Fig. 7. DAG partially restores exocytosis from permeabilised mast cells. RPMCs in suspension were permeabilised for the indicated time at pCa 8 and then stimulated as described in Fig. 6B. Unstimulated cells (pCa 7) released 2% of their total ß-hexosaminidase (ß-hex) activity. (A) Cells were permeabilised for 10 minutes in the absence (Ctrl) or presence of 0.3 U/ml PtdIns-PLC, with or without 20 µM Et-18-OMe as indicated. (B) In vitro activity of PtdIns-PLC against [2-³H]-PtdIns in the absence or presence of 20 µM Et-18-OMe. Boiled PtdIns-PLC refers to 0.3 U/ml of the enzyme inactivated at 110°C for 15 minutes. (C) Indicated DAG analogues were applied for 10 minutes before stimulation. (D) Combined effect of PtdIns-PLC and SigD. Mast cells were permeabilised in the presence of 0.3 U/ml PtdIns-PLC and 100 µg/ml SigD for 10 minutes as indicated; refers to the difference in ß-hex release between incubations with and without SigD. (E) Effect of PtdIns-PLC on the IC₅₀ of neomycin. Cells were permeabilised in the presence of the indicated concentration of neomycin with or without 0.3 U/ml PtdIns-PLC for 5 minutes before stimulation. Data are normalised to the levels of secretion observed without neomycin, which were 92±4% of the total cellular ß-hex activity in the presence of PtdIns-PLC, and 72±4% in its absence. Data are means ± s.e.m. (n=3) for stimulated cells (for inactive cells at pCa 7, n=1).