Prevention of alphaII(b)beta3 activation by non-steroidal antiinflammatory drugs

We have studied the effect of non-steroidal antiinflammatory drugs (NSAIDs) on alphaII(b)beta3 integrin activation and platelet aggregation. NSAIDs such as meloxicam, piroxicam, indomethacin and aspirin, but not aceclofenac or diclofenac interfered with the activation state of alphaII(b)beta3. NSAIDs that inhibited alphaII(b)beta3 activation were also able both to partially inhibit platelet primary aggregation and to accelerate platelet deaggregation. These effects of NSAIDs were not dependent on cyclooxygenase inhibition. The results obtained indicate that some NSAIDs exert a specific action on alphaII(b)beta3 activation, and provide an additional mechanism that accounts for their beneficial effects in diseases in which platelet activation is involved.     

Effects of lysolecithin on aggregation of human platelets induced by arachidonic acid and A23187 role of prostaglandin intermediary metabolism.

Lysolecithin at non-cytotoxic concentrations (30–500 uM) was found capable of completely inhibiting the $\text{aggregation of human platelets}$ induced by arachidonic acid in the absence of any effect upon total platelet production of malondialdehyde, an end-product of platelet $\text{prostaglandin intermediary metabolism}$, and to inhibit platelet aggregation stimulated by the calcium ionophore, A23187. As the induction of platelet aggregation by arachidonic acid is dependent upon an intact prostaglandin biosynthetic pathway while that of A23187 is not and since lysolecithin-induced inhibition of arachidonic acid-stimulated platelet aggregation was evident in the absence of an effect upon platelet malondialdehyde production, it is suggested that lysolecithin inhibits the platelet release reaction and irreversible aggregation by a mechanism separable from a major affect upon prostaglandin intermediary metabolism.     

Effects of C-reactive protein on platelet function. III. The role of cAMP, contractile elements, and prostaglandin metabolism in CRP-induced inhibition of platelet aggregation and secretion.

It was previously demonstrated that C-reactive protein (CRP) inhibits platelet aggregation and release reactions, activation of platelet factor 3, and platelet-dependent clot retraction. Multiple considerations including selective inhibition of secondary wave aggregation suggested that CRP exerted its inhibitory effects by interfering with the release of endogenous ADP. In the present investigation, CRP was found by direct assay to inhibit the release of endogenous ADP and/or serotonin concomitant with inhibition of platelet aggregation stimulated by ADP, epinephrine, thrombin, and AHGG. CRP did not induce an increase in the basal level of platelet cAMP, suggesting independence of a direct effect upon this mediator system. Furthermore, CRP did not inhibit the aggregation and secretion induced by the antibiotic ionophore A23187, suggesting the absence of a direct effect upon the activation of platelet contractile elements. By contrast, CRP did inhibit both thrombin-induced release of malondialdehyde, a prostaglandin endoperoxide nonprostanoate endproduct, and platelet aggregation induced by the prostaglandin endoperoxide precursor arachidonic acid. These data, therefore, raise the possibility that CRP inhibits platelet reactivities by interfering with an aspect of porstaglandin metabolism, and that this occurs subsequent to the hydrolytic accumulation of arachidonic acid and prior to the movement of calcium from the platelet dense tubules. These studies support the concept that CRP serves to modulate platelet reactivities during acute inflammatory reactions.     

Lack of influence of the COX inhibitors metamizol and diclofenac on platelet GPIIb/IIIa and P-selectin expression in vitro

BACKGROUND: The effect of non-steroidal anti-inflammatory drugs (NSAIDs) for reduced platelet aggregation and thromboxane A2 synthesis has been well documented. However, the influence on platelet function is not fully explained. Aim of this study was to examine the influence of the COX-1 inhibiting NSAIDs, diclofenac and metamizol on platelet activation and leukocyte-platelet complexes, in vitro. Surface expression of GPIIb/IIIa and P-selectin on platelets, and the percentage of platelet-leukocyte complexes were investigated. METHODS: Whole blood was incubated with three different concentrations of diclofenac and metamizol for 5 and 30 minutes, followed by activation with TRAP-6 and ADP. Rates of GPIIb/IIIa and P-selectin expression, and the percentage of platelet-leukocyte complexes were analyzed by a flow-cytometric assay. RESULTS: There were no significant differences in the expression of GPIIb/IIIa and P-selectin, and in the formation of platelet-leukocyte complexes after activation with ADP and TRAP-6, regarding both the time of incubation and the concentrations of diclofenac and metamizol. CONCLUSIONS: Accordingly, the inhibitory effect of diclofenac and metamizol on platelet aggregation is not related to a reduced surface expression of P-selectin and GPIIb/IIIa on platelets.     

Comparison of verapamil and nifedipine in thrombosis models

Calcium blockers and calmodulin antagonists have been reported to inhibit the aggregation of blood platelets in vitro. In the present study, the effects of two calcium blockers, verapamil and nifedipine, were compared in several rodent thrombosis models. In rat and mouse platelet-rich plasma, preincubation with either verapamil or nifedipine had a dose-dependent inhibitory effect on collagen-induced aggregation (P less than 0.01). The concentration required for 50% inhibition of rat platelet aggregation was 0.91 X 10(-4) M for verapamil and 1.77 X 10(-4) M for nifedipine. In in vivo thrombosis models in mice, acute pretreatment with nifedipine had a significant, dose-dependent protective effect (P less than 0.05). At a dose of 500 micrograms/kg, nifedipine inhibited thrombotic sudden death provoked by arachidonic acid, a thromboxane agonist (U46619), or a combination of collagen and epinephrine. In vivo platelet depletion induced by U46619 was also inhibited by this calcium blocker. Thus, nifedipine is protective against a variety of thrombotic stimuli, and its antiplatelet aggregatory effect apparently extends to the in vivo situation. In contrast, no in vivo antithrombotic activity was observed for verapamil. Two additional calcium blockers, perhexilene and diltiazem, and three calmodulin antagonists, W-7, chlorpromazine, and trifluoperazine, were also tested in the U46619-induced thrombotic sudden death model. Of these, only diltiazem (5 and 10 mg/kg) had an acute protective effect.     

Nitric oxide and cardiovascular effects: new insights in the role of nitric oxide for the management of osteoarthritis

Nitric oxide (NO) is an important mediator in both health and disease. In addition to its effects on vascular tone and platelet function, it plays roles in inflammation and pain perception that may be of relevance in osteoarthritis. Many patients with osteoarthritis take nonsteroidal anti-inflammatory drugs (NSAIDs) long term for pain control. Over recent years concern has been raised about the possible cardiovascular side effects of NSAIDs. The reasons for this possible increased cardiovascular risk with NSAIDs are not yet entirely clear, although changes in blood pressure, renal salt handling and platelet function may contribute. Recently, drugs that chemically link a NSAID with a NO donating moiety (cyclo-oxygenase-inhibiting NO-donating drugs [CINODs]) were developed. NO is an important mediator of endothelial function, acting as a vasodilator and an inhibitor of platelet aggregation, and having anti-inflammatory properties. The potential benefits of CINODs include the combination of effective analgesic and anti-inflammatory actions with NO release, which might counterbalance any adverse cardiovascular effects of NSAIDs. Effects of CINODs in animal studies include inhibition of vasopressor responses, blood pressure reduction in hypertensive rats and inhibition of platelet aggregation. CINODs may also reduce ischemic damage to compromised myocardial tissue. In addition, endothelial dysfunction is a recognized feature of inflammatory arthritides, and therefore a drug that might provide slow release of NO to the vasculature while treating pain is an attractive prospect in these conditions. Further studies of the effects of CINODs in humans are required, but these agents represent a potential exciting advance in the management of osteoarthritis.     

Effect of t-butyl hydroperoxide on liver microsomal membranes and microsomal calcium sequestration

In vitro exposure of hepatocytes or liver microsomes to t-butyl hydroperoxide resulted in a marked decrease of liver microsomal calcium pump activity. Decreased calcium pump activity was dependent upon both concentration and time. Liver microsomes could be protected from this effect by glutathione or dithiothreitol. In addition to decreased calcium pump activity, exposure of liver microsomes to t-butyl hydroperoxide produced a concentration-dependent aggregation of microsomal membrane protein as determined by polyacrylamide gel electrophoresis. Inhibition of microsomal calcium pump activity was observed when intact hepatocytes were incubated, in vitro, with t-butyl hydroperoxide. However, aggregation of microsomal membrane protein was not observed when hepatocytes were incubated with t-butyl hydroperoxide. The effects produced by exposure of liver microsomes to this compound do not appear to be a complete model of actions of the compound on intact cells.     

Effect of etmozin on thrombocyte aggregation capacity

It was shown in in vitro experiments that etmozin at a concentration of 100 micrograms/ml significantly suppressed (by 21%) platelet aggregation induced by ADP, but it had no effect on platelet aggregation induced by arachidonic acid. In in vivo experiments etmozin was found to cause a marked suppression of tendon collagen-induced platelet aggregation in the doses 2-5 mg/kg having antiarrhythmic activity. Under suppressed platelet aggregation induced by indomethacin, the prostaglandin biosynthesis blocker etmozin displayed no antiaggregation effect. It is suggested that etmozin effects on ADP release from platelets play the main role in the mechanism of its antiaggregation action.     

Effects of Welsh onion extracts on human platelet function in vitro

Welsh onion has been consumed for prevention of cardiovascular disorders. However, its underlying mechanisms are still unclear. This study investigated whether Welsh onion extracts can alter human platelet function (ie, platelet adhesion, aggregation, and thromboxane release). To clarify the underlying mechanisms, we also measured the intracellular calcium ([Ca2+]i) and cyclic nucleotide levels in platelets. Our results showed that 1) boiled extracts directly induced platelet aggregation in a dose-dependent manner; 2) raw extracts inhibited platelet adhesion and ADP-evoked platelet aggregation, while boiled extracts enhanced them; 3) raw green extract suppressed ADP-stimulated platelet [Ca2+]i elevation and thromboxane production, whereas boiled green extract enhanced them; 4) raw green extract elevated platelet cAMP level, whereas boiled green extract had no effect on cAMP level. Furthermore, the boiled green extract, but not the raw extract, induced pronounced platelet morphological changes. In conclusion, raw extracts of Welsh onion inhibit platelet function in vitro while boiled extracts activate platelets.     

Effects of novel plant antioxidants on platelet superoxide production and aggregation in atherosclerosis.

Superoxide anion is produced in human platelets predominantly by Nox2-dependent NADPH oxidases. In vitro experiments have shown that it might play a role in modulating platelet functions. The relationship between platelet superoxide production and aggregation remains poorly defined. Accordingly, we aimed to study superoxide production and aggregation in platelets from subjects with significant cardiovascular risk factors (hypertension, hypercholesterolemia, smoking and diabetes mellitus) and from control individuals. Moreover, we studied the effects of novel polyphenol-rich extracts of Aronia melanocarpa (chokeberry) berries on platelet function in vitro. Superoxide production was significantly increased in patients with cardiovascular risk profile when compared to controls, while platelet aggregation in response to either collagen or thrombin were borderline higher, and did not reach statistical significance. Interestingly, no relationship was observed between platelet aggregation ex vivo and platelet superoxide production in either of studied groups. No correlation was found between endothelial function (measured by FMD) and platelet aggregation ex vivo either. Polyphenol-rich extracts of A. melanocarpa berries caused a significant concentration dependent decrease in superoxide production only in patients with cardiovascular risk factors, while no effect was observed in the control group. A. melanocarpa extracts abolished the difference in superoxide production between risk factor patients and controls. A. melanocarpa extracts exerted significant concentration dependent anti-aggregatory effects in both studied groups, which indicated that these effects may be independent of it's ability to modulate superoxide production. The anti-aggregatory effects of chokeberry extracts were similar irrespective of aggregation inducing agent (collagen or thrombin). Moreover, they appear to be independent of platelet NO release as NOS inhibition by L-NAME did not lead to their abrogation.     

广西眼镜蛇毒L-氨基酸氧化酶体内外抗血小板聚集实验研究

目的 研究广西眼镜蛇中提取的L-氨基酸氧化酶(L-amino acid oxidase)在人体外及家兔体内的抗血小板聚集作用.方法 用比浊法测定广西眼镜蛇毒L-氨基酸氧化酶对二磷酸腺苷(ADP)、胶原、凝血酶、花生四烯酸(AA)在人体外及家兔体内引起的血小板聚集率的影响.结果 实验中,能明显抑制二磷酸腺苷(ADP)、胶原、凝血酶、花生四烯酸(AA)引起的血小板聚集,并呈明显的正相关.结论 广西眼镜蛇毒L-氨基酸氧化酶在体内外均有较强的抗血小板聚集活性.     

Effect of dipyridamole on prostaglandin generation by human platelets and vessel walls

These experiments were conducted to determine the effects of dipyridamole on human platelet aggregation, platelet thromboxane A2 (TXA2) and human vessel wall prostacyclin (PGI2) generation. Dipyridamole in varying concentrations (5 to 50 micrograms/ml) had no direct effect on ADP-induced platelet aggregation in vitro, but it potentiated PGI2-induced platelet aggregation inhibition at these concentrations. Dipyridamole also inhibited arachidonic acid-induced platelet TXA2 generation at these concentrations. In continuously perfused umbilical vein segments, dipyridamole treatment resulted in stimulation of PGI2 release determined by bioassay and by measurement of its stable metabolite 6-keto-PGF1 alpha. Minimum concentration of dipyridamole causing PGI2 release was 50 micrograms/ml. These in vitro studies suggest that anti-thrombotic effects of dipyridamole in man are mediated mainly by potentiation of PGI2 activity and to some extent by TXA2 suppression. Stimulation of PGI2 release by human vessels may not be seen in usual therapeutic concentrations.     

Effects of gaseous anesthetics and ultrashort and short-acting barbiturates on human blood platelet free cytosolic calcium: relevance to their effects on platelet aggregation.

The effects of elevated pressures (to 6 atmospheres absolute (ATA)) of nitrous oxide (N2O) and of xenon (Xe), and barbiturates on platelet free cytosolic calcium ([Ca2+]i) and platelet aggregation were studied. N2O inhibited the ADP-induced rise in [Ca2+]i whereas Xe had no effect. Neither affected basal levels. Pentobarbital and methohexital had little effect on basal or stimulated levels in the presence or "absence" of extracellular Ca2+; but both, at concentrations > 10(-4) M, inhibited platelet aggregation induced by adenosine diphosphate. Thiopental increased basal and stimulated [Ca2+]i when extracellular Ca2+ was present, but not when it was absent, and displayed a bimodal effect with low and high doses being more active than intermediate ones. It also potentiated aggregation. Methitural displayed similar, but nonsignificant, effects. These patterns held for all agents whether or not acetylsalicylic acid was present. Pentobarbital and methohexital inhibited phorbol myristate acetate aggregation in low extracellular calcium and no potentiation was seen with thiopental. In the absence of extracellular Ca2+, no potentiation was observed in stimulated platelets. Potentiation of aggregation previously reported for Xe does not involve increased Ca2+ uptake and did not occur in the absence of extracellular Ca2+. A common mechanism of action for these agents cannot be inferred from their effects on platelet aggregation or [Ca2+]i, as their pharmacological profiles differ markedly. It is evident that their inhibitory properties in this cell are not dependent on extracellular Ca2+, whereas the potentiation observed with pentobarbital, and formerly with Xe, is so dependent.     

Calcitonin gene-related peptide-mediated antihypertensive and anti-platelet effects by rutaecarpine in spontaneously hypertensive rats

We have previously reported that Chinese traditional medicine rutaecarpine (Rut) produced a sustained hypotensive effect in phenol-induced and two-kidney, one-clip hypertensive rats. The aims of this study are to determine whether Rut could exert antihypertensive and anti-platelet effects in spontaneously hypertensive rats (SHR) and the underlying mechanisms. In vivo, SHR were given Rut and the blood pressure was monitored. Blood was collected for the measurements of calcitonin gene-related peptide (CGRP), tissue factor (TF) concentration and activity, and platelet aggregation, and the dorsal root ganglia were saved for examining CGRP expression. In vitro, the effects of Rut and CGRP on platelet aggregation were measured, and the effect of CGRP on platelet-derived TF release was also determined. Rut exerted a sustained hypotensive effect in SHR concomitantly with the increased synthesis and release of CGRP. The treatment of Rut also showed an inhibitory effect on platelet aggregation concomitantly with the decreased TF activity and TF antigen level in plasma. Study in vitro showed an inhibitory effect of Rut on platelet aggregation in the presence of thoracic aorta, which was abolished by capsazepine or CGRP(8-37), an antagonist of vanilloid receptor or CGRP receptor. Exogenous CGRP was able to inhibit both platelet aggregation and the release of platelet-derived TF, which were abolished by CGRP(8-37). The results suggest that Rut exerts both antihypertensive and anti-platelet effects through stimulating the synthesis and release of CGRP in SHR, and CGRP-mediated anti-platelet effect is related to inhibiting the release of platelet-derived TF.     

Small molecular weight heat shock-related protein, HSP20, exhibits an anti-platelet activity by inhibiting receptor-mediated calcium influx

We have shown that Hsp20, one of small molecular weight heat shock protein, which is present at a high concentration both in vascular smooth muscle cells and in circulating blood in patient with vascular disease, strongly inhibits platelet aggregation in vitro and ex vivo. To clarify the mechanism, we investigated the effect of Hsp20 on free calcium concentration in human platelet cytoplasm using fura 2. Hsp20 inhibited thrombin-induced calcium influx without affecting calcium release from intracellular calcium stores. The degree of inhibition is well-correlated with that of suppression of thrombin-induced platelet aggregation by this substance. Hsp20 also inhibited the elevation of cytoplasmic free calcium level triggered by collagen, but not that by A-23187. In contrast, Hsp28, another type of small molecular weight Hsp, failed to affect the cytoplasmic free calcium level. These findings suggest that Hsp20 inhibits the receptor-mediated calcium influx of platelets without affecting calcium release from intracellular calcium stores, leading to its anti-platelet activity.     

Inhibition of platelet aggregation and arachidonate metabolism in platelets by procyanidins

The effects of procyanidins on platelet aggregation and arachidonate metabolism in platelets were studied. Nine procyanidins were used in this investigation. Procyanidins B-2-S, EEC and C-1 significantly induced the inhibition of platelet aggregation, and the potency of inhibition was comparable with aspirin. Procyanidin B-2-S was used as a representative of procyanidins for further studies on the effect on arachidonate metabolism. In arachidonate metabolism by fatty acid cyclooxygenase pathway, B-2-S inhibited TXB2 and HHT formation by intact platelets treated with exogenous arachidonic acid. It also inhibited TXB2 formation measured by a specific radioimmunoassay when the cells were challenged with calcium ionophore A23187. In cell-free system, B-2-S inhibited both TXB2 and 12-HETE bioxynthesis in platelet microsome and cytosol, respectively. The inhibitory effect on thromboxane biosynthesis might explain the inhibitory effect of procyanidins on platelet aggregation.     

Effects of atrial natriuretic factor on human platelet function

We examined the hypothesis that atrial natriuretic factor (ANF), a substance with known vasorelaxant activities, shares with other vasodilators the property of inhibiting platelet function. Aggregation of citrated platelet-rich plasma (PRP) from 23 healthy volunteers induced by ADP, adrenaline, arachidonic acid, collagen, gamma-thrombin, the endoperoxide analogue U-44069, serotonin, the calcium ionophore A-23187 or platelet aggregating factor was measured after incubation of PRP with ANF for 3 minutes at concentrations of 4 X 10(-9), 4 X 10(-8) and 4 X 10(-7) M or vehicle as control. ANF decreased ADP-induced aggregation significantly (P less than 0.02), but only at the highest concentration used and to a minor extent (control: 73.6 +/- 11.2%; after ANF 4 X 10(-7) M: 60.0 +/- 17.1%, mean +/- S.D., n = 39) by a selective inhibitory effect on the secondary wave; neither aggregation by all other agents tested nor thromboxane B2 generation induced by ADP and adrenaline was altered by incubation with ANF. Although ANF thus has detectable effects on ADP-induced platelet aggregation in vitro, these data suggest that ANF is unlikely to be a physiologically significant modulator of platelet function.     

Adenylic dinucleotides produced by CD38 are negative endogenous modulators of platelet aggregation

Diadenosine 5',5'-P1,P2-diphosphate (Ap2A) is one of the adenylic dinucleotides stored in platelet granules. Along with proaggregant ADP, it is released upon platelet activation and is known to stimulate myocyte proliferation. We have previously demonstrated synthesis of Ap2A and of two isomers thereof, called P18 and P24, from their high pressure liquid chromatography retention time, by the ADP-ribosyl cyclase CD38 in mammalian cells. Here we show that Ap2A and its isomers are present in resting human platelets and are released during thrombin-induced platelet activation. The three adenylic dinucleotides were identified by high pressure liquid chromatography through a comparison with the retention times and the absorption spectra of purified standards. Ap2A, P18, and P24 had no direct effect on platelet aggregation, but they inhibited platelet aggregation induced by physiological agonists (thrombin, ADP, and collagen), with mean IC(50) values ranging between 5 and 15 mum. Moreover, the three dinucleotides did not modify the intracellular calcium concentration in resting platelets, whereas they significantly reduced the thrombin-induced intracellular calcium increase. Through binding to the purinergic receptor P2Y(11), exogenously applied Ap2A, P18, and P24 increased the intracellular cAMP concentration and stimulated platelet production of nitric oxide, the most important endogenous antiaggregant. The presence of Ap2A, P18, and P24 in resting platelets and their release during thrombin-induced platelet activation at concentrations equal to or higher than the respective IC(50) value on platelet aggregation suggest a role of these dinucleotides as endogenous negative modulators of aggregation.     

Inhibitory effects of total flavones of Hippophae Rhamnoides L on thrombosis in mouse femoral artery and in vitro platelet aggregation

Total flavones of Hippophae Rhamnoides L (TFH) are extracted from Sea buckthorn, a Chinese herbal medicine. Sea buckthorn has antioxidant, anti-ulcerogenic and hepato-protective actions, and its berry oil is reported to suppress platelet aggregation. Though it is frequently used for patients with thrombosis, the likely mechanism(s) and effects of TFH on thrombogenesis remain unclear. Thus, we have investigated the effect in-vivo of TFH on thrombogenesis and in vitro on platelet aggregation, comparing them to those of aspirin.We measured thrombotic occlusion time in a mouse femoral artery thrombosis model by the photochemical reaction between intravenously injected rose bengal and green light irradiation. In vitro platelet aggregation in whole blood was measured by single platelet counting. Thrombotic occlusion time was 8.5 +/- 0.6 min in the control group. TFH at a dose of 300 micro g/kg, intravenously administered 15 min before the rose bengal injection, significantly prolonged it to 11.6 +/- 1.0 min (P < 0.05), a similar effect on in-vivo thrombogenesis to that of aspirin. TFH at a concentration of 3.0 micro g/ml significantly (P < 0.01) inhibited in vitro platelet aggregation induced by collagen (2 micro g/ml) in a concentration dependent manner, in contrast TFH did not affect aggregation induced by arachidonic acid (80 micro M) and ADP (0.3 micro M).The results of the present study, in which TFH prevented in-vivo thrombogenesis, probably due to inhibition of platelet aggregation, suggest a possible clinical approach for the prevention of thrombosis.     

Molecular mechanism of thromboxane A(2)-induced platelet aggregation. Essential role for p2t(ac) and alpha(2a) receptors.

Thromboxane A(2) is a positive feedback lipid mediator produced following platelet activation. The G(q)-coupled thromboxane A(2) receptor subtype, TPalpha, and G(i)-coupled TPbeta subtype have been shown in human platelets. ADP-induced platelet aggregation requires concomitant signaling from two P2 receptor subtypes, P2Y1 and P2T(AC), coupled to G(q) and G(i), respectively. We investigated whether the stable thromboxane A(2) mimetic, (15S)-hydroxy-9, 11-epoxymethanoprosta-5Z,13E-dienoic acid (U46619), also causes platelet aggregation by concomitant signaling through G(q) and G(i), through co-activation of TPalpha and TPbeta receptor subtypes. Here we report that secretion blockade with Ro 31-8220, a protein kinase C inhibitor, completely inhibited U46619-induced, but not ADP- or thrombin-induced, platelet aggregation. Ro 31-8220 had no effect on U46619-induced intracellular calcium mobilization or platelet shape change. Furthermore, U46619-induced intracellular calcium mobilization and shape change were unaffected by A3P5P, a P2Y1 receptor-selective antagonist, and/or cyproheptadine, a 5-hydroxytryptamine subtype 2A receptor antagonist. Either Ro 31-8220 or AR-C66096, a P2T(AC) receptor selective antagonist, abolished U46619-induced inhibition of adenylyl cyclase. In addition, AR-C66096 drastically inhibited U46619-mediated platelet aggregation, which was further inhibited by yohimbine, an alpha(2A)-adrenergic receptor antagonist. Furthermore, inhibition of U46619-induced platelet aggregation by Ro 31-8220 was relieved by activation of the G(i) pathway by selective activation of either the P2T(AC) receptor or the alpha(2A)-adrenergic receptor. We conclude that whereas thromboxane A(2) causes intracellular calcium mobilization and shape change independently, thromboxane A(2)-induced inhibition of adenylyl cyclase and platelet aggregation depends exclusively upon secretion of other agonists that stimulate G(i)-coupled receptors.