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. 2019 Aug 7;10:1008. doi: 10.3389/fphys.2019.01008

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Copyright © 2019 Xiong, Baker and Pietrantonio.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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BMLK3 preincubation (5 min) with dosages of TKSM14 before agonist addition, enhances its antagonistic activity in the calcium bioluminescence assay. (A) The kinetic responses of BMLK3 cells to the agonist (1 μM FFFSWGa) after being incubated with various concentrations of TKSM14 (1–100 μM) or blank solvent for 5 min. The response was recorded for 30 s at 1 s intervals by the Clariostar^TM plate reader (BMG LABTECH). The kinetic response is represented as the percentage of the maximal bioluminescence units (BU) of the positive control (PC = blank buffer + agonist, blue curve) in each assay (Mean ± SEM, n = 9–10 wells from three independent assays). (B) The total cellular response in each well is expressed as the percentage of the average BU (per second) obtained in the 30 s relative to the average response units to the PC in 30 s. Three independent assays were performed. Responses from each assay were calculated as the average of individual responses from 3-4 pseudo-replicate wells (Y axis: mean ± SEM). Statistical differences in inhibitory activities between different treatment were analyzed by one-way ANOVA (n = 3) followed by Tukey’s multiple comparison test (p ≤ 0.05) with GraphPad 6.0 (GraphPad Software, La Jolla, CA, United States). Bars with different letter superscripts indicate significant differences.