Chapter 2

Neurophilosophy

• No separation of mind and brain

• Mind, body and soul are not different things

• Monism

• W. Richie Russell (from Brain Memory Learning: A Neurologist's View, 1959)

Cell Theory

All tissue is composed of microscopic units called cells

• Theodor Schwann (1839)

Golgi Stain

cell body (soma)/perikaryon

Perikaryon

central region containing nucleus

Golgi’s Drawings

showed neurons in a reticulum (network), hence “Reticular Theory”

• rat hippocampus

The Neuron Doctrine

The principle that individual neurons are the elementary signalling elements of the nervous system

• Santiago Ramon y Cajal (1852-1926)

Cajal’s contribution:

• Neural circuitry

• Neurons communicate by contact, not continuity

Neuron Doctrine:

• Neurons adhere to cell theory

• Use of Golgi stain

Franz Nissl (~1885)

• Medicine

• Pathologist

Nissl Stain

• Fixation – alcohol

• Cresyl violet

• Nissl bodies - rER

Histology

The microscopic study of tissue structure

Advances in Fixation Procedures: The Nissl Stain (1800s)

• Stains rER in the cell body and dendrites

• Facilitates the study of cytoarchitecture in the CNS

• Does not produce much detail about the structure of the neuron

What is the relationship between Nissl bodies and the rough endoplasmic reticulum (rER)?

• Nissl bodies are the neuron-specific, visible form of rough endoplasmic reticulum + ribosomes

• They are crucial for producing the proteins neurons need to function

Cell Body (Soma)

Nucleus

• typically gives rise to a single axon

Neurites

• Axons

• Dendrites

Unipolar

• Single process (invertebrate)

• Specialized segments

Bipolar

• Dendrites carry info to the cell body

• Axon transmits it to other cells

Multipolar

Dominate vertebrate nervous system

Single Neurite

Unipolar

Two or More Neurites

• Bipolar - two neurites

• Multipolar- more than two

Microscopy

• Light microscope

• Electron microscope

• Fluorescence microscope

• Confocal microscope

• Two-photon excitation microscope (2P scope)

Classifying Neurons

Based on dendritic and somatic morphologies:

• can be spiny or aspinous

Further Classification:

By connections within the CNS (e.g.):

• Primary sensory neurons,

• Motor neurons,

• Interneurons

Based on neurotransmitter type:

• e.g., Cholinergic = Acetycholine at synapses

Stellate Cells

star-shaped

Pyramidal Cells

pyramid-shaped

Dendrites

• “Antennae” of neurons

• Dendritic tree Synapse—receptors

• Dendritic spines:

  • Small protrusions of membrane

  • Postsynaptic (receives signals from axon terminal)

Classification Based on Gene Expression

Differences between neurons occur at the genetic level

• Green Fluorescent Protein

  • GFP

  • Jellyfish

  • Transgenic mice can now be created in which genes are inserted into a cell under the control of a particular promoter that is distinct from that cell type

Function of Glia

non-neuronal cells in the nervous system that support, protect, and nourish neurons, maintaining homeostasis and modulating neural signalling

Types of Glial Cells

• oligodendrocytes

• schwann cells

• astrocytes

The Myelinating Glia

• Oligodendroglia (in the CNS)

• Schwann cells (in PNS)

• Insulate axons

Node of Ranvier

• Region where the axonal membrane is exposed

• Channel proteins in the node

• Action potential regenerates

Astrocytes

• Most numerous glia in the brain

• Influence neurite growth

• Regulate the chemical content of the extracellular space

  • Remove substances

  • Release substances

Astrocytes-neuron Communication: Metabolism

• Store glucose as glycogen

• Supply neurons with an alternative form of energy

  • Glucose metabolized

  • Take up glucose from vessels

  • Breakdown glycogen and supplies lactate and pyruvate to neurons (converts to energy)

Microglia

• Phagocytic

• Injury and inflammation

• Harm or help?

  • Pro-inflammatory and anti-inflammatory properties and signals

  • Pro-inflammatory cytokines can trigger neuronal cell death

• Not only injury: Stress

• Model of reactive microgliosis driving neurotoxicity in Parkinson’s Disease

Microglia Activation and Alzheimer’s Disease

• Cagnin et al. (2001)The Lancet

• [11C]-PK11195: Peripheral BZP binding site present on activated microglia

• AD: Entorhinal, temporoparietal, and cingulate cortex

Prototypical Neuron

The SOMA:

• Cytosol: Watery fluid inside the cell

• Organelles: Membrane-enclosed structures within the soma

• Cytoplasm: Contents within a cell membrane (e.g., fluid, organelles)

The NUCLEUS:

• Chromosomes

• Genetic information

Cytosol

Watery fluid inside the cell

Organelles

Membrane-enclosed structures within the soma

Cytoplasm

Contents within a cell membrane (e.g., fluid, organelles)

Chromosomes

Genetic information

DNA

deoxyribonucleic acid (the blueprint):

• Same genetic information can be found in cells throughout the body

• But more of the total genetic info encoded in DNA is expressed in the brain (10-20x more than liver or kidney cells)

Gene

A sequence of DNA that encodes a single polypeptide or protein

Watson and Crick (1953)

Based on the X-ray crystallography data of Franklin:

• Two intertwined (polynucleotide) chains (double helix)

• Connected together by bonding of hydrogen atoms

DNA Backbone

Forms the structural framework of DNA:

• consists of alternating deoxyribose sugar and phosphate groups linked by strong phosphodiester bonds

Bases

Purine and pyrimidine bases are on the inside of the double helix:

• Purine base on one chain is always hydrogen-bonded to a pyrimidine on the other chain

• Purine and pyrimidine bases selectively bond

  • Thymine -> Adenine

  • Cytosine -> Guanine

Pyrimidines

Thymine and Cytosine

• Smaller

Purines

Adenine and Guanine

• Larger

Hydrogen Bond Stability

T → A

• 2 hydrogen bonds

C → G

• 3 hydrogen bonds

• The hydrogen bonds are the weakest links

• So many base pairs that it doesn’t spontaneously unravel

• Heat (boiling) can separate the double helix into two complimentary chains (denaturation)

Gene Expression

• ‘reading of DNA’

• Product is proteins

• happens in the nucleus

Transcription

Assembling the RNA info of the gene:

• happens in the nucleus

• RNA molecules are synthesized from the DNA template by RNA polymerase

• RNA processing

• mRNA (messenger) carries the information from the nucleus to the cytoplasm

*GENES are sequences of coding (exon) and non-coding (intron) nucleotide triplets

RNA Processing

Splicing

• happens in the nucleus

Translation

Assembling of proteins from amino acids

• Happens in the cytosol/cytoplasm

Protein Synthesis

• Construction of proteins

• Occurs in the cytoplasm

• DNA never leaves nucleus

Promoter Region

• Transcription is initiated

• RNA polymerase would bind to this region to initiate transcription

• Controlled by transcription factors

Terminator Region

RNA polymerase recognizes the signal to end transcription

Introns

Segments of the gene that do not code for proteins

Exons

Segments of the gene that DO code for proteins

RNA Splicing

The processing of removing introns and splicing together exons

• Sometimes exons are also removed:

  • Alternative splicing

  • One gene can produce multiple proteins

In the RNA transcript:

Uracil replaces Thymine

The Cytoplasm

mRNA transcripts leave nucleus through pores

Rough Endoplasmic Reticulum (rER)

• Major site for protein synthesis

• In the cytoplasm

• Membrane-bound proteins

• Threaded through the membrane of rER

rER ‘Nissl Bodies’

more in neurons than any other cell type

Free Ribosomes

free

Polyribosomes

several ribosomes floating free in cytoplasm, connected by mRNA strand

Translation: Ribosomes

• Protein synthesis in neurons

• Free Ribosomes

• Rough ER (rER):

  • Membrane bound proteins

  • Threaded through membrane of rER

Translational Processes

• Ribosomes begin to read the RNA from the 5’ end

Codon and tRNA:

• 3 nucleotide bases code amino acids

Smooth ER and Golgi Apparatus

• In the Soma

• Sites for preparing/sorting proteins for delivery to different cell regions (trafficking) and regulating substances

Protein Destiny

Each nerve cell makes only 3 classes of proteins:

1. Proteins synthesized in the cytosol- and stay there

2. Proteins synthesized in the cytosol but later incorporated into the nucleus and mitochondria

3. Proteins synthesized in association with membrane systems

   1. Remain attached to the membrane of rER, the GA (and vesicles)

   2. Remain in the organelle (eg. rER or GA etc) - not attached to membrane

   3. Transported by means of vesicles from the GA or other organelles - can become secretory products (e.g. neuropeptides!)

Epigenetics

The study of changes in gene expression without changing the genotype of the organism:

• Change in phenotype without a change in genotype

• Environmental influences that can turn genes on or off

• These changes may and/or may not be heritable

• Ex. Highly nurtured rats grow up to be calm adult rats

Altering Levels of Anxiety in Rats

The amount of licking a rat pup receives from its mother can alter the expression of glucocorticoid receptors (GR)

Histone Remodeling

Involves modifications to a histone protein (around which DNA is coiled) and can either decrease or increase gene expression.

DNA Methylation

Involves the attachment of a methyl group to DNA and tends to reduce the expression of adjacent genes.

CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)

A new technique that enables researchers to modify specific genes inside the cell:

• Application of a technique that is part of the way bacteria protect themselves from invading viruses

• Allows researchers to target individual genes for deletion and replacement with a different gene

• May be useful to fix broken genes, like those in Huntington disease or Tay-Sachs disease

Mitochondrion (1 um)

• Site of cellular respiration

• ‘Inhale’ pulls in pyruvic acid and oxygen

• Krebs cycle

• ATP → cell’s energy source

• ‘Exhale’ →17 ATP molecules released for each pyruvic acid molecule

The Cytoskeleton

• Not static

• Internal scaffolding of neuronal membrane

• Three “bones”

  • Microtubules (20 nm)

  • Neurofilaments (10 nm)

  • Microfilaments (5 nm)

Microtubules

20 nm

Neurofilaments

10 nm

Microfilaments

5 nm

The Axon

• Axon hillock (beginning)

• Axon proper (middle)

• Axon terminal (end)

Axon Hillock

beginning

Axon Proper

middle

Axon Terminal

end

Differences between axon and soma:

• ER does not extend into axon

• Protein composition:

  • Unique (different from the soma)

Differences between the cytoplasm of axon terminal and axon:

• No microtubules in the terminal

• Presence of synaptic vesicles

• Abundance of membrane proteins

• Large number of mitochondria

The Synapse

Where a neruon makes synaptic contact-innervation

Presynaptic

The axon terminal

Postsynaptic

• Axo-dendritic

• Axo-somatic

• Axo-axonic (rare)

Synaptic Cleft

Synaptic transmission:

• Electrochemical signal

• Electrical-chemical-electrical events

Modification of the synapse can occur in many forms:

• Plasticity

• Drug actions

• Toxins

Axoplasmic Transport

Membranes and secretory proteins are actively transported:

• Synthesis - cell body

• Travel (up to 0.5 - 1 m/day)

FAST Axonal Transport

• Large particles move in a ‘saltatory’ manner

• Radioactive studies

• Dependent on ATP (energy-consuming)

• Independent of the cell body:

  • Axon will not survive

  • Transport can still occur

• Anterograde

• Retrograde

• Microtubules → tract

• The motor molecule → kinesin

SLOW Axonal Transport

• Depends on the filaments that make up the cytoskeleton

• Cytoskeletal elements, soluble proteins, and some metabolic enzymes

Two components:

• Slow: 0.2-2.5 mm/day

  • Carries: components of the neurofilaments & parts of microtubules

• Fast: 2x as fast as the slow

  • Some actin (microfilament)

Does not occur in the retrograde direction

Axoplasmic Transport: Vesicles

• Proteins are synthesized and incorporated in ER and GA

• Fast axoplasmic transport to the terminal

• Vesicles and precursors reach synapse-exocytosis

• Degraded portion is taken back to cell body-fast retrograde

• Degraded material is recycled

Kinesin

The motor molecule:

Form cross-bridges:

• Microtubule-associated protein

• Little feet walking along the microtubule:

  • Two motor heads and two neck linkers tether to organelle/vesicle

Motor movement of kinesin:

• Motor head has ADP molecule bound

• Binding of motor head to microtubule causes ADP release

• Molecule of ATP now binds, triggers neck linker zipper action onto the core

• Throws the second motor head forward to further bind to the microtubule (ATP hydrolyzed back to ADP)

• Active process: 1 ATP molecule step/2 ATP molecules cycle

Retrograde Fast Transport

• Returning materials to the cell body:

  • Degradation

  • Reuse

• Packaged in large membrane-bound organelles (lysosomal system)

• Slow (though still fast)

  • 1/2 to 1/3 as fast as anterograde

• Dynein → motor molecule

  • Microtubule-associated protein

• Viral infections and labelling

• Tracing with Horseradish peroxidase (labels neurons)

Tau (Microtubule Associated Protein)

The primary role is maintaining the stability of microtubules:

• interacts with tubulin proteins and promotes the formation of microtubules

Tau Protein and Alzheimer’s Disease

• Tau protein and hyperphosphorylated tau

• Neurofibrillary tangles (NFT) are the most associated with cognitive decline in AD