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PSYCH 478 > Topic 3: Research Methods in Pharmacology > Flashcards

Topic 3: Research Methods in Pharmacology Flashcards

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1
Q

What are neurobiological techniques?

A

research in neuropsychopharmacology can span the entire spectrum of neuroscience research

human behavior, animal behavior, brain function, circuit function, neuron/glia function, synaptic function, biochemical function

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2
Q

What does in vivo mean?

A

analyses performed in a living organism

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3
Q

What does in vitro mean?

A

in glass, analyses performed in a test tube

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4
Q

What does ex vivo mean?

A

analyses performed in live tissue removed from an organism

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5
Q

What does in situ mean?

A

analyses performed in context to living tissue but typically after dissection

tissue is no longer living

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6
Q

What is stereotaxic surgery?

A

apparatus for specific positioning of instruments/syringe into the brain

stereotaxic atlas gives coordinates to target specific regions

can be used to give very precise injections, microsurgeries, lesions

place head in fixed position

can target small parts of the brain without seeing them

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7
Q

What is microdialysis?

A

technique for collecting fluid from a live and awake animal

analysis of neurotransmitter levels, signaling molecules, drugs, etc. in vivo

collect fluid from awake animals

osmosis causes liquid in brain to enter tube and leave so we can measure

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8
Q

What is chromoatography?

A

chromatographic techniques seperate molecules based on their size, charge, relative polarity, or specific interactions

compounds can then be detected, quantified, or further analysed

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9
Q

What are the different analytes of chromatography?

A

proteins: column chromatography (size exclusion, cation/anion exchange, affinity)

proteins/peptides: high performance liquid chromatography (HPLC) often coupled to mass spectroscopy (HPLC-MS)

neurotransmitters: HPLC, gas chromatography (GC)

drugs: HPLC, GC, thin layer chromatography (TLC)

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10
Q

What is electrophysiological recording?

A

a variety of techniques for measuring electrical activity in organisms, tissues, or cells

useful in vivo, ex vivo, or in vitro

classic studies by Hodgkin and Huxley on squid giant axon

can put electrode in specific areas

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11
Q

What are in vivo electrical recordings?

A

more specific than an EEG

implantable electrodes connected to an external recorder

can measure activity in specific regions during normal behavior

typically measure field activity (relatively large number of neurons)

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12
Q

What is intracellular recording?

A

measures activity across the membrane of a single cell

whole-cell (sharp electrode)
patch-clamp (smaller areas)

can be used to isolate single ion channels for measurement

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13
Q

What are molecular methods of research?

A

a variety of techniques have been used over the years to locate neurotransmitters and receptors in the brain

early techniques (sometimes used today) used in radioactive isotope labelled drugs or antibodies to visualize drug binding: autoradiography, radioimmunoassay

these have been largely supplanted by immunofluorescent and transgenic techniques

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14
Q

What are immunological techniques?

A

antibodies are proteins produced by the adaptive immune system

antibodies have specific binding sites for recognition of foreign antigens

antibody binding is high affinity and is responsible for targeting foreign bodies for destruction

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15
Q

What is innate immunity?

A

immunity can be exploited experimentally to produce antibodies against biologically relevant proteins

typically an isolated protein is injected into a rabbit (mouse, goat, donkey, chicken), an immune response generated, and antibodies are isolated from the rabbit serum

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16
Q

What is immunofluorescence?

A

antibodies can be conjugated to fluorescent molecules and used to identify specific proteins in cells of the CNS

tells us where in the brain that protein is

usually attach fluorescent tag to antibodies

immunohistochemistry is the detection of proteins in tissue

immunocytochemistry is the detection of proteins in cells (typically cells cultured in vitro)

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17
Q

What is confocal microscopy?

A

specialized technique used in fluorescence microscopy to filter out-a-plane light

allows imaging of an optical “slice” of tissue

can be used to generate 3D images of biological structures

3D image and multiple layers

fancy, high-powered microscope

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18
Q

What is 2-photon microscopy?

A

uses infrared laser (700-900 nm) to image living tissue

IR light can penetrate ~10 mm through brain tissue

doesn’t have 3D or multiple levels

most simple fluorescent microscope

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19
Q

What is enzyme-linked immunosorbent assay (ELISA)?

A

detects an immobilized antigen using an enzyme-linked antibody

higher sensitivity

lets us know how much antigen there is based on antibodies

20
Q

What is western blotting?

A

detects an immobilized antigen using an enzyme or fluorophore linked antibody

higher throughput

21
Q

How is gene expression evaluated?

A

RNA expression (transcripts) can be measured as a proxy for protein expression

RNA is translated into proteins but protein is the functional unit of biological systems

numerous processes can modify or alter translation so RNA =/= protein

22
Q

What is fluorescence in situ hybridization (FISH)?

A

binds a fluorescent probe in tissue sections to show which cells may be expressing a gene of interest

same as protein but just targeting RNA

have animal do activity, see what RNA were active

23
Q

What is quantitative polymerase chain reaction (qPCR)?

A

uses the PCR process to identify specific transcripts from tissue homogenates

24
Q

What are cDNA microarrays?

A

can identify thousands to millions of transcripts in a single sample

usually comparative between two samples (e.g. control vs drug treated)

25
Q

What are structural techniques of brain imaging?

A

can reveal differences in gross anatomy in various pathological conditions

26
Q

What are functional techniques of brain imaging?

A

can reveal changes in brain function with between pathological conditions or during specific tasks

27
Q

What is magnetic resonance imaging (MRI)?

A

uses the phenomenon of nuclear magnetic resonance to generate structural images of living tissue

ideal imaging technique

cannot say anything about function

imaging modes can be varied for normal anatomy, pathology, or grey/white matter differences

28
Q

What is positron emission tomography?

A

PET uses positron-emitting radiotracers to analyze brain receptors or metabolism

radioligands for specific receptors can be used to reveal binding in different brain regions

tissue uptake of 2-deoxy-2-[18F]fluoro-D-glucose (FDG) is proportional to glucose uptake and hence activity

29
Q

What is single photon emission computed tomography?

A

uses gamma-emitting radiotracers to analyze brain blood flow

tissue uptake of “fake oxygen” is proportional to blood flow and hence activity

mostly used for experimental not diagnosis

30
Q

What are connectivity measures?

A

fMRI is used to measure functional connectivity by correlating changes in activity during tasks between regions (fcMRI)

effective connectivity is established by information flow between regions

structural connectivity is established using DTI

31
Q

What is genetic engineering?

A

transgenic mic are a commonly used technique for assessing genetic contributors to disease or for determining functions of specific genes

common models use genetic knockouts (KO mice), knock-ins, or selective or conditional knockouts

knock out a gene and see what happens

32
Q

What is optogenetics?

A

specific bacterial proteins function as light-gated ion channels

expression of opsins can be used to non-invasively stimulate or inhibit neuronal function in vivo

van be combined with specific genetic techniques to target different neuronal populations

can be used with fiber optics to alter activity involved in behavior in live, free-moving animals

offers several improvements over conventional electrical stimulation including a less invasive procedure, and cell- or region-specific modulation of activity

33
Q

How are animal models used in behavioral pharmacology?

A

animal models are used to mimic human behaviors and pathologies

use of animals presents challenges for psychopharmacology as there is no such thing as a depressed or schizophrenic mouse

behavioral tests are designed to provide measurable proxies for human behaviors

34
Q

What is construct validity?

A

similarity between the methods by which the model is induced and the etiology of disease

genetic and environmental factors

replicate the cause of the disease

35
Q

What is predictive validity?

A

utility of the model to predict drug effects

assumes similarities in effect are based on shared mechanism

how well drug works

does the drug work the same in animals and humans

36
Q

What is face validity?

A

ability of the model to recreate key features of a disease

anatomical, behavioral, and neurochemical features of disease

how similar are symptoms

37
Q

What is the Morris water maze?

A

Morris water maze tests spatial memory

animals are tested for their ability to recall the location of a hidden submerged platform in a pool

decreased time to find the target in repeated trials indicates spatial memory

38
Q

How are mazes used to test learning and memory in animal models?

A

mazes are useful for measuring memory in animals

many possible configurations

the radial 8-arm maze tests spatial memory

39
Q

How does novel object recognition test learning and memory in animal models?

A

novel object recognition tests can evaluate episodic memory

animal is habituated with two objects

in a repeat trial one object is replaced

time spent investigating the novel object is indicating of episodic memory

if they have episodic memory they spend more time on new thing

no episodic memory mice will spend equal time on both

40
Q

How is anxiety tests in animal models?

A

anxiety tests are common measures for depression

open field test places animal in a novel open box environment

mice have both strong exploratory drive and fear of open spaces

movement tracked over a defined time period

decreased time spent in open vs peripheral areas is indicative of anxiety

amenable to automated analysis

if anxious, decreased time in the middle and increased time in the sides

41
Q

What is the elevated plus maze test for animal models?

A

plus maze consists of two open arms and two closed arms

entry into open arms is indicative of novelty-seeking behavior

depressed/anxious animals spend more time in closed arms

42
Q

What is fear-conditioning in animal models?

A

animals are presented with a signal (light or tone) followed by an unescapable foot shock

later trials measure the fear response (freezing) to the conditioned stimulus

fear-conditioned startle is an alternative in which the conditioned stimulus if followed by a startling noise

even when given opening they will stay, learned helplessness model of depression

43
Q

What is reward-operant conditioning in animal models?

A

animals are readily trained to obtain rewards (food or drugs) in response to operant behaviors (lever press)

useful for developing models of substance abuse (operant self-administration)

44
Q

What is conditioned place preference?

A

Pavlovian conditioning paradigm

drug injection (unconditioned stimulus) paired with a neutral environmental stimulus to create a conditioned stimulus

2-3 chamber conditioning box

45
Q

What is the three phase conditioned place preference protocol?

A

pre-conditioning (habituation): open exploration, 15 minutes each for 3 days

conditioning: alternate injections of drug and vehicle in alternate chambers

post-conditioning (test): open exploration, time spent in each chamber measured

PSYCH 478 (9 decks)

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