Vulnerability of expression reduced sert mice to learned helplessness
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VULNERABILITY OF EXPRESSION REDUCED SERT MICE TO LEARNED HELPLESSNESS. By Jane Belyavskaya Mentor: Jeff Muller Affiliation: Columbia University (Sackler Institute of Neurobiological Sciences. Introduction. Serotonin: - key modulatory transmitter located in the

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Vulnerability of expression reduced sert mice to learned helplessness


By Jane Belyavskaya

Mentor: Jeff Muller

Affiliation: Columbia University (Sackler Institute of Neurobiological Sciences



  • Serotonin:

    - key modulatory transmitter located in the

    Central Nervous System

    - Implicated in the regulation of certain

    developmental, behavioral, and

    physiological processes

  • Many receptor types that mediate the effects of serotonin

  • The activation of these receptors is regulated almost completely by serotonin transporters (SERTs)

Sert protein that acts to clear serotonin from extracellular regions of the brain

SERT: protein that acts to clear serotonin from extracellular regions of the brain

Present research and use in everyday life

Present Research and Use in Everyday Life

  • Behavioral Changes in Serotonin Transporter Deficient Mice suggest that SERTs are necessary in the development of the brain – lack of them causes greater vulnerability to serious depression and anxiety, as well as greater prevalence of risk- taking

  • Modulation of SERT function can have important effects on neuropsychiatric functioning

    - SSRI’s = serotonin selective reuptake

    inhibitors  enhance serontenergic

    transmissions and decreases depression and


Taking it a step further

Taking it a step further…

  • There are different genetic variations of the SERT promoter gene

    - s-allele:  suggested in the expression

    of less SERT’s, and therefore more

    serotonin left in the extracellular

    regions of the brain

     also suggested in the prevalence of

    depression and anxiety among those who

    possess this gene (at odds with SSRI’s)

     has let do the conclusion that serotonin

    reuptake is necessary during development as

    shown but fundamental KO experiment

    - l-allele: suggested in the expression of

    less SERT

G x e interaction between the gene and environment

G x E (Interaction Between the Gene And Environment)

  • Critical to normal development and mature function, including mental faculties and health

  • Search for causes of mental health problems has recognized the importance of such interaction

  • A human variation in the serotonin transporter gene has been shown to be associated with increased depression only when accompanied by a history of significant stressful life events, such as job loss, divorce, etc.

Relationship with the midbrain

Relationship With the Midbrain

  • Amygdala:

    - Arousal

    - Controls automatic

    responses associated

    with fear

    - Emotional Responses

    - Hormonal Secretions

  • Hippocampus:

    - Consolidation of New Memories

    - Emotions

    - Navigation

    - Spatial Orientation

  • Prefrontal Cortex  Responsible for the executive functions which include:

    - mediating conflicting thoughts

    - making choices between right and wrong

    - predicting future events

    - governing social control

Vulnerability of expression reduced sert mice to learned helplessness

People with at least one short SERT gene end up with smaller amygdalae, and even more dramatically, smaller cingulate cortices as well. Therefore there is a prevalent different relationship between the structures.

Control loop for fear

Control Loop For Fear

Genotypes in mice

Genotypes In Mice

Mice do not have distinguishable promoter regions and therefore no s and l alleles, so complete testing among these alleles is not really possible

The next best thing:

- WT mice (with two functioning SERT genes)

act as the representation of the l-allele

- HEZ [heterozygous] mice (with one

functioning and one KO SERT gene) act as

the s- allele



Part 1: Establish a standard for how s-allele humans react in the presence of stress as compared to l-allele patients

Part 2: How this aforementioned G x E interaction affects neuronal activity among previously implicated parts of the brain


Part 1: In the presence of stress, low expression of serotonin transporters within the brain by a promoter region causes greater susceptibility towards it impacts

Part 2: Neuronal firing within the brain is seriously affected by the amount of SERT expression is noted, especially in those areas most affected by stressful events (i.e. the amygdala, prefrontal cortex, and hippocampus)


The stressor inescapable shock learned helplessness

The Stressor : Inescapable Shock/Learned Helplessness

  • Stressor established within 3 sessions:

    - each consisting of 70 shocks per mouse

    - each on average 15 seconds apart

    - on average 3 seconds in duration

    - Measure of current = 6 milliamps

    - No where to escape

  • Then, moved into a container with a door and shocked again – those who had been properly stressed and were in a condition of learned helplessness had a greater latency of escape

  • Cause the mice to feel completely powerless to change his or her circumstances for the better.

  • The result of learned helplessness is often severe depression and extremely low self-esteem.

Adult inescapable shock stress design

Adult Inescapable Shock Stress Design



Behavioral testing

Behavioral Testing

  • Novelty Induced Hypophagia

  • Novelty Suppressed Feeding : showed a “trend” that suggests that there would have been more evident results if the sample size had been larger

  • Forced Swim Test

Open field

Mice were placed into Plexiglas activity chambers equipped with infrared beams to detect horizontal activity and vertical activity

Activity of mice was recorded for 30 min.

Inescapable shock reduced total locomotion of the mice

I.S. stress increased the proportion of total locomotion spent in the center region of the open field

Open Field

Shock escape

Shock Escape

Mice place into box w/ two compartments separated by a guillotine

At the beginning: subject experience slight shock

The amount of time it took the mouse to escape to the other chamber was measured.

Real story : expected and warranted results achieved

The I.S. HET mice took longer to escape to another chamber than did the I.S. HET

Expected significant different between the Controlled WT/HET and the stressed WT/HET

Protein staining to indicate active neurons within the brain

Protein Staining to Indicate Active Neurons Within the Brain

  • All brains were sliced, stained, and mounted on slides in the course of the past few months

  • The Stain : FOS-B

  • Using a high-end microscope supplied with a camera, we will be either counting manually or with an automated program on the computer

  • The amygdala, prefrontal cortex, and hippocampus are the main focus of these ventures

Expected results

Expected Results

  • Considering that control loop for fear, the expected outcome of presence of immediate early gene of FOS-B

  • There should be less firing in the prefrontal cortex in HET mice than WT in the presence of stress

  • On the other hand, there should be more firing in the amygdala of HET mice than WT in the presence of stress since the prefrontal cortex has failed to calm down the “anxiety” within that region

Discussion why is this important

Discussion: Why is this important?

  • This is the first time that the interaction between serotonin transporter expression levels and adult stress can be modeled in mice.

  • If it is proven that the amygdala in s-allele humans fires more in the presence of stress then in l-allele, then it would lay more groundwork for the study of how the alleles themselves influence the amygdala.

  • How does less serotonin reuptake cause more neuron firing in the amygdala?

Further research

Further Research

  • Learned Helplessness is only one of the many psychological situations found within humans and simulated by Inescapable Shock

  • Though this is common, other stressors can also be experimented with in order to find out how different psychiatric disorders affect behavior and neuronal activity within the brain, or rather how genotypes influence environmental interaction

  • Ex. Social Defeat, Prenatal Separation


Willeit M, Stastny J, Pirker W, Praschak-Rieder N, Neumeister A, Asenbaum S et al. No evidence for in vivo regulation of midbrain serotonin transporter availability by serotonin transporter promoter gene polymorphism. Biol Psychiatry 2001; 50: 8–12.

Caspi A, Moffitt TE. Gene–environment interactions in psychiatry: joining forces with neuroscience. Nat Rev Neurosci 2006; 7: 583–590.

Ansorge MS, Zhou M, Lira A, Hen R, Gingrich JA. Early-life blockade of the 5-HT transporter alters emotional behavior in adult mice. Science 2004; 306: 879–881.

Williams RB, Marchuk DA, Gadde KM, Barefoot JC, Grichnik K, Helms MJ et al. Serotonin-related gene polymorphisms and central nervous system serotonin function. Neuropsychopharmacology 2003; 28: 533–541.


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