Desensitization of TP receptor-mediated platelet activation has been studied extensively. Liel, et al (154) and Murray and FitzGerald £155) carried out the first studies of homologous desensitization of normal TP receptors. They observed that exposure of platelets to TP receptor agonists resulted in attenuation of subsequent agonist-induced platelet aggregation and a reduction in receptor sites identified by radioligand binding. Okwu, et al (107) found that shape change, as well as aggregation, induced by I-BOP was inhibited following incubation of human platelets with U46619. Binding of both agonist and antagonist radioligands was decreased following homologous desensitization (107,154,155). Murray and FitzGerald (155) also provided evidence that TP receptor desensitization resulted in receptor-G protein uncoupling. Studies carried out in human astrocytoma cells indicated that homologous desensitization of TP receptors occurred in two phases (156). The early phase of desensitization to STA2 involved uncoupling of receptors from G proteins, while a reduction of binding sites occurred later. Spumey (157) observed that desensitization of transfected mouse TP receptors was dependent upon the level of receptor expression achieved. Prior exposure (10 min.) to U46619 inhibited subsequent U46619-induced IPj formation and calcium flux ~50% in mouse mesangial cells expressing low levels of mouse TP receptors, while these indicators of PLC-P_activation were very little affected in cells with high levels of receptor expresssion.
Dom and Davis (158) found that PKC activation was necessary for TP receptor desensitization. Subsequent studies of the mechanism of desensitization revealed evidence of TP receptor phosphorylation (See IV. TP RECEPTOR FUNCTION, E. TP RECEPTOR PHOSPHORYLATION below) that was in part mediated by PKC. Studies of CHRF-288 cells indicated that TP receptor internalization also occurred following agonist exposure (159). Desensitization of receptor-effector coupling, that occurred early (Tl/2 ~1 hour) following agonist exposure, was separable from receptor internalization, that occurred later. [125I]-BOP binding sites decreased 57%, without a change in binding affinity, after 24 hour incubation with U 46619. Both internalization and degradation were found to be necessary for maximal desenstization of TP receptors (159).
Transfected TP-a and TP-P receptors in CHO cells demonstrated different patterns of homologous desensitization. TP-a receptors, overexpressed in fibroblasts, decreased in number, as did native platelet receptors, following 24 hour exposure to I-BOP, but TP-P receptors increased following similar long-term exposure to I-BOP (160). Activation of PKC uncoupled calcium mobilization from TP-P receptors to a greater extent than from TP-a_receptors (160). These differences cannot be attributed to differences in phosphorylation, since both isoforms transfected into HEK cells were phosphorylated similarly on agonist stimulation (60).
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