rat as an animalmodelofschizophrenia:a - ETH E

Diss. ETH N° 14042
Long-termeffects of social isolation in the
rat as an animal model of schizophrenia: a
behaviouraland neurobiological
investigation
A dissertationsubmitted to the
Swiss FederalInstitute of Technology Zürich
For the degree of
Doctor of Natural Sciences
Presented by
Isabelle Weiss
Biologist
DEA, ULP Strasbourg
Born November 26* 1974
Citizen of Strasbourg(France)
and Boudevilliers (Switzerland)
Acceptedon recommendation of
Prof. Dr. Joram Feldon, examiner
Prof. Dr. Michael Koch, coexaminer
Dr. Christopher Pryce, coexaminer
2001
Resume
maladie neuropsychiatriquemultifactorielle dont
l'apparition semble etre associee ä des facteurs genetiques, envfronnementaux et
neuro-developpementaux. Le role de chacun de ces facteurs dans le
developpementde la maladie reste cependant en grande partie incompris. Pour ces
raisons, l'etude de modeles animaux presentant des deficits de type
schizophrenique a recu une attention grandissante au cours de ces vingt dernieres
La
Schizophrenie est
une
annees.
L'inhibition du reflexe de sursaut (IRS) reflete la reduction du reflexe de
sursaut ä un Stimulus intense lorsque ce Stimulus est immediatement precede par
un Stimulus d'une intensite plus faible. Des etudes cliniques ont mis en evidence
que certains patients schizophrenes presentent un deficit d'IRS. Ce deficit preattentionel a ete associe ä une perte des capacites de filtrage sensori-moteurchez
Des deficits d'IRS similaires etant egalement observes chez le rat
apres activation du Systeme dopaminergique mesolimbique, le paradigmed'IRS a
ete propose comme modele animal des dysfonctionnements attentionnels observes
ces
patients.
chez certains patients schizophrenes.La similarite des parametres experimentaux
utilises pour l'IRS chez l'homme et le rat, par exemple 1'intensite de Stimulation,
suggere un substrat neurologique sous-jacent commun. L'etude de modeles
animaux de deficits d'IRS a donc recu un interet considerable en recherche preclinique, afin d'expliquer les mecanismes neurobiologiques de la Schizophrenie, et
de developper de nouvelles substances pharmacologiques ä des fins
therapeutiques.
manipulations
pharmacologiques (injections systemiques, infusions intracerebrales) ou
chirurgicales (lesions corticales ou sous corticales). Ces modeles sont toutefois
invasifs et, de plus, restreignent les etudes ä un Systeme de neurotransmission
specifique ou ä une region cerebrale particuliere. En outre, dans le cadre du
developpement de nouveaux traitements, les manipulations pharmacologiques
augmentent le risque de reactivite croisee et tendent ä selectionner des
homologues pharmacologiques (par exemple, un antagoniste dopaminergique va
annuler les effets sur l'IRS induits par un agoniste dopaminergique). C'est pour
Chez le rat,
un
deficit d'IRS peut etre obtenu ä la suite de
10
contoumer ces limitations que de nouveaux types de modeles animaux ont ete
recemment developpes. Ces modeles sont bases sur la theorie neuro-
Schizophrenie et consistent ä manipuler l'environnement
social des animaux pendant leur developpement.
L'isolement social (IS) represente l'une de ces manipulations
developpementale
de la
(1 rat par
cage) durant la periode developpementale allant du sevrage (21 jours d'äge) ä l'äge
adulte. Les effets d'une teile privation sociale sont generalement evalues ä l'äge
environnementales. II consiste ä elever des rats
en
conditions isolees
adulte et sont compares avec ceux d'animaux contröles eleves dans
environnement social (2-4 rats par cage).
un
L'objectif principal de ce travail de these est d'etudier les effets ä long terme
de TIS sur les capacites de filtrage sensori-moteurchez le rat. De plus, les profus
comportementaux,neuroendocrinologiques et neurochimiques des rats isoles sont
examines, dans le but de mieux comprendre les mecanismes neurobiologiquesqui
dysfonctionnements d'IRS observes chez ces animaux.
Le premier chapitre replace mon travail de these dans son contexte
theorique en discutant et resumant les connaissances actuelles sur l'utilisation des
sous-tendentles
manipulations
environnementales pour l'induction de deficits d'IRS. Les
experiences presentees dans les quatre chapitres suivants etudient l'implication de
differents parametres experimentaux dans l'induction de deficits d'IRS apres IS,
dans le but d'augmenter la robustesse et la reproductibilite de ce modele. Un
premier essai en ce sens est decrit au Chapitre 2. Cette etude examine l'influence
d'un environnementappauvri (cages ä fonds grillages) durant la periode d'IS, sur
la reponse d'IRS de rats adultes. De maniere surprenante, l'elevage de rats dans
des cages ä fonds grillages supprime l'expression de deficits d'IRS chez les rats
isoles (Cf. Chapitre 2). En consequence, les experiences suivantes sont toutes
effectuees avec des animaux eleves sur sciure. La possible influence du cycle
circadien sur l'expression de l'IRS chez des rats sociaux est presentee au Chapitre
3. Les resultats montrent que l'IRS reste stable, que les animaux soient testes lors
de leur phase diume ou nocturne. Dans le Chapitre 4, des rats mäles issus de trois
souches couramment utilisees dans les etudes d'IS (c'est ä dire Wistar, SpragueDawley et Lister hooded) sont testes pour leur capacite ä montrer des deficits
d'IRS apres IS. Seuls les isoles de la souche Sprague-Dawleymontrent un deficit
d'IRS robuste et stable (Cf. Chapitre 4). Cette souche de rats est donc utilisee
pour toutes les etudes suivantes.
11
Afin de renforcer davantage le modele d'IS, une experience combinee est
effectuee, au cours de laquelle des animaux nouveau-nes sont separes de leur mere
4h par jour puis eleves en IS apres sevrage (Cf. Chapitre 5). Aucun effet additif
de la Separation maternellen'est observe sur la reponse d'IRS des rats isoles. Cette
etude, egalement conduite avec des rats femelles, permet d'etudier l'influencedu
sexe sur la reponse d'IRS. A l'inverse des mäles, les femelles isolees ne montrent
pas de deficits d'IRS comparees ä des femelles groupees et ceci independamment
de la manipulation effectuee avant le sevrage (Cf. Chapitre 5). Enfin, differents
aspects du comportement et de la neurobiologie des rats sont etudies afin de
comprendre les mecanismes neurologiques impliques dans le dysfonctionnement
des capacites de filtrage sensori-moteur apres IS. Ainsi, le Chapitre 6 etudie la
sensibilite ä l'amphetamine (agoniste dopaminergique indirect) de mäles et
femelles isoles. Enfin, le Chapitre 7 examine l'expression comportementale et
neuro-endocrinologique d'etats de stress et d'anxiete chez les rats isoles. Pour ce
faire, la reactivite des animaux ä des situations nouvelles et potentiellement
stressantes ou anxiogenes est testee dans Popen field, l'elevated plus maze ou le
paradigme de freezing. De plus, la liberation d'hormones de stress (ACTH et
corticosterone) est mesuree en conditions basales, ainsi qu'apres la presentation
repetee de Stimuli sonores de forte intensite.
les resultats presentes dans cette these demontrent que le
deficit d'IRS obtenu apres IS est un modele fragile. La vulnerabilite du modele
repose sur sa dependance ä de nombreux parametres methodologiques, comme la
souche, le sexe des animaux et les conditions de cages lors de l'elevage en IS. De
plus, meme lorsque ces parametres experimentaux sont correctement contröles, le
deficit d'IRS induit par TIS reste fragile. Cette fragilite reduit considerablement la
En
conclusion,
fiabilite du modele pour la selection et le
developpementde nouvelles substances
pharmaceutiques aux proprietes anti-psychotiques. Neanmoins, ce modele pourrait
permettre une meilleure comprehension des alterations physiologiques et
neurochimiques induites par une experience stressante vecue precocementdans la
vie de l'individu. De telles connaissances sont indispensablespour comprendre les
mecanismes neurobiologiques impliques dans une maladie psychiatrique teile que
la
Schizophrenie.
SUMMARY
Schizophrenia is a multifactorial neuropsychiatric disorder for which several
causes have been suggested,
including genetical, environmental and
neurodevelopmental alterations. However, these factors are mostly hypothetical
and their contribution to the disease remains mainly unexplained. Therefore, the
study of animal models has received considerable attention in the past twenty
years. Prepulse inhibition (PPI), the normal reduction of the startle reflex to an
intense Stimulus when this Stimulus is immediately preceded by a Stimulus of
weaker intensity, is impaired in Schizophrenie patients. This impafrment of preattentive processes has been associated with a loss of sensorimotor gating abilities.
Because PPI deficits have been also observed in rats following activation of the
dopaminergic mesolimbic system, the PPI paradigm has been proposed as an
animal model of the attentional impairments seen in Psychiatric diseases such as
schizophrenia. The fact that the PPI experimental parameters are very similar
between humans and rodents suggests that the neurobiological mechanisms
underlying the PPI response can be similar across species. Therefore, the study of
animal models of deficient PPI in pre-clinical research has received considerable
interest, largely because these models can provide valuable knowledge regarding
the neurobiology of schizophrenia and can also be used for developing
antipsychotic drugs.
animals,
PPI deficits
by means of pharmacological (i.e.,
systemic injeetion, intracerebral infusion) or surgical (i.e., lesion) manipulations.
However, these invasive models bias the research towards a specific
neurotransmitter system or brain region. Furthermore, in the context of drug
Screening, the pharmacological model automatically implicates drug-drug
interactions and tends to select pharmacological homologues. More recently, in
line with the neurodevelopmental hypothesis of schizophrenia,another category of
In
can
be induced
animal models has been developed. These models are based on manipulations of
the social environment. One of these environmental manipulations, i.e., social
isolation (SI), consists of rearing rats alone in single cages during the
14
adulthood. Commonly,
the effects of such social deprivation are evaluated in adulthood and compared
with those of control subjects reared in a social environment (2-4 rats in a cage).
developmental period from weaning (21 days of age) to
goal of the present thesis was to examine the long-term effects of
SI on sensorimotor gating abilities of adult animals. Furthermore, the behavioural,
neuroendocrinological and neurochemical profiles of social isolates were
investigated in an attempt to better characterise the neurobiologicalmechanisms
underlying the PPI alterations observed in these animals. The first chapter
provides the theoretical background for my thesis work by discussing the current
knowledge on the utilisation of environmental manipulations to disrupt PPI. The
experiments presented in the following four chapters were aimed at improvingthe
robustnessand reproducibility of the inductionof PPI deficits following SI. A first
attempt to obtain a robust Sl-induced PPI disruption is described in Chapter 2.
The main
This study examined the influence of a sterile rearing environment during the SI
period (i.e., cages containinggrid-floors versus sawdust) on the subsequent PPI
response of male Wistar rats. Surprisingly, rearing rats in cages containing gridfloors prevented the expression of PPI deficits in isolates (see Chapter 2).
Therefore, the following experiments were all performed using animals reared in
sawdust cages. The possible influence of the light-dark cycle on the expression of
PPI was investigated in control rats and is presented in Chapter 3. The results
revealed that the PPI response remained stable across the circadian cycle. In
Chapter 4, male rats from the three strains commonly used for studies on SI (i.e.,
Wistar, Sprague-Dawley and Lister hooded) were tested for their suitability to
show Sl-induced PPI deficits. Only isolated rats from the Sprague-Dawley strahl
demonstrated a robust and reliable PPI deficit (see Chapter 4). The following
experiments were consequently conducted with rats from the Sprague-Dawley
strain.
combined experiment was performed,
in which the animals were separated from their mothers 4 hours per day for the
entire pre-weaning period and then isolated after weaning (see Chapter 5). No
'additive' effects of maternal Separation and SI were observed on the PPI response
To further strengthen the SI model,
a
of the rats. The latter study was also conducted with female rats to examine the
influence of gender on the PPI response. The results showed that in contrast to
males, isolated females did not demonstrate a PPI deficit as compared with
grouped females, irrespective of the pre-weaning manipulation (see Chapter 5).
Finally, further aspects of the behaviour and the neurobiology of socially isolated
animals were studied to elucidate the underlying brain mechanisms possibly
15
involved in the disruption of sensorimotor gating abilities. In Chapter 6, the
sensitivity of isolated male and female rats to an indirect dopaminergic agonist
(i.e., amphetamine) was investigated. The behavioural and neuroendocrine
expression of stress/anxiety states in isolated rats was examined in a large study
presented in Chapter 7. The behavioural reactivity of the rats to novel and
potentially stressfulsituations was investigated in the open field, the elevated plus
maze and the freezing paradigm. Moreover, the hormonal release of stress
hormones (ACTH and CORT) were measured in basal and stress conditions (see
Chapter 7).
In conclusion, the findings presented in this thesis suggest that the
disruption of PPI following SI is a very fragile model that requires careful
considerationof a number of methodological parameters, such as strain, gender,
and caging condition, which can significantly affect the size of the SI effect. The
presented in this thesis further demonstrate that even when all these
parameters are correctly controlled, the Sl-induced PPI deficit is still not robust.
Although the SI model may require further refinements, the fragility of its effect
results
considerablyweakens its reliability for drug Screening. Nevertheless,this
model provides promising features for understanding the physiological and
neurochemical changes producedby early stressful experience, which may prove
to be helpful in elucidating the neurobiology of schizophrenia.
on
PPI