| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2006: ¥300,000 (Direct Cost: ¥300,000)
Fiscal Year 2005: ¥3,400,000 (Direct Cost: ¥3,400,000)
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| Research Abstract |
Animal brain is too complex to record entirely their activity at cellular level. Ascidian larva has extremely simple brain comprising only 100 neurons although the animal shares common basic body plans with vertebrates. I attempted to measure the neural activities of an ascidian, Ciona intestinalis, whole larva. I applied a Ca^2+ indicator protein, Cameleon, for this purpose, because it can be expressed selectively in certain cells by a proper promoter.
To set up the measurement system, I budgeted for buying color 3CCD camera. Reduction of budget compelled me to change my plan. I bought a high sensitive monochrome camera instead. I tried to measure FRET imaging by the camera but eventually arrived at the conclusion that the camera was hard to detect true Ca^2+ change. Therefore, I made experiments at RIKEN WAKO by using color 3CCD camera with collaboration from Dr. Miyawaki.
1. Ca^2+ imaging of the muscles
To test Ca^2+ imaging with Cameleon, I applied it to the muscle cells firstly. I could detect Ca^2+ transient with the firing of muscle cells. The larval muscle cells are not fused as those of vertebrates. The Ca^2+ imaging demonstrated that Ca^2+ signals were transmitted from anterior to posterior muscle through the gap junctions between muscle cells.
2. Ca^2+ imaging of the photoreceptors
I found that Ciona larval photoreceptors decreased Ca^2+ upon light stimuli as those of vertebrates. This is a first-ever in vivo Ca^2+ imaging in animal photoreceptors. Elucidation of the signal transduction of Ciona larva is a critical issue to understand the evolution of the vertebrate eyes. This measurement system is very useful for the study.
3. Ca^2+ imaging of cholinergic neurons
The larva has 5 pair motor neurons in the motor ganglion. Ca^2+ imaging showed that the 3rd cholinergic neurons show more active than other neurons, suggesting that the 3rd cholinergic neurons function as pace makers.
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