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[answered] Biology Experiment 5 Extracellular Recordings and Measuring


Please do the Q1 to Q5 in the attachment, thank you

Extracellular Recordings and Measuring Field potentials using MEA chips

Q1

a. Go to the following website

http://vlab.amrita.edu/index.php?sub=3&brch=43&sim=135&cnt=2 108

b. Click on the ?Animation?. Participate in a full animated walk-through to complete the feel for a real LFP recording. You will have to click/move using the computer mouse to pass through the various stages of the experiment.

c. Finish ?Self-evaluation? and get a screenshot for your answers, and provide it as solution to exercise 1.

Q2

Extracellular Recordings and Measuring Field potentials using MEA chips

Action potential propagation

  • Sketch the membrane voltage as measured in A and B.
  • What is the amplitude of a typical extracellularly measured signal?
  • Is the resultant waveform biphasic or triphasic?
Extracellular Recordings and Measuring Field potentials using MEA chips

Q 3) The amplitude of the measured voltage is different for intracellular and extracellular recordings. Explain clearly why.

Q 4) The extracellular recording changes polarity. Why does this happen? Why does this not happen with intracellular recordings?

Q 5) Why does LFP only capture slower currents, i.e., only subthreshold activity. Why does it not capture spikes or high frequency activity?



Biology Experiment 5 Extracellular Recordings and

 

Measuring Field potentials using

 

Microelectrode array (MEA) chips

 

V.Guntu, S.S.Nair, University of Missouri 1 Introduction to Extracellular Recording (adapted from CNS Tutorial 2012)

 

? Electrical signals from individual cells

 

are acquired by inserting a conductive

 

microelectrode near the cell

 

membrane, which then provides a

 

source or sink for the trans-membrane

 

currents associated with an action

 

potential.

 

? Each spike appears as a stereotyped

 

waveform on the voltage trace

 

recorded from the microelectrode.

 

? Neural tissue in the brain is dense

 

enough that a probe inserted at

 

random into the gray matter will lie

 

close to many cell membranes, and

 

will couple to currents across all of

 

them.

 

Figure 1: Extracellular Recording (from CNS 2012 Tutorial)

 

V.Guntu, S.S.Nair, University of Missouri 2 Different stages of getting the desired signal

 

? The signal from the microwire (Fig 1) is amplified and band-pass

 

filtered and the firing of the nearby neurons appears as spikes on top

 

of background activity.

 

? Spikes are detected using an amplitude threshold and then sorted

 

according to their shapes.

 

? For neurons close to the electrode tip -about 50 to 100 microns

 

the signal-to-noise ratio is good enough to distinguish the activity of

 

each single unit .

 

? For more distant neurons upto about 150 microns- spikes can be

 

detected but the difference in their shapes is masked by the noise

 

(multi-unit activity).

 

? Spikes from neurons further apart cannot be detected and they

 

contribute to the background noise activity.

 

V.Guntu, S.S.Nair, University of Missouri 3 Illustration of Extracellular Recording at single cell level Figure 2: IR Drop (From Axon Guide) ? In an extracellular recording

 

experiment: the current I that

 

flows between parts of a cell

 

through the external resistance

 

R produces a potential

 

difference ?V, which is usually

 

less than 1 mV .

 

? As the impulse propagates, I

 

changes and, therefore, ?V

 

changes as well.

 

V.Guntu, S.S.Nair, University of Missouri 4 Volume conductor theory

 

? A : when the axon is at rest, the

 

membrane potential is uniform,

 

and no current flows

 

? B: Current will flow when a

 

segment of the membrane is

 

depolarized. The flow is inward

 

at the depolarized region

 

(?sink?) and outward at adjacent

 

regions, which acts as a

 

(?source?) of the current sink.

 

Figure 3: Current flow around an axon (Wikipedia) V.Guntu, S.S.Nair, University of Missouri 5 Model of sources and sinks predicts that a triphasic wave will be recorded

 

from an isolated axon

 

? A . As the action potential approaches the

 

region underneath the electrode, that

 

membrane serves as a source, and the

 

electrode sees a positive potential relative

 

to a distant indifferent electrode.

 

? B. When the action potential reached the

 

membrane underlying the membrane, the

 

electrode records a negative potential.

 

? C. As the action potential continues down

 

the axon, the membrane under the

 

electrode once again acts as a source, and

 

as a consequence, the electrode records a

 

positive potential.

 

Figure 4: Recording triphasic waveform from an axon (Wikipedia)

 

V.Guntu, S.S.Nair, University of Missouri 6 How does the extracellular signature of action

 

potentials depend on neuronal morphology

 

? Amplitude is (i) roughly

 

proportional to sum of crosssectional areas of dendrites

 

connected to soma, (ii)

 

independent of membrane

 

resistance Rm, ? (check)

 

? Spike width increases with

 

distance from soma, i.e., highFigure 5 - Above : Different cells

 

frequency dampening also with

 

with different morphologies,

 

simple ohmic extracellular

 

Left : Characteristics of Extracellular

 

medium.

 

Signals

 

(Adapted from CNS Tutorial 2012)

 

V.Guntu, S.S.Nair, University of Missouri 7 Advantages and Limitations of Extracellular Recordings

 

? Extracellular recordings are attractive. Why?

 

? They produce a (whole) lot of data with ?moderate? tissue damage.

 

? They potentially provide sub-millisecond resolution of neuronal

 

spiking activity.

 

? They are relatively cheap to implement.

 

? But ...

 

? They require a lot of processing.

 

? They cannot provide some very important pieces of information like

 

the neuronal cell type, the sub-threshold activity, the morphology of

 

the recorded cells.

 

V.Guntu, S.S.Nair, University of Missouri 8 Local Field Potential (Adapted from Scholarpedia)

 

? The local field potential (LFP) refers to

 

the electric potential in the

 

extracellular space around neurons.

 

? A. Array of 100 silicon electrodes with

 

400?m inter-electrode distance.

 

? B.Plan of Implementation (black dots)

 

? As Distance from microwire increases

 

the recordings will pick up slower

 

activity. We can record action

 

potentials from unit recordings as

 

shown.

 

? C. Typical recordings obtained.

 

? Ecog: Electro corticogram Figure 3: Local field potentials recorded in humans using an array of extracellular electrodes.

 

V.Guntu, S.S.Nair, University of Missouri 9 Typical Data Analysis after measurement using

 

microelectrode array (MEA) chip Figure 4: Recorded Signal (From CNS Tutorial 2012) ? Neuronal recordings split into two

 

frequency bands:

 

? High-frequency band (>~ 500 Hz):

 

Multi-unit activity(MUA),

 

measures spikes in neurons

 

surrounding electron tip

 

? Low-frequency band (<~300 Hz):

 

Local field potential(LFP), measures

 

subthreshold activity

 

? LFP represents the summed

 

excitatory and inhibitory signals

 

and other types of slow activity

 

such as voltage dependent

 

oscillations or spike after potentials

 

? ?slower? processes, synapses, leak

 

currents, capacitive currents etc. V.Guntu, S.S.Nair, University of Missouri 10 The simplest model of LFP (Adapted from Scholarpedia) ? Assumptions: LFP is generated by transmembrane currents, Neurons

 

are embedded in perfectly resistive medium, Electrical conductivity

 

(?) and permittivity (?) of the extracellular medium are constant and

 

independent of frequency.

 

? Electrical Potential generated Is V(r)

 

Where V(r) is the extracellular potential at a position r in extracellular space, Io is the current source,

 

and Ir-roI is the absolute distance between r and the position of the current source ro The potential resulting from a set of current sources is given by : This expression can be used to calculate the LFP resulting from a

 

network of neurons or complex morphologies.

 

V.Guntu, S.S.Nair, University of Missouri 11 Frequency filtering properties of LFPs

 

? The fact that action potentials contribute little to the LFP may be

 

explained by frequency-filtering properties of the extracellular

 

medium.

 

? If the medium acts as a low-pass filter, it may attenuate more severely

 

the high frequencies (greater than ? 100 Hz), such as that produced

 

by action potentials, while the attenuation will be less severe for

 

lower frequencies such as synaptic events.

 

? As a consequence, an extracellular electrode at a given position will

 

"see" only the action potentials for cells immediately adjacent to the

 

electrode.

 

? On the other hand, the low-frequencies of the LFP will be a

 

compound signal of slower events (such as synaptic events) from a

 

population of cells around the electrode.

 

V.Guntu, S.S.Nair, University of Missouri 12 BIO LAB 5 ASSIGNMENT (5 questions)

 

Q1 : LFP using MEA

 

a. Go to the following website

 

http://vlab.amrita.edu/index.php?sub=3&brch=43&sim=135&cnt=2

 

108

 

b. Click on the ?Animation?. Participate in a full animated walk-through

 

to complete the feel for a real LFP recording. You will have to

 

click/move using the computer mouse to pass through the various

 

stages of the experiment. c. Finish ?Self-evaluation? and get a screenshot for your answers, and

 

provide it as solution to exercise 1.

 

V.Guntu, S.S.Nair, University of Missouri 13 Q2: Action potential propagation

 

? Sketch the membrane voltage as

 

measured in A and B.

 

? What is the amplitude of a typical

 

extracellularly measured signal?

 

? Is the resultant waveform biphasic or

 

triphasic? V.Guntu, S.S.Nair, University of Missouri 14 Q 3) The amplitude of the measured voltage is different for intracellular

 

and extracellular recordings. Explain clearly why.

 

Q 4) The extracellular recording changes polarity. Why does this happen? Why does this not happen with intracellular recordings?

 

Q 5) Why does LFP only capture slower currents, i.e., only subthreshold activity. Why does it not capture spikes or high frequency activity? V.Guntu, S.S.Nair, University of Missouri 15 References

 

? Axon guide

 

? Modeling and interpretation of extracellular potentials

 

? Neurophysiology Virtual Lab: http://vlab.amrita.edu/?sub=3&brch=43

 

? Principles of Extracellular Single- Unit Recording. CNS2012 Tutorial

 

? Scholarpedia : http://www.scholarpedia.org/article/Local_field_potential

 

? Wikipedia : http://en.wikipedia.org/wiki/Multielectrode_array

 

Video References:

 

http://www.youtube.com/watch?v=E8l3_kAUe0w

 

http://www.youtube.com/watch?v=RTgoRmDJvmE

 

V.Guntu, S.S.Nair, University of Missouri 16

 


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