Version History
This page contains a detailed description of how the Nested RF Protocols have evolved over time. It includes detailed descriptions of the contents of the different protocol versions, a timeline of changes and tabular comparisons of the different parameters used in each version.
Descriptions of Protocol 1 Versions
P1 protocol (Dec 2024 - January 2025) - RNAi experiments
- Area of arena presented: Left half of the arena screen (LHS). 17:112 pixels in x (width) and 1:48 pixels in y (height). [96 x 48 pixels area]
- Background intensity: (6/15) - gs_val = 4.
- Number of repetitions: 4
- Flash sizes: 12 x 12 pixels and 6 x 6 pixels
- Flash contrasts: both ON (15/15) and OFF (0/15)
- Flash duration: 200ms
- Flash overlap: 0% (non-overlapping grid)
- Interval between flashes: 50ms
- Location:
protocols/LHSwithin thenested_RF_stimulusGitHub repository. - Protocol:
protocol1_10kHz_4reps_12px_6px_LHS_2sbkg_200msfl_50msint_12-13-24_14-33-03
NoteResults file:
“Spring2025_RNAi_Protocol2_Analysis”. Data: 2024_12_12 to 2025_01_23. See the file “RNAi_exp_log”.
P1 protocol (April 2025 - May 2025) - TmY3, TmY18, TmY5a, Tm5Y
- Area of arena presented: Right half of the arena screen (LHS). 97:196 pixels in x (width) and 1:48 pixels in y (height). [96 x 48 pixels area]
- Background intensity: (6/15) - gs_val = 4.
- Number of repetitions: 4
- Flash sizes: 12 x 12 pixels and 6 x 6 pixels
- Flash contrasts: both ON (15/15) and OFF (0/15)
- Flash duration: 200ms
- Flash overlap: 0% (non-overlapping grid)
- Interval between flashes: 50ms
- Location:
protocols/RHSwithin thenested_RF_stimulusGitHub repository. - Protocol:
protocol1_10kHz_4reps_12px_6px_RHS_2sbkg_200msfl_50msint_04-08-25_08-08-42andprotocol1_10kHz_4reps_12px_6px_RHS2_2sbkg_200msfl_50msint_81_180_05-05-25_16-18-66
NoteResults file:
“Exp_Log_NestedRF_v2”. Data: 2025_04_08 to 2025_05_09
P1 protocol (July 2025 - Nov 2025) - T4/T5 experiments
- Area of arena presented: Left half of the arena screen (LHS). 17:112 pixels in x (width) and 1:48 pixels in y (height). [96 x 48 pixels area]
- Background intensity: (4/15) - gs_val = 4.
- Number of repetitions: 4
- Flash sizes: 12 x 12 pixels and 6 x 6 pixels
- Flash contrasts: both ON (15/15) and OFF (0/15)
- Flash duration: 200ms
- Flash overlap: 0% (non-overlapping grid)
- Interval between flashes: 150ms
- Location:
protocols/bkg4/LHSwithin thenested_RF_stimulusGitHub repository. - Protocol:
protocol1_10kHz_4reps_12px_6px_LHS_2sbkg_200msfl_150msint_07-10-25_15-45-60
NoteResults file:
“Summer2025_MARCM_p2_analysis” and “Autumn2025_MARCM_p2_analysis”. Data: 2025_07_12 to 2025_11_13
P1 protocol (Sep 2025 - Dec 2025) - pharmacology experiments
- Area of arena presented: Left half of the arena screen (LHS). 17:112 pixels in x (width) and 1:48 pixels in y (height). [96 x 48 pixels area]
- Background intensity: (4/15) - gs_val = 4.
- Number of repetitions: 4
- Flash sizes: 6 x 6 pixels
- Flash contrasts: both ON (15/15) and OFF (0/15)
- Flash duration: 200ms
- Flash overlap: 0% (non-overlapping grid)
- Interval between flashes: 150ms
- Location:
protocols/bkg4/LHSwithin thenested_RF_stimulusGitHub repository. - Protocol:
protocol1_10kHz_4reps_6pxONLY_LHS_2sbkg_200msfl_150msint_11-14-25_13-47-30
NoteResults file:
“Autumn2025_T4T5_Pharma”. Data: 2025_11_18 to 2025_12_22
Parameter Overview
Protocol 1 Parameter Comparison
Side-by-side comparison of all Protocol 1 versions, highlighting the key parameter differences.
| Parameter | Early Testing | RNAi (Dec-Jan) | DS (Apr-May) | T4/T5 (Jul-Nov) | Pharmacology (Nov-Dec) |
|---|---|---|---|---|---|
| Date range | Nov 27 - Dec 12, 2024 | Dec 13, 2024 - Jan 23, 2025 | Apr 8 - May 9, 2025 | Jul 10 - Nov 2025 | Nov 14 - Dec 2025 |
| Screen side | LHS | LHS | RHS | LHS | LHS |
| Pixel range (x) | 17:112 | 17:112 | 97:196/81:180 | 17:112 | 17:112 |
| Flash sizes | 12px + 6px | 12px + 6px | 12px + 6px | 12px + 6px | 6px only |
| Flash duration | 200ms | 200ms | 200ms | 200ms | 200ms |
| Inter-flash interval | 50ms | 50ms | 50ms | 150ms | 150ms |
| Background intensity | gs_val=4 (6/15) | gs_val=4 (6/15) | gs_val=4 (6/15) | gs_val=4 (4/15) | gs_val=4 (4/15) |
| Flash contrasts | ON + OFF | ON + OFF | ON + OFF | ON + OFF | ON + OFF |
| Repetitions | 4 | 4 | 4 | 4 | 4 |
| Sampling rate | 20 kHz | 10 kHz | 10 kHz | 10 kHz | 10 kHz |
ImportantKey Protocol 1 Changes Over Time
- Dec 2024: Changed from 20kHz to 10kHz sampling rate
- Apr 2025: Switched from LHS to RHS for DS Probe experiments
- Jul 2025: Increased inter-flash interval from 50ms to 150ms; changed background from 6/15 to 4/15
- Nov 2025: Removed 12px flashes for pharmacology experiments (6px only)
Descriptions of Protocol 2 Versions
P2 protocol (Dec 2024 - Jan 2025) - RNAi experiments
- Area of arena presented: [30 x 30 pixel area]
- Background intensity: (6/15) - gs_val = 4.
- Number of repetitions: 3
- Only ONE contrast - either ON (15/15) or OFF (0/15)
- Square flashes presented on 50% overlapping grid.
Sequence of stimuli in the protocol:
- 4 pixel square flashes - 160ms duration - 340ms interval
- 4 pixel square flashes - 80ms duration - 170ms interval
- 4 pixel wide bar sweeps - 28 dps
- 4 pixel wide bar sweeps - 56 dps
NoteResults file:
“Spring2025_RNAi_Protocol2_Analysis”. Data: 2024_12_12 to 2025_01_23. See the file “RNAi_exp_log”.
P2 protocol (April 2025 - May 2025) - TmY3, TmY18, TmY5a, Tm5Y
- Area of arena presented: [30 x 30 pixel area]
- Background intensity: (6/15) - gs_val = 4.
- Number of repetitions: 3
- Only ONE contrast - either ON (15/15) or OFF (0/15)
- Square flashes presented on 50% overlapping grid.
Sequence of stimuli in the protocol:
- 10s background at the start and between repetitions (added 2025_05_06 onwards)
- 4 pixel square flashes - 160ms duration - 340ms interval
- 4 pixel square flashes - 80ms duration - 170ms interval
- 4 pixel wide bar sweeps - 28 dps
- 4 pixel wide bar sweeps - 56 dps
NoteResults file:
“Exp_Log_NestedRF_v2”. Data: 2025_04_08 to 2025_05_09
P2 protocol (July 2025 - Sep 2025) - T4/T5 experiments - SUMMER
- Area of arena presented: [30 x 30 pixel area] The 6 x 6 pixel flashes are presented in a 33 x 33 pixel square area - larger than the 30 x 30 pixel square area for the 4 x 4 pixel flashes - to get an even number of flashes on a 50% overlapping grid.
- Background intensity: (4/15) - gs_val = 4.
- Number of repetitions: 3
- Only ONE contrast - either ON (15/15) or OFF (0/15)
- Square flashes presented on 50% overlapping grid.
- 1s interval between each bar sweep
Sequence of stimuli in the protocol:
- 10s background at the start and between repetitions
- 4 pixel square flashes - 160ms duration - 440ms interval
- 6 pixel square flashes - 160ms duration - 440ms interval
- 4 pixel wide bar sweeps - 28 dps
- 4 pixel wide bar sweeps - 56 dps
NoteResults file:
“Summer2025_MARCM_p2_analysis”. Data: 2025_07_12 to 2025_09_12
P2 protocol (Oct 2025 - Nov 2025) - T4/T5 experiments - AUTUMN
- Area of arena presented: [30 x 30 pixel area] The 6 x 6 pixel flashes are presented in a 33 x 33 pixel square area - larger than the 30 x 30 pixel square area for the 4 x 4 pixel flashes - to get an even number of flashes on a 50% overlapping grid.
- Background intensity: (4/15) - gs_val = 4.
- Number of repetitions: 3
- Only ONE contrast - either ON (15/15) or OFF (0/15)
- Square flashes presented on 50% overlapping grid.
- 1s interval between each bar sweep
Sequence of stimuli in the protocol:
- 10s background at the start and between repetitions
- 3s background interval between square flashes and sweeps, between sweep speeds and between bar flash speeds. (Added 2025_10_21).
- 4 pixel square flashes - 160ms duration - 440ms interval
- 6 pixel square flashes - 160ms duration - 440ms interval
- 4 pixel wide bar sweeps - 28 dps
- 4 pixel wide bar sweeps - 56 dps
- 4 pixel wide bar sweeps - 168 dps
- 4 pixel wide bar flashes - 80ms duration - 920ms interval
- 4 pixel wide bar flashes - 14ms duration - 486ms interval
The bar flashes are presented in different random orders for each repetition. The bar flash stimuli contains 11 different positions. From 2025_10_15 - 2025_10_23, the “central” bar flash is positioned in the middle of the 30 x 30 pixel area, then the 5 positions either side are this bar moved by 1 pixel increments and decrements from this position in the 8 different orientations. From 2024_10_23 onwards, these 5 other positions are the central bar moved by 2 pixel increments and decrements.
NoteResults file:
“Autumn2025_MARCM_p2_analysis”. Data: 2025_10_15 to 2025_11_17
P2 protocol (Nov 2025 - Dec 2025) - T4/T5 pharmacology experiments
- Area of arena presented: [30 x 30 pixel area]
- Background intensity: (4/15) - gs_val = 4.
- Number of repetitions: 3
- Only ONE contrast - either ON (15/15) or OFF (0/15)
- 1s interval between each bar sweep
- NO SQUARE PIXEL FLASHES PRESENTED IN P2.
Sequence of stimuli in the protocol:
- 10s background at the start and between repetitions
- 3s background interval between sweep speeds and between bar flash speeds.
- 4 pixel wide bar sweeps - 28 dps
- 4 pixel wide bar sweeps - 56 dps
- 4 pixel wide bar sweeps - 168 dps
- 4 pixel wide bar sweeps - 250 dps
- 4 pixel wide bar sweeps - 500 dps
- 4 pixel wide bar flashes - 80ms duration - 920ms interval
- 4 pixel wide bar flashes - 14ms duration - 486ms interval
There are 88 bar flash stimuli. 8 different orientations. Bar centred on the 30 x 30 pixel square and then moved by 2 pixel steps in either direction along this axis.
NoteResults file:
“Autumn2025_T4T5_Pharma”. Data: 2025_11_18 to 2025_12_22
Protocol 2 Parameter Comparison
Side-by-side comparison of all Protocol 2 versions.
| Parameter | RNAi | DS Probe | T4/T5 Summer | T4/T5 Autumn | Pharmacology |
|---|---|---|---|---|---|
| Date range | Dec 12, 2024 - Jan 23, 2025 | Apr 8 - May 9, 2025 | Jul 12 - Sep 12, 2025 | Oct 15 - Nov 17, 2025 | Nov 18 - Dec 22, 2025 |
| Background intensity | 6 | 6 | 4 | 4 | 4 |
| Repetitions | 3 | 3 | 3 | 3 | 3 |
| Stimulus area | 30x30 px | 30x30 px | 30x30 px (33x33 for 6px) | 30x30 px (33x33 for 6px) | 30x30 px |
| Contrast | ON or OFF | ON or OFF | ON or OFF | ON or OFF | ON or OFF |
Square Flash Parameters
| Parameter | RNAi | DS Probe | T4/T5 Summer | T4/T5 Autumn | Pharmacology |
|---|---|---|---|---|---|
| 4px flashes | Yes | Yes | Yes | Yes | No |
| 4px duration | 160ms, 80ms | 160ms, 80ms | 160ms | 160ms | - |
| 4px interval | 340ms, 170ms | 340ms, 170ms | 440ms | 440ms | - |
| 6px flashes | No | No | Yes | Yes | No |
| 6px duration | - | - | 160ms | 160ms | - |
| 6px interval | - | - | 440ms | 440ms | - |
Bar Sweep Parameters
| Parameter | RNAi | DS Probe | T4/T5 Summer | T4/T5 Autumn | Pharmacology |
|---|---|---|---|---|---|
| 14 dps | No | No | No | No | No |
| 28 dps | Yes | Yes | Yes | Yes | Yes |
| 56 dps | Yes | Yes | Yes | Yes | Yes |
| 168 dps | No | No | No | Yes | Yes |
| 250 dps | No | No | No | No | Yes |
| 500 dps | No | No | No | No | Yes |
| Inter-sweep interval | - | - | 1s | 1s | 1s |
Bar Flash Parameters
| Parameter | RNAi | DS Probe | T4/T5 Summer | T4/T5 Autumn | Pharmacology |
|---|---|---|---|---|---|
| 80ms bar flashes | - | - | - | Yes (920ms interval) | Yes (920ms interval) |
| 14ms bar flashes | - | - | - | Yes (486ms interval) | Yes (486ms interval) |
| Number of positions | - | - | - | 11 | 11 |
| Position spacing | - | - | - | 1px (Oct 15-23), 2px (Oct 23+) | 2px |
Timing Parameters
| Parameter | RNAi | DS Probe | T4/T5 Summer | T4/T5 Autumn | Pharmacology |
|---|---|---|---|---|---|
| 10s background at start | No | Yes (from May 6) | Yes | Yes | Yes |
| 10s background between reps | No | Yes (from May 6) | Yes | Yes | Yes |
| 3s background between stimuli | No | No | No | Yes (from Oct 21) | Yes |
ImportantKey Protocol 2 Changes Over Time
- May 6th 2025: Added 10s background intervals between repetitions amd at the start of the protocol.
- July 2025: Changed the background intensity from 6/15 to 4/15.
- Oct 2025: Added 168 dps sweeps and bar flashes (80ms and 14ms) to the protocol.
- Oct 2025: Introduced a 3s background interval between stimulus blocks for easier data parsing.
- Oct 23rd 2025: Changed bar flash position spacing from 1px to 2px increments.
- Nov 2025: Removed square flashes for pharmacology experiments, focusing solely on bar sweeps and flashes. Added 2 faster speeds.
Timeline Overview
The following table provides a chronological overview of all protocol versions used, linking dates to experiment types and data locations.
Protocol 1 Timeline
| Date Range | Experiment Type | Key Changes | Data Location |
|---|---|---|---|
| Nov 27 - Dec 12, 2024 | Early testing | Initial version, 20kHz | protocol1/LHS/protocol1_4reps_12px_6px_LHS_2sbkg _200msfl_50msint_11-27-24_15-02-95/ |
| Dec 13, 2024 - Jan 23, 2025 | RNAi experiments | 10kHz sampling, 50ms interval, LHS | protocol1/LHS/protocol1_10kHz_4reps_12px_6px_LHS_2sbkg_ 200msfl_50msint_12-13-24_14-33-03/ |
| Apr 8 - May 4, 2025 | DS Probe (TmY3, TmY18, TmY5a, Tm5Y) | Switched to RHS | protocol1/RHS/protocol1_10kHz_4reps_12px_6px_RHS_2sbkg_ 200msfl_50msint_04-08-25_08-08-42/ |
| May 5 - May 9, 2025 | DS Probe (continued) | RHS v2 (pixels 81-180) shifted to more central part of arena | protocol1/RHS/protocol1_10kHz_4reps_12px_6px_RHS2_2sbkg_ 200msfl_50msint_81_180_05-05-25_16-18-66/ |
| Jul 10 - Nov 2025 | T4/T5 MARCM | 150ms interval, bkg4 | protocol1/bkg4/protocol1_10kHz_4reps_12px_6px_LHS_2sbkg_ 200msfl_150msint_07-10-25_15-45-60/ |
| Nov 14 - Dec 2025 | Pharmacology | 6px flashes only | protocol1/bkg4/protocol1_10kHz_4reps_6pxONLY_LHS_2sbkg_ 200msfl_150msint_11-14-25_13-47-30/ |
Protocol 2 Timeline
| Date Range | Experiment Type | Key Changes | Data Location |
|---|---|---|---|
| Dec 12, 2024 - Jan 23, 2025 | RNAi experiments | 160ms/80ms flashes, 28/56 dps sweeps | protocol2/data/RNAi_ttl_2025/ |
| Apr 8 - May 5, 2025 | DS Probe | Same as RNAi version | protocol2/data/DS_v2/ |
| May 6 - May 9, 2025 | DS Probe | Added 10s background between reps | protocol2/data/DS_v2/ |
| Jul 12 - Sep 12, 2025 | MARCM summer | Added 6px flashes, 440ms intervals | protocol2/data/T4T5_Summer2025/ |
| Oct 15 - Nov 17, 2025 | MARCM autumn | Added 168 dps sweeps, bar flashes | protocol2/data/T4T5_autumn2025/ |
| Nov 18 - Dec 22, 2025 | Pharmacology | No square flashes, 5 sweep speeds | protocol2/data/Octopamine/, protocol2/data/CGP54626/ |
NoteHow to verify the version if unsure
For definitive confirmation, check the metadata.mat or currentExp.mat files within the experiment folder, which contain the exact parameters used.