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New Biological Frontiers Illuminated by Molecular Sensors and Actuators
Tuesday Speaker Abstracts
Spying Neurotransmitter Release by New Genetically-encoded Indicators
Yulong Li
.
Peking University, P.R.China, Beijing, China.
The nerve communication relies on the vesicular release of various neurotransmitters from
presynaptic terminals. The various neurotransmitters are chemical diverse, but universally
concentrated into small synaptic vesicles (SVs, diameter 40-50 nm) before their discharge upon
the nerve activities. How high the concentration of the transmitters can reached inside the SVs?
How to visualize their release, especially in vivo? When and how long would they act on
postsynaptic neurons? We have developed various genetically-encoded sensors aiming to answer
those questions. First, we have generated voltage sensors that specifically targeted to the SVs.
These sensors allow us, for the first time, to measure the membrane potential across the SVs,
unapproachable by conventional electrophysiological means. Because the uptake of
neurotransmitter are determined by the proton-electrochemical gradients, with the known
intravesicular pH and membrane potential, we could estimate the steady state neurotransmitter
concentration. Secondly, to aid visualization of transmitter release in vivo, we have generated
super pH sensitivity fluorescent protein (pHlamingo) with large apparent stokes shift.
pHlamingo’s excitation and emission peaks are at 460 nm and 610 nm, respectively. Its pH
dependent fluorescent emission (pH7.5/pH5.5) is 20% higher than pHluorin. When targeting to
SVs, pHlamingo’s convenient excitation peak and superior pH sensitivity allows it to report the
release of neurotransmitter release, especially for the in-vivo 2-photon microscopy settings.
Finally, we have constructed sensors that can convert the ligand-dependent structural
rearrangement of receptors to the folding of a fluorescent protein. When targeting to the neuronal
cell surface, these sensors can sense transmitter release such as acetylcholine. We anticipate that
the further optimization of the new generation sensors would provide critical help to deepen our
understanding the regulation of neurotransmitter release in health and disease.
Imaging Intraorganellar Ca
2+
at Subcellular Resolution Using CEPIA
Junji Suzuki
1
, Kazunori Kanemaru
1
, Kuniaki Ishii
2
, Masamichi Ohkura
3
, Yohei Okubo
1
,
Masamitsu Iino
1
.
1
The University of Tokyo, Bunkyo-ku, Tokyo, Japan,
2
Yamagata University, Iida-nishi,
Yamagata, Japan,
3
Saitama University, Sakura-ku, Saitama, Japan.
See abstract: Pos-43 Board 43