To asses any possible effects of stressful conditions experienced by the parental generation (G0) on generation 1 (G1) flies on day 0 of the assay the air volume for the 60 experimental flies was reduced by ca. 1/7 by positioning the cotton plug ~ 10mm above the food surface. For the 30 flies of the control group the cotton plug was not positioned differently. Crowding took place 48h on 25°C. On day 2 the control flies were transferred into a big vial with fresh yeast. The experimental group was anesthetized with CO2 and separated to the usual number of flies per vial, 20 females and 10 males, and also transferred into a big vial with fresh yeast. After 24h all the flies in the vials were discarded. On day 12 newly hatched G1 flies were collected and incubated for 24h at 25°C before the ATR treatment.

To differentiate the effects on learning behaviour of G1 flies that experienced stress but their parents have not been stressed (group 1), G1 flies whose parents have been stressed and also experienced stress themselves (group 2), G1 flies that have never experienced stress wether their parents nor themselves (group 3) and G1 flies whose parents have been stressed but they haven´t experienced stress (group 4), following steps were taken.

For the ATR treatment, on day 13, four small vials without yeast were prepard with ATR. The collected males from the experimental and the control group, were again separated into 2 different vials each. the 4 different vials represent the following four groups:

Group_1: control group; stressed

Group_2: experimental group; stressed

Group_3: control group; not stressed (Positive control)

Group_4: experimental group; not stressed

After again 48h incubation at 25°C all 4 groups were prepared and tested in the Joystick.

Joystick experiments

The following are the pooled results of seven days of testing over the last two months.