PSB 3340 Research Methods in Neuroscience

reductionist approach

reductionist approach

scientific strategy to break a system down into increasingly smaller parts to understand it

observe & manipulate

What are the 2 categories of experiment?

methods of observational studies

humans: questionnaires, life history, postmortem tissue, brain scans, EEG

animals: neurotransmitter levels, physiological properties of neurons

methods of manipulative experiments

humans: FDA-approved drugs, treatments

animals: somatic manipulations, behavioral interventions, test non-FDA approved drugs

operant learning

used to study food consumption/obesity, liquid consumption, drug addiction, learning

Rats show consistent self-administration of cocaine. When cocaine became unavailable, they stopped pressing the button as often, but it never reached 0. After cocaine was available again, they pressed the button even more times.

Describe the rat operant learning experiment.

Neurocircuitry

aims to find what circuits are controlling behavior and how they are connected

TTX & Lidocaine

both of these chemicals block voltage-gated Na+ channels

stereotaxic atlas

map of the brain; relies on two landmarks: Lambda & bregma; brain areas are located by their coordinates

anterograde labeling

uses molecules taken up by the cell and then transported to the axon tips; uses anterogradely transported AAVs (viruses)

cell body —> axon (forward)

retrograde labeling

uses molecules taken up in the axon terminals and transported to the cell bodies; Cholera Toxin b (Cub), retrogradely transported AAVs

axon —> cell body (backward)

microdialysis

1. Uses stereotaxic surgery to implant cannulae (hollow metal tube) in a brain region of interest

2. Descend probe and perfuse artificial CSF across a membrane that allows diffusion of neurotransmitter

used in experimental animals and humans w/ seizure disorders

immunohistochemistry

1. A brain section is covered w/ antibody-containing solution

2. Antibodies attach to target cell

3. Unattached bodies are washed off

4. Labeled cells are visible

western blotting

1. Gel electrophoresis

2. Transfer proteins in samples to a “membrane”

3. Detect only protein of interest using antibodies against it

4. Visualize w/ detection kit

knock-out animals

eliminate one gene from the genotype and look for phenotypic differences; typically mice

Will knocking out X protein change Y behavior?

Viral over-expression

insert a target gene into viral DNA so that the target gene is overexpressed instead of a pathogenic gene

gene silencing techniques

uses RNAi via siRNA to interfere w/ transcription of genes w/ complementary sequence; can design and synthesize siRNA for target proteins

optogenetics

uses viral vectors to overexpress ion channels that can then influence membrane potential

ChR2

artificial light gated cation (+ ion) channel; blue light —> depolarization

HR

artificial light-gated Cl- channel; yellow light —> hyperpolarization

electrophysiology (“ephys”)

detect electrical activity which is amplified and interpreted by computer

Live animal: microelectrode placed out neuron to detect activity OR multi-electrode recording

Brian slice: patch-clamp OR intacellular; can manipulate receptors

macrolectrode recordings

detect PSPs from a population of neurons immediately below the skull; EEG

seizure

instead of activity being desynchronized across regions, there is too many action potentials, so there is a synchronization of electrical activity; “spike-and-wave” pattern

event-related potentials (ERP)

large potential shifts caused by discrete stimuli; sensory-evoked and auditory-evoked

CAT or CT scan

uses X-rays to map tissue density to show brain structure

MRI

uses magnets to produce images of brain structure

fMRI

Blood Oxygen Level-Dependent (BOLD) signal indicates metabolic activity; active brain regions require more oxygen

DTI

movement of water molecules in bundles of axons will be parallel to the the axon that make up the bundles; rainbow image

MEG

measures tiny magnetic fields given off by active neurons

high spatial resolution

provide a lot of structural detail

low temporal resolution

inability to track dynamic changes in brain activity well

transcranial magnetic stimulation

briefly stimulates discrete cortical regions w/ magnets

possible treatment for substance abuse; current treatment for anxiety/depression

behavioral pharmacology

involves the administration of drugs and observing behavior

animals: systemic and intracranial

humans: objective behavioral changes, self-reported systems, changes in brain activity

in situ hybridization

requires the use of a probe with a fluorescent label that will ONLY GLOW if the probe has binded to the target DNA (hybridization); requires the use of a flurorescent microscope

qPCR

done in tissue from a brain region, in a tube; similar to in situ hybridization