ABSTRACT Glaucoma is a leading cause of permanent vision loss worldwide, but the mechanisms of damage are not fully understood. The retinal ganglion cells (RGC) and their axons transmit visual information from the retina to the brain, and these axons pass out of the eye through the scleral canal at the optic nerve head (ONH), which is spanned by a fenestrated connective tissue structure known as the lamina cribrosa (LC). The preponderance of evidence suggests that the RGC axons are damaged in the laminar region of the ONH in glaucoma. One of the most consistent glaucoma risk factors is elevated intraocular pressure (IOP), although the “safe” IOP threshold varies widely among individuals. While there is some evidence that IOP fluctuations contribute to glaucoma, prior studies have been hampered by the absence of continuous IOP measurement. Retrobulbar cerebrospinal fluid pressure (CSFP) surrounding the optic nerve partially counteracts IOP at the LC through the translaminar pressure (TLP=IOP-CSFP). Retrospective clinical studies have suggested that higher CSFP (and low TLP) is protective for glaucoma and low CSFP (and high TLP) increases glaucoma risk, after accounting for the effects of IOP. In addition, since the LC bears the bulk of the pressure load in the ONH due to its high stiffness relative to the surrounding neural tissues, LC thickness plays a critical role in the distribution of TLP in the ONH via the translaminar pressure gradient (TLPG = TLP/LC thickness), adding a morphological component to TLP. Hence, the goal of this project is to test the hypotheses that IOP, TLP, and TLPG fluctuations independently contribute to eye-specific susceptibility to glaucoma onset and progression after accounting for differential mean IOP in fellow eyes, and confirm the recent finding that CSFP and IOP are coupled via neural pathways. In this project, we will perform mechanical compliance testing to quantify LC deformations in vivo in response to controlled acute TLP challenge, in an animal model of unilateral glaucoma instrumented with continuous IOP, CSFP, TLP, and TLPG telemetry. We will then determine the relationships between axonal and visual function loss per unit of differential mean IOP in fellow eyes and 1) transient and diurnal IOP fluctuation, 2) TLP and TLPG (mean and fluctuation) and 3) LC deformations in response to acute TLP challenge, measured while the eye is normal and after glaucoma onset and progression. Impact: If results show that IOP fluctuations, TLP and/or TLPG contribute to glaucoma pathogenesis and progression in addition to mean IOP, new therapeutic approaches could be developed to modulate these factors to treat glaucoma.