Fig. 5.

Involvement of PKA and Trk receptor in BDNF-induced Ser916/898 phosphorylation of RasGRF1. (A, B) Cortical neurons at 3 DIV were pretreated with 10 μM H-89 (A) or 100 nM K252a (B) for 45 min, and then treated with 50 ng/mL of BDNF for 30 min. Phosphorylated RasGRF1 (Ser916/898) was analyzed by immunoblotting with the anti-pSer916/898 antibody. (C, D) Densitometry analysis was performed, and p-RasGRF1/total RasGRF1 ratio was determined. Relative levels of p-RasGRF1/total RasGRF1 compared to vehicle-treated cells are shown. Data are presented as the means ± SEM of four independent preparations, *p < 0.05; **P < 0.01; ***P < 0.001; ns, not significantly different. (E) Cortical neurons at 3 DIV were treated with 50 ng/mL of BDNF for 15 min with or without pretreatment with 100 nM K252a for 45 min. Then, the intracellular cAMP concentration was measured using the Cyclic AMP XP assay kit. Relative cAMP levels compared to vehicle-treated cells are shown. Data are presented as the means ± SEM of four independent preparations, ns, not significantly different.