W3 L5 211 Flashcards
For humans, brain interaction needs to be non invasive for most part, but for structural organization need invasion and rely on animals and experimental ablation. “Lesion method”
Experimental Ablation (Lesion Methods)
▪ Experimental ablation
A method used to investigate brain function by destroying part of the brain and evaluating the animals subsequent behaviour.
▪ The area of the brain that is damaged is called the brain lesion.
▪ The goal of lesion methods is to discover what functions are performed by different regions of the brain and how these functions are organised to produce complex behaviour (i.e functional organisation of the brain).blocalization of function.
▪ Neural circuits in the brain perform functions (or a set of functions).
▪ Functions contribute to the performance of a behaviour.
Experimental approaches to understanding behaviour
▪ Invasive physiological research methods
(lesion, electrical stimulation, neurophysiological recording)
▪ Behavioral paradigms that assess constituent cognitive processes (visuospatial attention, working memory, reversal learning)
▪ Pharmacological research methods
(drug adminstration, chemical lesions, chemical activity of the brain)
▪ Genetic engineering
(gene knockout, genetic replacement)
▪ Visualising the living human brain (PET, FMRI)
▪ Neuropsychological testing (WCST, delayed response)
Due to vast array, different approaches may procudece different results or complementary findings, need to choose wisely. Answer right question.
Wa to create brain lesions: Brain Stereotaxy
No two brains of any given species are identical.
The skull is made up of several bones that join together and form sutures. The junction at which the sutures meet at the front of the head is called bregma. The junction at the at which the sutures meet at the back of the head is called lambda.
Stereotaxic surgery is a form of surgical intervention. It uses a three dimensional coordinate system to locate a target brain region and perform the lesion.
A stereotaxic atlas contains drawings that correspond to brain sections taken at varying distances from bregma. 3d system.
Stereotaxic apparatus for performing brain surgery in rats
Adjusting knobs
▪ Inject excitotoxins to create selective lesions to specific brain areas
▪ Create neurochemical lesions (e.g. deplete acetylcholine in the frontal cortex)
▪ We can use it to place wires to stimulate neurons
Different lesion méthods
Excitotoxic lesions: Like using nmda
▪ An excitatory amino acid (e.g. ibotenic acid, kainic acid, N-methyl-d-
aspartic acid) is injected into the target region of the brain.
▪ The chemical destroys cell bodies (the somas) by stimulating them to
death.
▪ This method is highly selective; it destroys cell bodies only, and spares
axons of other neurons that pass nearby (fibres of passage).
▪ This method can create selective neurochemical lesions thereby depleting dopamine, serotonin, norepinephrine or acetylcholine in specific brain areas. Damaging terminals.
Aspiration lesions
▪ The brain area of interest is removed by suction (or aspiration) using a fine-
tipped hand-held glass pipette.
▪ Can be performed under visual guidance.
▪ This method can sometimes damage underlying white matter and major
blood vessels.
▪ This is a nonselective method because it removes cell bodies and fibres of
passage.
Radiofrequency lesions
▪ Small subcortical lesions (i.e.regions located beneath the cortex) are made by passing a radiofrequency current through a stainless steel wire that is insulated except the tip.
▪ The wire is guided stereotaxically.
▪ The electric current produces heat that destroys the cells in the region surrounding the tip of the wire.
▪ The size and shape of the lesion is determined by the duration and intensity of the current.
Sham lesions (operated controls)
▪ When we create anatomical or neurochemical lesions, we can potentially cause additional damage simply by inserting the electrode or injector into the brain.
▪ For experimental comparisons, a group of animals will undergo the same experimental procedure WITHOUT the excitotoxin, frequency or aspiration.
▪ Animals with sham lesion serve as a control group.
NOTE: Most lesions are permanent. Temporary (or reversible) lesions can be made by infusing a local anaesthetic (e.g. lidocaine) which blocks action potentials, or infusing a GABA agonist (e.g. muscimol) which is an inhibitory neurotransmitter.
Optogenetics (photostimulation)
“New sexy method”
▪ Photostimulation uses light to artificially activate cells
▪ The light activates a light sensitive protein which excites the cell that expresses that
protein.
▪ E.g. When blue light hits channelrhodopsin-2 (ChR2), a protein that controls a cation channel, the channel opens enabling the influx of cations thereby depolarizing the membrane.
▪ When yellow light hits natronomonas pharaonis halorhodopsin (NpHR), a protein that controls an anion transporter, the transporter enables the influx of anions thereby hyperpolarizing the membrane.
Histological Methods (1): Nissl Staining? Golgi Staining?
Histological methods allow:
1. Location of structures
2. How structures are connected
3. Identify areas of the brain that are damaged (verify the lesion)
▪ Histological stains allows us to visually identify specific substances within and outside a cell. Key to neurobiology, so we can actually see something meaningful.
▪ The most commonly used stain is cresyl violet which stains cell bodies.
▪ The dark regions represent cell bodies. The light regions represent axons or fibre bundles.
Histological Methods (2):Golgi Staining
The golgi stain allows us to visually identify the external structure of neurons.
The neuron silhouettes are revealed in great detail but the internal structure of the cells is invisible.
Electron microscopy allows us to see small anatomical structures
(e.g. synaptic vesicles and details of cell organelles) using a special
electron microscope.
An electron photomicrograph showing the cross section of a synapse
A colour enhanced electron photomicrograph showing a neuron cell body (green) studded with terminal buttons (orange).
Tracing Neural Connections
Brain regions do not work in isolation. They receive inputs from different structures and send outputs to other brain regions. The integration of these inputs and outputs leads to a network that requires activation for the appropriate behaviour.
Anterograde labeling (tracing efferent axons)
▪ Used when tracing a path of axons
projecting ‘away’ from the cell bodies.
▪ The method employs chemicals such as PHA-L (an anterograde tracer) which is injected into the brain region of interest.
▪ The PHA-L molecules are taken up by the cell bodes and dendrites and transported down the axon to the terminal button. Use immunocytochemistry to see it.
Retrograde labeling (tracing afferent axons)
▪ Used when tracing a path of axons projecting
‘toward’ a particular brain area.
▪ The method employs chemicals such as fluorogold (a retrograde tracer) which is injected into the brain region of interest.
▪ The fluorogold molecules are taken up by the terminal buttons and transported back to the cell bodies.
Reciprocal relationships or bidirectial interactions would shout both in theory, unlike unidirectional one from thalamus to frontal cortex.
Transsynaptic labelling with pseudorabies virus (PRV) allows us to trace chains of connected neurons
See textbook for a bit more ideas.
Remember it is transynaptic.
Allows temporal distribution percentage of virus spreading happening, if taken at different times.
Visualising the Living Human Brain (I) - CT
Computerised Tomograpy (CT)
▪ A computer assisted X-ray procedure used to visualise the brain.
▪ The patient lies with his or her head positioned in the centre of a large cylinder.
▪ An X-ray beam is projected through the head to an X-ray detector.
▪ The X-ray bean scans the head from all angles.
▪ A computer translates the information received from the X-ray detector into a series of pictures of the skull and brain.
MRI
Magnetic Resonance Imaging (MRI)
▪ Does NOT use X-rays. Instead, it uses strong magnetic fields.
▪ The patient lies with his or her whole body positioned in the centre of a large cylinder.
▪ When the body is in a strong magnetic field, the hydrogen atoms in the body spin with a particular orientation.
▪ A radiofrequency wave passed through the body makes the hydrogen atoms emit their own radiowaves.
▪ The computer is tuned to detect radiation from hydrogen atoms because these atoms are present in different concentrations in different tissues.
▪ The result is a high spatial resolution, three- dimensional image of the brain.
Visualising the Living Human Brain (III) - PET
Positron Emission Tomography (PET)
▪ Measures the metabolic activity of specific brain regions.
▪ The person is injected with radioactive 2-DG (2- deoxyglucose) into the carotid artery.
▪ 2-DG is similar to glucose. It is taken up by active energy consuming molecules of the brain.
▪ However, unlike glucose, 2-DG cannot be metabolised, so it is broken down.
▪ As the radioactive 2-DG molecules decay, they emit positrons which are detected by the scanner.
▪ The computer determines which regions of the brain have taken up the radioactive 2-DG and produces a picture of the brain showing different activity levels.
PET scan of human brain showing brain at rest (top row) and when clenching right fist (bottom row). 2- DG uptake is indicated by greater yellow/orange/red.
fMRI Functional Magnetic Resonance Imaging (fMRI)
▪ The most recent advance in imaging technology.
▪ It produces images of the increase in oxygen flow in the blood to active areas of the
brain.
▪ Has four major advantages over other imaging tools:
a) Nothing has to be injected;
b) Provides both structural and functional information in the same image; c) High spatial resolution;
d) Produces 3D images of the whole brain.
Imp. Area of high statistical value in red.
Recording neural activity (4)
Different perceptions and behavioural responses involve different patterns of neural activity. These patterns of activity can be recorded. Recordings that are chronic are made over an extended period of time. Recordings that are made acutely are made for a relatively short period of time.
Microelectrodes
▪ Very fine electrodes made of thin wires.
▪ Have a fine tip that can record the electrical activity of a single neuron (known as single-unit recording).
▪ Electrodes are implanted in the brains of animals using stereotaxic surgery.
▪ The wires are connected to a socket and the animals can be ‘plugged in’ to a recording system.
Macroelectrodes
▪ An electrode used to record activity of a large number of neurons in a particular region of the brain.
▪ The electrode measures post synaptic potentials of millions of cells located in the region of the electrode.
▪ Electrodes come in different forms (e.g. unsharpened wires or metal discs attached to the skull which conduct electricity).
▪ The electrodes are attached to an amplifier which records an electroencephalogram (EEG) which provides a measure of gross activity in the brain.
▪ EEG’s provide a diagnostic tool with which particular states of consciousness or types of cerebral atrophy are associated with specific patterns of EEG waveforms.
Some typical EEG’s and their psychological correlates
Stimulating Neural Activity (3)
Sometimes we need to artificially change the activity of specific regions of the brain to see the effects of these changes on behaviour.
Electrical stimulation
▪ Involves passing an electrical current through a wire inserted into the brain.
Chemical stimulation
▪ Involves injecting a small amount of excitatory amino acid into the brain (e.g. glutamic acid stimulates glutamate receptors).
Microiontopheresis
▪ Determines the effect of transmitter substances (or drugs) on the activity of individual neurons.
▪ It uses a a multibarreled micropipette which discharges small quantities of drug when an electrical current is passed through the micropipettes.
▪ The recording electrode detects the activity of the cell exposed to the drug.
Transcranial Magnetic Stimulation (TMS)
Neural activity induces magnetic fields that can be detected by a special device call magnetoencephalagram.
Similarly, magnetic fields can be used to stimulate neurons by inducing electrical currents in brain tissue.
TMS uses a coil of wires (arranged like the figure 8) to stimulate neurons on the surface of the cerebral cortex.
The cross point of the figure 8 is the place where stimulating takes place.
Pulses of electricity send magnetic fields that activate neurons.
