Histological Evaluation of Diabetic Neurodegeneration in the Retina of Zucker Diabetic Fatty (ZDF) Rats

In diabetes, retinal dysfunctions exist prior to clinically detectable vasculopathy, however the pathology behind these functional deficits is still not fully established. Previously, our group published a detailed study on the retinal histopathology of type 1 diabetic (T1D) rat model, where specific alterations were detected. Although the majority of human diabetic patients have type 2 diabetes (T2D), similar studies on T2D models are practically absent. To fill this gap, we examined Zucker Diabetic Fatty (ZDF) rats - a model for T2D - by immunohistochemistry at the age of 32 weeks. Glial reactivity was observed in all diabetic specimens, accompanied by an increase in the number of microglia cells. Prominent outer segment degeneration was detectable with changes in cone opsin expression pattern, without a decrease in the number of labelled elements. The immunoreactivity of AII amacrine cells was markedly decreased and changes were detectable in the number and staining of some other amacrine cell subtypes, while most other cells examined did not show any major alterations. Overall, the retinal histology of ZDF rats shows a surprising similarity to T1D rats indicating that despite the different evolution of the disease, the neuroretinal cells affected are the same in both subtypes of diabetes

Rate of regrowth of damaged retinal ganglion cell axons regenerating in a peripheral nerve graft in adult hamsters

The rate of regrowth of ganglion cell axons regenerating into a peripheral nerve graft implanted into the retina of adult hamster was measured, utilizing the method of retrograde labelling by horseradish peroxidase. The fastest regrowing axons were found, after an initial delay of 4.5 days, to extend at about 2 mm/day in the graft. The role of the cell body in controlling the rate of axonal regeneration was briefly discussed.link_to_subscribed_fulltex

Regrowth of retinal ganglion cell axons into a peripheral nerve graft in the adult hamster is enhanced by a concurrent optic nerve crush

Transplantation of a segment of peripheral nerve to the retina of the adult hamster resulted in regrowth of damaged ganglion cell axons into the graft, with the fastest regenerating axons extending at 2 mm/day after an initial delay of 4.5 days (Cho and So 1987b). In this study, the effect of making 2 lesions on the same axon (the conditioning lesion effect) on the regrowth of ganglion cell axons into the peripheral nerve graft was examined. When a conditioning lesion (first lesion) was made by crushing the optic nerve 7 or 14 days before the peripheral nerve grafting (the second lesion) to the retina, the distance of regrowth achieved by the fastest regenerating axons in the graft, measured at the 7th post-grafting day, was lower than in animals with a peripheral nerve grafted to a normal eye. This indicated that in contrast to the situation in peripheral nerve axons (Forman et al. 1980) and goldfish optic axons (Edwards et al. 1981), the conditioning lesion was unable to enhance the regrowth of mammalian retinal ganglion cell axons. However, when crushing of the optic nerve was followed immediately by peripheral nerve grafting, an enhancement in axonal regrowth could be observed. The initial delay time before the axons extended into the peripheral nerve graft was reduced by 1 day while the rate of elongation of the fastest regrowing axons in the graft apparently remained unchanged. Moreover, the shortening of the initial delay could still be observed even when the sequence of performing the 2 lesions was reversed. From these data, it was concluded that the classical conditioning lesion effect was not responsible for the enhancement observed. Rather it was suggested that changes in the intra-retinal environment brought about by crushing of the optic nerve might account for it.link_to_subscribed_fulltex

De novo formation of axon-like processes from axotomized retinal ganglion cells which exhibit long distance growth in a peripheral nerve graft in adult hamsters

Damaged axons in the central nervous system of the adult mammal can be stimulated to regenerate extensively into a peripheral nerve graft. It was generally believed that the new axonal sprouts which extend into the graft arose from the injured proximal axonal stumps. However, when the retinal ganglion cells of the adult hamster were axotomized by crushing the optic nerve and the proximal axonal stump was not in direct apposition to the graft, a new axon-like process could be seen to be emitted from either the cell soma or dendrite and extended in the graft for at least 1-2 cm. This axon-like process was distinct from the original injured axon which could still be seen to course towards the optic disc in the retina. Evidently, even a fully differentiated central nervous system neuron of the adult mammal retains a great degree of morphological plasticity so that if the original axon is discouraged to regrow after injury, other parts of the neuron can act as favourable sites for the sprouting of a new axon-like process.link_to_subscribed_fulltex

Sprouting of axon-like processes from axotomized retinal ganglion cells is influenced by the distance of axotomy from the cell body and the mode of transplantation of the peripheral nerve

It is a well known fact that the proximity of an axonal lesion from the cell body influences the degree of neuronal survival; a lesion close to the cell body leads to more severe cell death and vice versa. On the other hand, experiments involving transplantation of a peripheral nerve (PN) to various central nervous system (CNS) regions to induce axonal regeneration have suggested that axonal regrowth is more vigorous when the grafting is performed closer to the cell body. It is not clear, however, whether it is the distance of the site of axotomy or the location of the trophic source (PN graft) or both from the cell body which dictates the vigorousness of axonal regrowth. Using either a model of transplantation of a PN to the retina or implantation of a short PN into the vitreous body of the eye of the adult hamster, we have demonstrated that sprouting of axon-like processes from retinal ganglion cells (RGCs) depends on the distance of axotomy from the cell body when the PN graft is maintained at a constant distance from the cell body. Moreover, it was found that the distance of axotomy at which sprouting of axon-like processes could be induced was different for the 2 paradigms: with the intravitreal PN model, sprouting was observed even after intracranial ON cut whereas it was absent in the PN grafting-to-retina paradigm. This suggests that extrinsic influence (in this case an intravitreal PN) can overcome to a certain extent the growth-suppressive effects due to a certain extent the growth-suppressive effects due to a long distance of axotomy.link_to_subscribed_fulltex

Characterization of the sprouting response of axon-like processes from retinal ganglion cells after axotomy in adult hamsters: A model using intravitreal implantation of a peripheral nerve

Peripheral nerves provide a favourable environment for damaged CNS axons to sprout and regenerate. It has also been demonstrated that retinal ganglion cells respond to a peripheral nerve segment grafted to the retina by emitting axon-like processes from the somatodendritic compartment into the graft. The factors influencing the pattern of sprouting of axotomized retinal ganglion cells were explored in this study by implanting a short segment of peripheral nerve, which did not come into contact with the retina, into the vitreous body of an eye whose optic nerve was concurrently crushed. Silver staining was used to assess the morphology of the retinal ganglion cells which underwent sprouting. Some retinal ganglion cells were induced to sprout axon-like processes; these emerged primarily from dendrites and less frequently from the soma or intraretinal axon. Implantation of a nonviable graft (freezed-thawed) elicited only minimal sprouting. These results suggest that diffusible factors secreted by cells in the graft are a possible stimulus to sprouting in axotomized retinal ganglion cells. Examination of the pattern of dendritic sprouting indicates that sprouting was most intense (in terms of number of sprouts per cell) at early times post-axotomy. Moreover, a differential pattern of development of sprouts arising from individual primary dendrites of the same cell was observed; sprouts tend to arise from all primary dendrites initially but as the post-axotomy time increased, retraction of sprouts from some primary dendrites occurred. Concomitant with this retraction, however, there was an increase in the number of sprouts on those primary dendrites which were still in the active phase of sprouting. Selective stabilization of sprouts by extrinsic factors may account for this phenomenon. Changes in the area and outline (irregularity) of the somata of retinal ganglion cells with sprouts from two weeks to two months after optic nerve crush could be correlated temporally with the intensity of sprouting from the dendritic tree, suggesting that during sprouting, intrinsic mechanisms coordinate the responses of different cellular compartments. In contrast to extensive ectopic sprouting of axotomized retinal ganglion cells in the presence of an intravitreal graft, when a long peripheral nerve segment is grafted to the cut optic nerve, there is extensive axonal regeneration into the graft from retinal ganglion cells, most of which did not exhibit ectopic sprouting. Thus, a hierarchy of sprouting sites within a neuron seems to exist, with the damaged axonal tip being the most favoured site, followed by the dendrites, and then the intraretinal axon. The soma appears to be the least preferred compartment for sprout emission.link_to_subscribed_fulltex

Morphological changes of retinal ganglion cells regenerating axons along peripheral nerve grafts: A Lucifer yellow and silver staining study

The morphology of the retinal ganglion cells (RGCs) with their axons regenerating along a peripheral nerve graft at different post-grafting periods was studied by the intracellular injection of Lucifer yellow (LY) and silver staining methods. Several morphological features which were observed on developing RGCs, but not mature RGCs, have also been observed in the regenerating RGCs studied by the intracellular injection of LY. These morphological features observed on the regenerating RGCs included intraretinal axonal branches and collaterals, spine-like processes on the dendrites and soma, and short processes on the soma. These results suggest that damaged mammalian RGCs may be able to recapitulate certain cellular events which occur during normal development provided the regenerating cells are given the proper stimulus and a favorable environment for regrowth. From the results of both LY injection and silver staining experiments, it was found that the dendrites of the regenerating RGCs were, in general, much simpler than that of control Type I RGCs. However, regenerating RGCs with different degree of dendritic complexity could be observed in all post-grafting periods studied, and the dendritic complexity seems to decrease continuously with the increase in the post-grafting time. These results suggest that the ability to regenerate an axon is not closely related to dendritic responses and the peripheral nerve does not seem to be able to prevent the deterioration and retraction of the dendrites.link_to_subscribed_fulltex