Neur 602 Cellular Neuroscience

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Neur 602Cellular Neuroscience

• Tuesdays, 130 420 pm
• Instructor
• Dr. Kim "Avrama" Blackwell
• Krasnow Institute, Room 105
• Office Hours Tues 430 530 pm or by
  appointment
• Contact avrama_at_gmu.edu, 993-4381
• Important Dates
• Last Day to Add Sept 15, 2009
• Last Day to Drop Oct 2, 2009

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Reading for Neur 602

• Books
• Required Byrne and Roberts, From Molecules to
  Networks, 2nd Edition, Academic Press, 2009
• Recommended The Neuron, Levitan Kaczmarek
• Other reading
• Journal articles to be downloaded from web

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Goals of Neur 602

• Intense introduction to the structure and
  function of individual neurons
• how nerve cells integrate and transmit signals
• biochemical properties of single neurons
• electrical and chemical communication between
  neurons
• Provide foundation for more in-depth study of any
  aspect of cellular neuroscience

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Brief Syllabus of Neur 602
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Brief Syllabus of Neur 602
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Course Requirements for Neur 602

• Synopsis of assigned papers 10 of Grade
• Math Problem sets - application of neuroscience
  equations 10 of Grade
• Midterm 40 of Grade
• Final 40 of Grade
• Make up exams are not allowed, unless a doctors
  note is provided

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Neur 602 - Class format

• Weekly lectures will be divided in two portions
  of approximately 1hr 15 min, with a 5 min break
  in the middle.
• Lectures will be followed by discussion of the
  paper assigned the previous week, or other
  activities.
• Cell phones must be turned off or to silent mode

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HONOR CODE

• All students are expected to uphold the GMU honor
  code, which prohibits Cheating, Plagiarism,
  Stealing, and Lying.
• The subsequent text is from http//www.gmu.edu/fac
  staff/handbook/aD.html
• In this class, working together on homework is
  permitted, so long as such working together does
  not take the form of copying

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HONOR CODE

• A. Cheating encompasses the following
• 1.The willful giving or receiving of an
  unauthorized, unfair, dishonest, or unscrupulous
  advantage in academic work over other students.
• 2.The above may be accomplished by any
  means whatsoever, including, but not limited to,
  the following fraud, duress, deception, theft,
  trick, talking, signs, gestures, copying from
  another student, and the unauthorized use of
  study aids, memoranda, books, data or other
  information.
• 3.Attempted Cheating.

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HONOR CODE

• B. Plagiarism encompasses the following
• 1.Presenting as one's own the works, the
  work, or the opinions of someone else without
  proper acknowledgement.
• 2.Borrowing the sequence of ideas, the
  arrangement of material, or the pattern of
  thought of someone else without proper
  acknowledgement.
• Plagiarism includes copying as little as a
  sentence or paragraph from published work, or an
  internet webpage without putting the material in
  quotes and giving the source.

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Responsible Conduct in Science

• Beyond the GMU Honor code, the following is not
  tolerated in science
• Fabrication making up data
• Falsification Altering data (there are gray
  areas here)
• Plagarism using others words, ideas or data
  without attribution
• Youll hear more about this in Neur 604 Ethics
  in Scienitific Research

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Falsification

• After analyzing data to generate results,
  removing some data points that are questionable
• If questionable (animals health, uncooperative
  human subjects, other conditions), remove data
  prior to analysis.
• Selecting "best" data to support hypothesis
• What about best data used as figures in paper?
• Pay attention to this aspect of papers

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Methodology

• Consider these points as you read papers
• Descriptive experiments
• Purpose is the build up knowledge base
• Where does neuron project
• What is structure of protein
• What is distribution of neurotransmitter
• Induction
• Develop hypotheses about cause and effect

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Methodology

• Hypothesis testing Essential for NIH grants
• Experiments support (not prove) or disprove
  hypotheses
• Analysis
• Due to variability, can only provide statistical
  probability that effect not due to chance
• Due to highly reduced, often non-physiological
  conditions, limited applicability

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Methodology

• Information presented in this class developed
  from thousands of experiments
• If X is observed under several conditions
  (different species, preparations, paradigms),
  then X may be real.
• If X is consistent with most of our other
  observations, then X may be real
• As new methodologies developed, some truths are
  revealed to be false

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Neuroscience

• Term first used in 1960s
• Included disciplines used to study brain function
  and pathology
• Anatomy-cell shape, connectivity
• Physiology-bioelectric properties
• Biochemistry-subcellular molecules
• Psychology - behavior
• Later, molecular biology and imaging added

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Nervous System Function and Behavior

• In all animals
• Obtain information from the environment
• Process information
• Store information
• Generate behavior
• Additional functions documented in humans
• Feelings
• Aspirations
• Abstract thought
• How and where are these functions performed?

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Systems Level OrganizationAnatomy

• Where are nervous system function performed?
• Each component can be further subdivided!

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Systems Level Organization

• Nervous System has additional subdivisions
• Sensory, motor, cognitive, intrinsic
• Five developmental divisions of CNS
• Neur 601 Map functional subdivisions to
  anatomical subdivisions
• All parts, independent of subdivisions, are
  composed of neurons (and glia)
• Thus, we are studying neurons

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Levels of Organization
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Focus of this Class

• Single Neurons
• Information Processing
• Integration of inputs
• Generation of outputs
• Plasticity
• Some molecules
• Intracellular signalling
• Single ion channels
• Some small networks

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Neuronal function

• Process incoming signals, and transmit signals
• Signals arrive via Synapses
• End of one neuron makes specialized contact with
  membrane of another neuron.
• Signals propagate down dendrites to soma
• Channels in membrane affect how signals are
  combined in dendrites
• Action potentials are generated
• All-or-none signal which travels rapidly down
  axon
• AP causes transmitter release at the end of the
  neuron
• Transmitter carries signal to the next neuron for
  processing.

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Relevance to Neuroscience Disciplines

• Molecular Neuroscience
• Function of molecules important to neurons, such
  as ion channels, signaling pathways
• Computational Neuroscience
• Deeper understanding of ion channel and synaptic
  channel function
• Neuroanatomy and systems neuroscience
• Information processing is shaped not only by
  neuronal connectivity, but also individual neuron
  activity
• Cognitive Neuroscience
• What is producing that BOLD signal???
• Behavioral Neuroscience
• Deeper understanding of action of pharmaceuticals

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Which Neuron Properties are Important for
Behavior?

• Networks / Connectivity / topographic maps
• Can connectivity explain behavior?
• Morphology
• Information processing is influenced by shape
• Electrical Properties
• Information processing influenced by membrane
  channels
• Intracellular signalling

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Brain is not homogeneous structure

• Morphological diversity of neurons
• size few microns to tens of microns
• axons none, or up to 1-2 meters
• dendrites none, or vast branching pattern
• Diversity in Connectivity of Circuits
• Origin, number of incoming fibers
• Destination, number of outgoing fibers

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Brain is not homogeneous structure

• Diversity in Neuron Communication (synapses)
• Electrical synapses (gap junctions)
• Chemical synapses
• Presynaptic vesicles contain different chemicals
• Postsynaptic receptors have different channels

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Brain is not homogeneous structure

• Diversity in Membrane Channels
• Permeability e.g. Na vs K
• Densities
• Voltage or ligand dependent properties
• Intracellular signaling
• Enzymes activated by substances such as calcium
• Enzymes modify different membrane proteins

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End of IntroductionBeginning of Lecture

• Beginning students of neuroscience justifiably
  could find themselves confused Fundamental
  Neuroscience, by Zigmond, Squire et al., Chapter
  1, page 3

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Neurons

• Fundamental unit in the nervous system
• Specialized cell
• Highly active secretory cell
• Highly polarized
• Distinct domains
• Extent defined by Plasma Membrane

Dendrites
Soma
Axon
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Plasma Membrane

• Bilayer of phospholipid molecules
• Composition
• Phosphatidyl serine
• Phosphatidyl choline
• Phosphatidyl inositol bisphosphate
• Substrates in certain enzymatic reactions
• Will be discussed in neuromodulation

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Plasma Membrane

• Function
• Separates intracellular and extracellular
  contents
• Maintains concentration gradient
• Keeps organelles Inside
• Electrical insulator
• Prevents flow of current (charged ions)
• Capacitor
• Allows charge imbalance, which allows electrical
  field

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Plasma Membrane

• Anchors integral proteins
• Enzymes
• Channels
• Other proteins
• Exocytosis
• Release of vesicles of neurotramitters

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Neuron Structure

• Soma
• Cell Body or perikaryon
• Contains Nucleus Transcription and protein
  synthesis
• Functions in Signal integration
• Contains major cytoplasmic organelles

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Soma Organelles

• Nucleus
• Most prominent organelle
• Contains DNA - genetic material
• High level of transcription in neurons
• Ribosomes free or ER associated
• Protein translation

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Soma Organelles

• Rough Endoplasmic Reticulum
• Intracytoplasmic membrane
• Ribosomes attached
• Functions in Protein Translation
• Golgi apparatus
• Intracytoplasmic membrane
• Post-translation modification of proteins
• Packaging of proteins for transport

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Soma Organelles

• Smooth Endoplasmic Reticulum
• Calcium storage and release
• Smooth because no ribosomes
• Mitochondria
• Energy Production
• Calcium storage
• Apoptosis - programmed cell death

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Nissl Substance
Endoplasmic Reticulum, and Free Ribosomes
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Nissl Substance

• Parallel rows of rough endoplasmic reticulum
• Higher density than remainder of cell
• Membranous framework for other components
• Reticulum consists of tubules, strings of
  vesicles, numerous large and flat cisternae
• Polysomal Rosettes
• Clusters of ribosomes, "free" of ER

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Neuron Structure

• Dendrites
• Long, thin, tubular structures arising from soma
• Structure maintained by cytoskeleton
  (intracellular) and adhesion molecules (surface
  interactions)
• Continuously branch and taper distally
• Multiple dendrites and branches provide huge
  increase in surface area
• Receive hundreds to thousands of inputs
• Convergence (inputs from many other neurons)
• Develops after axon
• Varying numbers and types of channels

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Protein Synthesis in Dendrites

• Dendrites contain Nissl substance
• Some proteins synthesized predominantly in
  dendrites
• Microtubule-associated proteins
• Some mRNA excluded from dendrites
• Some proteins targeted to dendrites
• e.g. Those associated with post-synaptic density

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Dendrites

• Surface is sometimes covered with spines
• Spiny versus non-spiny dendrites
• Spines are input structures of dendrites
• Small protrusions of dendritic membrane
• 1-2 mm length
• lt 1 mm diameter
• Narrow "neck" attached to dendritic shaft
• Wider, spherical "head" at distal end
• Dramatically increase dendritic surface area

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Spine Ultrastructure

• Microtubles and neurofilaments in dendritic shaft
  do not extend into spine
• Fine, indistinct filaments of Actin and a or b
  tubulin
• Dendritic polyribosomes are clustered at base of
  spine in association with ER
• Local synthesis of spine proteins
• Spine apparatus
• Tubular cisterns that are extension of dendritic
  smooth ER into the spine

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EM of Dendritic Spine
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Spines

• Prominent feature is asymmetric synapse on distal
  aspect
• Presynaptic element
• Specialized part of Presynaptic neurons axon
• Postsynaptic element
• Specialized part of postsynaptic neuron
• Usually part of soma or dendrite
• Cleft
• Narrow space between two closely apposed elements
• Spanned (stabilized) by adhesion molecules

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Synapse

• Pre-synaptic element
• Vesicles and thickening of membrane
• Post-synaptic element
• Thickening of membrane, Few vesicles
• If large, may have discontinuity perforated
• Symmetric synapse (usually dendritic)
• Both sides equally thick, usually inhibitory
• Asymmetric synapse
• Post-synaptic is thicker, usually excitatory

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Byrne Figure 1.3 Dendritic spines (S) and
synapses in the human brain. narrow spine necks
(asterisks) emanating from the main dendritic
shaft (D). Cisterns of the spine apparatus
visible in lower panel spine. Big arrow
postsynaptic densities of asymmetric excitatory
synapses. Axonal boutons (B). A perforated
synapse marked with small double arrow in lower
left panel. Right symmetric inhibitory synapses
(arrowheads). In this case the axonal boutons (B)
contain some ovoid vesicles compared.
mitochondria (m). Scale bar 1 µm.
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Neuron Structure

• Axon
• Longer, thinner tubular structures
• Do not taper
• Minimal branching
• Output structure
• Allow for communication over long or short
  distances
• Divergence to many other neurons

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Axon

• Specialized for information transfer between
  nerve cells
• Thin tube-like process that arises from cell body
• Travels for distances ranging from mm to meters
• Between 0.1 to 20 mm diameter in vertebrates
• Each structural feature designed to optimize
  information transfer

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Axon Structural Features

• Axon Hillock
• Axon Initial Segment
• Branching structure
• Axon Terminal
• En passant boutons
• Myelin

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Saltatory Conduction

• Myelin insulation increases current flow axially,
  decreases leak across membrane
• Current reaches Node of Ranvier
• Causes small depolarization
• Activates sodium channels
• Regenerative action potential produces large
  current
• Current flows axially to next Node

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Axon Hillock

• Region of cell where axon originates
• Deficiency of Nissl substance
• No protein synthesis here
• Microtubules and neurofilaments begin to align

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Axon Hillock

• Region where material is "sorted"
• Cytoskeletal elements, synaptic vesicle
  precursors, mitochondria are committed to axon
• RER, dendritic microtubule associated proteins,
  free polysomes are excluded from axon
• Molecular mechanism of sorting is not known
• No "sizing" barrier evident

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Initial Segment

• Region of axon adjacent to hillock
• Microtubules form characteristic bundles called
  fascicles
• Distinctive plasma membrane
• Electron-dense "coating" separated by 5-10 nm
  from inner surface
• Highest density of membrane channels
• Most action potentials originate here

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Branching

• Child branches have comparable diameter as parent
  branch
• Branches can be proximal
• Cortical axons may have "recurrent" branch that
  innervates local area, and another branch that
  travels far
• Branches can be distal
• Motorneurons branch to innervate many muscle
  fibers

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Axon terminals

• Specialized area where neurotransmitter is
  released
• Multiple terminals may innervate a single neuron
• Terminals may contact widely dispersed neurons
  (ramified)
• Recurrent collaterals
• Projections to same layer as cell body
• En passant synapses - in passing
• Axonal protrusions forming synapses prior to the
  terminal

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Myelin

• Produced by glia
• Oligodendrocytes in CNS
• Schwann cells in PNS
• Surrounds many, but not all axons
• Absent from local circuit neurons
• Prominent for long range connections
• Acts as electrical insulator
• Increases conduction speed

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Myelin - EM
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Myelin

• Tight wrapping of cell membrane around axon
• Cytoplasm of glial cell is gradually squeezed out
  as cell wraps around
• As many as 20 wraps
• Result is concentric layers of closely apposed
  membrane
• Alternating light and dark bands on EM

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Myelin in PNS

• A single Schwann cell is 1 mm wide, wraps single
  axon
• Long axons have multiple Schwann cells
• Gaps between Schwann cells
• Nodes of Ranvier
• Several micrometers wide
• Huge effect on speed of action potential
  transmission

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Unrolled Schwann Cell
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Myelin in PNS

• Schwann cells secrete extracellular matrix
  components
• Associated collagens (produced by fibroblasts)
  prevent compression damage to nerves passing
  between muscles or around joints
• Respond to injury (as astrocytes), remove debris
  after injury

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Myelin in CNS

• Oligodendrocytes wraps several axon segments
• Reduce number of glia required
• Have nodes of Ranvier (similar to PNS)
• Lamellae are 30 thinner than in PNs
• Minimal extracellular space or matrix
• Do not respond to injury

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Myelin in PNS

• Major integral membrane protein is P0
• One immunoglobulin-like domain
• One transmembrane segment
• Highly charged cytoplasmic domain
• Membrane phospholipid head groups are highly
  charged
• P0 neutralizes charge and allows close apposition
  of membrane layers

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Myelin in PNS

• Major integral membrane protein is proteolipid
  protein (PLP and DM-20)
• Four transmembrane segments
• Axonotropic function
• Replacing PLP with P0 in rodent enhances axonal
  degeneration
• Demyelination and subsequent axonal degeneration
  is cause of multiple sclerosis