REGULATION OF PHOSPHOLIPASE-D ACTIVITY IN NEUROBLASTOMA-CELLS

The regulation of phospholipase D was studied in human neuroblastoma c ells using phosphatidylethanol as a marker of the enzyme activity. Car bachol induced phospholipase D activity in SH-SY5Y cells. Muscarinic a ntagonists inhibited the response with potencies suggesting that musca rinic M(1) receptors are responsible for the activation. In permeabili zed SH-SY5Y cells, both the carbachol- and GTP gamma S-induced Peth fo rmation was inhibited by GDP beta S, indicating that both responses ar e mediated via a G-protein. The protein kinase C inhibitors, bisindoly lmaleimide and staurosporine significantly inhibited the carbachol-ind uced Peth formation whereas H7 had no effect. Thus, the cholinergic ac tivation of phospholipase D in SH-SY5Y cells is probably mediated via a direct receptor-G-protein coupling but an involvement of protein kin ase C cannot be excluded. Calmidazolium, a calmodulin antagonist, indu ced an increase in phosphatidylethanol formation in both SH-SY5Y and I MR-32 cells. This effect was inhibited by genistein and tyrphostin, in dicating a tyrosine kinase dependent pathway for phospholipase D activ ation in neuroblastoma cells.