Retinal ganglion cell neuroprotection by an angiotensin II blocker in an ex vivo retinal explant model

PURPOSE: An ex vivo organotypic retinal explant model was developed to examine retinal survival mechanisms relevant to glaucoma mediated by the renin angiotensin system in the rodent eye. METHODS: Eyes from adult Sprague Dawley rats were enucleated immediately post-mortem and used to make four retinal explants per eye. Explants were treated either with irbesartan (10 µM), vehicle or angiotensin II (2 μM) for four days. Retinal ganglion cell density was estimated by βIII tubulin immunohistochemistry. Live imaging of superoxide formation with dihydroethidium (DHE) was performed. Protein expression was determined by Western blotting, and mRNA expression was determined by RT-PCR. RESULTS: Irbesartan (10 µM) almost doubled ganglion cell survival after four days. Angiotensin II (2 µM) reduced cell survival by 40%. Sholl analysis suggested that irbesartan improved ganglion cell dendritic arborisation compared to control and angiotensin II reduced it. Angiotensin-treated explants showed an intense DHE fluorescence not seen in irbesartan-treated explants. Analysis of protein and mRNA expression determined that the angiotensin II receptor At1R was implicated in modulation of the NADPH-dependent pathway of superoxide generation. CONCLUSION: Angiotensin II blockers protect retinal ganglion cells in this model and may be worth further investigation as a neuroprotective treatment in models of eye disease.