RETINAL GANGLION CELL DYSFUNCTION AND SYNAPTIC ALTERATIONS ARE SUBTYPE-SPECIFIC IN A MOUSE MODEL OF OCULAR HYPERTENSION

Glaucoma is a group of neurodegenerative disorders characterized by retinal ganglion cell (RGC) loss with optic nerve cupping and visual field defects. Previous studies have suggested that there may be specific types of RGCs that are more vulnerable to elevated intraocular pressure (IOP). The purpose of this study is to determine the effects of laser- induced ocular hypertension (LIOH) on various subtypes of RGCs at different time points after IOP elevation. IOP was elevated unilaterally using laser photocoagulation of the limbal and episcleral vessels of adult CD-1 mouse eyes and measured by rebound tonometry. Multielectrode array recordings were performed 3, 7, 14 and 30 days after IOP elevation. Based on their response to a square-wave stimulus, RGCs were classified into: OFF-transient (OFF-T), OFF-sustained (OFF-S), ON-transient (ON-T), and ON-sustained (ON-S). Light response properties were quantified. In addition, retinas were biolistically transfected with YFP- tagged PSD95 and prepared for whole-mount retina immunohistochemistry using SMI-32 to label OFF-T and ON-S RGCs, as well as CtBP2 to label presynaptic ribbons. After LIOH, the IOP of the treated eye was elevated as early as 6 hours and returned to baseline by 1 week. Spontaneous activity was quantified for all subtypes: OFF-T RGCs showed the earliest decrease 14 days after IOP elevation, ON-S and OFF-S after 30 days, whereas activity of ON-T RGCs was unchanged within the observation time. We recorded the light responses of these RGC subtypes to Gaussian white noise stimulation, and used reverse-correlation methods to evaluate the spatial structure of their receptive field (RF). A significant decrease in the RF size was found only in OFF-T RGCs, as early as 7 days after IOP elevation. Morphologically, all alpha-like RGCs displayed reduction of PSD95 puncta across their dendritic arbor, 7 to 14 days after IOP elevation. In parallel to postsynaptic changes on RGCs, presynaptic ribbon density declined in the inner plexiform layer (IPL), with the greatest reduction observed within the OFF sublamina. Finally, examination of alpha-like RGC density by SMI-32 immunostaining revealed that the proportion of OFF-T RGCs that were died by 14 days after IOP elevation is greater than that of ON-S RGCs. Taken together, these data suggest that IOP elevation differentially affects various RGC subtypes. Neurons stratifying in the OFF sublamina undergo the highest reduction of excitatory synapses prior to cell death. Among alpha-like RGCs, OFF-T RGCs exhibit the earliest loss of function, whereas ON-T RGCs exhibit relative resistance to elevated IOP.