Figure 2. Deoxygedunin activates TrkB and protects neurons from apoptosis.

(A) Deoxygedunin activates TrkB in primary hippocampal neurons. Hippocampal neurons were treated with 500 nM gedunin derivatives for 30 min and neurons were fixed and immunostained with rabbit polyclonal anti-p-TrkB (816) (1∶100) and anti-MAP2. The nuclei were stained with DAPI. BDNF and a few gedunin derivatives selectively triggered TrkB phosphorylation in neurons. (B) Deoxygedunin triggers TrkB activation in primary neurons. Rat cortical neurons were treated with various concentrations of deoxygedunin for 30 min. Neuronal lysates were subjected to immunoblotting analysis by mouse monoclonal anti-p-TrkB (817)(1∶20,000). Equal amount of TrkB was loaded (anti-TrkB from Biovision, 1∶1,000) (lower panel). (C) Deoxygedunin provokes Akt and Erk1/2 activation in primary neurons in a dose and time-dependent manner. Mouse monoclonal anti-TrkB 817 was used at 1∶20,000 dilution. (D) K252a blocks deoxygedunin's agonistic effect on TrkB. Cortical neurons were pretreated with K252a (100 nM) for 30 min, followed by BDNF (100 ng/ml) or deoxygedunin (500 nM) for 30 min. Cell lysates were analyzed by immunoblotting. (E) Deoxygedunin activates TrkB in mouse brain. Deoxygedunin (5 mg/kg) was intraperitoneally injected into mice and mouse brains were dislocated at different time points. Brain lysates were analyzed by immunoblotting.