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We want to know how the brain works during perception and behaviors. Zebrafish larva
can recognize the paramecium at 5 days postfertilization. The fish finds a paramecium, approaches the
paramecium, and catch it. We aimed to visualize brain activity during perception of a paramecium.
In order to make this possible, we developed tools and methods in the model vertebrate,
zebrafish. One important tool is GCaMP. GCaMP was invented by Dr. Nakai and Dr. Ohkura at
Saitama University in 2001. GCaMP contains M13, GFP, and calmodulin sequences. When calcium
ions bind to the calmodulin, conformational change occurs and GFP fluorescence increases.
Because calcium influx occurres when a neuron is activated, we can use GCaMP to detect neuronal
activity. Further, in this study, we developed a new GCaMP, which is more sensitive than
the old GCaMP. Another important technique is the Gal4-UAS system. Gal4 binds to UAS,
and activates transcription of a gene placed downstream of UAS. In my lab, we are doing
gene trap screens and generated more than 1,000 transgenic fish lines that express Gal4
in different tissues.
Each tank has a different line.
Animals, including humans and fish, have common features in the brain structure. One of such
features is the visuotopic map (retinotopic map). What it means is that neurons in the
brain are spatially organized so that the visual world is mapped on a part of the brain.
This is well-established notion, but no one has ever demonstrated this in real time in
a natural condition. In this study, we wanted to visualize this using our new GCaMP in prey
capture behavior. In fish, the optic tectum is the center for visual information processing.
So, first, we looked at our database and chose one Gal4 line, in which the gal4 was expressed
in the optic tectum. On the other hand, we generated UAS:GCaMP7a transgenic fish. By
mating that Gal4 fish with UAS:GCaMP7a fish, we expressed GCaMP7a in the optic tectum of
the larvae. The experiment is straightforward. We let a paramecium swim around a larva, and
do the calcium imaging using a fluorescence microscope. Here is what you see in the recording.
Changes in fluorescence intensity is shown in pseudo-colors. Colors in the eye balls
are artifacts caused by image processing due to eye movements. Neuronal signals are observed
in the optic tectum. We could also record from a free-swimming larva which captured
a paramecium. We visually demonstrated that the paramecium was represented on the visuotopic
map as predicted by anatomical studies. And activation in the anterior part of the optic
tectum was correlated with the occurrence of the prey capture behavior. In this study,
we imaged neuronal activity in the initial step of prey capture. Using this method that
we have developed, now we are trying to visualize neuronal activity that underlies cognitive
process.