The results of the HB test show difference between the CO2, and Air and N2. However, under a high concentration of CO2, the larvae cease cardiac activity.. The average time for the movement ending under high concentration of CO2 is 57 sec. Even under a

1
THE EFFECTS OF CO2 AND HYPOXIA ON THE PHYSICAL
BEHAVIOR AND HEART RATE IN DROSOPHILA
LARVAE NICOLAS H. BADRE AND ROBIN L.
COOPER DEPARTMENT OF BIOLOGY, UNIVERSITY OF
KENTUCKY, LEXINGTON, KY 40506-0225

• CONCLUSION
• Under a high concentration of N2, the larvae have
  a decreased locomotion which seems to increase
  with time.
• Under a high concentration of CO2, the larvae
  have a even stronger decrease in locomotion which
  leads to a stopping of movements.
• Under a high concentration of N2, the larvae does
  not react by stopping its BWM.
• Under a high concentration of CO2, the larvae
  cease movement in less than minute.
• Under a high concentration of N2 or with air, the
  larvae do not have any kind of cardiac arrest.
• Under a high concentration of CO2, the larvae
  cease cardiac activity.
• The larvae needs a recovery time close to a
  minute to regain cardiac activity after a being
  in an environment high in CO2.
• CO2 provokes to the larvae behaviors, which are
  different to the one caused by hypoxia and, which
  are similar to the one of anesthesia.
• In 67 of the CO2 exposure, the larvae will go
  in shell position before stopping its cardiac
  activity.

Heart Beats
RESULTS Body Wall Movements
INTRODUCTION AND BACKGROUND Carbon dioxide is
commonly used as an anesthesia for adult
Drosophila melanogaster, however, the mechanism
of its actions is unknown. This is important, as
it might lead to the discovery of new types of
insecticides that would be innocuous to plants
and plant eaters. Since mosquitoes have been
shown to have sensory structures that detect
carbon dioxide, we postulated that Drosophila
must also contain similar types of receptors
because they share the same kind of environment.
Laval insects have never been examined for carbon
dioxide sensory neurons. Previous experiment
supposed that carbon dioxide affected larvae the
same way than humans a increase in body fluid
acidity causing different behaviors such as
anesthesia (Sillans and Biston, 1979). Those
experiments also showed that carbon dioxide had
different effects to hypoxia as a high
concentration of carbon dioxide and oxygen could
also cause anesthesia (Sillians et al., 1969).
However, this current research has an objective
to find sensory neurons on the larvae capable of
detecting the CO2.
The results of the HB test show difference
between the CO2, and Air and N2. However, under a
high concentration of CO2, the larvae cease
cardiac activity.. The average time for the
movement ending under high concentration of CO2
is 57 sec. Even under a high concentration of N2
or with air, the heart beat does not stop as with
CO2. Another figure determined by this
experiment is the time needed for the larvae to
recover its cardiac activity once the CO2
injection is stopped and that the container is
opened. The average for the recovery time of the
larvae is 59.6 sec. (This figure is only
available for the CO2 as the larvae would only
stop their cardiac activity under high
concentrations of CO2).
METHODOLOGY We tested Canton S, the common
wild-type laboratory strain of Drosophila
melanogaster. This experiment focused on larvae
at the beginning of the wandering phase of the
third instar. Many of the techniques used in this
experiment have already used in Cooper and
Neckameyer (1999). Each larva was in a sealed
agar plate with carbon dioxide injected into the
container. Part I Body wall movements (bwm)
Heart Beats (HB) We injected CO2 in the sealed
container for a period of 10 minutes, after which
the container was opened. We recorded the bwm for
the first and last two minutes. If at any time
bwm or the HB stopped, the time would be
recorded. If the HB stopped, the time when the HB
started again once the container was open, would
be recorded. The objective of this test is to
quantify the difference between CO2 and hypoxia
in the larvae using common features of the
animal. Part II The reaction of the larvae
with the CO2 In our effort to identify particular
characteristics of the larval response to carbon
dioxide, we designed several terms to quantify
those responses. Shell position designates larvae
which are in a curved position. Elongated
position designates larvae which are flaccid and
which look longer than usual. Contracted position
designated larvae which had returned to their
normal shape after being in elongated position.
Those response were tested by placing the larvae
under anesthesia for approximately 5 minutes and
recording the different behaviors of the larvae
during the first minutes and the minutes
following the end of the CO2 injection. This test
has for objective to understand and detail the
reaction of the larvae to CO2. We repeated the
experiment with N2 to make sure that the results
were specific to CO2. We also had a control,
recording the natural bwm and HB of the larvae,
without the injection of any gas. Here is a
model of the experiment. CO2 and N2 are
represented on the left and the control on the
right.
The results of those bwm tests show difference in
all three conditions. The normal (air) average
number of body wall movements for the first two
minutes is 76 bwm. However, with the N2, the
results were twice smaller with an average of
35.4 bwm for the first two minutes. Furthermore,
with CO2, the results were very small with an
average of 10 bwm which is 7 times smaller. The
normal (air) average number of body wall
movements for the last two minutes is 98.2 bwm.
However, with the N2, the results were five times
smaller with an average of 19.4 bwm for the first
two minutes. Furthermore, with CO2, the larvae
stopped moving so the average for the last two
minutes is 0 bwm. The trend is that, under a
high concentration of N2, the larvae have a
decreased locomotion which seems to increase with
time. For CO2, the result seem comparable to N2,
except that the effects are stronger as the
results are smaller for the first two minutes and
inexistent for the last two.
Physical Behavior
Current Research Objectives Determining where
the CO2 receptors are located on the larvae by
inhibiting certain part of the body at detecting
the presence of CO2. Understanding which parts
of the larvae are still active/responding while
the animal is under anesthesia. Finding out how
the larvae knows when to stop its anesthesia mode
to return to a normal activity. Explaining the
benefit of having such quick responses to
CO2. Recording the activity of the sensory
neurons by electrophysiology.
shell position
elongated position
contracted position
The physical behavior test showed that in 67 of
time, the larvae would go in shell position
before stopping its cardiac activity. During the
other cases, the larvae would directly go in
elongated position. In all of the trials (100),
the larvae would go in elongated position once
the HB was stopped. When the injection of CO2 was
stopped, the larvae would have many different
behaviors which did appears in the same order
every time. However, slight head turns and mouth
hooks movements were very common behaviors in the
first three to four minutes. It took on average
4.63 minutes for the larvae to gain back their
common size contracted position. Once the larvae
is in contracted position, it usually does a
couple of non-motional bwm. Before starting its
natural motion, the larvae often turns its head
left and right and then starts its motion. The
average time for the larvae to come back to a
normal behavior after five minutes of exposition
to CO2 is 8.8 minutes.
REFERENCES Biston J, Sillans D (1979) Studies on
the anesthetic mechanism of carbon dioxide by
using Bombyx mori larvae. Biochimie 61,
nº2153-156 Sillans D, Esteve J, Legay JM (1969)
C.R. Acad. Sci., 269 1209-1212 Cooper RL,
Neckameyer WS (1999) Dopaminergic neuromodulation
of motor neuron activity and neuromuscular
function in Drosophila melanogaster. Comp
Biochem Physiol B 122199-210.
The results of this bwm test show difference for
the CO2. Under a high concentration of N2, the
larvae does not react by stopping its bwm.
However, under a high concentration of CO2, the
larvae cease movement in less than minute. The
average time for the movement ending under high
concentration of CO2 is 40 sec.