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. 2011 Jul 14;6(7):e22246. doi: 10.1371/journal.pone.0022246

Figure 1. HEK293-TRPV1 cells respond to hypertonic stimuli in a temperature-dependent fashion.

Figure 1

(A) Immunostaining of HEK293-TRPV1 and HEK293 cells with anti-TRPV1 antibody. Signals for TRPV1 were predominantly observed at the plasma membrane specifically in HEK293-TRPV1 cells. Bar, 40 µm. (B) Cell response to capsaicin. Representative pseudo-color images of HEK293-TRPV1 cells at 2 min before (upper left) and 1 min after (upper right) a drop of addition of 1 µM capsaicin to the chamber at 36°C. Elevations of the intracellular calcium concentration ([Ca2+]i) are shown by an increase in the ratio of F340/F380 (see color bar). Representative traces of the fluorescence ratio of single cells (lower graph) upon application (arrowhead) of capsaicin; HEK293-TRPV1 (solid line) and control HEK293 (dotted line). Bar, 40 µm. (C) Temperature-dependent sensitivity to osmotic stimuli. Representative pseudo-color images of [Ca2+]i in HEK293-TRPV1 cells at 2 min before (upper left) and 4 min after (upper right) perfusion with a hypertonic solution (350 mOsm) at 36°C. The temperature was maintained throughout the respective recording. Representative single cell traces of the fluorescence ratio at a temperature of 24, 30, 36, or 40°C (lower graph). The line on the top indicates the timing of the change from 300 mOsm (gray) to 350 mOsm (black). The response of HEK293-TRPV1 cells to hypertonic stimuli increased noticeably with the experimental temperature up to 36°C. Bar, 40 µm. (D) Summary of the change in the fluorescence ratio during the perfusion with the hypertonic solution at various temperatures in HEK293-TRPV1 (filled bars) and HEK293 (open bars) cells. Data are differences between fluorescence ratios 2 min before and 4 min after the change of the solution. The maximal sensitivity of the HEK293-TRPV1 cells to the hypertonic stimulation was observed at 36°C. Values are the mean ± SEM. HEK293-TRPV1: n = 147 (24°C), n = 121 (30°C), n = 249 (36°C), n = 106 (40°C). HEK293: n = 110 (24°C), n = 132 (30°C), n = 109 (36°C), n = 101 (40°C).