D609-phosphatidylcholine-specific phospholipase C inhibitor attenuates thapsigargin-induced sodium influx in human lymphocytes

Previously, we reported that the phosphatidylcholine-specific phospholipase C (PC-PLC) inhibitor tricyclodecan-9-yl xanthogenate (D609) potentiates th apsigargin-induced Ca2+ influx in human lymphocytes. In the present study w e examined the effect of D609 on the thapsigargin-induced Na+ entry. We fou nd that the early phase of the thapsigargin-induced increase in the intrace llular Na+ concentration (approx. 1-2 min after stimulation) was attenuated after preincubation of lymphocytes with D609. By contrast, thapsigargin-in duced Na+ influx was not affected in the presence butan-1-ol, which inhibit s phosphatidylcholine-specific phospholipase D (PG-PLD). The thapsigargin-i nduced Na+ influx could be mimicked by PC-PLC exogenously added to the lymp hocyte suspension, whereas addition of PC-PLD had no effect. In addition, t hapsigargin stimulated formation of the physiological PC-PLC products, diac ylglycerol. Cell-permeable diacylglycerol analogue, dioctanoyl-glycerol (DO G), produced time- and concentration-dependent increase in the intracellula r Na+ concentration. Both thapsigargin- and DOG-induced Na+ increases were not affected in the presence of Na+/H+ antiport inhibitor, HOE609, or Na+/C a2+ antiport inhibitor, dimethylthiourea, as well as in the presence of Co2 + and Ni2+, which block store-operated Ca2+ entry. By contrast, markedly re duced thapsigargin- and DOG-induced Na+ influx were noted in the presence o f flufenamic acid, which blocks the non-selective cation current (I-CRANC). In conclusion, our results suggest that diacylglycerol released due to the PC-PLC activation contributes to the thapsigargin-induced Na+ entry. (C) 2 000 Elsevier Science Inc. All rights reserved.