Density of optic terminals in half tecta of goldfish with compressed retinotectal projections

If the caudal half of the tectum in goldfish is removed, the projection from the entire retina is compressed onto the remaining half tectum. The percentage increase in density of optic terminals and synaptic contacts in such half tecta compared to that in tecta with normal projections ranged from 20–112% and 19–109% respectively. Both measures had similar percentage increases in each individual fish, with the proportion of terminals with more than one synaptic contact remaining constant.Tectal sites available for optic termination are flexible so that a higher density is allowed in response to increased optic innervation.     

Ultrastructure of reorganising visual projections in half tecta of carp kept in constant light

In carp with the caudal half of the tectum cut off the projection of the whole contralateral visual field, mapped electrophysiologically, becomes compressed on to the remaining half tectum during the succeeding months. Ultrastructural study of tecta in various stages of reorganisation shows initially a profuse and widespread sprouting of unmyelinated axons in the optic nerve and external grey layers. This occurs before any compression of the projection is detectable electrophysiologically. After physiological compression is completed the bundles of unmyelinated fibres are greatly reduced and many myelinated axons are present. We conclude that compression initially involves the widespread growth of axon sprouts with the retention and myelination of a selected few when new connections are formed. Contrary to previous reports, exposure to constant light did not always delay or prevent the reorganisation of the visual projection. The widespread occurrence of optic terminal degeneration in both operated and normal tecta of fish kept in constant light suggests that light-induced retinal degeneration may have over-ridden the effect of constant light by providing vacant synaptic sites in the tectum for growing axonal sprouts.     

The transneuronal transport of horseradish peroxidase in the visual system of the frog, Rana pipiens

During the course of experiments designed to study synaptic relationships between the terminals of retinal axons and the various cell populations in the optic tectum of the frog, Rana pipiens, we found that neurons in many of the retinorecipient nuclei, including the tectum, are labeled transneuronally following injections of horseradish peroxidase into the optic nerve. In the optic tectum, particular cell groups are labeled to the extent that their dendrites as well as their somas are filled with reaction product while other cell types, which, on the basis of the location of their somas or dendrites, seem equally likely to receive direct retinal projections, remain free of label. Electron microscopic investigation of the optic tectum reveals that the label is confined to pre- and postsynaptic processes. These results suggest that the transneuronal transport depends on a transfer of horseradish peroxidase from presynaptic terminals to postsynaptic cells rather than on a widespread diffusion of the enzyme through the neuropil followed by a selective uptake by particular cell groups. These results also suggest that only some of the tectal cell groups which receive direct retinal projections may be transneuronally labeled. The transneuronal transport of horseradish peroxidase is useful since it reveals the morphology as well as the location of at least some of the retinorecipient cells. Moreover, the robust nature of this phenomenon makes the frog a good choice for future studies of the mechanism of transneuronal transport.     

Schwann cells in the regenerating fish optic nerve: evidence that CNS axons, not the glia, determine when myelin formation begins

Fish optic nerve fibres quickly regenerate after injury, but the onset of remyelination is delayed until they reach the brain. This recapitulates the timetable of CNS myelinogenesis during development in vertebrate animals generally, and we have used the regenerating fish optic nerve to obtain evidence that it is the axons, not the myelinating glial cells, that determine when myelin formation begins. In fish, the site of an optic nerve injury becomes remyelinated by ectopic Schwann cells of unknown origin. We allowed these cells to become established and then used them as reporters to indicate the time course of pro-myelin signalling during a further round of axonal outgrowth following a second upstream lesion. Unlike in the mammalian PNS, the ectopic Schwann cells failed to respond to axotomy and to the initial outgrowth of new optic axons. They only began to divide after the axons had reached the brain. Shortly afterwards, small numbers of Schwann cells began to leave the dividing pool and form myelin sheaths. More followed gradually, so that by 3 months remyelination was almost completed and few dividing cells were left. Moreover, remyelination occurred synchronously throughout the optic nerve, with the same time course in the pre-existing Schwann cells, the new ones that colonised the second injury, and the CNS oligodendrocytes elsewhere. The optic axons are the only common structures that could synchronise myelin formation in these disparate glial populations. The responses of the ectopic Schwann cells suggest that they are controlled by the regenerating optic axons in two consecutive steps. First, they begin to proliferate when the growing axons reach the brain. Second, they leave the cell cycle to differentiate individually at widely different times during the ensuing 2 months, during the critical period when the initial rough pattern of axon terminals in the optic tectum becomes refined into an accurate map. We suggest that each axon signals individually for myelin ensheathment once it completes this process.     

Ganglion Cells with Sustained Activity in the Fish Retina and Their Possible Function in Evaluation of Visual Scenes

Neuroscience and Behavioral Physiology - Background extracellular spike activity of single ganglion cells was recorded from axon terminals in the optic tectum of living immobilized fish. The sizes...     

Progress of topographic regulation of the visual projection in the halved optic tectum of adult goldfish.

1. The patterns of re-established visual projections on to the rostral half-tectum are studied following excision of the caudal tectum at various intervals after section of either the contralateral optic nerve or the ipsilateral optic tract in adult goldfish. 2. The pattern of a newly restored retinotectal projection depends on the duration of the post-operative period given to the halved tectum before it is re-innervated by regenrating optic fibres from the retina. 3. When the duration is such that regenerating optic fibres invade the denervated rostral half-tectum at about 40 days or longer after excision of the caudal tectum, the remaining half-tectum is able to accommodate incoming optic fibres not only from the appropriate temporal hemi-retina but also from the foreign nasal hemiretina in an orderly compressed topographic pattern. 4. If the surgical operations are timed so that the halved tectum receive regenerating optic fibres earlier than 33 days after excision of the caudal tectum, the halved tectum initially accommodates only those optic fibres originating from the temporal half of the retina at this early stage. 5. This normal (uncompressed) pattern of the newly regenerated visual projection, however, eventually changes into an orderly compressed pattern at a later period. Post-operative dark-deprivation of the operated fish has no significant effect on the temporal transition. 6. The temporal transition from an initially normal pattern into an orderly compressed pattern may reflect the time course of progressive and systematic changes involved in topographic regulation of the halved tectum into a whole.     

Effects of post-operative visual environments on reorganization of retinotectal projection in goldfish.

1. Possible influence of different visual environments on the reorganization of retinotectal projection was studied with neurophysiological mapping methods following excision of the caudal half of the optic tectum in adult goldfish. 2. Post-operative light-deprivation showed no significant effects: in the absence of visual input, the visual projection from the whole retina because compressed on to the remaining rostral half-tectum in correct retinotopic order within 4 months, regardless of whether the contralateral optic nerve was left intact, or severed and then allowed to regenerate. 3. When the operated goldfish were continually exposed to visual stimuli without any dark period (post-operative dark-deprivation), two different results were observed: if the optic nerve was sectioned, in addition to excision of the caudal tectum, an orderly field compression was observed within 70 days in the re-established retinotectal projection; on the other hand, if the optic nerve was left intact, the dark-deprived fish retained the original connexions between the remaining rostral half-tectum and the temporal hemiretina without showing any sign of field compression for up to 253 days. 4. When the dark-deprived fish was then transferred into darkness, the suppressive effect disappeared: a compression of the retinotectal projection was induced within 2 or 3 weeks after the transfer. 5. Histological preparations of the fish brains showed consistent morphologic changes in the laminar structure of the remaining half-tectum. The stratum opticum and the stratum fibrosum et griseum superficiale merged together to form a new layer which contained an intricate network of thick fibre bundles.     

Effect of AF64A on the cholinergic systems of the retina and optic tectum of goldfish

Summary AF64A, a presumed selective cholinergic neurotoxin has been used to study the effect on cholinergic systems of the goldfish retina and optic tectum. Toxin injection in the vitreum and in the optic tectum caused a selective decrease of choline acetyltransferase activity in both areas, while no significant decrease of glutamate decarboxylase and D-3H aspartate uptake were observed at different times after the injections. The effect was particularly dramatic in the retina of long term-injected animals, where choline acetyltransferase dropped to practically zero level. The ultrastructural analysis showed selective degeneration of some neurons in the amacrine and ganglion cell layer of the retina as well as of synaptic terminals and neuronal cell bodies in the optic tectum. The results favour a selective cholinotoxicity of AF64A in fish nerve tissue at doses substantially higher than those found to have additional unselective effects in mammals.Supported by a grant from the Italian Ministry of Education     

Retinotectal reorganization in goldfish--II. Effects of partial tectal ablation and constant light on the retina.

In a proportion of small fish, after removal of half the tectum, exposure to constant light induces compression of the retinotectal projection. In contrast, it prevents compression in large fish. Loss of rod photoreceptors occurs in both small and large fish kept in constant light. This was not sufficient to induce compression. In large fish removal of half the tectum followed by exposure either to light/dark or to constant light has no consistent effect on the number of retinal ganglion cells. In small fish the removal of half the tectum alone causes a loss of ganglion cells in the eye projecting to the operated tectum. This in itself was not associated with compression. In half the small fish, exposure to constant light following the removal of half the tectum produced an even greater loss of ganglion cells in the eye projecting to the operated tectum. This increased loss of ganglion cells may be sufficient to induce compression, probably by creating vacant synaptic sites in the tectum. That only a proportion of fish show increased loss of cells correlates with the finding that only a proportion undergo compression in constant light.     

A pattern of optic axons in the normal goldfish tectum consistent with the caudal migration of optic terminals during development

Optic axons were cut in the goldfish optic nerve or tectum, filled with horseradish peroxidase and traced in tectal wholemounts. Many of them ran in conspicuous fascicles which curved across the tectum. Axons from central nasal retina, which ran in the most rostral fascicles, turned abruptly as they left these fascicles; ran caudally in a diffuse, parallel array for up to half the tectal length; and passed beneath more caudal fascicles to innervate the caudal half-tectum. Axons from peripheral nasal retina ran in the most caudal fascicles and terminated near their turning-points. Axons from temporal retina entered the tectum at its rostral margin and ran caudally from their points of entry to innervate the rostral half-tectum. The resultant pattern was entirely consistent with the proposal that a slow caudal migration of optic terminals compensates during normal development for disparate modes of retinal and tectal growth.     

Quantitative analysis of optic terminal profile distribution within the pigeon optic tectum

Two regions of the pigeon optic tectum, caudoventral and rostrodorsal, were examined electron microscopically in an attempt to clarify quantitatively previous findings on the distribution of optic afferent terminal profiles within the retino-receptive zone. Major terminations of optic axons were found in sublaminae IIb, c and d of both the caudoventral and the rostrodorsal tectum. Multiple peaks of such terminal concentrations occur in sublayers a, b and c: in the rostrodorsal field they lie largely centrally within sublayers b and c, whereas in the caudoventral tectum they lie at the a/b, b/c and c/d boundaries. The distribution to sublayer IId tends to be maximal within its superficial and deep extremities, especially in the caudoventral tectum. Sublayer f contains a more noticeable concentration of optic terminals in the caudoventral than in the rostrodorsal tectum. The retinal input is distributed more nearly equally between sublayers a, b and c combined and sublayer d in the rostrodorsal tectum than in the caudoventral field, where sublayer d is the major target. Other differences exist between the two regions: the caudoventral tectum shows a higher density of terminal profiles/unit area than the rostrodorsal tectum, and, in the deeper parts of the retino-receptive zone, a higher density per unit volume; in general the profiles in deeper sublayers are also larger than those in the rostrodorsal tectum. Numbers of synaptic thickenings (as profiles) per terminal profile show less clear differences between tectal regions. Interlaminar differences in the synaptic thickening/terminal profile ratios are more obvious: in sublamina IIb especially, terminal profiles with multiple synaptic thickenings are more common than elsewhere.The results are discussed in relation to previous morphological and electrophysiological studies and an attempt is made to resolve some of the discrepancies in the previous studies.     

Distribution of GABA immunoreactivity in the optic tectum of the frog: a light and electron microscopic study

GABA immunoreactivity was studied in the optic tectum of the frog, Rana esculenta, by postembedding immunohistochemical methods at the light and electron microscopic levels. Nearly one-third of the total population of tectal cells appeared to be GABA-immunoreactive. The proportion of stained neurons was highest in layer 9 (61%), and they occurred less frequently in layers 7 (21%) and 6 (27%). Stained perikarya represented a population of small neurons with a diameter of 8-10 microns. Large cell bodies in layer 7 or at the top of layer 6, and cells of origin of the mesencephalic trigeminal tract in layer 2, were devoid of labelling. Axon terminals and dendrites displaying immunoreactivity for GABA were observed in all of the plexiform layers. On the basis of ultrastructural characteristics two types of GABA-positive axon terminals and two variations of GABA-immunoreactive dendrites were distinguished. Synaptic relations of GABA-immunoreactive and GABA-negative axons as well as dendrites were also studied. Besides a wide variety of axodendritic synapses, dendrodendritic synaptic appositions were also revealed. The results suggest that various inhibitory mechanisms are involved in tectal circuits, which have to be incorporated into future neuronal models concerning visual information processing in the optic tectum of the frog.     

Neuronal nitric oxide synthase in the olfactory system,forebrain, pituitary and retina of the adult teleost Clarias batrachus

Immunocytochemical application of antibodies against nNOS to the brain sections of Clarias batrachus revealed intense immunoreactivity in several olfactory receptor neurons (ORNs), in their axons over the olfactory nerve, and terminals in the olfactory glomeruli. Several basal cells in the olfactory epithelium showed NOS immunoreactivity. Application of post-embedding immunoelectron microscopy showed nNOS labeled gold particles in apical cilia, dendrites and soma of the ORNs and also in the axon terminals in the glomeruli of the olfactory bulb. nNOS containing fibers were also encountered in the medial olfactory tracts (MOTs). Bilateral ablation of the olfactory organ resulted in total loss of nNOS immunoreactivity in the fascicles of the olfactory nerve layer and also in the MOT. nNOS immunoreactivity was seen in several cells of the nucleus preopticus (NPO) and their axons that innervate the pituitary gland. Some cells in the floor of the tuberal area were stained positive with nNOS antibodies. nNOS immunolabeled cells were seen in all the three components of the pituitary gland with light as well as post-embedding immunoelectron microscopy. While several nNOS immunoreactive fibers were seen in rostral pars distalis, a much limited fiber population was seen in the proximal pars distalis. In addition, conspicuous immunoreactivity was noticed in some ganglion cells in the retina and in some fibers of the optic nerve traceable to the optic tectum. The NO containing system in this fish appears to be similar to that in other fishes.     

Transport of RNA along the optic pathway of the chick: An autoradiographic study

Summary Autoradiograms of the optic tectum were made 8 days after monocular injection of ³H-uridine in 3 day-old chicks. Electron microscopic analysis of grain density over morphological structures in various layers of the optic tectum contralateral to the injected eye, was carried out. This revealed that the highest concentration of labeled RNA was within the axons of the more superficial layers of the tectum. A lesser density of grains was observed over glial cells, neuronal perikarya and dendrites. In addition, the bulk of the total grains counted were over axons. In lower tectal layers, radioactivity was predominantly over unmyelinated axons and glial elements. High molecular weight RNA appears to be transported within the axons of the optic nerve toward their terminations which are largely in the outer layer of the optic tract.     

Monoaminergic markers in the optic tectum of the domestic chick

The avian optic tectum has become a reliable model system to study the basic mechanisms that underlie the computation of visual stimuli. Many aspects of its cytoarchitecture, chemoarchitecture, connectivity and development are thoroughly characterized. However, knowledge about its monoaminergic innervation is still incomplete. As a prerequisite to understand a possible functional role of the monoaminergic neurotransmitters, the serotonergic, noradrenergic, and dopaminergic innervation of the optic tectum as well as the distribution of serotonin 2A receptors, the dopamine- and cAMP-regulated phosphoprotein DARPP-32 and calbindin D-28K was studied in domestic chicks by immunohistochemical techniques. Serotonergic, noradrenergic, and tyrosine hydroxylase positive axons and axon terminals were present in all layers of the optic tectum. Generally, the highest densities of serotonergic, noradrenergic, and tyrosine hydroxylase positive fibers were found in the superficial tectal layers 1-8, whereas only moderate densities of serotonergic, noradrenergic, and tyrosine hydroxylase positive fibers became obvious in the deep tectal layers 9-15. Serotonergic fibers were particularly abundant in layers 4, 5a and 7 and serotonin 2A receptors in layer 13. Noradrenergic fibers were densest in layers 4 and 5a, whereas tyrosine hydroxylase positive fibers showed a slightly different distribution pattern with additional dense labeling in layer 7. As revealed by double-labeling immunohistochemistry, serotonergic fibers were closely related to the cell bodies of calbindin-positive horizontal cells in layer 5b and tyrosine hydroxylase positive fibers often contacted DARPP-32+ dendritic shafts in layers 9 and 10. These findings indicate that the catecholaminergic innervation of the optic tectum consists of a noradrenergic and a dopaminergic component and that the noradrenergic, serotonergic, and dopaminergic system may be potentially involved in the modulation of retinal input in the superficial layers of the optic tectum as well as in the modulation of tectal output via the deep tectal layers.     

Expression of multiple class three semaphorins in the retina and along the path of zebrafish retinal axons.

Retinal ganglion cells (RGCs) extend axons that exit the eye, cross the midline at the optic chiasm, and synapse on target cells in the optic tectum. Class three semaphorins (Sema3s) are a family of molecules known to direct axon growth. We undertook an expression screen to identify sema3s expressed in the retina and/or brain close to in-growing RGC axons, which might therefore influence retinal-tectal pathfinding. We find that sema3Aa, 3Fa, 3Ga, and 3Gb are expressed in the retina, although only sema3Fa is present during the time window when the axons extend. Also, we show that sema3Aa and sema3E are present near or at the optic chiasm. Furthermore, sema3C, 3Fa, 3Ga, and 3Gb are expressed in regions of the diencephalon near the path taken by RGC axons. Finally, the optic tectum expresses sema3Aa, 3Fa, 3Fb, and 3Gb. Thus, sema3s are spatiotemporally placed to influence RGC axon growth.     

The ramifications and terminals of optic fibres in layers 2 and 3 of the avian optic tectum: a golgi and light and electron microscopic anterograde tracer study

The ramification patterns and terminals of optic fibres in layers 2 and 3 of the optic tectum were studied in Golgi-stained and immunolabelled preparations made from the brains of chicks and pigeons. The different neuronal structures of layers 2 and 3 were also examined. In Golgi preparations, two types of optic fibre were found both in chick and pigeon tectum according to their thickness and terminal branching patterns. The same types of optic fibres were also found to be present in the anterograde tracer experiments after iontophoresis of biotinylated dextran amine into the optic nerve. The varicose terminals of thin fibres mostly terminated on terminal dendritic sections of radiate and pyramidal-like neurons, contacting them on their apical dendrites. The medium-thick fibre terminals in layer 2 mainly established synapses with horizontally extending dendrites, which may therefore be contacts with inhibitory local circuit neurons. The medium-thick optic fibre bushy-like arborisation in layer 3 established synapses with larger dendrites and also stem dendrites. Their terminals formed groups with different dendritic profiles, some of which were partly covered by glial processes, and/or were located among converging dendrites. The presence of these glomerular-like synapses in layer 3 proves that the optic terminals in layer 3 also take part in the transmission of optic impulses to the nucleus rotundus.     

Position-encoding molecules and the development of retinal maps

Molecules that encode position in the retina and a principal target, the optic tectum of frogs, fish and chicks, or its mammalian homologue, the superior colliculus, are hypothesized to control the development of topographic connections by restricting the growth of retinal ganglion cell axons to topographically correct locations in the target. The normal topographic targeting of developing retinal axons is consistent with this concept in frogs and fish, and in chicks it is consistent at the coarse level of matching appropriate retinal and tectal halves. In rats, however, molecules postulated to encode rostral-caudal position do not effectively guide or restrict the growth of retinal axons to topographically appropriate regions of the SC, but seem to contribute to map formation by promoting topographic specificity in branch and arbor formation.     

Growth hormone and its receptor in projection neurons of the chick visual system: retinofugal and tectobulbar tracts

Recent studies have shown the presence of growth hormone (GH) in the retinal ganglion cells (RGCs) of the neural retina in chick embryos at the end of the first trimester [embryonic day (E) 7] of the 21 day incubation period. In this study the presence of GH in fascicles of the optic fiber layer (OFL), formed by axons derived from the underlying RGCs, is shown. Immunoreactivity for GH is also traced through the optic nerve head, at the back of the eye, into the optic nerve, through the optic chiasm, into the optic tract and into the stratum opticum and the retinorecipient layer of the optic tectum, where the RGC axons synapse. The presence of GH immunoreactivity in the tectum occurs prior to synaptogenesis with RGC axons and thus reflects the local expression of the GH gene, especially as GH mRNA is also distributed within this tissue. The distribution of GH-immunoreactivity in the visual system of the E7 embryo is consistent with the distribution of the GH receptor (GHR), which is also expressed in the neural retina and tectum. The presence of a GH-responsive gene (GHRG-1) in these tissues also suggests that the visual system is not just a site of GH production but a site of GH action. These results support the possibility that GH acts as a local growth factor during early embryonic development of the visual system.     

Functional development of the visual system in normal and protein deprived rats II. Morphometric and biochemical studies on adult optic nerve

The optic nerve of normal (C) and protein deprived (PD) adult rats was examined by morphometry and biochemistry. The mean cross-sectional area of the optic nerve was reduced by 15% and the number of axons per unit area increased by 17% in the PD rats. Calibre spectrum analysis of axons revealed a reduction in median diameter from 0.49 μm in controls to 0.45 μm in PD rats. The number of axons with a diameter larger than r μm was reduced by 35% in PD rats. These reductions were probably due to a general reduction in size, since the calculated total number of axons in the optic nerve was almost identical in C and PD rats (126 times 10³ and 124 times 10³, respectively). The increased packing density of axons in the nerve was not only due to thinner axons. The biochemical measurements showed a marked reduction in myelin basic protein in the optic nerves of PD rats, without an alteration in the composition of the total protein. This confirms the persistant hypomyelination which has been reported previously in other malnutrition models. The possible relations between the structural and biochemical changes affecting optic nerve fibres and physiological findings on cortical visual evoked response and on optic nerve in vitro in PD rats are discussed.