Figure 2.

Inhibition of focal adhesion kinase (FAK) 925 Tyr-P abrogates CB₁-stimulated ERK2 204 Tyr-P in N18TG2 cells. Cells were treated with 0.01 μM WIN55212-2 (WIN) at 37°C for 1, 2, 5, 10, or 20 min (m). (A,B) Cell lysates were analyzed using western blots and representative blot images and analysis of FAK 925 Tyr-P (normalized to total FAK levels) are shown. *p < 0.01 indicates significantly different from basal/time 0 using Student’s t-test. (C–I) Cells were pretreated for 15 m with 10 nM FAK inhibitor [Y11 or PF 573228 (PF)] before treatment with 0.01 μM WIN. Cell lysates were analyzed using western blots and representative blot images and analysis of (C,D) FAK 397 Tyr-P, (E,F) FAK 925 Tyr-P, and (G–I) ERK2 204 Tyr-P (normalized to total FAK or ERK2 levels) are shown. Data are reported as mean ± SEM of (B) the % change over basal FAK 925 Tyr-P, (D,F,H) the % of vehicle-treated FAK 397, FAK 925 and ERK2 204 Tyr-P at the same time point, and (I) the % of basal/time 0 from three separate experiments. For 2C-I, significance was assessed using One Way ANOVA followed by Dunnett’s multiple comparisons posthoc test [*p < 0.01 indicates significantly different from vehicle-treated (at the same time point); **p < 0.05 indicates significantly different from basal/time 0]. For each dataset, cells were cultured and experiments were completed on at least three separate occasions.