Evaluation of fluorescent dyes for measuring intracellular glutathione content in primary cultures of human neurons and neuroblastoma SH‐SY5Y

  title={Evaluation of fluorescent dyes for measuring intracellular glutathione content in primary cultures of human neurons and neuroblastoma SH‐SY5Y},
  author={Jordi Ponce Sebasti{\`a} and Rosa M. Crist{\`o}fol and Manuela Mart{\'i}n and Eduard Rodr{\'i}guez-Farr{\'e} and Coral Sanfeliu},
  journal={Cytometry Part A},
Reduced glutathione (GSH) protects cells against oxidative injury and maintains a range of vital functions. To study GSH content in human neuronal cell cultures, thiol‐sensitive fluorescent techniques requiring a small number of cells may be of great value, but their GSH specificity has not been established in these cells. 

High Throughput Method for Assessment of Cellular Reduced Glutathione in Mammalian Cells.

This study provides a fast, simple method for the high throughput screening of GSH in a widely available 96-well format and addresses the pitfalls associated with fluorescent compounds in cell culture and proper data normalization.

Probing Cell Redox State and Glutathione-Modulating Factors Using a Monochlorobimane-Based Microplate Assay

A microplate assay with monochlorobimane (MCB) as an available fluorescent probe for GSH, although poorly detected in the microplate format is proposed, providing a useful methodology for the evaluation of cell redox status probed through relative GSH content and responsiveness to both supplemented thiols and variation in oxygen pressure.

Functional Induction of the Cystine-Glutamate Exchanger System Xc - Activity in SH-SY5Y Cells by Unconjugated Bilirubin

It is suggested that bilirubin can modulate the gluthathione levels in neuroblastoma cells through the induction of the System Xc −, and this renders the cell less prone to oxidative damage.

Determination of glutathione and glutathione disulfide in biological samples: an in-depth review.

γ-Tocotrienol does not substantially protect DS neurons from hydrogen peroxide-induced oxidative injury

It is suggested that γT3 pre-treatment are not sufficient to protect DS neurons from H2O2-induced oxidative assault, instead induced the apoptosis process.

Loss of glutathione homeostasis associated with neuronal senescence facilitates TRPM2 channel activation in cultured hippocampal pyramidal neurons

GSH plays a physiologically relevant role in the regulation of TRPM2 currents in hippocampal pyramidal neurons, and this interaction may play an important role in aging and neurological diseases associated with depletion of GSH.

Recent developments in methods intracellulary localizing glutathione within plant tissues and cells (a minireview)

A specific antibody, which recognizes t-GSH, was used in order to demonstrate the distribution of glutathione in different cell compartments for light and electron microscopical investigations.



Characterization of Hydrogen Peroxide Toxicity in Cultured Rat Forebrain Neurons

It is demonstrated that H2O2 is a potent and effective neurotoxin that produces oxidative stress, as well as apoptotic neuronal death, in cultured rat forebrain neurons.

Glutathione levels in primary glial cultures: Monochlorobimane provides evidence of cell type‐specific distribution

It seems that owing to the greater intracellular concentration of reactive oxygen and nitrogen species to which microglia are subjected, especially under conditions of inflammation, this cell type is fortified with higher GSH levels as a means to combat oxidative and nitrosative stress.

Evaluation of glutathione‐sensitive fluorescent dyes in cortical culture

Neurons experienced a dramatic decline in GSH levels relative to astrocytes between 5–6 DIV, as shown by a loss of neuronal staining with CMAC, CMAC‐blue and MCB, indicating specificity for GSH.

Direct evidence for glutathione as mediator of apoptosis in neuronal cells.

Synthesis of the Antioxidant Glutathione in Neurons: Supply by Astrocytes of CysGly as Precursor for Neuronal Glutathione

The data suggest the following metabolic interaction in glutathione metabolism of brain cells: the ectoenzyme γ-glutamyl transpeptidase uses as substrate the glutATHione released by astrocytes to generate the dipeptide CysGly that is subsequently used by neurons as precursor for glutathion synthesis.

Quantitative analysis of cellular glutathione by flow cytometry utilizing monochlorobimane: some applications to radiation and drug resistance in vitro and in vivo.

Heterogeneity in populational GSH levels was observed under certain circumstances, such as release from hypoxia, and the significance of this heterogeneity is discussed in regard to the induction of gene amplification and drug resistance by transient Hypoxia.

Evaluation of methods for measuring cellular glutathione content using flow cytometry.

It is recommended that flow cytometric GSH assays should routinely include a sample of cells that have been depleted of GSH in order to determine the extent of background labeling, and monobromobimane is the agent of choice.

Metabolism and functions of glutathione in brain

Glutathione metabolism in brain

The main focus of this short review is recent results on glutathione metabolism of brain astrocytes and neurons in culture, which show that these two types of cell prefer different extracellular precursors for glutATHione.