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Neuroscience
気相で、嗅覚による嗅覚受容体活性化の In Vitro モニタリング リアルタイム
気相で、嗅覚による嗅覚受容体活性化の In Vitro モニタリング リアルタイム
JoVE Journal
Neuroscience
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JoVE Journal Neuroscience
Real-time In Vitro Monitoring of Odorant Receptor Activation by an Odorant in the Vapor Phase

気相で、嗅覚による嗅覚受容体活性化の In Vitro モニタリング リアルタイム

Full Text
7,521 Views
09:53 min
April 23, 2019

DOI: 10.3791/59446-v

Claire A. de March1, Yosuke Fukutani1,2, Aashutosh Vihani1,3, Hitoshi Kida1,4, Hiroaki Matsunami1,3,5,6

1Department of Molecular Genetics and Microbiology,Duke University Medical Center, 2Department of Biotechnology and Life Science,Tokyo University of Agriculture and Technology, 3Department of Neurobiology,Duke University Medical Center, 4Department of Mechanical Systems, Engineering,Tokyo University of Agriculture and Technology, 5Institute of Global Innovation Research,Tokyo University of Agriculture and Technology, 6Duke Institute for Brain Sciences,Duke University

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Overview

This study presents a novel method for real-time in vitro monitoring of odorant receptor activation using vapor phase odorant stimulation. Previous in vitro systems have largely relied on liquid phase stimulation, potentially limiting physiological relevance. The method provides insights into the activation kinetics of odorant receptors and their role in odor perception.

Key Study Components

Area of Science

  • Neuroscience
  • Olfactory biology
  • Cell culture techniques

Background

  • Odorant receptors respond to volatile odorant molecules, crucial for the sense of smell.
  • Existing in vitro methods primarily use liquid phase odorants, which may not accurately mimic natural conditions.
  • This research addresses gaps in current methodologies by allowing for vapor phase stimulation.

Purpose of Study

  • To develop a method that facilitates the detection and discrimination of volatile odors in vitro.
  • To enable real-time monitoring of odorant receptor activation.
  • To bridge the gap between in vitro and in vivo approaches in olfactory research.

Methods Used

  • Utilized cell culture techniques to grow cells in M10 PSF medium at physiological conditions.
  • Involves transfection of cells with specific odorant receptor plasmids to study their activation.
  • Key steps include careful cell preparation, including trypsinization and centrifugation.
  • Measurements of luminescence provide data on receptor activation upon exposure to odorants.

Main Results

  • The method allows for precise monitoring of odorant receptor responses to specific odorants like acetophenone.
  • Findings illustrate the kinetics of odor perception and potential roles of mucosal metabolic enzymes.
  • Successfully demonstrated the capability to detect and discriminate odors in a controlled in vitro setting.

Conclusions

  • This innovative approach enhances the understanding of olfactory receptor dynamics in a more physiologically relevant context.
  • The findings enable further research into the development of odorant receptor-based biosensors.
  • Implications extend to understanding mechanisms of odor detection, which could inform broader olfactory studies.

Frequently Asked Questions

What are the advantages of using vapor phase stimulation?
Vapor phase stimulation mimics natural conditions, providing a more physiologically relevant method to study odorant receptors compared to liquid phase approaches.
How is the biological model implemented in this study?
Cells are cultured in specific media and transfected with plasmids encoding odorant receptors to investigate their responses to volatile stimuli.
What types of data are obtained from this method?
The methodology provides luminescence data indicating the activation levels of odorant receptors in response to odorant exposure.
How can this method be applied to other research areas?
This technique can potentially be adapted for various studies involving sensory receptor activation and biosensor development across biological systems.
What are the key limitations or considerations for this study?
Care must be taken to ensure that experimental conditions, such as temperature and CO2 levels, remain constant to obtain accurate results in receptor activation studies.

生理的には、嗅覚受容体は匂い分子気相に吸入によってアクティブ化されます。ただし、ほとんどの in vitro システムは、液相嗅覚刺激を利用します。嗅覚刺激による気相匂い物質受容体活性化のリアルタイムでの in vitro の監視を可能にする方法を紹介します。

当社のプロトコルは、臭気受容体のパネルで揮発性臭気の検出と識別を測定することができます。この方法は、臭気受容体ベースのバイオセンサーを開発する可能性を開きます。この技術により、気相の臭気分子に対する臭気受容体活性化のリアルタイムインビトロモニタリングが可能になります。

それはインビトロとインビボアプローチの間のギャップを埋める。この方法は、粘膜代謝酵素の役割を含む、臭気知覚につながる事象の運動学を理解するために使用することができる。まず、原稿に従ってM10とM10 PSFを準備します。

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神経科学 問題 146 蒸気の刺激 嗅覚受容体 リアルタイム アクティブ化 匂い分子 組合せのコード 機能試金

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