Neural Plasticity / 2011 / Article / Fig 2

Research Article

Diacylglycerol Signaling Underlies Astrocytic ATP Release

Figure 2

OAG-dependent calcium oscillations in cultured astrocytes are a result of ATP release. (a) During the first imaging period ( 𝑑 1 ) a baseline response to 100 μM OAG is established in astrocyte cultures. Data are shown as individual cell traces of 𝐹 / 𝐹 0 (fold change) over time (minutes). Blocking astrocytic ATP receptors during the second imaging period ( 𝑑 2 ) with a combination of the P 2 receptor inhibitors PPADS and Suramin greatly reduces the response to 100 μM OAG. This blockade is reversible, after washout the response to 100 μM is reestablished in the third imaging period ( 𝑑 3 ). Diagonal lines between imaging session indicate a wash period of 15 minutes occurring between imaging sessions. (b) Addition of the enzyme apyrase, which degrades ATP to ADP, and ADP to AMP, during 𝑑 2 abolishes the astrocytic response to 100 μM OAG. (c) Quantification demonstrates a significant decrease in the percentage of astrocytes responding with calcium oscillations to OAG stimulation with either the blockade of ATP receptors (PPADS/Suramin; control 𝑛 = 5 , PPADS/Suramin 𝑛 = 8 ; 𝑑 2 𝑃 = 0 . 0 2 ) or the degradation of extracellular ATP (Apyrase; control 𝑛 = 5 , apyrase 𝑛 = 4 ; 𝑑 2 𝑃 = 0 . 0 2 ). (d) ATP assays performed with the luciferin-luciferase reaction demonstrate the release of ATP from cultured astrocytes treated with OAG, compared to controls with DMSO added (DMSO 𝑛 = 2 9 , OAG = 2 7 , 𝑃 = 0 . 0 0 1 ).
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