演題詳細
Poster
マイクロコンタクトプリンティング法による培養マウス海馬神経細胞の軸索誘導
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.