1. Screen for neurons involved in innate olfactory
attraction behavior in Drosophila
Methods
Pavel Morales, Emily Leung, Daniel Jong, Susy M. Kim, Hui-hao Lin, Jing W. Wang
University of California, San Diego
Conclusions
X102
The ability to make decisions is dependent on assigning attractive and
aversive values to different stimuli. Flies show robust innate attraction and
aversion behaviors to different odors. In the antennal lobe, specific
glomeruli are hardwired to behavioral olfactory attraction and aversion.
What additional neural circuits in the brain make one like or dislike a
particular odor? We addressed this question using a high throughput
olfactory four-field paradigm, which quantifies odor attraction in fruit
flies. Candidate GAL4 driver lines that targeted different regions of the
Drosophila brain were silenced to see if any were critical for behavioral
attraction.
We measured the fruit flies’ odor attraction by using a high throughput four-
field olfactometer paradigm (Figure 1). Vinegar odor is introduced to one
quadrant, while air is run through the other 3 quadrants. The path each fly
took is recorded and collected.
Data
Sample traces (Figure 2) depicting fly responses to no odor(left) and vinegar
(right).
We silenced neurons labeled by selected drivers with a temperature
sensitive shibirets2 transgene. Flies were tested at the non-permissive
temperature (32⁰C). From this analysis, only two lines exhibited
significant differences in behavioral attraction . The performance indices
of all drivers tested are shown as a bar graph in Figure 3.
PrGal4
Traces depicting fly movements in the four field olfactometer. Responses
to vinegar odor in for X102/Shibirets flies (Figure 9a), Or42b/ Shibirets
flies (Figure 9b, our positive control group), Shibirets/ + flies (Figure 9c,
our negative control group).
The X102 Gal4 driver strongly labels
neurons in the lateral horn with weaker
expression in other neuropils such as the
ventrolateral protocerbrum (see arrow and
Z-stack projection of X102 Gal4 driving
GFP, Figure 7).
Data
We verified Gal4 drivers that emerged from the
screen by testing them with additional controls.
The Pr-Gal4 driver labels neurons in multiple
neuropils including the lateral horn and
ventrolateral protocerbrum (see arows and Z-stack
projection of Pr-GAL4 driving GFP in the
Drosophila brain. (Figure 4.).
Vinegar
We silenced neurons labeled by the Pr-GAL4
driver using a UAS-Shibire(ts2) transgene.
Flies were tested at both the permissive
temperature (25⁰C) and heat shock conditions
(32⁰C).
The bar graph (Figure 5) summarizes
performance indices for both control and
experimental groups at the permissive and non-
permissive temperatures.
We silenced neurons labeled by X102
driver using a UAS-Shibirets2 transgene.
Flies were tested at the non-permissive
temperature (32⁰C).
The bar graph (Figure 8) summarizes
performance indices for both control and
experimental groups at the non-permissive
temperature.
No-odor
Four-Field Olfactometer
Figure1.
Figure 2.
Screen ing for Neurons involved in Behavioral Attraction
Traces depicting fly movements in the four field olfactometer are
displayed below. Responses to vinegar odor in control Pr-Gal4/+
(Figure 6a) and Pr-Gal4/Shi (Figure 6b) flies are compared in side-by-
side.
Figure5.
Figure 6a. Figure 6b.
Figure8.
Figure 9a. Figure 9b. Figure 9c.
Figure4.
It is of interest that both X102 and Pr-Gal4 drivers target neurons in the
lateral horn, a brain region theorized to mediate innate olfactory
behaviors. Thus, our data supports this popular theory.
Introduction
Figure7.
We evaluated the odor attraction ability in flies by measuring the path and
amount of time it takes for flies stay in the odor quadrant. The performance
index is defined as (2p1/2 – 1) x 100%, in which p is the fraction of time the fly
spends in the odor quadrant between 0 and 60 s after odor application.
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PerformanceIndex
X102
Or42b
Shibire