retinotopic projection after optic nerve section), recognize position-specific differences in cell surface membranes of their target, the tectum opticum. On a surface consisting of alternating stripes of membranes from rostral and caudal tectum, temporal axons accumulate on membranes derived from their retinotopically related rostral tectal half. Nasal axons grow randomly over both types of membranes. Nasal and temporal axons can elongate on both rostral and caudal membranes. A quantitative growth test, however, revealed that caudal membranes are less permissive substrates for the outgrowth of temporal axons than rostral membranes, and than rostral or caudal membranes for nasal axons.     

Retinotectal plasticity in Xenopus: Anomalous ipsilateral projection following late larval eye removal

The left eye was removed from Stage 56 Xenopus tadpoles. Two to 9 months after metamorphosis, electrophysiologic analysis showed that the surviving (right) eye mediated a normal visual field projection to the left (contralateral) optic tectum. In addition, a peripheral region of the same retina innervated the entire right (ipsilateral) tectum. Primary evidence that indicates this anomalous ipsilateral projection was due to direct retina-to-tectum innervation comes from singleunit analysis, latency measurements, and tectal lesion studies. Thus, the peripheral retina simultaneously connected in much different patterns to the two optic tecta, solely on the basis of the presence (in the left tectum) or absence (in the right tectum) of central retinal fibers. This documents a role for fiber-fiber interaction (such as repulsion or competition) acting in combination with fiber-tectum interactions in the formation of the retinotectal map.