PSYC 444 Lecture Notes - Lecture 7: Knockout Mouse, Adenosine A2A Receptor, Adenosine Receptor
14 May 2018
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PSYC 444 – LECTURE 7
Phenotype: any observable/measurable trait or behavior
• Example phenotypes: hair color, height, IQ
• The goal is to determine the relative contributions to the phenotype made by genes, environment, and random factors
• Some random factors are observable, others are not
(left): Traditionally, it was hardest studying the DNA and genome and easiest studying phenotypes. However, in the last ten years, this has flipped.
(right): Now, due to advances in technology, it is the phenotype that is hardest to measure
WHAT ARE SOME OF THE WAYS WE CAN MEASURE SLEEP?
Measurement of the phenotype is critical
• How are the phenotypes operationally defined and measured?
• What are some of the strengths and caveats of this method?
Example measurement methods:
• Questionnaires: easy to use but retrospection and self-reports are less reliable and prone to bias
• Actigraphy and video recordings
• Polysomnography: requires more work but are more objective
It is important, especially in genetics, to stop and think about what measurements are being used and how this impacts the findings made.
DO OUR GENES INFLUENCE HOW WE SLEEP?
Yes, they do; a better question is to what extent?
EEG profiles are variable between people
• Brain activity measured during sleep is used to determine the amount of high frequency small waves and low frequency big waves
• The power spectral profile content is different between individuals
The EEG fingerprint is relatively stable. Environmental changes have little effect within an individual.
• Like a fingerprint because it is a very individualized personal profile of how brain activity is distributed
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Changing the amount of sleepiness (sleep deprived or well-rested) has no effect on distribution of brain activity
• Even with sleep-deprivation, individuals are more like themselves (rows) than other people (columns)
There is very little change in spectral morphology, despite large developmental changes and decreases in total spectral power
Below are 4 subjects: 2 teens and 2 children
• Measured EEG spectral profile over development and found much less change in power spectrum compared to dramatic developmental changes.
ESTIMATION OF HERITABILITY
Heritability of a trait: proportion of observable differences between individuals that is due to genetic differences
• The trait-like characteristics of sleep EEG can be studied using twin studies
• Twin studies involve comparison of either monozygotic (one egg; identical) twins who share 100% of their genes or dizygotic (two eggs; non-identical)
twins who share 50% of their genes
o Note that dizygotic twins are genetically comparable to siblings born at the same time
Heritability estimate: proportion (0.0-1.0) of phenotypic variance that is due to genetics
• Twin studies can be used to determine contributions made by genetics and environment on the variance, or difference in phenotype
o With monozygotic twins, there is more genetic control
o With dizygotic twins, there is more environmental control
DZ: some similar, others not (different shades of orange)
MZ: very similar (all orange)
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MZ are much more similar in EEG profile than DZ twins, suggesting that power spectral analysis during sleep is highly attributable to genetics
• Heritability is 0.959, meaning over 90% of the variance between individuals is due to genetics
• Intra-class correlation coefficients (ICC) is 0.934, meaning there is very high within-pair similarity between twin pairs
SUMMARY
Need to be very careful about measurement of sleep phenotypes
• Each measurement has strengths and weaknesses that influence the interpretation of genetic studies
Many sleep phenotypes, particularly EEG phenotypes, appear to be trait-like (stable) and have a strong genetic basis
• Resistant to change with changes in sleep (environment)
• Twin studies suggest many EEG traits are heritable
WHICH GENES INFLUENCE SLEEP? HOW?
Proteins are the functional component of genes
Which genes (and therefore what proteins) are important for regulating sleep?
The Human Genome: a large scale international project that catalogued and described the entire human genome
Many techniques have been used to determine the genes involved in sleep-related phenotypes:
1. Reverse genetics: genetically modified organisms
2. Forward genetics: Quantitative Trait Loci (QTL) mapping; human linkage and association studies
3. Genetics of sleep disorders: narcolepsy, RLS, FFI
1. REVERSE GENETICS
Involves addition or deletion experiments: take model organism, delete or add the gene of interest, then examine corresponding changes
Reverse because you work backwards from the gene of interest to determine what it does
Reverse genetic studies are commonly carried out in mice.
Studies in transgenic/KO mice hae otiuted sigifiatl to ou udestadig of the egulatio of sleep ueous sste, iludig…
• Monoamine neurotransmitters
• Ion channels
• Cytokine pathways (IL1, IK10, TNF)
• Circadian genes (clock, Bmal1, Per1/2, Cry1/2, Npas2)
Transgenic (knockout) mice: inactivation or removal of a gene results in both the mouse and its offspring(s) no longer having the active form of protein translated
from the gene of interest.
Example I: Adenosine A2A receptor
It is known that caffeine affects our sleep; it is a stimulant that makes it difficult to sleep for some amount of time
• The pathway by which the effects of caffeine are mediated was determined by using KO mice
Activity profile: amount of activity (wheel-running behavior in mice) changes over the course of 24 hours
• Following caffeine injection (i.p.), there is a big spike in activity.
• WT: normal, wild-type mice
• KO: knockout mice with either A2A or A1 receptor inactivated
To determine the specific adenosine receptor involved, different receptors were knocked out in different mice.
• In A2A mice, the white and black dots have a very similar profile; the spike following injection is absent
o This suggests that the A2A receptor was, in part, responsible for mediating the effect of caffeine
• On the other hand, in A1 mice, the effect is still present
o Comparable white and black dot profile to wild-type mice
Thus, the adenosine A2A, not A1, receptors mediate the arousal effect of caffeine.
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
The goal is to determine the relative contributions to the phenotype made by genes, environment, and random factors. Some random factors are observable, others are not (left): traditionally, it was hardest studying the dna and genome and easiest studying phenotypes. However, in the last ten years, this has flipped. (right): now, due to advances in technology, it is the phenotype that is hardest to measure. Questionnaires: easy to use but retrospection and self-reports are less reliable and prone to bias. Polysomnography: requires more work but are more objective. It is important, especially in genetics, to stop and think about what measurements are being used and how this impacts the findings made. Brain activity measured during sleep is used to determine the amount of high frequency small waves and low frequency big waves. The power spectral profile content is different between individuals. Environmental changes have little effect within an individual.
