In vitro and in silico analysis of the vascular effects of asymmetrical N,N-bis(alkanol)amine aryl esters, novel multidrug resistance-reverting agents.

Asymmetrical N,N-bis(alkanol)amine aryl esters (FRA77, GDE6, and GDE19) are potent multidrug resistance (MDR) reversers. Their structures loosely remind that of the Ca(2+) antagonist verapamil. Therefore, the aim of this study was to investigate their vascular activity in vitro. Their effects on the mechanical activity of fresh and cultured rat aorta rings on Cav1.2 channel current (I Ca1.2) of A7r5 cells and their cytotoxicity on A7r5 and EA.hy926 cells were analyzed. Docking at the rat α1C subunit of the Cav1.2 channel was simulated in silico. Compounds tested were cytotoxic at concentrations >1 µM (FRA77, GDE6, GDE19) and >10 µM (verapamil) in EA.hy926 cells, or >10 µM (FRA77, GDE6, GDE19) and at 100 µM (verapamil) in A7r5 cells. In fresh rings, the three compounds partly antagonized phenylephrine and 60 mM K(+) (K60)-induced contraction at concentrations ≥1 and ≥3 µM, respectively. On the contrary, verapamil fully relaxed rings pre-contracted with both agents. In cultured rings, 10 µM GDE6, GDE19, FRA77, and verapamil significantly reduced the contractile response to both phenylephrine and K60. Similarly to verapamil, the three compounds docked at the α1C subunit, interacting with the same amino acids residues. FRA77, GDE6, and GDE19 inhibited I Ca1.2 with IC50 values 1 order of magnitude higher than that of verapamil. FRA77-, GDE6-, and GDE19-induced vascular effects occurred at concentrations that are at least 1 order of magnitude higher than those effectively reverting MDR. Though an unambiguous divergence between MDR reverting and vascular activity is of overwhelming importance, these findings consistently contribute to the design and synthesis of novel and potent chemosensitizers.

Nitric oxide modulation of interleukin-1[beta]-evoked intracellular Ca2+ release in human astrocytoma U-373 MG cells and brain striatal slices.

Intracellular Ca(2+) mobilization and release into mammal CSF plays a fundamental role in the etiogenesis of fever induced by the proinflammatory cytokine interleukin-1beta (IL-1beta) and other pyrogens. The source and mechanism of IL-1beta-induced intracellular Ca(2+) mobilization was investigated using two experimental models. IL-1beta (10 ng/ml) treatment of rat striatal slices preloaded with (45)Ca(2+) elicited a delayed (30 min) and sustained increase (125-150%) in spontaneous (45)Ca(2+) release that was potentiated by l-arginine (300 microm) and counteracted by N-omega-nitro-l-arginine methyl ester (l-NAME) (1 and 3 mm). The nitric oxide (NO) donors diethylamine/NO complex (sodium salt) (0.3 and 1 mm) and spermine/NO (0.1 and 0.3 mm) mimicked the effect of IL-1beta on Ca(2+) release. IL-1beta stimulated tissue cGMP concentration, and dibutyryl cGMP enhanced Ca(2+) release. The guanyl cyclase inhibitors 1H-[1,2, 4]oxadiazole[4,3-a] quinoxalin-1-one (100 microm) and 6-[phenylamino]-5,8 quinolinedione (50 microm) counteracted Ca(2+) release induced by 2.5 but not 10 ng/ml IL-1beta. Ruthenium red (50 microm) and, to a lesser extent, heparin (3 mg/ml) antagonized IL-1beta-induced Ca(2+) release, and both compounds administered together completely abolished this response. Similar results were obtained in human astrocytoma cells in which IL-1beta elicited a delayed (30 min) increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) (402 +/- 71.2% of baseline), which was abolished by 1 mm l-NAME. These data indicate that the NO/cGMP-signaling pathway is part of the intracellular mechanism transducing IL-1beta-evoked Ca(2+) mobilization in glial and striatal cells and that the ryanodine and the inositol-(1,4,5)-trisphosphate-sensitive Ca(2+) stores are involved.

Interleukin-1 beta stimulation of 45Ca2+ release from rat striatal slices.

1. Previous observations that centrally injected interleukin-1 beta (IL-1 beta) into rabbits induces a sustained rise in cerebrospinal fluid (CSF) Ca2+ concentration ([Ca2+]) as well as fever, prompted us to undertake an in vitro study to corroborate the in vivo results and gain insight as to the source and mechanism of IL-1 beta-induced Ca2+ mobilization. 2.IL-1 beta treatment of rat striatal slices preloaded with 45Ca2+ elicited a rise in spontaneous 45Ca2+ release which was dose-dependent, delayed in onset and of extended duration. At concentrations of 1, 5 and 10 ng ml-1, the 45Ca2+ efflux increased by 6.3 +/- 1.3 (s.e. mean), 33.4 +/- 5.0 and 159 +/- 10.5% respectively. 3. At 1 microgram ml-1, the specific IL-1 receptor antagonist, IRAP, antagonised the effect induced by, 10 ng ml-1 IL-1. 4. Caffeine 10 mM,which failed to release calcium on its won, potentiated IL-1-elicited 45Ca2+ release. 5. Perfusion with a Ca(2+)-free medium obtained by use of excess EGTA (3 mM) or in the presence of the Ca2+ channel blocker, nifedipine (3 x 10(-8 M) abolished the potentiating effect of caffeine without affecting the IL-1-induced 45Ca2+ release. 6. Preincubation of slices for 4 h with Bordetella pertussis toxin (PTX, 1.3 micrograms ml-1) did not change the pattern of Ca2+ efflux in response to IL-1. 7. In conclusion, these data indicate that IL-1 stimulates calcium release from brain tissue by a specific, receptor-mediated mechanism which partly depends on extracellular calcium but does not involve a PTX-sensitive G protein as part of the transducing signal.

Increase of extracellular brain calcium involved in interleukin-1 beta-induced pyresis in the rabbit: antagonism by dexamethasone.

1. This study investigates the role of extracellular brain calcium in the hyperthermia induced by interleukin-1 beta (IL-1 beta). 2. Intracerebroventricular (i.c.v.) injection of IL-1 beta (12.5 ng kg-1) in rabbits caused a prompt and sustained rise in cerebrospinal fluid (CSF) Ca2+ concentration ([Ca2+]) followed by enhanced prostaglandin E2 (PGE2) release and hyperthermia. 3. A linear and significant correlation was observed between the increase in [Ca2+] induced by IL-1 beta and the rise in body temperature. 4. Ventriculo-cisternal perfusion with artificial CSF containing the calcium chelator EGTA (1.3 mM) blocked the IL-1-induced PGE2 release and countered the febrile response. 5. I.c.v. administration of dexamethasone (Dex) (2.4 and 24 micrograms kg-1) 100 min prior to IL-1 beta, dose-dependently antagonized the cytokine-induced Ca2+ increase, the PGE2 release and the febrile response. 6. These results suggest that changes in extracellular brain calcium are involved in the regulation of body temperature. In this light, the antipyretic action of Dex may be related to its effect on Ca2+ uptake.

Potentiation of mitochondrial Ca2+ sequestration by taurine.

The effects of taurine (2-aminoethanesulphonic acid) and its analogues, 2-aminoethylarsonic acid, 2-hydroxyethanesulphonic (isethionic) acid, 3-aminopropanesulphonic acid, 2-aminoethylphosphonic acid, and N,N-dimethyltaurine, were studied on the transport of Ca2+ by mitochondria isolated from rat liver. Taurine enhanced Ca2+ uptake in an apparently saturable process, with a Km value of about 2.63 mM. Taurine behaved as an uncompetitive activator of Ca2+ uptake, increasing both the apparent Km and Vmax values of the process. This effect was not modified in the presence of cyclosporin A (CsA). N,N-Dimethyltaurine also stimulated Ca2+ uptake at higher concentrations, but there was no evidence that the process was saturable over the concentration range used (1-10 mM). Aminoethylarsonate was a weak inhibitor of basal Ca2+ uptake, but inhibited that stimulated by taurine in an apparently competitive fashion (Ki = 0.05 mM). The other analogues had no significant effects on this process. Taurine either in the presence or the absence of CsA had no effect on Ca2+ release induced by 200 nM ruthenium red. Thus, the mechanism of taurine-enhanced Ca2+ accumulation appears to involve stimulation of Ca2+ uptake via the uniport system rather than inhibition of Ca2+ release via the ion (Na+/Ca2+ and/or H+/Ca2+) exchangers or by taurine modulating the permeability transition of the mitochondrial inner membrane. Overall, these findings indicate an interaction of taurine with an as yet unidentified mitochondrial site which might regulate the activity of the uniporter. The unique role of taurine in modulating mitochondrial Ca2+ homeostasis might be of particular importance under pathological conditions that are characterised by cell Ca2+ overload, such as ischaemia and oxidative stress.

2,5-Di-t-butyl-1,4-benzohydroquinone induces endothelium-dependent relaxation of rat thoracic aorta.

The aim of this work was to clarify the mechanism by which 2,5-di-t-butyl-1,4-benzohydroquinone (BHQ) induces relaxation of rat thoracic aorta. In particular, the role of endothelium-derived nitric oxide (NO) was investigated. BHQ concentration dependently (0.1-10 microM) relaxed rat aorta rings precontracted with phenylephrine. This effect was dependent on the intactness of the endothelium, suppressed by preincubation with 100 microM N(omega)-nitro-L-arginine methyl ester and antagonised by 3-30 microM methylene blue. The 10 microM BHQ-induced relaxation, however, was followed by the gradual and slow return to phenylephrine-induced tone. Superoxide dismutase (250 U/ml) increased the BHQ-induced relaxation, while preincubation with 3 mM diethyldithiocarbamate inhibited it in a time-dependent fashion. BHQ gave rise to superoxide anion formation which was markedly inhibited by the addition of superoxide dismutase (250 U/ml), either in the presence or in the absence of aorta rings. The non-specific blocker of Ca2+ channels, Ni2+, concentration dependently attenuated the BHQ relaxing effect. BHQ did not modify the relaxation induced by the NO donor 3-morpholino-sydnonimine in endothelium-deprived rings. In conclusion, BHQ induces endothelium-dependent relaxation and gives rise, by auto-oxidation, to the formation of superoxide anion. The former effect results from the enhanced synthesis of NO rather than from its enhanced biological activity; NO synthase is presumed to be stimulated by BHQ-induced activation of Ca2+ influx through Ni2+-sensitive Ca2+ channels.

On the mechanisms of the antispasmodic action of some hindered phenols in rat aorta rings.

The antispasmodic effects of 3-t-butyl-4-hydroxyanisole (BHA) and some structurally related compounds were investigated in endothelium-intact rat aorta rings. Nordihydroguaieretic acid (NDGA), BHA, 3,5-di-t-butyl-4-hydroxyanisole (DTBHA), 2,6-di-isopropyl phenol (propofol) and 2,2'-dihydroxy-3,3'-di-t-butyl-5, 5'-dimethoxydiphenyl (DIBHA) did not cause relaxation when added at the plateau of phenylephrine-evoked contraction, nor did they affect the concentration-relaxation curve for acetylcholine in precontracted rings. In rings depolarised with physiological salt solution (PSS) containing 40 mM K(+), NDGA, BHA, DTBHA, 2, 5-di-t-butyl-1,4-benzohydroquinone (BHQ), propofol and nifedipine, but not DIBHA, inhibited the contraction induced by cumulative addition of Ca(2+) (0.05-10 mM) in a concentration-dependent manner; this inhibition was inversely related to the Ca(2+) concentration. In 40 mM K(+) PSS, 25 nM nifedipine blocked the 1 mM Ca(2+)-induced contraction, whereas 50 microM DTBHA, NDGA, BHA, BHQ and propofol significantly antagonised it by 84.4%, 73.0%, 52.8%, 45.6% and 35.7%, respectively. In the presence of 1 microM methyl-1,4-dihydro-2, 6-dimethyl-3-nitro-4-(2-trifluoromethylphenyl)-pyridine-5-carboxylate (Bay K 8644), the response to Ca(2+) did not differ from control values with nifedipine and BHQ, was partially restored with DTBHA and NDGA, and was not affected with BHA and propofol. Nifedipine markedly inhibited (85.2%) the Ba(2+)-induced contraction and this effect was totally reversed by Bay K 8644. BHA and DTBHA showed antispasmodic activity (45.3% and 43.1%, respectively) which was partly reversed by Bay K 8644. In contrast, Bay K 8644 did not affect the inhibition exerted by BHQ, NDGA and propofol (69.5%, 53. 3% and 46.1%, respectively). Nifedipine, BHA, DTBHA, propofol and NDGA inhibited the contractile response to 1 mM Ca(2+) of aorta rings depolarised with 40 or 80 mM K(+) PSS to a similar extent. Cromakalim inhibited the Ca(2+)-evoked contraction only in 30 mM K(+) PSS and BHQ only in 80 mM K(+) PSS. DIBHA had no effect on this model. Cromakalim, but not BHA, stimulated 86Rb(+) efflux from ring preparations. In 80 mM K(+) PSS containing 1 microM nifedipine, only papaverine affected the phenylephrine-induced contraction. Moreover, when the rings were preincubated with 1 mM Ni(2+), the response to phenylephrine in the presence of BHQ was significantly reduced. In conclusion, we propose that BHA may non-specifically inhibit Ca(2+) influx at the plasmalemma level rather than affect the function of K(+) channels, Ca(2+) release from intracellular stores or endothelium-dependent relaxation.

Inhibition of interleukin-1 beta-induced pyresis in the rabbit by peptide 204-212 of lipocortin 5.

The intracerebroventricular administration of interleukin-1 beta (12.5 ng/kg) in rabbits caused a prompt rise of prostaglandin E2 concentration (+ 632.6 +/- 243.9%) in the cerebrospinal fluid followed by hyperthermia (+ 1.61 +/- 0.14 delta degrees C). The intracerebroventricular administration of an anti-inflammatory nonapeptide (amino acids 204-212, SHLRKVFDK) derived from lipocortin 5, thereafter referred to as lipocortin 5-(204-212)-peptide, inhibited in a significant manner both the increase in cerebrospinal fluid [prostaglandin E2] and the febrile response induced by the cytokine. This inhibitory effect is probably due to interference by the peptide with phospholipase A2 activity. A control peptide (FKRVHDLKS) formed by the same amino acids in a randomly shuffled sequence had no effect. These results show that, in addition to the anti-inflammatory effect previously reported, the peptide 204-212 of lipocortin 5 possesses, like glucocorticoids, anti-pyretic activity. The research on lipocortin-derived peptides may lead to the development of novel anti-inflammatory and anti-pyretic compounds.

Cytochrome P450-dependent N-dealkylation of L-deprenyl in C57BL mouse liver microsomes: effects of in vivo pretreatment with ethanol, phenobarbital, beta-naphthoflavone and L-deprenyl.

The monoamine oxidase inhibitor L-deprenyl [(-)-deprenyl, selegiline] is an effective therapeutic agent for improving early symptoms of idiopathic Parkinson's disease. It appears to exert this action independently of its inhibition of monoamine oxidase B (MAO-B) and some of its metabolites are thought to contribute. Cytochrome P450 (CYP) activities are known to give rise to L-deprenyl metabolites that may affect the dopaminergic system. In order to clarify the interactions of L-deprenyl with these enzymes, C57BL mice were treated with L-deprenyl, ethanol, phenobarbital or beta-naphthoflavone to induce different CYP isozymes. After preincubation of L-deprenyl with liver microsomes from control or treated mice, the metabolites were analysed by a GLC method. L-deprenyl (10 mg/kg i.p. for 3 days) caused a significant decrease in total CYP levels (0.315+/-0.019, L-deprenyl; 0.786+/-0.124, control, nmol/mg protein) and CYP2E1-associated p-nitrophenol hydroxylase activity (0.92+/-0.04 vs. 1.17+/-0.06 nmol/min/mg). Both phenobarbital and ethanol increased the N-depropynylation activity towards L-deprenyl that leads to the formation of methamphetamine (4. 11+/-0.64, phenobarbital; 4.77+/-1.15, ethanol; 1.77+/-0.34, control, nmol/min/mg). Ethanol alone increased the N-demethylation rate of L-deprenyl, that results in formation of nordeprenyl (3.99+/-0.68, ethanol; 1.41+/-0.31, control, nmol/min/mg). Moreover, the N-dealkylation pathways of deprenyl are inhibited by 4-methylpyrazole and disulfiram, two CYP2E1 inhibitors. None of the other treatments modified L-deprenyl metabolism. These findings indicate that mainly CYP2E1 and to a lesser extent CYP2B isozymes are involved in L-deprenyl metabolism. They also suggest that, by reducing CYP content, L-deprenyl treatment may impair the metabolic disposition of other drugs given in combination regimens.

HPLC determination of inorganic cation levels in CSF and plasma of conscious rabbits.

An HPLC method is described using conductimetric detection for the quantitative determination of sodium, potassium, magnesium, and calcium in cerebrospinal fluid (CSF) and plasma of conscious rabbits. This method enabled the four cations to be estimated with rapidity, sensitivity, accuracy, and precision. The mean millimolar concentrations +/- SD found in CSF and (plasma) of 15 untreated animals were as follows: sodium, 146.96 +/- 17.84 (135.06 +/- 20.11); potassium, 3.32 +/- 0.56 (4.57 +/- 1.03); magnesium, 0.90 +/- 0.20 (0.72 +/- 0.13); and calcium, 1.47 +/- 0.19 (3.32 +/- 0.59).

Calcium changes in rabbit CSF during endotoxin, IL-1 beta, and PGE2 fever.

New Zealand rabbits were chronically incannulated in the lateral ventricle and cisterna magna to assess the hypothesis that calcium concentration (Ca) of cerebrospinal fluid (CSF) varies during fevers of diverse origin. In normothermic and febrile animals recovering from surgery, CSF Ca was positively and significantly correlated to rectal temperature (TR). IV injection of E. coli endotoxin and ICV injection of human recombinant interleukin 1 beta (hrIL-1 beta) induced a TR rise of 1.7 +/- 0.3 degrees C (mean +/- SEM) and 1.45 +/- 0.25 degrees C, respectively, accompanied by significant increases in CSF Ca. After endotoxin administration, maximal Ca increases ranged between 0.21 and 0.48 mM above basal values in individual animals (p < 0.01), whereas after administration of hrIL-1 beta increases were 0.17 and 0.25 mM (p < 0.05). Acetylsalicylic acid (ASA) countered the fever induced by both endotoxin and hrIL-1 beta administrations and concomitantly antagonized the Ca increase in CSF. HrIL-1 beta-derived nonapeptide was characteristically devoid of pyrogenic effect and did not modify CSF Ca. Although ICV injection of prostaglandin E2 (PGE2) increased TR by 2.1 +/- 0.77 degrees C, it failed to have any effect on CSF Ca. Differently from the other Ca enhancers, PGE2, however, increased CSF protein concentration (protein). These findings suggest that brain calcium metabolism plays a role in fever development and that prostaglandin involvement is only engaged once changes in CSF calcium concentration have taken place.

N-Dealkylation of chlorimipramine and chlorpromazine by rat liver microsomal cytochrome P450 isoenzymes.

The role of different cytochrome P450 isozymes (CYP) in the N-demethylation of chlorimipramine and chlorpromazine has been investigated in liver microsomes from rats by studying the effects of multiple subchronic doses of chlorimipramine, chlorpromazine, phenobarbital and beta-naphthoflavone on the N-demethylation of ethylmorphine, mono-N-demethyl-chlorimipramine and chlorpromazine and on the hydroxylation of aniline. With control microsomes, CYP-dependent metabolism of chlorimipramine and chlorpromazine (100 nmol; 30 min incubation) resulted in the formation of predominantly chlorimipramine (46.5 +/- 4.9 nmol) whereas chlorpromazine (14.1 +/- 0.9 nmol) accounted for only part of the overall metabolism of chlorpromazine. Multiple doses of chlorimipramine increased the capacity of microsomes to N-demethylate ethylmorphine (9.8 +/- 0.73 and 6.08 +/- 0.06 nmol min(-1) (mg protein)(-1) for chlorimipramine-treated and control rats, respectively) as well as itself (4.65 +/- 0.25 and 3.10 +/- 0.33 nmol min(-1) (mg protein)(-1), respectively). Multiple doses of chlorpromazine induced aniline-hydroxylase activity (1.11 +/- 0.16 and 0.94 +/- 0.06 nmol min(-1) (mg protein)(-1) for chlorimipramine and control microsomes, respectively) but the capacity to N-demethylate itself was unchanged. Phenobarbital treatment induced ethylmorphine N-demethylation activity, but did not affect N-demethylation activity, towards chlorimipramine and chlorpromazine. In control microsomes the N-demethylation capacity of chlorimipramine or chlorpromazine (0.160 +/- 0.025 and 0.015 +/- 0.003 nmol min(-1) (mg protein)(-1), respectively) was one order of magnitude lower than that of chlorimipramine or chlorpromazine. The capacity to N-demethylate either chlorimipramine or chlorpromazine was increased by treatment with either phenobarbital or beta-naphthoflavone. In control microsomes, sulphaphenazole markedly inhibited both chlorimipramine-N-mono- and di-N-demethylation, whereas quinidine markedly inhibited the rate of formation of chlorpromazine. The CYP2C and CYP2D subfamilies seem to be involved in the mono N-demethylation of chlorimipramine and chlorpromazine, respectively. Moreover the CYP1A and CYP2B subfamilies might participate in the N-demethylation of either chlorimipramine or chlorpromazine. This could have important implications in the clinical use of chlorimipramine and chlorpromazine in view of the genetic polymorphism of CYP2C and CYP2D isozymes in man.

Rat tissue concentrations of chlorimipramine, chlorpromazine and their N-demethylated metabolites after a single oral dose of the parent compounds.

A single oral dose of 90 mg kg-1 chlorimipramine or chlorpromazine, corresponding to 54.5 or 55.9 mumol, respectively, was given to male Sprague-Dawley rats and concentrations of parent drugs and their N-desmethyl metabolites measured by gas chromatography in plasma and major organs (brain, liver, lung, kidney, heart, spleen and peritoneal fat). In the case of chlorimipramine, N-desmethyl metabolite levels were consistently higher than those of the parent drug for the entire observation period of 24 h in all tissues except fat, while lower N-desmethyl metabolite/parent compound ratios were observed for chlorpromazine. N-Desmethyl metabolite kinetics of chlorimipramine appeared to be elimination-rate limited, while those of chlorpromazine were formation-rate limited. In all analysed organs, the maximu detectable drug+metabolite concentrations accounted for only 2.3 and 4.6% of the initial dose of chlorimipramine and chlorpromazine. Chlorpromazine treatment gave rise to an area under the total amount-time curve (AUC0-24) for parent drug+metabolites, 3.9-fold that for chlorimipraine. Closer scrutiny discloses a conversion ratio of parent compound to N-desmethyl metabolite of 1.1 for chlorpromazine and of 2.2 for chlorimipramine, indicating the greater efficiency of chlorimipramine metabolism in all compartments. The expected high conversion index found in the liver (2.3) reaches its maximum of 5.4 in the lung. Fractional data analysis of chlorimipramine and chlorpromazine distribution patterns revealed greater organ transfer for the N-desmethyl metabolites than for the more stably-located parent compounds. The N-desmethyl metabolites of chlorimipramine apparently moved from liver to lung, kidney and spleen, whereas N-desmethylchlopromazine moved preferentially to the brain and lung tissue. This single dose study of chlorimipramine and chlorpromazine kinetics, highlights the two distinct dispositional processes at work in the rat in all likelihood, attributable to different absorption patterns, to a slower metabolism and, thus, to the longer persistence of chlorpromazine.

Effects of taurine and some structurally related analogues on the central mechanism of thermoregulation: a structure-activity relationship study.

There is large body of evidences on the role of taurine in the central mechanisms of thermoregulation in mammals, but it is not clear, whether the hypothermic effect of taurine depends on its interaction with GABA receptors or with a specific receptor. In order to answer this question, we have performed a structure-activity relationship study by using both in vitro and in vivo preparations. MicroM amounts of taurine or each of 20 analogues were injected intracerebroventricularly in conscious, restrained rabbits while rectal temperature was recorded. Receptor-binding studies, with synaptic membrane preparations from rabbit brain were used to determine the affinities of these compounds for GABA(A) and GABA(B) receptors. Furthermore, the interaction with presynaptic GABA and taurine uptake systems was studied using crude synaptosomal preparations from rabbit brain. Among the compounds tested, (+/-)-cis-2-aminocyclohexanesulfonic acid, induced hypothermia, but did not interact with GABA(A) and GABA(B) receptors neither did it affect GABA and taurine uptake, thus suggesting that its effect on body temperature is not mediated by the central GABAergic system. Interestingly, the trans-isomer was devoid of effects either in vivo or in vitro. In order to explain (+/-)-cis-2-aminocyclohexanesulfonic acid-induced hypothermia, a stereoscopic model was produced showing its possible interactions with a putative taurine brain receptor.

Synthesis of taurine analogues from alkenes: preparation of 2-aminocycloalkanesulfonic acids.

(+/-)trans 2-Aminocyclohexanesulfonic acid and (+/-)trans 2-aminocyclopentanesulfonic acid were prepared from cyclohexene and cyclopentene respectively by sulfur monochloride addition, followed by oxidation to 2-chlorosulfonic acid and substitution of chlorine.

The possible role of taurine and GABA as endogenous cryogens in the rabbit: changes in CSF levels in heat-stress.

We investigated whether heat-stress induced hyperthermia could enhance release of both endogenous taurine and GABA from nerve cells into the extracellular compartment, thus acting like endogenous cryogens. Conscious rabbits were exposed for 1 hr to 40 degrees C (heat stress) while cerebrospinal fluid (CSF) and plasma osmolality and the CSF concentrations of some cations, proteins as well as those of taurine and GABA were determined. Heat stress-induced hyperthermia was accompanied by a significant rise in CSF and plasma osmolality, CSF calcium, taurine and GABA levels. It is suggested that during heat stress taurine and GABA are released in the extracellular space of brain tissues in higher amounts, as compared to control conditions, to counteract the resulting hyperthermia, thus acting as cryogenic agents.

The mitochondrial permeability transition and taurine.

Perturbed cellular calcium homeostasis has been implicated in both apoptosis and necrosis, but the role of altered mitochondrial calcium handling in the cell death process is unclear. Recently we found that taurine, a naturally occurring amino acid potentiates Ca2+ sequestration by rat liver mitochondria. These data, which accounted for the taurine antagonism on Ca2+ release induced by the neurotoxins 1-methyl-4-phenylpyridinium plus 6-hydroxy dopamine previously reported, prompted us to investigate the effects of taurine on the permeability transition (PT) induced experimentally by high Ca2+ plus phosphate concentrations. The parameters used to measure the PT were, mitochondrial swelling, cytochrome c release and membrane potential changes. The results showed that, whereas taurine failed to reverse changes of these parameters, cyclosporin A completely reversed them. Even though these results exclude a role in PT regulation under such gross insult conditions, they cannot exclude an important role for taurine in controlling pore-opening under milder more physiological PT-inducing conditions.

Pharmacokinetics of diphenhydramine in healthy volunteers with a dimenhydrinate 25 mg chewing gum formulation.

This pharmacokinetic study evaluated diphenhydramine in the plasma of healthy volunteers after a single 25 mg oral dose of dimenhydrinate (diphenhydramine theophyllinate), corresponding to 12.7 mg diphenhydramine, in a chewing gum formulation. Seven volunteers (4 men, 3 women; age: 26.3 +/- 1.2 years; body weight: 63.1 +/- 4.1 kg; height: 172.4 +/- 4.6 cm) chewed the gum for 1 h. Blood samples (10 ml) were collected at different time intervals up to 24 h. Blood plasma was subsequently processed and analyzed for diphenhydramine content using a GLC method and an NPD detector. Analytical data revealed the following kinetic parameters: AUC(0-24h): 155.2h x ng x ml(-1); AUC(0-infinity): 195.3 h x ng x ml(-1); Mean resident time: 16 h; t(1/2): 10 h; C(max): 14.5 ng x ml(-1); t(max): 2.6 h; and plasma clearance: 9.0 ml x min(-1) x kg(-1). This study indicates that the pharmaceutical formulation employed provided sustained plasma concentrations of diphenhydramine, presumably sufficient to support its clinical efficacy towards motion sickness owing to the almost complete (> 95%) release by the formulation of the active principle. Moreover, the maximal concentrations of diphenhydramine attained in plasma were much lower than the concentration threshold needed to produce drowsiness.

Toxicity assessment on trophoblast cells for some environment polluting chemicals and 17ß-estradiol.

The identification of reproductive toxicants is a major scientific challenge for human health. We investigated the effects of a selected group of environmental polluting chemicals mostly provided with estrogenic activity on the human trophoblast cell lines BeWo and HTR-8/SVneo. Cells were exposed for 24h to various concentrations (from 0.1 pM to 1 mM) of atrazine (ATR), diethylstilbestrol (DES), para-nonylphenol (p-NP), resveratrol (RES) and 17 ß-estradiol (E2) and assayed for cell viability and human beta-Chorionic Gonadotropin (ß-hCG) secretion. Decrease of cell viability as respect to control, vehicle-treated, cultures was obtained for all chemicals in the concentration range of 1 µM-1 mM in both cell types. A parallel decrease of ß-hCG secretion was observed in BeWo cells, at 1 µM-1 mM concentrations, with the only exception of ATR which caused an increase at concentrations up to 1mM. ß-hCG release was also unexpectedly inhibited by ATR, DES, p-NP and RES at non-toxic (pM-nM) concentrations. These findings raise concern about the negative, potential effects of various environmental polluting chemicals on pregnancy success and fetal health.