Welcome to the final week of cellular mechanisms of brain function
We've come a long way since we began this course
where we discussed lipid membranes and protein ion channels
until last week when we discussed
the methods for studying the mouse brain during behavior.
As we think about behavior,
It becomes apparent that we need to understand motor control.
Behavior is nothing but muscle contractions
and the control of those muscular contractions
is the movement that we call behavior.
And so, now we'll begin thinking about motor control
and as we think about motor control
it becomes apparent that sensory input to motor control
is one of the mayor impacts.
We govern our lives, our movements, our behaviors
by thinking about the incoming sensory information.
And so, sensory input to motor control centers
is a key way that determines behavior.
Equally important is to think about the converse
that motor control affects sensory input to the brain.
So, for example, if we think about how we look at the world around us
we make head and eye movements
and we select regions of interest in the visual field
that we'd like to pay attention to.
And so, as a first order approximation we choose the visual information,
that falls on the retina and that flows into the brain.
Equally, if we think about our sense of touch, it's very obvious
that the way that we move our hands and our fingertips
determines the tactile sensory information that we obtain,
so in order to get the information about the texture of an object
we need to stroke that object with our figertips
and it's the movement of the finger
that generates the flow of sensory information.
So the sense of touch and vision are dominated to a large extent
by self-generated movements
that form the so-called <i>basis of active sensing</i>
where we select the type of sensory information
we want to obtain by making specific goal-directed movements.
So we can think of the basis of behavior
as a sensorimotor loop
which neither has a beginning nor an end
but we can enter it at any point we want.
We can think about neuronal computations that, of course, are influenced
by sensory information that's been gathered
and that generates motor output behavior
and that motor output, of course, interacts with the real world.
We might move our hands against an object
or move our eyes and then the visual field will change
and give rise to different sensory input
that then, in turn, influences the neuronal computation.
And in that way we have sensorimotor loops
that are the most important determinants of behavior.
In this video we'll consider the Mouse Whisker System
which is one of the key systems being analyzed at the moment
for active sensing.
Mice are nocturnal animals, they live in tunnels
and one way in which they can obtain spatial information
about their immediate surroundings is by using their whiskers
which they actively move around to explore their enviroment.
And the first thing I'd like to show you is a behavioral paradigm
where we can see just how they use their whiskers
to get special information.
In the so called gap crossing task
that was developed by Hutson and Masterton
a mouse is placed on an elevated plataform.
It's in the dark under infrared illumination
and what the mouse needs to do
is to identify the location of a target platform,
where it needs to jump over and then when it does so,
it receives a reward.
The gap between this two platforms is so large that the only way
that the mouse can identify the location of this target platform
is by using its whiskers to reach across this gap.
We're gonna look at a movie where we'll film from above
and we'll see the silhouette of the mouse with its whiskers
stands on one platform and reaches across space
to touch a target platform
where it needs to jump to to obtain its reward.
So, here you see the mouse touching the target platform
and when it's built up sufficient confidence,
it jumps and now it's enjoying its snack.
now that all happen rather quickly, so, let's look at that
slowed down twenty times and zoomed in on this area
where the animal's whiskers are contacting the target platform.
Now the individual movements of the whiskers become obvious
and you'll see that most of the time the whiskers move at the same time
both left and right and they move in concert with each other
but sometimes they become out of phase and they move individually
and if one begins to quantify the movements of the individual whiskers
you can actually see that each whisker
is individually under its own motor control.
So, clearly, if we're gonna understand
how the animal perceives its enviroment here
and gets sensory information under these conditions
it becomes important to look at the mechanisms driving motor control.
So, how <i>does</i> the mouse control its whisker movements?
There are two important sets of muscles
that are involved in generating whisker movements.
There are so-called intrinsic muscles shown here in green
that are located entirely within the whisker pad
and attached to each whisker is its own individual intrinsic muscle,
it attaches at the base of one follicle
and at the whisker behind it at the top.
When this muscle contracts the whisker pivots
around its insertion point here in the skin
and is rotated anteriorly in a protraction movement,
so the wisker moves forwards
through the contraction of this intrinsic muscle
and is pivoting around its insertion point.
So, the intrinsic muscle here, drives whisker protraction
as the animal reaches out in front of it to touch an object.
The other important group of muscles are the so-called extrinsic muscles
that are anchored onto bone outside the whisker pad
and when they contract the pull the whiskers backwards.
So, this extrinsic muscles here are relatively superficial
in the whisker pad,
when they contract they pull the whiskers backwards
causing whisker retraction.
So, we have intrinsic muscle that generates protraction
and we have extrinsic muscle that generates retraction of the whisker.
These muscles are under neuronal control.
So, each muscle is innervated by an axon from the so-called motor neuron
and these motor neurons release neurotransmitter onto the muscle.
And so, the junction between neurons and muscle is a synapse,
a chemical synapse very similar to the synapses we've been thinking about
within the central nervous system.
At the neuromuscular junction the neurotransmitter
that's being released is called acetylcholine.
And acetylcholine is released from synaptic vesicles
just like in the nervous system
and it acts upon postsynaptic ligand-gated ion channels
so called nicotinic acetylcholine receptors.
So the acetylcholine is released from the motor neuron axon,
it activates the ligand-gated ion channel, the nicotinic acetylcholine receptor.
That causes an influx of cations, Sodium importantly,
causes a postsynaptic depolarization and muscle is also excitable.
So in fact, the muscle gets depolarized and fires an action potential.
Just like a neuron would.
That action potential in the muscle
causes a rise in cytosolic calcium concentration
and that rise in calcium is what then drives
the biochemical reactions that cause muscular contraction.
So, action potentials here in the motor neurons
drive muscle contraction and can then move the whiskers
forwards or backwards and of course the same is true
across all the muscles in our body.
Acetylcholine is the neurotransmitter at the neuromuscular junction.
So now in order to understand how these whiskers are moved
we then clearly need to know
where the neurons are that innervate these muscles.
In order to localize where these circled motor neurons are,
the ones that directly innervate muscle,
we can use viral tools,
highly genetically engineered viruses
like the rabies virus here that's been modified
through genetic engineering by Ian Wickersham and Ed Callaway.
If we inject a red variant of rabies virus,
expressing so called mCherry fluorescent protein
into the extrinsic muscle, that then gets taken up by the axonal junctions,
retrogradely transported along the axon and then it labels the locations
of the neurons that project here where the motor neurons are.
We can also inject rabies virus into the intrinsic muscle
and label then the motor neurons that innervate the intrinsic muscle.
We can have two different variants.
We can have a red one for the extrinsic, a green one, G green fluorescent protein
injected into intrinsic muscle, so these are the pictures here:
the whisker pad, the injections of virus here into the whisker pad
labelling the axons here of the motor neurons
and here into the extrinsic muscle labelling again the motor neurons.
If we now cut sections here of the brainstem,
we can then see the location of where the motor neurons are.
So here's one of these coronal sections here taken through the brainstem
and here ipsilateral to the same side where the muscles are
we see the locations of the cell bodies of the motor neurons.
So these are now cholinergic neurons so they release acetylcholine
here onto the muscle
and this is the cell body and also, of course, they have dendrites
all around here that are more difficult to see.
You can also see that the location
of the motor neurons for the extrinsic muscle
are in a slightly different location
to the motor neurons for the intrinsic muscle.
And this is an individual example
and this is a group representation
of where in general we find the extrinsic
and the intrinsic groups of motor neurons.
So here you can see one of the general principles
of organization of motor neurons.
And that is that the motor neurons that innervate a given muscle type
are located close to each other, in a cluster.
So these motor neuron pools innervate different muscles
and that's typical throughout the spinal cord and brainstem.
That the cholinergic motor neurons that innervate a given muscle
are located close to each other in a tightly segregated region.
And here the two different muscles that cause retraction
or protraction of the whisker are located in different areas
of the brainstem,
in the facial nucleus, where all the motor neurons are
that govern facial movements.
Now, in order to understand what controls action potential firing
in these motor neurons,
we need to look and see
where do they receive their synaptic inputs.
So, like all other neurons, they have dendrites
and they receive glutamatergic and GABAergic and glycinergic input
distributed across their somatodendritic arborization
and the reason that these guys fire action potentials
is because of the synaptic input that they receive.
So we need to know what are the presynaptic neurons
to these motor neurons.
And the presynaptic neurons to motor neurons
are called premotor neurons.
These are the ones that give synaptic input onto motor neurons.
And we can use a trick with the rabies virus
to label the presynaptic neurons of the motor neurons.
So, as before, we inject the rabies virus to infect the motor neurons
and then we complement them with the missing factor
that allows them to jump just one synapse back
and then that labels the neurons that send synaptic input
onto the motor neurons.
And it turns out that premotor neurons
are widely distributed across the brain in many different brain regions.
And that's true for the whisker system but it's true for all of our muscles.
There are many presynaptic neurons to any given motor neuron.
One of the hotspots for the location of premotor neurons
is hidden here, immediately behind the facial nucleus
where the motor neurons are located.
And here we see a couple of sections
in coronal sections, immediately posterior
to the facial nucleus.
And you'll see that the red dot here correspond to the locations
of cell bodies of premotor neurons that innervate the extrinsic muscles
of these, generate retraction movements.
And here we've injected the rabies virus into the intrinsic muscle
and these are then the premotor neurons that drive protraction,
contraction of the intrinsic muscles and forward movement of the whiskers.
And you'll see there's an obvious difference
in the lay out of the neurons innervating
the retraction and the protraction whisker muscles.
The retraction premotor neurons are located here
in the spinal trigeminal nucleus,
whereas the neurons that innervate the motor neurons
that give rise to protraction movements,
are set in here, in the reticular formation of the brainstem.
And so there are two separate locations for where premotor neurons are located.
So one would expect then that if we stimulated these neurons
that would drive action potential firing in the facial nucleus motor neurons
that innervate the extrinsic muscle and cause retraction of the whisker.
And so if we now put a stimulation electrode here
into the spinal trigeminal nucleus and we drive current through that,
it turns out indeed, it does cause a retraction of the whisker.
This is now a horizontal section through the brain somewhere around here
where we see the horizontal section through the reticular formation
and spinal trigeminal here, more laterally.
We would expect that stimulating this area here
might cause whisker protraction.
There are many cells here, premotor neurons that innervate
the intrinsic motor neurons that drive protraction of the whisker.
And indeed, if we put a stimulation electrode here
in the reticular formation and stimulate it electrically
we see that the whisker indeed moves forwards.
Compatible with our premotor mapping.
Now, there are many sensory motor loops at the level of the brainstem.
So sensory information, both from the whisker system
and from many other types of sensory inputs
impinge upon these premotor neurons.
And in general, there's a great deal of local circuitry
in the brainstem and the spinal cord that gives rise to reflex movements
and even more complex sensorimotor contractions.
However, if we're interested in volitional movement
then typically this is something that results from cortical activity.
So, if we're thinking about volitional control of whisker movements
we might be interested in the cortical inputs
to these premotor neurons.
In general, cortical neurons don't connect strongly to motor neurons
and most of the impact of the neo cortex upon movement
is indirectly through complex brainstem and spinal cord circuits
through interacting with the premotor neurons.
So here, we then introduce an anterograde label for the axons
into the whisker sensory cortex and into the whisker motor cortex.
Two areas that we think are intimately involved
in processing and generating whisker movements.
If we inject this virus into the sensory cortex,
we can see the axons down here in the brainstem.
If we cut a coronal section then, we can look and see that the axons
are coming in here from the sensory cortex and largely innervating
this spinal trigeminal interpolaris region,
exactly where the extrinsic premotor neurons are located.
So you might imagine then that if we stimulate sensory cortex
that would release glutamate in this area, stimulate these neurons here
and that should then cause a retraction of the whiskers.
If we now put the virus to label the axons into the motor cortex,
we then cut sections here in the brainstem and we look and see
in these sections where the axon's terminating,
we see that there's a great deal axon that's terminating here,
in the reticular formation.
And that's where there's a high density of neurons that are involved
in protracting whiskers.
So you might imagine that stimulating this area of motor cortex
might then cause the whisker to move forwards.
So we can test this hypothesis by putting channelrhodopsin
in these two different brain areas.
Here we've injected a virus to express channelrhodopsin
in the sensory cortex.
Here we've injected channelrhodopsin into the motor cortex.
This is now the fixed brains that have been removed
from the animal after diffusion.
While the animal was alive, we put blue light
onto this spot of the cortex and saw
that indeed generated a whisker retraction.
So stimulation of sensory cortex causes the whisker to move backwards
while stimulation of motor cortex causes the whisker to move forwards,
but it does so in a more complex way,
it's a rhythmic forwards and backwards movement.
Very much like what we saw during the movie when the animal
was performing the gap crossing task.
It's this exploratory rhythmic palpation of its surroundings.
So as a summary when we think about whisker motor control by the cortex,
we can see that the sensory cortex sends axons down to the brainstem,
innervates spinal trigeminal interpolaris nucleus.
That has premotor neurons that innervate facial nucleus motor neurons.
And those facial nucleus motor neurons
are the ones that are involved in retracting the whisker.
So, activity in the sensory cortex causes the whisker to move backwards
and that could be viewed as a negative feedback signal.
And perhaps, that's useful if we think about, say ourselves
that are exploring a dark room, we're moving our arms around
and when we contact the wall the first thing we might want to do
is to stop moving our hand forwards.
We can't bring that hand through the wall, so we need a negative feedback signal
that prevents or stops that movement.
So that might be one of the purposes of the sensory signals and sensory cortex,
is to cause a feedback signal to driving whisker retraction.
On the other hand, the whisker motor cortex
does the opposite, it tries to get more sensory information
and moves the whiskers forwards
to try and get more sensory information to flow in.
And the way it does that is by its axons
that project down to the reticular formation,
here there are complex neuronal circuits
that probably act as central pattern generators
that then innervate the rhythmic whisker protraction neurons
that are the motor neurons for the intrinsic muscle
that then drive the whisker to move forwards and backwards.
So, frontal cortex is involved in trying to get more sensory information
and the sensory cortex seems to be involved in a negative feedback signal.
So there are two distinct types of cortical control signals
for whisker movements.
So, having seen what the cortex can do when we artificially stimulate it,
it's also interesting to see what types of signals occur from the periphery
and whether we can follow through sensorimotor loops.
And in particular here, what we do is we deliver a stimulus
onto one particular whisker.
The C2 whisker we have a piece of metal attached to this
and there's a magnetic coil underneath.
We can generate a brief one-millisecond pulse onto this whisker
that then activates the signalling pathway.
First, glutamatergic synapse in the brainstem.
Second, glutamatergic synapse in the thalamus.
Then the information is processed here in the primary somatosensory cortex.
After we deliver a whisker stimulation, some tenths of a millisecond later,
we can see the activity here in S1, it spreads within S1,
and it also sends a signal here to the motor cortex.
And we believe that's a direct monosynaptic projection
from somatosensory cortex to motor cortex.
What you see here is that we're also filming the behavior
of the animal while we measure the activity in the brain here,
with the voltage sensitive imaging technique that we've described before.
The animal's sitting still at the beginning of the trial,
when we deliver the stimulus but by the time we've seen this activity
in sensory and motor cortex, afterwards the animal in this trial
is moving its whisker, and what we think it's happening
is that the animal is sitting here waiting, waiting and suddendly
there's this unexpected sensory input that arrives on the whisker.
And the animal might wonder "what was it in my environment
that touched my whisker?" and so it stops moving its whisker around
actively trying to explore, looking for the stimulus
that arrived on its whisker.
And of course, there is none
because it's an artificial magnetic stimulus
that we've delivered.
And it seems that the underlying neuronal activity
at least at the level of the cortex, is a large response in sensory cortex
that might drive a small whisker.
Retraction followed by activation of motor cortex
that might then drive forwards and backwards movements
as the animal actively explores it's environment
looking for what stimulated it.
Now it turns out that there's considerable trial to trial variability
especially in the neo cortex.
So even though one gives the same sensory stimulus
time after time again,
each time there's a slightly different response
on the neo cortex.
And probably tha relates to context-dependent processing
of sensory information.
So here is another trial where we stimulate the animal
sitting still at the beginning of the stimulus
but in this case,
the animal doesn't make any exploratory self-generated movement afterwards.
It simply ignores the stimulus and sits still.
If we now look and see what happened in the brain on this trial,
we'll see that there's actually a relatively small response.
There's a little bit that happens here in the sensory cortex
and nothing in the motor cortex.
These are just two individual trials but we can select for all trials
where the animal makes a sensory evoked whisker movement,
and we can select for all trials where the animal makes no movement
and we can see in general how the pattern of neuronal activity occurs.
And you'll see that on the trials where the animal
begins to move its whiskers and explore its environment,
that's accompanied by large activation in sensory cortex
and also substantial activation here in the motor cortex.
Whereas on trials where the animal doesn't make any movements,
it just simply ignores the stimulus,
we'll see that there's a sensory response in primary somatosensory cortex,
it's smaller in amplitude,
and there's nothing activating the motor cortex.
So, at least at the level of correlations, we can see
that there's sensorimotor activity when the animal begins
to actively explore its environment driven by a sensory stimulus
and there's other trials where the animal ignores that stimulus
and that's accompanied by very little activity in the neo cortex.
We can also go and see what happens from the other perspective.
See what happens as the animal moves its whisker and contacts an object.
So, here we're tracking whisker position and looking at membrane potential,
below in black,
and at some stage there's gonna be a drop of this pole, it drops down
and then the animal's whisker is now contacting that object
and as it contacts the object, there are obvious
membrane potential fluctuations that correlate with sensory responses
as the animal touches repeatedly this pole that's been put in its way.
And if the animal needs to localize where this pole is,
clearly it needs to take into account the motor position of where the whisker is
as well as the timing of that contact in order to extract the position
of where that pole is.
Here is another example of a different cell
recorded where each time the whisker is touching an object
it's shaded in gray and you see every gray shaded area here
correlates with a membrane potential depolarization
in the cell that's being recorded.
And here there are some action potentials
that are being fired in response of this particular contact here.
So we can see that there's active encoding
of each individual touch response
in the primary somatosensory cortex of the mouse.
So there's a clear signal in the brain
that corresponds to each active touch as the animal contacts its environment.
Now we need to be able
to put those sensory and motor signals together
in order, for example, to extract the position of the object.
So it's interesting to note
that there's a substantial motor input to sensory cortex
as well as across the sensory cortex that sends input to motor cortex
so they're bidirectionally coupled.
Here we just look at one aspect,
that the motor control signals arrive in sensory cortex
this is the axonal innervation
of primary somatosensory cortex.
And it seems to come in two layers,
There's an upper layer of innervation here
that arrives at the surface of the brain in Layer 1
and there's also deeper innervation.
If we now focus our attention here, on the superficial innervation of Layer 1
that's particulary interesting
because it contacts the distal dendrites of the excitatory neurons in this region
and it's actively controlled by the GABAergic inhibitory neuron
the somatostatin Martinozzi cells
that send their axons here into the upper layer of the neo cortex.
So the input for motor cortex
in terms of its impact upon the dendritic integration here
on the excitatory neurons is regulated by how much GABA
is being released by the somatostatin cells.
When the animal's sitting still and not moving its whiskers,
these somatostatin cells are firing quite actively.
So they are inhibiting these distal dentrites
and it may be that the motor input makes relatively little impact
upon the excitatory cells.
However, when the animal begins to move its whiskers,
the somatostatin cells hyperpolarize, they stop firing action potentials
and now, the GABAergic inhibition that was strong before,
is now taken out and the motor input can now make a substantial input
onto the excitatory cells.
So, there's an interesting regulation of this distal dendritic input
by the somatostatin cells
and that then might allow for motor interactions
onto the excitatory neurons.
That then might help the animal interpret the sensor information
coming in during active touch for example,
by combining motor signals with the upcoming sensory input
that also arrives, of course, in sensory cortex.
So in this video we've seen
that if we're interested in understanding behavior,
we absolutely need to investigate motor control.
Motor control and movements are the very basis of any behavior
and so, that's an important starting point.
We've seen that sensory input to the motor control systems
is extremely important and forms the first way
in which our motor output is regulated.
We have various sensorimotor feedbacks that drive reflexes for example,
and at higher levels we can imagine
that sensory input often initiates our movements and certainly
helps us guide us through any action that we whish to perform.
From the other point of view, our movements and motor control
make a big difference to the type of sensory information that we obtain.
So we actively touch objects, we make eye movements
to select things that we want to analize in higher resolution.
So active processing of sensory information is extremely important
both in terms of the information that we obtain and also in terms
of selecting for specific types of sensory information.
So in this video we've largely looked
at what are probably hardwired genetically programed
sensorimotor loops and sensory interactions.
In the next video, we're going to see
how we can use reward-based learning
to change sensorimotor interactions and drive goal-directed behaviour.
And that is essential as a starting point for understanding sensory perception.