Changes in retinal responses and morphology induced by elevated intraocular pressure in mice

To determine time courses of retinal structural and functional changes in wild type mice following the induction of ocular hypertension by anterior chamber injection of polystyrene beads. Induction of ocular hypertension: 6 week old C57BL/6J female mice were anesthetized and had the left eye dilated. A 30 gauge needle was used to puncture the cornea. One of the following mixes was injected over 90 seconds via a glass micropipette attached to a Hamilton syringe and inserted into the anterior chamber through the corneal incision. Mix 1 contained one microliter of a fixed combination of 1 and 6 micron diameter dyed polystyrene beads (Polysciences, Inc.) followed by 3 microliters of Provisc (Alcon, Inc.). Mix 2 contained 2 microliters of the same bead combination followed by 2 microliters of Provisc. Mix 3 contained 2 microliters of PBS followed by 2 microliters of Provisc. The right eye was untreated. IOP was measured with a rebound tonometer (Tonolab) three times a week for 6 weeks.Histology: Six weeks after IOP induction, freshly enucleated eyes were placed optic nerve down in 8% neurobiotin for 20 minutes and washed with oxygenated Ames solution for several light-dark cycles. Retinas were fixed, blocked, incubated with secondary antibody, and whole-mounted. Images were acquired with a confocal microscope and processed in Photoshop.ERGs: Scotopic a- and b- waves and positive and negative scotopic threshold responses (STRs) were measured at 1, 2, 3, 4, and 6 weeks after IOP induction according to previously published protocols. Mix 1 (n = 11) and Mix 2 (n = 18) caused an average increase in cumulative IOP (sum of all measured IOPs) of 35% and 36%, respectively. 28 of 29 animals (97%) had an increased cumulative IOP in the treated eye compared to the untreated eye. Mix 1 and Mix 2 had an average IOP of 12.8mmHg and 12.9mmHg, respectively. Control (untreated) eyes had an average IOP of 9.3mmHg. Only 3 animals (10%) had a measured IOP of 30mmHg or greater at any point. Mix 3 (n = 4) caused an average increase in cumulative IOP of 8% and no animals had an IOP of 30mmHg or greater at any point. Animals that were injected with beads showed a reduction in neurobiotin-positive retinal ganglion cells (RGCs) of 15.7% when compared to untreated control eyes (n = 3). The a- and b-wave amplitudes of bead treated (n = 9), PBS treated (n = 4), and control eyes were essentially the same at all tested light intensities and time points. Analysis of the pSTR and nSTR is currently ongoing. Injection of either volume of polystyrene beads into the mouse anterior chamber results in a reliable, chronic, mild elevation of IOP with a low incidence of IOP spikes. Like human glaucoma, this technique leads to RGC loss without reduction of outer retinal function.