Different reasons could account for the different patterns of loss in RPC perfusion density and RNFLT in the EG eyes examined in this study. First, it is quite possible that RNFL axons have different susceptibilities to changes in vascular perfusion in different eyes. Second, despite our best efforts to consistently image the same retinal regions, it is also possible that alterations in RPC perfusion occurred outside of the regions that were imaged at a given timepoint or outside of the annulus analyzed in this work. Furthermore, these observed patterns were influenced by our method for determining the timepoint of first change (i.e., the time when a given parameter first fell outside of its confidence interval limits
and remained outside of these limits). While the focus of this study was on early experimental glaucoma, it could have been possible for an ONH or retinal parameter to have rebounded back within the confidence interval in an EG eye if each experiment continued fully to “end stage.” We believe the likelihood for this occurrence is low for the vast majority of eyes as 7 of 9 EG eyes experienced significant neuropathy (i.e., had a mean percentage loss of RNFLT between 13% and 62%) by the end of the experiment. In addition, with the exception of one animal that was followed for 14 days after the first change in RPC density (OHT-87), all other animals were followed for anywhere from 49 to 295 days after the first change in RPC density, and no rebound was observed. In addition, different results could have been found if we only considered the timepoint at which a given parameter first fell outside of the confidence interval and did not require that the parameter remain outside of the interval. For example, RPC perfusion density measured in the superotemporal sector within OHT 81 (
Fig. 4j) first fell outside of its confidence interval at day 14, but rebounded back within the confidence interval at a later timepoint (day 70) before dropping back outside and remaining outside of the confidence interval at day 105. Due to our more conservative criterion (i.e., requiring the parameter to also remain outside of the confidence interval), the reported time of the first change was 105 days. However, without this criterion, the initial change would have occurred at day 14, which would have been before the first measured change in RNFLT in the superotemporal sector (
Fig. 4i) at day 49. The rebound in mean ALCSD, mean MRW, and RPC perfusion density observed in the superotemporal sector of the EG eye at day 70 could likely have been due to a lower pre-imaging session IOP (18 mm Hg) at this time point relative to the previous session (38 mm Hg), possibly resulting in an associated hysteresis in structural measures. In addition, it could be plausible that the capillaries imaged at later timepoints contained both RPCs and deeper capillaries that were subsequently visible due to losses in overlying RPCs. Although great effort was taken to ensure that only the most superficial capillaries were imaged at each timepoint, it is possible that the most superficial capillary network that was visible at later timepoints in an EG eye with a reduced RNFL could have been a deeper capillary plexus. Also, it is important to recall that perfusion images only show vessels that are actively perfused. If capillaries are nonperfused because of an ischemic event or blockage upstream of the capillary blood supply that later resolves, capillaries could reperfuse and be imaged at a later time.
60 Decreased autoregulation of the capillary network could also result in transient perfusion decreases. It has been shown (using Doppler laser flowmetry) that there is decreased autoregulation of blood flow in the major vasculature in the ONH in eyes with glaucoma.
61 Some eyes could be more susceptible to decreased vascular autoregulation and vascular instabilities could be more prominent in these eyes, leading to alterations in perfusion over time. In particular, patients with normal tension glaucoma could have more perfusion alterations as many vascular risk factors have been associated with normal tension glaucoma.
5,6,9,10,62,63 It is important to note that a pressure-based model of experimental glaucoma was used in this study. Although we know of no non-human primate models of normal tension glaucoma, further research could examine RPC perfusion over time in human patients with normal-tension glaucoma.