Abstract
Purpose :
The calcium (Ca2+) selective transient receptor potential vanilloid 1 (TRPV1) channel is an important integrator of Ca2+ signaling. Ca2+ influences the excitability of retinal ganglion cells (RGCs) in a subtype-selective manner in response to visual stimuli. Downstream, Ca2+ interacts with molecules such as the Ca2+/calmodulin-dependent phosphatase, calcineurin, that help to maintain Ca2+ homeostasis. Calcineurin is associated with neurodegenerative diseases and in an experimental model of glaucoma, calcineurin inhibition affords neuroprotection to RGCs. Here, we aim to show how inhibiting calcineurin directly affects RGC light-evoked responses in C57 wild type (WT) and TRPV1 knockout (Trpv1-/-) mice.
Methods :
Eyes from WT and Trpv1-/- mice were enucleated, and retinas were dissected out in a dark room under red light. Retinas were placed in a physiological chamber and constantly perfused with Ames’ medium with 20mM glucose. Recording pipettes were filled with (in mM): 125 K-gluconate, 10 KCl, 10 HEPES, 10 EGTA, 4 Mg-ATP, 1 Na-GTP, and 0.1 ALEXA 555. Whole-cell current clamp signals were amplified and digitized at a sampling rate of 50kHz. The calcineurin inhibitor, FK506, was bath applied in increasing concentrations from 10nM to 100nM and was thoroughly washed out prior to the application of each subsequent dose.
Results :
Our investigation focused on alpha ON-sustained (αON-S), alpha OFF-sustained (αOFF-S), and alpha OFF-transient (αOFF-T) RGCs. For αON-S RGCs, FK506 did not significantly change the light-evoked response compared to baseline in WT (101%) or Trpv1-/- (87%) mice. In αOFF-S responses decreased with increasing concentrations of FK506. Light-evoked responses following 100nM FK506 were decreased to 19% of baseline in WT (p=0.001) and 34% of baseline in Trpv1-/- (p<0.001) mice. For αOFF-T, we found a significant increase in the light-evoked responses for WT (119%, p=0.03) mice compared to baseline. However, in Trpv1-/- αOFF-T RGCs no change was observed (103%). For all cell types analyzed, FK506 elicited similar responses between WT and Trpv1-/- mice.
Conclusions :
Results presented here highlight how different RGC subtypes utilize Ca2+ signaling to initiate spike outputs in response to light. Modulation of the Ca2+ signaling pathway using calcineurin inhibitor FK506, elicits RGC subtype specific changes in light-evoked responses that is preserved after ablation of functional TRPV1 channels.
This is a 2020 ARVO Annual Meeting abstract.