of light into chemical signals through a phenomenon called the phototransduction cascade. Understanding this complex chemical chain of chemical
activations and structural changes in molecules
caused by photoexcitation is key to harnessing
their power to activate neurons, and even after
reaching competence in this domain Deisseroth
struggled for years to achieve the results in his
core domain, “behaving” mammals.
But other methods not using microbial opsins
hadn’t been practical or adopted by other labs
either. The core goal remained to be solved.
Deisseroth’s microbial opsin research came after
he had concluded a decidedly different pursuit: a
residency in clinical psychiatry. The draw to this
discipline had motivated his desire to develop technologies, and to find a different way of understanding and possibly treating disorders of the brain. It
was also, he explains, a valuable exposure to the
variety of expressions of brain activity dysfunction.
His residency, simultaneous with a fellowship under
Prof. Robert Malenka at Stanford, had exposed him
not only to psychiatric theory, but also to real people who demonstrated disorders of the brain daily
without any real hope of improvement. He grew
to appreciate the vastness of the divide between
neuroscience and clinical psychiatry.
“That was really transformative for me, to get
that experience of seeing and treating people with
different psychiatric diseases. To witness their
suffering and to see how different their reality was,
it showed me how vast was the set of unknowns,”
says Deisseroth. Unlike much of his work in the
laboratory and in the operating room, psychiatry
doesn’t easily create an approachable path, he says.
“There’s a lot of mystery there. Inadequate treat-
ments are common, and there’s no way of getting
into the complex structures of the human brain.”
But what psychiatry did do was give Deisseroth
extra drive in his opsin research. In 2004, he formed
the Deisseroth Lab with the intention of making
microbial opsins useful in neuroscience. He enlisted
several talented graduate students to his group,
including Feng Zhang, who has since gone on to
further fame with his discovery of genome editing
with the CRISPR-Cas9 system. In a flurry of work
over the next year, Deisseroth’s team achieved a
breakthrough that resulted in a 2005 paper pub-
lished in Nature Neuroscience. Deisseroth had, the
previous year, found that microbial opsins could
be safely expressed and functional in the cellular
membranes of mammalian neurons in culture. This
major step was made possible by learning how to
leverage gene delivery methods—basically infecting
brain cell cultures—to effectively express opsins and
place them in a position to stimulate mammalian
neurons using high-speed photoswitching.
Not long after, Deisseroth and his team did just
that, delivering flashes of blue light to cultured
rodent neurons in a petri dish. The team was
surprised at how well it worked when all of the re-
quired elements were brought together, including
molecular, viral, optical and electrophysiological
methods. But this was only in a dish, and was
not “optogenetics” yet, as Deisseroth would soon
call control of specified events in defined cells in
behaving animals with millisecond precision. As
with any new method, says Deisseroth, a number
of bugs had to be ironed out. “We had to overcome
about five or six major challenges to really make
optogenetics work,” says Deisseroth. “The goal
with the early research was slow to be achieved,
to show that light sensitivity can be conferred,
by microbial opsins, to defined cells in behaving
mammals via a robust, practical approach.”
In 2010, Deisseroth’s team eventually solved
the key problems of opsin targeting, expression in
mammals and deep brain light delivery. Optoge-
netics gained traction in research settings as more
robust opsins were developed. “Now, we’ve gotten
even better at guiding light,” says Deisseroth.
Today, clones of effective opsins are sent to
thousands of laboratories, and development of
better tools continues.
As optogenetics developments continued, Deisseroth tackled another longstanding problem:
visualizing structural detail throughout the entire
intact brain. High-resolution information about
complex biological systems is difficult to obtain.
Optical solutions rarely penetrate more than a few
tenths of a millimeter in living samples, and spe-
cial sample preparation techniques are required
with seemingly inevitable sample damage.
Deisseroth’s dream was to do what had been
thought impossible—creating transparent tissue
without disrupting it or losing information. He re-
cruited to his laboratory, as a postdoctoral fellow,
Kwanghun Chung, a chemical engineer by train-
ing. With the help of Chung and other colleagues,
Deisseroth’s team converted fully intact brain
tissue into an optically transparent form, sup-
ported by a nanoporous and macromolecule-per-
meable hydrogel. Cross-linked to 3-D network
of hydrophilic polymers, this transformed tissue
retains almost of all of the brain’s original neural
wiring information and biomolecules, including
neurotransmitters, nucleic acids and subcellular
structures. His 2013 paper describing the method
goes on to describe the use of lightsheet microsco-
py to image the transformed samples, as well detail
potential clinical tissue applications.
Although developed more than 100 years ago,
lightsheet microscopy had not been widely adopted
in biology until a few years ago. Because CLARITY
removes lipids, it allows the “sheet” of laser light to
scan through the sample with far less scattering.
“One of the issues that we faced early on in de-
veloping CLARITY was the fact that commercially
available light-sheet systems were engineered to
analyze samples at about 10- to 20-μm resolution.
For neuroscience, we needed to study individ-
ual axons, which meant an order of magnitude
increase in resolution, so we had to build our
own system an order-of-magnitude better,” says
Deisseroth, and this achievement became known
as “CLARITY-optimized lightsheet microscopy”.
Compared with his work to develop to optoge-
netics tools, the pace of development for CLAR-
ITY has been far more rapid and more quickly
applied by others. He credits this progress in part
to the availability of better instrumentation, but
also with the effectiveness of the team he brought
together to make it happen.
Deisseroth is quick to stress that neither
technology represents a treatment in its own
right, even when they do enter clinical settings.
But the potential, in fundamental research and
commercial domains, is considerable. In 2013, the
U.S. government committed hundreds of millions
of federal dollars to map the tens of billions of
neurons in the human brain. One of the BRAIN
Initiative’s top advocates, Deisseroth is now one of
the 15 advisory committee members. Innovations
like CLARITY and optogenetics tools will likely
contribute significantly to progress in this effort.
In 2013, Deisseroth’s team published a paper that described
the first successful effort to remove lipids from brain tissue,
leaving neurons and proteins intact for 3-D fluorescent
imaging. Image: Stanford Univ.