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13/10/2014 (Added to site)
Author(s): Tolstykh G. P.; Beier H. T.; Roth, C. C.; Thompson, G. L.; Ibey, B. L.

600 ns pulse electric field-induced phosphatidylinositol_4,5- bisphosphate depletion

Journal: Bioelectrochemistry, 100/1 (2014), pp. 80-87
DOI: 10.1016/j.bioelechem.2014.01.006
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Abstract: The interaction between nsPEF-induced Ca2 + release and nsPEF-induced phosphatidylinositol4,5-bisphosphate (PIP2) hydrolysis is not well understood. To better understand this interrelation we monitored intracellular calcium changes, in cells loaded with Calcium Green-1 AM, and generation of PIP2 hydrolysis byproducts (inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG)) in cells transfected with one of two fluorescent reporter genes: PLCδ-PH-EGFP or GFP-C1-PKCγ-C1a. The percentage fluorescence differences (ΔF %) after exposures were determined. Upon nsPEF impact, we found that in the absence of extracellular Ca2 + the population of IP3 liberated during nsPEF exposure (ΔF 6% ± 3, n = 22), is diminished compared to the response in the presence of calcium (ΔF 84% ± 15, n = 20). The production of DAG in the absence of extracellular Ca2 + (ΔF 29% ± 5, n = 25), as well as in cells exposed to thapsigargin (ΔF 40% ± 12, n = 15), was not statistically different from cells exposed in the presence of extracellular calcium (ΔF 22 ± 6%, n = 18). This finding suggests that the change in intracellular calcium concentration is not solely driving the observed response. Interestingly, the DAG produced in the absence of Ca2 + is the strongest near the membrane regions facing the electrodes, whereas the presence of extracellular Ca2 + leads to a whole cell response. The reported observations of Ca2 + dynamics combined with IP3 and DAG production suggest that nsPEF may cause a direct effect on the phospholipids within the plasma membrane.


Keywords: calcium    electric pulse    nanopore    Phospholipase C    PIP2 depletion   

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