Figure 4. Blocking Y5R inhibits pro-survival effects of BDNF in NB cells.
A. SY5Y/TrkB cells were cultured for 24h in 1% FBS media and then treated with BDNF (1ng/ml), with or without 1h pre-incubation with Trk and Y5R antagonists (both at 10−6M). Cell viability was measured 24h later by MTS assay. B. SY5Y/TrkB cells were cultured and treated with BDNF and Trk or Y5R antagonists, as above. 2h after BDNF administration, cells were additionally treated with doxorubicin (1μg/ml) or etoposide (2.75 μg/ml). Cell extracts for detection of activated p44/42 MAPK by Western blot were collected 1h later, while cell viability was measured by MTS assay 24h later. C. SK-N-BE(2) cells were cultured as described for panel A, pre-incubated for 1h with Trk or Y5R antagonists at concentrations 10−6M, and then treated with BDNF (1ng/ml) followed by cisplatin (0.5μg/ml). 24h later, cell viability was measured by MTS assay and apoptosis was assessed by caspase 3/7 activity assay.