DOI: 10.5061/dryad.j9kd51crg
title: Photopic flicker optoretinography captures the light-driven length modulation of photoreceptors during phototransduction

description: In this study, we used an inhibitor of phosphodiesterase 6 (PDE6) to
 examine the impact of changes in the conformation of the PDE6 protein on
 the light-induced process responsible for altering the length of the outer
 segments of photoreceptor cells in both human and rodent eyes. We employed
 a imaging method called spatiotemporal optical coherence tomography, which
 ensures high contrast and phase stability within the strongly scattering
 photoreceptor– Retinal Pigment Epithelium complex. Using this approach, we
 recorded nanometer-scale changes in human cones and rods in response to
 photopic flicker stimulation and observed length changes in rodent rods
 under scotopic conditions following a single pulse of light, in the
 absence or presence of sildenafil, which inhibits the catalytic activity
 of PDE6. Our findings are consistent with the interpretation that during
 phototransduction conformational changes in PDE6 structure, which occur on
 an angstrom scale, are amplified to the nanometer scale due to the unique
 structure of the photoreceptor outer segments and sequential stimulation.
 This finding opens up possibilities for the informed use of photopic
 flicker optoretinography measurements as a diagnostic tool, as the
 observed nanometer-scale changes in rod and cone dimensions as a function
 of light stimulus can now be directly linked to molecular events involved
 in the phototransduction pathway.
description: # Photopic flicker optoretinography captures the light-driven length
 modulation of photoreceptors during phototransduction **Folder:
 Figure2.zip contains 1 file:** **Figure2_data.mat** -Variable named
 "experimentalResults" contains structs with measurement
 descriptions and results (cell named “ORG”). "ORG" cell
 contains, ORG response (nm), high-pass filtered response (nm), amplitude
 spectrum (nm), frequency-axis (Hz), time-axis (s), and finally amplitude
 (nm) and phase of the spectrum at stimulus frequency. -Variable
 "t" contains the time axis (s). -Variables "ORG*"
 contain recorded ORG signals (nm) at given stimulus values. -Variables
 "*Comp*" contain extracted components ("slow",
 "fast", "flicker") with values given in nm at
 different stimulus luminances.   **Folder: Figure3.zip contains 1 file:**
 **Figure3_data.mat** -Variables named "subject (1-6)" contain
 measurement results for different subjects. In each of the structs, there
 are measurement descriptions and results (cell named “ORG”). The “ORG”
 cell contains, response (nm), high-pass filtered response (nm), amplitude
 spectrum (nm), frequency-axis (Hz), time-axis (s), and finally amplitude
 (nm) and phase of the spectrum at stimulus frequency. -Variable named
 "frequency_characteristics" contains six rows representing each
 subject. Two columns contain frequency values (Hz) and average amplitudes
 of ORG signal (nm) for each subject respectively. -Variable
 "dataFit" contains fitted components.   **Folder: Figure4.zip
 contains 1 file:** **Figure4_data.mat** -Variables named
 "subject(1-6)" contain measurement results for different
 subjects. Inside each of the subject's results are cells representing
 different depths with respect to COST (with value given from -8px to 8px
 with 2px step). Each of these cells contains structs with measurement
 descriptions and results (in a similar way to Figure 2 and Figure 3).
 -Variable named "frequency_characteristics" contains six rows
 representing each subject. Two columns contain frequency values (Hz) and
 average amplitudes of ORG signal (nm) for each subject respectively. There
 are nine cells for each subject as the signal was calculated at 9 depths
 (-8px to 8px with 2px step).   **Folder: Figure5.zip contains 14 files:**
 Files related to the manuscript Fig. 5(C) and 5(D). More specifically, it
 contains the following files: -         
 Figure5_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForAllMice.fig: Matlab figure file plotting individual single pulse optoretinography response curves - ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) versus time from stimulus (s) – from a total of 48 datasets from five albino mice, of which 24 before (in red) and 24 after (in cyan) Sildenafil administration. -          Figure5_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForMouse1.fig: Matlab figure file plotting individual single pulse optoretinography response curves - ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) versus time from stimulus (s) – from a total of 10 datasets from albino mouse #1, of which 4 before (in red) and 6 after (in cyan) Sildenafil administration. Thicker plot lines highlight measurements taken at least 2 days after the thinner plot lines. -          Figure5_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForMouse2.fig: Matlab figure file plotting individual single pulse optoretinography response curves - ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) versus time from stimulus (s) – from a total of 12 datasets from albino mouse #2, of which 6 before (in red) and 6 after (in cyan) Sildenafil administration. Thicker plot lines highlight measurements taken at least 2 days after the thinner plot lines. -          Figure5_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForMouse3.fig: Matlab figure file plotting individual single pulse optoretinography response curves - ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) versus time from stimulus (s) – from a total of 11 datasets from albino mouse #3, of which 5 before (in red) and 6 after (in cyan) Sildenafil administration. Thicker plot lines highlight measurements taken at least 2 days after the thinner plot lines. -          Figure5_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForMouse4.fig: Matlab figure file plotting individual single pulse optoretinography response curves - ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) versus time from stimulus (s) – from a total of 5 datasets from albino mouse #4, of which 3 before (in red) and 2 after (in cyan) Sildenafil administration. -          Figure5_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForMouse5.fig: Matlab figure file plotting individual single pulse optoretinography response curves - ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) versus time from stimulus (s) – from a total of 10 datasets from albino mouse #5, of which 6 before (in red) and 4 after (in cyan) Sildenafil administration. Thicker plot lines highlight measurements taken at least 2 days after the thinner plot lines. -          Figure5_BoxPlot_DeltaROSspeed_Before_And_AfterSildenafil_ForAllMice_ExcludingDatasets7-44-45-51.fig: Matlab figure file showing a Box plot (obtained with the boxplot Matlab function) of the ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image004.png) from 44 of the 48 selected datasets from the five albino mice, before (in red) and after (in cyan) Sildenafil administration. This figure corresponds to the Manuscript Figure 5(D). -          Figure5_DataForFig5.mat: Matlab file, including three variables (matrices): o    TimeFromStimulusIn_s: matrix of time vectors (each containing 290 datapoints) for each of the 48 datasets (48x290 double), representing the time from the stimulus in seconds, *i.e.*, the x axis for each of the DeltaROS_vs_time figures. o      DeltaROSin_nm: matrix of vectors (each containing 290 datapoints) representing ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image006.png) values in nanometers, *i.e.*, the y axis, for each corresponding time point of the previous matrix, for each of the 48 datasets (48x290 double). o    oplchangevstimevstimesincesildenafilv6: matrix presenting a lookup table between the first column, representing the series index, and the second column, representing the dataset index. This variable is used by the following Matlab script to display the data as in: All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForAllMice.fig -       Figure5\_ AllGreenDay4toDay9: Excel spreadsheet showing ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image008.png)(D2:D59) and other reliability metrics for all 58 datasets that provided good STOC-T image quality -       Figure5\_ SelectedDay4toDay9Excl38to43: Excel spreadsheet showing: ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image008.png)(E2:E49) and other reliability metrics for the 48 datasets that yielded viable ORG signals with properly suppressed breathing artifacts. -       Figure5\_ SelDay4to9Excl38to43&7,44,45,51: Excel spreadsheet showing ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image008.png)(E2:E45) and other reliability metrics for the 44 datasets where the ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image008.png)estimation was reliable. -       Figure5\_ Selected Day4to9Excl38to43Plots: Excel spreadsheet showing ![](file:///C:/Users/grazy/AppData/Local/Temp/msohtmlclip1/01/clip_image002.png) (G2:IV49) versus time from stimulus (G1:IV1) for the 48 datasets that yielded viable ORG signals with properly suppressed breathing artifacts. Unlike the matrix DeltaROSin_nm in the DataForFig5.mat Matlab file, the number of time points in the spreadsheet was limited to 250, instead of 290   -          Figure5_MatalbScript_for_All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForAllMice.m: Matlab script to display the data as in: All_Selected_DeltaROS_vs_time_Before_And_AfterSildenafil_ForAllMice.fig. It requires access to DataForFig5.mat -          Figure5_MeanAndStd_DeltaROS_vs_time_Before_And_AfterSildenafil_ForAllMice.fig: Matlab figure showing rod-mediated, single-pulse, scotopic ORG recordings before (red) and after (cyan) Sildenafil administration. The centerline represents the mean and the shaded area represents +/- one standard deviation. This figure corresponds to the Manuscript Figure 5(C).   **Folder: Figure6.zip contains 5 files:** \- Figure6_A_VEPS: Excel spreadsheet contains (rows 2-97) the averaged responses in μV (VEP) for each mouse at time 0 minutes (control without Sildenafil) and every 30 minutes up to 210 minutes after Sildenafil injection. Time points without and after sildenafil administration (0 min, 30 min, 60 min, 90 min, 120 min, 150 min, 180 min and 210 min) are provided in column A. For each timepoint (0 min, 30 min, 60 min, 90 min, 120 min, 150 min, 180 min and 210 min) timings of responses measurements are provided in seconds in row 1 and ranges from -0.296 s to 1 s. \- Figure6_B: Excel spreadsheet for panel B contains the response amplitudes in μV for each mouse and electrode recorded every 30 minutes. Time points without and after sildenafil administration (0 min, 30 min, 60 min, 90 min, 120 min, 150 min, 180 min and 210 min) are provided in row 1.  - Figure6_C: Excel spreadsheet for panel C contains the ORG slopes and VEP Amplitudes data without sildenafil (VEP 0 and ORG 0) and 90 min after sildenafil injections (VEP 90 and ORG 90). VEP Amplitudes are in μV; ORG slopes are in mm/s. \- Figure6_D: Excel spreadsheet for panel D, contains the data of ORG slopes and VEP Amplitudes as a percentage of control recordings. ORG slopes and VEP Amplitudes data without sildenafil (VEP 0 and ORG 0) and 90 min after sildenafil injections (VEP 90 and ORG 90). \- Figure6_E: Excel spreadsheet for panel E. Column A, Amplitude Drop in %; Column B, ORG slope in %.