Morphology Study Of Two Animal Models About Hereditary Retinal Disease

Hereditary retinal diseases are kinds of severe diseases which can causeblind in clinical. And animal models are very important for studying thesediseases. Our laboratory found about two animal models for these two diseases,which are congenital stationary night blindness model rats (CSNB rats) andretinal degeneration fast model mice (rdf mice). The mutation gene of CSNB ratsis Cancna1f gene, and its inheritance is X1inked recessive; The mutation gene ofrdf mice is Pde6b gene, which is autosomal recessive genetic animal model. Wemainly observed that during the stage of development, the changes of retinalhorizontal cells in CSNB rats, and the changes of optic nerve myelin in rdf mice,then discussed the possible mechanism. We try to provide basic experiment basisfor studying hereditary retinal diseases, retinal signal circuit and conduction ofneural signal.Methods1. Wild type SD rats and CSNB rats were divided into different groupsaccording to different time points (postnatal day15、30and60, PND15、PND30 and PND60）to sacrifice, and which follow the rule of randomization.2. Utilized pathological methods, immunohistochemistry methods,immunohistofluorescence methods, wholemount immunocytochemistry methodsand western blotting methods to compare the change of retinal horizontal cells inCSNB rats with wild rats at the same ages.3. Wild type mice and rdf mice were divided into different groups accordingto different treatments(rearing under a12h light/dark cycle or rearing in totaldarkness) and the time points（PND7，PND14，PND28） to sacrifice, and therewere4groups according to different treatments: wild type kunming mice group,rdf mice group, wild type kunming mice group rearing in darkness and rdf micegroup rearing in darkness, and then every group divided into3subgroupsaccording to different time points: PND7，PND14and PND28. Total dark roomwas established, and mice rearing in darkness were reared in it from PND0.4. Utilized pathological methods, immunocytochemistry methods andelectron microscopy methods to observe the change of optic nerve diameter,number of myelinated fibers, myelinated nerve fiber diameter and myelinthickness in wild type mice，rdf mice, and mice rearing in darkness at the sameages.Results1. Results of study of CSNB rats:In CSNB rats, the number of retinal horizontal cells, the density ofhorizontal cells and the level of calbindin protein decreased when compared withage-matched wild type rats at PND15, PND30and PND60. In wild type rats,there was no obvious change in the number of retinal horizontal cells and thedensity of horizontal cells at different time points, but in CSNB rats, the number of retinal horizontal cells and the density of horizontal cells decreased fromPND15to PND60. In wild type rats, the level of calbindin protein increased fromPND15to PND60, but in CSNB rats there was no obvious change in the level ofcalbindin protein at different time points. The axons of horizontal cells at CSNBrats were sparse；Apoptosis can be detected between OPL and ONL at CSNB ratsretina；In CSNB rats, the thickness of OPL decreased when compared withage-matched wild type SD rats at PND15, PND30and PND60（P <0.05）.2. Results of effect of retinal degeneration and light on mice optic nerve myelindevelopment:Optic nerve diameter: In every group, there was a rapid age-dependentincrease in the optic nerve mean diameter. There was no significant differencebetween wild type KunMing mice and wild type KunMing mice reared indarkness at PND7, PND14and PND28. The average optic nerve diameter in therdf mice group was signifcantly decreased compared with age matched wild typeKunMing mice group. For rdf mice reared in darkness, the mean diameter of theoptic nerve was significantly increased when compared with age matched rdfmice group, but compared with age matched wild type KunMing mice groupreared in darkness, no significant differences were observed.The mean number of myelinated nerve fiber: for PND14and PND28wildtype KunMing mice, the mean myelinated nerve fiber of the optic nervesignificantly decreased when compared with wild type KunMing mice reared indarkness. For PND14and PND28rdf mice, the mean myelinated nerve fiber ofthe optic nerve was significantly more when compared with wild type KunMingmice. At PND14and PND28, for rdf mice reared in darkness, the meanmyelinated nerve fiber of the optic nerve was significantly less when comparedwith wild type KunMing mice reared in darkness and when compared with rdf The mean diameter of myelinated nerve fiber: at PND14and PND28, for thewild type KunMing mice, the mean diameter of myelinated nerve fiber wassignificantly larger when compared with wild type KunMing mice reared indarkness. For the rdf mice, the mean diameter of myelinated nerve fiber wassignificantly smaller when compared with age matched wild type KunMing mice.At PND14and PND28, for the rdf mice reared in darkness, the mean diameter ofmyelinated nerve fiber was significantly smaller when compared with rdf micebut significantly larger when compared with wild type KunMing mice reared indarkness.The mean thickness of myelinated nerve fiber:(1) For small axons, the meanthickness of nerve axon in rdf mice was significantly thicker when comparedwith wild type KunMing mice. For wild type KunMing mice reared in darkness,it was significantly thicker when compared with wild type KunMing mice.(2)For large axons, the mean thickness of nerve axon in rdf mice was significantlythinner when compared with wild type KunMing mice. For wild type KunMingmice reared in darkness, it was significantly thinner when compared withKunMing mice.(3) Both for the small and large axons, at PND14the rdf micereared in darkness, the mean thickness of nerve axon was significantly thinnerwhen compared with rdf mice and significantly smaller when compared with wildtype KunMing mice reared in darkness, but at PND28the rdf reared in darknessmice, it was significantly thicker when compared with rdf mice and significantlylarger when compared with wild type KunMing mice reared in darkness.ConclusionOur study found that, the number of retinal horizontal cells in CSNB rats decreased when compared with age-matched wild rats, and the number decreasedfrom PND15to PND60, which suggested that congenital stationary nightblindness can casuse the deficit of retinal horizontal cells and signal transduction.The development of optic nerve myelin disturbed in the rdf mice and lightdeprivation mice, but light deprivation can relieve the effect of retinaldegeneration, which suggested that light deprivation and retinal degeneration canretard optic nerve myelogenesis to some extent, but interestingly light deprivationcan protect optic nerve myelogenesis after retinal degeneration at earlydevelopment stages.The experiment results indicate that genetic disease which caused by geneticdefects can influence the visual circuit and visual signal transmission, and thiseffects are not just limited to direct target tissue and cell structure, but also affectother parts of the visual circuit because visual circuit is a complex and integratedstructure. The experiment results further suggest that, the vision developmentstage is the important stage in which diseases progress and visual circuitdevelopment, which suggests that it is critical for treatment in this period and thetherapeutic effect may be more significant.