The relationship between intracellular second messengers and platelet secretion.
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Adenosine Triphosphate↗
Biomedical subjects
M C Scrutton
Publications and source records attributed to M C Scrutton.
Mammalian platelet adrenoceptors.
Mammalian platelets vary widely in their responses to catecholamines in part because these agonists can act via excitatory alpha- and inhibitory beta-adrenoceptors. In the absence of antagonists, adrenaline enhances the response of rabbit platelets to an excitatory agonist, e.g. adenosine-5'-O-(1-thiodiphosphate) (ADP-alpha-s) acting at another receptor, but has no effect on the response of rat or guinea pig platelets to such an agonist. In the presence of a beta-adrenoceptor antagonist, adrenaline enhances the response of rat, but not guinea-pig platelets to ADP-alpha-S and the extent of the enhanced effect on rabbit platelets is increased. In the presence of an alpha-adrenoceptor antagonist, adrenaline inhibits the response of rabbit and rat platelets to ADP-alpha-S but has no such effect on the response of guinea-pig platelets. Studies using selective agonists and antagonists demonstrate that the excitatory response of rat platelets to adrenaline is mediated by an alpha 2-adrenoceptor, and the inhibitory response of rabbit platelets to adrenaline by a beta 2-adrenoceptor as is the case in other species which have been examined. The mean alpha 2-adrenoceptor density on human, rabbit, rat and guinea-pig platelets as assessed using [3H]-yohimbine as radioligand is obtained as 258, 270, 42 and less than 10 receptors per platelet. The mean beta 2-adrenoceptor density on human, rabbit, rat and guinea-pig platelets as assessed using (-)-[125I]-iodocyanopindolol is obtained as 66, 14, 41 and less than 5 receptors per platelet. The nature of the effect of adrenaline on the response of mammalian platelets to other excitatory agonists, e.g. ADP-alpha-S, appears therefore to be largely determined by the absolute number of alpha 2-adrenoceptors and by the relative content of alpha 2- and beta 2-adrenoceptors on these cells for the species which have been examined in this analysis.
Animals↗
Effects of alpha 2-adrenoceptor agonists and of related compounds on aggregation of, and on adenylate cyclase activity in, human platelets.
A range of 2-(,5-dihydroimidazolyl)-benzene, -quinoline, and -quinoxaline derivatives and 2-morpholino-4-catechol have been characterized as agonists or partial agonists for human platelet aggregation; and for inhibition of adenylate cyclase by measurement of their effect on platelet [cyclic-3',5'-AMP]. Antagonist activity for these compounds versus adrenaline as agonist has also been assessed for these two responses. The compounds can be divided into 4 groups. Group I contains compounds that are agonists for both responses; group II, compounds that are agonists for inhibition of adenylate cyclase but antagonists for the aggregatory response; group III, compounds that are agonists for the aggregatory response but are antagonists for inhibition of adenylate cyclase by adrenaline; and group IV, compounds that are antagonists for both responses. In group I the EC50 values for induction of aggregation are not significantly different from the EC50 values for inhibition of adenylate cyclase except for 2-morpholino-4-catechol which is significantly more potent as an inhibitor of adenylate cyclase. In group IV a linear correlation is observed between the K1 values for the two responses for 8 compounds but 2 other compounds do not conform to this correlation. The data are not consistent with a model in which a single chi 2-adrenoceptor mediates both the aggregatory response and inhibition of adenylate cyclase and hence support a model in which unique chi 2-adrenoceptors mediate these two responses.
Adenosine Diphosphate↗
Some properties of the human platelet vasopressin receptor.
Aggregation of human platelets by vasopressin is potently inhibited by 1-[beta mercapto-(beta, beta'-cyclopentamethylene propionic acid)]-L-arginine vasopressin, a selective vasopressor (V1) antagonist. 1-Desamino-8-D-arginine-vasopressin, a selective anti-diuretic (V2) agonist failed to induce aggregation and acted as a weak antagonist. Vasopressin analogues which lacked the N-terminal amino group or which contained an uncharged amino acid residue at position 8 acted as partial agonists for the human platelet. The response to such partial agonists could be enhanced by increasing the cytosolic Ca2+ concentration but not by altering the level of cyclic-3', 5'-AMP. These observations provide further evidence indicating that the platelet vasopressin receptor is of the V1 sub-type.
Adenosine Diphosphate↗
The role of 1-O-alkyl-2-acetyl-sn-glyceryl-3-phosphorylcholine (AcGEPC) and palmitoyl-lysophosphatidate in the responses of human blood platelets to collagen and thrombin.
Desensitisation of human blood platelets to the effects of 1-O-alkyl-2-acetyl-sn-glyceryl-3-phosphorylcholine (1-O-alkylAcGEPC) and palmityl-lysophosphatidate by pre-incubation with these agonists has no effect on the aggregatory or secretory responses to collagen but causes 30-40% inhibition of these responses to thrombin in aspirin-treated platelets. The effects of 1-O-alkylAcGEPC and palmitoyl-lysophosphatidate are not additive. The results are not consistent with the proposal that 1-O-alkylAcGEPC or lysophosphatidate are the mediators for the responses to collagen observed when prostaglandinendoperoxide synthesis is prevented, although they may play some role in the responses to thrombin under these conditions.
Adenosine Triphosphate↗
Synergistic interaction and platelet inhibitory agonists.
Marked synergistic interaction is observed between two platelet excitatory agonists neither of which cause inhibition of adenylate cyclase. No synergistic interaction is observed between the platelet inhibitory agonists PGI2, PGD2 and adenosine nor between ADP and adrenaline when acting as inhibitory agonists for adenylate cyclase. The studies suggest that synergistic interaction between platelet agonists may be restricted to the excitatory responses.
Adenosine↗
The sub-cellular localisation of pyruvate carboxylase and of some other enzymes in Aspergillus nidulans.
The sub-cellular localisation of enzymes has been defined by latency analysis, and fractionation by differential centrifugation, in cell-free extracts prepared from the mycelium of Aspergillus nidulans by growth in the presence of 2-deoxyglucose followed by treatment with a mixture of beta-glucuronidase, sulphatase and beta-glucanase and exposure to N2 cavitation at 5.2 PMa. In such extracts pyruvate carboxylase and NAD-dependent and NADP-dependent glutamate dehydrogenases are exclusively localised in the cytosol whereas all the other enzymes studied have sub-cellular localisation patterns similar to those described for mammalian liver. Electrophoretic analysis has established the presence of unique mitochondrial and cytosolic isoenzymes for many of the enzymes, e.g. NAD--malate dehydrogenase, NADP--isocitrate dehydrogenase, glutamate/oxaloacetate transaminase, fumarase, which show a marked extent of incomplete latency and the presence of significant activity in the mitochondrial and cytosolic fractions prepared by differential centrifugation. A novel method is described for detection of citrate synthase activity following electrophoresis of the cell-free extract. Application of this method confirms the absence of a unique cytosolic isoenzyme of citrate synthase and hence shows that citrate synthase activity detected in the soluble fraction results from damage to the mitochondria during isolation. A scheme is proposed on the basis of these data to describe the organisation of lipid and amino acid synthesis from glucose in an organism which possesses a cytosolic pyruvate carboxylase.
Aspergillus nidulans↗
Binding of [3H]-dihydroalprenolol and [3H]-acetobutolol to human blood platelets is not related to occupancy of beta-adrenoceptors.
Binding of [3H]-dihydroalprenolol to human platelet lysates is inhibited by (+/-)-propranolol and (+/-)-butoxamine, but less effectively by (+/-) practolol. (-)-Isoprenaline causes no significant inhibition of binding where stimulation of adenylate cyclase can be shown. Binding of [3H]-acetobutolol is also inhibited by (+/-)-propranolol. "Specific" binding of [3H]-dihydroalprenolol and [3H]-acetobutolol defined by (+/-) propranolol shows a non-classical saturation curve. 50% maximal binding is observed in the range 15-25 mM. The extent of "specific" binding is 2-fold greater for [3H]-dihydroalprenolol. Similar and rapid rates of binding of [3H]-dihydroalprenolol are observed at 4 degrees C and 20 degrees C. No stereoselectivity is observed for inhibition of [3H]-dihydroalprenolol binding by (+) and (-)-propranolol. Binding of [3H]-dihydroalprenolol and [3H]-acetobutolol may relate to the lipophilic character of these radioligands and does not represent interaction with beta-adrenoceptors.
Acebutolol↗
Platelet beta-adrenoceptors.
Inhibition by isoprenaline of the aggregatory response of human and rat platelets induced by various excitatory agonists is blocked by beta 2-adrenoceptor antagonists. beta 1-Adrenoceptor antagonists are ineffective. beta 2-Adrenoceptor agonists cause inhibition of the response of human platelets to various excitatory agonists. The maximal extent of inhibition is less than that observed for isoprenaline. beta 1-Adrenoceptor agonists fail to cause detectable inhibition of this response. Neither beta 1 nor beta 2-adrenoceptor agonists cause inhibition of the response of rat platelets to excitatory agonists. Only beta 2-adrenoceptor agonists block the inhibitory response to isoprenaline. The extent of inhibition by isoprenaline is a function of the excitatory agonist used and in human platelets is correlated with the ability of that agonist to suppress elevated platelet cyclic adenosine 3',5'-monophosphate (cyclic AMP) levels. Inhibition by isoprenaline is prevented in the presence of an inhibitor of adenylate cyclase. Isoprenaline increases platelet cyclic AMP levels with an EC50 similar to that required to observe inhibition of the aggregatory response. These data indicate that human platelets carry beta 2-adrenoceptors whose occupancy causes inhibition of the response to excitatory agonists as a consequence of elevation of platelet cyclic AMP. The beta-adrenoceptor present on rat platelets also appears to be of the beta 2-subtype.
Adrenergic beta-Agonists↗
Effect of 781094, a new selective alpha-adrenoceptor antagonist, on the aggregatory responses of human blood platelets and on binding of [3H]-dihydroergocryptine to these cells.
781094 (2-(2(1, 4-benzodioxanyl))-2-imidazoline hydrochloride) is a potent competitive inhibitor of the aggregatory responses of human platelets induced by adrenaline (pA2 = 7.3) and UK-14304. 781094 is a more potent inhibitor of the pro-aggregatory response to clonidine than of that to methoxamine. The alpha 2/alpha 1-adrenoceptor selectivity ratio is 11.4. 781094 is a potent inhibitor of the binding of [3H]-dihydroergocryptine to platelet lysates. 781094 has no effect on the aggregatory responses of human platelets induced by adenosine-5'-pyrophosphate (ADP), 5-hydroxytryptamine, thrombin, U-46619, or arginine-vasopressin provided that the platelet-rich plasma does not exhibit enhanced responsiveness. In some instances high concentrations of 781094 potentiate the aggregatory response to collagen. In platelet-rich plasma which exhibits enhanced responsiveness, 781094 causes partial inhibition of the aggregatory responses to 5-hydroxytryptamine, ADP and vasopressin at concentrations similar to those required to inhibit the response to adrenaline. 781094 acts as a specific antagonist for the responses mediated by human platelet alpha-adrenoceptors and exhibits moderate selectivity for the alpha 2-adrenoceptors on this cell.
Adrenergic alpha-Antagonists↗
Synergistic responses and receptor occupancy in rabbit blood platelets.
Several observations indicate that simultaneous receptor occupancy is necessary for generation of a synergistic response of rabbit platelets to two excitatory agonists. First, an aggregatory response to adrenaline induced by prior addition of 5HT reverses rapidly unless stabilised by inhibition of 5HT uptake. The extent of response correlates with the extracellular 5HT concentration. Second, addition of an ADPase following adrenaline enhances the rate of disaggregation if the response to this agonist has been induced by prior addition of ADP. Third, disaggregation is induced, or enhanced, if an antagonist selective to the inducing agonist is added following completion of the induced aggregatory response. These data suggest marked lability in the mechanisms responsible for generation of synergistic responses.
Adenosine↗
Desensitisation in human and rabbit blood platelets.
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15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
Agonist selectivity and second messenger concentration in Ca2+-mediated secretion.
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Acetylglucosaminidase↗
Depressed responsiveness to adrenaline in platelets from apparently normal human donors: a familial trait.
Decreased responsiveness to adrenaline has been observed in five apparently normal unrelated human donors. In four of the donors this trait is inherited. Three of the donors, as well as their affected relatives, also exhibited depressed responsiveness to collagen and vasopressin but normal responsiveness to ADP and thrombin. The other two affected donors exhibit normal responsiveness to most other agonists. Normal responsiveness can be restored in all instances either by incubating the platelet-rich plasma at 20 degrees C or by addition of a low concentration of the divalent cation ionophore, A-23187. All affected platelets which have been examined have ATP and ADP contents, cholesterol to phospholipid ratios, and phospholipid class compositions within the normal range. Both the resting level of cyclic-3'5'-AMP and the ability of adrenaline to prevent elevation of cyclic-3',5'-AMP levels by prostaglandin E1 are normal. Mixing experiments demonstrate the absence of a circulating inhibitor of platelet function and suggest that the defect resides in the platelets. We conclude that the depressed responsiveness of human platelets to adrenaline may result from a defect in Ca2+ mobilization to the cytosol.
Adenosine Diphosphate↗
Pyruvate carboxylase from Aspergillus nidulans. Regulatory properties.
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Acetyl Coenzyme A↗
Direct evidence for a role for Ca2+ in amine storage granule secretion by human platelets.
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Blood Platelets↗
Positive interaction between agonists in the aggregation response of human blood platelets: interation between ADP, adrenaline and vasopressin.
ADP, adrenaline and vasopressin interact positively as agonists in aggregating human blood platelets in vitro. This interaction is maximal if the addition of two of the agonists is separated by 10--20 s but decreases rapidly at longer intervals especially at low agonist concentrations. The agonist concentrations at which positive interaction gives full aggregation are significantly less than those required for such a response to each agonist alone. The lowest concentrations at which adrenaline and vasopressin interact positively are at least two orders of magnitude greater than the normal blood concentrations of these hormones, and at least an order of magnitude greater than the concentrations achieved in pathological states. Specifically antagonizing the adrenaline and ADP receptors showed that the response was to the second agonist added to the system. An inhibitor of intracellular Ca2+ movement (tetracaine) is equally effective in blocking the responses generated by a single agonist or by interaction of two agonists. Inhibitors which increase cyclic-3',5'-AMP concentration (adenosine, prostaglandin E1, dipyridamole) are more effective against the response to a single agonist than that to agonist interaction. These data suggest that positive agonist interaction results from effects on the concentrations of second messengers within the platelet rather than from a direct interaction on the membrane receptors or the transmembrane coupling mechanisms.
Adenosine Diphosphate↗
Interaction of selective alpha-adrenoceptor agonists and antagonists with human and rabbit blood platelets.
1 The selectivity of alpha-adrenoceptors mediating the pro-aggregatory response of human and rabbit platelets to adrenaline and the conditions required to permit expression of an aggregatory response to partial agonists at these alpha-adrenoceptors have been studied.2 Yohimbine causes effective blockade of the pro-aggregatory responses whereas indoramin and prazosin are ineffective.3 The clonidine analogue, UK-14304, is nearly as effective as adrenaline in inducing an aggregatory response in human platelets and a pro-aggregatory response in rabbit platelets. Cross-tachyphylaxis between adrenaline and UK-14304 has been demonstrated.4 Clonidine is a weak agonist for the pro-aggregatory response of rabbit platelets and in some donors for the aggregatory response of human platelets.5 Methoxamine induces a pro-aggregatory response in human platelets which is blocked by indoramin or prazosin but not by yohimbine. No such response to methoxamine is observed in rabbit platelets.6 The divalent cation ionophore, A-23187, induces an aggregatory response to clonidine (in platelets from a non-responsive donor), phenylephrine and methoxamine in human platelets and to adrenaline, UK-14304 and clonidine in rabbit platelets. A secretory response to clonidine is also induced by A-23187 in human platelets.7 The adenylate cyclase inhibitor, SQ-22536, is ineffective in either inducing a response to the alpha-agonists or potentiating the effect of A-23187.8 The aggregatory responses to adrenaline and UK-14304 in rabbit platelets and to clonidine in human and rabbit platelets, which can be induced by A-23187, are blocked by yohimbine but not by prazosin or indoramin.9 From these studies we conclude that the pro-aggregatory responses of human and rabbit platelets to adrenaline are mediated primarily by alpha(2)-adrenoceptors. The presence of alpha(1)-adrenoceptors on human platelets is confirmed but these receptors do not appear to be present on rabbit platelets. The conditions required for expression of an aggregatory response to partial agonists at the human and rabbit platelet alpha-adrenoceptors implicate an increase in cytosolic Ca(2+) concentration as a key event in stimulus-response coupling but do not indicate such a role for depression of cyclic adenosine-3',5'-monophosphate concentration.
Adrenergic alpha-Agonists↗