Date of Award

12-2025

Degree Name

Master of Science

Department

Biological Sciences

First Advisor

Cindy Linn, Ph.D.

Second Advisor

Pamela Hoppe, Ph.D.

Third Advisor

John Spitsbergen, Ph.D.

Access Setting

Masters Thesis-Open Access

Abstract

Experiments were performed to demonstrate electrical recovery in glaucoma-induced adult mice after neurogenesis of retinal ganglion cells (RGCs). Newly derived RGCs were induced following eye drop treatment with a selective alpha7 nicotinic acetylcholine receptor agonist, PNU-282987. Both sexes of transgenic (RlbpCre-ER (CRE+) and Rosa-tdTomato (tdTomato+)) adult mice were utilized to trace newly generated RGCs originating from Müller glia (MG). Experimental mice were injected with hypertonic saline into the episcleral vein of anesthetized mice to significantly increase intraocular pressure (IOP) and induce glaucoma-like conditions. One month after the injection and after significant death of RGCs occurred, mice began daily eye drops containing PNU-282987 for two weeks. In addition, weekly ERGs were recorded using the Celeris Diagnosys ERG System before inducing glaucoma, after the procedure to induce glaucoma, and after the glaucoma-inducing procedure followed by PNU-282987 treatment. After treatments, animals were euthanized, retinas were removed, processed with antibodies against RFP and Thy 1.2, counterstained with DAPI, and viewed with a Nikon confocal microscope. Cell counts were obtained under all conditions and compared to internal controls. Hypertonic saline injections elicited a significant increase in IOP within 3 weeks, which corresponded with a significant loss of RGCs. However, the loss of RGCs was eliminated if PNU-282987 treatment began after RGCs death occurred, supporting the hypothesis that PNU-282987 induced neurogenesis of RGCs. Triple labeled retinal sections from transgenic mice demonstrated that new Thy1.2+ RGCs were generated from MG. In ERG recordings, the amplitude of PhNR, pERG and pVEP recordings significantly decreased after hypertonic saline injection. All these recordings returned to control values when PNU-282987 treatment began after inducing glaucoma. Results from these studies support the hypothesis that PNU-282987 induces neurogenesis in glaucoma-induced adult mice and elicits functional recovery in ERG waveforms associated with RGC activity.

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