Agonist-specific regulation of inositol phosphate metabolism in cardiac endothelial cells.

The actions of nucleotides and hormones at endothelial cell (EC) receptors are known to result in the release of ATP that acts as a local hormone to facilitate release of mediators such as NO and PGI2. Stimulation of ECs with the P2Y1 receptor agonist 2-MeSATP leads to the rapid release of Ca2+ from stores consistent with a role for inositol trisphosphate (Ins-1,4,5-P3) in mediating the action of extracellular nucleotides. Guinea pig ECs were grown in primary culture. [3H]d-myo-inositol (30 Ci/mmol) labeling studies revealed maximal incorporation of radioactivity into [3H]Ins-1,4,5-P3 when glucose in the labeling buffer was lowered to 1 mM and non-radioactive inositol was added at 10 microM. Stimulation of EC for one sec led to the dose-dependent accumulation of [3H]Ins-1,4,5-P3 as well as [3H]IP4, [3H]IP5, and [3H]IP6. Unexpectedly, the metabolism of [3H]Ins-1,4,5-P3 to IP1 was disparat in stimulated versus un-stimulated cells. In [3H]d-Ins labeled stimulated EC or in homogenates derived from unlabeled, stimulated EC, dephosphorylation of [3H]Ins-1,4,5-P3 led to the exclusive formation of [3H]Ins-4-P1. Addition of on-nucleotide agonists such as bradykinin gave the same results suggesting that the dephosphorylation pathway for IP formation in EC is dependent on agonist stimulation and may be correlated with regulation of agonist responsiveness or heretofore unrecognized actions of IP isomers in stimulated versus unstimulated cells.