Behavioral Neuroscience Introduction to Neuroscience:

Lecture 4: Animal models of social disorders

Tali Kimchi Department of Neurobiology Tali.kimchi@weizmann.ac.il

* Presentation Materials for Personal Use ONLY

♂ ♀ XY

XX

Estradiol &

Progesterone

Testosterone &

Estradiol Organization

Dimorphism of the brain: differentiation and activation

-

Bipotential gonad

SRY gene

+

Ovary Testis

Estradiol &

Progesterone

Sex-specific behaviors

Activation Testosterone &

Estradiol

Organization

(permanent changes)

Factors regulating sexual dimorphic (reproductive) behaviors

-Sex hormones (organization and activation stage)

- Sex chromosomes (X/Y)-linked genes

- Imprinting genes (maternal and paternal genes

- Environmental factors

Exposure different levels of testosterone in uterus on female/male behavior

High level of testosterone

in the surrounding

Low level of testosterone

in the surrounding High level of testosterone

in the surrounding

Low level of testosterone

in the surrounding

Rayan and Vandernbergh 2002; Neuroscience and Bioheavioral Reviews

Exposure to testosterone in uterus on female behavior

Rayan and Vandernbergh 2002; Neuroscience and Bioheavioral Reviews

Effect of prenatal stress on sexual dimorphism in brain and behavior

The Medial Preoptic Area (MPOA) is activated by testosterone and is essential to the activation of male sexual behavior

Castration Microinjection of testosterone into the MPOA

Reinstate sexual behavior

Abolish of sexual behavior

MPOA lesion Abolish of sexual behavior

♂ ♀

Anderogen Receptor expression in the MPOA

Intact male Sexual behavior Increase neuronal firing rate in the MPOA

Sex-specific pheromone stimuli or mating behavior

Induce c-fos in MPOA of both males and females (c-fos is immediate early gene, indirect molecular marker of neuronal activity)

Shah et al 2004; Neuron

SDN-POA control male

SDN-POA stressed male

Anderson et al 1985; Brain Research

Effect of prenatal stress on sexual dimorphism in rat brain

ES: Environment stress (change in lighting/ temperature)

NS: nutritional stress (50% of total food of control males)

Mueller and Bale 2008; JNS

C: control;

Prenatal stress during (E) early, (M) mid or (L) late gestation

Suspension tail assay Force swim test

Mueller and Bale 2008; JNS

The control of pheromone signals on dimorphic behaviors

Chemical (odor) signals that are emitted by animals to communicate information to their own species

What are pheromones?

What are pheromones?

Pheromone signals are largely involved in the regulation of social and reproductive behaviors in most animals (including in human)

Attracting mate partner Pups recognition Male territoriality

Primer effects: induce sequence of slow long-lasting physiological and neuroendocrine responses

Releaser effects: induce relatively rapid, fixed, behavioral responses

Ultrasonic vocalization in the presence of female

Aggressive behavior toward intruder male

Mating behavior

Pheromone effects in rodents

Aggressive behavior of lactating female

Maternal behavior (e.g. pups retrieval)

Bruce effect: Recently mated female will return to estrous if exposed to strange male (pregnancy block)

Whitten effect: Induction of estrous in group-housed females by exposing to male (urine)

Lee-Boot effect: Grouping several (8-12 individuals) females in a cage results in suppression of their estrous cycles

Vandenbergh effect: Puberty acceleration caused by exposure to male, during female development.

Puberty-delay caused by group-housed females.

Endocrine effects: Intact male exhibit LH surge Following exposure to female mice. Female exhibit LH surge in response to male or its bedding

The olfactory systems

Liman et al. 1999

TRPC2 expression in the VNO

VNO

OB

Detection of chemosensory signals in mice

Typical male-female reproductive behaviors

♂

♀

Aggressive behavior Sexual behavior

TRPC2 mutant female (brown) with normal male (black)

Sexual behavior of TRPC2-KO lab female

Kimchi et al. 2007; Nature

♂

♀ WT

WT

TRPC2-KO

Male-typical sexual behavior in TRPC2-KO lab females

An

ima

l m

ou

nti

ng

(%

) M

ou

nti

ng

tim

e (

se

c)

Pe

lvic

th

rus

tin

g (

se

c)

♀

Kimchi et al. 2007; Nature

Female

Male

Social behavior under semi-natural environment

control

mutant

Maternal Behavior

Failure to discriminate between male and female

?

Male-typical sexual behavior (courtship and mounting behaviors)

Female mutant

Normal male

Female-typical behavior (maternal behavior)

Summary-part 1: Behavioral phenotype of TRPC2-KO lab females

Old model

♀

♂ ♂

New model

♀

4 mutant males

2 control (normal) males

Kimchi et al. 2007; Nature

Stowers et al 2002; Science

Social and sexual behaviors of male mutant mice

Failure to discriminate between male and female

?

Aggressive behavior

Normal testosterone level

Model: Pheromone inputs repress neuronal circuit for male-typical sexual behavior in females

♂ Sex-specific

pheromone signals

♀ Sex-specific pheromone signals

Dulac and Kimchi, 2007; CONB

♂

Pheromone inputs repress maternal behavior in males

TrpC2-/- mutant male retrieve newborn pups back to nest

TrpC2-/- mutant males exhibit reduce male-typical (infanticide) behavior and elevated female-typical (pup nursing) behavior

Attacking pups Nursing pups

Social and sexual behaviors of male mutant mice

Failure to discriminate between male and female

?

Aggressive behavior

Normal testosterone level

Female-typical behavior (pup caring / nursing behavior)

Donaldson and Young 2008; Science

Oxytocin and Vasopressin

Isoleucine

Phenylalanine

Argenine

Leucine

Neurophysiology of Oxytocin (OXT) and arginine vasopressin (AVP)

Oxytocin KO mouse model

Ferguson et al 2001; J Neuroscience

Summery:

OTKO mouse fail to habituate to, or recognize, a stimulus mice even

after repeated exposures

The deficit in social recognition in OTKO mice represents a defect in

the initial processing of olfactory cues and not in the recall of the

previously stored memory

OT must be present in the MeA during the initial social exposure for

the proper processing of the olfactory information and the

development of the social memory

Oxytocin effects on humans

The role of OT and VPA in social behavior of Bi-parental rodent

The neuronal basis of pair bonding

Pair bonding and social behavior in voles

Prairie voles

Highly social

Monogamous

Spend more than 50% of

their time interacting with

other prairie vole

• Montane voles

• Avoid social contact except for

the purpose of mating

• Polygamous

• Spend only around 5% of their

time socially interacting.

Hemanth et al 2006 Physiology

Prairie voles have high levels of OT receptor in the nucleus accumbens and the basolateral amygdala relative to montane voles

Montane voles have high levels of receptors in the lateral septum.

Prairie voles have high densities of

the V1a subtype of the AVP receptor in the ventral pallidum and the medial amygdala compared with montane voles.

Montane voles have much higher levels of receptors in the lateral septum than do prairie voles.

Oxytocin and parental behavior in Prairie voles

Oxytocin receptor level

High

Parental behavior

Low

% o

f fe

male

show

ing n

urt

uring b

ehavio

r

Ross and Young, 2009

Prairie vole

Montane vole

Oxytocin and partner preference in Prairie voles

Transgenic mice expressing

the V1aR of the prairie vole

Young et al 1999; Nature

Vasopressin receptor and pair bonding in Prairie voles and Montane voles

V1aR expression

Prairie

Montane

Hammock & Young, 2005 Science

Meadow voles (Microtus pennsylvanicus) Prairie voles (Microtus ochrogaster)

Young et al. 1999 Nature

♂

Promiscuous social behavior

Monogamous social behavior

“A single gene can turn the Don Juan of voles into an attentive home-loving husband”. BBC news

The “Fidelity gene”

Walum et al 2008, PNAS

?

The genetic basis for pair bounding

Vasopressin R1a

The use of mouse models to study neuropsychiatric disorders (e.g. autism spectrum disorders)

Kanner L. Autistic disturbances of affective contact.

Nervous Child 2, 217-250 (1943)

2011-2013 DSM-V: classified better the symptoms (criteria of diagnostics)

• Complex genetic disorders (i.e. interactions between many genes and environmental factors)

• Few hundreds of genes were associated with autism

• No biomarker for autism diagnostic: diagnostic is based only behavioral symptoms (~1.5-3 years old)

2014: 1 in 68 children

(1 in 42 boys and 1 in 189 girls)

Autism Spectrum Disorder

Must meet criteria A, B, C, and D:

A. Persistent deficits in social communication and social interaction

across contexts, not accounted for by general developmental delays, and

manifest by all 3 of the following:

1. Deficits with social initiation and responses

2. Deficits in non-verbal communication

3. Deficits in social awareness and insight, as well as with the broader

concept of social relationships

B. Restricted, repetitive patterns of behavior, interests, or activities as

manifested by at least 2 of 4 symptoms:

1. Atypical speech, movement, and play

2. Preocupations with objects or topics

3. Rituals and resistance to change

4. Atypical sensory behaviors

C. Symptoms must be present in early childhood (but may not become fully

manifest until social demands exceed limited capacities)

D. Symptoms together limit and impair everyday functioning.

American Psychiatric Association DSM-5 Development (May, 2013)

http://depts.washington.edu/dbpeds/Screening%20Tools/DSM-5(ASD.Guidelines)Feb2013.pdf

http://depts.washington.edu/dbpeds/Screening%20Tools/DSM-5(ASD.Guidelines)Feb2013.pdf

Stoner et al 2014 New England Journal of Medicine

The use of mouse models to study neuropsychiatric disorders (e.g. autism spectrum disorders)

Recent mouse history

W.E. Castle C.C. Little

Fancy mouse breeding - Asia, Europe

(last few centuries)

Retired schoolteacher Abbie Lathrop

collects and breeds these mice

Granby, MA – 1900

Castle, Little and

others form most

commonly used

inbred strains

from Lathrop stock

(1908 on)

• Mammalian system - close to human

(genetically, physiologically and morphologically)

• Small and easy to handle

• Easy to house and breed

• Inexpensive

• Can be genetically manipulated

• There is a lot of biological knowledge (e.g. Jax lab database, MGI, Allen brain atlas)

Mouse sequence reveals great similarity with the human genome

Extremely high conservation: 560,000 “anchors”

Mouse-Human Comparison

both genomes 2.5-3 billion bp long

> 99% of genes have homologs

> 95% of genome “syntenic”

Steps towards a Transgenic Model

Working hypothesis

Gene Construct

Insertion into an early embryonic stage

Screening for transgenic animals

Profiling of expression pattern

Phenotyping

Model Validation / Experimentation

Transgenic Animals: Definition

Mutant animals carrying experimentally introduced foreign

genetic elements in all their cells, including the germline

Transgenic:

Introduction of foreign or altered gene: transgenic Over-expression, mis-expression, dominant-negative

Normal allele also present - product from two alleles

Knockout:

Replace normal with mutant allele: Gene knock-out - removal of a part of or a whole gene

No normal allele - product of manipulated allele only

Transgenic and Knockout Animals

Institute of Laboratory

Animal Science

University of Zurich

Analyse the progeny

by PCR or Southern blotting

Generating Transgenic Mouse Model

• Transgenic construct: gene size • Varied expression levels: random integration, copy number • F0 mice may be mosaic • Lethality due to transgene integration or expression

Generating Knockout Mouse Model

Social and reproductive, Stress-anxiety, Learning and memory, Orientation,

Motor and cognition skills, sensory perception, etc

Behavioral phenotyping of the transgenic mouse model

Knockout mouse model

Voineagu et al 2010; Nature

Autism

Schizophrenia

Autism &

Schizophrenia

McCellan & King 2010; Cell

Common genetic basis to Autism and Schizophrenia

Silverman et al 2010;

Nature Reviews Neuroscience

Silverman et al 2010; Nature Reviews Neuroscience

Studying autistic-related behaviors in mouse models

Autism Spectrum Disorders

Main characteristics:

Social and Communication Deficits

Fixed Interests (Rigidity in habits)

Stereotypic (repetitive) Behaviors

Symptoms must be present in early childhood

Social behavior test for mice

Cognitive rigidity Wet T-maze assay

Stereotypic behavior

Cryan and Holmes, 2005

Suggested behavioral assays in animal models relevant to anxiety/stress disorder

Examples of automated behavioral phenotyping systems for rodents to evaluate locomotion, anxiety, memory