Retinal ganglion cells (RGCs) are the output neurons of the retina. All our visual experiences derive from signals generated from RGCs traveling down through the optic nerves. This thesis aims to investigate the signaling of RGCs in both physiological and pathological conditions. Our first aim was to characterize the morphology of amacrine cells coupled with alpha RGCs (αRGCs). As myopia is a prevalent abnormal eye conditions and defocus is the main trigger factor, our second aim was to investigate the effect of focused/defocused image projection on the signaling of αRGCs. As atropine is widely used to control myopic progression in children, our third aim was to explore the effect of low-does atropine on the signaling of αRGCs.
Retinitis pigmentosa is a blinding neurodegenerative disease. The fourth aim was to explore the unmasking effect of PTX on signaling of αRGCs in rd10 mouse. The last part focused on glaucoma, the leading cause of visual loss. We aimed to characterize the effect of elevated IOP on αRGCs activities in the mouse model.
Wild-type C57BL/6 mice, the KCNG-YFP mice, rd10 mice, and/or Cx36-knockout mice were used in the study. Dye-injection (Neurobiotin and 594 mixed with Popro1) and confocal microscopy were used to investigate the morphology of neurons in the mouse retina. For electrophysiology, spikes or current responses of RGCs were recorded. Either 525nm full-field light or pattern light projected by an Organic Light-Emitting Diode (OLED) micro-display was used as light stimulation.
Study 1. The ON and OFF αRGCs, which were fluorescence-labeled in the KCNG-YFP mice, were coupled with at least two types of ACs. Most coupled ACs were wide-field ACs.
Study 2. Custom-made light patterns were projected as focus/defocused images onto the retina, resulting in generation of different defocused statuses. Light-evoked current responses of αRGCs were recorded by patch clamp. Myopic and hyperopic defocus led to a decrease of the light-evoked current response in αRGCs.
Study 3. Approximately 2 μM atropine was detected in the retina after external application of 800 μM (equal to 0.05% topical application) atropine. Low-dose atropine (1 μM) did not change the morphology of the αRGCs. Most physiological properties were unchanged after 0.05-10 μM atropine application. However, ON responses were induced in OFF αRGCs by the low to high concentrations of atropine, which might block the GABA pathway and affect the signaling of the retina.
Study 4. The morphology of αRGCs in P46 rd10 and wide-type mice did not show significant difference. PTX increased the excitatory postsynaptic current and decreased the inhibitory postsynaptic currents of αRGCs. PTX induced light-evoked spike responses in 47.5% of tested αRGCs, mainly by blocking the GABAA receptors. Glycine and dopamine receptor antagonists
could not induce light-evoked spike responses. PTX application could improve the ERG and optokinetic responses in P41 rd10 mice.
Study 5. The high IOP mouse model was produced by anterior injection of 1 μm and 6 μm microbeads mixtures. The IOP was increased by 3.28 ± 1.41 mmHg 7 days after injection, and this elevation could persist for up to 28 days after injection. RGC number decreased, and light sensitivity of OFF αRGCs increased under 4-week IOP elevation.
Alpha RGCs are coupled with wide-field ACs. Electrical activities of RGCs could reflect different pathological conditions. Defocused stimuli decreased the inputs of αRGCs to inhibit the firing, which might be the first step of the retina sensing the defocus, providing clues to prevent myopia in electrophysiology. The mild change of atropine on αRGCs signaling might suggested the mechanisms of side effects in clinical administration. Removing inhibition unmasked light responses of αRGCs derived from retained cones, suggested a therapeutic clue for RP and other neurodegenerative diseases. Enhanced excitability hypothesized in our
glaucomatous model suggested an early change before RGCs degeneration.
Overall, these observations of signaling of RGCs in physiological and pathological conditions have important consequences for our understanding of the neural information processing in the retina. The RGCs signaling suggested very early pathological changes and mechanisms in myopia, RP, and glaucoma, providing us biophysical mechanisms to target in exploring preventive strategies for these eye diseases.
|Date of Award||16 Feb 2022|
|Supervisor||Chi Wai Do (Co-supervisor) & Feng Pan (Supervisor)|