Figure 5.

Voltage-gated calcium channel (VGCC) subtype contribution to Ca²⁺ transients monitored by synGCaMP6f. (A) Consecutive application of ω-conotoxin GVIA (2 μM; N-type Ca²⁺ channel blocker), ω-agatoxin IVa (0.4 μM; P/Q-type Ca²⁺ channel blocker) and nifedipine (20 μM; L-type Ca²⁺ channel blocker) was employed to evaluate the contribution of different VGCC subtypes to fluorescence changes induced by trains of 10 APs (50 Hz). Color-coded traces show averaged responses from 239 ROIs in nine neurons before (black) and after sequential addition of blockers as indicated. (B) Independent set of experiments (n = 235/11) similar to (A) but with altered order of application and addition of SNX-482 (0.5 μM; R-type Ca²⁺ channel blocker). (C,D) Bar diagrams summarizing the relative contribution of Ca²⁺ channel subtypes from experiments in (A,B). Both diagrams confirm that N- and P/Q-type VGCCs contribute mostly and almost equally to presynaptic calcium transient in hippocampal neurons. (E) Histogram presenting the wide variability of VGCC subtype contributions to Ca²⁺ transients at the level of individual presynaptic boutons (n = 239/9) as evaluated from the recordings in (A,C). A few percent of the boutons even had more than 50% nifedipine-sensitive L-type Ca²⁺ current.