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SHR Neuro Krebs Kardio Lipid

Eroglu, E; Rost, R; Bischof, H; Blass, S; Schreilechner, A; Gottschalk, B; Depaoli, MR; Klec, C; Charoensin, S; Madreiter-Sokolowski, CT; Ramadani, J; Waldeck-Weiermair, M; Graier, WF; Malli, R.
Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells.
J Vis Exp. 2017; 174(121): [OPEN ACCESS]
PubMed PUBMED Central FullText FullText_MUG

 

Autor/innen der Med Uni Graz:
Bischof Helmut
Blass Sandra
Depaoli Maria Rosa
Eroglu Emrah
Gottschalk Benjamin
Graier Wolfgang
Klec Christiane
Madreiter-Sokolowski Corina
Malli Roland
Ramadani Jeta
Rost René
Waldeck-Weiermair Markus
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Abstract:
Nitric Oxide (NO•) is a small radical, which mediates multiple important cellular functions in mammals, bacteria and plants. Despite the existence of a large number of methods for detecting NO• in vivo and in vitro, the real-time monitoring of NO• at the single-cell level is very challenging. The physiological or pathological effects of NO• are determined by the actual concentration and dwell time of this radical. Accordingly, methods that allow the single-cell detection of NO• are highly desirable. Recently, we expanded the pallet of NO• indicators by introducing single fluorescent protein-based genetically encoded nitric oxide (NO•) probes (geNOps) that directly respond to cellular NO• fluctuations and, hence, addresses this need. Here we demonstrate the usage of geNOps to assess intracellular NO• signals in response to two different chemical NO•-liberating molecules. Our results also confirm that freshly prepared 3-(2-hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-propanamine (NOC-7) has a much higher potential to evoke change in intracellular NO• levels as compared with the inorganic NO• donor sodium nitroprusside (SNP). Furthermore, dual-color live-cell imaging using the green geNOps (G-geNOp) and the chemical Ca(2+) indicator fura-2 was performed to visualize the tight regulation of Ca(2+)-dependent NO• formation in single endothelial cells. These representative experiments demonstrate that geNOps are suitable tools to investigate the real-time generation and degradation of single-cell NO• signals in diverse experimental setups.

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