Translator: Carmela Napolitano Reviewer: Denise RQ
You're pretty still, waiting for my talk, despite a few applause.
So you think you are doing nothing.
But, actually, one part of your body is extremely active,
actually using up to 20% of your energy.
That part of your body, of course, is your brain.
You are smelling odors, you are feeling the temperature,
you are looking at the slides, you're listening to me.
Eventually, you may be understanding what I'm saying, and, hopefully,
if I'm really lucky, you'll actually remember part of what I'm saying.
All these incredible functions we know are performed
by this absolutely fantastic machine we have in the head, that's called brain.
And despite its complexity, and its utmost importance,
and also, unfortunately, the fact that it's so affected in certain diseases,
we still know little about the brain, It's still very much of a black box.
What I want to tell you today is that, in fact, using light,
researchers are starting to know more and more about the function of the brain.
What I'm going to present you is
how, using a variety of imaging methods and lights,
we can actually look at what's inside this thing that's a human brain.
So, that's a human brain.
About 1.5 kilos of mainly water, 70% water, so 30% of tissue.
Not much, right?
But this not much is actually doing so many fantastic things:
creating arts, writing books, thinking,
making also some not so nice things, but that's another story.
We are going to see how we can look inside that using light.
To look inside the brain we need models, of course.
It would be fantastic to look directly into the human brain.
Unfortunately, I'm not so sure many of you would be ready
to give me a little piece of their brain to do experiments.
Maybe? Somebody? No. You see.
So, we need models. We did two types of models.
We did conceptual models, and we did experimental models.
So, conceptual models mean models of learning, memory, and brain function.
What's been really fascinating me from the very beginning
is this incredible function that has a brain to store information,
process that information, and use it to change our behaviour
as a function of what's happening outside, in the outer world.
There is one easy protocol, one easy concept of memory
that researchers have used a lot.
- we're going to talk about it a little - It's called spatial memory.
What is spatial memory?
When you came here for the first time, some of you probably got lost,
probably it took you a while to find the right room.
We all know that when you'll come back the second time, it's going to be easier.
And, probably, the third time,
you will come directly to this room without getting lost
because you've memorized from cues,
from indices in the environment how to come here.
So that's called spatial memory.
To work on that memory, to understand how it's actually working,
we need experimental models,
and the experimental models many, many researchers use are mice,
because mice learn very well, and we can actually study that.
We are using mainly two models. One of them is called the Y maze.
You cannot see [it] very well here, but you'll see it on the Internet.
This Y maze has cues on the side.
When the mouse is put at the maze's entry with a reward only in one of the arms,
initially the mouse doesn’t know there is a reward.
It's going to get lost, it's going to go away.
It's a catastrophe.
And then, little by little, as we work more and more and more,
the second time is better, she's nearly there, and that's it.
She found it. She found the cheese.
And she's going to memorize that, thanks to the indices in the environment.
That's spatial memory.
Another way to do that is to use what's called the Morris water maze.
What's the Morris water maze? It's a tank of water.
Mice hate water; the only thing they want to do when they're in water
is just find a spot where they can get out of the water to get dry.
Researchers are using that, putting the mice in the tank of water.
There is a little platform so that the mice can find the platform
and escape from the water.
Around this water tank there are cues, you know, diagrams and things
so that the mouse actually sees where she is,
and she can know where is the platform when she's found it.
And then she can memorize the location of this platform.
So, typically, we can then track the movement of the mice.
We measure the length, the amount of time it takes the mouse to find the platform.
As the mouse improves its performance, we can see how long it takes the mouse
to actually remember well where was the platform.
That's a very, very efficient way to actually measure memory.
So that's an example.
First, we teach the mice how to find the platform with a transparent liquid,
then we use milk in the same tank
so that the mouse doesn't know where is the platform.
So it finds it initially by random.
But then, thanks to the cues which are surrounding the tank of liquid,
she can remember where is it and then accelerates her finding.
That is a very nice paradigm. We can use that also to test for diseases.
Here, what you're going to see is, on the left side, a wild type normal mouse
and on the right side, a mouse affected with a disease
that's very close to the human Alzheimer disease.
That's a very good Alzheimer model.
These mice have been trained.
You're going to see that left mouse has learned where is the platform,
it finds the platform very rapidly;
within a few seconds, it goes back to the platform
looking at surrounding clues.
But on the right side, the Alzheimer mouse doesn't remember where is the platform.
It has forgotten. It has been unable to remember where was the platform.
That's what we want to study, really, in molecular terms,
we want to study what's the molecular, the cellular correlates of this function.
We all know that the brain information processing is mainly done by neurons.
Neurons are not the major type of cells in the brain.
The major type of cells in the brain are called ilia.
Glia play a very important role.
My colleagues who work on glia make sure I say every time that glia is important.
But even though it's very important,
it's neurons which are really doing information processing.
And that's because neurons form a very interconnected network
with billions of cells, each connecting with 10,000 of other cells.
All together is a humungous number of synapses,
those zones of contact between neurons that are present in the brain.
Those things are very small, very numerous, very complex,
and that's what we want to look at.
So, that's a spine, with a synapse, two cells connecting one to the other.
It is really thought now, really believed pretty strongly
that these synapses are the main location of storage of information in the brain.
Understanding how it works, how you can store information is very important.
The problem is that that is very small, it's so tiny.
They are only a micron size. That means the one twentieth of a hair.
We want to study those, possibly in live brain,
to understand how they are organized.
We know quite a bit about their function.
In this movie here, what you're going to see is
the basic summary of what we know about synaptic transmission in the basal state.
You have those synapses, site of contact between two cells
and then, on the bottom cell, you have the receptors,
and on the top cell, you have the neurotransmitter.
When there is an action potential that comes in,
the neurotransmitter is released in the synaptic cleft in-between the cells
and then activates the receptors in the post-synaptic cell,
transmitting information and that's the basis of synaptic transmission.
What you see on this movie is that receptors are not immobile,
they move, they move all the time,
and that's something very important we're going to come back to, actually.
So, let's go back to our synapse, the scheme of the synapse.
You see those receptors, those green and red entities.
To understand how a synapse works, how it stores memory,
we want to be able to look at those receptors in live cells,
if possible, in the live brain, if possible, even in the live animal.
To do that, what we do is
that we label those receptors with bulbs, light bulbs, they are called fluorophores,
to look at them, to try to see how they are organized.
But what I told you is that those synapses are very, very small
and there's a physics law you may remember from your school days
that's called limit of diffraction, the law of diffraction,
that tells you that you cannot look at an object which is smaller,
you can't differentiate objects which are about half the light wavelength.
It means that, although you have a single receptor, which would be a point source,
actually, the spot of the light that it gives you is very big,
it is nearly the size of a synapse.
So, all together, you cannot distinguish two receptors which are close by,
and, in fact, within the synapse, if you label them
and look at them all together, what you see is very blurry.
This image on the left, this spine which is completely blurry
because it is blurred by our diffraction limit.
Actually, very recently, only in the last few years,
researchers, with the help of chemists, physicists, and opticians, and biologists
have developed a whole new series of experimental ways to look at objects
which are smaller than the size, than the limit of diffraction.
That's called optical nanoscopy.
It's been awarded a Nobel prize to three people last year
for the development of super-resolved fluorescence microscopy.
What's the idea? The idea, in fact, is pretty simple.
It is to look at each object one by one instead of looking at them all together.
This is actually a technique that had been developed already,
nearly 100 years ago, by Neo-impressionists.
That's called pointism.
You can actually reconstruct a full image just by using several dots
and, little by little, you reconstruct the full image.
Well, we do the same thing for synapses.
We're going to label individual fluorophores,
localize with very high precision those individual fluorophores,
and do that over and over again.
From the first image on the left,
as we localize each single fluorophore, one by one,
we can get a much clearer picture of the object.
That's an example of a neuron, visualized with classical light microscopy.
It's pretty fuzzy. You see the synapses, but they are not very precise.
You cannot certainly get any information about the organization of the receptors.
And then, if you turn on this method to look at molecules one by one,
you actually start seeing something which is much more crisp.
Now you can really distinguish what's happening inside the synapse,
that's the nanoscopic scale organization of receptors inside the synapse.
But that's still fixed, that's still dead tissue,
and what we want to do is to look at that in live cells.
We can actually do that. We can track the movement of these individual receptors.
First, we can reconstitute in 3D the image of the synapse,
- pretty image -
but, more interestingly, we can track the movements of receptors.
All these little red balls are individual receptors for neurotransmitters
that we track in a real live neuron, in real time, to get their movement.
That's a big finding that we made more than ten years ago,
receptors are not immobile, but are actually moving all the time.
That's the basal state.
What happens when you learn, what happens when you store a memory?
Well, I think we've progressed in that direction, again, using a model,
using a model of neurons, and stimulating neurons electrically
to mimic what's happening when you learn something
and looking at what happens to the movement of these receptors.
That's what is represented here.
In this movie you see the same type of neurons,
we're going to follow the same type of receptors,
but this time, we are going to apply an electric stimulation,
as if you were stimulated by my talk and learning something.
What you see when the red bar happens
is that receptors actually get immobilized very quickly.
They get immobilized, they get accumulated in the synapse,
and that is, we think, the cellular basis of memory:
accumulating more receptors to strengthen the strength of these synapses.
So, in this movie there is a summary of what we think happens during learning.
There is a strong activity from the pre-synaptic cell,
a lot of neurotransmitters are released in the synaptic cleft
that activates a complex enzymatic cascade that we start to know pretty well,
and then, receptors are modified in their properties
and they get stabilized, they get accumulated,
and there is more and more receptors
that are present in front of the site of transmitter release
so that the synapse gets stronger.
That is called cellular memory,
and that's probably the substrate for when you learn something.
What's the future? Now we can look at the synapse memory.
We would like to be able to modify it, we'd like to be able to modify memories.
That will become possible very soon,
thanks to the development of a variety of other tools, genetic tools
which are called channelrodopsin.
Those are molecules that can get activated by light
to either inhibit or stimulate neurons.
You can put those molecules into live mice,
and using fiber optics - such as this thing,
exactly the same thing that you put in your mice here -
you can actually change the memory of the mice.
The dream is probably, one day, we'll be able to do that in humans,
to cure diseases and to modify brain activity.
It's a bit scary.
Of course, as always, we are scientists, there is a good side and a bad side.
But it's clear, the future of neuroscience is extremely bright thanks to light.
And I thank you very much.
(Applause)