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PMID: 6089753 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Tight coupling of thrombin-induced acid hydrolase secretion and phosphatidate synthesis to receptor occupancy in human platelets.

The Biochemical journal ·Vol. 222 ·No. 1 ·1984-08-15 ·Pages 157-67

Holmsen H, Dangelmaier CA, Rongved S

Abstract

Human platelets incubated with [32P]Pi and [3H]arachidonate were transferred to a Pi-free Tyrode's solution by gel filtration. The labile phosphoryl groups of ATP and ADP as well as Pi in the metabolic pool of these platelets had equal specific radioactivity which was identical to that of[32P]phosphatidate formed during treatment of the cells with thrombin for 5 min. Therefore, the 32P radioactivity of phosphatidate was a true, relative measure for its mass. The thrombin-induced formation of[32P]-phosphatidate had the same time course and dose-response relationships as the concurrent secretion of acid hydrolases. 125I-alpha-Thrombin bound maximally to the platelets within 13s and was rapidly dissociated from the cells by hirudin; readdition of excess 125I-alpha-thrombin caused rapid rebinding of radioligand. This binding-dissociation-rebinding sequence was paralleled by a concerted start-stop-restart of phosphatidate formation and acid hydrolase secretion. [3H]Phosphatidylinositol disappearance was initiated upon binding but little affected by thrombin dissociation and rebinding. ATP deprivation caused similar changes in the time courses for [32P]-phosphatidate formation and acid hydrolase secretion which were different from those of [3H]phosphatidylinositol disappearance. The metabolic stress did not alter the magnitude (15%) of the initial decrease in phosphatidylinositol-4,5-bis[32P]phosphate, but did abolish the subsequent increase of phosphatidylinositol-4,5-bis[32P]-phosphate in the thrombin-treated platelets. It is concluded that in thrombin-treated platelets (1) phosphatidate synthesis, but not phosphatidylinositol disappearance, is tightly coupled to receptor occupancy and acid hydrolase secretion in platelets, (2) successive phosphorylations to phosphatidylinositol-4,5-bisphosphate is unlikely to be the main mechanism for phosphatidylinositol disappearance, and (3) only a small fraction (15%) of phosphatidylinositol-4,5-bisphosphate is susceptible to hydrolysis.

MeSH Terms
Adenosine Diphosphate/blood Adenosine Triphosphate/blood Antimetabolites/pharmacology Blood Platelets/drug effects,metabolism Dose-Response Relationship, Drug Hirudins/pharmacology Humans Hydrolases/blood Phosphates/blood Phosphatidic Acids/biosynthesis,blood Phosphatidylinositols/blood Receptors, Cell Surface/metabolism Receptors, Thrombin Thrombin/metabolism,pharmacology
Chemicals
Antimetabolites Hirudins Phosphates Phosphatidic Acids Phosphatidylinositols Receptors, Cell Surface Receptors, Thrombin Adenosine Diphosphate Adenosine Triphosphate Hydrolases Thrombin
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Holmsen H
Dangelmaier C A
Rongved S
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58 references, click to expand
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Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2220157
Published
1984-08-15
Pages
157-67
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1144156
Subset
IM
Grants
NHLBI NIH HHS · HL 14217 · United States
External Links
PubMed source DOI: 10.1042/bj2220157 PMC Full Text View journal details
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