Discussion: experiment-summary

[Ogul-suray]

Hi all,

I have some questions about the experimental setup and data that I need clarified for my dissertation. Any help would be appreciated.

1. SLAP2 Pixel Resolution
What does each pixel in SLAP2 correspond to anatomically? Specifically:

• Does 1 pixel capture multiple synapses?
• Or do multiple pixels cover a single synapse?

2. Mouse Strain Information
What mouse strains were used across all sessions?
I see that Mouse ID: 794237 is recorded as C57 wild-type, but other mice (e.g., Mouse ID: 796630) have no strain information. Can I assume all sessions for all subjects (mice) used C57 wild-type mice?

3. DFF Measurements — Glutamate vs Calcium
What exactly is the DFF measuring in the glutamate and calcium sessions?
I noticed that the emission wavelength for session SLAP2_776270_2025-07-02_11-50-06 is listed as 510nm (green) (circled in red). Can someone clarify what indicator this corresponds to?

4. Tuning Block Structure and Recording Gaps
Two related questions:
a) Block structure:
For sessions with Mouse ID: 794237, I only see two tuning blocks (TB1 and TB2) with one oddball block between them. Is this the intended structure for these sessions?
b) Recording gaps:
For sub-801381 session 2025-09-26, I found a recording gap from 3180s to 3600s, which falls directly on tuning block 3. Is this expected?
Similarly, for sub-801381 session 2025-10-02, there's a 360-second gap from 3090s to 3450s, meaning tuning block 3 only covers 162 trials.

5. Preferred Orientation Scatter Plot
I wanted to check that my approach for quantifying preferred orientation changes between tuning blocks is appropriate.
The challenge is that orientation data is circular -- e.g., 157 degrees is actually close to 0 degrees (or 180 degrees), but on a standard scatter plot it appears as a large difference.

My current approach:
Circular distance is computed for each ROI using: distance = min(|theta_2 - theta_1|, 180 - |theta_2 - theta_1|). This accounts for periodicity (e.g., 157 degrees vs 2 degrees gives 25 degrees, not 155 degrees). I then report descriptive statistics on these distances -- mean +/- SD if normally distributed (Shapiro-Wilk), or median (IQR) if not. I am not performing a hypothesis test on these distances because the ROIs come from the same neuron of the same mouse, so they are not independent observations. Additionally, OLS linear regression is fitted to TB1 vs TB2 preferred orientations, reporting R-squared and slope as descriptive measures of tuning stability (R-squared close to 1 with slope close to 1 indicates stable tuning). The hypothesis testing will be done at the population level across mice rather than within a single session.

Does this descriptive approach (circular distance with mean +/- SD or median/IQR, plus regression R-squared and slope) seem appropriate for the within-session analysis?

6. Stimulus Type
Are the gratings used in all sessions static or drifting?

7. Grating Coordinate System
Which coordinate system is used for the grating orientations?

"Compass" system: 0° = horizontal bar moving upward is considered to be moving at 0° and angles increase in a clockwise manner
Cartesian system: 0° = vertical bar moving rightward, angles increase counterclockwise

(This is important for interpreting and reporting my results correctly.)

I hope to hear from you soon and thank you in advance for any clarification.

Kind regards,
Ogulsuray