Elevated Pressure Induces TRPV1-Mediated Ca2+-Dependent Apoptosis of Retinal Ganglion Cells

Abstract

Purpose:: Vision loss in glaucoma is associated with sensitivity to intraocular pressure and the death of retinal ganglion cells (RGCs). Loss of RGCs in glaucoma suggests they could express an intrinsic mechanism for mediating sensitivity to pressure. We asked whether transient receptor potential vanilloid-1 receptor (TRPV1) could mediate this sensitivity, because of its role in mechanosensation and calcium (Ca²⁺)-dependent death in other neurons.

Methods:: We examined expression of TRPV1 by RGCs in whole rat retina using PCR, ISH, ICC and western blot. We measured pressure-induced changes in TRPV1 expression in RGCs isolated from rat retina by immuno-magnetic separation and exposed to elevated hydrostatic pressure. We compared survival of RGCs at ambient or elevated pressure following treatment with the TRPV1 agonist capsaicin and the TRPV1 antagonist iodo-resinferatoxin (IDX). To measure pressure-induced changes in the accumulation of intracellular Ca²⁺ caused by elevated pressure and TRPV1 activation, we quantified changes in the Ca²⁺ dye Fluo-4.

Results:: TRPV1 localization in the RGC axon and dendrites appears in node-like clusters. This pattern was similar in primary cultures of RGCs, where 24 hours of elevated pressure induced a 16-fold increase in TRPV1 expression that preceded a 35% loss of RGCs. Treatment with various doses of IDX (10mM-100pM), diminished pressure-induced loss of RGCs and TUNEL reactivity by as much as 30% and 52%, respectively. In contrast, various doses of capsaicin (10mM-100nM) at ambient pressure mimicked the effects of elevated pressure, decreasing RGC density by as much as 50% and increasing TUNEL reactivity by 2-fold. Exposure to elevated pressure for just one hour increased the intensity of Fluo-4 label in cultured RGCs by 4-fold that was reduced by 40% following treatment with 10nM IDX. Treatment with 1mM capsaicin for one hour at ambient pressure also induced an increase in Ca²⁺ accumulation (2.5-fold) that was reduced by 62% with IDX treatment.

Conclusions:: Our data suggest that activation of TRPV1 likely contributes to the susceptibility of RGCs to pressure-induced apoptosis through an increase in intracellular Ca²⁺.

Keywords: apoptosis/cell death • ganglion cells • calcium