Control of inositol polyphosphate-mediated calcium mobilization by arachidonic acid in pancreatic acinar cells of rats.

1. The patch-clamp technique of whole-cell current recording was applied to single, enzymatically isolated, rat pancreatic acinar cells to investigate the current responses evoked by internal perfusion of inositol polyphosphates (InsPx). The InsPx were included in the solution filling the recording pipette and inositol 1,4,5-trisphosphate (Ins(1,4,5)P3; 10 microM) evoked transient current responses generally of less than 1 min duration, inositol 2,4,5-trisphosphate (Ins(2,4,5)P3; 10 microM) evoked smaller current transients while inositol 1,3,4,5-tetrakisphosphate (InsP4; 10 microM) evoked no detectable current response. However, in the presence (in external bathing solution) of the phospholipase A2 inhibitor 4-bromophenacyl bromide (4-BPB; 8 microM) all three of the InsPx now evoked prolonged current responses lasting for several minutes. The current responses to all three InsPx were abolished by inclusion of the Ca2+ chelator EGTA (5 mM) in the internal, pipette-filling solution indicating that the responses are calcium dependent and reflect the effect of the InsPx in increasing intracellular Ca2+. Inositol 1,3,4,5,6-pentophosphate (InsP5) induced no current response when tested up to 20 microM in the presence or absence of 4-BPB. 2. The potentiating effect of 4-BPB on the InsPx-induced current responses was not mimicked by application of arachidonic acid (AA) oxidation inhibitors; indomethacin (20 microM), nordihydroguaiaretic acid (20 microM) or proadifen (SKF525A, 100 microM). The effects of 4-BPB were countered however, by the inclusion of 2 microM AA in the external solution. The results suggest that the 4-BPB potentiates the response by inhibiting the activity of phospholipase A2, thereby reducing the formation of AA. 3. In the presence of 4-BPB (8 microM) the InsPx-evoked responses were dose dependent with an increase in both the amplitude and speed of onset with increasing concentrations. In the presence of 4-BPB InsP4 was as efficient as Ins(1,4,5)P3 both in terms of speed of onset and amplitude of responses; the efficacy and dissociation constant (Kd) for both of these InsPx were the same at 1 microM and 45 nM respectively. Ins(2,4,5)P3 was always less effective, with an efficacy and Kd of 10 microM and 750 nM respectively. 4. If 4-BPB was applied after the current responses evoked by the InsPx were over, or if guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) was included in the recording pipette then the phospholipase inhibitor gave rise to an additional, prolonged, current response.(ABSTRACT TRUNCATED AT 400 WORDS)