Minimized cell usage for stem cell-derived and primary cells on an automated patch clamp system

Becker, Norbert; Stoelzle, Sonja; Göpel, Sven; Guinot, David; Mumm, Patrick; Haarmann, Claudia; Malan, Daniela; Bohlen, Heribert; Kossolov, Eugen; Kettenhofen, Ralf; George, Michael; Fertig, Niels; Brüggemann, Andrea

doi:10.1016/j.vascn.2013.03.009

Cited by 47 publications

(22 citation statements)

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“…By reducing the complexity of the process, they also make patch clamping accessible to more users, regardless of previous experience in electrophysiology [170]. The use of platforms including IonWorks Quattro [171], PatchXpress [172], Patchliner [173], SynchroPatch [174] and Opatch [175] has been mainstream for the analysis of recombinant cell lines overexpressing single ion channels, and they are now being trialled for use with more complex hPSC-CMs [24], [175], [176], [177] with varying degrees of success (Table 4). …”

Section: Towards Industrial Phenotyping Of Hpsc-cmsmentioning

confidence: 99%

Cardiomyocytes from human pluripotent stem cells: From laboratory curiosity to industrial biomedical platform

Denning

Borgdorff

Crutchley

et al. 2016

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

241

255

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Cardiomyocytes from human pluripotent stem cells (hPSCs-CMs) could revolutionise biomedicine. Global burden of heart failure will soon reach USD $90bn, while unexpected cardiotoxicity underlies 28% of drug withdrawals. Advances in hPSC isolation, Cas9/CRISPR genome engineering and hPSC-CM differentiation have improved patient care, progressed drugs to clinic and opened a new era in safety pharmacology. Nevertheless, predictive cardiotoxicity using hPSC-CMs contrasts from failure to almost total success. Since this likely relates to cell immaturity, efforts are underway to use biochemical and biophysical cues to improve many of the ~ 30 structural and functional properties of hPSC-CMs towards those seen in adult CMs. Other developments needed for widespread hPSC-CM utility include subtype specification, cost reduction of large scale differentiation and elimination of the phenotyping bottleneck. This review will consider these factors in the evolution of hPSC-CM technologies, as well as their integration into high content industrial platforms that assess structure, mitochondrial function, electrophysiology, calcium transients and contractility. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.

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Section: Towards Industrial Phenotyping Of Hpsc-cmsmentioning

confidence: 99%

Cardiomyocytes from human pluripotent stem cells: From laboratory curiosity to industrial biomedical platform

Denning

Borgdorff

Crutchley

et al. 2016

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

241

255

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“…These human neurons are a consistent cell population expressing primarily GABAergic and glutamatergic receptors, but have also been used to measure voltage-dependent Na + and K + channels (Becker, et al, 2013). The relative expression of GABA A R subunits in iCell Neurons was determined by qRT-PCR.…”

Section: Introductionmentioning

confidence: 99%

Characterization of GABA A receptor ligands with automated patch-clamp using human neurons derived from pluripotent stem cells

Yuan

Witzigmann

Cook

et al. 2016

Journal of Pharmacological and Toxicological Methods

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Introduction Automated patch clamp is a recent but widely used technology to assess pre-clinical drug safety. With the availability of human neurons derived from pluripotent stem cells, this technology can be extended to determine CNS effects of drug candidates, especially those acting on the GABAA receptor. Methods iCell Neurons (Cellular Dynamics International, A Fujifilm Company) were cultured for ten days and analyzed by patch clamp in the presence of agonist GABA or in combination with positive allosteric GABAA receptor modulators. Both efficacy and affinity were determined. In addition, mRNA of GABAA receptor subunits were quantified by qRT-PCR. Results We have shown that iCell Neurons are compatible with the IonFlux microfluidic system of the automated patch clamp instrument. Resistance ranging from 15-25 MΩ was achieved for each trap channel of patch clamped cells in a 96-well plate format. GABA induced a robust change of current with an EC50 of 0.43 μM. Positive GABAA receptor modulators diazepam, HZ166, and CW-04-020 exhibited EC50 values of 0.42 μM, 1.56 μM, and 0.23 μM, respectively. The α2/α3/α5 selective compound HZ166-induced the highest potentiation (efficacy) of 810% of the current induced by 100 nM GABA. Quantification of GABAA receptor mRNA in iCell Neurons revealed high levels of α5 and β3 subunits and low levels of α1, which is similar to the configuration in human neonatal brain. Discussion iCell Neurons represent a new cellular model to characterize GABAergic compounds using automated patch clamp. These cells have excellent representation of cellular GABAA receptor distribution that enable determination of total small molecule efficacy and affinity as measured by cell membrane current change.

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“…Recently, iPS neurons (CDI) have been used successfully in both conventional electrophysiology and on the Patchliner. 26,28 In these cells, a voltage-gated Na C channel, K C channel and the ligand-gated ion channel, the GABA A receptor, could be recorded (Fig. 2C, D).…”

mentioning

confidence: 93%

“…2,25 Included in this is the evaluation of the use of pluripotent stem cell applications for cardiovascular risk assessment. So far, the Patchliner has been used successfully to record from mouse embryonic stem cells (CorAt, Axiogenesis), 10 human iPS cells from CDI, 14,26 Axiogenesis (Cor.4U) 26 ), and Cellectis ( Fig. 2A, B).…”

mentioning

confidence: 99%

Novel screening techniques for ion channel targeting drugs

et al. 2015

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Minimized cell usage for stem cell-derived and primary cells on an automated patch clamp system

Cited by 47 publications

References 9 publications

Cardiomyocytes from human pluripotent stem cells: From laboratory curiosity to industrial biomedical platform

Cardiomyocytes from human pluripotent stem cells: From laboratory curiosity to industrial biomedical platform

Characterization of GABA A receptor ligands with automated patch-clamp using human neurons derived from pluripotent stem cells

Novel screening techniques for ion channel targeting drugs

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