Getting back to our electrophysiology
session, we're on session 2 and we're going to focus more
on neuronal potentials. And these neuronal potentials
are the electrical activity that we're trying to record
from our electrodes in the brain. So when we sample the brain, there are
many different ways of sampling it. We could do structural imaging,
we could do blood flow measures, we could do radio-active particles,
magnetic fluxes and electrical potentials. And there are positive and negative thing
s
to each of these sampling methods. However, I will argue that
the electrical potentials are recording from the individual neurons in the brain
and not trying to see a mass activity. We can actually go down to the single
neuron and see how it's involved in a particular behavior,
such as moving your arm, moving your leg, how you learn to move your arm,
how you learn how to move your leg, and that these other measures,
they do have some advantages because some of them are noninvasive,
for instanc
e, structural imaging, very, very informative to help us identify different structures
in the brain to find abnormalities. However, you can't see activities here.
Within these different structures, there are multiple functions, so there are
no lines saying this part of the thalamus is involved in motor function
and this part's involved in vision. There are no lines here. However, at the single unit
electrical level, we can go in and sample these various parts of the brain and say:
"Oh, this is a
motor area, this is a non motor area."
We can look at blood flow. This is an example of what's
called functional MRI. And with functional MRI,
we can use various things to look at blood flow
from the imaging technique. So it's pretty valuable. We actually can identify general regions
involved in arm movements. We can have the patients play tasks
involved in learning and see what parts of the brain are getting more blood
flow during that process. However, one of the drawbacks is, it
gives you a
general area and secondly, inhibitory mechanisms in the brain
require energy just like excitatory. So it doesn't tell you if that area
is inhibited or excited. At a single unit, with electrophysiology, you can actually
see if the neurons firing more or less. So you get a concept of inhibition
versus excitation. This just tells you what part of the brain
is utilizing energy. We can use radioactive particles
that go into the brain and bind. Here's an example of fluor gold. And we can look at metab
olism,
for instance, essentially energy utilization as well. This is PET imaging, again,
a noninvasive technique. You can go in and look
right after a seizure and say, is there a part of the brain
that's more active than it normally is if you compare it
to a non seizure time period? So it does have value. However, it doesn't give you that
individualized value. What is this
part of the brain exactly doing? And is it excited? Is it inhibited? It doesn't give you that information. However, it does
provide important
information for certain diagnostics. We also have what is called Mag. Here's an example. There's the shielding
and within the shielding or hundreds of small inductors
that can measure magnetic flux. So yet and a noninvasive method, you
put your head in here, you have a seizure in there, you can pay a behavioral task,
look in on arm function, for instance. And what will happen
is, as the part of the brain involved in that function will have a change in its magnetic properties,
w
hich then are captured. And we can reformat
that back into the brain to know where that flux in magnetic field was.
Again, not highly localized. You can't tell
what the single neurons are doing. And is that flux due to inhibition
or excitation? The other problem with these,
especially mag, is localizing where the activity is because the power
of the magnetic field shrinks by distance. And the closest thing to these inductors is
the surface of the brain sometimes is very difficult to know
if it's
deep in the brain or if it's the cortex of the brain. So it becomes very difficult to localize
with this technique. And again, I'm going back to single units. I am biased,
but I wanted to bring this concept to you about how I
look at the scales of recording. And this is a very simplistic way
of looking at it. So imagine that the macro context
is a really big speaker or microphone and you place it
in the middle of the cafeteria. What you're going to pick up with that
microphone is the talking in
the hum
of everybody in the room so what you can get the concept of
is the basic hum in the room. When we go down
to the local field potential. Now instead of a big microphone
in the middle of the room, we put a microphone
between two tables. Now we can discern maybe not individual
speakers at the two tables, but basically the information
of those two tables, the hum of those two tables, now eliminating
the whole hum of the cafeteria. And when we go down
to the actual individual recording the
single unit recording,
we can hear each of the individuals at the table now
and then we can move our microphone from table to table and sample individuals
at the different tables so we can understand
how each of those individuals now contribute
to this whole harmony of the brain. And that's an important concept. We're going down and down,
higher and higher resolution in our information
and down to the individual. And then now for the first time,
the electrode really capture that whole domain fro
m what the whole room is doing to
what the individuals are contributing. And we can imagine
if we just take epilepsy, maybe there are two epileptic cells
that you can record from at an individual table
and you're like: "Wow, they're involved
in the epilepsy." Once we make the microphone bigger
and bigger, our ability to see these two abnormal people or cells
is diminished because we get the hum of all this overriding
what you can hear from the individual. So my hope is for epilepsy
that we're g
oing to find out a whole bunch about how individual cells
may look normal, but yet we don't see that same abnormality
at macro level. And this may provide great insights
on change in how we diagnose foci. We have this big macro contact
recording the whole room when we're trying
to find some abnormality, and it may be just washed out
by the number of people in that room. As I told you,
we have the sodium potassium pumps. It creates a charge differential. So that's called rest potential. When we h
ave a ligand come in and open
channels, that change occur, more potentials
to move along the cell membrane. They could be excitatory,
as in this case or inhibitory. Oftentimes inhibitory mechanisms require
actually more energy than an excitatory mechanism, because they involve
what are called second messengers. So the neurotransmitter binds,
causes some change in a protein
that then affects this protein, this effector protein
then opens some channel. More complicated, these are called
G-coupled
proteins, they're inhibitory. So here's an example of more energy
being utilized to cause an inhibitory process
than the cause and excitatory process. Both of them utilize energy. When you record local field potentials, you're primarily
recording this type of activity, and it's mirroring the dendritic activity
that we're recording. When we do single unit physiology,
we look at an action potential. We're looking at what that neuron is
outputting. We're at macro recordings, we are looking
what's h
appening at the input of the neuron, not at the output,
uniquely distinct types of information. So as I said, we can put neurotransmitters
that cause or stimulate
which cause a depolarization of the cell. And if it depolarizes enough
and that activity gets to the axon hillock, it will generate an action potential. We can also hyper polarize the cell. That means inhibit it. So if we stimulate in a negative way, we can actually inhibit
the cell, prevent it from firing. The neurotransmitters do bot
h. So they're coming in. Some are exciting, some are inhibiting,
and that activity is traveling through the dendrites
to the cell body, to the axon hillock. And it summates
the positives and negatives. And if you have equal positive or
negative, you get zero changing current. If you have more excitatory or
depolarizing activity than hyper polarizing and it's sufficiently large enough,
it will generate an action potential. I talked about tempering
spatial summation, so if enough information acti
vates the axon hillock,
that activity traverses the axon in the form of an action potential down
to the synapses, the action potential then cause the synapses to release drugs
or neurotransmitters onto the next cells,
whether excitatory or inhibitory. And then they do their summation
and project to the next neurons, either an action potential
or they don't fire at all. So this is the shape of an action
potential, and it's an idealized shape. In fact, different cells in the brain
have different s
hapes of their action potentials, and that's because they have
different distributions of channels. And so let's just look at what's happening
here. The differential in charge
between the inside and the outside of the neuron is
is at about -70 millivolts. Some neurotransmitter comes
in, it opens some of the sodium channels
at the X on the on the cell membrane. And if a sufficiently large enough of them
come in and the current gets high enough, it will trigger
the rest of the sodium channels to o
pen. And then you get an all or nothing event. It will shoot all the way up. Here's your action potential
and during the different phases of your action potential,
different channels are opening. So at first the sodium channels are opening,
causing a positive change in the voltage. But as all the sodium channels
are opening, they reach some point where now all them have opened
and some of them start to close again. Also, at the same time, these potassium
channels open around the peak, which then
starts
to make the charge go negative. And in fact,
all the sodium channels have now closed. They're all going, the potassium is coming
in, reversing the polarity. And then down here, it's hyper polarized,
which means it's below its resting potential. And this hyper polarization is actually necessary
to reset all these sodium channels. We have the sodium channels
which drive the voltage up. They shut down and the potassium then
invades and then the voltage goes down. And that hyper polarization
period
makes the proteins of the sodium channels go back until they're ready
to open phase again. And then the resting potentials here,
interestingly enough, after this hyper polarization period,
this neuron is more likely to fire because we'll say 100% of its sodium
channels are available to open. So you can actually activate
a neuron right after this hyper polarization and it's more likely to fire
than any time period after. The whole growth of our recordings
is to get clean recordings. And w
e can see each of these inflections
is a neuron firing. And we could see even in the same
recording, there are different neurons. So we have this neuron here
who fires higher than this neuron here. To get this type of recording,
we have to have the right type of contact close to the neuron
and void of environmental artifact. We'll talk about that as we go along. But I wanted to point out that,
as I had said, neurons fire differently,
their shape is different. And I told you
the action potential
is different. And we could see
this is one part of the thalamus here, the dorsal thalamus, how it fires. This is a VIM of the thalamus, it fires differently. We can see even
it has two different types of cells: it has a more sustained firing and it has another type of neurons
that kind of burst fires. Then when we get down further
in the thalamus, we get into a different type
of firing pattern. So these different patterns of firing,
the shapes of the neurons all allow us to know
what part of the
thalamus that we are in. There are different factors
that lead to the ability to record these spikes in this fashion. And if we don't meet those factors, it looks more like a real potential
rather than a single unit. And we can't discern individuals. We certainly won't be able to see that
this bursting pattern relative to this and that comes down to the dimensions
of the contacts, the impedance
and what we're actually recording from. I just have these three different things
here. This is a sing
le unit
electrode used for deep brain stimulation. The tip on it is roughly
30 micron exposure. This is what we record single units
for when we go in for DBS cases and we can record those nice, crisp recordings
that you saw in the prior slide. However, if we go to scalp, we have these
really big contacts on the surface recording from thousands of units,
they're not close and some of them may be deep so it becomes harder and harder
to isolate individual function. Now we're going to look at a DIXI
implant
and we can see these very, very nice electrodes
with the metal contacts. Each of these dark spots
is a macro contact. Now we're within the brain,
we're closer to the population. We're listening to the cafeteria here and we're listening
to the individuals at the tables here. And it requires different dimensions
of the contact and different impedances The impedances on these macros
are far lower than they are in the micro. So examples,
local fields or EEG, ECog, SEEG, With with single uni
t,
there are three different types. Most of what I'm showing you
is all extracellular activity, but there's also intracellular.
Right now I don't I don't believe there's any way of doing intracellular in human. What we do in these experiments
is we put a glass pipette inside of the neuron
and we only record from that one neuron. The patch technique, we actually put the glass pipette to the membrane of the neuron
and we put a little vacuum and we're only recording the activity
of that membrane,
one neuron. Extra-cellular, we have neurons outspaced
and our contacts between them and we're seeing the electrical flux
recording from the extra-cellular space. The problem is as you get lower,
lower resolution with extra-cellular, I can discern maybe three neurons
in a recording. Intra-cellular, you're only seeing one neuron at a time
and with patch, you only see one neuron at a time. The next level of extraction is what's called multi-unit,
and that's where you can see inflections. But your r
ecording is not good enough
to isolate individual neurons from those inflections. So you may have three neurons,
but the spikes all kind of look alike and you can't separate them. So this extra-cellular recording
gives you that ability to separate. And typically what the difference is
is the impedance of the electrode or your ability to get close
enough to the neurons which separate true single unit, extra-cellular
from multi-unit. Here's an example of a cell attached,
this the membrane. You can
see the cell body
as being invaginated a little bit
because the glass is pushed on it. This is an example
of an extra-cellular contact. Here's an individual neuron. Here's another individual cell body
of a neuron, another one and another one. So if we look at the single unit contact
coming in to do the extra-cellular recording, we are able to get close to this neuron.
So the spike we detect from this neuron will be larger than the spike
we detect from this one, and much larger than what we
cou
ld detect from this. So we might be able to discern these three neurons
from our recording based on height. However, this guy might be too far away
and we're not able to separate it. If you don't have a good enough electrode, a high impedance,
a good enough impedance electrode, what you do is record from all three
and they all kind of look the same. The heights are almost together. It's very hard
to separate the individual cells. But I want to bring your attention here. I told you that the micro
contact
exposure the tip is only 30 microns in size, the cell body of a neurons about 10.8. And here's an example of our electrode. It's 15.9 micron cross-section. So we're actually on the order
of the size of the neuron when we're recording with our electrode. One other thing to point out here, our
macro contact is two millimeters in size. So if you were to have that on the screen it would cover
everything here in relation to this image. So imagine how many neurons
surrounding a macro contact
versus the fine resolution
you get of a single unit contact. So what we're measuring here,
these sharpened sections are individual action potentials. The height, I said, is determined
by how close you are to the neuron. Actually,
the voltage drops off at one of our square in relation the magnetic field
drops off over one over cubed. So a small change in distance
is a huge drop in magnetic field, whereas electrical fields drop off at r squared. Underlying
each of these inflections is an action po
tential
of a slightly different morphology, depending on the cell. The firing pattern,
cells have different firing patterns. Obviously,
if let's say you're in a motor region and you move your arm,
this pattern of firing will change. It may go up or down in intensity. I'm blowing up this picture
to show you scale. On the bottom here, we have two and quarter
a second of recording. You can see this neuron is firing
quite fast. Neurons can fire 40,
50, 60 spikes a second, down to two, three.
If I b
low up just a quarter second, we can see here
I was able to separate this spike, which is labeled green,
here it is again firing, labeled green From the other spike in the recording,
which is this one labeled yellow. And what we can see is they kind of
look the same, the yellow ones and the green ones
kind of look the same. And that's because when they fire
an action potential, it's a very consistent, reproducible,
logical flux. So spikes,
I don't sit here and blow up orders and then say this i
s neuron one
and this is neuron two. We have very sophisticated software
to help do this. So down beneath here we can see a trace
and we can see there's spiking activity. It's not the best picture,
but then we have a computer algorithm that can separate it
into an individual neuron. In this case,
it separated this recording into one, two, three, four,
five different types of neurons. We do go back and check to make sure it's a reasonable way
of separating the neurons. Now, you can imagine when
I started doing physiology,
we recorded one electrode at a time. With this new MME we have, we can record, depending on the model,
up to 12 recordings at a time. There are other types of electrodes
where you can record hundreds of single channels at a time,
so we need computers to help us. There's
no way we can monitor all 100 channels. How do we source 100 channels? We'd be in the lab just doing that
all the time instead of the analysis. So we've developed algorithms
to help us go through this
process. I'm not going to get too much into data
analysis. What I will do is show in later segments
some of the type of studies that you can do in a single unit. And that pretty
much is the end of this session.
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