Neurons and glia

Question 1

What important step is needed in order to slice/section the brain?

Answer 1

It needs to be stored and firmed up by paraformaldehyde.

Without it, the brain is a similar softness to raw chicken (if we put a bit of pressure on it, it deforms, so we can’t get clean slices).

Question 2

How can we slice a brain using a microtome?

Answer 2

You embed the brain in wax in a particular orientation, then you mount it in the microtome and slice it

Question 3

How can we slice a brain using a cryostat?

Answer 3

You freeze the brain, then slice it in the cryostat

Question 4

What do we need in order to see small slices?

Answer 4

Staining methods

Question 5

What does nissl stain do?

Answer 5

Stains RNA using purple dye

Question 6

What does golgi stain do?

Answer 6

Uses silver chromate and allows us to see the cell body and also the dendritic tree, as well as the beginning of the axons to some extent.

Question 7

Why can you draw all entire cells with the Golgi technique?

Answer 7

With the Golgi technique you can draw entire cells, but you can only do it because so few cells actually stain.

Question 8

What do solvents do in staining?

Answer 8

Get rid of the myelin to increase the clarity (make the brain more transparent)

Question 9

What is biocytin?

Answer 9

If you end up with a thin enough sharp enough point you can infuse the cell with a tracer. A classic one which is used a lot is biocytin.

Question 10

How does biocytin work?

Answer 10

The cells are so small that you still would be unable to see the cell, so you have to take your biocytin and visualise it in a way that magnifies its existence. The classical technique you’ll find in a lot of papers is immunocytochemistry.

Question 11

How does immunocytochemistry work?

Answer 11

You raise a set of antibodies against your tracer, and then raise antibodies against your antibodies so you can put layers of chemicals around your original tracer molecules. To your outer layer of immunoglobins, you add something, like horseradish peroxidase which will catalyse a histochemical reaction which will deposit a black solid substance within the cell wherever the biocytin was.

Question 12

What is one other benefit of using an intracellular injection?

Answer 12

You can record multiple cells with the same electrode

Question 13

What is one downfall of intracellular injections?

Answer 13

They won’t give you long range connections. You can inject an extracellular label into the tissue, which will be taken up by nerve cells and synaptic signals. Then you will be able to see where axons from that one piece of tissue link up. This extracellular label will also be taken up by extracellular terminals. You can see where these axons came from, their shape, and their dendritic pattern.

Question 14

Summarise how electron microscopy works?

Answer 14

• Uses a beam of electrons and a camera in place of light rays and the observer’s eye
• Ultrathin sections, 30-60nm
• Magnification >100,000x
  Resolution to <0.5nm

Question 15

Where are inhibitory/modulatory synapses found?

Answer 15

On dendritic shafts and soma, excitatory ones on the spine heads.

Question 16

How do synapses increase their strength to learn a new skill or lay down a new memory?

Answer 16

They change in the size, shape and number of spines

Question 17

Summarise some features of microtubules?

Answer 17

• Run throughout the cell
• Vital for the transport of materials from the cell body, e.g. structural proteins, neurotransmitter-associated proteins, and organelles (e.g. mitochondria)
• And transport them back to the cell body (signalling proteins, debris and used materials)

Question 18

Summarise some features of actin microfilaments?

Answer 18

• Actin supports the spines and causes the change of shape in the spines as you learn something new to make bigger synapses
• Guides the growth of axons and dendrites, making sure they go to the right place and adopt the right shape
• Produce growth cones at the tip of a growing dendron

Question 19

What happens to abnormal Tau proteins?

Answer 19

Abnormal Tau proteins form dense, neurofibrillary tangles associated with Alzheimer’s disease

Question 20

What can we do with the fluorescent genes we isolate from jellyfish?

Answer 20

With modern techniques you can put it into an animal and instruct the genes to only activate that protein under very specific circumstances

We can get the cells to make their own label, so you don’t have to cut the brain in sections. If the brain doesn’t have to be cut, and you can make the brain transparent, then you can end up with a block of transparent tissue, and in it all of the cells of a particular type are glowing. They connect up in the way they’ve always connected up.

Question 21

Where are glial cells made?

Answer 21

These were all made by the neural tube by the same stem cells that make nerve cells, but later in development they switch from nerve cells to glial cells

Question 22

What do astrocytes do?

Answer 22

• Control the movement of materials into and out of the tissue
• Endothelial cells in the capillary have big gaps. In the brain they have tight junctions, so the transport can be very tightly controlled. Some smaller lipid-soluble molecules have diffuse through, but most things can’t. it is the astrocytes which cause the capillaries to form the blood-brain barrier in the nervous system
• CSF runs down along the tracks of the capillaries into the neural tissue from the thin layer of the meninges, and these cells pump fluid through the neuropil. That flow of fluid is believed to remove debri and waste products from the neuropil. It happens when you are asleep. We think it has a protective effect against things like Alzheimer’s
• Create a blanket around the nerve cells and control electrolyte levels in the intervening extracellular space. They remove used neurotransmitters in the fast synapses.
• They link together in tight chain gaps, and can transport substances over a distance, from the capillaries to the nerve cells. They build precursors up to complex molecules and break things down in the opposite direction

Question 23

What do neurolemmocytes do?

Answer 23

• Myelinate the axons running down the peripheral nerves

- Only works on one axon at a time

Question 24

What are microglia?

Answer 24

Immune cells that migrate into the CNS very early in development

Question 25

What do microglia do?

Answer 25

• Helps direct the development of neurones, and constantly monitor their and other cells’ health thereafter
• Become amoeboid and travel to areas of injury and infection
• Engulf and eliminate microbes, damaged cells, and other particulate matter
• Secrete factors essential for recovery and repair