Article Figures & Data
Figures
- Fig. 1.
Nonlinear relationship between EPSC amplitude and [Ca2+]o. The cooperative involvement of several Ca2+ ions in the release of each vesicle of neurotransmitter is demonstrated by the nonlinear relationship between [Ca2+]o and EPSC amplitude.A, AMPA EPSCs recorded at several different [Ca2+]o. Each trace is the average of 10–50 EPSCs. B, Amplitude of individual EPSCs (same cell as in A) plotted against stimulus number. Each [Ca2+]o is bracketed by an epoch in 2 mm [Ca2+]oto ensure recording stability. C, Average EPSC amplitude plotted against [Ca2+]o on log–log axes. Error bars indicate ±1 SD. The solid line is the optimally fitted Dodge–Rahamimoff equation. The fit was poor and could be rejected (p > 0.05).D, Same data as in C, showing the optimally fitted Hill equation (solid line) with a Hill coefficient of 3.7. This equation provided an adequate fit (p < 0.05). The degree of cooperativity was independently estimated at 3.8 by fitting a power function over the 0.4–1.2 [Ca2+]o range (dashed line).
- Fig. 2.
Cd2+ does not change the steepness of the Ca2+ dose–response curve. The competitive Ca2+ channel antagonist Cd2+ shifts the Ca2+dose–response curve to the right in a dose-dependent manner.A, Linear log plot of the normalized EPSC amplitude versus [Ca2+]o for three individual cells recorded in normal bath solution (filled circles) and in the presence of 2 μmCd2+ (open squares) and 4 μm Cd2+ (open triangles). Each point is the ensemble average amplitude ± 1 SD. The solid lines are Hill equation fits to the data. B, Cd2+increased the EC50 for Ca2+. Average EC50 values are shown in control solution (n = 9) and in the presence of Cd2+ (2 and 4 μm;n = 5). Error bars indicate SEM.C, Cd2+ did not broaden or change the steepness of the dose–response curve. Log–log plot of normalized EPSC amplitude versus [Ca2+]o is shown for the same three cells as in A. Solid linesshow power function fits to the data. D, Cd2+ had no effect on cooperativity. Average cooperativity (NP) over the 0.4–1.2 mm [Ca2+]o range in control solution (n = 9) and 0.8–2 mm[Ca2+]o range in the presence of Cd2+ (2 and 4 μm;n = 5). Error bars indicate SEM.
- Fig. 3.
Dose–response relationships for EPSCs recorded in the presence of ω-CTx and ω-Aga. A, AMPA EPSCs recorded at several different [Ca2+]oin the presence of ω-CTx. Each trace is the average of 5–20 EPSCs. B, Amplitude of individual EPSCs (same cell as in A) plotted against stimulus number. Each [Ca2+]o is bracketed by an epoch in 2 mm [Ca2+]o to ensure recording stability. C, AMPA EPSCs recorded at several different [Ca2+]o in the presence of ω-Aga. Each trace is the average of 5–20 EPSCs.D, Amplitude of individual EPSCs (same cell as in C) plotted against stimulus number.
- Fig. 4.
ω-CTx and ω-Aga broaden the Ca2+ dose–response curve. Selective Ca2+ channel blockers shift the Ca2+ dose–response curve to the right and broaden it. A, Linear log plot of the normalized EPSC amplitude versus [Ca2+]o for three individual cells in normal bath solution (filled circles) and in the presence of ω-CTx (open triangles) and ω-Aga (open squares). Each point is the ensemble average amplitude ± 1 SD. The solid linesare Hill equation fits to the data. B, Log–log plot of normalized EPSC amplitude versus [Ca2+]o for two individual cells recorded in the presence of ω-CTx (open triangles) or ω-Aga (open squares). Each point is the ensemble average amplitude ± 1 SD. The solid linesare power function fits over the [Ca2+]o range from 0.8 to 2 mm. C, The broadening of the dose–response curve by selective toxins reduces the cooperativity (NP). AverageNP is shown in control (n = 9) and in the presence of either ω-CTx (n = 7) or ω-Aga (n = 4). The significant reduction in NP in the presence of selective toxin is consistent with a nonuniform distribution of Ca2+ channel subtypes across presynaptic terminals. Error bars indicate SEM. *Statistical significance (p < 0.05).
- Fig. 5.
A synaptic model predicts the Ca2+ dose–response curve observed in the presence of ω-CTx, ω-Aga, and Cd2+. The modified Dodge–Rahamimoff equation, which incorporates a sublinear relationship between [Ca2+]it and [Ca2+]o, provides an accurate description of the Ca2+ dose–response curve.A, Log–log plot of EPSC amplitude versus [Ca2+]o is shown for an individual cell. The solid line is the optimally fitted modified Dodge–Rahamimoff equation, which represents a good fit (p < 0.05). B, Log–log plot of average normalized EPSC amplitude versus [Ca2+]o under four different experimental conditions: control (filled circles; n = 9), ω-Aga (0.5 μm; filled squares; n= 5), ω-CTx (1 μm; filled triangles;n = 5), and Cd2+ (2 μm; open circles, n = 7). EPSC amplitudes from individual cells were normalized to the amplitude recorded at 2 mm[Ca2+]o in the absence of blockers.C, A model based on the modified Dodge–Rahamimoff equation (see inset and Results) accurately predicted the observed dose–response curves in B. Theoretical dose–response curves are shown in control and in the presence of Ca2+ channel blockers. Inset, Schematic overview of the model. Presynaptic terminals were divided into three classes: one with only P/Q-type Ca2+channels (QQ), one with only N-type Ca2+ channels (NN), and one with both channel subtypes (NQ).
