Relationship between AQP5 membrane trafficking and hydrostatic pressure regulation in the rat lens

Abstract

Lens transparency and refractive properties are maintained by the directional movement of ionic and water fluxes generated by the microcirculation system which, in the absence a of blood supply, deliver nutrients, remove waste products, and control the volume of the lens. In this system, Na+ and water enter the lens at the anterior and posterior poles via an extracellular pathway associated with lens sutures, which direct the fluxes towards the lens core. These fluxes then flow to the surface via an intracellular pathway mediated by gap junctions, before exiting the lens at the equator, completing the circulation. In the mouse lens, movement of water through the gap junction pathway generates an intracellular hydrostatic pressure gradient that is tightly regulated by a dual feedback system, initiated by the Transient Receptor Potential Vanilloid (TRPV) channels, TRPV1 and TRPV4. In the lens, water is transported across the membrane via the aquaporins (AQP), AQP1, AQP0 and AQP5. Of these three AQPs, only AQP5 was regulated by translocation. In the rat lens, a large pool of AQP5 channels were observed that could translocate out of the membranes, with reduced zonular tension, in the influx anterior and efflux zones. These discoveries led to my hypothesis that changes of the surface hydrostatic pressure are associated with the trafficking of the AQP5 water channel. To test this hypothesis, I combined surface hydrostatic pressure measurements and immunolabeling experiments of AQP5 expression and applied them to the rat lens. Using a microelectrode-based pressure measurement system, I first confirmed that the surface hydrostatic pressure of the rat lens, like the mouse lens, is regulated by TRPV1 and TRPV4 mechanoreceptors producing a biphasic increase and decrease, respectively, in the surface hydrostatic pressure. In addition, I found that the biphasic increase was abolished in lenses that were pre-treated with a TRPV4 inhibitor followed by the application of a TRPV1 activator, resulting in a sustained increase of surface hydrostatic pressure, and this sustained increase was mimicked by the application of pilocarpine. Using immunolabeling analysis, I found that incubation of rat lenses in TRPV1/4 modulators resulted in a rapid removal of AQP5 from the membranes in fibre cells localised in the anterior influx and efflux zones, and this removal of AQP5 was mimicked by the application of pilocarpine. Taken together, my results suggest that modulation of the water permeability in the influx anterior and efflux zones is mediated by the removal of AQP5 from the membranes, and may regulate the changes of the surface hydrostatic pressure of the lens. My findings present a modified model of the microcirculation system by incorporating the membrane trafficking of AQP5, which is hoped to provide a better understanding of the changes to lens optics that cause the onset of presbyopia in middle age, and cataract in the elderly.