The document summarizes a study that investigated whether neurodegeneration alone can cause schizophrenia-like behaviors in mice. The study ablated neural stem cells in transgenic mice using ganciclovir over 56 days. Behavioral tests revealed no significant differences between control and ablated mice in prepulse inhibition, locomotor activity after PCP/amphetamine, time spent in open arms of elevated plus maze, or floating behavior in Porsolt swim test. This suggests neurodegeneration alone may not cause schizophrenia. However, other studies that also applied stressors found ablation did produce schizophrenia-like behaviors, so stress may be a contributing factor.

1. Redetermining the Role of Neurodegeneration in Schizophrenia
Megan Burrey
UCSB Summer Sessions Research Mentorship Program 2012
and
San Marcos High School, Santa Barbara, CA
Mentor: Chris Knight
Department of Psychology
University of California, Santa Barbara
Abstract
Schizophrenia is a psychiatric disorder associated with symptoms such as hallucinations
and pathologies in the structure of the brain. Research indicates that antipsychotic treatment
reverses these abnormalities, potentially by stimulating neurogenesis. This suggests that
decreases in neurogenesis might be involved in the expression of schizophrenic-like behaviors.
To investigate this possibility, the neural stem cells were ablated in mice by administration of
ganciclovir for fifty-six days. Results revealed no significant differences between control and
ablated mice when tested for schizophrenic-like behaviors through four tests: the elevated plus
maze, Porsolt swim test, prepulse inhibition test, and PCP- or amphetamine-induced locomotor
activity. These results suggest that neurodegeneration alone may not be the sole cause of
schizophrenia.
Introduction
Schizophrenia is a chronic psychiatric disorder that affects approximately 1% of
Americans (NIMH, 2009). The symptoms of schizophrenia fall into three categories: positive,
negative, and cognitive. Positive symptoms are unusual thoughts or perceptions and include
hallucinations, delusions, and movement disorders. Negative symptoms, such as lack of interest

2. and speaking very little, are characteristics of the disease that take away from everyday life.
Cognitive symptoms include poor decision-making skills, the inability to focus, and memory
deficits. Additionally, many schizophrenics experience depression and anxiety which contributes
to the 10% lifetime incidence of suicide in schizophrenia (David A. Lewis, 2000).
Research regarding the biological aspects of schizophrenia has identified structural brain
abnormalities in schizophrenic patients. These include increased ventricular volume, decreased
hippocampal volume, and decreased whole brain volume (Wright et al., 2000). It has been
reported that these abnormalities can be reversed by antipsychotic drug treatment. However, the
nature of these effects remains unclear (Chakos et al., 1998). Reversals of the increased
ventricular volume, decreased hippocampal and whole brain volume in schizophrenic patients
has previously been attributed to changes in cell volume. The recent discovery, however,
established that the central nervous system possesses the ability to produce new neurons
throughout adulthood and has led researchers to address and examine the possible role of
neurogenesis in the etiology and treatment of psychiatric disorders such as schizophrenia
(Malberg et al., 2000).
A study led by Iwata (2008) investigated if radiation exposure, which ablates neural stem
cells, leads to changes in animal behaviors relevant to schizophrenia. To test its hypothesis, the
Iwata lab ablated the neural stem cells in rats through irradiation and observed the rats’ behaviors
post-irradiation. Neurochemical, behavioral and immunohistochemical studies were performed
three months after irradiation. After three months, the total numbers of BrdU-positive cells in
both the subventricular zone (SVZ) and subgranular zone (SGZ) zones of irradiated rats were
60% lower than those of control rats. Behavioral abnormalities such as social interaction deficits,
working memory deficits, and auditory sensory gating deficits were observed in the irradiated

3. rats (Iwata, 2008). These results suggest that irradiation in adulthood causes behavioral
abnormalities relevant to schizophrenia, and that reduction of adult neurogenesis by irradiation
may be associated with schizophrenia-like behaviors in rats.
The Hayashi lab (2008) studied how high levels of stress during adolescence affects
one’s susceptibility to developing psychiatric disorders in adulthood. Hayashi and his colleagues
examined the role of postnatal neurogenesis during adolescence, a period between three to eight
weeks of age, in rodents to restrict postnatal neurogenesis in the hippocampus. Electrical
footshock stress was given to the mice at eight weeks old. X-irradiated mice subjected to
electrical footshock stress during adolescence expressed decreased locomotor activity in the
novel open field, and exhibited prepulse inhibition deficits in adulthood (Hayashi, 2008). This
suggests that mice with unusually high levels of stress during adolescence, which leads to
decreased postnatal neurogenesis, are more likely to develop mental disorders as adults.
The results from the Iwata (2008) and Hayashi (2008) experiments demonstrate that the
ablation of neural stem cells in mice and rats causes the rodents to express schizophrenic-like
behaviors. Unlike the Iwata and Hayashi labs, the neural stem cells of our mice will be ablated
with the drug ganciclovir rather than irradiation. Killing the neural stem cells by means of
irradiation causes an inflammatory response that does not occur with transgenic mouse models.
Irradiation kills a fewer neural stem cells than using transgenic mice. When transgenic mice are
bred, at least 90% of the neural stem cells die. When ablation occurs through irradiation, only
60% of the neural stem cells are killed. It is expected that our transgenic mice will exhibit
schizophrenic-like behaviors in four behavioral tests: the elevated plus maze, prepulse inhibition
test, Porsolt swim test, and locomotor activity test.

4. Methods
Animals
Transgenic GFAP-tk mice were generated by breeding hemizygous GFAP-tk female
mice obtained from Jackson Laboratory (Bar Harbor, ME) with nontransgenic males. Mice were
between eight and ten weeks of age at the start of drug or saline treatment.
GCV Infusion
Animals were anesthetized with isoflurane. Canciclovir (.02 mg/kg/d) in physiological
saline or saline for control groups was infused unilaterally for fifty-six days into the lateral
ventricle using a subcutaneously implanted osmotic minipump (Alzet, model 1004) with the
canula implanted at +1.0 A/P, +0.5 M/L, and -2.50 D/V to bregma.
Prepulse Inhibition
To measure sensorimotor gating in mice, a prepulse inhibition test was employed.
Animals were placed in sound attenuated startle chambers (SR-LAB, San Diego Instruments).
The procedure employed was reported previously (Szumlinkski et al., 2005). The startle program
consisted of six trial types: startle pulse (st110, 100 dB/40 milliseconds), low prepulse stimulus
alone (st74, 74 dB/20 milliseconds), high prepulse stimulus alone (st90, 90dB/20 milliseconds),
stimulus tones st74 or st90 delivered 100 milliseconds before the presentation of a startle
stimulus (pp74 and pp90, respectively), and no stimulus (background noise only, st0). St100, st0,
pp74, and pp90 trials were given five times. The average interval between trials was fifteen
seconds, and trial types were given in random order, and the background noise of each chamber

5. was 70dB. For statistical analyses, the percent inhibition of the 110dB stimulus by the 74dB and
90dB prepulse stimuli were analyzed.
Locomotor Testing
To determine whether the ablation of the neural stem cell population increases locomotor
activity after NMDA inhibition or stimulant administration, animals were tested for PCP-induced
locomotor activity on the first day of behavioral testing and for amphetamine-induced locomotor
activity on the second day. Mice were placed in plexiglass containers and allowed to habituate
for half an hour prior to testing. Following the habituation session, animals were injected with
saline and allowed to move freely for thirty minutes. Following the saline session, mice were
inhected with either PCP (5 mg/kg) or amphetamine (2 mg/kg) and allowed to move freely for
one hour. Total movement (m) was measured in the saline and drug sessions using Any-maze
software (Wood Dale, IL). All injections were given intraperitoneally (i.p.).
Elevated Plus Maze
The elevated plus maze was a white plexiglass maze with two closed arms (surrounded
by black walls ten inches high) and two open arms. The arms were fifty-one inches from arm to
arm, two inches wide, and rested sixteen inches from the ground on a metal platform. Animals
were placed in the center of the four arms and allowed to move freely for fifteen minutes. An
observer calculated the total time with all four limbs of the animal on the open arms during each
session.
Porsolt Swim Test

6. On the first day of testing, mice were placed separately in plastic buckets filled with
water and allowed to swim for fifteen minutes. During the session, behavior was manually
scored every twenty seconds, and was categorized under one of two behavioral profiles.
Swimming behavior was defined as mobility of the front or back paws, and floating behavior
was defined as immobility of both the front and back paws. The latency to float for each animal
was also recorded. On the next day, the animals were placed in buckets were allowed to swim for
five minutes. Scoring was performed as described above.
Results
To analyze the effects of neural stem cell ablation on prepulse inhibition, a between-
within ANOVA was performed to analyze the effect of drug treatment, genotype, and sex on
across the startle tone levels, prepulse inhibition 70dB and prepulse inhibition 90dB. The results
indicated no significant interaction between PPI, sex, genotype, and treatment, F(1,23) = .167, p
= .985, no interaction between PPI, genotype, and treatment, F(1,23) = .078, p = .998, no
significant interaction between PPI, sex, and treatment, F(1,23) = .252, p = .958, and no
interaction between PPI, sex, and genotype, F(1,23) = .231, p = .966.
A two-way ANOVA for drug treatment and genotype on locomotor activity after PCP
administration indicated no significant effect, F(1,23) = 2.55, p = .124. There were no significant
effects of genotype, F(1,23) = 1.38, p = .25, or drug condition, F(1,23) = .228, p = .637 on PCP
induced locomotor activity. A two-way ANOVA was also performed to analyze the effect of
drug condition and genotype on amphetamine induced locomotor activity. The results revealed
no significant effect, F(1,23) = .142, p = .71. There were no significant effects of genotype,

7. F(1,23) = .25, p = .622, or drug treatment condition, F(1,23) = 1.277, p = .27 on amphetamine
induced locomotor activity. The alpha level was set at .05 for all tests.
A two-way ANOVA for the elevated plus maze test revealed no significant effect of
treatment and genotype on time spent on the open arm of the apparatus, F(1,23) = 1.33, p = .260.
Results indicated no significant effects of genotype, F(1,23) = .19, p = .67, or drug treatment,
F(1,23) = .01, p = .94 on time spent on the open arm. The alpha level was set at .05 for all tests.
A two-way ANOVA was performed to analyze the effect of drug treatment and genotype on
floating incidents in the Porsolt swim test. The data revealed no significant effect, F(1,23) = .34,
p = .57. Results indicated no differences in floating incidents between genotypes, F(1,23) = .04,
p = .841, or drug treatments, F(1,23) = .08, p = .78.
Discussion
The major finding of our most recent study is that neurodegeneration in the brains of
mammals is not a direct cause of psychiatric disorders such as schizophrenia. We also found that
mouse models are not completely accurate in determining whether neurodegeneration causes
schizophrenia because neurogenesis greatly differs in human and animal brains, and mice cannot
be diagnosed with schizophrenia. The transgenic mice showed no significant difference from the
control mice in the behavior tests used to model schizophrenia. Our tests showed that ablation of
the neural stem cell population in mice for fifty-six days does not lead to deficits in behaviors
relevant to animal models of psychosis or neuropsychiatric disease.
When given an amphetamine or an NMDA receptor antagonist, the mice did not display
schizophrenic-like behaviors as expected. Had the mice expressed schizophrenic-like behaviors,

8. their mobility would have increased considerably, as measured by the Any-Maze software.
Instead, their locomotor activity remained unchanged (as compared to controls). The transgenic
mice given the Porsolt Swim Test showed latency to float times which were not significantly
different from the control mice. We expected the transgenic mice to express depression and
hopelessness, common side effects of schizophrenia, by floating rather than swimming. Both the
control and transgenic mice were placed in the center of the Elevated Plus Maze, and both
behaved similarly and did not display schizophrenic-like behavior by hiding on the arms with
walls. If the transgenic mice had stayed on either of the two enclosed arms, that would have
represented increased anxiety, a characteristic of schizophrenia.
The last experiment, the Prepulse Inhibition Test, is designed to determine if an animal
subject is able to adapt to the presentation of a tone which precedes a louder startle tone. The
transgenic mice were expected to be more paranoid and startle despite being exposed to a
warning tone before the startle than the control mice, but behaved similarly and did not display
schizophrenic-like behaviors. All four tests gave reason to believe that the ablation of stem cells
in mice, which simulates the neurodegeneration caused by schizophrenia, does not cause mice to
express schizophrenia-related behaviors.
Tests run by other scientists, however, have expressed data contrary to ours. DeCarolis
and Eisch (2009) investigated whether hippocampal neurogenesis is an appropriate target for
treating mental illness such as PTSD, Alzheimer’s Disease, and schizophrenia. DeCarolis and
Eisch recognized that mental illness, in general, is marked by decreased hippocampal volume,
structure, and function. They hypothesized that targeting adult hippocampal neurogenesis could
reverse atrophy in the brain and relieve cognitive symptoms such as mood dysregulation and
learning and memory deficits. Their research suggested that neurogenesis can be promoted by

9. methods such as sexual intercourse, because sex steroids emit considerable influences in stress
responses and on the dentate gyrus physiology; environmental enrichment; and exercise, because
it releases endorphins and is an effective treatment for depression. DeCarolis and Eisch
concluded that improving overall health is beneficial to humans with mental illness and can
improve hippocampal health (DeCarolis, 2009). While their theory is well-researched and well-
thought-out, we conducted multiple experiments and none suggested that neurodegeneration has
any effect on psychiatric disorders.
Another study (Iwata 2008) involved the ablation of the stem cells in rats. Deficits in
movement, prepulse inhibition, and other tests were observed after psychostimulant
administration, but not NMDA antagonist administration. A potential explanation for Iwata’s and
our different results might be we ablated the stem cells differently--the Iwata lab exposed the rats
to irradiation, which causes inflammation in the brain and could be responsible for the changes in
the rats’ behavior. The Iwata lab also used rats, not mice, which may have caused variations in
the experiments because different species respond differently to various biological
manipulations.
Hayashi (2008) investigated if high levels of stress during adolescence can cause mental
illness as an adult. It is known that neurogenesis is involved with the postnatal development of
the brain, but Hayashi and his colleagues wanted to determine whether adolescents under stress
are more susceptible to psychiatric diseases in adulthood. They studied the role of postnatal
neurogenesis in adolescent, or three- to eight-week-old, mice by X-irradiating them at four
weeks old to restrict hippocampal neurogenesis. Electrical footshock stress, or FSS, was
administered when the mice were eight weeks old. X-irradiated mice that underwent FSS during
adolescence expressed decreased locomotor activity, a characteristic of schizophrenia, and high

10. levels of paranoia in adulthood. These experiments suggested that lower rates of neurogenesis
during adolescence increase the risk for mental illness such as schizophrenia during adulthood.
The experiments run in our lab suggest that the ablation of stem cells in mice does not
cause them to express schizophrenic-like behaviors. Studies conducted in other labs, however,
have presented contrasting information. In the studies previously described, the stem cells in the
brains of their animal subjects were eliminated, which caused the animals to express
characteristics of schizophrenia such as paranoia and anxiety. A possible explanation for the
varying results between our and other studies could be we only ablated the stem cells, and the
other studies also incorporated psychological stressors. This suggests that, at least in mice, the
ablation of neural stem cells is not sufficient to induce changes in behaviors relevant to animal
models of schizophrenia, but may make the animals more vulnerable to the effects of high stress
levels. Our study used the transgenic GFAP-tk mouse model, in which an inflammatory response
does not make it as difficult to interpret the results as in the irradiation experiments. One
potential follow-up to our study could be to expose our mouse models to a psychological stressor
as well as ablation.
Acknowledgements
I first thank the UCSB Research Mentorship for giving me a generous scholarship, which
allowed me to participate in the program. I also thank my parents, who supported me financially
and also enabled me to participate. I would also like to thank my mentor, Christopher Knight,
who taught me nearly everything I know about neuroscience and sparked my passion for this
field.

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