Figure 3. BDNF inhibition of protein rephosphorylation by GSK3 arrests dextran uptake.

(a) CGNs or (c) CGNs transfected with either wild-type (DynI_WT) or S774A dynamin I mutant (DynI_S774A) were placed in incubation medium for 10 min before stimulation with a train of 800 action potentials (80 Hz) in the presence of 50 μM tetramethylrhodamine dextran. (b) CGNs or (d) CGNs transfected with either wild-type (DynI_WT) or S774A dynamin I mutant (DynI_S774A) were placed in incubation medium for 10 min before a priming stimulus of 50 mM KCl (1 min). CGNs were then repolarized for 10 min before stimulation with 800 action potentials (80 Hz) in the presence of 50 μM tetramethylrhodamine dextran. BDNF (100 ng ml⁻¹), LY294002 (LY, 10 μM) or CT99021 (CT, 2 μM) were present throughout the experiment where indicated. Dextran was perfused away immediately after stimulation in both cases. (a,b) The number of dextran puncta expressed as a percentage of control corrected for the unstimulated background±s.e.m., (a) Ctrl and BDNF n=5; Student’s t-test P=0.69, (b) Ctrl n=10, BDNF n=4, BDNF/LY n=4, LY n=7, BDNF/CT n=3, CT n=3 one-way analysis of variance (ANOVA). *P<0.05, **P<0.01 compared with control. (c,d) The average number of dextran puncta per 100 μm of transfected neuron corrected for unstimulated background ±s.e.m., (c) both n=3; Student’s t-test P=0.006, (d) DynI_WT n=5, all others n=3, one-way ANOVA. ***P<0.001 compared with DynI_WT.