Graft–host connections in long-term full-thickness embryonic rabbit retinal transplants. Invest Ophthalmol Vis Sci

PURPOSE. To establish neiironal connections in the rod and cone pathway between laminated rabbit retinal transplants and the host retina. METHODS. Fourteen adult rabbits received a complete full-thickness embryonic transplant. After survival times of 3 to 10 months, the retinas were studied under light microscope and with immunohistochemistry. Antibodies against protein kinase C (PKC), parvalbumin, and calbindin were used to label rod bipolar cells, All amacrine cells, and cone bipolar cells, respectively. The A155 antibody was used to label ganglion cells. RESULTS. The transplants displayed laminated morphology with layers parallel to the host retinal pigment epithelium. In the oldest specimens (10 months after surgery), laminated layers of graft and host approached each other and almost reconstructed the normal retinal appearance. The ganglion and cone bipolar cells of the host survived well, as was seen with AI35 and calbindin double-labeling. Connections between cone bipolar cells in the graft and ganglion cells in the host were not common. PKC-labeled rod bipolar cells and parvalbumin-labeled All amacrine cells of host and graft showed sprouting activity directed toward an intermediate plexiform layer located between the graft and host. In specimens double-labeled with PKC and parvalbumin, this intermediate plexiform layer was seen to contain numerous PKC-and parvalbumin-labcled processes. Direct connections between rod bipolar and All amacrine cells in host and graft were seen in the 10-month specimens. CONCLUSIONS. Full-thickness embryonic transplants survive for at least 10 months, and normal laminated morphology develops. Host and graft fuse and together contribute nerve cell processes to an intermediate plexiform layer. Direct graft-host contacts are also present between neiironal types that in the normal retina participate in the rod pathway. (Invest Opbthalmol Vis Sci. 1999; 40:126-132) R etinal transplantation offers the possibility of restoring visual function in eyes affected by degenerative disease. Different techniques have been used, and reports on the survival of transplants are numerous. 1 " 4 To serve its purpose, however, the retinal transplant must form functional contacts with the host retina. Reports on such graft-host connections in retina-to-retina transplantation are few. Contacts between lacZ-labeled photoreceptor and host retinal cells have been investigated, 5 but the results were inconclusive, because the origin of their postsynaptic targets have not been established. Sprouting activity has been reported in Dil-labeled grafts, r> but to our knowledge, no study has shown connections between selectively labeled neurons of the graft and host

Retinal function and histopathology in rabbits treated with Topiramate.

Purpose To evaluate retinal function and histopathology in rabbits treated orally with the antiepileptic drug topiramate. Methods Six rabbits were treated with a daily oral dose of topiramate during a period of eight months. Six rabbits receiving water served as controls. Blood samples were analyzed for determination of topiramate serum levels in order to ensure successful drug exposition. Standardized full-field electroretinograms (ERGs) were performed before treatment and then at 2, 3 and 8 months during the treatment period. After terminating treatment the rabbits were sacrificed and the morphology of the sectioned retina was studied. Results After eight months of treatment the fullfield ERG demonstrated normal rod function in treated and control rabbits, but the light adapted 30 Hz flicker b-wave amplitude was significantly reduced in the treated rabbits. This was the case for both the light adapted (Wilcoxon signed ranks test, P=0.046) and the dark adapted (Wilcoxon signed ranks test, P=0.028) 30 Hz flicker response from the treated rabbits. Retinal immunohistology revealed a severe accumulation of GABA in amacrine cells and in the inner plexiform layer in 4 of 6 treated rabbits compared to the controls. Conclusions Topiramate, orally administrated to rabbits, may cause a significant reduction of the retinal function demonstrated by the reduced b-wave amplitude in the full-field ERG, as well as changes in immunohistology characterized by a severe accumulation of GABA in the inner retina. The retinal dysfunction and the morphological changes indicate that topiramat may damage the retina, similarly to vigabatrin (another antiepileptic drug)

Retinal glial cell immunoreactivity and neuronal cell changes in rats with STZ-induced diabetes

PURPOSE: To study whether diabetes could influence glial cells, retinal neurons, and pigment epithelial cells and if so, to evaluate whether any changes could be influenced by aminoguanidine (AG) or probucol (PB). METHODS: Streptozocin (STZ)-induced diabetic male Wistar rats and age-matched control rats were fed a normal diet, addition of AG in the drinking water (0.5 g/l for diabetic and 1.0 g/l for control rats) or PB in the pellets (1 % w/w) for one or six months. Paraffin embedded retinal sections were incubated in the primary antibodies GFAP, calbindin, RPE65, and Hu, for glial, horizontal, pigment epithelial, and ganglion cells, respectively, and in fluorescent secondary antibodies. RESULTS: One month after STZ injection, GFAP immunoreactivity was sparse, but after six months it was prominent in glial cells in 5/5 diabetic and 1/7 control retinas (p = 0.015). Neither AG, nor PB influenced this immunoreactivity. Numbers of retinal pigment epithelial cells and cells in the ganglion cell layer, were similar at one and six months of diabetes. By time, the number of horizontal cells decreased (p < 0.001) and branching and numbers of their terminals were reduced (p < 0.001). CONCLUSION: Diabetes for six months resulted in increased glial cell immunoreactivity, and by age, horizontal cell numbers and branching of their terminals decreased, morphological patterns that were unaffected by AG or PB. The numbers of retinal pigment epithelial cells and cells in the ganglion cell layer were unaffected both by age and diabetes

Dose-related changes in retinal function and PKC-alpha expression in rabbits on vigabatrin medication : Effect of vigabatrin in the rabbit eye.

BACKGROUND: To investigate, in a rabbit model, the effect of two different doses of vigabatrin (VGB) on retinal function and morphology. METHODS: Twenty-nine rabbits of mixed strain were divided into two groups, receiving either high-dose (n = 15) or low-dose (n = 14) oral VGB treatment (cumulative dose 29.8 +/- 2.9 g and 14.2 +/- 0.6 g respectively). Ten rabbits receiving water served as control animals. The rabbits underwent three baseline ff-ERG measurements before initiation of VGB medication and two ff-ERG registrations during treatment, after 8 and 12-14 weeks respectively. At the end of the study, the expression of protein kinase C-alpha (PKC-alpha), gamma amino butyric acid (GABA) A receptors, vimentin, glial fibrillary acidic protein (GFAP) and peanut agglutinin (PNA) was examined in retinal sections from all rabbits. RESULTS: In animals of the high-dose group, the ff-ERG measurements revealed a significant decrease of isolated rod b-wave amplitudes, combined rod-cone b-wave amplitudes and amplitudes of oscillatory potentials (OPs); OP1, OP2 and OP3. In the low-dose group, the b-wave amplitudes of combined rod-cone responses as well as OP2 and OP3 were significantly reduced. PKC-alpha labeling demonstrated a dose-related translocation of the enzyme in rod bipolar cells, also revealing a significant decline of the number of PKC-alpha labeled rod bipolar cells in treated animals. Vimentin labeling showed a dose-related deviant labeling pattern of Müller cells, with strikingly low labeling intensity in the outer parts of the cells; in the outer limiting membrane (OLM) as well as the outer nuclear layer (ONL). Labeling with antibodies against GABA A receptors and GFAP, as well as PNA staining, revealed no differences between treated animals and controls. CONCLUSIONS: In this study, VGB medication was associated, in a dose-related manner, with a decrease of ff-ERG amplitudes as well as with altered protein expression in rod bipolar cells and Müller cells, suggesting alterations of inner retinal function. The dose-related morphological and electrophysiological changes indicate a retinal pathology that may explain the constricted visual fields seen in some patients treated with VGB

Alterations in electroretinograms and retinal morphology in rabbits treated with vigabatrin

PURPOSE: To determine whether long-term treatment with the anti-epileptic drug vigabatrin causes damage to rabbit retina.METHODS: Five rabbits were treated continuously with a daily dose of vigabatrin solution per orally during a period of 1-8 months. Two rabbits receiving water were used as controls. Repeated full-field electroretinograms (every two weeks) were assessed during this period. Vigabatrin serum concentration was repeatedly measured for securing successful drug administration. After termination of treatment the rabbits were sacrificed and the morphology of the sectioned retina was studied.RESULTS: In all rabbits treated with vigabatrin the serum analyses repeatedly demonstrated elevated drug concentration. Full-field electroretinograms demonstrated normal rod function in all treated rabbits, but reduced cone function in two of the five treated rabbits verified by 30Hz flicker stimulation. Morphologic studies of the sectioned retina demonstrated GFAP immunoactivity of the glial cells localized in the retinal periphery in all five treated rabbits, one of which had staining also in the centrally localized glial cells. The treated rabbits also demonstrated a weaker GAD staining in the IPL and less positive amacrine cells, compared to the controls. Only two treated rabbits had normal GABA staining while three had an enhanced GABA immunoreactivity and undistinguishable fibers in the IPL. In three out of five treated rabbits the Müller cells were short, stubby and fragmented, with swollen endfeet.CONCLUSION: This study demonstrates changes in histopathology caused by vigabatrin in an animal model, which has not been reported previously. We have found that vigabatrin orally administrated to rabbits does not affect rod function but may reduce cone function in the full-field electroretinogram, which is similar to the previously reported vigabatrin effect on the human ERG. The results indicate that vigabatrin may damage or influence, at least one cell type in the rabbit retina

Heat shock protein 70 (HSP70) after laser thermotherapy of an adenocarcinoma transplanted into rat liver.

The heat shock proteins (HSPs) HSP70 and gp96 from necrotic tumour cells are considered to function as chaperones in presenting tumour antigens. We therefore studied HSP70 and immune cells in a transplantable carcinoma in the liver of rats after interstitial laser thermotherapy (ILT). Experiments were performed in Wistar FU rats using a dimethyl-hydrazine-induced adenocarcinoma implanted into the left lateral lobe of the liver. Rats were randomized to one of the following groups: a) ILT of tumour, b) sham ILT, or c) control. ILT was suboptimal and was performed at a steady-state temperature of 43 degrees C at the tumour margin for 30 minutes. Rats were killed 15 minutes, 5 hours, 10 hours, 15 hours or 12 days after treatment. Double immunohistochemistry was performed for HSP70 and ED1 macrophages or CD8 lymphocytes, and ELISA for serum concentrations of HSP70. After ILT, there was an increase of HSP70 immunoreactivity in tumours as compared to sham ILT. At the same time, tumour cells affected by ILT showed a shift of HSP70 from the cytoplasm to the nucleus with a peak at 10 hours. Few CD8-positive cells were found. There was an increase of tumour-infiltrating ED1 macrophages after ILT as compared to sham ILT at 10-15 hours after treatment. HSP70 was present in ED1 macrophages significantly more frequently after ILT than after sham ILT, and this was true both for HSP70 localized to the surface and the cytoplasm of the macrophage. There was a significant increase in serum HSP70 during the first 15 hours after ILT. In conclusion, laser thermotherapy resulted in increased HSP70 immunoreactivity within tumours and HSP70 shifts from cytoplasm to nucleus. Furthermore, it resulted in increased numbers of tumour-infiltrating macrophages and an increased presence of HSP70 in the membrane and cytoplasm of these macrophages