Decreased Reactive Oxygen Species Signaling Alters Glutamate Receptor Transport to Synapses in C. elegans AVA Neurons

Abstract

Reactive oxygen species (ROS) are chemically reactive molecules normally produced during cellular respiration. High ROS levels negatively impact forms of synaptic plasticity that rely on changes in the number of ionotropic glutamate receptors (iGluRs) at synapses. More recently, we have shown that physiological increases in ROS reduce iGluR transport to synapses by acting on activity-dependent calcium signaling. Here, we show that decreasing mitochondria-derived ROS decrease iGluR transport albeit in a calcium-independent manner. These data demonstrate differential regulatory mechanisms by elevated or diminished ROS levels which further support a physiological signaling role for ROS in regulating iGluR transport to synapses.

Figure 1. Diminished ROS levels decrease GLR-1 transport out of the cell body via a calcium-independent mechanism.

A) Representative images showing expression of contents of transgenic arrays used. Scale bar = 5 µm. B) GLR-1::mCherry transport. Top: Single time points from 50 s of continuous imaging (100 ms/frame). Two GLR-1-containing vesicles (blue and green numbers) are pointed out for three timepoints. Bottom: Kymograph derived from 30 s of a 50 s image stream depicting position (x-axis) of GLR-1 transport vesicles over time (y-axis). Scale bar = 5 µm. C and E) Representative kymographs depicting GLR-1 transport vesicles (black lines) as they are transported through the length of the neurite (x-axis) over time (y-axis). Scale bar = 5 µm. D) Quantification of transport events in control ( lin-15(n765ts) X; glr-1(ky176) III; csfEx63 , n=15) and worms overexpressing catalase (CTL-2(OX)) in the AVA ( lin-15(n765ts) X; glr-1(ky176) III; csfEx65 , n=11, **:p=0024, two-tailed Student’s t-test). F) Quantification of transport events from untreated (n=18) and mitoTEMPO treated (n=20) worms ( glr-1(ky176) III; akIs201 , ***:p=0.0008, two-tailed Student’s t-test). G) Representative traces of changes in GCaMP6f fluorescence in the AVA cell body over 60 s in vivo normalized to baseline fluorescence (DF/F _min ) from untreated (n=35) and mitoTEMPO treated (n=35) worms ( lin-15(n765ts) X; csfEx62 ). H) Average DF/F _min normalized to untreated controls. I) Total activity (sum of fluorescence values above baseline divided by baseline) normalized to untreated controls. J) Average baseline of these groups normalized to untreated controls.