Effects of halothane on transport of 5-hydroxytryptamine by platelet membranes.

Na+-dependent uptake of 5-HT (5-hydroxytryptamine) into plasma membrane vesicles derived from bovine blood platelets and ATP-dependent 5-HT uptake into storage vesicles in platelet lysates were measured. Na+-dependent uptake was temperature-dependent, inhibited by imipramine and exhibited Michaelis-Menten kinetics (apparent Km, 0.12 +/- 0.02 microM; Vmax. 559 +/- 54 pmol/min per mg of protein. Halothane had no effect on Na+-dependent transport of 5-HT in plasma-membrane vesicles. ATP-dependent 5-HT transport into storage granules also exhibited Michaelis-Menten kinetics (apparent Km 0.34 +/- 0.03 microM; Vmax. 34.3 +/- 1.7 pmol/min per mg of protein) and was inhibited by noradrenaline (norepinephrine), but not by imipramine. Exposure of the granules to halothane resulted in a progressive decrease in Vmax. The results demonstrate a possible site for disruption of platelet function by anaesthetics.     

Influence of histone-chondroitin sulphate A complexes on blood platelet aggregation

The influence of histones and their complexes with chondroitin sulphate A on platelet aggregation was studied. Histones F1 and F2a1 and complex histone F1-chondroitin sulphate A do not induce platelet aggregation by themselves but they decreased threshold aggregating concentration of ADP whereas complex histone F2a1-chondroitin sulphate A increased it. The results of our study may point out to the possibility of the examined substances taking part in disturbances of the platelet function.     

Influence of liposomal local anesthetics on platelet aggregation in vitro

We assessed the effect of local anesthetics (LA) from different families such as esters (benzocaine), linear aminoamides (lidocaine) and cyclic aminoamides (bupivacaine) on the platelet aggregation induced by ADP. Liposomal formulations of the three LA, prepared with egg phosphatidylcholine:cholesterol alpha-tocopherol, were also tested. The three LA were able to inhibit platelet aggregation induced by ADP, in the following order: bupivacaine > lidocaine > benzocaine. After encapsulation into liposomes the inhibitory effect increased for all anesthetics studied, showing that aggregation tests could be used to assess the toxicity of new drug formulations.     

Mechanism of imipramine inhibition of platelet 5-hydroxytryptamine transport.

Plasma membrane vesicles isolated from porcine blood platelets take up approximately 8 to 15 pmol of [3H]imipramine per mg of membrane protein. This apparent binding requires Na+ in the external medium and is reversed by 5-hydroxytryptamine and fluoxetine. The apparent KD for imipramine uptake is 23 nM, which agrees well with the KI for competitive inhibition of 5-hydroxytryptamine transport by imipramine. In contrast to 5-hydroxytryptamine transport, imipramine uptake is not dependent on transmembrane Na+ and K+ gradients and is insensitive to ionophores such as nigericin and gramicidin which dissipate these gradients. Although 5-hydroxytryptamine rapidly and competitively displaces imipramine from membrane vesicles, imipramine does not cause 5-hydroxytryptamine efflux and inhibits 5-hydroxytryptamine exchange. These results are consistent with the proposal that imipramine binds to the substrate site of the 5-hydroxytryptamine transporter but cannot be transported.     

Effect of 5-HT2-selective agonists on cat platelet aggregation

The effects of the 5-HT2-sselective agonists 1-(4-bromo-2, 5-dimethoxyphenyl) -2- aminopropane (DOB) and 1- (2, 5-dimethenyl4-iodophenyl) -2- aminopropane (DOI) on cat platelet aggregation were investigated and compared with those produced by serotonin (5-HT) and a positional isomer of DOB (i.e., isoDOB). Serotonin, DOB, and DOI enhanced the aggregation of platelets induced by a suboptimal concentration of ADP. This effect was completely inhibited by pre-incubation of the platelet suspension with the 5-HT2-selective antagonist ketanserin. IsoDOB, an isomer of DOB with a very low affinity for central 5-HT2 binding sites, was inactive in the platelet aggregation assay. The present results are consistent with the proposed role of 5-HT2 receptors in serotonin-induced platelet aggregation     

Effect of nucleoside 5'-(glycosyl pyrophosphates) on platelet aggregation

The effect of the nucleoside 5′-(glycosyl pyrophosphates) on the aggregation of human blood platelets has been studied in vitro. Devoid of any aggregating effect, ADP-ribose was shown to act as a moderate inhibitor on the ADP-induced aggregation. After a preincubation without stirring, the inhibitory effect of ADP-ribose increased and total inhibition was reached after 20 min. On the contrary, ADP-mannose caused platelet aggregation in citrated plasma. During the course of a 20-min incubation, ADP-mannose progressively lost the aggregating effect and showed total inhibition. At first, ADP-glucose showed a slight aggregating effect, but after a 20-min incubation it also became inhibitory. The UDP- and GDP-sugars have no effect on aggregation, and thus may be used in the study of glycoprotein biosynthesis by platelets.     

Inhibition of platelet aggregation by acyl-CoA thioesters

The aggregation of platelets induced by ADP, collagen, or epinephrine in human platelet-rich plasma is inhibited by long chain acyl-CoA thioesters. Palmityl-CoA exerts a concentration dependent inhibition of collagen-induced aggregation of the primary and secondary waves of ADP-induced aggregation. Palmitlyl-CoA also inhibits the secondary wave of epinephrine-induced aggregation but has no effect on the primary wave. The inhibitory effect of palmityl-CoA can be reversed by addition of excess ADP and cannot be attributed to a detergent action.     

Reversal of platelet aggregation by aortic microsomes

The microsomal fraction of dog aortas inhibited human platelet aggregation induced by arachidonic acid, ADP, or thrombin. When aortic microsomes were added to a preparation of irreversibly aggregated platelets, the aggregates dispersed after 4–6 minutes. The fact that aortic microsomes inhibit platelet aggregation induced by ADP suggests that its effect is probably on the cellular function of platelets and not in direct competition against thromboxane A₂.     

Inhibition of cyclooxygenase activity and platelet aggregation by epoxyeicosatrienoic acids. Influence of stereochemistry

Certain epoxyeicosatrienoic acids (EETs) that were not cyclooxygenase substrates were effective cyclooxygenase inhibitors. Both (+/-)-14,15-cis-EET and (+/-)-8,9-cis-EET inhibited purified enzyme at concentrations from 1 to 50 microM; (+/-)-11,12-cis-EET was ineffective at concentrations below 100 microM. For the case of 14,15-cis-EET, only the (14R,15S)-stereoisomer was active. Other isomers including (14S,15R)-cis-EET, (14R,15R)-trans-EET, (14S,15S)-trans-EET, and the erythro and threo vicinal 14,15-diols were inactive. In addition to their effects on isolated enzyme preparations, cyclooxygenase activity in platelet suspensions, reflected by thromboxane B2 formation, was also inhibited by (14R,15S)-cis-EET and (+/-)-8,9-cis-EET but not by the other isomers. Thus potency and stereospecificity requirements were maintained for cyclooxygenase within intact platelets. Unlike the stereospecific inhibition of the cyclooxygenase enzyme, platelet aggregation induced by arachidonic acid was inhibited by all EET isomers at concentrations from 1 to 10 microM with no evident stereospecificity. Inhibition of aggregation was not uniformly associated with inhibition of thromboxane B2 formation; ordinarily, these two parameters correlate closely. This dissociation was not maintained for another biochemical process involved in platelet activation. For instance, there was a uniform correlation between inhibition of phosphorylation of a 40-kDa platelet protein and inhibition of aggregation. Our results suggest that effects of EET may originate from either stereospecific or nonspecific mechanisms. Definition of such mechanisms may be important to appreciate any physiological relevance of these substances.