Controlled Elevation of Intraocular Pressure (CEI): Physiologic and Ocular Responses in the Mouse

Abstract

Purpose : To develop a protocol for prolonged anesthesia in mice and evaluate optic nerve axon injury in response to 4 hours of controlled elevation of intraocular pressure (CEI).

Methods : C57BL/6NCrl male retired breeder mice (8 – 10 months old) were anesthetized with 1.5% isoflurane with 100% O₂ for 4 hours, using active evacuation of expired gas and anesthetic, hydrated with intraperitoneal, warm lactated ringers (0.5 ml) with 5% dextrose at the start and end of the anesthesia, and temperature maintained with a warm water therapy pad. Oxygen saturation (O₂), heart rate (HR) and respiratory rate (RR) were monitored throughout the experiment with a Mouse Ox Plus and blood pressure (BP) with a CODA tail cuff manometer. For CEI, one eye was cannulated with polyurethane tubing (tip diameter, 0.006 inch) connected to a reservoir containing balanced salt solution and elevated to 60 mmHg, verified by rebound tonometry.Ten days after exposure, mice were perfusion-fixed with glutaraldehyde to optimize morphology. Six masked observers graded optic nerve cross sections for injury on a scale from 1 (normal) to 5 (>50% of axons degenerating). Mean grades for 4 groups of eyes [(1) experimental CEI; (2) fellow; (3) anesthesia alone; and (4) naïve (no anesthesia or cannulation)] were calculated and compared by ANOVA.

Results : Mean physiologic values over the course of anesthesia (N= 13) were: O₂ = 91 ± 2% (SEM), HR = 571 ± 7 and RR = 156 ± 13. Systolic, diastolic and mean BPs (N = 4) were 92± 14, 71±11 and 77 ± 12 mmHg, respectively. One animal died during the experiment and one failed to recover following anesthesia. Mean (± SEM) optic nerve injury grades were: 2.03 ± 0.41 for CEI eyes (N=7); 1.23 ± 0.16 for fellow eyes (N = 7); 1.09 ± 0.03 for naïve eyes (N = 8); and 1.06 ± 0.03 (N = 6) for anesthesia only eyes. ANOVA analysis identified the CEI group as the only one to have a significantly higher injury grade (CEI vs naïve, P = 0.02). No significant differences were noted between control, anesthesia only and naïve groups (p>0.99, all comparisons). Physiologic parameters in animals with greater CEI injury were similar to those with minimal injury.

Conclusions : Prolonged anesthesia in mice requires careful attention to animal physiology. With this, a 4-hour exposure to elevated IOP can produce variable, but significant optic nerve injury.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.