N/B genetics 3

• ◦ Immediate Early Genes are genes that code for transcription factors that affect the expression of other genes.
• ◦ Some of these genes appear to be related to changes in synaptic strength and possibly memory.
• ◦ The low expression of these genes during sleep may be associated with thte fact that learning and memory formation are largely absent in this state.

Zif268 : It has a distinct pattern of expression in the brain, and its induction has been shown to be associated with neuronal activity. Several studies suggest it has a role in neuronal plasticity

• ARC: Changes in Arc mRNA and/or protein are correlated with a number of behavioral paradigms including fear conditioning. The mRNA is notably upregulated following electical stimulation in LTP-induction procedures such as high frequency stimulation (HFS), and is massively and globally induced by maximal electroconvulsive shock (MECS).
• The Arc transcript is the first known IEG that is entirely dependent upon activation of the mitogen-activated protein kinase or MAP kinase (MAPK) cascade, a pathway important for regulation of cell growth and survival.

nPas4: Early IEG that induces other IEGS.



C-Fos: Neuroscientists measure expression of c-fos as an indirect marker of neuronal activity because c-fos is often expressed when neurons fire action potentials. Upregulation of c-fos mRNA in a neuron indicates recent activity

It's an image analysis software package that accepts multispectral microscope images that can channel one or more IEG images and show the signal associated with each nuclear component and the amount in the area around it. It's an effective way to look at/ trace IEGs and map neural circuits.

• Npas4 expression is selectively induced by neuronal activity in vitro and by learning in vivo.Npas4 mRNA and protein in cultured hippocampal neurons is selectively induced by membrane depolarization and dependent on Ca2+ influx.
• Npas4 regulates a learning-specific transcriptional program in CA3 that includes many well-known activity-regulated genes, which suggests that Npas4 is a master regulator of activity-regulated gene programs and is central to memory formation.

◦ It regulates the activity-dependent pathway of inhibitory neuronal connections and transcription in the CA3 required for long term fear memory.

CA3 hippocampus

• Anxiety disorders

• PTSD can be modeled through cued fear conditioning.


• General anxiety disorders can be modeled through contextual fear conditioning.
• 

Phobias can be modeled through innate fear conditioning.

• ◦ Through KOs mice can model PTSD,
• ◦ General anxiety disorder (deficient fear memories), panic attacks (pup retrieval) and
• ◦ phobia (innate fear).

• Repetitive movements/behavior, cerebellum related
• Lack of social brain-failure to imitate, understand emotions, underdeveloped social skills and trustworthyness -(fails sallyanne test), amygdala related.
• Spectrum disorders
• Eye contact/facial recognition not well developed

• In the test process, after introducing the dolls, the child is asked the control question of recalling their names (the Naming Question). A short skit is then enacted; Sally takes a marble and hides it in her basket. She then 'leaves' the room and goes for a walk. Whilst she is away, and therefore unbeknownst to her, Anne takes the marble out of Sally's basket and puts it in her own box. Sally is then reintroduced and the child is asked the key question, the Belief Question: 'Where will Sally look for her marble?'
• To pass, the children have to show that Sally has her own beliefs that may not correlate with reality.Children under the age of four, along with most autistic children (of older ages), will answer "Anne's box," seemingly unaware that Sally does not know her marble has been moved.

• ◦ Positive symptoms (disordered thought, hallucinations, delusions) are DIFFICULT to analyze.


• ◦ Some negative functions can be studied (
Social withdrawal -Blunted affect -Deficit in working memory -Attentional deficits)
• 1. working memory deficits (PFC)
2. attention deficits (striatum; sensory motor gaiting)

PTSD/Anxiety have the best animal models available because we know which neural circuits are involved and there are behavioral paradigm that allow studying fear behaviors very well

• ◦ Schizophrenia is hard to model in mice (hard to model positive symptoms).
• ◦ Autism is mostly diagnosed with behavioral symptoms that are hard to model in mice (social behaviors) and can't be modeled in one single mouse line.

• ◦ 2 DA systems are disturbed in schizophrenia.
• ◦ 1. Increase in activity of the mesolimbic pathway (subcortical- amygdala, striatum) would account for the POSITIVE SYMPTOMS.
• ◦ 2. Decreased activity of the mesocortico connections in the PFC accounts for NEGATIVE SYMPTOMS.

• ◦ 1. Target the PFC and isolate specific genes
• ◦ 2. Target the dopaminergic system and: overexpress D2 receptor in striatum, inactivate the D1 recepto, inactivate NMDA receptors to reduce D1 responsiveness.

• ◦ Caused by defect at the nucleus
• ◦ Methylation of cytosine causes a disruption of MeCP2 and transcription is blocked- change of signaling pathway.
• ◦ Causes lots of genes to shut down via methylation (when genes develop they demethylate- stops development).

• ◦ function as a part of the gamma-secretase intramembrane protease complex that cut the APP
• ◦ Forms the active site of complex that produces amyloid beta.
• ◦ Over expression causes accumulation of amyload beta into plaques.

• 1. Amyolid plaques: B-amyloid is a peptide made from APP, when cut from protein fail to leave brain and instead clump together and form amyloid plaques.
• 2. Neurofibrillary Tangles: Made of tau proteins (MAPs), when tau detaches from MT and is hyerphophoralated, it then pairs up with other detached tau's to form tangles, the microtubule disintergrates and azon transport is disrupted which cause neruonal death.
• 3. Shrinking of the brain: coretx and hippocampal neurons die, shrinking the brain. Venricles also enlarge.

• ◦ Memory loss!
• ◦ Also disorientation, impairment of judgment and problem solving, deterioration of language abilities, motor complications, dementia.

• ◦ Early onset is before 65.
• ◦ Late onset is after 65.

• Hippocampal,
• and cortex neurons.

B-amyloid is a peptide made from APP, when it from a protein fails to leave the brain and instead clump together and form amyloid plaques.

◦ About 1% result from autosomal dominant inheritance of an age dependent trait.

◦ They form the active site on the complex that produces amyloid beta.

• ApoE is involved in deposition and clearance of plaques. Polymorphisms are a risk because 2 of the 3 are dysfunctional and if it's not working properly plaques will accumulate.
• ApoE can bind AB and localizes to senile plaques, raising the possibility that this protein plays a role in AB clearance.

• ◦ IEG/transcription factor
• ◦ It is a synaptically induced enzyme.
• ◦ Glutamate activates NMDA receptors that allow Ca2+ flow which eventually induces Cox2.
• ◦ Induced by synaptic activity (neural activity/learning)

◦ Thy1 promoter was used because it is shown postnataly and avoids potential developmental deficits. 

An antibody was used because it will show human and murine cox2 expression and demonstrates continued high expression of TG cox2 in aged animals.

• ◦ Cox-2 results found can be 'applied' to humans as well.
• ◦ One long-term goal of this type of work is to one day devise pharmaceuticals to prevent problems (in this case AD).
• ◦ These must work in humans so it is best to test all biochemical/molecular models with the human protein.
• ◦ Another more practical answer is that by using the human protein you can easily tell where it is being (over)expressed because of that one antibody they had that would bind only the human protein.

Older mice were used, to relate tests to the human model as efficiently as possible (AD prevalent in older people).

• ◦ Speed of swimming (no physical deficits)
• ◦ Straight alley
• ◦ Visual discrimination (normal sight)
• ◦ To eliminate any possible outlying reasons for results. To prove that it is the mental deficit that was performed causing the results.

• ◦ 470nm blue lights, activated with Cre recombinase on site
• ◦ Na+ pump activated

• ◦ 580nm yellow light
• ◦ Cl pump activated

• ◦ Optogenetics – using light and opsin proteins to control neuronal activity in vivo
• ◦ Done with high precision