NSC Research Methods

Neuroscience

The study of the nervous system

Basic Neurosciences

Molecular Neuroscience, Cellular Neuroscience, Systems Neuroscience

Applied Neurosciences

Behavioral, Cognitive, Developmental, Clinical

Molecular Neuroscience

Study of specific molecules and their effects/significance within regulating functional circuitry in the PNS or CNS (typically interested in specific genes/proteins)

Cellular Neuroscience

Study of neurons on a cellular level (morphology & physiological properties such as membrance trafficking, synthesis, transport of proteins, and synaptic plasticity)

Systems Neuroscience

Identifying how neurons form networks, work together in complex networks

Behavioral Neuroscience

Biology of nervous system pertaining to some observable ability

Cognitive Neuroscience

Biology of nervous system pertaining to some unobservable ability

Developmental Neuroscience

Changes over time in the biology of nervous system pertaining to any ability or behavior

Clinical

Atypical functioning in the biology of nervous system pertaining to any ability or behavior

What regions of the brain are involved in playing ping pong? (Pick an applied & basic)

Behavioral & Systems

What type of neuron carries signals leaving the brain to the muscles? (Pick applied & basic)

Behavioral Cellular

How does Acetylcholine from a nerve trigger a muscle fiber to contract? (Pick applied & basic)

Behavioral Molecular

Three Features of Science

Follow Systematic Empiricism, Address an Empirical Question/Hypothesis, Be Public Knowledge

Psudeoscience

Violate a feature of science AND be claimed to be scientific

Theory

a coherent explanation or interpretation of one or more phenomena

Hypothesis

a specific prediction about a new phenomenon that should be observed if a particular theory is accurate

Operationalization

the meaning of a scientific concept depends on the procedures used to establish it, so that each concept can be defined by a single observable and measurable operation

Strong Inference

the process of organizing questions and hypotheses to build structured, logical sequences of experiments to work through understanding a phenomenon

Structural Neuroimaging

Techniques that show the brain’s static physical structure at one point in time (essentially a picture)

Functional Neuroimaging

Techniques that show the brain’s activity over time

Spatial Resolution

how accurately the measured activity is localized within the brain (how small can you see)

Temporal Resolution

how closely the measured activity corresponds to the timing of the actual neuronal activity (shows different rates)

Structural Imaging Techniques

sMRI, DTI, CAT/CT

Functional Imaging Techniques

fMRI, fNIRS, PET, EEG

sMRI > What is the data, what does it look like, how do we use the data?

Radio frequency pulses of atoms, a structural image of the brain is produced, used to view the locations and measure metrics of the brain

What does a sMRI measure exactly?

Hydrogen Decay rates

Hydrogen Decay

Process of atoms naturally re-orienting to magnet, releasing an RF signal that we measure

Decay Rate

Speed of H decay differs across tissues (caused by diff. interactions w/ environment) which we used to differentiate tissues

Process of sMRI on H

No magnet: randomly positioned > magnet on: mostly aligned > radio pulse: move orthogonally (90 degrees) > protons relax: release radio signal

The stronger the sMRI magnet, the better the…

image clarity

What is the unit of measurement for sMRI?

T = Tesla (unit of strength of magnetic field)

Common Metrics of Interest in sMRI

Grey matter thickness, volume, or density > white matter volume

Voxels

Volumetric pixels

What does DTI stand for?

Diffusion Tensor Imaging

DTI > what is the data, what does the data look like, how do we use the data

Diffusion of water molecules throughout the brain (location and strength of axons) > a “highway map” of the brain’s white matter tracts are produced > used to assess structural connectivity of the CNS (where is the fluid and in what direction is it going?)

Mean Diffusivity (DTI)

how much fluid is there

Fractional Anisotropy (DTI)

how uniform is the flow of the fluid

What does CAT stand for?

Computerized Axial Tomography

CAT > what is the data, what does the data look like, how is the data used

X-rays > creates stacked-up 2D X-ray scans of the brain > used for basic visual assessment of the brain’s structures

sMRI: Invasion, Spatial Resolution, Expense

Noninvasive, high spatial resolution, extremely expensive

DTI: Invasion, Spatial Resolution, Expense

Noninvasive, high spatial resolution, extremely expensive

CAT: Invasion, Spatial Resolution, Expense

Noninvasive but higher risk (radioactive), high spatial resolution, typically cheaper > doesn’t give a lot of data and not as great for research

What does fMRI stand for?

Functional Magnetic Resonance Imaging

fMRI > what is the data, what does the data look like, how is it useda

BOLD-influenced radio frequency pulses of atoms (focus on H atoms in the blood stream) > produces a map of blood flow changes over time > assesses the roles and relationships of regions of the brain

Process of fMRI

Stimulus > Neuronal activity (when they increase their energy, they need more blood flow) > neurovascular coupling > haemodynamic response > detection by MRI scanner > fMRI BOLD response (increased blood flow which we assume is related to neural activity)

What is a potential complication with fMRI?

The haemodynamic response takes about 5-7s which could mean there is another potential stimulus directly after the intended stimulus

How does the fMRI process changes in blood flow?

Sees the changes in oxygenated blood

What changes in the oxygenated blood are observed?

Deoxyhemoglobin is paramagnetic > alters the effect of MRI RF/magnets on H > H in deoxygenated blood doesn’t have uniform decay rates > RF signal from a voxel with more deoxygenated blood will be noisier and weaker (signal gets better means more oxygenated blood)

BOLD Imaging

Blood-Oxygen Level Dependent Imaging > a voxel’s signal intensity (consistent intense signal in voxel = oxygenated blood)

fMRI Spatial and Temporal Resolution

Spatial = mm, Temporal = sec

What does fNIRS stand for?

Functional Near-Infrared Spectroscopy

fNIRS > what is the data, what does it look like, what does it measure

NIR light intensity (measures infrared light intensity) > generates map of blood flow changes, assesses roles and relationships of regions

Process of fNIRS

Stimulus > Neuronal Activity > Neurovascular coupling > Haemodynamic response > Detection by fNIRS device > Output

What does fNIRS use to measure activity?

Optodes (sources and detectors), on head > measures how much NIR light bounces from cortex

Why is NIR light used in fNIR?

NIR has predictable absorption levels > can pass through all tissues but blood which it bounces off of > the amount of light depends on how oxygenated the blood is

What does PET stand for?

Positron Emission Tomography

PET > what is the data, what does the data look like, how is it used

Gamma rays > creates a heat map of a blood-borne radioactive tracer > assesses the roles of regions in the brain

Process of PET

Stimulus > Neuronal activity > neurovascular coupling > haemodynamic response > detection by PET device > output

PET Spatial and Temporal Resolution

Spatial: mm, Temporal: min

What does EEG stand for?

Electroencephalography

EEG > what is the data, what does it look like, how to we use it

Electrical activity > creates a representation of the brain’s overall real-time response to a stimulus > investigates temporal order of events in brain

Naming Structure of ERPs

Event Related Potentials > P = positive, N = Negative, # = ms after stimulus (ex: N400 = Negative 400 ms after stimulus)

EEG Spatial and Temporal Resolution

Spatial: Lobes > Temporal: ms (real time essentially)

fMRI: Invasion, S/T Res., Exp. Mobility

Noninvasive, high s/t res (mm & sec), extremely expensive, immobile

fNIRS: Invasion, S/T Res., Exp., Mobility

Noninvasive, moderate s (cm), very high t (ms), extremely cheap, mobile

PET: Invasion, S/T Res., Exp., Mobility

Highly invasive (injection of radioactive material), high s (mm), low t (min), expensive, immobile

EEG: Invasion, S/T Res., Exp., Mobility

Noninvasive, terrible s (lobes), real-time t, extremely cheap, mobile (actually measuring activity of neurons)

Cellular Imaging Techniques

MRI (Single-Cell), PET (Single-Cell), Histology, Multiphoton Microscopy

MRI (Single-Cell) > what is the data, what does it look like, what is it used for

Radio frequency pulses of atoms > produces structural image of brain w/ certain cells highlighted > determines the locations of and migration of individual cells in vivo

What does SPION stand for?

Superparamagnetic Iron-Oxide Nanoparticles

What can you use SPION for?

SPION has a coating and is guided by an external magnet to their target cell > allows these cells to show up on an MRI as they have a high magnetic properties and the H have different decay rates

PET (Single-Cell): Data, Looks like, Uses

Gamma rays > heat map of certain cells, determines locations of and migration of individual cells in vivo (tag cells w/ something radioactive)

What is FDG and what is it used for?

Fluorodeoxyglucose is a radioactive sugar-like compound > radiolabeling through bathing cells, single cell dispensing, IV injection (sees where cells went), PET/CT imaging & intracardiac injection (where the cells ended up)

Mesoporous Silica Nanoparticles

Infuse Ga-68 into MSN’s > infuse MSN’s into cells of interest > separate and inject single cells into subject > record cell locations/migrations

Histology: Data, Looks like, Uses

Chemical staining of cells/components of cells, depends on technique, to determine locations of individual cells in ex vivo brain tissue

Histology Stages

Fixing > Drying & Clearing > Inclusion & Cutting > Staining > Mounting > Microscopy

Histology Techniques

Nissl, Luxol Fast Blue, Golgi, Immunhistochemistry

Nissl Stain & Target

Fluorescent compunds (Cresyl Violet) > “nissl bodies” (rough endoplasmic reticulum) in soma of neurons/glia (looking at cell bodies)

Luxol Fast Blue Stain & Target

the fluorescent compund “luxol fast blue” & myelin

Golgi Stain & Target

Potassium chromate & random cells in a block of tissue (about 5%) (way to look at whole cell) > can stain to see type of neuron & density of neurons in an area

Immunohistochemistry Stain & Target

Antibodies (bind), Substrates (color) & proteins (frequency in area) > very effective at identifying locature of proteins and neurons

Microscopy > Data, Looks like, Uses

Depends on technique, depends on technique, assesses cellular-level questions on structure/location/function in vivo/ ex vivo

Microscopy Techniques

Calcium Imaging, Multiphoton Microscopy

What does GECI stand for?

Genetically Encoded Calcium Indicators

What do GECI’s do?

observe influx of calcium to understand firing (measures the activity of neurons in vivo)

How do GECI’s work?

fluorescence of GECI’s increases when more Ca++ flows through them > causes the neurons to flash that can be seen through microscopy > microscope is attached to subject

Viral vs Transgenic Expression (GECI’s)

Viral: targeted area, designed viruses can carry GECI’s & target a certain area & leave GECI’s behind | Transgenic: all over, can genetically encode GECI’s in organism

What does GECI’s measure?

They measure light intensity > see when neurons activate (not their location though) (not taking images)

What is Multiphoton Microscopy?

Imaging structure/function of fluorescent cells at femtosecond speed using NIR light in vivo (actual images of neurons)

Multiphoton Micrscopy: How it works & target

shoots light and sees how much light bounces back > targets endogenous fluorescent proteins/substances & injected fluorescent compounds > used to look at layers in the voxel (structural data)

What can Multiphoton Calcium Imaging?

Can measure where the activity is & how active the neurons are (when they activate and where & what they look like)

MRI (Single-Cell): Vivo, Inv., Exp., S Res.

In Vivo, noninvasive, extremely expensive, high s (mm) (no t res.)

PET (Single-Cell): Vivo, Inv., Exp., S Res

In Vivo, invasive, expensive, high s res (mm) (no t res)

Histology: Vivo, Inv., Exp., S Res

Ex Vivo, Sacrifice, Scopes are main cost, depends on microscope (no t res)

Microscopy: Vivo, Inv., Exp., S Res

In/Ex Vivo, Invasive/Sacrifice, Scopes are main cost, Multi. have extremely high s res (micrometers) & calcium imaging = real-time

Electrical Data Techniques

Extracellular, Intracellular (Patch Clamp)

Chemical Data Techniques

PET (Molecular), MRS

Electrical Data: Data, Looks like, Uses

Electrical activity, real-time changes in electrical charge/space between neurons, assesses temporal aspects of neurons’ activity

Extracellular vs Intracellular Electrical Data

E: in between neurons, I: attached to neuron (both measure electrical charge of a small area in the brain)