Antigen-mediated stimulation of hydrolysis of inositol phospholipids in RBL-2H3 cells is dependent of the extent are rate of aggregation of the plasma membrane receptors for IgE. The hydrolysis is associated with an increase in concentration of free Ca2+ (Ca2+)i and histamine secretion. The pattern of phosphoinositide response and dependency on external calcium varies markedly with the type of stimulatory legand used. With the exception of aggregated ovalbumin some stimulation of hydrolysis of inositol phospholipids and small increases in (Ca2+)i were induced by DNP24BSA or higher oligomers of IgE. Whatever receptor aggregation system is employed, the mobilization of intracellular calcium is an insufficient stimulus for secretion. All stimulants at optimal concentrations produced comparable responses in the presence of calcium. The addition of calcium to the medium markedly amplifies the phosphoinositide response and causes the release of histamine. Separation of the inositol phosphate by HPLC has shown that upon stimulation with antigen at least eleven inositol phosphate are produced. The metabolic pathway involved in the production of the various inositol phosphates, as determined by studies with cell extracts, was as follows: Inositol 1,4,5-trisphosphate is converted to inositol 1,4- bisphosphate (subsequently to inositol 1 monophosphate) and to inositol 1,3,4,5-tetrakisphosphate. Inositol 1,3,4,5- tetrakisphosphate was converted back to inositol, 1,4,5- trisphospate and to inositol 1,3,4-trisphosphate which, in turn, was degraded to inositol 3,4-bisphosphate and to a lesser extent inositol 1,3-bisphosphate. These were subsequently degraded to inositol 1- or 4-monophosphate. In studied with patch clamped single cells, both inositol 1,4,5-trisphosphate and inositol 1,3,4,5- tetrakisphosphate when injected into cells activated the same kind of ion-conducting channels as those observed with antigen stimulation.
