Neurons and Glia Flashcards
What substance is used to harden the brain?
Paraformaldehyde
What is the tool used to slice the brain?
Microtome
What type of sections is the brain sliced into?
Sagittal and Coronal
Imbed the brain in different orientations and get thin slices
What is used to freeze the brain and why?
Crysostat is used to freeze the brain and then section it
What type of tissue cannot refract light?
Thin tissue so no contrast can be seen
What are the two different staining methods?
Nissel Stain
Golgi Stain
What is Nissel Staining?
- First stain developed
- Stains RNA with purple dye so pyramidal images obtained
What is Golgi staining?
- Uses silver chromate
- Labels some neurons
- Not very effective at initially labelling neurons but once labelled, does it in its entirety
What is the gap junction theory?
One neuron coupled to another by membrane proteins that span both cells.
Why are gap junctions important?
- Charge flows through the neurons to another directly
- Heart has gap junctions
- Some cells in the brain communicate via gap junctions: astrocytes
Which cells in the brain communicate via gap junctions?
Astrocytes
What is the synaptic theory?
When AP arrives at the synapse, it converts electrical signal to chemical signal.
Why is the synaptic theory important?
- Chemical signal interacts with receptors post-synaptically
- Enables influx/efflux
- Generation of second AP
What is fluorescent neurohistology?
- Use of particular light frequencies shined through a sample -> if the sample expresses green fluorescent protein (GFP), use blue light. If it emits red light, it can be detected as a light source
What is removed from the brain to allow more transparency?
To allow for clarity - fatty tissue (myelin) is removed from the brain
How is green fluorescent protein obtained?
- Jellyfish express GFP
- Genetic code for GFP was identified
- Inserted into organisms and these were made fluorescent also
- They can express multiple fluorescent proteins and express many different wavelengths of light
What are opsins?
Light emitting proteins
What are the two ways opsins can be targetted to specific cells?
Viral delivery
Cre/lox technology
Explain the viral delivery of fluorescent proteins
- Sequence GFP genetic code
- Take DNA for GFP and package into the virus
- Take out viral DNA
- Fluorescent protein DNA code is inserted into viral DNA code along with a promoter
- The promoter determines the cell types the virus can infect
- The virus infects the neurons and inserts the GFP code into the DNA of the neuron.
- The DNA transcripts and translates the GFP DNA to produce the fluorescent protein
Which promoter is required to target which cell types?
CMV/CAG -> All neurons and glia
GFAP -> Glia only
hSYN -> Neurons only
CamKII -> Excitatory neurons only
Explain the use of cre/lox technology for fluorescent proteins
- Cre recombinase is an enzyme that recognises loxP sites on DNA
- When the Cre enzyme encounters this region, it depends on the orientation of the site on what happens:
- > Inversion: if two sites facing each other - gene is flipped, reverse order
- > Deletion: if facing the same way: completely cuts out the gene in between them
- > Translocation: if sites on different strands of DNA, cuts and swaps them.
Why can different modifications take place when using cre technology?
Different genetic modifications take place dependent upon loxP location/orientation
Why is cre/lox method favoured over the viral method?
It is more selective than viral method and can target subtypes of cells.
Describe the structure of the prototypical neuron
- The neuronal ‘body’ containing K+ rich cytosol which is important for establishment of resting membrane potential and generation of ATP.
- The nucleus for DNA replication and DNA transcription
- Endoplasmic reticulum for RNA translation
- Golgi apparatus for protein folding
- Mitochondria as the powerhouse of the cell
What type of receptors do dendritic proteins contain and what does this mean for neurotransmitter release?
Ligand gated and GPCR expressed on dendrities; neurotransmitter is released presynaptically onto dendrities postsynaptically so need receptor sites
What type of receptors do axonal proteins have and why?
Voltage gated ion channels in the nodes of Ranvier to allow AP propogation
Structure of microtubules in neurons
- Relatively large (20nm diameter)
- Run the length of neurites: dendrites and axons
- Tubulin composition which is spiral shaped
Function of microtubules in neurons
- Vital for transport of materials from the cell body such as structural proteins, neurotransmitters and organelles
- Also transport towards the cell body such as signalling proteins, debris and used material
Structure of actin microfilaments in neurons
- 5nm diameter which is the same as the neuronal membrane
- Numerous in neurites
- Actin composition
Function of actin microfilament in neurons
- Provides support, helping to maintain the shape of the cell body and neurites
- Plays a vital role in neural embryonic growth and helping to shape axons and dendrites
- Actin skeleton within the filopodia grows or shrinks in response to chemical guidance signals
What are tau proteins?
Neurofilament protein that binds together the cytoskeletal elements
What can pathological problems with tau proteins cause?
Alzheimer’s and Parkinson’s disease
What are intermediate filaments?
- Composed of five proteins: NFL, NFM, NFH, internexin and peripherin
- 10nm in diameter
- Protein combinations dependent upon neuronal cell type and development stage
What is the axon hillock?
Where EPSP and IPSP summate and an AP is fired/inhibited
How long are axons?
1mm to over 1 metre in length
What do axon branches form?
They form collaterals these are the main branches away from the main axon. They are at right angles to the main axon
What is the terminal bouton?
The end of the axon terminal
A bouton en passant
A synapse along the length of an axon
Where are synaptic boutons?
They can be anywhere along the axon, they do not have to be at the end of the dendrite. Lots of axonal connections happen within the dendritic tree
What does the axon terminal contain?
- Terminal bouton
- Many mitochondria as an energy rich process
- Synaptic vesicles
Function of synaptic vesicles
signal proteins on the surface so when the AP arrives, senses signal and receives instruction to fuse with the synapse membrane
Why don’t axons have ribosomes?
Do not have ribosomes as they need space for mitochondria and vesicles
What transports the vesicles?
Kinesin
How does the axon receive other necessary things?
Via transport down the cytoskeleton and microtubules by kinesin proteins
What are Kinesin proteins?
ATP activated proteins that ‘walk’ along microtubules carrying whatever the synaptic bouton needs
What is anterograde transport?
From cell down the axon
What is retrograde transport?
From bouton back to soma
Why is retrograde transport used?
Useful for unwanted components as they need breaking down
How are neurons classifed?
Classified by structure and gene expression
What part of the structure is looked at to classify neurons?
- Number of neurities (axons and dendrites)
- Dendrities - dendritic tree formation; presence of spines; single dendrite not receiving from a single synapse
- Connections: primary sensory neurons, motor neurons and interneurons
- Axon length
What part of gene expression is looked at to classify neurons?
- Types of proteins
- Types of neurotransmitter: GFP can be tagged to a certain protein and neurons expressing that protein are then revealed
How are glia different to neuronal cells?
- Difficult to measure responses
- Come from the same precursors as neuronal cells
- Same amount of glial cells as there are neuronal cells
- Generate resting membrane potential the same as nerve cells but DO NOT generate action potential
- Small in size
- Communicate in slow waves of K+ concentration
What are the 3 types of glia cells?
- Astrocytes
- Schwann cells
- Oligodendrocytes
Describe the structure and function of astrocytes
- Control the movement of material into and out of the tissue
- Make contact with the capillaries which induces them to form the BBB
- Washes waste products from brain by driving the flow of fluid in and out
- Regulate contents of the extracellular space
- Express receptors
- Releases neurotransmitter
- Control the biochemical environment of the nerve cell
How do astrocytes control the biochemical environment around a nerve cell?
- Form a blanket around the soma and dendrities
- Control electrolyte levels in the intervening extracellular space
- Remove used neurotransmitters from synaptic clefts
- Control synaptic development and function
- Can be activated upon synaptic transmission of neurotransmitter
Difference between oligodendrocytes and schwann cells
Oligodendrocytes myelinate neurons in the CNS and schwann cells in the PNS
How do the oligodendrocytes and schwann cells myelinate the cell?
They wrap their own extracellular membrane around the nerve cell, leaving layers of the phospholipid membrane
Function of oligodendrocytes and schwann cells
- Ensure that the nerve cell is well insulated and does not leak ions
- Speeds up conduction
- Secrete growth factors and inhibitory factors that control the axon regeneration after injury
What does damage to oligodendrocytes cause?
Multiple sclerosis
Why is it important that axons are myelinated?
- Acts as an insulation
- Stops dissipation of current
- Speeds up AP
Difference between microglia and glia
- Not actual glial due to them arising from the mesoderm rather than the neural tube
What are microglia?
Immune cells that migrate to the CNS early in development
Function of microglia
- Help the development of neurons and monitor neuronal health
- Become amoneboid and travel to areas of injury
- Engulf and eliminate microbes, damaged cells and other matter
- Secrete factors essential for recovery and repair
- Help to fight Alzheimer;s
- Do the function of immune cells as they are not able to cross the BBB
- Present in deeper tissue
- Perform immunological functions such as act phagocytes and invading pathogens; regulate apoptosis; nurture formation of new cells - neurogenesis
What is neurogenesis?
The formation of new cells
How does neuroglia check the health of nerve cells?
Health checks by interacting with the dendritic spines to make sure everything is working properly - if it isn’t working it induces apoptosis
Other types of non-neuronal cells
Ependymal cells line the ventricles
Vascular cells deliver oxygen and remove CO2
