Electrophysiological variability in the SH-SY5Y cellular line

Santillo, Silvia; Moriello, Aniello Schiano; Maio, Vito Di

doi:10.4149/gpb_2013071

Cited by 23 publications

(25 citation statements)

References 9 publications

(15 reference statements)

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“…Differentiated and non‐differentiated cells varied in terms of current amplitudes and action potential shapes. Non‐differentiated cells do not show spontaneous activity and can only be depolarized when the membrane potential is clamped to −90 mV (Santillo, Schiano Moriello, & Di Maio, 2014). Upon differentiation, cells become more excitable, fire spontaneous action potentials (Toselli, Tosetti, & Taglietti, 1996), and become mechanically stiffer as an indication of more ion channels on the membrane (Halitzchi et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

Substrate stiffness effects on SH‐SY5Y: The dichotomy of morphology and neuronal behavior

et al. 2020

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Like many other cell types, neuroblastoma cells are also known to respond to mechanical cues in their microenvironment in vitro. They were shown to have mechanotransduction pathways, which result in enhanced neuronal morphology on stiff substrates. However, in previous studies, the differentiation process was monitored only by morphological parameters. Motivated by the lack of comprehensive studies that investigate the effects of mechanical cues on neuroblastoma differentiation, we used SH-SY5Y cells differentiated on polyacrylamide (PA) gels as a model. Cells differentiated on the surface of PA hydrogels with three different elastic moduli (0.1, 1, and 50 kPa) were morphologically evaluated and their electrophysiological responsiveness was probed using calcium imaging. Immunodetection of neural marker TUJ1 and p-FAK was used for biochemical characterization. Groups with defined stiffness that are matching and nonmatching to neural tissue extracellular matrix were used to distinguish biomimetic results from other effects. Results show that while cells display morphologies that do not resemble neurons on soft substrates, they are in fact electrophysiologically more responsive and abundant in neuronal marker TUJ1. Our findings suggest that while neuronal differentiation occurs more efficiently in microenvironments mechanically mimicking neural tissue, the SH-SY5Y model demonstrates morphologies that conflict with neuronal behavior under these conditions. These results are expected to contribute considerable input to researchers that use SH-SY5Y as a neuron model.

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Section: Introductionmentioning

confidence: 99%

Substrate stiffness effects on SH‐SY5Y: The dichotomy of morphology and neuronal behavior

et al. 2020

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“…The American Journal of Human Genetics 108, 1-10, April 1, 2021 3 KO#2, and KO#9 lines for 7 days with RA, allowing for neurite formation ( Figure S3). 22 Image-based analysis revealed that neurites were formed in all three lines ( Figure 2B). We then estimated the average neurite length in each line by counting intersections between neurites and test lines of a superimposed frame of fixed size 23 applied on neuronal cultures as described previously.…”

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confidence: 97%

“…When stimulated with retinoic acid (RA), wild-type (WT) SH-SY5Y cells acquire neuron-like phenotypes such as neurite formation, electrical excitability, and expression of neurotransmitters and neurotransmitter receptors. 22 We targeted NCDN in WT SH-SY5Y cells by using CRISPR/Cas9 and expanded two independent clones with distinct homozygous NCDN deletions, one spanning 16 bp (c.1458_1473del16 [GenBank: NM_014284.3]; SH-SY5Y NCDND16/D16 ), assigned KO#2, and one spanning 1,158 bp; (chr1: 36,027,723-36,028,880; SH-SY5Y NCDND1158/D1158 ), assigned KO#9. The 16 bp deletion (KO#2) is located within exon 5 of NCDN and the 1,158 bp deletion (KO#9) spans intron 3 to exon 5 ( Figure S1).…”

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confidence: 99%

Monoallelic and bi-allelic variants in NCDN cause neurodevelopmental delay, intellectual disability, and epilepsy

Fatima

Hoeber

Schuster

et al. 2021

The American Journal of Human Genetics

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Neurochondrin (NCDN) is a cytoplasmatic neural protein of importance for neural growth, glutamate receptor (mGluR) signaling, and synaptic plasticity. Conditional loss of Ncdn in mice neural tissue causes depressive-like behaviors, impaired spatial learning, and epileptic seizures. We report on NCDN missense variants in six affected individuals with variable degrees of developmental delay, intellectual disability (ID), and seizures. Three siblings were found homozygous for a NCDN missense variant, whereas another three unrelated individuals carried different de novo missense variants in NCDN. We assayed the missense variants for their capability to rescue impaired neurite formation in human neuroblastoma (SH-SY5Y) cells depleted of NCDN. Overexpression of wild-type NCDN rescued the neurite-phenotype in contrast to expression of NCDN containing the variants of affected individuals. Two missense variants, associated with severe neurodevelopmental features and epilepsy, were unable to restore mGluR5-induced ERK phosphorylation. Electrophysiological analysis of SH-SY5Y cells depleted of NCDN exhibited altered membrane potential and impaired action potentials at repolarization, suggesting NCDN to be required for normal biophysical properties. Using available transcriptome data from human fetal cortex, we show that NCDN is highly expressed in maturing excitatory neurons. In combination, our data provide evidence that bi-allelic and de novo variants in NCDN cause a clinically variable form of neurodevelopmental delay and epilepsy, highlighting a critical role for NCDN in human brain development.

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“…'Maximal conductance' determines the magnitude of the current that would flow if all the channels were open, a value proportional to the number of channels in the membrane. High levels of variability in kinetics have been observed in many studies Bekkers et al (1990); Finkel et al (2006); Feigenspan et al (2010); Golowasch (2014); Santillo et al (2014); Altomare et al (2015); Annecchino and Schultz (2018). A previous study Lei et al (2019b) analysed hERG current kinetics recorded simultaneously in 124 cells using an automated high-throughput patch-clamp machine.…”

Section: Introductionmentioning

confidence: 95%

Accounting for variability in ion current recordings using a mathematical model of artefacts in voltage-clamp experiments

Lei

Clerx

Whittaker

et al. 2019

Preprint

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Mathematical models of ion channels, which constitute indispensable components of action potential models, are commonly constructed by fitting to whole-cell patch-clamp data. In a previous study we fitted cell-specific models to hERG1a (Kv11.1) recordings simultaneously measured using an automated high-throughput system, and studied cell-cell variability by inspecting the resulting model parameters. However, the origin of the observed variability was not identified. Here we study the source of variability by constructing a model that describes not just ion current dynamics, but the entire voltage-clamp experiment. The experimental artefact components of the model include: series resistance, membrane and pipette capacitance, voltage offsets, imperfect compensations made by the amplifier for these phenomena, and leak current. In this model, variability in the observations can be explained by either cell properties, measurement artefacts, or both. Remarkably, by assuming that variability arises exclusively from measurement artefacts, it is possible to explain a larger amount of the observed variability than when assuming cell-specific ion current kinetics. This assumption also leads to a smaller number of model parameters. This result suggests that most of the observed variability in patch-clamp data measured under the same conditions is caused by experimental artefacts, and hence can be compensated for in post-processing by using our model for the patch-clamp experiment. This study has implications for the question of the extent to which cell-cell variability in ion channel kinetics exists, and opens up routes for better correction of artefacts in patch-clamp data.

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Electrophysiological variability in the SH-SY5Y cellular line

Cited by 23 publications

References 9 publications

Substrate stiffness effects on SH‐SY5Y: The dichotomy of morphology and neuronal behavior

Substrate stiffness effects on SH‐SY5Y: The dichotomy of morphology and neuronal behavior

Monoallelic and bi-allelic variants in NCDN cause neurodevelopmental delay, intellectual disability, and epilepsy

Accounting for variability in ion current recordings using a mathematical model of artefacts in voltage-clamp experiments

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