Delayed post-ischemic conditioning significantly improves the outcome after retinal ischemia

Abstract

In previous studies, it was shown that post-conditioning, a transient period of brief ischemia following prolonged severe ischemia in the retina, could provide significant improvement in post-ischemic recovery, attenuation of cell loss, and decreased apoptosis. These studies showed that post-conditioning effectively prevented damage after retinal ischemia when it was instituted early (within 1 h) in the post-ischemic period. While post-ischemic conditioning holds high promise of clinical translation, patients often present late after the onset of retinal ischemia and therefore immediate application of this anti-ischemic maneuver is generally not feasible. In this study, we examined the hypothesis that application of a post-conditioning stimulus at 24 h or greater following the end of prolonged ischemia would decrease the extent of ischemic injury. Ischemia was induced in rat retina in vivo. Recovery after ischemia followed by 5 min of post-conditioning brief ischemia 24 or 48 h after prolonged ischemia was assessed functionally (electroretinography) and histologically at 7 days after ischemia and post-conditioning or sham post-conditioning. We found that the brief ischemic stimulus applied 24, but not 48 h after prolonged ischemia significantly improved functional recovery and decreased histological damage induced by prolonged ischemia. We conclude that within a defined time window, delayed post-ischemic conditioning ameliorated post-ischemic injury in rats. Compared to earlier studies, the present work demonstrates for the first time the novel ability of a significantly delayed ischemic stimulus to provide robust neuroprotection in the retina following ischemia.

Figure 1

Stimulus-Intensity response in post-conditioned retinae for 24 h post-C vs 24 h sham post-C. Double normalized (corrected for the non-ischemic eye and for diurnal variation from baseline to 7 days after post-conditioning) ERG data for a-, b-wave, oscillatory potentials (sum of root mean square, OP RMS) and P2 over a range of flash intensities from −1.02 to 1.40 log cd-s/m². The data were recorded at baseline (prior to ischemia) and at 7 days after post-conditioning; post-C was applied 24 h after ischemia. ERG data for a- (A), b-wave (B), OP RMS(C) and P2 (D) over a range of flash intensities suggests that 24 h delayed post-C was functionally protective. Solid lines with diamonds = 24 h sham post-C (n = 11). Solid lines with squares = 24 h post-C (n = 15).

Figure 2

Stimulus-Intensity response in post-conditioned retinae for 24 h post-C vs 24 h sham post-C. Absolute ERG data for a-, b-wave, oscillatory potentials (sum of root mean square, OP RMS) and P2 over a range of flash intensities from −1.02 to 1.40 log cd-s/m². The data were recorded at baseline (prior to ischemia) and at 7 days after post- conditioning; post-C was applied 24 h after ischemia. ERG data for a- (A), b-wave (B), OP RMS(C) and P2 (D) over a range of flash intensities suggests that 24 h delayed post-C was functionally protective. Solid lines with diamonds = 24 h sham post-C (n = 11). Solid lines with squares = 24 h post-C (n = 15).

Figure 3

(A) Representative ERG traces at 0.87 log cd.s/m² flash intensity for the a- and b-wave (top), P2 (middle) and the OPs (bottom) for both 24 h post-C and 24 h sham post-C groups. (B) Representative ERG traces at 0.87 log cd.s/m² flash intensity for the a- and b-wave (top), P2 (middle) and the OPs (bottom) for both 48 h post-C and 48 h sham post-C groups.

Figure 4

Representative histopathological images of hematoxylin and eosin-stained retinae in 4 µm thick sections for each of the experimental groups. These sections were prepared from retinae removed from the rats at 7 days following delayed post-C. Arrows indicate layers demonstrating cell loss and asterisks denote regions of inflammatory cell infiltration. Below each representative retina is a close-up of the RGC layer.

Figure 5

Stimulus-Intensity response in post-conditioned retinae for 48 h post-C vs 48 h sham post-C. Double normalized (corrected for the non-ischemic eye and for diurnal variation from baseline to 7 days after post-C) ERG data for a-, b-wave, oscillatory potentials (sum of root mean square, OP RMS) and P2 over a range of flash intensities from −1.02 to 1.40 log cd-s/m². The data were recorded at baseline (prior to ischemia) and at 7 days after post-conditioning; post-C was applied 48 h after ischemia. ERG data for a- (A), b-wave (B), OP RMS(C) and P2 (D) over a range of flash intensities suggests that 48 h delayed post-C was not functionally protective. Solid lines with diamonds = 48 h sham post-C (n = 4). Solid lines with squares = 48 h post-C (n = 4).

Figure 6

Stimulus-Intensity response in post-conditioned retinae for 48 h post-C vs 48 h sham post-C. Absolute ERG data for a-, b-wave, oscillatory potentials (sum of root mean square, OP RMS) and P2 over a range of flash intensities from −1.02 to 1.40 log cd-s/m². The data were recorded at baseline (prior to ischemia) and at 7 days after post-conditioning; post-C was applied 48 h after ischemia. ERG data for a- (A), b-wave (B), OP RMS(C) and P2 (D) over a range of flash intensities suggests that 48 h delayed post-C was not functionally protective. Solid lines with diamonds = 48 h sham post-C (n = 4). Solid lines with squares = 48 h post-C (n = 4).