Figure 5. The G-protein action does not involve activation of heterotrimeric G-proteins.

A, AlF₄⁻, a potent activator of heterotrimeric G-proteins, stimulated [Ca²⁺]_i oscillation which was accompanied by Ca²⁺-dependent exocytosis. The pipette solution contained AlF₄⁻ (100 μM) and 100 μM indo-1 but no BAPTA. B, when the intracellular Ca²⁺ store was depleted, AlF₄⁻ failed to trigger any exocytosis. To deplete the intracellular Ca²⁺ store, the cell was bathed continuously in a solution containing 1 mM EGTA (no added Ca²⁺) plus the SERCA inhibitor BHQ (1 μM). It was then challenged with GnRH (100 nM) for 3 min. About 5 min following the removal of GnRH, the whole-cell recording began. Note that [Ca²⁺]_i remained near 100-200 nM throughout the time course of the recording, suggesting that most of the intracellular Ca²⁺ store was depleted by this experimental manipulation. Same pipette solution as in A. C, the depletion of the intracellular Ca²⁺ store does not affect the GTP-γ-S triggered Ca²⁺-independent exocytosis. The cell was subjected to the same experimental procedure as described in B. Intracellular dialysis of GTP-γ-S still triggered robust exocytosis. The pipette solution contained 500 μM GTP-γ-S and 10 mM BAPTA. Note that in both B and C, the cells were continuously bathed in a Ca²⁺-free solution containing BHQ (1 μM).