PSYC 310 Lecture Notes - Lecture 16: C-Fos, Basolateral Amygdala, Dopamine Receptor
7 May 2018
School
Department
Course
Professor
PSYC 318
Behavioural Neuroscience II
January 15th, 2018
Lecture 3/24: Social Reward II
• Reading 1 basically an anatomy paper and loss of function paper
o Experiments blocked/removed oxytocin receptors in specific brain areas to see effect in
NAc
o Authors didn’t describe when anything normally happens: when do cells normally spike in
the brain?
o There’s not much serotonin release in NAc
o Every cell in NAc has 1000+ glutamate inputs from all over: if you’re saying oxytocin
affects serotonin which then affects glutamate, then what glutamate inputs experience
LTD when oxytocin was released?
▪ Complex system, glutamate comes from all over in tons of synapses receiving
numerous inputs, so the overall model of reducing glutamate inputs is a little too
generalizing
o It is hard to draw strong conclusions about behavioral significance about oxytocin
signaling without more information
• Paper 2 research question: Do midbrain dopamine neurons (many of which project to the NAc)
play a causal role in promoting social behavior?
o Dopamine in the NAc particularly (but in many areas of the brain) is known to be involved
in reinforcement learning, or the likelihood that you’ll repeat actions based on their
consequences.
▪ People have determined that natural rewards activate dopamine neurons,
particularly the ones projecting to the NAc
▪ We think NAc involved in lots of reinforcement mechanisms, so logically it could
be involved in reinforcement of social behavior
• First sub. question: Are dopamine neurons active during social interactions?
o We could drill a hole in skull, plant metal wires, a week later record electrical activity
along those wires and assume that is dopamine activity
o Issue: challenging to know precisely which kind of neurons you’re recording from
o Just because we find spiking activity near those metal wires doesn’t necessarily mean
that’s dopamine neurons firing, because these dopamine neurons and intermingled with
many other types of cells in the midbrain
o How can we determine precisely when genetically-identified dopamine neurons
fire?
▪ Approach: put fluorescent indicator of neural activity in dopamine neurons, so
they only became fluorescent when they started spiking, we would then collect
light from the midbrain
• We need a special protein that glows with every action potential
• Is there a good way to make neurons glow when they fire action
potentials?
o Yes: when neurons fire action potentials they have a change in
their membrane potential. People have created fluorescent
proteins that glow more or less at different membrane potentials:
they’re called voltage-sensitive fluorescent proteins
o Voltage changes quite rapidly, and there are numerous technical
challenges associated with recording these diff membrane
potentials
find more resources at oneclass.com
find more resources at oneclass.com
• During action potentials, calcium enters the cell and raises intracellular
calcium levels for tens of milliseconds
o With every action potential calcium enters cell.
o Main reason calcium is used as good indicator of neural activity
is because calcium concentrations in neurons are kept extremely
low: if there’s ever an increase in permeability of calcium ion
then tons of calcium comes in very quickly
o If we can have fluorescent indicator of calcium levels it should
correlate well with spiking activity
• Biologists have engineered strings of DNA that encode synthetic proteins
glowing whenever they bind to calcium known as fluorescent calcium
indicators
o Trick is to selectively get dopamine neurons to express this
synthetic DNA so they glow whenever calcium levels rise
▪ GCaMP is a genetically encoded calcium indicator
• Can be made from DNA in cells
• It glows whenever it is bound to calcium
• GCaMP was made by fusing together three different natural proteins:
GFP, calmodulin and M13 peptide sequence from another protein
o When they did this, GFP no longer fluoresced unless calcium-
bound, when the natural glow was restored
• People have been trying to modify GCaMP protein ever since
• If a cell expresses GCaMP it glows brighter whenever calcium levels
rise, when action potentials occur
o Target GCaMP to dopamine cells
▪ Authors want to make dopamine neurons in midbrain express GCaMP protein so
that they glow whenever an action potential fires
▪ The simplest way to do this would be using a virus to deliver GCaMP DNA to
cells in the midbrain, infecting all midbrain cells
• To limit expression to just dopamine cells they would need to include a
gene promoter region that only dopamine cells would read
• We know that dopamine cells are the only ones in the brain that make
enzymes needed to make dopamine, one is tyrosine hydroxylase (TH)
• We know TH gene and preceding DNA but don’t fully understand gene
promoter region
• Typically 2000 base pairs before the gene in genome is the promoter
region, but when we take this we don’t get perfect expression in practice
o Promoter for TH enzyme presumably comes before normal TH
gene
o We tried to remove all viral DNA (sometimes used AAV virus)
▪ This virus can hold about 6000 base pairs of DNA, but
2000 of them are needed for the virus to function, so we
have 4000 base pairs to work with and add in DNA
o GCaMP protein itself is about 2000 base pairs, leaving us 2000
left: so we take last 2000 base pairs of DNA string before TH, the
promoter region
o But this isn’t enough promoter region to achieve specificity, for
that you’d need 4000-5000
▪ Authors can’t get selective expression of GCaMP in dopamine cells using just a
virus, it is too small to include all DNA instructions so just dopamine cells would
read it
▪ So they use tyrosine hydroxylase-cre mouse (TH-cre mice)
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
Lecture 3/24: social reward ii: reading 1 basically an anatomy paper and loss of function paper, experiments blocked/removed oxytocin receptors in specific brain areas to see effect in. Ltd when oxytocin was released: complex system, glutamate comes from all over in tons of synapses receiving numerous inputs, so the overall model of reducing glutamate inputs is a little too generalizing. Is there a good way to make neurons glow when they fire action potentials: yes: when neurons fire action potentials they have a change in their membrane potential. It glows whenever it is bound to calcium. Gfp, calmodulin and m13 peptide sequence from another protein: when they did this, gfp no longer fluoresced unless calcium- bound, when the natural glow was restored, people have been trying to modify gcamp protein ever since. Iegs) that tend to be read and turned into protein in response to any elevations in normal activity.
