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K.–S. Cho, D. Chen; Abnormal Segregation of Regenerating Axons at the Optic Chiasm in Bcl–2 Transgenic Mice . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1579.
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In mammals, injured optic nerve usually does not regenerate. We recently showed that over–expressing Bcl–2 gene in neurons supports robust optic nerve regeneration from the nerve to the brain in the postnatal stage. However, most regenerating fibers enter the ipsilateral side of the brain and terminate at the visual targets. It will be interesting to understand the underlying mechanisms that prevent the majority of regenerating fibers entering the contralateral side of the optic chiasm. Axon guidance molecule, EphB1, has been reported to have a key role in the ipsilateral projection of the optic nerve in development. The pattern of EphB1 expression changes significantly in the postnatal retina. In this study, we sought to determine whether EphB1 plays a role in axon guidance following optic nerve regeneration in postnatal Bcl–2 transgenic mice.
Optic nerve crush procedure was performed in Bcl–2 transgenic mice that were maintained at a C57BL/6J genetic background. Age–matched C57BL/6J mice were used as controls. Immediately following optic nerve crush, an anterograde tracer cholera toxin B subunit conjugated with fluorescein or rhodomine (CTB) was injected into the vitreous cavity to label regenerating axons. Mice were sacrificed at 4 days post–crush and perfused with saline and 4% paraformaldehyde. The optic nerve and brain sections were prepared, and axon projection trajectory after entering the optic chiasm was examined. To determine EphB1 expression in the retina, mice received optic nerve crush and sham–operation were examined. Total retinal RNAs were extracted, and the levels of EphB1 expression were determined using semi–quantitative reverse transcriptase–polymerase chain reaction. Moreover, mice carrying Bcl–2 transgene and simultaneously, lack of EphB1 gene, were generated, and optic nerve regeneration in this mouse line was studied.
We noted that the majority of regenerating axons in postnatal Bcl–2 transgenic mice stayed on the ipsilateral side of the chiasm and the brain. No regeneration was observed after optic nerve injury in wild type mice. Following optic nerve injury, significant increase in EphB1 expression was observed in retinal tissues, as determined by RT–PCR.
The majority of regenerating axons projected to the ipsilateral brain targets, correlating with the increased expression of EphB1 in their retinas after optic nerve injury in postnatal Bcl–2 transgenic mice. These data suggest that EphB1 may be a key player in guiding regenerating axon to the ipsilateral brain targets in the postnatal stage.
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