Fig. 2.

S1P, LPA, and thrombin induce InsP production through PLCε. Cultured WT and PLCε KO astrocytes were serum-starved overnight in the presence of [³H]inositol. (A) Cells were treated for 30 min with vehicle (BSA), 5 μM S1P, 10 μM LPA, 0.5 unit/ml thrombin, or 500 μM carbachol in the presence of LiCl before isolation of [³H]InsPs. Data are reported as cpm of agonist-stimulated InsP production (cpm of agonists − cpm of vehicle). Cpm of vehicle were 290 ± 26 and 183 ± 10 for WT and KO cells, respectively. Data are averaged from five experiments performed in triplicate on at least two different astrocyte culture preparations and are presented as means ± SEM. †, P < 0.001, one-way ANOVA. (B) Time course of thrombin- and carbachol-induced InsP production. Cultured WT (■) and KO (▴) astrocytes were serum-starved overnight in DMEM without FBS containing [³H]inositol and challenged with 0.5 unit/ml thrombin or 500 μM carbachol in the presence of LiCl for the indicated times before isolation of [³H]InsP. Data are reported as cpm recovered per well in triplicate and are averages from two independent experiments.