A few months ago I finished my first screen for DA-neuronal populations relevant in the photopreference shift. I tried a small collection of TH-Gal4s with interesting results that you could see here. To validate those results, I am currently searching for other Gal4s labeling the same groups of neurons. Here you can see the outcome of the first experiments for one of those lines.

Trying to define subsets of neurons related to the photopreference shift, I started to test the different OA and DA drivers available. Here you can see two OA-GAL4s used in Burke et al 2012:

My results:

The following plot shows the photopreference of two LexA lines presumably labeling TbH neurons. Only wing-clipped flies are shown.

26-09-2016, new experiments added.

In order to study the role of OA and DA in photopreference I am constantly looking for new drivers that label subpopulations of these groups. I recently found a TbH driver from the QF system, and wanted to know how representative of the TDC2-G4 neurons was. I have established a QUAS-mCherry;TbH-QFs line (Magenta) and crossed it with TDC2-G4;UAS-GFP (Green). In both cases, what is shown is the endogenous expression.

This is an update of the T-maze experiments for flies with OA and DA neurons manipulated.

1. TH-G4s x shi^TS.  Except for 2 groups the N is at least 5.

2. TbH-lexAs x shi^TS.  Last year I tried to complete this experiment and several problems appear. Now I am starting to do it again and it seams that it works. N=4.

3. I am starting to test the same TbH lines crossed to TrpA1. So far, not much to say.

After finishing the first round of the screen to find out the DA substrate of the photopreference, I decided to change the genetic background of the GAL4s and UAS. Now all of them are in WTB background.

Here are the first results. It is only an n of 1.

Update:

As a control for my T-Maze experiments I decided to test if treated flies were able to fly at the same conditions they were tested in the T-Maze (32°C).

As in the T-Maze experiments, I put the flies 15 min at 32°C before testing them at the same temperature. The, I measure the duration of the first spontaeous flight (Blue). If within 5min the fly doesn’t start to flight, I gently remove the steroform ball from the legs (Red), simulating a takeoff.

Details from 0 s to 200 s.

As a control for my T-Maze experiments I decided to test if treated flies were able to fly at the same conditions they were tested in the T-Maze (32°C).

As in the T-Maze experiments, I put the flies 15 min at 32°C before testing them at the same temperature.

The dashed line at 11 min indicates when a T-Maze experiment would have ended.

Almost all flies tested flew more than the time required to perform a T-Maze experiment.

In previous experiments, I found one TβH(lexA)>shi^TS  combination that recapitulated TDC2>shi^TS T-Maze results (https://lab.brembs.net/2015/09/looking-for-the-da-oa-neurons-involved-in-phototactic-flexibility/). Here I present the expression pattern of those two TβH-lexA drivers used.

TβH⁵⁴⁹⁵⁴-LexA (Brain)

TβH⁵⁴⁹⁵⁴-LexA (VNC)

TβH⁵⁴⁰⁷⁵-LexA (Brain & VNC)

Last week I continued the experiments for the optogenetics. These are my results:

and these are Lena’s results:

Compared to Lena’s results, my results are a bit too positive.