Antiplatelet effects of protopine isolated from Corydalis tubers

Protopine inhibited the aggregation and ATP release of rabbit platelets induced by ADP, arachidonic acid, PAF, collagen and ionophore A23187. Although the platelet aggregation caused by thrombin was not inhibited by protopine (100 micrograms/ml), the release reaction was partially suppressed. In rabbit platelet-rich plasma, protopine also inhibited the platelet aggregation caused by ADP, arachidonic acid, PAF and collagen. The thromboxane B2 formation of washed platelets caused by arachidonic acid, collagen, ionophore A23187 and thrombin was suppressed by protopine. Protopine inhibited the intracellular calcium increase caused by arachidonic acid in quin-2/AM loaded rabbit platelets. In the presence of indomethacin, the intracellular calcium increase caused by collagen and PAF was completely suppressed by protopine, and the intracellular calcium increase caused by thrombin was partially inhibited. The phosphoinositides breakdown caused by collagen and PAF was inhibited by protopine, but that by thrombin was not affected significantly. Protopine did not cause the elevation of cyclic AMP level of platelets. It is concluded that the antiplatelet effects of protopine is due to inhibition on thromboxane formation and phosphoinositides breakdown and then lead to the decrease of intracellular calcium concentration.     

Inhibition of platelet thromboxane formation and phosphoinositides breakdown by osthole from Angelica pubescens

Osthole, isolated from Chinese herb Angelica pubescens, inhibited platelet aggregation and ATP release induced by ADP, arachidonic acid, PAF, collagen, ionophore A23187 and thrombin in washed rabbit platelets. It showed a weak activity in platelet-rich plasma. Osthole inhibited the thromboxane B2 formation caused by arachidonic acid, collagen, ionophore A23187 and thrombin in washed platelets, and also the thromboxane B2 formation caused by the incubation of lysed platelet homogenate with arachidonic acid. The generation of inositol phosphates in washed platelets caused by collagen, PAF and thrombin was suppressed by osthole. These data indicate that the inhibitory effect of osthole on platelet aggregation and release reaction was due to the inhibition of thromboxane formation and phosphoinositides breakdown.     

Effect of the calcium-entry blocking agent nifedipine on activation of human platelets and comparison with verapamil

The effects of the calcium-entry blocking agent nifedipine on the activation of human platelets by various agonists has been studied and compared with verapamil. Like verapamil, nifedipine inhibited platelet aggregation and secretion caused by collagen, the second phase of ADP-induced aggregation, and aggregation caused by the ionophore A23187. Both agents inhibited the formation of TXB2 from endogenous arachidonate, whereas only nifedipine inhibited platelet aggregation and decreased TXB2 formation caused by exogenous arachidonate without inhibiting uptake. These results indicate that both calcium-blocking agents may be inhibiting the release of arachidonate in platelets by phospholipases, and that nifedipine also inhibits the formation and action of thromboxane A2 in platelets. Epinephrine-induced aggregation was inhibited by low concentrations of verapamil while nifedipine only inhibited aggregation by epinephrine at much higher concentrations. It is suggested that low concentrations of verapamil inhibit epinephrine-induced aggregation by interacting with platelet alpha-adrenergic receptors, and that higher concentrations of both calcium-blocking agents inhibit platelet responses to other aggregating agents by preventing intracellular calcium mobilization.     

Effect of cobra venom phospholipase A2 on platelet aggregation in comparison with those produced by arachidonic acid and lysophophatidylcholine

Cobra venom phospholipase A₂ induced a biphasic effect on washed rabbit platelets. The first phase was a reversible aggregation which was dependent on stirring and extracellular calcium. The aggregation and thromboxane B₂ formation were inhibited by indomethacin, mepacrine, tetracaine and imipramine, while PGE₁ and sodium nitroprusside inhibited only the aggregation, but not the thromboxane B₂ formation. The second phase was an inhibitory effect on platelet aggregation induced by arachidonic acid, PAF, ADP or collagen but not that by thrombin or ionophore A23187. The longer the incubation time of cobra venom phospholipase A₂ with platelets, the more the inhibitory effect. The aggregating and anti-aggregating effects could be overcome by bovine serum albumin. Lysophosphatidylcholine (Lyso-PC) and arachidonic acid showed synergistic inhibition in platelet aggregation. Lyso-PC decreased thromboxane B₂ formation in platelets formed by collagen. The inhibitory effect of Lyso-PC on platelet aggregation was more marked at lower calcium concentrations. It is concluded that the aggregating effect of exogenous addition of venom phospholipase A₂ is due to thromboxane formation and the antiplatelet effect is similar to those produced by arachidonic acid and lysophosphatidylcholine.     

Inhibition of platelet aggregation by some flavonoids.

The inhibitory effects of five flavonoids on the aggregation and secretion of platelets were studied. These flavonoids inhibited markedly platelet aggregation and ATP release of rabbit platelets induced by arachidonic acid or collagen, and slightly those by platelet-activating factor. ADP-induced platelet aggregation was also suppressed by myricetin, fisetin and quercetin. The IC50 on arachidonic acid-induced platelet aggregation was: fisetin, 22 microM; kaempferol, 20 microM; quercetin, 13 microM; morin, 150 microM less than IC50 less than 300 microM. The thromboxane B2 formations were also inhibited by flavonoids in platelets challenged with arachidonic acid. Fisetin, kaempferol, morin and quercetin antagonized the aggregation of washed platelets induced by U46619, a thromboxane A2/prostaglandin endoperoxides mimetic receptor agonist. In human platelet-rich plasma, quercetin prevented the secondary aggregation and blocked ATP release from platelets induced by epinephrine or ADP. These results demonstrate that the major antiplatelet effect of flavonoids tested may be due to both the inhibition of thromboxane formation and thromboxane receptor antagonism.     

Inhibition of rabbit platelet aggregation by 1,4-naphthoquinones

The effects of four 1,4-naphthoquinone derivatives on the aggregation of rabbit platelets were examined. All the four 1,4-naphthoquinone derivatives inhibited the platelet aggregation of washed rabbit platelets induced by thrombin (0.1 U/ml) and the IC50 is: 2-chloro-3-methyl-1,4-naphthoquinone (CMN), 5 micrograms/ml; 3-methyl-5,8-dihydroxy-1,4-naphthoquinone, 13 micrograms/ml; 5,8-dihydroxy-1,4-naphthoquinone, 18 micrograms/ml; 3-methyl-1,4-naphthoquinone (vitamin K3), 53 micrograms/ml. CMN was the most potent in inhibiting the aggregation and release reaction induced by ADP, arachidonic acid, PAF, ionophore A23187, collagen and thrombin in a dose-dependent manner in washed platelets, platelet-rich-plasma and whole blood. The thromboxane B2 formation caused by collagen and ionophore A23187 was inhibited by CMN. However, the thromboxane B2 formation by arachidonic acid was markedly increased. The platelet inhibitory effect of CMN could not be antagonized either by raising the concentrations of extracellular Ca++ or by wash out. The phosphoinositides breakdown induced by thrombin was inhibited by CMN. Phospholipids (PE, PC, PI) could slightly antagonize the antiplatelet effect of CMN. It is concluded that the inhibitory effect of CMN on rabbit platelet aggregation may be due to the inhibition of phosphoinositides breakdown caused by the inducers.     

HA-29: an inhibitor of thromboxane A2 formation with antagonism of thromboxane A2/prostaglandin endoperoxide receptor in rabbit platelets.

HA-29, 2-[(3-methoxyphenyl)methyl]-pyrano[2,3-c]pyrazol-6(1H)-one, was investigated for its inhibitory mechanism of action in washed rabbit platelets. This compound inhibited the aggregation and ATP release of rabbit platelets induced by arachidonic acid and collagen in a concentration-dependent manner, without affecting those induced by ADP, PAF and thrombin. Prolongation of the incubation time of HA-29 with platelets did not cause further inhibition and the aggregability of the agent-treated platelets could be restored after washing of platelets. The concentration-response curve of U-46619-induced platelet aggregation was shifted to the right by HA-29 in a concentration-dependent manner, but the maximal aggregation was suppressed by HA-29. The pA2 and pA10 values of HA-29 on U-46619-induced platelet aggregation were 4.26 and 3.58, respectively, with a slope value of -1.4. The U-46619-induced aggregation was markedly disaggregated by HA-29 even it was added 5 min after U-46619. HA-29 inhibited the secondary aggregation and ATP release, but not the primary aggregation of human platelet-rich plasma induced by ADP and epinephrine. Thromboxane B2 formation caused by arachidonic acid, collagen and thrombin was markedly suppressed by HA-29. HA-29 also inhibited the formation of prostaglandin D2 caused by arachidonic acid. HA-29 inhibited almost completely the formation of inositol monophosphate caused by U-46619, but not that by collagen or thrombin. HA-29 did not affect U-46619-induced contraction of rat aorta. It is concluded that the antiplatelet effect of HA-29 is due to the inhibition of thromboxane A2 formation and blockade of thromboxane A2/prostaglandin endoperoxide receptor.     

In vitro effects of ethanol on rabbit platelet aggregation, secretion of granule contents, and cyclic AMP levels in the presence of prostacyclin

Ethanol, at physiologically tolerable concentrations, inhibits platelet responses to low concentrations of collagen or thrombin, but does not inhibit responses of washed rabbit platelets stimulated with high concentrations of ADP, collagen, or thrombin. However, when platelet responses to high concentrations of collagen or thrombin had been partially inhibited by prostacyclin (PGI2), ethanol had additional inhibitory effects on aggregation and secretion. These effects were also observed with aspirin-treated platelets stimulated with thrombin. Ethanol had no further inhibitory effect on aggregation of platelets stimulated with ADP, or the combination of ADP and epinephrine. Thus, the inhibitory effects of ethanol on platelet responses in the presence of PGI2 were very similar to its inhibitory effects in the absence of PGI2, when platelets were stimulated with lower concentrations of collagen or thrombin. Ethanol did not appear to exert its inhibitory effects by increasing cyclic AMP above basal levels and the additional inhibitory effects of ethanol in the presence of PGI2 did not appear to be brought about by further increases in platelet cyclic AMP levels.     

In vitro effects of acetaminophen and its analogues on human platelet aggregation and thromboxane B2 synthesis

We examined the effect of acetaminophen and the structural analogues 2,6-dimethylacetaminophen, 3,5-dimethylacetaminophen, and N-acetyl-p-benzoquinone imine on human platelet aggregation, 14C-serotonin secretion, and thromboxane B2 synthesis. Preincubation with 1 mM acetaminophen for 2 min completely inhibited arachidonic acid- and collagen-stimulated platelet aggregation. Thromboxane B2 production and 14C-serotonin secretion by arachidonic acid-stimulated platelets also were completely inhibited. Preincubation of platelets with 1 mM 3,5-dimethylacetaminophen inhibited collagen and arachidonic acid-induced aggregation and arachidonic acid-stimulated thromboxane B2 synthesis, while treatment with 2,6-dimethylacetaminophen did not inhibit aggregation and blocked thromboxane B2 formation to a much lesser degree. Preincubation with 1 mM N-acetyl-p-benzoquinone imine inhibited arachidonic acid-induced aggregation and 14C-serotonin secretion but had no effect on arachidonic acid-induced thromboxane B2 formation and collagen-induced platelet aggregation.     

Inhibitory effect of hexapeptide (RGRHGD) on platelet aggregation

The B chain of beta-bungarotoxin 1-6 sequence, RGRHGD, presents the highest local average hydrophilicity measured by Kyte and Doolittle modeling analysis. The RGRHGD holds parts of both RGD and KGD peptides, which have been reported as having high binding affinity to GPIIb-IIIa. The present study evaluates whether the synthesized hexapeptide, RGRHGD, has an antiplatelet effect and further elucidates the possible mechanisms of action. RGRHGD dose-dependently inhibited rabbit platelet aggregation and adenosine triphosphate release induced by arachidonic acid, collagen, platelet-activating factor, thrombin, or U46619 with the IC50 range of 82.7 to 510 microg/mL. The platelet thromboxane B2 formation induced by collagen or thrombin was also significantly decreased by RGRHGD, but there was no effect on arachidonic acid-induced thromboxane B2 formation. In addition, RGRHGD also inhibited the rise of intracellular calcium level stimulated by arachidonic acid, collagen, or thrombin in Fura 2-AM-loaded platelets. The adenosine 3',5'-cyclic monophosphate level of washed platelets was not affected by RGRHGD. In conclusion, these data indicate that the inhibitory effect of RGRHGD on platelet aggregation may be due to the attenuation of thromboxane A2 formation and intracellular calcium mobilization. In addition, this study may provide a useful method of finding potential therapeutic agents by using molecular modeling analysis.     

Inhibition of platelet aggregation by ethanol in vitro shows specificity for aggregating agent used and is influenced by platelet lipid composition

Ethanol at physiologically tolerable concentrations inhibited platelet aggregation in vitro in a relatively specific way, which may be influenced by platelet membrane lipid composition. Aggregation to collagen, calcium ionophore A23187 and thrombin (low doses) were often markedly inhibited by ethanol, adrenaline and ADP responses were little affected, and aggregation to exogenous arachidonic acid was actually potentiated by ethanol. Aggregation to collagen, thrombin and A23187 was inhibited more by ethanol in platelets enriched with saturated fatty acids than in those enriched with unsaturated fats. Platelets enriched with cholesterol showed increased sensitivity to ADP, arachidonate and adrenaline but this increase in cholesterol content did not appear to influence the inhibition by ethanol of platelet responses. The results suggest that ethanol may inhibit aggregation by an effect on membrane fluidity and/or calcium mobilization resulting in decreased activity of a membrane-bound phospholipase.     

Effect of amino sugars on platelet aggregation and on fibrinogen binding

The amino sugars glucosamine, galactosamine and mannosamine (30 mM) inhibited aggregation of human or rabbit platelets induced by ADP, collagen, thrombin, PAF or high concentrations of sodium arachidonate. 125I-fibrinogen binding during ADP-induced aggregation, and release of amine storage granule contents were also inhibited. Increasing the calcium concentration of the suspending medium to 5 mM did not overcome the inhibitory effect on the release reaction. The amino sugars deaggregated rabbit platelets that had been aggregated by ADP, collagen or thrombin, but deaggregated human platelets readily only when ADP was used as the aggregating agent. Fibrinogen-induced aggregation of chymotrypsin-treated platelets was blocked by the amino sugars. They did not inhibit platelet adherence to a collagen-coated glass surface, nor affect release of granule contents from the adherent platelets. Aggregation and release induced by low concentrations of sodium arachidonate or the divalent cation ionophore A23187 were potentiated, indicating that the effects of the amino sugars on platelets are more complex than simple inhibition of the lectin-like activity that becomes available on the surface of platelets that have undergone the release reaction. One of the effects of the amino sugars, however, is interference with the binding of fibrinogen to platelets. The effects of the amino sugars are shared by other primary amines.     

Two antiplatelet agents from Magnolia officinalis

Magnolol and honokiol are two position isomers isolated from the bark of Magnolia officinalis. Both inhibited the aggregation and ATP release of rabbit platelet-rich plasma induced by collagen and arachidonic acid without affecting that induced by ADP, PAF or thrombin. Aggregation of washed platelets was more markedly inhibited than that of platelet-rich plasma, while the aggregation of whole blood was least affected by both inhibitors. Thromboxane B2 formation caused by collagen, arachidonic acid or thrombin was in each case inhibited by magnolol and honokiol. The rise of intracellular calcium caused by arachidonic acid or collagen was also suppressed by both agents. Collagen-induced intracellular calcium increase in the presence of indomethacin was suppressed by magnolol. It is concluded that the antiplatelet effect of magnolol and honokiol is due to an inhibitory effect on thromboxane formation and also an inhibition of intracellular calcium mobilization.     

Effects of Ethanol on Platelet Responses Associated with Adhesion to Collagen

Adhesion of platelets to collagen in damaged blood vessels or ruptured atherosclerotic plaques is important in hemostasis and arterial thrombosis. Adhesion to collagen results in secretion of granule contents and formation of thromboxane A₂; thromboxane A₂ and released ADP synergistically promote aggregation around platelets adherent to collagen. Ethanol inhibits collagen-induced platelet aggregation, secretion, arachidonate mobilization, and thromboxane A₂ formation but does not inhibit platelet adhesion to de-endothelialized rabbit aortae. We investigated whether ethanol affects the initial signalling events and responses of platelets adherent to collagen, independent of the actions of secondary agonists. Suspensions of washed human platelets, labelled by incorporation of [³H]oleate into phospholipids, were used to measure platelet adhesion to collagen by a filtration method; studies were done in the presence of an ADP-removing system and blockers of receptors for thromboxane A₂, platelet-activating factor, serotonin, and fibrinogen. Ethanol (87 mM) did not affect the rate or extent of platelet adhesion to collagen or secretion of [¹⁴C]serotonin from prelabelled platelets adherent to collagen, but ethanol did inhibit thromboxane A₂ formation. Previous studies showed that ethanol does not affect platelet stimulation by arachidonate, leading to the suggestion that reduced mobilization of arachidonate, rather than inhibition of its conversion to thromboxane A₂, is responsible for inhibition by ethanol of thromboxane A₂ formation. Here, we show by a gel mobility shift assay and immunoblotting, that ethanol delays the collagen-induced increase in the phosphorylation of cytosolic phospholipase A₂, the enzyme responsible for arachidonate mobilization. However, ethanol has no effect on collagen-induced tyrosine phosphorylation of phospholipase Cγ2, determined by immunoprecipitation and immunoblotting. Thus, ethanol's effect on signal transduction in collagen-adherent platelets occurs distal to phosphorylation of phospholipase Cγ2 but proximal to phosphorylation of cytosolic phospholipase A₂.     

Inhibitory mechanisms of dantrolene on platelet aggregation

The antiplatelet effect of dantrolene and possible inhibitory mechanisms were studied in rabbit platelets. Preincubation of rabbit washed platelets with dantrolene (50-300 microM) inhibited the platelet aggregation and adenosine triphosphate release induced by arachidonic acid (100 microM), collagen (10 microg/mL), or thrombin (0.1 U/mL) in a dose-dependent manner. The thromboxane B2 formation caused by collagen or thrombin was inhibited by dantrolene, while formation of thromboxane B2 and prostaglandin D2 induced by arachidonic acid were not inhibited. In addition, the formation of phosphoinositide breakdown and the rise of intracellular calcium level induced by collagen or thrombin were also inhibited in a dose-dependent manner by dantrolene in the presence of indomethacin (2 microM). However, the platelets cyclic AMP level was not affected by dantrolene. In conclusion, the present study demonstrates that dantrolene inhibits platelet activation mainly due to suppression of phosphoinositide breakdown.     

Antiplatelet actions of some coumarin compounds isolated from plant sources.

Xanthyletin, xanthoxyletin, suberosin (all from Citrus grandis), aurapten (from Severinia huxifolia) and poncitrin (from Poncirus trifoliata) were isolated and their chemical structures were characterized to be coumarin compounds. All these coumarin compounds except xanthyletin inhibited the aggregation and ATP release of rabbit platelets induced by arachidonic acid, collagen, ADP, platelet-activating factor (PAF) or U46619 (a thromboxane A2 analog). Thrombin-induced ATP release, but not the aggregation, was also inhibited by these compounds. Xanthyletin inhibited only platelet aggregation induced by arachidonic acid and collagen, while poncitrin inhibited that caused by PAF more markedly than other coumarin compounds. The thromboxane B2 formation in washed platelets caused by arachidonic acid and collagen was suppressed by these coumarin compounds. The phosphoinositide breakdown caused by collagen and PAF was also inhibited by these compounds. They did not affect fibrinogen-induced aggregation of elastase-treated platelets. These antiplatelet actions were immediate, reversible by washout and independent on the incubation time (except suberosin). Antiaggregating effect was also studied by an electrical impedance method and the inhibitory effect of coumarin compounds on the whole blood and platelet-rich plasma was much less than that of platelet suspension in the aggregation induced by collagen, PAF and ADP. It is concluded that the antiplatelet actions of these coumarin compounds are due to the inhibition on thromboxane A2 formation and phosphoinositides breakdown.     

Suppression by beta-lactam antibiotics of thromboxane A2 generation and arachidonic acid release in rabbit platelets in vitro

High concentrations of latamoxef and some other beta-lactam antibiotics suppressed thromboxane A2 generation as determined from the thromboxane B2 level in the in vitro aggregation of rabbit platelets in agonist-induced stimulations. In aggregation induced with a low concentration of collagen, the suppression of thromboxane B2 generation correlated well with the suppressions of aggregation and serotonin (5-HT) release; collagen produced thromboxane A2-dependent aggregation and 5-HT release as judged from the inhibitory action of indomethacin. Latamoxef also suppressed thromboxane B2 generation when platelet stimulation was induced by thrombin or platelet activating factor at concentrations at which it did not affect either aggregation or 5-HT release. However, latamoxef did not affect platelet responses including thromboxane B2 generation induced by exogenous arachidonic acid or calcium ionophore, A23187. Beta-lactam antibiotics also interfered with arachidonic acid release from the membrane phospholipids of platelets which had been prelabelled with [3H]arachidonic acid and aggregated with collagen. These results suggest that in the in vitro aggregation of platelets, beta-lactam antibiotics interfere with some of the receptor-stimulated processes which lead to arachidonic acid release from the membranes and this, in turn, suppresses thromboxane B2 generation.     

Inhibition of thrombin- and collagen-induced phosphoinositides breakdown in rabbit platelets by a PAF antagonist--denudatin B, an isomer of kadsurenone

Denudatin B, an isomer of kadsurenone, was isolated from Magnolia fargesii. It inhibited the aggregation and ATP release of washed rabbit platelets caused by platelet-activating factor (PAF) in a concentration-dependent manner. The IC50 on PAF (2 ng/ml)-induced aggregation was about 10 micrograms/ml. High concentration of denudatin B (greater than 50 micrograms/ml) also inhibited the aggregation and ATP release of platelets caused by ADP, collagen, arachidonic acid and thrombin. However, shape change of platelets still existed. Prolongation of the incubation time with platelets could not cause further inhibition, and the aggregability of platelets could be restored after denudatin B was washed out from platelets. Thrombin-induced thromboxane B2 formation was almost completely suppressed. In the absence of extracellular calcium (EGTA 1 mM), ATP release caused by thrombin was inhibited. Thrombin-induced rise of the intracellular calcium concentration was suppressed by denudatin B, but not by BN52021 or kadsurenone. The generation of inositol phosphate in washed platelets caused by collagen, PAF and thrombin was also suppressed. The data indicate that PAF antagonist denudatin B has nonspecific antiplatelet action at high concentration by inhibiting phosphoinositides breakdown induced by collagen and thrombin.     

Pharmacological profile of G619, a new platelet aggregation inhibitor.

G619, a 4-OH-isophthalic acid derivative, was studied for its capacity to inhibit platelet aggregation. G619 dose-dependently inhibited U46619, collagen, ADP, PAF, thrombin and epinephrine-induced platelet aggregation in vitro. The IC50 values for inhibition of U46619-induced human and rabbit platelet aggregation were 39 and 43 microM, respectively. G619, at 100 microM, inhibited high concentration collagen (10 micrograms/ml)-induced aggregation of rabbit platelets pretreated with indomethacin and increased the level of cAMP in washed rabbit platelets by 30% (p less than 0.01 vs basal). However, G619, did not inhibit fibrinogen binding to GPIIb/IIIa receptor, phosphodiesterase, U46619-induced contractile responses on canine saphenous vein or rabbit aorta, calcium-induced vasoconstriction and thrombin or PAF-induced elevation of [Ca++]i in platelets in vitro. In vivo, the U46619-induced maximal thrombocytopenia in rats was reduced from 40% (vehicle) to 22% and 18% by 10 and 30 mg/kg of G619 i.v., respectively. G619 (30 mg/kg) had no effect on the U46619-induced vasopressor response or sudden death in rats, and had no effect on TxB2 formation. Our results indicate that G619 is a broad-spectrum platelet aggregation inhibitor and may have its effect on a common mechanism for platelet aggregation besides an effect on the thromboxane A2 receptor.     

Influence of granulocyte elastase-like proteinase (ELP) on platelet functions

The influence of granulocyte elastase-like proteinase (ELP) on platelet functions was investigated. ELP inhibited the platelet aggregations induced by a wide variety of agonists. The inhibition was marked in the case of receptor-mediated agonists such as thrombin, ristocetin, etc. It was moderate with the pervading agonist, arachidonic acid, and mild with the bypassing agonist, Ca²⁺ ionophore A23187. ELP inhibited the release of thromboxane A2 from platelets in the case of the platelet aggregation induced by thrombin. On the other hand, ELP did not inhibit the release of thromboxane A2 from platelets in the platelet aggregation induced by arachidonic acid or Ca²⁺ ionophore A23187. ELP suppressed the release of serotonin from platelets induced by thrombin, while it did not markedly suppress the release of serotonin induced by Ca²⁺-ionophore A23187. Treatment of platelets with ELP resulted in a slight increase of intraplatelet cAMP levels. These results suggest that ELP acts on receptors and inhibits platelet functions. As a results, ELP markedly inhibits the platelet functions such as aggregation or release of serotonin or thromboxane A2 stimulated by receptor-mediated agonists. ELP slightly elevates the CAMP level in the platelets, resulting in the mild inhibition of the platelet functions stimulated by the pervading agonist, arachidonic acid, or the bypassing agonist, Ca²⁺-ionophore A23187.