L11 Flashcards
what are 2 pathological hallmarks of AD
plaques and tangles
what does amyloid precursor protein usually get cleaved by
alpha and gamma secretase
what does amyloid beta do
This is toxic and it clumps together to form plaques. These kill all the cells around them (these are also toxic in their soluble form)
what happens to Tau in AD
TAU proteins usually stabilise microtubules.
These proteins become phosphorylated and then the microtubules fall apart and the TAU gets clumped together
what are the 3 stages of Tau mediated neurodegeneration
- Hyperphos of tau leading to disassembly of MTs causing axonal transport
insufficiency - Tau aggregates in
axons/dendrites which congest axonal
transport - Tau pathology
transmitted
synaptically
describe the chronological relationships among Alzheimer’s disease
pathology and clinical symptoms
both Tau phosphorylation and beta amyloid start happening in the preclinical phase
however Tau phosphorylation starts happening before BA but happens slowly
BA generation happens latter but more rapidly
what are the similarities between AD and diabetic neuropathy
they both disrupt protein structure –
disrupt structure =
disrupt function causing clinical symptoms
once symptoms appear it maybe too late for recovery
what does the nissl stain, stain
rER (therefore soma in neurons)
Nissl was found to be selective for
RNA and so represents large stacks
of rough endoplasmic reticulum
what are proteins involved in, in neurons
axonal transport and phospholipid bilayer (the proteins in it)
describe the Phospholipid bilayer
hydrophobic ends in middle zone
hydrophilic ends –intra/extracellular
compartment
!not permeable to ions!
- it is this reason that neurons have so many proteins as the need so many ion channels
what are the 2 types of channels
a simple channel
a complex folded structure composed of subunits (same/different)
all cells have proteins in the membrane. why in neurons do they produce significantly more than other cells
neurons have a huge membrane area
´Epithelial cell
- 20µm soma diameter
- membrane area 1,256µm2
neuron
- 20µm soma diameter
- membrane area 250,000µm2
therefore it is 200x greater membrane area
to make these proteins there is lots of RER
also have specialised vesicles
why are membrane proteins are essential components of
neuronal structure
- are essential for neuronal function therefore proteins need to be in the ‘right’ place and any change may result in dysfunction
what hold the transmembrane protein in the correct place in the membrane
Transmembrane protein –> scaffolding protein –> actin mesh
what technique is used to identify and locate proteins
immunohistochemistry
what does immunohistochemistry tell us
where the protein is, where it is held/located (cellular level)
how does immunohistochemistry work
first you would take a protein of interest from a rat and inject it into a goat. The goats immune system would then produce antibodies to this foreign object (protein of interest). This is called the primary antibody
We then produce a secondary antibody which is just a generic goat antibody (because the primary is specific to the protein of interest so we don’t know what it is made up of. The secondary we do therefore we are able to produce a morker to see the secondary antibody
what is the point of the primary antibody when we have the secondary antibody
We don’t just send the general goat antibody (secondary) because the protein of interest might be in a hard place to get to therefore you want a specific antibody to be produced so that it is alse to reach it.
The second reason why we do it this way is so that we can amplify the signal
what is ankG
it is a scaffolding protein in the proximal axon and nodes
what is Caspr
- transmembrane protein
- member of neurexin family
- cell adhesion
This is in very specific locations
what is MAP2
- microtubule associated protein
- absent in most astrocytes
MAP 2 is specific for dendrites
what is PO
- Myelin protein
why is actin often yellow in immunohistochemistry
Dendritic spine morphology (green)
F-actin (red)
Merged red and
green are shown in yellow.
because actin is what is in the dendritic spines
why do we want to know protein location
because it helps to understand their function
why do we use immunohistochemistry
Using Immunohistochemistry we can identify multiple proteins in
multiple locations -> precision of function and a range of
functions
what is found at a chemical synappes
presynaptic = specialised membrane with active zones where neurotransmitter is released
cleft = matrix fibrous proteins
postsynaptic = specialised membrane with receptors for neurotransmitters
therefore the synapse is a complex membrane specialisation
what does the presynaptic membrane contain
- Calcium channels
- Docking proteins
- Transporter proteins
- Reuptake transporter
- Specific vesicle proteins
what does the postsynaptic membrane contain
Voltage gated channels
Transmitter-gated ion channels
what is an example of why we have postsynaptic densities
NMDA receptor
NMDA receptor- transmitter gated ion channel
it has a complex protein - post synaptic density
PSD-95 - scaffold protein, under the plasma membrane for the clustering of receptors, ion channels etc.
what are some examples of cleft proteins
Trans-synaptic proteins
Mind the gap (MTG) - binding protein
cell adhesion molecules
where is mind the gap protein secreted from
Secreted presynaptically and crosses the gap to the post synaptic density
what do cell adhesion molecules interact with
interact with molecules on
surface of adjacent cell
why do we have local protein synthesis
because if everything was made in the soma and then transported down the axon (anterograde by kinesin) then it would take forever
where does local protein synthesis happen
at the post syanptc membrane
how does local protein synthesis happen
The repressed mRNA gets attached onto the motor protein and via kinesin it is attached to the actin network and is transported to where it needs to go
the mRNA then goes to the polyribosomes which make the protein
what is Fluorescence in situ hybridization (FISH)
you insert a florescent probe which is complimentary to the sequence of interest
then you can see where the protein is being made showing the localization of
the mRNA at presynaptic sites,
what can identify newly synthesised proteins form mature proteins
PLA labeling
how many synapses are there in dendrites per micrometer
4
how many receiving synapses per neuron
10,000
how many output synapses per neuron
100,000
why do we have local protein synthesis
morphological complexity, the volume, and the capacity of the neurons
Remember it takes a day to get from the soma to the axon therefore we need local protein synthesis
Therefore it is about getting that processing of information within minutes not hours/days
how many different proteins are there at a synapse
2500
why does the brain use so much energy
because of the protein demand
