title: Real-time electrochemical recording of dopamine release under optogenetic stimulation.
abstract: Dopaminergic PC12 cells can synthesize and release dopamine, providing a good cellular model for investigating dopamine regulation. Optogenetic stimulation of channelrhodopsin-2 provides high spatial and temporal precision for selective stimulation as a powerful neuromodulation tool for neuroscience studies. The aim of this study is to measure dopamine release from dopaminergic PC12 cells under optogenetic stimulation using electrochemical recording of self-assembled monolayers modified microelectrode with amperometric measurement in real time. The activation of PC12 cells under various optogenetic stimulation schemes are characterized by measuring single-cell Ca(2+) imaging. After 10 seconds of optogenetic stimulation, the evoked intracellular Ca(2+) level and dopamine current of channelrhodopsin-2-transfected PC12 cells were 1.6- and 3.5-fold higher than those of the control cells. The optogenetic stimulation effects on Ca(2+) influx and dopamine release were 81% and 63% inhibition by using a Ca(2+) channel antagonist Nifedipine. The results indicate that optogenetic stimulation can evoke voltage-gated Ca(2+) channel-dependent dopamine exocytosis from PC12 cells in a cell specific, temporally precise and dose-dependent manner. This proposed dopamine recording system can be developed to be a good cell model for dopamine regulation and drug screening in vitro, or dopaminergic cell implantation therapy in vivo using optogenetic stimulation in a precise and convenient way.
authors.affiliation: Department of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan ; Institute of Basic Medical Sciences, National Cheng Kung University, Tainan, Taiwan ; Medical Device Innovation Center, National Cheng Kung University, Tainan, Taiwan.
authors.forename: Wen-Tai
authors.initials: WT
authors.lastname: Chiu
authors.affiliation: Department of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan.
authors.forename: Che-Ming
authors.initials: CM
authors.lastname: Lin
authors.affiliation: Department of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan.
authors.forename: Tien-Chun
authors.initials: TC
authors.lastname: Tsai
authors.affiliation: Department of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan.
authors.forename: Chun-Wei
authors.initials: CW
authors.lastname: Wu
authors.affiliation: Department of Marine Biotechnology and Resources, National Sun Yat-Sen University, Kaohsiung, Taiwan.
authors.forename: Ching-Lin
authors.initials: CL
authors.lastname: Tsai
authors.affiliation: Institute of Clinical Medicine, National Cheng Kung University, Tainan, Taiwan.
authors.forename: Sheng-Hsiang
authors.initials: SH
authors.lastname: Lin
authors.affiliation: Department of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan ; Medical Device Innovation Center, National Cheng Kung University, Tainan, Taiwan ; National Applied Research Laboratories, Taipei, Taiwan.
authors.forename: Jia-Jin Jason
authors.initials: JJ
authors.lastname: Chen
mesh_terms: D000818:Animals; D015374:Biosensing Techniques; D002118:Calcium / Q000378:metabolism; D000075402:Channelrhodopsins; D004298:Dopamine / Q000378:metabolism*; D055664:Electrochemical Techniques; D005089:Exocytosis / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008839:Microelectrodes; D062308:Optogenetics; D016716:PC12 Cells; D051381:Rats
chemical_list: D000075402:Channelrhodopsins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D002118:Calcium; D004298:Dopamine