This study considers the role that multiple, brief IOP insults can have on retinal neurons. The effect of one, two, and four 15-minute IOP insults on photoreceptor amplitude (Rm
P3 ) was not severe enough to differentiate the insults from each other
(Fig. 2) . In contrast, the postreceptor components (P2 and nSTR) were better indicators of multiple challenge, consistent with a greater susceptibility of the inner retina to IOP elevation compared with the outer retina.
1 2 31 32
Functional recovery after IOP insults becomes slower as the number of insults increases
(Figs. 3 4) . This outcome provides in vivo evidence that repeated IOP challenge temporarily exacerbates neuronal dysfunction with recovery. Anatomic damage has been reported after repeated pressure insult in an ex vivo model.
8 In contrast, Kim et al.
33 did not find any cumulative b-wave dysfunction after three IOP episodes. This negative outcome may arise because the b-wave (P2) shows less cumulative compromise compared with our STR measure.
Figure 8Ashows that functional recovery becomes further delayed with more insults. However, this effect is less pronounced for the P2 (unfilled) than the nSTR (filled;
P < 0.05). Alternatively, Kim et al.
33 used a longer (1 hour) interval between IOP challenges, which allowed full b-wave recovery before subsequent insults. Our short intervals (10 minutes) provided only partial b-wave recovery before the next challenge
(Fig. 3C) . Short intervals may place greater stress on cellular reserves needed for recovery, thereby fostering cumulative dysfunction. Consistent with this possibility, our data for two insults
(Fig. 3B) , which had a longer interval and allowed full recovery, showed no cumulative effect
(Fig. 3A) .
It is unclear whether the IOP-related dysfunction observed in this study occurred through ischemic or mechanical processes. It is thought that both factors may occur simultaneously at high IOP because studies
34 35 have shown that blood vessels and axoplasmic flow are compromised by IOP-induced deformation of the lamina cribrosa. According to the ischemic theory, ocular perfusion becomes compromised when IOP reaches a level that disrupts normal retinal blood flow, producing an ischemic challenge.
36 On the other hand, high pressure may act directly on retinal neurons. Stretch-activated channels on ganglion cells have been shown to open in response to mechanical force (e.g., IOP), triggering potassium efflux.
37 Moreover, mechanical stretching of neuronal membranes can activate N-methyl d-aspartate receptors,
38 which, in turn, increases intracellular calcium. Although calcium influx can lead to neuronal apoptosis in longstanding or severe IOP elevation,
39 we did not find any sustained functional loss (
Fig. 1 , bottom row). We postulate that exposure to four 15-minute insults is too brief to induce apoptosis. Specifically, any changes in intracellular potassium and calcium may be too low to induce apoptosis but still high enough to dampen cellular excitability and glutamate release. Such a mechanism has been recently described in cultured midbrain dopaminergic neurons after 5 minutes of ischemia.
40 This mechanism will minimize the possibility for glutamate excitotoxicity at the expense of neurotransmission, thus manifesting as a transient functional deficit.