The in vitro effects of nicotine and cotinine on prostacyclin and thromboxane biosynthesis.

The comparative effects of nicotine and cotinine on the biosynthesis of prostacyclin (PGI2) and thromboxane A2 (TXA2) in the horse aorta and platelet microsomes were studied. TXB2 and 6-keto PGF1a stable metabolites of TXA2 and PGI2 respectively were determined by radioimmunoassay. TXA2 production in the presence of either nicotine or cotinine treatment was not altered. However, a dose dependent inhibition of PGI2 biosynthesis, and a dose dependent stimulation of PGI2 biosynthesis, was observed in the presence of nicotine and cotinine respectively. Moreover, cotinine (10b3 M) was able to prevent the inhibitory effect of nicotine on PGI2 synthetase when preincubated with horse aorta microsomes. It appears that cotinine, the major nicotine metabolite resulting from a breakdown process, could be useful for the organism, at least for the cardiovascular system.

Binding of a thromboxane A2 (TXA2)/prostaglandin H2 (PGH2) receptor antagonist to rabbit and pig heart membrane protein.

This study aimed at testing the hypothesis that the binding sites for TXA2/PGH2 are present and different in the heart as compared to platelets and blood vessels. Kinetic studies on the thromboxane binding to protein of membrane preparations from rabbit and pig hearts were carried out using [125I]-PTA-OH, a potent specific thromboxane receptor antagonist. The following points are stressed: 1. the binding sites to 125I-PTA-OH were shown to be present in the heart membrane whatever the adopted experimental conditions were i.e. the temperature (4 or 30 degrees C) used for incubation the protein fractions under study: either the 105,000 g supernatant or the 200,000 g supernatant of the solubilized pellet (105,000 g) the animal species: rabbit and pig 2. the radioligand binding was rapid, saturable and reversible 3. the kinetically determined Kd's were in the picomolar range--11 and 14 pM for the rabbit and pig heart membrane preparation respectively--instead of the nanomolar range found in other tissues of the nanomolar range found in other tissues--27-39 nM and 2 nM for human platelets and bovine artery endothelial cells respectively- using the same thromboxane receptor antagonist.

Modifications in the TXA(2) and PGI(2) plasma levels and some other biochemical parameters during the initiation and development of non-insulin-dependent diabetes mellitus (NIDDM) syndrome in the rabbit.

Having developed a non-insulin-dependent diabetes mellitus (NIDDM) syndrome model in the rabbit using Wirsung duct ligation, it appeared interesting to use it to study the relationship between glycemia and the plasma levels of TXA(2)and PGI(2), and of some other biochemical parameters such as cholesterol, triglycerides, alkaline phosphatase and transaminases. A comparative study was carried out in the sham-operated rabbits (controls, C) and those having their pancreatic duct ligatured (NIDDM, D) at 15, 30, 40, 50 and 60 days post-ligation. On the 40th days, whereas in the controls, glycemia was 1.17 +/- 0.04 g.l(-1), it reached a maximum of 4.62 +/- 0.76 g.l(-1)(25.40 mM) in the NIDDMs. No significant modification was observed either in cholesterolemia or in triglyceridemia in either group. The GOT and GPT were highly increased, from 11.50 +/- 4.00 IU. l(-1)and 27.00 +/- 1.50 IU.l(-1)(C) to 37.50 +/- 5.64 IU.l(-1)(P<0. 001) and 58.50 +/- 7.50 IU.l(-1)(D) (P<0.001) in the NIDDM group, suggesting that hyperglycemia occurred simultaneously with the degeneration of the pancreatic tissue. In parallel, in D rabbits, the plasma levels of TXB(2)and 6 keto PGF(1alpha)were augmented to 68.22 +/- 6.20 pg.ml(-1)versus 22.49 +/- 5.74 pg.ml(-1)(C) (P<0.001), and 127.11 +/- 14.39 pg.ml(-1)versus 48.65 +/- 4.51 pg.ml(-1)(C) (P<0. 001) respectively. Statistical studies showed a significant correlation (P<0.05 and <0.02) between glycemia and the biosynthesis of eicosanoids under study. Moreover, 25 mM was found to be the threshold level of glucose excess essential to increase the TXA(2)and PGI(2)biosynthesis significantly. This supports the results obtained by other authors studying the action of glucose on phospholipase activity and consequent eicosanoid production.

Pharmacophore requirements in new series of pyridazinyl alkanoic acids, N-[(pyridazin-2-yl) alkyl] succinyl and glutaryl amides, inhibitors of thromboxane A2 biosynthesis.

New series of 5-benzyl-6-methyl-4-oxo pyridazin-2-yl alkanoic acids, N-[(pyridazin-2-yl)alkyl] succinyl and glutaryl amides have been synthesized and evaluated in vitro as TXA2 biosynthesis inhibitors. The experiments were carried out using arachidonic acid (32.8 microM) as a substrate and horse platelet microsomes as sources of TXA2 synthase. The presence of TXB2, a stable metabolite of TXA2, was determined by RIA. The potency of active compounds (1.10(-4) < IC 50 < 1.10(-6) M) greatly depends on the length of the chain at the N-2 position on the pyridazine ring. Furthermore, enzyme inhibition in vitro is increased with the presence of a halogen atom on the aromatic moiety of the benzyl group at C-5. Compound 4f having a pentanoic side chain and a 4-fluoro benzyl moiety was the most active derivative with an IC50 value of 6.69 x 10(-6) M. Molecular modelling studies were done on all the synthesized pyridazinones and on prostaglandin H2 (PGH2) suggesting spatial features and volumes of TXA2 synthase pharmacophore mode in these series of derivatives.

Thromboxane A2 biosynthesis inhibitors: synthesis and evaluation of pyrazolotriazinyl alkanoic acids.

A novel series of (6-aryl-4-oxo-pyrazolo 2,3-d] [1,2,5] triazin-3-yl) alkanoic acids was synthesized and evaluated in vitro as thromboxane A2 (TXA2) biosynthesis inhibitors. The experiments were carried out using arachidonic acid (32.8 microM) as a substrate and horse platelet microsomes (HPM) as sources of TXA synthetase. TXB2, a stable breakdown product of TXA2, was determined by radioimmunoassays (RIA). The substances under study, at concentrations ranging from 1.10(-6) M to 1.10(-4) M, significantly inhibited the biosynthesis of TXA2 in vitro. This activity was found to be dose-dependent, the potency of which could be related to structural features of the molecules. Compound 3b, bearing a butanoic side chain in the 3-position and a 4-chloro phenyl ring in the 6-position of the bicyclic system, was the most active derivative in in vitro enzyme inhibition (ID50 = 2.81 x 10(-5) M). Comparison of the spatial configurations of prostaglandin H2 (PGH2 and 3b displayed a good correlation between essential structural moieties of both molecules. In addition, conceptual model for the PGH2 and TX synthetase interactions was applied to compound 3b.

Toxicity and effects on the central nervous system of a Cerbera odollam leaf extract.

The immediate and delayed toxicity of Cerbera odollam leaf extract was studied in mice. Under the experimental conditions adopted, using macroscopic and microscopic examinations, Cerbera odollam leaves appeared to be relatively devoid of the marked toxicity found in seeds, a common source of poisoning. At doses smaller than the maximal dose never lethal (14.5 g/kg i.p.), the leaf extract decreased mice spontaneous motor activity significantly, increased the reaction time to a thermal stimulus, reduced the duration of pentylenetetrazole-induced tonic seizures and mortality, and potentiated sodium pentobarbital-generated hypnotic effects.

Toxicological studies of deltamethrin.

Deltamethrin [( S]-alpha-cyano-3-phenoxybenzyl-cis-(1R,3R)-3-(2,2-dibromovinyl+ ++) (2,2-dimethyl-cyclopropane-carboxylate], is the most potent insecticide known at the present time. But it is also one of the most toxic pyrethroids for vertebrates. The toxicity study of deltamethrin was performed on mice and rats, and on anaesthetized dogs, the administration route being either oral or intravenous. The oral LD50 of deltamethrin suspended in 10% gum-arabic solution was 5.54 +/- 1.29 g/kg p.o. in male mice and 3.45 +/- 1.27 g/kg p.o. in female mice. In rats and anaesthetized dogs, deltamethrin at high concentrations by the oral route engendered neither mortality nor signs of intoxication. When dissolved in glycerol formal and given intravenously, the LD50 of deltamethrin was as low as 3.44 +/- 0.67 mg/kg in anaesthetized dogs. Values obtained for the toxicity of deltamethrin varied not only with the animal species and sex involved, but also with the administration routes and solvents used. Administered orally, it was 100 times less toxic when suspended in gum-arabic solution than dissolved in oil or organic solvent. Whatever the animal species, sex, administration routes and solvents employed, the poisoning symptoms of deltamethrin are identical, i.e. salivation, ataxia and choreoathetotic movements with rolling convulsions, appearing within 7 h after administration. No cellular alteration was detectable by means of optical microscopy of excised organs.

Pharmacological effects of deltamethrin on the central nervous system.

Pharmacological exploration of the central nervous system carried out by means of a series of tests confirms the affinity of 3-(2,2-dibromovinyl)-2,2-dimethylcyclopropanecarboxylic acid alpha-cyano-3-phenoxybenzyl ester (delta-methrin) for the nervous system and reveals the following activities of deltamethrin: Deltamethrin has no neuroleptic or sedative action. It inhibits motility, sense of balance and inquisitiveness of treated animals. It potentiates chloral-induced hypnotic action but not that of pentobarbital. Not only does it have no antagonistic action on convulsivant agents: electric shock, pentetrazol, strychnine, but it stimulates their toxicity and prolongs the convulsive seizures. The fact that deltamethrin acts on different convulsivant agents (i.e. pentetrazol and strychnine) with various sites of action suggests that its activity is located in both the cortical and the medullary centres. Deltamethrin reveals the latent toxicity of tryptamine used in the non-toxic dose range; this predicts that deltamethrin might have an IMAO (inhibitor of monoamine oxidase) activity. Experiments carried out under the conditions adopted showed that deltamethrin acts on various sites.

Creation of a strain of genetically obese-hypertensive rats.

A strain of genetically obese-hypertensive rats (SHR-fa/fa) was created by transferring the fatty/fa gene of hyperlipaemic obese non-inbred rats into the genome of an SHR inbred strain by five successive crossings of SHR-fa+ brother-sister matings. SHR-fa/fa rats were heavier than their SHR littermates. They showed a severe hypertension, their systolic arterial blood pressure being higher than that previously found in genetically hypertensive rats. Their blood glucose content was not significantly different from that of their SHR littermates but their plasma triglycerides were increased by more than 500 per cent. While obesity and hypertension occurred from the 5th week following the rat's birth, the increase in blood triglycerides was manifest earlier.

Effect of a deproteinized blood extract on respiratory and haemodynamic modifications induced by hypoxia in the anaesthetized animal.

The effects of a deproteinized blood extract on the modifications of respiration, general metabolism and systemic haemodynamics induced by hypoxia were studied. Under the experimental conditions adopted, the deproteinized blood extract moderated the reactions of the organism which are exacerbated by hypoxia, that is increase in oxygen consumption and stimulation of the cardiac performances. The mechanism of action of the deproteinized blood extract has been discussed.

Taurine and icosanoids in the heart.

Experiments were carried out on non-working isolated rabbit hearts perfused by Tyrode solution: the effects of Taurine introduced into the coronary circulation were studied on the biosynthesis of the anti-thromboxane synthetase factor ("FATS") and on the TXA2 and PGI2 synthetase activities of cardiac tissue. The effects of Taurine were simultaneously studied on the biosynthesis of TXA2 and PGI2 in vitro. Experiments performed under the adopted conditions have shown that in vitro Taurine did not significantly modify the biosynthesis of TXA2 and PGI2; ex vivo Taurine did not change the biosynthesis of "FATS" but inhibited both TXA2 and PGI2 synthetase activities of the cardiac tissue: Taurine was more active on the TXA2 synthetase activity than on the PGI2 one. Thus Taurine promoted the formation of vasodilator and antiaggregating PGI2 at the expenses of vasoconstrictor and proaggregating TXA2. This could at least partly explain the beneficial effects of Taurine in the physiopathology of the heart.

Comparative study of the biosynthesis of PGE2, PGF2 alpha and TXA2 by different organs of genetically hypertensive (SHR) and obese-hypertensive (SHR-fa/fa) rats.

As an experimental model, we used 6-week-old genetically obese-hypertensive rats (SHR-fa/fa) which were obtained by transferring the fatty/fa gene of hyperlipaemic obese rats into the genome of the SHR strain: the SHR-fa/fa were bigger and more hypertensive than their SHR littermates. Studying the capacity of the hearts, kidneys, spleens, brains and lungs to synthesize PGE2, PGF2 alpha and TXA2, enabled us to show that the hearts and lungs of SHR-fa/fa synthesized more PG than those of SHR; SHR-fa/fa brains generated less icosanoids than those of SHR; the amounts of PGE2 and TXA2 produced by the kidneys are similar in SHR and in SHR-fa/fa. From the experimental data we can infer that the introduction of the fatty/fa gene into the genome of SHR does not significantly alter the capacity of the kidneys to synthesize icosanoids; the more severe hypertension in the SHR-fa/fa would result from an increase in TXA2 biosynthesis by cardiac tissue which, at the same time, synthesized more PGE2, which could be a means of defence against hypertension. Moreover this genetical manipulation inhibited the icosanoid-synthesizing capacity of the brain which thus attenuated the central nervous system activity of the animals.

Study of the PG E2 synthetase activity of different regions of dog and rabbit hearts.

Prostaglandin E2 synthetase activity of the microsomal fraction from different parts of dog and rabbit heart was tested with 3H-arachidonic acid as substrate. PG E2 synthesized was separated and purified by TLC and determined by the radiometric method or by bioassay. In the experimental conditions adopted, it was shown that the heart tissue is endowed with an enzyme system capable of synthesizing PG E2 but this PG E2 synthetase activity is not uniformly distributed in the different parts of the heart. It is highest in the right atrium and the activity of the atria is higher than that of the ventricles. It is species-dependent. The closely similar repartition of PG E2 synthetase activity and sympathetic nerve endings strongly suggests that PG E2 modulates adrenergic neurotransmission in the heart.