Cell culture chamber with gas supply for prolonged recording of human neuronal cells on microelectrode array.
Typically, live cell analyses are performed outside an incubator in an ambient air, where the lack of sufficient CO2supply results in a fast change of pH and the high evaporation causes concentration drifts in the culture medium. That limits the experiment time for tens of minutes. In many applications, e.g. in neurotoxicity studies, a prolonged measurement of extracellular activity is, however, essential.



We demonstrate a simple cell culture chamber that enables stable culture conditions during prolonged extracellular recordings on a microelectrode array (MEA) outside an incubator. The proposed chamber consists of a gas permeable silicone structure that enables gas transfer into the chamber.



We show that the culture chamber supports the growth of the human embryonic stem cell (hESC)-derived neurons both inside and outside an incubator. The structure provides very low evaporation, stable pH and osmolarity, and maintains strong signaling of hESC-derived neuronal networks over three-day MEA experiments.



Existing systems are typically complex including continuous perfusion of medium or relatively large amount of gas to supply. The proposed chamber requires only a supply of very low flow rate (1.5ml/min) of non-humidified 5% CO2gas. Utilizing dry gas supply makes the proposed chamber simple to use.



Using the proposed culture structure on top of MEA, we can maintain hESC-derived neural networks over three days outside an incubator. Technically, the structure requires very low flow rate of dry gas supporting, however, low evaporation and maintaining the pH of the culture.
Tampere University of Technology, Biomedical Sciences and Engineering, BioMediTech, Korkeakoulunkatu 3, FI-33720, Tampere, Finland. Electronic address: joose.kreutzer@tut.fi.
Joose
J
Kreutzer
University of Tampere, Neuro-Group, BioMediTech, Lääkärinkatu 1, FI-33520, Tampere, Finland. Electronic address: laura.yla-outinen@uta.fi.
Laura
L
Ylä-Outinen
Tampere University of Technology, Biomedical Sciences and Engineering, BioMediTech, Korkeakoulunkatu 3, FI-33720, Tampere, Finland. Electronic address: antti-juhana.maki@tut.fi.
Antti-Juhana
AJ
Mäki
University of Tampere, Neuro-Group, BioMediTech, Lääkärinkatu 1, FI-33520, Tampere, Finland. Electronic address: mervi.ristola@uta.fi.
Mervi
M
Ristola
University of Tampere, Neuro-Group, BioMediTech, Lääkärinkatu 1, FI-33520, Tampere, Finland. Electronic address: susanna.narkilahti@uta.fi.
Susanna
S
Narkilahti
Tampere University of Technology, Biomedical Sciences and Engineering, BioMediTech, Korkeakoulunkatu 3, FI-33720, Tampere, Finland. Electronic address: pasi.kallio@tut.fi.
Pasi
P
Kallio
D002245:Carbon Dioxide / Q000008:administration & dosage; D018929:Cell Culture Techniques / Q000295:instrumentation*; D002478:Cells, Cultured; D003198:Computer Simulation; D053595:Embryonic Stem Cells / Q000502:physiology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007185:Incubators; D008839:Microelectrodes; D008959:Models, Neurological; D058953:Neural Stem Cells / Q000502:physiology; D009474:Neurons / Q000502:physiology*; D009994:Osmolar Concentration; D013997:Time Factors
D002245:Carbon Dioxide
Cell culture environment; Human embryonic stem cells; Long-term culture; MEA; Neuronal cells; Osmolarity; PDMS; pH