Ultrasonic Vocalizations during Intermittent Swim Stress predict proficiency

Ultrasonic Vocalizations during Intermittent Swim Stress predict proficiency

Ultrasonic Vocalizations during Intermittent
Swim Stress predict proficiency in Spatial Learning.

Van Hoogenstyn,
and Rob
Laura Castracane,
Drugan Department, University of New Hampshire, Durham, NH 03820, USA
Psychology Department, University of New Hampshire, Durham, NH 03820, USA


Depression is among the top five leading cause
of disability and disease burden throughout the world 1.
Untreated depression is the leading cause of suicide,
which validates ongoing research to better understand
the disorder2. Of the 121 million suffering world wide
from depression2, 30% have a treatment-resistant form
of the illness6. New preclinical research needs to
discover alternative pharmacotherapy to help those
people. The historical focus is on the depressionvulnerable group, but focusing on the more resilient
population of depression is a better approach..
Understanding the neurochemical correlates of
resilience may lead to novel insight into the
development of new pharmalogical treatments.
Human depressed patients suffer from spatial
learning and memory deficits, which are associated
with the hippocampus7. In previous studies, high stress
levels translated to enhanced hippocampal cell loss and
spatial memory deficits2. Consequently, those who are
vulnerable to depression may undergo hippocampal
change greater than a resilient subject.
There are many animal models of depression.
The exteroceptive model is more representative of the
human condition, in not assuming pathology.
Increasing evidence has shown that fundamental
mechanisms of neuronal change are disrupted in
animal models of stress, supporting the symptomology
of stress3. The Intermittent swim stress model of
depression has been shown to be a ecologically
relevant stressor3. The stress has been observed to
cause depression-like deficits in behavior. During the
ISS, some rats have been observed to emit ultrasonic
vocalizations 4. In a preliminary report using ISSinduced USVs of rats predicted whether they would be
resilient or vulnerable, based on their learning in a
subsequent swim escape task 4.
This study extended previous research that
concluded ISS-induced USVs forecast resilience in
instrumental learning.
The aim of the current study wass to investigate if ISS
induced USVs predict good performance in a
subsequent spatial learning
and memory task.



80 trials of ISS did not alter
spatial learning in the MWM.
However, there are individual
differences in USVs.

If High USVs during the ISS
procedure predict or forecasts
resilience, then we would expect
high USV calling rats to perform
better in the Morris Water maze
than the rats who do not call.

Of the 10 rats exposed to
stress (ISS), 6 rats vocalized,
the remaining four did not.

Day 1 Stress

Rats were randomly assigned to one of three groups: ISS
(n=10) or confined control (n=10)


ISS rats were exposed to 80 trials. Confined controls
were placed in the stress apparatus for the same 80
trials, but without water.


USVs were recorded during the ISS with a ultrasonic
microphone and the Labview data collecting program,
which allowed frequency and duration of calls to be

Figure 1: A line graph showing no significant difference
between the spatial learning scores the Control (non
stressed) and ISS group (stressed). 2 trial blocks were
plotted on the x-axis and time to find platform in
seconds was plotted on the y-axis.

Rats underwent 18 learning trials presented in 9 blocks
of two trials each.


Rats were placed in different cardinal directions (N, S, E,
W) for each trial and latency to find a hidden platform
was recorded. If the rat failed to find the platform in 60
seconds the animal was guided to the platform. Every rat
was left on the platform for 10 seconds at the conclusion
of each trial.


However, high vocalizers had
exceptional scores, showing
their resilience to stress.


1.Caspi, A., Sugden ,K., Moffitt T., Taylor A., et al (2003) Influence of life Stress on
Depression: Moderation by Polymorphism in the 5-HTT Gene. Science 301, 386392

Twenty-four hours after ISS, all rats were trained in the
Morris water maze in pairs by the experimenter blind to
group membership.


Stressed animals that did not
vocalize performed poorly in
spatial learning, requiring a
longer time to find the hidden

This verifies the high callers
brain needs to be looked at
more closely to find better
pharmalogical treatments.

Day 2Spatial


2.Castro J. E., Diessler, J., Varea E., Mrquez, C., Larson, M.H., (2011).Personality
traits in rats predict vulnerability and resilience to developing stress-induced
depression-like behaviors, HPA axis hyper-reactivity and brain changes in
pERK1/2 activity. Psychoneuroendocrinology, vol. 37, num. 8, p. 1209-23, 2012.

Figure 2: A scatter plot characterizing the USVs of the
rats in the ISS. Plotted are the number of USVs on the xaixs and average duration of vocalizations on the y-axis,
in ms, for each individual.

3.Christianson, J.P., Drugan, R.C. (2005). Intermittent cold water swim stress increases
immobility and interferes with escape performance in rat. Behavioural Brain
Research, 165,58-62
4.Drugan, R.C., Christianson, J.P., Stine,W.W., Soucy,D,P.(2009) Swim stress induced
ultrasonic vocalizations forecast resilience in rats, Behavioural Brain Research,
202, 142-145
5. Drugan,R.C., Skolnick, P, Paul.S., Crawley,J.N. (1989) A pretest procedure reliably
predicts performance in two animal models of inescapable stress. Pharmacology
Biochemistry and Behavior 33(3) 649-654

20 male Sprague-Dawley rats (CD Derived, Charles River Labs, NY),
each weighing 180-200 grams on arrival. Animals were housed for at
least 6 days before experimentation four to a cage with free access to
food and water. Following day 1 of procedures animals were
individually housed with food and water ad lib.

6. Krishnan, V. and Nestler, E.J., (2010) Linking Molecules to Mood: New Insight Into
the Biology of Depression. American Journal of Psychiatry;167:1305-1320

ISS Apparatus
ISS treatments were administered in Plexiglas cylinders (21 X 42 cm
DIA. X H.) with 0.64cm wire mesh on the bottom, suspended over a
tank of 15Cwater. On a swim trial, the cylinders were lowered into
the water to a depth of 20 cm. Space heaters blew warm air ( 36C)
into the cylinders during inter-trial-intervals. The swim stress
apparatus was controlled by a computer with Med-PC hardware and
software (Med-Associates, VT). The ultrasonic vocalizations were
recorded with a microphone and data was collected with the
program Labview.

Morris Water Maze (MWM) Apparatus
Spatial learning tests were conducted in a circular pool (122 cm in
diameter) filled with water to a depth of approximately 51 cm. Water
was made opaque with approximately 200 ml of white nontoxic liquid
tempera paint. The water temperature was maintained at 24C (+/1C). The hidden platform was painted white and constructed out of
PVC pipe and clear Plexiglas, with a diameter of 10 cm and a height
of approximately 50 cm. On each wall of the room different distinct
symbols were placed for orientation during procedures on day 2.
Some learning trials and all memory probes were recoded using
video camera from above, an HVS image tracking system and
computer analyzed the data.

ISS apparatus (Above Left)
Stress animals are lowered into water by the cylinders, confined are
lowered without water. All other conditions remain identical.

~Thank you Robert Drugan, Nate
Stafford, and Tim Warner for all of
your patience and guidance in the lab.

Morris Water Maze (MWM) (Top Right)
Picture of rat on hidden platform.
Diagram of MWM During A Hypothetical Learning Trial
(Lower Right)
Animals are placed at the outside of the pool at different cardinal
directions (N, S, E, W). Animal search for a hidden platform during
learning trials or are left to swim freely for 60 seconds during the
memory probe test.

Figure 3: A line graph showing rats who vocalized
during the ISS had significantly better scores than the
rats who did not emit vocalizations. Plotted are the 2
trial intervals on the x-axis and mean time to locate
platform on the y-axis.

~Funding was provided from Mr. Dana
Hamel through the Hamel Centers
~ All behavioral procedures were
reviewed and approved by the UNH
institutional Animal Care and Use
committee (IACUC).

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