Figure 7.

ProNGF-Δ9-13 at axon terminals induces greater apoptotic signaling at cell bodies than proNGF-KKE. ProNGF-Δ9-13, proNGF-KKE, or inactivated proNGF were applied to rat BFCN axon terminals for 15 min, followed by quantification of signaling factor activation at BFCN cell bodies. ProNGF-Δ9-13 (A) caused greater activation (phosphorylation) of the pro-apoptotic signaling factor, JNK, relative to proNGF-KKE (B) and inactivated proNGF (C). pJNK is shown in red, total JNK in green, and DAPI in blue. DIV, days in vitro. (D) Quantification of the images shown in A–C. Error bars represent SEM, n = 30 images/group taken from three chambers in three independent experiments. One-way ANOVA and post hoc Tukey test, *p < 0.05, ****p < 0.0001. (E) ProNGF-Δ9-13 or proNGF-KKE were applied to mouse p75^NTR exon III knockout BFCN axon terminals for 15 min, followed by quantification of JNK activation at BFCN cell bodies. Activation of the pro-apoptotic signaling factor, JNK, induced by proNGF-Δ9-13 was not different from that induced by proNGF-KKE, Student’s t-test, p > 0.05. Error bars represent SEM, n = 15 images/group taken from one chamber. (F) Immunocytochemistry was conducted on DIV8 rat BFCNs as described in Materials and Methods, with the elimination of primary antibodies. Minimal fluorescent signal was detected for each secondary antibody under these conditions. Alexa Fluor goat anti-rabbit 488 shown in green, Alexa Fluor goat anti-mouse 647 shown in red, DAPI shown in blue. Scale bars: 10 μm.