PMID- 35435901
OWN - NLM
STAT- MEDLINE
DCOM- 20220420
LR  - 20220507
IS  - 1940-087X (Electronic)
IS  - 1940-087X (Linking)
IP  - 181
DP  - 2022 Mar 31
TI  - High-Throughput Optical Controlling and Recording Calcium Signal in iPSC-Derived 
      Cardiomyocytes for Toxicity Testing and Phenotypic Drug Screening.
LID - 10.3791/63175 [doi]
AB  - Understanding how excitable cells work in health and disease and how that 
      behavior can be altered by small molecules or genetic manipulation is important. 
      Genetically encoded calcium indicators (GECIs) with multiple emission windows can 
      be combined (e.g., for simultaneous observation of distinct subcellular events) 
      or used in extended applications with other light-dependent actuators in 
      excitable cells (e.g., combining genetically encoded optogenetic control with 
      spectrally compatible calcium indicators). Such approaches have been used in 
      primary or stem cell-derived neurons, cardiomyocytes, and pancreatic beta-cells. 
      However, it has been challenging to increase the throughput, or duration of 
      observation, of such approaches due to limitations of the instruments, analysis 
      software, indicator performance, and gene delivery efficiency. Here, a 
      high-performance green GECI, mNeonGreen-GECO (mNG-GECO), and red-shifted GECI, 
      K-GECO, is combined with optogenetic control to achieve all-optical control and 
      visualization of cellular activity in a high-throughput imaging format using a 
      High-Content Imaging System. Applications demonstrating cardiotoxicity testing 
      and phenotypic drug screening with healthy and patient-derived iPSC-CMs are 
      shown. In addition, multi-parametric assessments using combinations of spectral 
      and calcium affinity indicator variants (NIR-GECO, LAR-GECO, and mtGCEPIA or 
      Orai1-G-GECO) are restricted to different cellular compartments are also 
      demonstrated in the iPSC-CM model.
FAU - Chang, Yu-Fen
AU  - Chang YF
AD  - LumiSTAR Biotechnology, Inc.; yu-fen.chang@lumistar.com.tw.
FAU - Su, Wan-Chi
AU  - Su WC
AD  - LumiSTAR Biotechnology, Inc.
FAU - Su, Chih-Chuan
AU  - Su CC
AD  - LumiSTAR Biotechnology, Inc.
FAU - Chung, Min-Wen
AU  - Chung MW
AD  - LumiSTAR Biotechnology, Inc.
FAU - Chang, Jin
AU  - Chang J
AD  - NEXEL Co., Ltd.
FAU - Li, You-Yi
AU  - Li YY
AD  - Institute of Pharmacology, College of Medicine, National Taiwan University.
FAU - Kao, Yi-Ju
AU  - Kao YJ
AD  - Institute of Pharmacology, College of Medicine, National Taiwan University.
FAU - Chen, Wen-Pin
AU  - Chen WP
AD  - Institute of Pharmacology, College of Medicine, National Taiwan University.
FAU - Daniels, Matthew J
AU  - Daniels MJ
AD  - Manchester Heart Centre, Manchester Royal Infirmary, Manchester University NHS 
      Foundation Trust; Division of Cardiovascular Sciences, Manchester Academic Health 
      Sciences Centre, University of Manchester; Division of Cell Matrix Biology and 
      Regenerative Medicine, University of Manchester.
LA  - eng
PT  - Journal Article
PT  - Video-Audio Media
DEP - 20220331
PL  - United States
TA  - J Vis Exp
JT  - Journal of visualized experiments : JoVE
JID - 101313252
RN  - 0 (Indicators and Reagents)
RN  - SY7Q814VUP (Calcium)
SB  - IM
MH  - *Calcium/analysis
MH  - Drug Evaluation, Preclinical
MH  - Humans
MH  - Indicators and Reagents
MH  - *Induced Pluripotent Stem Cells/chemistry
MH  - Myocytes, Cardiac/chemistry
MH  - Optogenetics
EDAT- 2022/04/19 06:00
MHDA- 2022/04/21 06:00
CRDT- 2022/04/18 12:12
PHST- 2022/04/18 12:12 [entrez]
PHST- 2022/04/19 06:00 [pubmed]
PHST- 2022/04/21 06:00 [medline]
AID - 10.3791/63175 [doi]
PST - epublish
SO  - J Vis Exp. 2022 Mar 31;(181). doi: 10.3791/63175.