Differential effects of heparin on inositol 1,4,5-trisphosphate binding, metabolism, and calcium release activity in the bovine adrenal cortex.
In a wide variety of cells, inositol-1,4,5-triphosphate is a second messenger that interacts with specific intracellular receptors and triggers the release of sequestered Ca2+ from an intracellular store. We have looked at the influence of heparin on the action and metabolism of inositol-1,4,5-triphosphate in the bovine adrenal cortex. Heparin blocked inositol-1,4,5-trisphosphate binding with half-maximal efficiency around 10 micrograms/ml. Scatchard analyses revealed that heparin did not change the affinity but decreased the number of available binding sites. The Ca2+-releasing activity of inositol-1,4,5-trisphosphate was monitored with the fluorescent indicator, fura-2. Heparin blocked this activity with half-maximal effeciency around 10 micrograms/ml. The effect of heparin could be overcome by a supramaximal dose of inositol-1,4,5-trisphosphate (25 microM). The activity of inositol-1,4,5-trisphosphate-3-kinase from bovine adrenal cortex cytosol was also studied. Heparin inhibited the activity of the kinase with a half-maximal effeciency around 0.4 microgram/ml. Lineweaver-Burk plots revealed that this potent effect was noncompetitive. Finally, we observed that heparin is without effect on inositol-1,4,5-trisphosphate-5-phosphatase (at concentrations as high as 2 mg/ml). These results are consistent with the suggestion that the binding sites for inositol-1,4,5-trisphosphate are the intracellular receptors responsible for the Ca2+-mobilizing effects of inositol-1,4,5-trisphosphate. These results also show that the kinase, the phosphatase, and the receptor are three different molecular entities, which are affected in a different manner by heparin.
Guillemette
G
G
Department of Pharmacology, Faculty of Medicine, University of Sherbrooke, Quebec, Canada.
Lamontagne
S
S
Boulay
G
G
Mouillac
B
B
D000302:Adrenal Cortex / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D006493:Heparin / Q000494:pharmacology*; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate; D007295:Inositol Phosphates / Q000378:metabolism* / Q000494:pharmacology; D000072181:Inositol Polyphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis; D010770:Phosphotransferases / Q000032:analysis; D017853:Phosphotransferases (Alcohol Group Acceptor); D013403:Sugar Phosphates / Q000378:metabolism*
D007295:Inositol Phosphates; D013403:Sugar Phosphates; D015544:Inositol 1,4,5-Trisphosphate; D006493:Heparin; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C051038:Inositol 1,4,5-trisphosphate 3-kinase; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; D002118:Calcium