Contribution of 2-arachidonoylglycerol to TRPV1 activation evoked by stimulation of phospholipase C-coupled histamine H₁ receptors in HEK293 cells.

<p>(A) Monoacylglycerol lipase (MAGL) expression in HEK293 cells, as shown by immunocytochemistry. Upper and lower left images show staining with human MAGL antibodies from Abcam and Pierce Biotechnology, respectively. Right hand images were obtained after incubation of the primary antibodies with respective blocking peptide. (B) Histamine (His, 100 µM) increased the content of 2-arachidonoylglycerol (2-AG) in HEK293 cells transiently expressing the phospholipase C-coupled histamine H₁ receptor. Incubation (30 min) with the MAGL inhibitor JZL184 (JZL, 10 µM) further increased the 2-AG content, whereas the diacylglycerol lipase (DAGL) inhibitor tetrahydrolipstatin (THL, 10 µM) prevented the histamine-induced 2-AG increase. The 2-AG content was normalized to the average 2-AG content induced by histamine in vehicle-treated HEK293 cells in each experiment. The results are presented as mean ± s.e.m (n = 6). One-way ANOVA followed by Bonnferroni's <i>post hoc</i> test was used for comparing groups of data. (C) Current-voltage relationships in HEK293 cells transiently co-expressing TRPV1 and the rat histamine H₁ receptor before (red trace) and after (black trace) exposure to histamine 100 µM in the absence and presence of JZL184 (10 µM), THL (10 µM), JZL184 plus THL (each 10 µM) or Cz (10 µM). The control trace shows the average current with error bars obtained from 119 cells. (D) The mean current amplitude elicited by histamine at a test potential of +100 mV in non-treated (n.t.) cells and in cells treated (at least 20 minutes) with enzyme inhibitors. Data are presented as mean ± s.e.m (n = 6–8). Kruskal-Wallis one-way ANOVA followed by Dunn's <i>post hoc</i> test was used for comparing groups of data. **<i>P</i><0.01, ***<i>P</i><0.001.