Purpose
To define the influences of IOP, anesthesia and anterior chamber cannulation on ONH gene expression in the CEI model following an 8 hour exposure to elevated IOP in rats.
Methods
Rats (N=72) were anesthetized with 2% isoflurane with one eye cannulated, connected to a reservoir filled with balanced salt solution and exposed (CEI model) to 8 hours of either 60 mmHg (60-8) or 20 mmHg IOP (20-8). Rats were sacrificed either immediately (0 survival) or after 3 days (3 day survival) and both ONH (cannulated and uncannulated fellow) processed for qPCR (≥8/group). A separate group of totally untreated ONH (Naive) were similarly analysed for baseline controls (100% values). Expression levels were normalized to the Naïve group and statistically compared to determine the effect of anesthesia, cannulation and IOP elevation on gene expression for mRNAs representing cell type markers, Il6-type cytokine signaling, extracellular matrix adhesion and cell proliferation. The latter groups were previously identified to respond maximally to early IOP-induced injury in both chronic model and initial CEI experiments.
Results
Tonolab readings (mean±sd), taken every 30 minutes, confirmed IOP targets: (60-8, 57.2±2.2 mmHg; 20-8, 22.1±2.3 mmHg; all uncannulated fellows, 8.3±1.1 mmHg). Historical mean TonoLab IOP readings in unanesthetized animals were 21.6±1.5 mmHg (N=20). For 60-8 ONH, IL6 and LiF messages (Table) were significantly elevated immediately after IOP exposure, and then normalized. Top2a, indicating cell proliferation, was elevated at 3 days. Axonal marker, Nefl, was significantly reduced at 3 days, suggesting either axonal injury or depressed axonal transport. Other cell type markers, as well as Epo (indicating ischemia), were not significantly affected. IMPORTANTLY, anesthesia alone ONH (uncannulated fellows, not shown in table) and cannulated ONH with IOP 20 mmHg (20-8), equivalent to normal unanesthetized IOP, experienced no significant alterations in any message level compared to Naive ONH.
Conclusions
Gene expression changes in key pressure-induced markers were consistent with our previous chronic model and CEI observations. The lack of significant effects of anesthesia or cannulation confirms that previously observed message responses are due to IOP elevation in the CEI model.
Keywords: 568 intraocular pressure •
430 astrocytes: optic nerve head •
637 pathology: experimental