so first of all it’s a great pleasure to be participating in this marvelous
be participating in this marvelous
be participating in this marvelous program what I’ll do it is to initially
program what I’ll do it is to initially
program what I’ll do it is to initially give you a short introduction as to what
give you a short introduction as to what
give you a short introduction as to what we do and then give you an example of
we do and then give you an example of
we do and then give you an example of current work using that technology so in
current work using that technology so in
current work using that technology so in every one of our cells of our body
every one of our cells of our body
every one of our cells of our body there’s an extremely complex instruction
there’s an extremely complex instruction
there’s an extremely complex instruction manual how complex is it it’s written
manual how complex is it it’s written
manual how complex is it it’s written the text is written in an alphabet of
the text is written in an alphabet of
the text is written in an alphabet of four letters ACTG
four letters ACTG
four letters ACTG rather than 26 letters and the entire
rather than 26 letters and the entire
rather than 26 letters and the entire sequence of the human genome as well as
sequence of the human genome as well as
sequence of the human genome as well as a mouse genome has now been determined
a mouse genome has now been determined
a mouse genome has now been determined it contains approximately three billion
it contains approximately three billion
it contains approximately three billion letters if we wrote out this text it
letters if we wrote out this text it
letters if we wrote out this text it would occupy a thousand volumes each a
would occupy a thousand volumes each a
would occupy a thousand volumes each a thousand pages long so it is an enormous
thousand pages long so it is an enormous
thousand pages long so it is an enormous text and knowing the sequence doesn’t
text and knowing the sequence doesn’t
text and knowing the sequence doesn’t tell us how it works
tell us how it works
tell us how it works it’s as if you had a new language and no
it’s as if you had a new language and no
it’s as if you had a new language and no rosetta stone to be able to interpret
rosetta stone to be able to interpret
rosetta stone to be able to interpret that language and so our test was to try
that language and so our test was to try
that language and so our test was to try to essentially functionally decipher
to essentially functionally decipher
to essentially functionally decipher this text and the approach was very
this text and the approach was very
this text and the approach was very simple what we wanted to do was to erase
simple what we wanted to do was to erase
simple what we wanted to do was to erase for example a small part of the text in
for example a small part of the text in
for example a small part of the text in every cell of your body and then ask
every cell of your body and then ask
every cell of your body and then ask what the consequences are
what the consequences are
what the consequences are that is if little finger disappeared
that is if little finger disappeared
that is if little finger disappeared then we know we were in the program for
then we know we were in the program for
then we know we were in the program for making a little finger so and thereby
making a little finger so and thereby
making a little finger so and thereby infer by the changes what the normal
infer by the changes what the normal
infer by the changes what the normal function of that part of the text is so
function of that part of the text is so
function of that part of the text is so that was the approach and in 1980 our
that was the approach and in 1980 our
that was the approach and in 1980 our submitted a grant to actually support
submitted a grant to actually support
submitted a grant to actually support this work and the response was very
this work and the response was very
this work and the response was very direct the project is impossible it’s
direct the project is impossible it’s
direct the project is impossible it’s not worth funding but I didn’t take no
not worth funding but I didn’t take no
not worth funding but I didn’t take no for an answer
for an answer
for an answer what we did was to actually take all the
what we did was to actually take all the
what we did was to actually take all the money from another project that we did
money from another project that we did
money from another project that we did have funded and sunk it into this
have funded and sunk it into this
have funded and sunk it into this project but unfortunately we had
project but unfortunately we had
project but unfortunately we had actually four years of funding so we had
actually four years of funding so we had
actually four years of funding so we had four years to get results and if we
four years to get results and if we
four years to get results and if we failed then my scientific career would
failed then my scientific career would
failed then my scientific career would be over but I think the one message of
be over but I think the one message of
be over but I think the one message of what I get across is that sometimes you
what I get across is that sometimes you
what I get across is that sometimes you have to take a risk you have to be
have to take a risk you have to be
have to take a risk you have to be trusting your gut and that’s actually my
trusting your gut and that’s actually my
trusting your gut and that’s actually my major message so we chose the mouse and
major message so we chose the mouse and
major message so we chose the mouse and I’ll come back as to why we chose the
I’ll come back as to why we chose the
I’ll come back as to why we chose the mouse as our organism or to investigate
mouse as our organism or to investigate
mouse as our organism or to investigate so let’s begin with the mouse and the
so let’s begin with the mouse and the
so let’s begin with the mouse and the first question you should ask yourself
first question you should ask yourself
first question you should ask yourself is why the mouse the mouse is a mammal
is why the mouse the mouse is a mammal
is why the mouse the mouse is a mammal and with respect to genes all mammals
and with respect to genes all mammals
and with respect to genes all mammals share many many genes for example humans
share many many genes for example humans
share many many genes for example humans and mice in terms of gene content are
and mice in terms of gene content are
and mice in terms of gene content are more than 99% the same so whatever we
more than 99% the same so whatever we
more than 99% the same so whatever we learn in the mouse is likely to be
learn in the mouse is likely to be
learn in the mouse is likely to be applicable to humans we have a because
applicable to humans we have a because
applicable to humans we have a because it’s a mammal we have a common
it’s a mammal we have a common
it’s a mammal we have a common developmental program physiology nervous
developmental program physiology nervous
developmental program physiology nervous system and so on and so forth now the
system and so on and so forth now the
system and so on and so forth now the starting point of our experiments are
starting point of our experiments are
starting point of our experiments are these cells these are mouse embryonic
these cells these are mouse embryonic
these cells these are mouse embryonic stem cells and they’re growing on a
stem cells and they’re growing on a
stem cells and they’re growing on a feeder layer which is supplying factors
feeder layer which is supplying factors
feeder layer which is supplying factors to keep these cells happy these cells
to keep these cells happy these cells
to keep these cells happy these cells are pluripotent that has given the right
are pluripotent that has given the right
are pluripotent that has given the right signals they can become anything skin
signals they can become anything skin
signals they can become anything skin cells muscle cells nerve neurons any
cells muscle cells nerve neurons any
cells muscle cells nerve neurons any cell of your body they’re capable of
cell of your body they’re capable of
cell of your body they’re capable of becoming they simply have to be told
becoming they simply have to be told
becoming they simply have to be told what to be and if we introduce these
what to be and if we introduce these
what to be and if we introduce these cells back into an early embryo and they
cells back into an early embryo and they
cells back into an early embryo and they participate in making all of the tissues
participate in making all of the tissues
participate in making all of the tissues of the mouse but most importantly also
of the mouse but most importantly also
of the mouse but most importantly also the germ cells sperm and eggs or eggs so
the germ cells sperm and eggs or eggs so
the germ cells sperm and eggs or eggs so that’s our starting point and these are
that’s our starting point and these are
that’s our starting point and these are the cells that we genetically manipulate
the cells that we genetically manipulate
the cells that we genetically manipulate by technology we call now gene targeting
by technology we call now gene targeting
by technology we call now gene targeting and this oblongs gene targeting so the
and this oblongs gene targeting so the
and this oblongs gene targeting so the top bar
top bar
top bar represents your favorite gene and the
represents your favorite gene and the
represents your favorite gene and the asterisks represents a modification in
asterisks represents a modification in
asterisks represents a modification in that sequence and that modification can
that sequence and that modification can
that sequence and that modification can be anything 10 letters
be anything 10 letters
be anything 10 letters 100 letters 10,000 letters a gene is
100 letters 10,000 letters a gene is
100 letters 10,000 letters a gene is roughly on average about 40,000 letters
roughly on average about 40,000 letters
roughly on average about 40,000 letters so we make the change in a test tube
so we make the change in a test tube
so we make the change in a test tube then we introduce this bar this gene
then we introduce this bar this gene
then we introduce this bar this gene back into the ESL nucleus and there it
back into the ESL nucleus and there it
back into the ESL nucleus and there it combines with a machine and then
combines with a machine and then
combines with a machine and then searches all three billion base pairs
searches all three billion base pairs
searches all three billion base pairs all three billion letters until it finds
all three billion letters until it finds
all three billion letters until it finds the exact same sequence and then matches
the exact same sequence and then matches
the exact same sequence and then matches up with it and then by a process called
up with it and then by a process called
up with it and then by a process called homologous recombination exchanges
homologous recombination exchanges
homologous recombination exchanges information with it so now in the living
information with it so now in the living
information with it so now in the living cell we have now changed that change
cell we have now changed that change
cell we have now changed that change that we created in the test tube is now
that we created in the test tube is now
that we created in the test tube is now in the genome of the living cell okay so
in the genome of the living cell okay so
in the genome of the living cell okay so then we can grow up those cells and
then we can grow up those cells and
then we can grow up those cells and reintroduce them into a pre-emptive Airy
reintroduce them into a pre-emptive Airy
reintroduce them into a pre-emptive Airy early embryo a pre implementation effort
early embryo a pre implementation effort
early embryo a pre implementation effort this is called a blastocyst and the
this is called a blastocyst and the
this is called a blastocyst and the cells down here are the cells that are
cells down here are the cells that are
cells down here are the cells that are going to make the embryo proper and here
going to make the embryo proper and here
going to make the embryo proper and here are the ES cells that we modified with a
are the ES cells that we modified with a
are the ES cells that we modified with a needle going in here and then they mix
needle going in here and then they mix
needle going in here and then they mix with these cells and then they’re
with these cells and then they’re
with these cells and then they’re getting all the signals to say what to
getting all the signals to say what to
getting all the signals to say what to become and then they make the
become and then they make the
become and then they make the appropriate tissue then we take that
appropriate tissue then we take that
appropriate tissue then we take that embryo implanted into a pseudopregnant
embryo implanted into a pseudopregnant
embryo implanted into a pseudopregnant Mouse they put a little slit in the back
Mouse they put a little slit in the back
Mouse they put a little slit in the back get to that allows an entry to the
get to that allows an entry to the
get to that allows an entry to the uterus put it right in there it attaches
uterus put it right in there it attaches
uterus put it right in there it attaches and then the Emperor starts to develop
and then the Emperor starts to develop
and then the Emperor starts to develop is born and now you have a mouse but
is born and now you have a mouse but
is born and now you have a mouse but we’ve changed its genetic composition so
we’ve changed its genetic composition so
we’ve changed its genetic composition so let me give you an example there’s a
let me give you an example there’s a
let me give you an example there’s a gene that we can isolate from bacteria
gene that we can isolate from bacteria
gene that we can isolate from bacteria that if we put their appropriate
that if we put their appropriate
that if we put their appropriate substrate turns the cells blue there’s
substrate turns the cells blue there’s
substrate turns the cells blue there’s another gene that we can isolate from a
another gene that we can isolate from a
another gene that we can isolate from a jellyfish that makes a protein that’s
jellyfish that makes a protein that’s
jellyfish that makes a protein that’s fluorescently green we take that gene
fluorescently green we take that gene
fluorescently green we take that gene and put it into a middle of a mouse gene
and put it into a middle of a mouse gene
and put it into a middle of a mouse gene and in this case is Hoxie 13 then
and in this case is Hoxie 13 then
and in this case is Hoxie 13 then wherever hoxsey 13 is functioning the
wherever hoxsey 13 is functioning the
wherever hoxsey 13 is functioning the cells will either turn blue or fur SNP
cells will either turn blue or fur SNP
cells will either turn blue or fur SNP green so that tells us that
green so that tells us that
green so that tells us that this gene is important for making
this gene is important for making
this gene is important for making whiskers it’s not only important for
whiskers it’s not only important for
whiskers it’s not only important for making whiskers but also nails and not
making whiskers but also nails and not
making whiskers but also nails and not only but in the process of putting this
only but in the process of putting this
only but in the process of putting this gene in the middle we destroy that gene
gene in the middle we destroy that gene
gene in the middle we destroy that gene and so now we can look at the mouse and
and so now we can look at the mouse and
and so now we can look at the mouse and see whatever it does this gene do
see whatever it does this gene do
see whatever it does this gene do functionally for these mice and that’s
functionally for these mice and that’s
functionally for these mice and that’s shown in the next slide so fortunately
shown in the next slide so fortunately
shown in the next slide so fortunately we always have two genes for most you we
we always have two genes for most you we
we always have two genes for most you we have two copies for most genes one that
have two copies for most genes one that
have two copies for most genes one that you acquired from your father one from
you acquired from your father one from
you acquired from your father one from your mother okay plus plus means two
your mother okay plus plus means two
your mother okay plus plus means two good copies that’s a starting point plus
good copies that’s a starting point plus
good copies that’s a starting point plus minus means we destroyed one of the
minus means we destroyed one of the
minus means we destroyed one of the genes and then we can breed these mice
genes and then we can breed these mice
genes and then we can breed these mice together brothers and sisters and then
together brothers and sisters and then
together brothers and sisters and then generate a 1/4 of the progeny will be
generate a 1/4 of the progeny will be
generate a 1/4 of the progeny will be minus minus that will have no oxy 13
minus minus that will have no oxy 13
minus minus that will have no oxy 13 that’s functional and what you can see
that’s functional and what you can see
that’s functional and what you can see this gene is important for making all
this gene is important for making all
this gene is important for making all body hair not only whiskers but all body
body hair not only whiskers but all body
body hair not only whiskers but all body hair not only that but it’s also
hair not only that but it’s also
hair not only that but it’s also required to make nails so you have
required to make nails so you have
required to make nails so you have defective nails so this is a normal nail
defective nails so this is a normal nail
defective nails so this is a normal nail and here’s a very defective nail now we
and here’s a very defective nail now we
and here’s a very defective nail now we can go to the human population and ask
can go to the human population and ask
can go to the human population and ask our other people or have alopecia
our other people or have alopecia
our other people or have alopecia completely bald and have defective nails
completely bald and have defective nails
completely bald and have defective nails and or we take a little bit of their
and or we take a little bit of their
and or we take a little bit of their blood look at their DNA and they have
blood look at their DNA and they have
blood look at their DNA and they have two defective mutant copies of Hoxie xxx
two defective mutant copies of Hoxie xxx
two defective mutant copies of Hoxie xxx so now we’ve closed the circle whatever
so now we’ve closed the circle whatever
so now we’ve closed the circle whatever we found in Mouse is also true in humans
we found in Mouse is also true in humans
we found in Mouse is also true in humans and this has been done now for thousands
and this has been done now for thousands
and this has been done now for thousands and thousands of genes let me move on to
and thousands of genes let me move on to
and thousands of genes let me move on to the next story you’re remodeling a
the next story you’re remodeling a
the next story you’re remodeling a psychiatric disorder
psychiatric disorder
psychiatric disorder it’s OCD and specifically OCD spectrum
it’s OCD and specifically OCD spectrum
it’s OCD and specifically OCD spectrum disorder called trichotillomania which
disorder called trichotillomania which
disorder called trichotillomania which is compulsive removing of body hair and
is compulsive removing of body hair and
is compulsive removing of body hair and what you’ll see is when this mouse moves
what you’ll see is when this mouse moves
what you’ll see is when this mouse moves there it is it’s removed all its body
there it is it’s removed all its body
there it is it’s removed all its body here it actually continues to do that
here it actually continues to do that
here it actually continues to do that until it actually has lacerations at
until it actually has lacerations at
until it actually has lacerations at that point we put it to sleep this is
that point we put it to sleep this is
that point we put it to sleep this is very similar to OCD obsessive-compulsive
very similar to OCD obsessive-compulsive
very similar to OCD obsessive-compulsive disorder and trichotillomania which is a
disorder and trichotillomania which is a
disorder and trichotillomania which is a Specter or a subgroup of OCD
Specter or a subgroup of OCD
Specter or a subgroup of OCD so here we have a behavior and so we go
so here we have a behavior and so we go
so here we have a behavior and so we go into the brain and ask what cells in the
into the brain and ask what cells in the
into the brain and ask what cells in the brain which neurons are responsible for
brain which neurons are responsible for
brain which neurons are responsible for this behavior and what we can do is
this behavior and what we can do is
this behavior and what we can do is again put GFP in the middle of be run by
again put GFP in the middle of be run by
again put GFP in the middle of be run by hots be eight this is the gene the room
hots be eight this is the gene the room
hots be eight this is the gene the room that’s involved and then ask where the
that’s involved and then ask where the
that’s involved and then ask where the green cells and here we got an enormous
green cells and here we got an enormous
green cells and here we got an enormous surprise the only green cells in the
surprise the only green cells in the
surprise the only green cells in the brain in the brain are not neurons but
brain in the brain are not neurons but
brain in the brain are not neurons but microglia
microglia
microglia and this is shown here CTL MMB marks
and this is shown here CTL MMB marks
and this is shown here CTL MMB marks marks microglia it’s an antibody and
marks microglia it’s an antibody and
marks microglia it’s an antibody and what we can see is here’s the merger of
what we can see is here’s the merger of
what we can see is here’s the merger of those and so these are yellow cells
those and so these are yellow cells
those and so these are yellow cells which is from green to red the other
which is from green to red the other
which is from green to red the other thing we can see is that not all
thing we can see is that not all
thing we can see is that not all microglia are have Hawks b8 because
microglia are have Hawks b8 because
microglia are have Hawks b8 because there are many other microglia here so
there are many other microglia here so
there are many other microglia here so there has to be actually two populations
there has to be actually two populations
there has to be actually two populations of microglia ones that have my ox b8 and
of microglia ones that have my ox b8 and
of microglia ones that have my ox b8 and the other ones that do not non ox being
the other ones that do not non ox being
the other ones that do not non ox being okay so what our microglia microglia are
okay so what our microglia microglia are
okay so what our microglia microglia are immune cells they’re like blood cells
immune cells they’re like blood cells
immune cells they’re like blood cells and they’re also made to fight
and they’re also made to fight
and they’re also made to fight infections for example in the brain when
infections for example in the brain when
infections for example in the brain when they’re in the brain if you have a
they’re in the brain if you have a
they’re in the brain if you have a stroke or a bacterial infection or a
stroke or a bacterial infection or a
stroke or a bacterial infection or a viral infection some cells are killed
viral infection some cells are killed
viral infection some cells are killed and microglia go there and clean up the
and microglia go there and clean up the
and microglia go there and clean up the mess and allow the brain to function
mess and allow the brain to function
mess and allow the brain to function again so that’s the role of microglia
again so that’s the role of microglia
again so that’s the role of microglia but what I’m telling you is a defective
but what I’m telling you is a defective
but what I’m telling you is a defective microglia that is the one that doesn’t
microglia that is the one that doesn’t
microglia that is the one that doesn’t have a functional sucksby aging is
have a functional sucksby aging is
have a functional sucksby aging is controlling behavior and when that first
controlling behavior and when that first
controlling behavior and when that first came out people just you know didn’t
came out people just you know didn’t
came out people just you know didn’t believe it so this made an enormous
believe it so this made an enormous
believe it so this made an enormous prediction we can do blood of
prediction we can do blood of
prediction we can do blood of transplantations because it’s blood
transplantations because it’s blood
transplantations because it’s blood cells so what we can do is to radiate
cells so what we can do is to radiate
cells so what we can do is to radiate our mutant mouse okay and then put good
our mutant mouse okay and then put good
our mutant mouse okay and then put good bone marrow from a normal Mouse back
bone marrow from a normal Mouse back
bone marrow from a normal Mouse back into that Mouse and that ask does the
into that Mouse and that ask does the
into that Mouse and that ask does the bone marrow transplant cure the mouse of
bone marrow transplant cure the mouse of
bone marrow transplant cure the mouse of the disease and here’s the actual
the disease and here’s the actual
the disease and here’s the actual experiment so here is a mouse this is a
experiment so here is a mouse this is a
experiment so here is a mouse this is a mutant Mouse lethally irradiated so it
mutant Mouse lethally irradiated so it
mutant Mouse lethally irradiated so it would die if we don’t put
would die if we don’t put
would die if we don’t put into him we put normal bone marrow into
into him we put normal bone marrow into
into him we put normal bone marrow into that mouse here is three weeks after the
that mouse here is three weeks after the
that mouse here is three weeks after the bone marrow transplant it’s already
bone marrow transplant it’s already
bone marrow transplant it’s already removed all its body hair but showed you
removed all its body hair but showed you
removed all its body hair but showed you this slide a week later you’d see lots
this slide a week later you’d see lots
this slide a week later you’d see lots of lacerations and then 12 weeks after
of lacerations and then 12 weeks after
of lacerations and then 12 weeks after bone marrow transplant which is what
bone marrow transplant which is what
bone marrow transplant which is what also takes in humans all the lacerations
also takes in humans all the lacerations
also takes in humans all the lacerations are healed all the hairs grow back and
are healed all the hairs grow back and
are healed all the hairs grow back and also here relax you’re looking how much
also here relax you’re looking how much
also here relax you’re looking how much time they’re spending grooming use a
time they’re spending grooming use a
time they’re spending grooming use a normal Mouse here’s a mutant mouse and
normal Mouse here’s a mutant mouse and
normal Mouse here’s a mutant mouse and now it’s the grooming time has gone back
now it’s the grooming time has gone back
now it’s the grooming time has gone back down because the all of those
down because the all of those
down because the all of those lacerations as a function because
lacerations as a function because
lacerations as a function because they’re over Burma they’re continually
they’re over Burma they’re continually
they’re over Burma they’re continually grooming that same place so and this
grooming that same place so and this
grooming that same place so and this mouse be completely cured of that
mouse be completely cured of that
mouse be completely cured of that neuropsychiatric disorder by a
neuropsychiatric disorder by a
neuropsychiatric disorder by a bone-marrow transplant we can do the
bone-marrow transplant we can do the
bone-marrow transplant we can do the opposite we can put mutant bone marrow
opposite we can put mutant bone marrow
opposite we can put mutant bone marrow into a normal Mouse and give it the
into a normal Mouse and give it the
into a normal Mouse and give it the pathology
pathology
pathology so it’s both necessary and sufficient so
so it’s both necessary and sufficient so
so it’s both necessary and sufficient so next thing we looked at is the source of
next thing we looked at is the source of
next thing we looked at is the source of my nibbly arm and the reason for that is
my nibbly arm and the reason for that is
my nibbly arm and the reason for that is that the dog mind the literature was
that the dog mind the literature was
that the dog mind the literature was that there’s only a single source all
that there’s only a single source all
that there’s only a single source all microglia come from what’s called
microglia come from what’s called
microglia come from what’s called primitive hemopoiesis
primitive hemopoiesis
primitive hemopoiesis blood cells are made in during
blood cells are made in during
blood cells are made in during embryogenesis both in humans and mice
embryogenesis both in humans and mice
embryogenesis both in humans and mice and a very complicated set of reactions
and a very complicated set of reactions
and a very complicated set of reactions they start in the yolk sac which is
they start in the yolk sac which is
they start in the yolk sac which is actually outside the embryo and then
actually outside the embryo and then
actually outside the embryo and then they go to the AGM and then they go to
they go to the AGM and then they go to
they go to the AGM and then they go to fetal liver and then they finally end up
fetal liver and then they finally end up
fetal liver and then they finally end up in the bone marrow
in the bone marrow
in the bone marrow so they transition from one population
so they transition from one population
so they transition from one population to the next population and then finally
to the next population and then finally
to the next population and then finally end up in the bone marrow there so we we
end up in the bone marrow there so we we
end up in the bone marrow there so we we had two sources we had two types of
had two sources we had two types of
had two sources we had two types of microglia and the question is is there a
microglia and the question is is there a
microglia and the question is is there a single source or two a couple of sources
single source or two a couple of sources
single source or two a couple of sources and if there are different sources what
and if there are different sources what
and if there are different sources what is different about them now in terms of
is different about them now in terms of
is different about them now in terms of what the dogma set is all microglia come
what the dogma set is all microglia come
what the dogma set is all microglia come from yolk sac and then directly go to
from yolk sac and then directly go to
from yolk sac and then directly go to the brain okay and that happens at
the brain okay and that happens at
the brain okay and that happens at what’s called nine and a half days of
what’s called nine and a half days of
what’s called nine and a half days of gestation told gestation period for a
gestation told gestation period for a
gestation told gestation period for a mouse is 19 and a half days okay not
mouse is 19 and a half days okay not
mouse is 19 and a half days okay not nine months but just 19 and a half
nine months but just 19 and a half
nine months but just 19 and a half very quick okay so let’s do the
very quick okay so let’s do the
very quick okay so let’s do the experiment the way we did the experiment
experiment the way we did the experiment
experiment the way we did the experiment is to label now GFP that green
is to label now GFP that green
is to label now GFP that green fluorescent protein and put that on all
fluorescent protein and put that on all
fluorescent protein and put that on all microglia so it that allows us to see
microglia so it that allows us to see
microglia so it that allows us to see everything then there’s another
everything then there’s another
everything then there’s another jellyfish protein they’re actually for
jellyfish protein they’re actually for
jellyfish protein they’re actually for Isis red and we put that into Hawks BA
Isis red and we put that into Hawks BA
Isis red and we put that into Hawks BA under the control of Hawks b8 and so
under the control of Hawks b8 and so
under the control of Hawks b8 and so those make red ones and so red and green
those make red ones and so red and green
those make red ones and so red and green together then make yellow ones and so
together then make yellow ones and so
together then make yellow ones and so again you see two groups you see green
again you see two groups you see green
again you see two groups you see green ones which if they’re totally green come
ones which if they’re totally green come
ones which if they’re totally green come from yolk sac and then if they’re yellow
from yolk sac and then if they’re yellow
from yolk sac and then if they’re yellow maybe same source of different source
maybe same source of different source
maybe same source of different source okay this is actually a live image in a
okay this is actually a live image in a
okay this is actually a live image in a live mouse using a two-photon
live mouse using a two-photon
live mouse using a two-photon multiphoton microscope so because see
multiphoton microscope so because see
multiphoton microscope so because see cells live and great resolution okay so
cells live and great resolution okay so
cells live and great resolution okay so the first thing we ask is when do we
the first thing we ask is when do we
the first thing we ask is when do we first see yellow cells in the brain at
first see yellow cells in the brain at
first see yellow cells in the brain at nine and a half days they’re station you
nine and a half days they’re station you
nine and a half days they’re station you see a wave of green cells going into the
see a wave of green cells going into the
see a wave of green cells going into the brain and so we can say everything’s
brain and so we can say everything’s
brain and so we can say everything’s green and then we don’t see any yellow
green and then we don’t see any yellow
green and then we don’t see any yellow cells until II twelve and a half so this
cells until II twelve and a half so this
cells until II twelve and a half so this is three and a half days later which is
is three and a half days later which is
is three and a half days later which is a long period in during development
a long period in during development
a long period in during development relative to a mouse because 19 and a
relative to a mouse because 19 and a
relative to a mouse because 19 and a half is totally gestation period and
half is totally gestation period and
half is totally gestation period and what we can see is they start here and
what we can see is they start here and
what we can see is they start here and then you continue p0 is birth and then
then you continue p0 is birth and then
then you continue p0 is birth and then they continue to get up and then eight
they continue to get up and then eight
they continue to get up and then eight days after birth now they reach pretty
days after birth now they reach pretty
days after birth now they reach pretty much a steady state of being about 30%
much a steady state of being about 30%
much a steady state of being about 30% of all microglia
of all microglia
of all microglia so there are two possibilities to
so there are two possibilities to
so there are two possibilities to explain this result one is that a
explain this result one is that a
explain this result one is that a subpopulation of green cells turn on
subpopulation of green cells turn on
subpopulation of green cells turn on Hawks b8 and then become yellow cells
Hawks b8 and then become yellow cells
Hawks b8 and then become yellow cells the other possibility is that there’s a
the other possibility is that there’s a
the other possibility is that there’s a completely new source coming from the
completely new source coming from the
completely new source coming from the outside the brain and then getting into
outside the brain and then getting into
outside the brain and then getting into the brain at E starting at eight twelve
the brain at E starting at eight twelve
the brain at E starting at eight twelve and a half days of gestation so those
and a half days of gestation so those
and a half days of gestation so those are the possibilities and we could
are the possibilities and we could
are the possibilities and we could address that simply by looking when is
address that simply by looking when is
address that simply by looking when is Hawks p8 actually made and what we can
Hawks p8 actually made and what we can
Hawks p8 actually made and what we can see is it’s being made in the yolk sac
see is it’s being made in the yolk sac
see is it’s being made in the yolk sac just like the other microglia that’s and
just like the other microglia that’s and
just like the other microglia that’s and then it’s still being made a little bit
then it’s still being made a little bit
then it’s still being made a little bit and then a
and then a
and then a I am still being made and then by the
I am still being made and then by the
I am still being made and then by the time it gets to feel deliver it’s no
time it gets to feel deliver it’s no
time it gets to feel deliver it’s no longer being made but in the brain it’s
longer being made but in the brain it’s
longer being made but in the brain it’s never made being made yeah so that rules
never made being made yeah so that rules
never made being made yeah so that rules out the first hypothesis can’t be a
out the first hypothesis can’t be a
out the first hypothesis can’t be a subgroup turning on oxb 8 because it’s
subgroup turning on oxb 8 because it’s
subgroup turning on oxb 8 because it’s never made it in the brain yeah so that
never made it in the brain yeah so that
never made it in the brain yeah so that cleared that situation the other thing
cleared that situation the other thing
cleared that situation the other thing we can ask is where are the yellow cells
we can ask is where are the yellow cells
we can ask is where are the yellow cells in this case we call it red cells
in this case we call it red cells
in this case we call it red cells being made in terms of Hema polices and
being made in terms of Hema polices and
being made in terms of Hema polices and what we can see is a small amount in the
what we can see is a small amount in the
what we can see is a small amount in the yolk sac now all of a sudden lots in the
yolk sac now all of a sudden lots in the
yolk sac now all of a sudden lots in the AGM and really a lot in the fetal liver
AGM and really a lot in the fetal liver
AGM and really a lot in the fetal liver so most here there’s an Eightfold
so most here there’s an Eightfold
so most here there’s an Eightfold increase essentially in numbers of cells
increase essentially in numbers of cells
increase essentially in numbers of cells and then here is actually a 272 fold
and then here is actually a 272 fold
and then here is actually a 272 fold increase in the fetal liver and then
increase in the fetal liver and then
increase in the fetal liver and then they go into the brain okay so normally
they go into the brain okay so normally
they go into the brain okay so normally the most of the population the green
the most of the population the green
the most of the population the green population goes directly from yolk sac
population goes directly from yolk sac
population goes directly from yolk sac to the brain
to the brain
to the brain this one goes from the yolk sac to the
this one goes from the yolk sac to the
this one goes from the yolk sac to the AGM to the fetal liver and then into the
AGM to the fetal liver and then into the
AGM to the fetal liver and then into the brain why is this important because as
brain why is this important because as
brain why is this important because as it going through these different organs
it going through these different organs
it going through these different organs it’s getting different signals and then
it’s getting different signals and then
it’s getting different signals and then different capacity and different
different capacity and different
different capacity and different functions so that then that second
functions so that then that second
functions so that then that second population is different from the other
population is different from the other
population is different from the other population so the next thing we asked is
population so the next thing we asked is
population so the next thing we asked is how similar are these two populations if
how similar are these two populations if
how similar are these two populations if there are two are they doing the same
there are two are they doing the same
there are two are they doing the same thing or not I’ll give you just one
thing or not I’ll give you just one
thing or not I’ll give you just one example so what we’re gonna see is we
example so what we’re gonna see is we
example so what we’re gonna see is we are using a laser to make damage they
are using a laser to make damage they
are using a laser to make damage they are BAM okay we kill a few neurons and
are BAM okay we kill a few neurons and
are BAM okay we kill a few neurons and now we see all of these microglia the
now we see all of these microglia the
now we see all of these microglia the yellow ones and green ones sending their
yellow ones and green ones sending their
yellow ones and green ones sending their pseudo parts to the that region to then
pseudo parts to the that region to then
pseudo parts to the that region to then correct what’s happening they’re getting
correct what’s happening they’re getting
correct what’s happening they’re getting rid of those dead cells okay but they
rid of those dead cells okay but they
rid of those dead cells okay but they get their red at the same time so that
get their red at the same time so that
get their red at the same time so that tells us that in terms of that function
tells us that in terms of that function
tells us that in terms of that function they’re identical how are they different
they’re identical how are they different
they’re identical how are they different if we look in the brain itself so this
if we look in the brain itself so this
if we look in the brain itself so this is a section of the brain this is the
is a section of the brain this is the
is a section of the brain this is the front part of the brain the back part of
front part of the brain the back part of
front part of the brain the back part of the brain and we’re interested in
the brain and we’re interested in
the brain and we’re interested in looking at
looking at
looking at microglia either red ones or green ones
microglia either red ones or green ones
microglia either red ones or green ones and we could look at the ratio
and we could look at the ratio
and we could look at the ratio and we can see here there’s a lot of
and we can see here there’s a lot of
and we can see here there’s a lot of rocks being microglia but not very many
rocks being microglia but not very many
rocks being microglia but not very many green microglia over here we have the
green microglia over here we have the
green microglia over here we have the opposite we have a lot of green
opposite we have a lot of green
opposite we have a lot of green microglia and then very little rocks
microglia and then very little rocks
microglia and then very little rocks being multiplied up now why is this
being multiplied up now why is this
being multiplied up now why is this interesting this is the primary and
interesting this is the primary and
interesting this is the primary and secondary motor cortex this is the
secondary motor cortex this is the
secondary motor cortex this is the anterior aspect of the striatum this is
anterior aspect of the striatum this is
anterior aspect of the striatum this is a lateral orbital cortex and by fMRI
a lateral orbital cortex and by fMRI
a lateral orbital cortex and by fMRI studies in humans this is known as the
studies in humans this is known as the
studies in humans this is known as the OCD circuit okay so that’s the circuits
OCD circuit okay so that’s the circuits
OCD circuit okay so that’s the circuits are required to do this behavior and
are required to do this behavior and
are required to do this behavior and then to monitor this behavior how much
then to monitor this behavior how much
then to monitor this behavior how much of it do you do and in OCD what this is
of it do you do and in OCD what this is
of it do you do and in OCD what this is all repetitive behavior and what’s
all repetitive behavior and what’s
all repetitive behavior and what’s happening if this circuit just goes wild
happening if this circuit just goes wild
happening if this circuit just goes wild and keeps on going and going rather than
and keeps on going and going rather than
and keeps on going and going rather than knowing when to stop for example
knowing when to stop for example
knowing when to stop for example patients continually wash their hands
patients continually wash their hands
patients continually wash their hands and then they finished and they think
and then they finished and they think
and then they finished and they think their hands are still dirty they
their hands are still dirty they
their hands are still dirty they continue to wash until they have
continue to wash until they have
continue to wash until they have lacerations it’s repetitive behavior
lacerations it’s repetitive behavior
lacerations it’s repetitive behavior going awry and what we see is that our
going awry and what we see is that our
going awry and what we see is that our Hawks b8 cells are right in the region
Hawks b8 cells are right in the region
Hawks b8 cells are right in the region to be doing what they should be doing
to be doing what they should be doing
to be doing what they should be doing that is controlling this behavior so and
that is controlling this behavior so and
that is controlling this behavior so and that’s where we are right now so here is
that’s where we are right now so here is
that’s where we are right now so here is our OCD circuit here are microglia
our OCD circuit here are microglia
our OCD circuit here are microglia somehow these microglia are controlling
somehow these microglia are controlling
somehow these microglia are controlling that circuit and we have no idea what
that circuit and we have no idea what
that circuit and we have no idea what the mechanism and there are many many
the mechanism and there are many many
the mechanism and there are many many possibilities an example is they could
possibilities an example is they could
possibilities an example is they could be producing chemicals cytokines
be producing chemicals cytokines
be producing chemicals cytokines chemokines neurotransmitters which then
chemokines neurotransmitters which then
chemokines neurotransmitters which then would be affecting the circuit the other
would be affecting the circuit the other
would be affecting the circuit the other is even more interesting there are all
is even more interesting there are all
is even more interesting there are all of these long pseudo parts that you
of these long pseudo parts that you
of these long pseudo parts that you sticking out and they’re actually very
sticking out and they’re actually very
sticking out and they’re actually very mobile they keep moving back and forth
mobile they keep moving back and forth
mobile they keep moving back and forth and in fact they’re so active then in a
and in fact they’re so active then in a
and in fact they’re so active then in a period of about 30 minutes they can
period of about 30 minutes they can
period of about 30 minutes they can actually and scan the entire brain
actually and scan the entire brain
actually and scan the entire brain surface okay and be able to sense what’s
surface okay and be able to sense what’s
surface okay and be able to sense what’s going on there and what you see is those
going on there and what you see is those
going on there and what you see is those pseudopods are moving around moving
pseudopods are moving around moving
pseudopods are moving around moving around then all of a sudden they stop
around then all of a sudden they stop
around then all of a sudden they stop and then the pseudopods go around a
and then the pseudopods go around a
and then the pseudopods go around a synapse at the connection between two
synapse at the connection between two
synapse at the connection between two neurons and completely as circulated and
neurons and completely as circulated and
neurons and completely as circulated and they could actually physically be
they could actually physically be
they could actually physically be monitoring how much traffic is going
monitoring how much traffic is going
monitoring how much traffic is going across that synapse okay so that’s
across that synapse okay so that’s
across that synapse okay so that’s another possibility and what’s nice is
another possibility and what’s nice is
another possibility and what’s nice is that as you’re going along the
that as you’re going along the
that as you’re going along the technology to be able to address these
technology to be able to address these
technology to be able to address these questions is coming in okay there’s a
questions is coming in okay there’s a
questions is coming in okay there’s a technology now known as optogenetics
technology now known as optogenetics
technology now known as optogenetics where you turn on the light and you can
where you turn on the light and you can
where you turn on the light and you can make a essentially a neuron be activated
make a essentially a neuron be activated
make a essentially a neuron be activated or we can actually make it pump another
or we can actually make it pump another
or we can actually make it pump another molecule and it becomes silent okay and
molecule and it becomes silent okay and
molecule and it becomes silent okay and if the reaction is so fast if it takes
if the reaction is so fast if it takes
if the reaction is so fast if it takes place in milliseconds we turned on the
place in milliseconds we turned on the
place in milliseconds we turned on the light stops sir I’m gonna turn off the
light stops sir I’m gonna turn off the
light stops sir I’m gonna turn off the light boom and then it’s all off again
light boom and then it’s all off again
light boom and then it’s all off again boom boom boom and that’s then
boom boom boom and that’s then
boom boom boom and that’s then information because that’s a frequency
information because that’s a frequency
information because that’s a frequency okay so then what we can ask you is we
okay so then what we can ask you is we
okay so then what we can ask you is we can turn on and off a neuron and then
can turn on and off a neuron and then
can turn on and off a neuron and then ask the microglia nearby is it
ask the microglia nearby is it
ask the microglia nearby is it responding in the same frequency if they
responding in the same frequency if they
responding in the same frequency if they are they’re talking to each other and if
are they’re talking to each other and if
are they’re talking to each other and if they’re talking to each other then we
they’re talking to each other then we
they’re talking to each other then we can ask what what is that language and
can ask what what is that language and
can ask what what is that language and so we have what now an entrant just an
so we have what now an entrant just an
so we have what now an entrant just an entry to see how microglia are
entry to see how microglia are
entry to see how microglia are controlling neural circuits and what’s
controlling neural circuits and what’s
controlling neural circuits and what’s surprising about this is that know why
surprising about this is that know why
surprising about this is that know why in the world would that microglia be
in the world would that microglia be
in the world would that microglia be controlling neural circuits and I think
controlling neural circuits and I think
controlling neural circuits and I think we have some ideas and that’ll be the
we have some ideas and that’ll be the
we have some ideas and that’ll be the next lecture thank you
next lecture thank you
next lecture thank you [Applause]
[Applause]
[Applause] [Music]
”
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