Thromboxane A2 activates phospholipase C in astrocytoma cells via pertussis toxin-insensitive G-protein.
The properties of thromboxane A2 (TXA2) receptors were examined in 1321N1 human astrocytoma cells. 9,11-Epithio-11,12-methanothromboxane A2 (STA2), a stable analogue of TXA2, stimulated the accumulation of inositol phosphates (IPs) with an EC50 of about 50 nM. The STA2-induced accumulation of IPs was inhibited concentration dependently by ONO3708, a TXA2 receptor antagonist, with an inhibition constant (Ki) of about 10 nM. Inositol trisphosphate (IP3) was accumulated more rapidly than inositol bisphosphate (IP2) in response to STA2. HPLC analysis indicated that inositol 1,4,5-trisphosphate accumulated in the presence of STA2. STA2 alone had no effect on the accumulation of IPs in membrane preparations but it potentiated the accumulation induced by GTP gamma S. [3H]SQ29548, a TXA2 receptor antagonist, bound specifically to TXA2 receptors, expressing a single binding site with a dissociation constant (Kd) of 10.9 nM. The competition curve for STA2 inhibition of [3H]SQ29548 binding was shifted to the right and was steeper in the presence of GTP gamma S. Pertussis toxin (IAP) elicited ADP-ribosylation of 41KD protein but had no effect on the sensitivity to GTP of the STA2 inhibition of SQ29548 binding or of STA2-induced accumulation of IPs. It is concluded from these results that the stimulation of TXA2 receptors results in activation of phospholipase C via a GTP binding protein and that the protein is not a substrate for IAP.
Nakahata
N
N
Department of Pharmacology, Fukushima Medical College, Japan.
Matsuoka
I
I
Ono
T
T
Nakanishi
H
H
D000244:Adenosine Diphosphate / Q000378:metabolism; D001254:Astrocytoma / Q000378:metabolism*; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002851:Chromatography, High Pressure Liquid; D004789:Enzyme Activation / Q000187:drug effects; D005231:Fatty Acids, Unsaturated; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006834:Hydrazines / Q000378:metabolism / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism; D037342:Pertussis Toxin; D010716:Phosphatidylinositols / Q000378:metabolism; D011982:Receptors, Prostaglandin / Q000378:metabolism; D017482:Receptors, Thromboxane; D012266:Ribose / Q000378:metabolism; D013928:Thromboxane A2 / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D019086:Bridged Bicyclo Compounds, Heterocyclic; D005231:Fatty Acids, Unsaturated; D006834:Hydrazines; D007295:Inositol Phosphates; D010716:Phosphatidylinositols; D011982:Receptors, Prostaglandin; D017482:Receptors, Thromboxane; D010566:Virulence Factors, Bordetella; D013928:Thromboxane A2; D000244:Adenosine Diphosphate; D012266:Ribose; C045749:SQ 29548; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins