CNAα and CDK5 regulate AP-stimulated Ca²⁺ influx at nerve terminals. A, Representative images of VAMP2–mCh and its corresponding Fluo5F kymograph image for 1AP stimulation in WT. B, Representative 1AP Fluo5F traces before (black) and after (red) EGTA-AM with CSA (blue) in WT (left) and CNAαKD (right) neurons. Arrow indicates 1AP point. C, Mean value of Fluo5F 1AP response amplitudes before and after EGTA-AM with/without CSA in WT and CNAαKD neurons: WT_{Before 1AP} = 2.92 ± 0.26, WT_{EGTA-AM 1AP} = 1.40 ± 0.17, WT_{EGTA-AM + CSA 1AP} = 0.89 ± 0.10 (n = 8), CNAαKD_{Before 1AP} = 1.38 ± 0.11, CNAαKD_{EGTA-AM 1AP} = 0.73 ± 0.06, and CNAαKD_{EGTA-AM + CSA 1AP} = 0.71 ± 0.09 (n = 9). D, Representative 1AP Fluo5F traces before (black) and after (red) EGTA-AM with roscovitine (Ros) (blue) in WT (left) and CDK5KD (right) neurons. Arrow indicates 1AP point. E, Mean value of Fluo5F 1AP response amplitudes before and after EGTA with/without roscovitine in WT and CDK5KD neurons: WT_{Before 1AP} = 2.76 ± 0.51, WT_{EGTA-AM 1AP} = 1.45 ± 0.26, WT_{EGTA-AM + Ros 1AP} = 1.90 ± 0.33 (n = 7), CDK5KD_{Before 1AP} = 3.81 ± 0.56, CDK5KD_{EGTA-AM 1AP} = 2.32 ± 0.35, and CDK5KD_{EGTA-AM + Ros 1AP} = 2.36 ± 0.37 (n = 8). F, Left, Representative 1AP Fluo5F traces before (black) and after (red) EGTA-AM with Ros and CSA (blue) in WT neurons. Right, Mean value of Fluo5F 1AP response amplitudes before and after EGTA-AM with/without Ros and CSA in WT neurons: WT_{Before 1AP} = 2.73 ± 0.19, WT_EGTA-AM-1AP = 1.57 ± 0.10, and WT_{EGTA-AM + Ros + CSA 1AP} = 1.63 ± 0.23 (n = 5). G, Left, Representative 1AP Fluo5F traces before (black) and after (red) EGTA-AM with Ros (blue) in CNAαKD neurons. Right, Mean value of Fluo5F 1AP response amplitudes before and after EGTA-AM with/without Ros in CNAαKD neurons: CNAαKD_{Before 1AP} = 1.87 ± 0.33, CNAαKD_{EGTA-AM 1AP} = 0.89 ± 0.14, and CNAαKD_{EGTA-AM + Ros 1AP} = 0.97 ± 0.20. (n = 5).