Figure 5. 2-APB (5 μM) enhances sustained elevation of [Ca²⁺]_i in the midpiece.

(a and b) Progesterone-induced responses at the midpiece under control conditions (a) and after application of 5 μM 2-APB (first arrow; b). Each plot shows six to nine representative single-cell traces and ΔF_mean (○-○) for all 25 (a) and 31 (b) cells in the experiment. (c) Dose-dependence of the effect of 2-APB pre-treatment on the sustained [Ca²⁺]_i signal. The amplitude of the sustained response (240 s after progesterone addition) in PHN (left-hand bars) and midpiece (right-hand bars) after exposure to 5 μM 2-APB (light grey bars) and 100 μM 2-APB (dark grey bars) was normalized to the amplitude of the parallel control (shown by a broken line). Each bar shows the means±S.E.M. of 11 (5 μM 2-APB) and six (100 μM 2-APB) experiments. *P<0.05; **P<0.005 compared with control. (d) Amplitude distribution of single-cell sustained [Ca²⁺]_i increases (240 s after progesterone addition). Open circles (○-○) show responses of 2-APB pre-treated cells (136 cells from five experiments), closed circles (●-●) show responses from 135 cells in the five parallel control experiments. (e) Progesterone-stimulated [Ca²⁺]_i elevation in the PHN (grey trace) and midpiece (black trace) of a 5 μM 2-APB pre-treated cell. 2-APB was added at the first arrow, 3 μM progesterone at the second arrow. An inflection in the rising phase of the midpiece trace occurs ~20 s after application of progesterone (arrowhead). (f) Progesterone-stimulated [Ca²⁺]_i elevation in the anterior flagellum (dark grey), PHN (light grey) and midpiece (black) of a 5 μM 2-APB pre-treated cell imaged at 10 Hz. Progesterone was applied at 7 s (arrow). The anterior flagellar response precedes responses in the other two compartments and the rising phase of the midpiece response shows an inflexion ~25 s after onset. (g) Image series of the same cell as (f), showing the delayed [Ca²⁺]_i rise in the midpiece. Numbers show time in seconds. Progesterone was applied at 7 s.