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開催日 2014/9/13
時間 11:00 - 12:00
会場 Poster / Exhibition(Event Hall B)

Microcontact printing of cell-adhesion molecules for directing axon-dendrite polarity of mouse hippocampal neurons in culture

  • P3-383
  • 高沖 英里 / Hidesato Takaoki:1 山本 英明 / Hideaki Yamamoto:2 桂林 秀太郎 / Shutaro Katsurabayashi:3 木村 康男 / Yasuo Kimura:4 平野 愛弓 / Ayumi Hirano-Iwata:1 庭野 道夫 / Michio Niwano:1,5 
  • 1:東北大院・医工・医工 / Graduate School of Biomedical Engineering, Tohoku Univ, Miyagi, Japan 2:東北大・学際フロンティア / FRIS, Tohoku Univ, Miyagi, Japan 3:福岡大薬 / Fac Pharm Sci, Fukuoka Univ, Fukuoka, Japan 4:東京工科大・コンピュータサイエンス / Sch Comput Sci, Tokyo Univ of Technol, Tokyo, Japan  5:東北大電気通信研 / RIEC, Tohoku Univ, Miyagi, Japan 

Nerve cells in dispersed culture have attracted attention as a simple model system for studying function of the nervous system. However, neurons grown on a conventional coverslip form a random network that is different from the structure of neuronal networks in vivo. In our study, we tried control of the position of cultured nerve cells and their axon-dendrite polarity by micropatterning asymmetric cell-adhesion domains on coverslips using the microcontact printing method.

Our micropattern consisted of a 30-μm circular island for soma adhesion, a 100-μm continuous line for axon growth, and a 50-μm dotted line (10-μm line separated by a 10-μm interval) for dendrite growth. Width of the lines were designed to be 5 μm. Using a mold fabricated by photolithography, a polydimethylsiloxane stamp was manufactured. Surface of the stamp was coated with a cell-permissive ink composed of poly-D-lysine and collagen, and the protein ink was transferred onto a coverslip. Neurons obtained from hippocampi of postnatal ICR mice (P1 or P2) were plated, and the coverslip was co-cultured with cortical astrocytes. After 3-5 days of culture, the cells were stained with Tau-1 antibody for axons and with MAP2 antibody for soma and dendrites.

Appropriate transfer of the micropattern was confirmed using FITC-labeled collagen. Neuronal culture on the micropatterned coverslip and subsequent immunostaining revealed that orientation of axon growth could be controlled with a probability of more than 60%. This result shows that axon-dendrite polarity of cultured nerve cells could successfully be controlled using the asymmetrically-shaped micropatterns.
Formation of synaptic contacts between two neighboring neurons are currently under investigation, and the results will be presented at the conference.

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