Purpose
Our previous studies have shown altered basal blood flow in the ONH to mildly increase during the early stages of EG and progressively decline with increasing axon loss, as measured by retinal nerve fiber layer thickness (RNFLT). We hypothesized that one of the cellular components underlying hemodynamic changes within the retrolaminar of the NHP optic nerve head (ONH) are astrocytes, whose processes encasing the blood vessels are known to modulate neurovascular coupling. Since, AQP4, a protein that is predominantly localized to astrocyte end-feet has been shown to regulate cerebral blood flow and calcium signaling, we quantified expression of AQP4 during different stages of EG.
Methods
Unilateral elevation of IOP was induced in 11 adult rhesus macaques (NHPs) by laser treatment to the trabecular meshwork. Retinal nerve fiber layer thickness (RNFLT) was measured by spectral-domain optical coherence tomography. With the resultant RNFLT % used to define early (RNFLT below 20% of baseline; n=6) from advanced (RNFLT greater than 20% of baseline; n=5) EG animals. Sagittal cryosections of the ONH from the NHPs were prepared for immunofluorescence and stained with an AQP4 antibody and imaged with a fluorescent microscope. Images were imported into Image J (NIH) for quantification analysis.
Results
In control, AQP4 presents as thread-like labeling parallel to the optic nerve bundles through-out the length of the retrolaminar (fig. 1A). The early EG samples show a labeling pattern similar to control, however, with a 44% reduction in pixel intensity compared to control (fig. 1B; P=0.04). The advanced EG display a labeling pattern that is distinct to that found in either the control or the early EG samples (fig. 1C). Further, this distinct labeling pattern is observable throughout the retrolaminar, with a 63% AQP4 expression increase over control (P=0.12).
Conclusions
In EG based on RNFLT loss, a two-phase pattern of AQP4 presentation within the ONH retrolaminar was observed. Specifically, AQP4 expression decreased 44% during the earliest EG stage (RNFLT <20% of baseline) followed by an increase of 63% compared to control in the advanced EG (RNFLT >20% of baseline). The specific role of AQP4 and the reason for these distinct expression changes on blood flow remain under investigation.
Keywords: 430 astrocytes: optic nerve head •
554 immunohistochemistry •
436 blood supply