Log EC50 Research Articles

Testosterone Alters Maternal Vascular Adaptations

Sex steroid hormones estradiol and progesterone play an important role in vascular adaptations during pregnancy. However, little is known about the role of androgens. Plasma testosterone (T) levels are elevated in preeclampsia, mothers with polycystic ovary, and pregnant African American women, who have endothelial dysfunction and develop gestational hypertension. We tested whether increased T alters vascular adaptations during pregnancy and whether these alterations depend on endothelium-derived factors, such as prostacyclin, endothelium-derived hyperpolarizing factor, and NO. Pregnant Sprague Dawley rats were injected with vehicle (n=12) or T propionate [0.5 mg/Kg per day from gestation day 15–19; n=12] to increase plasma T levels 2-fold, similar to that observed in preeclampsia. Telemetric blood pressures and endothelium-dependent vascular reactivity were assessed with wire-myograph system. Phospho-endothelial NO synthase and total endothelial NO synthase were examined in mesenteric arteries. Mean arterial pressures were significantly higher starting from gestation day19 until delivery in T-treated dams. Endothelium-dependent relaxation responses to acetylcholine were significantly lower in mesenteric arteries of T-treated dams (pD 2 [−log EC 50 ]=7.05±0.06; E max =89.4±1.89) compared with controls (pD 2 =7.38±0.04; E max =99.9±0.97). Further assessment of endothelial factors showed NO-mediated relaxations were blunted in T-treated mesenteric arteries (E max =42.26±5.95) compared with controls (E max =76.49±5.06); however, prostacyclin- and endothelium-derived hyperpolarizing factor-mediated relaxations were unaffected. Relaxation to sodium nitroprusside was unaffected with T-treatment. Phosphorylations of endothelial NO synthase at Ser 1177 were decreased and at Thr 495 increased in T-treated mesenteric arteries without changes in total endothelial NO synthase levels. In conclusion, increased maternal T, at concentrations relevant to abnormal clinical conditions, cause hypertension associated with blunting of NO-mediated vasodilation. T may induce the increased vascular resistance associated with pregnancy-induced hypertension.

Model Suite for Predicting the Aquatic Toxicity of α,β-Unsaturated Esters Triggered by Their Chemoavailability.

For a Michael-acceptor set of 45 α,β-unsaturated esters, the 2(nd) -order rate constant of reaction with glutathione, log kGSH , was modeled through the quantum chemical reaction barrier (ΔE(≠) ) employing methane thiol as model nucleophile. Regression of their 48-h toxicity toward the ciliates Tetrahymena pyriformis (log EC50 , 50 % growth inhibition) on log Kow (octanol/water partition coefficient) and log kGSH revealed a variation in the relative weights of hydrophobicity and electrophilic reactivity as determinants of the aquatic toxicity. The difference DKk =log Kow -log kGSH turned out as a suitable means for predictively discriminating between narcosis-level (DKk >3.0) and excess-toxic (DKk <2.0) compounds. In the intermediate DKk range (2.0≤DKk ≤3.0), both narcosis-level and reactive-toxicity models are applicable for predicting aquatic toxicity. As such, DKk represents the chemoavailability of Michael-acceptor esters, characterizing their likelihood for undertaking covalent reactions with thiol sites of endogenous peptides and proteins. At the same time, DKk introduces a straightforward way for characterizing the applicability domain of QSAR (quantitative structure-activity relationship) models for predicting the toxicity of Michael-acceptor esters. The resultant model suite comprising QSARs for reactive toxicity and baseline narcosis is triggered by the compounds' chemoavailability, and yields predictions superior to existing approaches.

Heparinized blood better preserves cellular energy charge and vascular functions of intraoperatively stored saphenous vein grafts in comparison to isotonic sodium-chloride-solution

Several studies have addressed the optimal storage conditions for vascular grafts during surgery. The results remain contradictionary. This may be attributed to the fact, that the various vascular beds have a different sensitivity to storage. We analyzed the impact of storage in isotonic saline solution (NaCl) or heparinized blood the vascular functions of human saphenous vein grafts. Special care was taken to choose storage conditions which are relevant for intraoperative storage of a saphenous vein graft in a setting of coronary artery bypass grafting with vein and internal mammary artery as grafts. Intraoperatively isolated V. saphena-segments (n = 36) were stored in NaCl or heparinized blood for approximately 30 minutes at room temperature. Subsequently, the segments were examined in a Mulvany-myograph. Following preconstriction with norepinephrine, concentration-relaxation curves were assessed for bradykinin and sodium-nitroprusside to assess developed vessel-wall tension as well as endothelium- and smooth-muscle-cell dependent vasorelaxation. The availability of adenosintriphosphate (energy charge) was determined based on liquid chromatography measurements of nucleotide tissue levels. Mean storage time was 27.4 ± 2.4 min in NaCl- and 26.3 ± 2.7 min in blood-group, respectively. After this period, receptor-dependent and-independent maximum of developed vessel wall tension was significantly reduced in NaCl-group (p = 0.05 and p = 0.045, respectively). Furthermore, the energy charge was significantly (p = 0.046) better preserved after blood storage (74 ± 1%) in comparison to NaCl-group (68 ± 2%). Endothelium-induced vasodilatation in response to bradykinin reached only 12.3 ± 2.5% in NaCl-group, but 19.3 ± 5.2% in blood-group (p = 0.033). Alike, EC50-concentration of bradykinin for half-maximal relaxation was significantly lower in blood- than in NaCl-group (log EC50 -7.08 ± 0.3 and -5.91 ± 0.4; respectively; p = 0.046). Endothelium-independent smooth muscle relaxation in response to sodium-nitroprusside was not different between both groups. Heparinized blood better preserves vascular contractile and endothelial functions of the saphenous vein graft. Storage in NaCl rapidly compromises vascular functions and impaires cellular energy. NaCl should no longer be recommended for intraoperative storage of harvested V. saphena grafts.

Differential Responses of Colletotrichum gloeosporioides and C. truncatum Isolates from Different Hosts to Multiple Fungicides Based on Two Assays.

Anthracnose is one of the most important postharvest diseases of many economically important crops worldwide. This study was conducted with the objective of investigating the sensitivity of Colletotrichum gloeosporioides and C. truncatum isolates to multiple fungicides with different modes of action. The study analyzed quantitative sensitivity data derived from conventional amended agar (AA) assays and qualitative spore responses obtained from a novel microtiter bioassay that is based on reduction of a viability dye, Alamar blue (AB). Generally, for AA assays, the percent growth inhibition (%RGI) increased with increasing concentration for all isolates and all fungicides, except for copper hydroxide. C. truncatum isolates reacted differently to increasing concentrations of the various fungicides depending on whether the isolates originated from pepper or papaya. C. truncatum from pepper had generally less %RGI than C. truncatum isolates from papaya. C. gloeosporioides isolates from papaya had generally higher %RGI than C. truncatum isolates for all concentrations tested for pyraclostrobin, chlorothalonil, and fosetyl-aluminum. C. gloeosporioides isolates from pepper had generally higher %RGI than C. truncatum isolates for all concentrations tested for most fungicides. In all cases, Colletotrichum sp. and fungicide had significant (P ≤ 0.001) effects on the log concentration of fungicide for which relative growth was inhibited by 50 and 90% (log EC50 and log EC90, respectively) calculated for all isolates, regardless of whether values were compared for only C. gloeosporioides isolates or only C. truncatum isolates. Correlation analyses of log EC50 and log EC90 values of all the isolates revealed a nonsignificant association for pyraclostrobin. In AB assays, all fungicides had an equivalent effect at inhibiting spore germination at the lower concentrations. According to binary logistic regression analyses, species, isolate, and fungicide concentration had significant predictive value in determining whether an AB test would be positive. Sequence alignments between C. gloeosporioides isolates and C. gloeosporioides f. sp. aeschynomene revealed no base substitutions at codons 198, 199, 200, and 240; however, sequence comparisons between C. truncatum isolates and C. gloeosporioides f. sp. aeschynomene revealed two codon changes located outside of the identified codon 198 or 200 associated with the benzimidazole-resistant phenotype of C. gloeosporioides isolates.

Abstract 337: Mechanisms of Reduced Aortic Relaxation Rate in Rats with Adenine-Induced Chronic Renal Failure

We have previously shown that adenine-induced chronic renal failure (A-CRF) in rats causes a pronounced reduction in the rate of relaxation of the thoracic aorta, but not of mesenteric arteries. The aim of this study was to elucidate the mechanisms causing a decreased rate of relaxation in the thoracic aorta of rats with A-CRF and to examine if this abnormality was specific for this vascular bed or comprised also other conductance arteries. Male Sprague-Dawley rats received either chow containing adenine (0.5% week 1-3, 0.3% w. 4-5, and 0.15% thereafter) or were pair-fed an identical diet without adenine (controls). After 8-13 weeks segments of thoracic aorta, abdominal aorta, common carotid artery and common femoral artery were analyzed ex vivo with wire myograph. Data are means±SD. Plasma creatinine levels were markedly elevated in A-CRF animals (306±139 vs. 31±3 μM, P&lt;0.001). The area under the curve for the initial 50 % reduction in force (reflecting relaxation rate) following wash-out of KCl was significantly elevated in A-CRF animals in the thoracic aorta (3.7-fold vs. controls, P&lt;0.01), abdominal aorta (5.6-fold vs. controls, P&lt;0.05), carotid artery (7.0-fold vs. controls, P&lt;0.05), but was not significantly affected in the femoral artery. Removal of endothelium did not affect relaxation rate significantly in A-CRF thoracic aortas. However, aortic relaxation rate increased significantly (P&lt;0.01), but was not normalized, in response to wash-out of KCl with Ca2+-free buffer. Thoracic aortas from A-CRF animals showed an increased sensitivity to KCl (log EC50 values -1.67±0.05 vs. -1.74±0.04 M in controls and A-CRF, respectively, P&lt;0.01) but an unaltered sensitivity to Ca2+ during concentration-response experiments. In conclusion, rats with A-CRF show a pronounced reduction in the rate of relaxation of larger conductance arteries localized proximal to the common femoral artery. This abnormality, which could contribute to increased arterial stiffness, is independent of the endothelium but partly dependent on extracellular Ca2+. The increased sensitivity to KCl in the presence of a normal sensitivity to Ca2+ in A-CRF aortas could indicate that vascular smooth muscle cells of A-CRF aortas are more depolarized but this needs to be investigated further.

Predicting Toxicity of Ionic Liquids in Acetylcholinesterase Enzyme by the Quantitative Structure–Activity Relationship Method Using Topological Indexes

A new topological index (TI) was proposed based on atom characters (e.g., atom radius, atom electronegativity, etc.) and atom positions in the hydrogen-suppressed molecule structure in our previous work. In this work, the TI was used for predicting the toxicity of ILs in acetylcholin esterase (log EC50 AChE) by the multiple linear regression (MLR) method. For ILs composed entirely of cations and anions, the TIs are calculated from cations and anions, respectively. The 221 ILs used in the MLR model are based on imidazolium (Im), pyridinium (Pyi), pyrrolidinium (Pyo), ammonium (Am), phosphonium (Ph), quinolinium (Qu), piperidinium (Pi), and morpholinium (Mo). The regression coefficient (R2) and the overall average absolute error (AAE) are 0.877 and 0.153, respectively.

QSAR, QSPR and QSRR in Terms of 3-D-MoRSE Descriptors for In Silico Screening of Clofibric Acid Analogues.

A series of 27 analogues of clofibric acid, mostly heteroarylalkanoic derivatives, have been analyzed by a novel high-throughput reversed-phase HPLC method employing combined gradient of eluent's pH and organic modifier content. The such determined hydrophobicity (lipophilicity) parameters, log kw , and acidity constants, pKa , were subjected to multiple regression analysis to get a QSRR (Quantitative StructureRetention Relationships) and a QSPR (Quantitative Structure-Property Relationships) equation, respectively, describing these pharmacokinetics-determining physicochemical parameters in terms of the calculation chemistry derived structural descriptors. The previously determined in vitro log EC50 values - transactivation activity towards PPARα (human Peroxisome Proliferator-Activated Receptor α) - have also been described in a QSAR (Quantitative StructureActivity Relationships) equation in terms of the 3-D-MoRSE descriptors (3D-Molecule Representation of Structures based on Electron diffraction descriptors). The QSAR model derived can serve for an a priori prediction of bioactivity in vitro of any designed analogue, whereas the QSRR and the QSPR models can be used to evaluate lipophilicity and acidity, respectively, of the compounds, and hence to rational guide selection of structures of proper pharmacokinetics.

Modelling defined mixtures of environmental oestrogens found in domestic animal and sewage treatment effluents using an in vitro oestrogen‐mediated transcriptional activation assay (T47D‐KBluc)

There is growing concern of exposure of fish, wildlife and humans to water sources contaminated with oestrogens and the potential impact on reproductive health. Environmental oestrogens can come from various sources including concentrated animal feedlot operations (CAFO), municipal waste, agricultural and industrial effluents. US EPA's drinking water contaminant candidate list 3 (CCL3) includes several oestrogenic compounds. Although these contaminants are currently not subject to any proposed or promulgated national primary drinking water regulations, they are known or anticipated to occur in public water systems and may require future regulation under the Safe Drinking Water Act. Using an in vitro transcriptional activation assay, this study evaluated oestrogens from CCL3 both individually and as a seven oestrogen mixture (fixed ray design) over a broad range of concentrations, including environmentally relevant concentrations. Log EC(50) and Hillslope values for individual oestrogens were as follows: estrone, -11.92, 1.283; estradiol-17α, -9.61, 1.486; estradiol-17β, 11.77, 1.494; estriol, -11.14, 1.074; ethinyl estradiol-17α, -12.63, 1.562; Mestranol, -11.08, 0.809 and Equilin, -11.48, 0.946. In addition, mixtures that mirrored the primary oestrogens found in swine, poultry and dairy CAFO effluent (fixed-ratio ray design), and a ternary mixture (4 × 4 × 4 factorial design) of oestrogens found in hormone replacement therapy and/or oral contraceptives were tested. Mixtures were evaluated for additivity using both the concentration addition (CA) model and oestrogen equivalence (EEQ) model. For each of the mixture studies, a broad range of concentrations were tested, both above and below environmentally relevant concentrations. Results show that the observed data did not vary consistently from either the CA or EEQ predictions for any mixture. Therefore, either the CA or EEQ model should be useful predictors for modelling oestrogen mixtures.

104 Enhancing endothelial insulin sensitivity reduces nitric oxide bioavailability: a role for NADPH oxidase-derived reactive oxygen species

IntroductionInsulin resistance at a whole body level and in the endothelium precedes type 2 diabetes. Insulin resistance at the level of the endothelium reduces bioavailability of the anti-atherosclerotic signalling radical...

Early life stress‐induced exaggerated AngII aortic vasoconstriction is transmitted to the F1 generation

Previously, we showed that maternal separation (MS), a model of early life stress, reduces creatinine clearance (Ccr) and enhances angiotensin II (AngII)‐induced aortic vasoconstriction via reduced nitric oxide (NO) buffering in male rats. We hypothesized that this phenotype would be transmitted to the first generation (F1) of male offspring from MS parents. Normally‐reared rats (12 weeks old) from either MS (MSF1) or control WKY parents were placed in metabolic cages. Food and water intake, body weight, and Ccr were all similar in control and MSF1 rats. Wire myography was performed to assess aortic vasoconstrictive responses to AngII in the presence and absence of the NO synthase inhibitor (L‐NAME, 100 μM, 15 min). Aorta from MSF1 compared to control rats showed greater sensitivity (log EC50: −8.51±0.14 vs. −7.93±0.12, p&lt;0.05) and exaggerated vasoconstriction to AngII (9.38±0.8 vs. 4.8±0.3 % KCl, p&lt;0.05). Exacerbated AngII‐induced constriction due the pre‐incubation with L‐NAME, was greater in aorta from control compared to MSF1 rats (delta % max constriction: 34.7±4.3 vs. 13.2±2.1, p&lt;0.05). KCl induced aortic vasoconstriction was similar in control and MSF1 rats. These data indicate that the MSF1 rats maintains the vascular phenotype with reduced aortic NO buffering to AngII‐induced vasoconstriction, thus, suggesting a role for an epigenetic mechanism(s).

Structural Alerts for the Excess Toxicity of Acrylates, Methacrylates, and Propiolates Derived from Their Short-Term and Long-Term Bacterial Toxicity

For eight acrylates, three methacrylates, and three propiolates as three subclasses of α,β-unsaturated esters, short-term and long-term bacterial toxicity with Vibrio fischeri was determined in terms of EC(50) (effective concentration 50%) values for the 30-min bioluminescence and 24-h growth inhibition. To this end, experimental exposure concentrations were corrected for volatilization through a thermodynamic model based on Henry's law constant of the compounds. Moreover, toxicity enhancements T(e) as the ratio of narcosis-predicted over actual EC(50) were determined and discussed in terms of underlying mechanisms of reaction of the electrophiles with endogenous nucleophiles such as glutathione (GSH) and proteins. Overall, log EC(50) [M] ranges from -2.28 to -3.70 (30 min) and from -2.80 to -5.28 (24 h), respectively, indicating a significantly larger sensitivity of the growth inhibition bioassay for the reactive toxicity of these Michael acceptors. The latter is also reflected in the observed toxicity enhancements, where log T(e) > 1 was obtained for only 5 of 14 30-min EC(50) values but for 11 of 13 24-h EC(50) values. Moreover, the average long-term to short-term difference in log T(e) is 1 unit for the acrylates and 0.7 units for both methacrylates and propiolates. Methacrylates exert narcosis-level toxicity except for the methyl derivative in the long-term assay. The log EC(50) (24 h) of a subset of 10 mostly excess-toxic acrylates and a propiolate correlates with their logarithmic rate constants of reaction with GSH, log k(GSH), significantly more than with log K(ow) (r(2) 0.76 vs 0.47), yielding a respective regression rms of 0.34 log units. For allyl and propargyl acrylate as well as propargyl methacrylate, the observed excess toxicity is likely caused by initial enzymatic hydrolysis and subsequent oxidation of the α,β-unsaturated alcohols to the respective carbonyls. The latter shows that in the context of nonanimal testing schemes such as for REACH, the metabolic capacity of in vitro screens requires attention.

Polysynaptic excitatory postsynaptic potentials that trigger spasms after spinal cord injury in rats are inhibited by 5-HT1B and 5-HT1F receptors

Sensory afferent transmission and associated spinal reflexes are normally inhibited by serotonin (5-HT) derived from the brain stem. Spinal cord injury (SCI) that eliminates this 5-HT innervation leads to a disinhibition of sensory transmission and a consequent emergence of unusually long polysynaptic excitatory postsynaptic potentials (EPSPs) in motoneurons. These EPSPs play a critical role in triggering long polysynaptic reflexes (LPRs) that initiate muscles spasms. In the present study we examined which 5-HT receptors modulate the EPSPs and whether these receptors adapt to a loss of 5-HT after chronic spinal transection in rats. The EPSPs and associated LPRs recorded in vitro in spinal cords from chronic spinal rats were consistently inhibited by 5-HT(1B) or 5-HT(1F) receptor agonists, including zolmitriptan (5-HT(1B/1D/1F)) and LY344864 (5-HT(1F)), with a sigmoidal dose-response relation, from which we computed the 50% inhibition (EC(50)) and potency (-log EC(50)). The potencies of 5-HT receptor agonists were highly correlated with their binding affinity to 5-HT(1B) and 5-HT(1F) receptors, and not to other 5-HT receptors. Zolmitriptan also inhibited the LPRs and general muscle spasms recorded in vivo in the awake chronic spinal rat. The 5-HT(1B) receptor antagonists SB216641 and GR127935 and the inverse agonist SB224289 reduced the inhibition of LPRs by 5-HT(1B) agonists (zolmitriptan). However, when applied alone, SB224289, SB216641, and GR127935 had no effect on the LPRs, indicating that 5-HT(1B) receptors do not adapt to chronic injury, remaining silent, without constitutive activity. The reduction in EPSPs with zolmitriptan unmasked a large glycine-mediated inhibitory postsynaptic current (IPSC) after SCI. This IPSC and associated chloride current reversed at -73 mV, slightly below the resting membrane potential. Zolmitriptan did not change motoneuron properties. Our results demonstrate that 5-HT(1B/1F) agonists, such as zolmitriptan, can restore inhibition of sensory transmission after SCI without affecting general motoneuron function and thus may serve as a novel class of antispastic drugs.

Correlation between hydrophobicity of N-alkylxanthine derivatives and their biological activities on guinea-pig isolated tracheal smooth muscle.

Cyclic (c) AMP phosphodiesterase (PDE) inhibitory activities of N-alkylxanthine derivatives (3-methyl-,3-ethyl-,3-propyl-,3-butyl-,1,3-dimethyl-,1-methyl-3-ethyl-,1-methyl - 3-propyl- and 1-methyl-3-butyl xanthines) and their relaxant effects on carbachol-induced contraction and on resting tone guinea-pig isolated tracheal smooth muscle have been investigated. The PDE inhibition constant (Ki) and the concentration producing 50% tracheal smooth muscle relaxation in-vitro (EC50) were determined. Significant correlations between the -log Ki values and the -log EC50 values on the carbachol-induced contraction or on the resting tone were found (r = 0.902 and 0.892). The apparent partition coefficient (P) between n-octanol and pH 7.4 phosphate-buffered saline (PBS) was measured as an index of hydrophobicity of the xanthine derivatives. There were significant correlations between log P and both -log EC50 values and between the log P and -log Ki values. These findings suggest that the cAMP PDE inhibitory activity of N-alkylxanthine derivatives contributes to the mechanism of bronchodilatory action, and that an increase in hydrophobicity of the xanthine molecule enhances the biological activity.

A trait database of stream invertebrates for the ecological risk assessment of single and combined effects of salinity and pesticides in South-East Australia

We compiled a database on a priori selected traits for South-East Australian freshwater macroinvertebrate families and used this data for the development of a biotic indicator for the detection of the effects of salinisation on freshwater communities (SPEAR salinity) and for the adaptation of the existing SPEAR pesticides index for South-East Australian taxa. The SPEAR salinity indicator showed a reasonably high relationship (0.38 ≤ r 2 ≤ 0.5) with salinity in terms of logarithmic electrical conductivity (log EC) using field biomonitoring data from 835 pools and riffle sites in Victoria and South Australia. Several other biotic indexes that were calculated for comparison purpose exhibited a lower relationship with log EC. In addition, SPEAR salinity was the only indicator that did not respond to other water quality variables and was therefore most selective. We used log EC data and modelled pesticide exposure for sites in Victoria in concert with SPEAR salinity and the existing SPEAR pesticides index to assess whether pesticides interacts with effects of salinity on invertebrate communities and vice versa. No interaction with pesticides was found for the effect of log EC on SPEAR salinity, whereas EC interacted with the estimated pesticide exposure on the invertebrate communities. To foster the development of further trait-based ecological indicators, we suggest a conceptual model that predicts response traits based on the disturbance regime and disturbance mode of action of the stressor. Biotic indicators based on a priori selected traits represent a promising biomonitoring tool even for regions where ecological information is scarce.

Comparative molecular field analysis to derive pharmacophore maps for induction doses of intravenous anaesthetic agents

BackgroundThe present study examines the molecular basis of induction of anaesthesia by i.v. hypnotic agents using comparative molecular field analysis (CoMFA). MethodsED50 induction doses for 14 i.v. anaesthetics in human subjects (expressed as molar dose per kilogram body weight) were obtained from the literature. Immobilizing potency data for the same 14 agents (expressed as the EC50 plasma free drug concentrations that abolish movement in response to a noxious stimulus in 50% patients) were taken from our previous publication. These data were used to form CoMFA models for the two aspects of anaesthetic activity. Molecular alignment was achieved by field-fit minimization techniques. The lead structure for both models was eltanolone. ResultsThe final CoMFA model for the ED50 induction dose was based on two latent variables, and explained 99.3% of the variance in observed activities. It showed good intrinsic predictability (cross-validated q2=0.849). The equivalent model for immobilizing activity was also based on two latent variables, with r2=0.988 and q2=0.852. Although there was a correlation between –log ED50 and –log EC50 (r2=0.779), comparison of the pharmacophore maps showed poor correlation for both electrostatic and steric regions when isocontours were constructed by linking lattice grid points, making the greatest 40% contributions; the relative contributions of electrostatic and steric interactions differing between the models (induction dose: 2.5:1; immobilizing activity 1.8:1). ConclusionsComparison of two CoMFA activity models shows only small elements of commonality, suggesting that different molecular features may be responsible for these two properties of i.v. anaesthetics.

Microwave-assisted synthesis of quinoline, isoquinoline, quinoxaline and quinazoline derivatives as CB2 receptor agonists

Quinoline, isoquinoline, quinoxaline, and quinazoline derivatives were synthesized using microwave-assisted synthesis and their CB1/CB2 receptor activities were determined using the [³⁵S]GTPγS binding assay. Most of the prepared quinoline, isoquinoline, and quinoxalinyl phenyl amines showed low-potency partial CB2 receptor agonists activity. The most potent CB2 ligand was the 4-morpholinylmethanone derivative (compound 40e) (-log EC₅₀ = 7.8; E(max) = 75%). The isoquinolin-1-yl(3-trifluoromethyl-phenyl)amine (compound 26c) was a high efficacy CB2 agonist (-log EC₅₀ = 5.8; E(max) = 128%). No significant CB1 receptor activation or inactivation was shown in these studies, except 40e, which showed weak CB1 agonist activity (CB1 -log EC₅₀ = 5.0). These ligands serve as novel templates for the development of selective CB2 receptor agonist.

Acute and Chronic Toxicity toward the Bacteria Vibrio fischeri of Organic Narcotics and Epoxides: Structural Alerts for Epoxide Excess Toxicity

The acute and chronic bacterial toxicity of 34 organic compounds comprising 19 baseline narcotics and 15 epoxides has been determined with regard to 30-min bioluminescence and 24-h growth inhibition in terms of EC50 (effective concentration 50%) values employing Vibrio fischeri. For the narcotics, linear regression of log EC50 on log Kow (octanol/water partition coefficient) yields r2 (squared correlation coefficient) and rms (root-mean-square error) values of 0.95 and 0.44 (30-min), and 0.94 and 0.34 (24-h), respectively. Employing the resultant baseline narcosis models, toxicity enhancement (Te) values were derived as a ratio of narcosis-predicted over experimental EC50 for the epoxides. For seven aliphatic epoxides, log Te was below 1 in both assays, indicating narcosis-range toxicity with regard to 30-min bioluminescence and 24-h growth inhibition. Concerning eight nonaliphatic epoxides, log Te values up to 2.4 were observed, reflecting excess toxicity through an enhanced electrophilic reactivity of the compounds. Here, however, the intercorrelation between both assays was very low (r2 = 0.09). The results are discussed in terms of electronic substituent effects activating an SN2-type epoxide reaction with nucleophilic protein sites and side-chain activation offering alternative electrophile-nucleophile reaction routes at side-chain sites, leading to respective structural alerts as indicators of excess toxicity. Surprisingly, 30-min bioluminescence appears to be slightly more sensitive to chemical stress than 24-h growth, which holds both for baseline narcotics and for most of the epoxides. This is also reflected by effective narcosis doses 50%, ED50, of 7.1 mmol/kg (30-min) and 7.7 mmol/kg (24-h) estimated from narcosis theory. Keeping in mind the different end points (bioluminescence vs growth) involved, this finding demonstrates that chronic toxicity is not always more sensitive than acute toxicity, calling for analyses with regard to further respective cases and associated mechanistic causes.

Acute toxicity and synergism of binary mixtures of antifouling biocides with heavy metals to embryos of sea urchin Glyptocidaris crenularis

Acute toxicity and synergism of four antifouling biocides (Irgarol 1051, dichlofluanid, tolylfluanid and Sea-Nine 211) and five heavy metals (Ni, Pb, Zn, Cd and Cu) are investigated using the sea urchin embryos of Glyptocidaris crenularis (G. crenularis) at six typical developmental stages, that is, 2-cell, 4-cell, 8-cell, blastula, gastrula and 4-arm pluteus. Our results show that the toxicity of the four biocides is in an order of Sea-Nine 211 > tolylfluanid > dichlofluanid > Irgarol 1051 and their -log EC(50) values at all stages are strongly linearly correlated with the 1-octanol/water partition coefficient (log P) values (correlation coefficients R > 0.72) indicating the importance of hydrophobicity for the embryonic toxicity. For the five heavy metals, the EC(50) ranges from 0.36 to 30.78 μM and the toxicity follows an order of Cu > Pb > Zn > Cd >Ni. The significant correlation (R > 0.79) between the -log EC50 and the bioconcentration factor (log BCF) values of metals also indicate that the bioaccumulation property of metals contributes to their aquatic toxicity. In addition, the joint effects of the biocides with the heavy metals in embryonic development are assessed by using a concentration addition model. Synergistic effects are observed in almost all 25 mixtures, showing that Cu yields the strongest while Ni the weakest synergistic toxic effects on the embryos development.

Experimental Physiology -Research Paper: The effect of cooling on the contractility of equine digital small lamellar arteries: modulating role of the endothelium

The equine hoof displays thermoregulatory functions, and the blood vessels lying under the hoof wall are temperature sensitive. The aim of this study was to investigate the effect of cooling on the contractile responses to α-adrenoceptor and 5-HT receptor stimulation in equine small lamellar arteries using wire myography. The role of the endothelium in the response mediated by 5-HT was also evaluated. Moderate cooling caused a reduction of the contraction induced by depolarizing Krebs solution (DKS, containing 118 mm KCl) and the maximal contraction caused by UK-14304 (α(2)-adrenoceptor agonist). The potency of methoxamine (α(1)-adrenoceptor agonist) was reduced by cooling [pD(2) (-log EC(50)) at 22°C, 5.7 (5.5-6.0) versus 30°C, 5.9 (5.7-6.1)]; however, the efficacy was maintained. The contractions evoked by sumatripan and α-methyl 5-HT (5-HT receptor agonists) were not modified by moderate cooling. In contrast, a cooling-enhanced contraction was observed in response to 5-HT [maximum response (E(max)) at 22°C, 213.2 ± 13% DKS versus 30°C, 179.9 ± 11% DKS]. Furthermore, this effect was more evident in endothelium-denuded lamellar arteries (E(max) at 22°C, 270.2 ± 26% DKS versus 30°C, 172.2 ± 20% DKS), suggesting a potential modulating role of the endothelium. The L-NAME/ibuprofen-resistant relaxation in response to carbachol was reduced by cooling in small lamellar arteries precontracted with 5-HT but not phenylephrine. Therefore, a moderate reduction of temperature modulates the reactivity of small lamellar arteries by enhancing the 5-HT-mediated contraction, but inhibits the α-adrenoceptor-mediated response. Furthermore, the endothelium of these blood vessels may play an important role in preventing excessive vasoconstriction in response to 5-HT and maintaining digital blood flow in cool environmental temperatures.

Prediction of metal cation toxicity to the bioluminescent fungus Gerronema viridilucens

A correlation between the physicochemical properties of mono- [Li(I), K(I), Na(I)] and divalent [Cd(II), Cu(II), Mn(II), Ni(II), Co(II), Zn(II), Mg(II), Ca(II)] metal cations and their toxicity (evaluated by the free ion median effective concentration, EC50(F)) to the naturally bioluminescent fungus Gerronema viridilucens has been studied using the quantitative ion character-activity relationship (QICAR) approach. Among the 11 ionic parameters used in the current study, a univariate model based on the covalent index (X(2) (m)r) proved to be the most adequate for prediction of fungal metal toxicity evaluated by the logarithm of free ion median effective concentration (log EC50(F)): log EC50(F) = 4.243 (± 0.243) -1.268 (± 0.125)· X(2) (m)r (adj-R(2) = 0.9113, Alkaike information criterion [AIC] = -60.42). Additional two- and three-variable models were also tested and proved less suitable to fit the experimental data. These results indicate that covalent bonding is a good indicator of metal inherent toxicity to bioluminescent fungi. Furthermore, the toxicity of additional metal ions [Ag(I), Cs(I), Sr(II), Ba(II), Fe(II), Hg(II), and Pb(II)] to G. viridilucens was predicted, and Pb was found to be the most toxic metal to this bioluminescent fungus (EC50(F)): Pb(II) > Ag(I) > Hg(I) > Cd(II) > Cu(II) > Co(II) ≈ Ni(II) > Mn(II) > Fe(II) ≈ Zn(II) > Mg(II) ≈ Ba(II) ≈ Cs(I) > Li(I) > K(I) ≈ Na(I) ≈ Sr(II)> Ca(II).