Figure 5.
N-type-specific slots are filled to capacity by endogenous N-type channels. A, Relative contributions of N-, P/Q-, and R-type channels to EPSC amplitude in neurons expressing EGFP alone (EGFP, n = 20), or EGFP with human CaV2 subunits (NWT, n = 23; NWT+PQWT, n = 11). The N-, P/Q-, or R-type Ca2+ channel blockers ω-CTx-GVIA (2 μm), ω-Aga-IVA (1 μm), and SNX482 (0.5 μm) were applied sequentially and cumulatively into the bath solution. In the presence of extra PQWT, overexpression of NWT could no longer increase the N contribution to synaptic transmission, indicating that N- and P/Q-type channels compete for a common subset of slots (*p < 0.05, NWT vs EGFP group; no significant difference between NWT+PQWT and EGFP groups). B, Cumulative distribution of overall EPSC amplitude at synapses expressing EGFP alone (n = 70), or EGFP with human CaV2 subunits (NWT, n = 61; NWT+PQWT, n = 18). The synaptic strength was comparable across all groups (p > 0.33, Kolmogorov–Smirnov test). C, Peak whole-cell Ba2+ current density through P/Q-type Ca2+ channels at +10 mV from hippocampal neurons expressing EGFP alone (EGFP, n = 19), or EGFP with human CaV2 subunits (NWT, n = 10; NWT+PQWT, n = 7). Overexpression of PQWT, either alone or with NWT, resulted in significant increase of whole-cell P/Q current density (***p < 0.001, relative to EGFP group). The magnitude of P/Q-current density was not significantly different between NWT+PQWT (50.1 ± 3.8 pA/pF) and the PQWT groups (58.0 ± 13.3 pA/pF) (Fig. 2 B) (p > 0.5). Overexpressing NWT in neurons did not alter P/Q-type current density. D, Peak whole-cell Ba2+ current density through N-type Ca2+ channels at +10 mV from the same neurons in C. Overexpression of NWT, either alone or with PQWT, significant increased the whole-cell N-type current density (**p < 0.01, relative to EGFP group). The magnitude of N-type current density was comparable between the NWT (32.8 ± 3.3 pA/pF) and the NWT+PQWT (35.1 ± 4.7 pA/pF; p > 0.6) groups. Error bars indicate SEM. E, Different relationship of N- and P/Q-type Ca2+ channels to slots in controlling neurotransmission at cultured hippocampal synapses. Our data indicate that there are at least two different kinds of presynaptic slots at excitatory hippocampal synapses. The N specific slots accept N-type channels selectively but reject P/Q-type Ca2+ channels. The PQ preferring slots prefer P/Q-type Ca2+ channels but also accept N-type when P/Q-type channels are missing or N-type are overexpressed. Note that terminology for slots does not consider R-type channels.