Fig. 3. High potassium- and NGF-mediated BDNF secretion from nnr5-TrkA, nnr5-Y794F and nnr5-Re794Y. Cells were perfused at a flow rate of 0.1 ml/min. Fractions were collected every 5 min. After collecting two fractions, a first stimulation by 50 mM KCl was initiated and four additional fractions collected. After a recovery of 30 min, cells were stimulated by 100 ng/ml NGF. BDNF concentrations were measured by two-site ELISA. Increased BDNF concentrations (expressed in pg/ml) occurred immediately after stimulation. The peak concentration occurred in the following fraction. The 5 min delay between the onset of stimulation and the peak of BDNF secretion is due to the dead volume of the perfusion system. NGF induced BDNF secretion in nnr5-TrkA and nnr5-Re794Y cells, but not in nnr5 or nnr5-Y794F cells. The values given represent the mean ± SE (n = 6).