Inhibition of phospholipase C activity in Drosophila photoreceptors by 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA) and di-bromo BAPTA

Cell Calcium. 2005 Dec;38(6):547-56. doi: 10.1016/j.ceca.2005.07.005. Epub 2005 Sep 2.

Abstract

In vivo light-induced and basal hydrolysis of phosphatidyl inositol 4,5-bisphosphate (PIP2) by phospholipase C (PLC) were monitored in Drosophila photoreceptors using genetically targeted PIP2-sensitive ion channels (Kir2.1) as electrophysiological biosensors for PIP2. In cells loaded via patch pipettes with varying concentrations of Ca2+ buffered by 4 mM free BAPTA, light-induced PLC activity, showed an apparent bell-shaped dependence on free Ca2+ (maximum at "100 nM", approximately 10-fold inhibition at <10nM or approximately 1 microM). However, experiments where the total BAPTA concentration was varied whilst free [Ca2+] was maintained constant indicated that inhibition of PLC at higher (>100 nM) nominal Ca2+ concentrations was independent of Ca2+ and due to inhibition by BAPTA itself (IC50 approximately 8 mM). Di-bromo BAPTA (DBB) was yet more potent at inhibiting PLC activity (IC50 approximately 1mM). Both BAPTA and DBB also appeared to induce a modest, but less severe inhibition of basal PLC activity. By contrast, EGTA, failed to inhibit PLC activity when pre-loaded with Ca2+, but like BAPTA, inhibited both basal and light-induced PLC activity when introduced without Ca2+. The results indicate that both BAPTA and DBB inhibit PLC activity independently of their role as Ca2+ chelators, whilst non-physiologically low (<100 nM) levels of Ca2+ suppress both basal and light-induced PLC activity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Drosophila Proteins / antagonists & inhibitors*
  • Drosophila Proteins / metabolism
  • Drosophila melanogaster / drug effects
  • Drosophila melanogaster / enzymology*
  • Egtazic Acid / analogs & derivatives*
  • Egtazic Acid / pharmacology
  • Patch-Clamp Techniques
  • Photoreceptor Cells, Invertebrate / cytology
  • Photoreceptor Cells, Invertebrate / drug effects*
  • Photoreceptor Cells, Invertebrate / enzymology*
  • Type C Phospholipases / antagonists & inhibitors*
  • Type C Phospholipases / metabolism

Substances

  • Drosophila Proteins
  • 5,5'-dibromo-1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid
  • Egtazic Acid
  • Type C Phospholipases
  • 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid