- Hello, I'm Cindy Forehand, Dean of the Graduate College
at the University of Vermont.
We're here this afternoon to welcome you to the first
of four university scholars lectures
for this academic year, in what is the 40th year
of our University Scholars Productions.
As always, we're here to celebrate the high value UVM places
on research, scholarship and creativity.
The University Scholar Award Program recognizes
outstanding faculty members each academic year
for sustained excellence in research,
scholarship and creative arts.
Today, we recognize one of this year's award recipients,
professor Gregory Holmes, a colleague in the Department
of Neurological Sciences within
the Larner College of Medicine.
Greg Holmes is chair of the Department
of Neurological Sciences and a Professor
of Neurological Sciences and Pediatrics.
He is an internationally renowned pediatric neurologist
with an expertise in pediatric epilepsy.
His research in the laboratory and clinic
has resulted in an understanding
of the pathophysiologic basis of cognitive impairments
and other comorbidities in children with epilepsy.
Dr. Holmes studied at Washington and Lee university
for his bachelor's degree and the University
of Virginia School of Medicine for his MD,
followed by residencies in Pediatrics at Yale
and Pediatric Neurology at Boston Children's Hospital
and Harvard Medical School, where in 1996,
he became Professor of Neurology
and was bestowed an honorary M.A degree.
Prior to becoming Chair of Neurological Sciences at UVM,
in 2013, Dr. Holmes was Inaugural Chair
of Neurology at Dartmouth Medical School.
His research is inspired by his patients and their families.
He has over 500 peer reviewed research articles,
books, book chapters and review articles.
He's enjoyed continuous funding for more
than 40 years from federal agencies, primarily the NIH.
Dr. Holmes served on the editorial boards of 12 epilepsy
and neurology journals and has been
on multiple NIH Study Sections and FDA Advisory Committees.
He is a past President of the American Epilepsy Society
and has received many honors, including research awards
from the American Epilepsy Society, NIH,
American Clinical Neurophysiology Society,
International League Against Epilepsy
and The Child Neurology Society.
From 2010 to 2012, he was a member
of the National Academy of Science sponsored committee
on the public health dimensions of the epilepsies,
culminating in the book Epilepsies Across The Spectrum.
On the lighter side, Dr. Holmes is known
for a large collection of ties featuring
the classic Disney character Mickey Mouse,
and also for a frame toilet seat hanging in his office,
which was gifted to him by a prior institution.
I am pleased now to introduce professor Gregory Holmes
for his inaugural university scholar lecture, titled,
Construction and Disruption of Spatial Memory
Networks During Development, attendees may submit questions
through the chat at any time throughout the lecture.
And professor Holmes will answer questions for about
10 minutes at the conclusion of the talk, professor Holmes.
- Okay thank you very much Dr. Forehand.
I really appreciate that wonderful introduction.
And again, I wanna reiterate, I'm just so proud
to receive this award, when you look at the number
of people over the last 40 years, who've gotten this award,
it's really nice to be in the same category
of my predecessors here, so I really do appreciate
the committee that picked me.
I also appreciate Dr. Rod Scott and Margaret Vizard
for writing a letter of nomination.
So again, I'm very happy to be here.
When I heard about this talk, I was told that I was gonna
be speaking in front of an empty auditorium
and that's not actually true, I have a five people here
and that's probably five more than I usually get it
at most of my lectures, I often have trouble getting my wife
to come to my lecture, so this to me is a big crowd.
So I appreciate you guys for being here.
So I'm gonna talk a little bit about epilepsy
and some of the research we are doing,
and you know, this is a broad audience.
So just a few definitions, a seizure is considered
the clinical manifestation of abnormal excessive
and hypersynchronized neuronal discharges
and focal or widespread brain areas,
resulting in an abnormal behavior.
Now what's fascinating about epilepsy
is that abnormal behavior can be quite varied.
It can be children just have simple staring attacks,
absence seizures or they can have more
cause severe generalized tonic-clonic seizures,
often called convulsions.
Now epilepsy is a spectrum of neurological disorders
characterized by spontaneous recurrent seizures.
So if you have a child like I did,
who had a febrile seizure,
that's not really considered a epilepsy.
'Cause that was a provoked seizures,
epilepsy is really unprovoked seizures.
Now I trained at a children's hospital in Boston
and ran the epilepsy program there for many years.
The person that founded the epilepsy program
was William Gordon Lennox, and a classic book in 1960.
He talked about the many myriad aspects of epilepsy
recognizing that this is a spectrum disorder.
What's interesting, he talked about mental impairment
in patients with epilepsy and in this figure here,
what you can see is that, he looked at
the lifetime accumulation of people
that had grand mal or generalized tonic-clonic seizures,
and found that if you had a thousand
or more during your lifetime, your chances
of having cognitive impairment were over 60%.
If you had petit mal or absence seizure,
the incidents of having cognitive impairment
was actually quite low.
So this is a pretty dramatic finding written in 1960.
What's been shown over the years is that epilepsy,
especially in young children can be quite devastating.
Now I will say at the onset that most kids
and most adults that have epilepsy actually do quite well,
but there's a subset of kids and that if they
start having seizures early in life
and they do not get controlled,
that means they're pharmacoresistant,
the medications do not work.
The chances of them having normal
cognitive function is very low.
So if you look at children, say at age two,
who do not achieve control of their seizures,
the chances of them having
cognitive impairment is really, really high.
Only a 20% will have an 80 score
on the Vineland, which is administered yearly.
So what we see here is that early life seizures,
especially if they're not controlled
with medications can be quite detrimental
and quite devastating to the children.
Now, if you started in an older age,
it really doesn't matter if you get control or not.
We usually don't see any cognitive
deterioration in those children.
Now there is a group of epilepsies
that are really devastating.
And these are called the epileptic encephalopathies.
And this refers to a group of disorders
in which there's unremitting, epileptic seizures.
These children have severe cognitive behavioral impairment,
and this occurs over and beyond what
can be expected from the underlying pathology.
So epilepsy is not normal so it's often caused
by a variety of different brain injuries
and those brain injuries themselves
can lead to cognitive impairment,
but there's indication at least in some children
that the seizures themselves can lead
to cognitive impairment or further the cognitive impairment.
In addition to frequent seizures, the children often go
from a normal EEG here to one that's quite disorganized,
very slow with a lot of epileptiform activity.
So epileptic encephalopathies are really
a devastating condition and it's tragic
to follow these children over time.
Now I've been working with a person named Susan Axelrod
for over 20 years, a couple of decades.
And Susan, you may recognize
that she's married to David Axelrod.
Susan's story began when she had a seven month old child,
Lauren who's pictured here and Lauren
was fine at birth, doing well and then developed epilepsy
and had very severe seizures.
And just did not do well at all.
She's now an adopt, she is in a group home
and she's functioning well, but she has a very low IQ
and has a lot of social difficulties
as well as intellectual problems.
So Susan, when she had this, she went to website,
she looked into the Epilepsy Foundation.
And what she read about epilepsy was that, you know,
it's a pretty benign condition, you know,
often in the epilepsy foundation website,
you would have these cute kids that probably
had petit mal absence seizures and looked absolutely fine.
And she, you know, she said,
this is not what I've been going through.
My child is not fine, my child is doing terrible.
And so she founded what's called CURE.
And this is an organization that's been incredible
over the years, emphasizing that epilepsy
is not a nice disorder to have,
looking at the number of people around the world
that have epilepsy, 50 million and also pointed out,
and this was coming about when she formed CURE that
it can be a lethal disease that patients can die,
either during a seizure, even in between seizures
at a much higher risk than the population in general.
So as Susan and I were talking one day,
we were at a grant meeting talking together
and she was talking about Lauren.
I was telling about my experience
with taking care of patients and almost simultaneously,
we both said at the same time, the problem with epilepsy
is that it really I'll say stinks,
but you know what, I'll not.
So it's a bad disorder, it's not nice to have.
Okay, so the problem that most of the kids
have with epilepsy that have cognitive problems
are ones of learning and memory.
So just a little primer on memory,
we break it down into declarative memory
and then non-declarative memory.
Declarative memory is like somatic memory,
this is like how you remember general information.
Episodic memory would be personally experienced events.
So declarative memory really comes from the part
of the brain called the hippocampus, which is here.
So this type of memory is hippocampo dependent.
It's often referred to as explicit memory.
And then there's non-declarative memory.
And this is information you cannot describe.
So this would be, you know, you learn to ride a bike
and you can remember how to ride the bike
and you can't describe how you learned
how to ride the bike, but you learned
how to ride the bike and habits.
And these non-declarative memory comes from
a part of the cerebellum and the basal ganglion, excuse me.
So there is a concept called infantile amnesia.
Excuse me, just a second here.
And this was actually described by Sigmund Freud.
Okay, thank you.
In which he noted that children, infants,
did not have much memory when they got older.
So there is infantile and there's childhood amnesia.
The infantile amnesia is described
as children before the age of two,
who cannot form autobiographical events
that happened in a spatial temporal context.
And then childhood amnesia is three to five years of age
where the children are remember less
than you would expect from simple forgetting.
So knowing this, I always tell my parents
that they're absolutely wasting their time
taking their four year old to Disneyland
and spending a fortune 'cause the kid's not gonna remember
anything about it when they're older.
So we can also look at memory, how it develops.
And so there's both an egocentric
and an allocentric spatial memory.
So egocentric, that refers to your body and you know,
you go to your right or you go to your left.
It's based on what you're doing.
Allocentric memory is that you look at objects
in the environment and then you figure out that
the where of the, what, where and when is based
on this information where you can encode it
and ascertain from these different cues
where the object really is, it has nothing to do with you,
it's these objects around you that you can then
put together to develop allocentric memory,
so you can remember the fire hydrant
is between the car and the bike here,
whereas egocentric, you just know that to find
the fire hydrant you have to go, right
or you have to go right or left.
You're not using any other cues around the environment.
Now I'm not gonna be cutting up baby,
babies have amnesia and that doesn't
mean babies aren't smart.
If you do a paired visual comparison test to assess memory,
what you do is you have the child look at two objects,
for example, that are the same,
then you substitute an object that's different.
And then you substitute another one, maybe different color,
different facial figuration and a normal baby,
will look at the novel object more than the familiar object.
And so babies can learn to remember things
during the first year of life,
but this isn't considered to be non-declarative in type,
so it doesn't depend upon the hippocampus.
Beautiful work by Rovee-Collier and colleagues,
who'd take babies and they had put them in a crib
and then they'd have a mobile and if the child came in,
they'd put a string on the child's leg,
connected to the mobile and if the child kicked
the leg, nothing would happen.
On day two, you come in and put the same thing
on the child and the child kicks the leg
and all of a sudden this mobile is moving.
And then your third day is you take it
and it doesn't move again when the child kicks.
So you compare the amount of kicking between
this day and this day and if they kick more on this day
than the first day, that means they remember
that kicking will cause the mobile to start moving around.
If you do the same thing with the train here,
you can just have the child push a button,
nothing happens next day, you push a button,
the train moves, third day he comes in
and he pushes the button, nothing happens,
subtract this from this and it demonstrates learning.
The child remembers that pushing the button
will cause the train to go forward.
And these are my kind of tests
because I think even I could handle them.
So another way to look at the development
of memory is to do a test and where you take a child
and you put them in a room and you have all these plates
on the ground and if the child picks the right plate,
there's a little toy or a piece of candy under the plate.
So the child has to go into the room
at different angles, at different entrances in other words,
and this particular experiment, they put a marker
on top of the plate in which the toy was located.
So all the child had to do is know to go right
to that marker, pick up the plate and he would get the toy.
The next step and that's local cue learning.
That's egocentric learning.
The next time you'd bring the child in
and you don't Mark any of the plates.
So the child has to remember where the plate is
based on cues around the room.
So this is an example of allocentric memory.
And so allocentric memory as I told you is hippocampus.
Local cue memory is non-hippocampus.
And it turns on at around a 43 to 46 months,
children learn to do the allocentric learning,
but not before then, another test, similar design,
fewer markers, if the plate is queued,
children learn that very early.
If the plate is not cued and you have
to use allocentric cues, it takes again 25, 43 months.
So we think that in children, the hippocampus kicks in
around 43 months and allows allocentric learning to occur.
So why is it this change that's occurring
in the hippocampus as for pretty much non-functioning
in young children, but develops in older children?
The brain is just an absolutely wonderful,
and we're gonna look at, during pregnancy,
you go from just a small number of cells
to a well-functioning developing hemisphere.
And a lot of that development is based on genes
as well as to a certain degree,
environment, that's okay, thanks.
Now as the child gets older, we have
what's called activity dependent plasticity.
So as you learn things, you increase excitation cells learn
to develop mature, connect with one another,
and learning drives that connectivity with maturation.
So there's environmental factors
that take place after birth.
So again, I don't want you to need
and learn a lot of anatomy, but I just wanna talk
a little bit about the hippocampus
'cause it's so important and memory and learning,
and keep in mind, there's a lot of information
that flows into the hippocampus, primarily from what
is called the entorhinal cortex of the brain.
And this flows into the hippocampus,
a number of different neuronal ensembles here,
code this information and then send this information out
to widespread areas of the brain.
So very, very important in learning and memory.
If you don't have a hippocampus,
if you remove both hippocampi, which was done surgically
on a patient named H.M. you have
devastating problems with memory.
So our lab has been very interested in this development
of memory and we use single cell recordings.
And what'd you do here is you take a rat
and you put in 64, 128 electrodes,
and you put them into the hippocampus
and then you can record single cells.
And when I say it records single cell,
people call this units and these are actually
action potentials that you're recording.
And the beauty of this system is you can rack
and run around and learn the tasks
while you're recording on a single cell level.
So we put a light on top of the rat's head here,
put them in a chamber and then through some nice techniques,
you can actually look at action potentials and wreak
and figure out how many cells
are firing these action potentials.
So we can identify using electrophysiological techniques,
individual cells that are causing those action potentials.
So these are pretty incredible cells because an investigator
named O'Keeffe put an animal once in a chamber.
The chamber had a cue card here.
So the animal could orient where he was in space.
And Keith was listening to these action potentials.
You can just turn up your amplifier
and you can actually listen to action potentials
and found that when the animal is running around,
there would be certain cells that fired in certain
locations and this is a cell that fires around five o'clock.
And these are the action potentials from the tetrodse.
Here's a cell that fires around noon
and you can see the action potentials.
And here's a cell that fires everywhere
and this would be an inter-neuron, a fast spiking cell,
which is a GABAergic or inhibitory cell.
So here we have a situation, a place cells in which
you have a single cell can help predict
where the animal is in space.
You take the animal out and bring them back the next day,
the animal goes to this area, the cell fires again.
So these are place cells an incredible discovery by O'Keefe
that led to him getting the Nobel Prize recently.
Now place cells are very interesting
because here's an animal running around.
And as he goes through these different areas,
certain place cells and colors here, fire.
Now and here's an example, when the animal
is what running on a linear track, you see
all these action potentials and they're lined up
in the order they fired and so the action potentials
are occurring as a rat goes into different locations.
Now, when rats stop and rest,
you see what's called a sharp wave-ripple.
And this is just a slow way with a lot of fast activity.
And what happens during a sharp wave-ripple,
the cells that fire while this animal is running fire again,
but they fire in rapid succession.
So the animal who's cells are firing is as animal's running
across different cells, different colors,
animal stops and they fire again spontaneously,
even though the animal's not moving.
Same thing with here, if the animal is awake,
when the animal goes to sleep, you get this replay of cells.
So this replay of cells is felt to represent or be
a physiological mechanism where one can consolidate memory.
So you're replaying this information from what you just did.
So and then there's another cell you need to know about,
in the entorhinal cortex, remember I told you
the entorhinal cortex flows into the hippocampus.
Now this is a single cell, but this cell fires
in a grid-like pattern and so they're called grid cells,
which then help the animal location in space,
and then feed into the hippocampus<
these are entorhinal cortex in the hippocampus
and you get these place cells on here.
So place is down here, grid cell here,
entorhinal cortex, hippocampus here.
So these cells are really pretty cool to look at.
And the thing is that these cells
are not firing just randomly.
The cells are firing based on brain oscillations.
And so we talk a lot about theta in rats,
and we have theta in humans, this is the frequency.
When rats running around, they have
this very nice theta activity.
When they rest, they have these sharp waves
and the EEG slows down while
they're trying to consolidate the memory.
So we can look at the cells firing and not only
will they fire and low space, but also how they fire
in relationship to the theta activity.
So this is called temporal coding,
the timing of the cell in relationship to the theta.
And so that's interesting, but who really caters?
Well, it's very important because the cells fire
on the theta dependent upon what the animal is doing.
For example, here's an animal that's just running
around chasing food and here's an animal
that's in a test called the active avoidance,
where the animal has to remember where the shock zone is.
He can't see it, but based on cues around the room,
the animal has to learn to avoid the shock zone
and the cells fire at different places there,
you can plot these out, you know, when you think thetas
it's 360 degrees and you can figure out
where the cells fire in relationship to the theta.
And when cells fire synchronously or fire on the same stage
if they are in different cells that can really affect how
much neurotransmitter is released and so it can be
a pretty powerful way to send messages from
one area of the brain to the other.
So what you can do when you're bored at night
is go out and buy yourself two metronomes
and then you take the metronomes and you put it on a board
and then you put some beer cans under the board.
If you're a faculty member at UVM
you would use Heady Topper, if you're an undergrad
of drinking age, you would probably buy Pabst Blue Ribbon.
But anyhow, you put these beer cans under here
and you set the metronomes off,
so they're not firing together.
What'll happen over time is that metronomes will,
through different torque pressures,
will start to become synchronous.
And as they become synchronous,
this board will move back and forth.
So there's a lot of power in two areas
that are in sync, can cause significant movement here
to neurons that are in sync, it can cause
increased neurotransmitter release
when they're in the same phase.
So this is a powerful mechanism
by which neural transmission occurs in the brain.
So we're back to development. And the rat,
this allocentric memory that occurs will depend
upon a lot of things coming in line, your rhythms,
theta rhythm, a faster rhythm, gamma rhythm,
which really drives input into the dendrites of cells,
your grid cells, boundary cell,
which I didn't talk about are cells that fire
at a boundary when the animal's in a cylinder or a square.
And then you have the sharp wave-ripples,
which is important for consolidation.
So, you know, I just think of these things
all coming together, being all very important.
And the timing of when they develop is important.
When I refer to P here, I'm talking about rats
that are postnatal day seven, postnatal day 21,
which is a current equivalent to P10 rat
is equivalent to about a term infant.
So think of this as a orchestra where everything
has to come together, timing has to be perfect.
And you can think of the percussion system here.
That percussion section is driving the theta activity.
The basses you can think of boundary cells here.
You can think of the string section,
and the violas, the violins, cellos,
perhaps representing grid cells and place cells.
And then you can think of the flute section,
the woodwind section, the brass section
as being the different types of rhythms
that have to come together.
So everything has to work beautifully in sync.
Timing is great and you get a beautiful music
from the orchestra and it's the same thing with the brain.
If everything is in sync, you have a child
or you have an animal that can learn and learn quickly.
Okay, so and again these are important developmental steps.
They occur in waves and the timing is absolutely critical
of when these different wave form
I talked about join up together and this maturation
of these different cells goes from an leads to,
in a child from egocentric memory to allocentric memory.
And we said earlier that allocentric memory
is around four or five years.
So we were talking about the hippocampus.
So this is the period where you really start
to see allocentric memory come into play.
So what about rats?
Well, it turns out that the equivalent to declarative memory
in children is the spatial cognition in rats.
And there's a number of ways you can look
at spatial cognition in rats and one
is the famous Morris water maze,
where you drop a rat into the water.
There's a platform here that's under the water.
The water is opaque, you put milk or latex paint
in the water, they can't see the platform.
Rats are great swimmers and they don't really like it.
So if they accidentally will find the platform.
If you keep doing the same test, as long as you have cues
around the room, allocentric cues,
the rat will quickly learn to swim right to the platform.
And that's called the Morris water maze.
One of the most famous and well-used tests
in spatial cognition in rodents
and the sight declarative memory.
So if you take rats and you give them a bunch
of seizures early in life in the newborn period,
and then you look at them versus controls,
we're talking about early life seizures here,
you can see and this is the number of trials here.
And we do the testing over days, four days typically.
And you can see that in early life seizures,
the animals really never learn the task.
And the control animals over time,
quickly learned the task here.
So early life seizures cause cognitive impairment.
And if you do the active avoidance test here,
so the rat can't see this, but this is avoidance,
he gets a shock if the rat enters here.
So if you take a control rat, the rat will quickly learn
using this dwell map here to stay away,
as far away from that shock zone as possible.
And this arena is rotating, so the animal
as he's setting in here is he knows,
he's getting close to that shock zone,
will turn around and run back.
So it's a rotating arena with a shock zone
and he has to use clues around the room
to know where that shock zone is.
Here's a rat that had early life seizures.
You can see just by the path the rat is taking,
he runs through that shock zone a lot.
Control animal stays away from it.
So this is another good test of spatial avoidance
and rats that have early life seizures
just have a great deal of difficulty learning
what that shock zone is.
So that's interesting, but what's going on physiologically?
I told you the importance of place cells,
theta coming in line, all these different
physiological parameters, well, if you take a rat
and you give them a bunch of seizures
and just very brief seizures actually,
and then you wait a while and then you test them
in the just the kind of single cell recording
just looking at place cells, you can see
that the figures on top here,
these place cells are very precise.
The dark colors are the higher firing rates,
and you can see that they're really precise
and the firing occurs in a very limited fashion.
If you look at rats that have had recurrent seizures,
the place maps are all over the place, very disorganized,
very poorly modulated, not very precise.
And they may actually even change position
from one section to the next, so as are looking
at four sections and that corresponds beautifully
with how they do in the Morris water maze.
So animals that have aberrant place cells
do very poorly in the Morris water maze.
So it's a good indication on a single cell level
of cognition and this is impaired
with rats with developmental seizures.
Now a very important thing is that a lot of people think
and this is true in adults, but not children.
And certainly adult rats and not pediatrics, not baby rats,
that you have a seizure, the rat convulses, they get blue,
they get cyanotic and you think, well,
maybe this is just due to cell loss, the cells die.
And that's why there's cognitive impairment.
And that's not the case and animal literature
suggested years ago that rats, baby rats,
will have frequent seizures and you cut the brains
and you look at them and you see no cell loss.
And so the implication was that seizures are harmless
in the developing animal because you don't see cell loss.
And it just shows the important cell loss is not everything.
And so animals can have significant deficits
without actually cells dying and what we
and some of our colleagues have shown is that you can get
with early life seizures, you can see, not cell loss,
but you can see changes in these neurons and decreases
in the synapses, the spines, the dendritic arborization here
and if you do functional MRI on these rats
with early life seizures, you can actually see changes
in how the current flow in this region occurs.
So here's a example of a diffusion tensor imaging,
looking at CA1 of the hippocampus,
this is part of the hippocampus.
This is a control rat, this is a rat
that's had a long seizure and you can see
that there's difference in the track profile
in these animals and how current flow
through these animals is different in these animals.
So these animals have impairment and we think a lot
of this impairment has to do with
that dendritic arborization that occurs.
And if you then look at oscillations in the hippocampus,
we see in the gamma region abnormalities in CA1
that correspond to the MRI findings as well.
So seizures are going something structurally to the brain,
but they're not killing brain cells.
So we've had recently and this is just from this year,
I've been very interested in what it is about seizures
and the developing animal that leads
to so much cognitive impairment and which leads
to aberrant place cells leads to spatial cognitive function.
And what is it about the seizure per se, that occurs?
What I told you earlier that there's this,
as you develop allocentric memory
in rodents and presumably humans,
these different processes come in line
and they're driven by brain oscillations.
So we've said, why not look at animals
during the critical period of spatial cognition
and this would be corresponding to a child as I mentioned,
that's when they develop allocentric memory,
we use the same age in rats, give them seizures
during the critical period and then when you follow
these animals, they are impaired cognitively.
But we said, what if we don't give them seizures?
What if we just changed their oscillatory pattern?
So here's a normal rat, very young rat,
and they have this beautiful theta rhythm.
We decided to take rats and we would use
this a technique called optogenetic-induction
in which you can stick what's called
a channelrhodopsin in the medial septum
and we can then control oscillations in the hippocampus.
And so what we did is not cause seizures, but we just said,
let's just throw in junk oscillations, nothing pathological,
normal oscillations, but not seizures.
So the process here is that you take a rat,
you put a electrode and you put a virus
which contains channelrhodopsin,
which will be taken up by the cells and in these cells,
when you shine a bright light,
a blue light, that cells will fire,
when you shine a yellow light, the cells will not fire.
So the medial septum is in the middle of the brain
and it projects to both of the hippocampi.
So it's a midline structure, goes to both hippocampi.
So we put electrodes in the medial septum,
we put electrodes in the hippocampus
and then we checked to make sure
that we were actually inducing changes
in the hippocampus with medial septum stimulation.
And so here, we had stimulated five, six, seven,
eight, nine, 10, 11, 12 hertz.
You can see in this spectrogram,
which shows the power of the wave form.
You can see very nice as you go up,
we're getting what we said we would get
and you get beautiful, what we would call driving
of the hippocampus so we can control
the hippocampal oscillations
through medial stapal stimulation.
Then we say, well let's take some rats in which
we can do this and let's just give them nonsense.
Let's just give them oscillations varying
from a 100 hertz to one hertz and you can see here,
this is over on at least an hour,
you can see different oscillatory patterns in the rat.
And then we test them in the active avoidance test
and here, so we're not giving seizures,
we're giving normal oscillations,
but in an aberrant temporal fashion.
And just to show you how beautiful this works,
the people in my lab are just fantastic
at getting quality, here's baseline, rat has temporal.
You start to stimulate then at six hertz,
this is just showing power here.
And this is an amplitude synchrony,
basically amplitude correlations,
but you can see it is six hertz,
You get exactly what you want, six hertz.
There's some normal rhythms in there as well.
12 hertz, you get beautiful 12 hertz,
and it's reflected here and in the spectrogram,
which shows the frequency, this is a crane rate
at six hertz and this is re occurring at 12 hertz.
What's interesting you can see
what's called harmonics as well.
So if you stimulate it six, you can see
also some activity at 12 and so on.
So it works, it works very, very well.
And lo and behold, we did this, these rats look fine.
You know, we did stimulate them for four days during
the critical period, then we put them in active avoidance.
And whether we gave them a one hour of stimulation
a day or five hours a day, these animals
had pretty significant impairment.
So these are number of shocks
and these are the trial numbers here.
And even though our ends were pretty small,
our statistics were so strong that we went ahead
and got this published and showing
that hippocampus oscillations during
the critical period of brain development results
in substantial deficits in spatial cognition.
And just to show you an example of this,
this is again, a dwell map, active avoidance here.
Here's the shock zone which the animal can't see,
these are animals that had no light stimulation,
or these are animals that had yellow light,
that will not activate the channelrhodopsin nothing happens.
But these are animals that had blue light and you can see
like this animal just ran through
the shock zone, pay no attention to it.
This animal had a different kind of strategy.
Just kind of did his own circle inside
and he was getting shocked a little less,
but was quite impaired in this testing.
So the question was that, although we thought
we were putting in normal oscillatory activity,
but just kind of random noise into the system,
could this cause any damage, could this be,
you know, could this be frying the neurons, et cetera?
But so we did some pretty nice histology here.
What I'm showing you here is the channelrhodopsin expressed
in the medial septum here and the medial septum, again,
projects to the hippocampus and you can see the axons
and the terminals of the medial septum neurons here.
This is CA1 of the hippocampus,
here this is a stain just to show cells.
Here's the dentate gyrus here, but these fibers
weren't right where we expected them to.
And this is a new N, Michelle Klok in the lab,
did a new N which shows neurons here.
And when you do extensive cell counting on these animals,
really, there's no cell loss.
So and this optogenetic stimulation just by kind
of putting a normal activity in at the wrong time,
it really can disrupt the development of spatial cognition
and lead to long-standing deficits in spatial cognition.
So we think that the body of the work recently
what we're doing is that it's not necessarily
the seizures that are causing the problems in these kids,
it's the abnormal rhythms that are going into the brain.
And as I mentioned earlier, the children
with the epileptic encephalopathies have terrible EEGs.
So 24 hours a day, seven days a week,
they're having aberrant activity.
They may not be having seizures,
but the brain is not, the temporal coding,
the information going to the hippocampus based
on the oscillatory activity is just not right.
So there's more and more evidence that the children
that have cognitive impairment with early life seizures,
it may be more related to the dysrhythmia of the brain,
the alterations and the oscillations,
more so than the actual seizures per se.
So the EEG may be more predictive of outcome
in these kids with the epileptic encephalopathies,
than the actual seizures themselves.
So again, the brain is a magical organ.
It's the timing is everything and I think that the brain
is a, you know, musical orchestra, where everything
has to come together and a perfectly timed pattern
for the children to develop a normal spatial cognition,
and which would be declarative memory in children.
So, in conclusion is I just wanna note,
and I hope with the one take-home message for you
is that epilepsy is far more than seizures.
Our children have cognitive impairment to a high degree,
the ones with severe seizures and early life seizures.
They're more prone for sleep disorders, depression,
attention deficit disorder, et cetera.
So it really is a spectrum disorder.
And we, as clinicians need to spend more time trying
to work on the cognitive and on the comorbidities
than we may need to do on the seizures.
Many parents have told me that the seizures
are just a small part of the problem with their child.
They have so many other issues they can handle the seizures,
but they have trouble handling the other things that occur.
And it's heartbreaking such as with Susan Axelrod's daughter
to see her decline so rapidly over time.
Children are particularly vulnerable
to the adverse effects of seizures, so timing is everything.
And so at a time when the brain is very plastic
and you're trying to develop the hippocampus
so that you can develop a declarative memory,
during that time you're being bombarded
with seizures or aberrant oscillations,
that can be really quite vulnerable.
And there's a critical period of time,
in work that we haven't done, but other people have done
is that if you can shut off activity
or cause abberent activity during
the critical period of time of spatial development,
and that in children would be during
the first four years of life and you have the insight
and do the same thing in an older child,
much less likely to have that cognitive impairment.
I told you earlier that children that are older
and start having seizures are much less likely
to be impaired than children that
are very young when they start having seizures.
So we really have to work hard on stopping
the seizures in the young children.
And I think the seizures may adversely effect the rhythms
of the brain and therefore result
in longstanding cognitive impairment.
So with that, I would like to thank people
for listening to this in the late afternoon
and I have so many people to thank.
But I'm just really concentrating this list on the people
I've been working with over the last couple
of years here at UVM in my lab here,
we work very close with Tallie Baram at UC Irvine,
I did a sabbatical in Paris and still maintain contact
and rich collaborations with the people there.
And then we've had post-docs, I had a post-doc
from the Ukraine who I'm still working with
and we just published a paper recently.
But all these people and as I mentioned before,
in the ceremony that, you know, I'm the type of person
that goes around and giving the talks,
getting the wards like this one, but these are the people
that do all the heavy lifting and are in the lab every day,
are working very hard and so I really have
a very good group, so with that, I thank you very much
and be glad to take some questions.
- Thank you, Dr. Holmes.
I wanna open this up right now for questions,
but I wanna first just say thank you very much.
And if you didn't have a room with only
the five people helping you do this,
you would hear us all applauding for you.
So I will take a second for you to appreciate that.
The questions will come from me,
they're being published from the audience.
So I have a beginning question for you,
comes from Jerry Herrera and it says,
very fascinating talk, I really enjoyed it.
Do kids with seizures, suffering cognitive impairment,
have any issues with control of bodily functions
like urination, voiding and bowel control?
- Yes, so that's often one of the major problems
with the children and those are the kids
that are usually pretty impaired.
So and yeah, the answer is yes.
And that's one of the things I always ask my parents
just talked to a patient today, you know,
and going through the developmental milestones,
the child has severe epilepsy and, you know,
you ask the parents about toilet skills and that
is so critical, so hard to take care
of some of these kids are so impaired,
but the toilet skills is the big thing.
It keeps them out of school, it's just a terrible thing.
And yes, it's a major problem with that.
There's also the issue of, you know, during seizures,
you can have, certainly one of the things
I always ask patients, did they urinate during the seizure
and you can even defecate during a seizure.
So it can be a problem, but usually
in the more impaired kids do we see it.
We also see there's a pretty high association
with epilepsy and autism and we're not saying
epilepsy causes autism, but they're comorbidities
and the children with autism tend
to have these toilet skills as well.
But that may be a different mechanism than some
of the kids with more cognitive complaints, yes.
- Great, thank you, there's another question coming through.
I don't have it yet, hold on, here it is,
from Cory Trcicia, are the phenotypes
that you observed sexually dimorphic,
is the epilepsy itself sexually dimorphic
either in incidents or severity?
- Really, really good question, that's a great question.
One of the things we're just doing in the lab,
and we just finished two with Rhys Niedecker is he did
a nice study and we're getting ready to publish it.
He's a medical student now, he spent a couple of years
in the lab, he's a first year medical student
and we deducted a study in which, well,
first of all at the NIH, we used to always be,
chauvinistically, we always used to use male rats,
'cause we said less variability and blah, blah, blah.
And that was really the wrong thing.
NIH said you now have to use sex as a biological variable.
And they were absolutely right.
So Rhys did a study in which we looked
at male rats and female rats pups that had seizures.
And it turned out that the female pups did much better
in outcome than the male pups that had seizures.
So I'm looking at this now clinically
because that's not been really reported
whether the prepubescent children,
whether you're male or female end up having
more severe seizures and worse outcome.
Our lab data would indicate and Rhys did
a great job in controlling for, you know,
the pain threshold and the active avoidance,
all the right things, looked at estrous cycles
in the adult animals and really didn't see much there.
But so, yeah, so even in prepubescent rats,
we were finding that there's a big difference
between male and females and it is much better
to be a female rat as far as outcome.
- Thank you, a follow-up on the voiding question.
Do you see this defective voiding in the rats
where you introduce optogenetic dysrhythmia?
- We do not, I shouldn't say that we didn't
really look very hard, it's not something
we observed spontaneously, but we did not see that.
But I can't say for sure, we really weren't monitoring that.
- Okay, I don't have another one at the moment,
but I do have a question to ask you myself.
So when you're looking at this disruption
of the rhythm by all this abnormal activity,
do you think it's the abnormal activity
and the abnormal rhythm or the loss
of the normal rhythm that's actually involved
in the disruption of the memory formation?
- Very good question and I don't know.
I think one of the things one could do
is just simply turn off the medial stuff.
It's been done before, actually in the mammillary bodies,
which flows into the medial septum.
And if you do that acutely in adult rats
and you test them in the water maze,
they cannot learn it when the theta has been stopped.
What hasn't been addressed yet is whether
if you knock off either developmentally,
would you have cognitive impairment later on?
And the answer is probably yes,
there's been one genetic study where they knocked
out a gene that was important
in synaptic scaffolding and connectivity.
And I forget the name of the gene and when they knocked
that off in the young rats and they actually
had decreased theta, those animals
did have impairment cognitively later on.
If they did it in older rats and knocked it out,
conditional knockout, it did not have any effects.
So it was during the critical period
that seemed to have an effect, so I would think that
it may be a combination of both, good point.
- Great, thank you, I have another follow-up on,
now we're gonna bounce back and forth,
the problem with chats is the follow-ups are delayed.
So the follow-up on the sexual dimorphism
is epilepsy in humans sexually dimorphic
with respect to incidents or severity?
- Yeah and because of what we found,
I have gone and started to look at that, I have it.
It's not clear from the clinical literature
whether that's true or not but it hasn't been really studied
that well and I think these large studies,
I haven't seen any significant
difference in cognitive outcome.
You know, one of the problems you get into
with clinical, when you try to go to the clinical
from the lab is that you get so much variability
in the clinical arena, different severities of seizures,
different age of onset, different medications
that have been given, different seizure types.
So it's often hard to dissect that out.
It's much easier to do it in an animal
where you have control of everything,
but it's something I'm looking at right now.
And again, based on Rhys' data.
- Great, thank you, another question from Karen Lounsbury,
somewhat similar to mine, is there a toxic effect of
the seizures themselves that leads to cognitive impairment
or is it the lack of normal neuronal activity?
Can you simulate the deficit with other toxins?
- Yeah, so, I mean, one of the things people have done
years ago is one condition that both in children and adults,
more so in adults, is called status epilepticus.
So if you cause a patient have a prolonged seizure,
and when I say prolonged, we're usually talking about
an hour or more of constant seizure activity,
those patients have pretty significant cell loss
and have cognitive impairment.
So people used toxins in the past, so one that we used
and other people used is pilocarpine kainic acid.
And so the question came up and that causes stutters.
So if you stick pilocarpine or kainic acid
in the hippocampus, the animals will have
a very severe seizure and they will have cell loss.
And the question was, well, is it due to the toxin
or is it due to the seizure?
So the professor I did my sabbatical with,
is Professor Ben-Ari in Paris answered that question.
So he gave kainic acid into the hippocampus
and he followed that with an injection of diazepam
or Valium which is a good drug to stop seizures.
What he saw was virtually no damage,
so it was a pilocarpine causing the seizure damage.
As far as how that occurs, most people think,
the most common theory is it's excitable toxicity.
So when you have this massive seizure,
you're releasing all this glutamate,
glutamate can be very toxic to cells
and can lead to cell death.
So that's the thinking there, but, you know,
you're not really asking this question,
but one point I actually really think should be made
is that and I didn't emphasize it enough,
is that whatever causes the seizures,
it has to be taken into consideration
of any cognitive impairment, so, you know,
children that have hypoxic-ischemic encephalopathy
and have birth asphyxia and they have seizures,
the seizures may contribute to cognitive impairment,
but those kids, the issue is the cause of the seizures,
other being the hypoxic-ischemic injury.
So etiology is clearly important
when you're seeing patients trying
to determine what's causing the epilepsy.
We'll give you a good idea of what the outcomes
are gonna be, but seizures can add
to that impairment or, you know, cause it
in people with like idiopathic epilepsy, but etiology,
the cause of the seizures is also very important.
- Great, thank you, I think we don't have another question
at the moment, I'm just gonna wait a second or two,
but I think that this is absolutely perfect timing
on a talk and question, so I thank you for that.
And hopefully you've enjoyed your time doing this
in this very strange presentation
to the crew, thank you very much.
- Okay, thank you very much.