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Psych 211 (neuro) > Animal research methods > Flashcards

Animal research methods Flashcards

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1
Q

What are the ways to study animal brains?

A
  • neuronal tracing
  • microelectrode EEG
  • electrical stimulation
  • pharmacological manipulation
  • optogenetics
  • calcium imaging
  • excitotoxic lesions
  • radio lesions
  • microdialysis
  • immunohistochemistry
  • cFos immunohistochemistry
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2
Q

What is experimental ablation?

A
  • lesion study
  • the removal or destruction of a portion of the brain
  • functions that can no longer be performed following the surgery are probably controlled by that brain region
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3
Q

What are radiofrequency lesions?

A
  • small lesions made by passing radiofrequency current through a metal wire that is insulated everywhere but the tip
  • burns cells around the tip of the wire
  • size and shape of lesion, determined by the duration and intensity of the current
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4
Q

What is a downside to radiofrequency lesions?

A
  • axons just passing through will also be burned
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5
Q

What is an excitotoxic lesion?

A
  • injection of a glutamate receptor agonist
  • cause so much excitation (and calcium influx) that the affected neurons undergo apoptosis
  • axons passing through are spared
  • permanent lesion
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6
Q

What is a sham lesion?

A
  • placebo procedure
  • duplicates all steps of producing a brain lesion except for the step that causes extensive brain damage
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7
Q

What is a reversible lesion?

A
  • temporary brain “lesion”
  • injecting drugs that block or reduce neural activity in area
  • permanent
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8
Q

What are common drugs to use for a reversible lesion?

A
  • voltage-gated sodium channel blockers (stops all action potentials, affects axons passing through)
  • GABA receptor agonists (hyperpolarize cell bodies, does not affect axons passing through)
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9
Q

What is the most direct measurements of neural activity?

A
  • made with metal wires placed in the brain
  • macroelectrodes
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10
Q

What are macroelectrodes?

A
  • thin metal wires with a fine tip
  • record the electrical activity of individual neurons (single-unit recordings)
  • used in behaving animals to record every action potential from a given neuron
  • record from hundreds of single neurons simultaneously
  • implanted in brain during stereotaxic surgery
  • wires connected to socket in animal’s head so that they can be ‘plugged in’ to a recording system at any time
  • permanently attached set of electrodes, with a connecting socket cemented to the skull
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11
Q

What are chronic electrical recordings?

A
  • made over an extended period of time
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12
Q

What are acute electrical recordings?

A
  • made over a relatively short period of time
  • often during surgery when the animal is anesthetized
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13
Q

What can we know when we manipulate neural activity?

A
  • how the activity of specific receptors or cell populations influence behaviour
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14
Q

What are ways to manipulate neural activity?

A
  • electrical stimulation
  • chemical stimulation
  • optogenetics
  • viral-mediated gene delivery (record + manipulate)
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15
Q

What is electrical stimulation?

A
  • passing an electrical current through a wire inserted into the brain
  • affect everything in the area (even passing)
  • fast stimulation frequencies counterintuitively produce the same behavioural effects as lesioning the brain area
  • stop action potentials
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16
Q

What is chemical stimulation?

A
  • drugs
  • administered through a guide cannula (hollow tube) implanted in a particular brain region
  • anesthetics to shut down all neural activity
  • receptor agonist/antagonist can be used (don’t affect passing)
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17
Q

What are optogenetics?

A
  • use light to depolarize and hyperpolarize neurons with millisecond precision
  • turn up or down the activity of specific cells or receptors
  • use of light to control neurons that have been made sensitive to light through the introduction of foreign DNA
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18
Q

How do optogenetics work?

A
  • foreign DNA provides instructions to make light-sensitive proteins
  • opsins we use to manipulate neural activity (optogenetic techniques) are often ion channels that open and close instantly in response to light
  • light sensitive protein (ion channel that opens in response to light)
  • take gene for protein
  • insert DNA into specific neurons in the brain
  • neurons communicate by firing
  • can cause neurons to fire by flashing light
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19
Q

What are opsins?

A
  • proteins that are activated by light
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20
Q

What are types of photosensitive ion channels that evolved in bacteria and algae?

A
  • ChR2
  • IC++ (designed by humans)
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21
Q

What is ChR2?

A
  • excitatory opsin
  • permeable to sodium ions
  • when activated with blue light, it depolarizes neurons, causing them to spike
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22
Q

What is IC++?

A
  • inhibitory light-gated ion channels
  • pass chloride and hyperpolarize neurons when activated by blue light
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23
Q

What do excitatory opsins do?

A
  • pulse light or leave it on to generate action potentials
  • depolarize neuron, causing them to spike
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24
Q

What do inhibitory opsins do?

A
  • continuous light delivery can prevent action potentials
  • like halorhodopsin
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25
Q

What is viral-mediated gene delivery?

A
  • remove DNA from virus
  • renders the virus “replication-deficient”
  • add foreign DNA to a virus
  • DNA that encodes proteins we want a cell to express
  • modified virus is injected, infect all cells in area
  • Once a virus gets its DNA into the nucleus of a cell, that cell will start to transcribe the viral DNA and make the associated proteins
  • section of DNA that encodes a fluorescent protein
  • Fluorescent proteins are used to identify which cells got infected
  • study function of neurons, use viruses to deliver DNA to them
  • make neurons express proteins that will change their resting membrane potential or generate action potentials
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26
Q

How does viral-mediated gene delivery work?

A
  • light sensitive protein (light gated ion channel)
  • take gene for protein
  • insert DNA into hollowed out virus
  • insert virus into brain
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27
Q

What is a virus?

A
  • type of DNA delivery system
  • replicate by injecting viral DNA into a host organism
  • contains instructions on how to make more virus
  • small infectious agent that replicates by injecting its DNA into normal cells
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28
Q

What is virus DNA?

A
  • instructions for how to make more virus
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29
Q

What is GCaMP?

A
  • protein
  • modified the fluorescent protein GFP
  • causing it to bind calcium and fluoresce much brighter when it does
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30
Q

What does measuring GCaMP do?

A
  • little calcium influx always occurs during action potentials
  • monitoring GCaMP fluorescence is good way to measure neural activity (in cells made to express GCaMP protein)
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31
Q

What are efferents?

A
  • outputs
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32
Q

What are afferents?

A
  • inputs
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33
Q

How do we trace neural connections?

A
  • retrograde labeling
  • anterograde labeling
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34
Q

What is retrograde labeling?

A
  • tracing afferent axons
  • What brain areas send their axons here?
  • label the cells that innervate (project to) a given region
  • retrograde tracers (chemicals like fluorogold)
  • fluorogold taken up by axon terminals and transported back to the cell body
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35
Q

What is anterograde labeling?

A
  • tracing efferent axons
  • Where do the axons from these cells go?
  • label where axons from a particular location go to
  • anterograde tracers (chemicals like PHA-L)
  • PHA-L is taken up by cell bodies and transported down to axon terminals
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36
Q

What is stereotaxic surgery?

A
  • surgical intervention that uses a stereotaxic apparatus
  • put something into a very specific part of the brain
  • inject drugs, viruses, or tracers (dyes) in brain
  • permanently implant things, like cannula, electrodes, or fiber optic cable
37
Q

What is a bregma?

A
  • junction where pieces of skull fuse together
  • used as a reference point for stereotaxic brain surgery
38
Q

Why is stereotaxic surgery commonly used for one-time injections of drug or virus?

A
  • Lesion a brain area
  • Lesion a specific type of cell in a particular brain area
  • To change gene expression
39
Q

Why else is stereotaxic surgery used?

A
  • Implant a guide cannula (allow for later infusions of drugs)
  • Implant microelectrodes
  • Implant fiber optic cables
40
Q

How do we measure fluctuations in neurotransmitter levels?

A
  • microdialysis
  • man-made fluorescent reporter proteins
41
Q

What is microdialysis?

A
  • old-fashioned approach
  • measuring changes in neurotransmitter levels in a brain region
42
Q

What is dialysis?

A
  • use of a semipermeable membrane to either deliver molecules to or measure the amount of molecules in some solution (or brain area)
  • takes time for the concentration of molecules to equilibrate across dialysis membrane
  • fastest sampling rate possible is once per minute
  • typically, once every 10 minutes
43
Q

What is a microdialysis probe?

A
  • small metal tube that holds dialysis tubing, which can be placed in an animal’s head
44
Q

What are man-made fluorescent reporter proteins?

A
  • design proteins, like receptors that become fluorescent when bound to neurotransmitter
  • use viral-mediated gene delivery to get neurons to express man-made fluorescent sensors
  • visualize neurotransmitter release in a living brain
45
Q

How would a man made fluorescent glutamate receptor work?

A
  • receptors fluorescence whenever glutamate binds to them
  • can see all places there’s glutamate release in brain in moment in time
46
Q

What is the short description of neuronal tracing?

A
  • Fluorescent molecule is injected to visualize afferents/efferents – identify inputs and outputs to a cell population
47
Q

What is the purpose of neuronal tracing?

A
  • Structure
48
Q

What are the pros of neuronal tracing?

A
  • Can create a circuit diagram of brain
49
Q

What are the cons of neuronal tracing?

A
  • Molecules don’t fill whole neuron
50
Q

What is the short description of microelectrode EEG?

A
  • Electrode in brain records electrical changes – observe activity changes with behaviour
51
Q

What is the purpose of microelectrode EEG?

A
  • Correlation
52
Q

What are the pros of microelectrode EEG?

A
  • High temporal resolution
53
Q

What are the cons of microelectrode EEG?

A
  • Can’t tell what you’re recording from
54
Q

What is the short description of electrical stimulation?

A
  • Electrode in brain delivers electrical current, stimulates nearby neurons – observe behavioural changes
55
Q

What is the purpose of electrical stimulation?

A
  • Causation
56
Q

What are the pros of electrical stimulation?

A
  • High temporal precision
57
Q

What are the cons of electrical stimulation?

A
  • Can’t tell what you’re stimulating
58
Q

What is the short description of pharmacological manipulation?

A
  • Drug delivered into brain region through guide cannula – alter cell activity in region
59
Q

What is the purpose of pharmacological manipulation?

A
  • Causation
60
Q

What are the pros of pharmacological manipulation?

A
  • Animals can be their own controls
61
Q

What are the cons of pharmacological manipulation?

A
  • Drug diffusion makes it not very spatially specific
62
Q

What is the short description of optogenetics?

A
  • Light sensitive ion channel expressed in cell population; shining light changes cell activity
63
Q

What is the purpose of optogenetics?

A
  • Causation
64
Q

What are the pros of optogenetics?

A
  • Temporal specificity; Can target specific cell populations
65
Q

What are the cons of optogenetics?

A
  • Nonphysiological
66
Q

What is the short description of calcium imaging?

A
  • Fluorescent molecule binds to calcium; can measure fluorescence as proxy of neural activity
67
Q

What is the purpose of calcium imaging?

A
  • Correlation
68
Q

What are the pros of calcium imaging?

A
  • Temporal specificity; Can target specific cell populations
69
Q

What are the cons of calcium imaging?

A
  • Can be difficult to handle so much data
70
Q

What is the short description of excitotoxic lesions?

A
  • Ionotropic glutamate receptor agonist causes excitotoxicity, killing cells in region
71
Q

What is the purpose of excitotoxic lesions?

A
  • Causation
72
Q

What are the pros of excitotoxic lesions?

A
  • Only affects cell bodies in region
73
Q

What are the cons of excitotoxic lesions?

A
  • Need an additional sham lesion group for control
74
Q

What is the short description of radio lesions?

A
  • Radio wave delivered through metal wire burns targeted brain area
75
Q

What is the purpose of radio lesions?

A
  • Causation
76
Q

What are the pros of radio lesions?

A
  • The neurons are dead for SURE
77
Q

What are the cons of radio lesions?

A
  • Destroys passing axons
78
Q

What is the short description of microdialysis?

A
  • Sample extracellular fluid through a dialysis membrane – quantify molecule concentration during behaviour
79
Q

What is the purpose of microdialysis?

A
  • Correlation
80
Q

What are the pros of microdialysis?

A
  • ID molecules present during behaviour
81
Q

What are the cons of microdialysis?

A
  • Poor temporal resolution (10+ mins)
82
Q

What is the short description of immunohistochemistry?

A
  • Localize proteins in brain tissue – fluorescent antibodies washed onto brain slice bind to protein of interest
83
Q

What is the purpose of immunohistochemistry?

A
  • Structure
84
Q

What are the pros of immunohistochemistry?

A
  • Can localize any protein you have the antibody for
85
Q

What are the cons of immunohistochemistry?

A
  • Cannot localize small molecules (neurotransmitters)
86
Q

What is the short description of cFos immunohistochemistry?

A
  • Recently active cells make cFos
  • Localize cFos in brain tissue as a correlate of recent activity
87
Q

What is the purpose of cFos immunohistochemistry?

A
  • Correlation
88
Q

What are the pros of cFos immunohistochemistry?

A
  • ID regions active during behaviour
89
Q

What are the cons of cFos immunohistochemistry?

A
  • Poor temporal resolution (post mortem)

Psych 211 (neuro) (22 decks)

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