2016 Fiscal Year Annual Research Report
| Project/Area Number |
16J07688
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| Research Institution | The Graduate University for Advanced Studies |
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Principal Investigator |
CHEN PEIーJU 総合研究大学院大学, 先導科学研究科, 特別研究員(DC1)
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| Project Period (FY) |
2016-04-22 – 2019-03-31
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| Keywords | insect / vision / optic lobe / photoreceptor / spectral opponency / histamine channel / synapse |
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| Outline of Annual Research Achievements |
This study aims at understanding the mechanism of spectral opponency at the initial stage of visual processing. Since synapse-like structures exist between photoreceptors of Papilio, and histamine is the transmitter in insect photoreceptors, I have hypothesized histaminergic channels exist between photoreceptors. Based on the “histamine hypothesis”, two specific antisera were raised against peptides specific to the Papilio xuthus-homologs of (Drosophila) histamine-gated chloride channels, PxHClA and PxHClB.
This year I have studied the possible distribution of histamine-gated chloride channels via immunohistochemical localization in the optic lope of Papilio. The anti-PxHClA immunoreactivity is associated with the plasma membrane of non-photoreceptor neurons, suggesting that PxHClA is responsible for synaptic transmission at the photoreceptor-LMC synapse. We also found that photoreceptors are immunopositive to anti-PxHClB, indicating possible inter-photoreceptor synaptic contacts: if so, the PxHClB may be involved in the photoreceptor spectral opponency. We further simulated the color discrimination ability based on two photoreceptors with different spectral sensitivities. This result suggests that wavelength discriminability would be better under an opponent mechanism.
Furthermore, the gene products of PxHClA and PxHClB were transfected to cultured cells and analyzed for their ion channel properties by whole-cell patch-clamp.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
The immunohistochemistry of anti-PxHClA and anti-PxHClB at both light microscopic (LM) and electron microscopic (EM) levels is currently being finalized. Overall, the present results stand as a strong evidence for our “histamine hypothesis”.
The patch-clamp experiments of characterizing the channel properties of artificially expressed PxHClA and PxHClB were carried out at the National Institute of Physiological Sciences (NIPS) under collaboration with Prof. Makoto Tominaga. The dose-response characteristics of PxHClA and PxHClB receptors supported these molecules are in fact gated by histamine and permeate chloride ions. We also observed both PxHClA and PxHClB receptors response to chemicals other than histamine.
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| Strategy for Future Research Activity |
Based on the immunohistochemical results, I am preparing a manuscript of identification and localization of the putative histamine-gated chloride channels in the eye of P. xuthus.
To characterize ion channel properties, other chemicals will be applied to artificially expressed PxHClA and PxHClB cells and analyzed by patch-clamp. Moreover, I will try slice recordings from native cells. Through comparing the property of transfected cells and those of native channels from photoreceptors and LMCs of living animals, we can further confirm the localization of putative channels.
In addition, I plan to attend the workshop of microelectrode techniques for cell physiology in Plymouth, UK and visit Prof. Belusic at University Ljubljana to learn electrophysiological techniques for experiments.
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