Presence Of ACh Research Articles

The Efficacy of I Na Block to Cardiovert Atrial Fibrillation Is Enhanced by Inhibition of I K1.

There is a need for more efficient pharmacological cardioversion of atrial fibrillation (AF). We tested the hypothesis that inhibition of I K1 significantly enhances the efficacy of I Na block to depress atrial excitability and to cardiovert AF. The study was conducted in canine isolated arterially perfused right atrial preparations with rim of ventricular tissue. AF was induced in the presence of acetylcholine (ACh; 0.5 µM). BaCl 2 (10 µM) was used to inhibit I K1 and flecainide (1.5 µM) to block I Na . Sustained AF (>45 minutes) was recorded in 100% atria (5/5) in the presence of ACh alone. Flecainide cardioverted AF in 50% of atria (4/8), BaCl 2 in 0% (0/5), and their combination in 100% (5/5). AF cardioversion occurred in 15 ± 9 minutes with flecainide alone (n = 4) and in 8 ± 9 minutes with the combination (n = 5). Following drug-induced AF cardioversion, AF was inducible in 4/4 atria with flecainide alone (≤5 minutes duration) and in 2/5 atria with the combination (≤30 seconds duration). Atrial excitability was significantly more depressed by combined versus monotherapies. There was little to no effect on ventricular excitability under any condition tested. Thus, inhibition of I K1 significantly enhances the efficacy of flecainide to depress atrial excitability and to cardiovert AF in our experimental setting. A combination of I Na and I K1 inhibition may be an effective approach for cardioversion of AF.

An elegant approach for the synthesis of multisubstituted imidazole via FeCl3/SiO2 catalyzed activation of acetals: a photophysical study of an imidazole-carbazole hybrid.

A simple and solvent-free catalytic system was developed for the direct conversion of multisubstituted imidazoles through the reaction of acetals and benzils with ammonium acetate/amines as the source of nitrogen. The reaction occurred under mild and benign conditions using FeCl3/SiO2 as a heterogeneous catalyst without the requirement of any toxic organic solvents. The easy preparation and recyclability of the catalyst allows the reaction to be simple and highly efficient, resulting in very good yields of imidazoles. Novel imidazole-carbazole hybrid compounds were also synthesised by adopting the present methodology. Single crystal X-ray diffraction study indicated the presence of a CH⋯π supramolecular interaction that renders effective molecular packing in the solid state. The steady-state and spectro-dynamic behaviours of these hybrid molecules were investigated, and it was revealed that a solvent-dependent excimer-coupled ICT phenomenon guided excited state photophysics. Very unusual excimer lifetime was noticed in the solid state of this bis-heterocyclic compound owing to the stacking of molecules via CH⋯π interaction, as evident from the X-ray studies.

A κ-OR Agonist Protects the Endothelial Function Impaired by Hyperuricemia Through Regulating the Akt/eNOS Signal Pathway.

To investigate the effects of κ-OR agonist on hyperuricemia rats and injured endothelial function, as well as the underlying mechanism. A hyperuricemia model was established on rats. The endothelial protective effects of U50,488H were evaluated and compared to the controlled groups. The protein levels of eNOS, p-eNOS, Akt, and p-Akt were determined using western blot analysis. ELISA was employed to measure the expression of soluble ET-1, ICAM-1, TNF-α, and NO in cell supernatants and rat serum samples. Cell migration and the artery tension were determined by in vitro functional assays. The suppressed production of ET-1, ICAM-1, and NO in the hyperuricemia rats was promoted by the treatment of U50,488H, which was reversed by the co-administration of nor-BNI. P-eNOS/eNOS and p-Akt/Akt were up-regulated by the incubation of serum from hyperuricemia rats, which was down-regulated by the introduction of U50,488H. The vascular tension of vessels incubated with U50,488H was higher than the baseline in the presence of ACh, which was lower than baseline in the presence of SNAP. U50,488H significantly promoted the release of ET-1, ICAM-1, and NO, and inhibited the release of TNF-α from endothelial cells and the migration ability of neutrophils in the presence of hyperuricemia rat serum, which were reversed by the co-incubation with nor-BNI, Akt inhibitor or L-NAME. U50,488H protected the endothelial function impaired by hyperuricemia through regulating the Akt/eNOS signal pathway.

Supramolecular Synthon Identification in Azelaic Acid – Isonicotinamide

Supramolecular synthons are the smallest structural units that contain the information inherent in building large molecular assemblies. Here, we apply solid-state NMR methods for the identification of the synthon in a cocrystal of Azelaic acid [(CH2)7(COOH)2]– Isonicotinamide [C6H6N2O]. It is observed that the spin-lattice relaxation time of protons in the cocrystal is long, implying that the choice of experiments, particularly those that involve low-sensitivity nuclei, could be time consuming. It is noticed that the two dimensional 1H double quantum (DQ) – 1H single quantum (SQ) correlation experiment at very fast MAS (60 kHz) is able to provide information on proton-proton proximities that leads to an understanding of the synthon structure. Experiments such as the CP-Reverse CP and 1H–13C HETCOR have been used to obtain spectral assignment which was additionally confirmed by first-principle GIPAW NMR chemical shift calculations. Present study indicates the basic unit to be a heterosynthon in Azelaic acid (AZA) – Isonicotinamide (INIC) cocrystal formed by two different modes of intermolecular connectivity, viz. acid–pyridine and acid–amide hydrogen bonding interactions. The presence of a CH…O type bond involving a hydrogen of INIC is also indicated from the observation of a larger chemical shift of the corresponding proton. The experimental protocol presented in this paper is expected to be a useful tool for the estimation of the basic structural arrangement of the synthons in a rapid fashion.

MnO2/DNAzyme-mediated ratiometric fluorescence assay of acetylcholinesterase.

Acetylcholinesterase (AChE) is an essential serine hydrolase associated with neurodegenerative diseases and serves as a biomarker. In this work, a ratiometric fluorescence strategy for the sensitive assay of AChE activity is developed based on the decomposition of MnO2 by the enzymatic hydrolysis product and DNAzyme-mediated strand cleavage. The sensing system is termed as MnO2/DNAzyme. DNAzyme locked by H2 strands is inactive in the absence of AChE. The fluorescence emission of fluorescein amidite (FAM)-labeled DNA (DNA-F) at 518 nm is quenched by neutral red (NR) and the fluorescence of NR at 632 nm is simultaneously enhanced due to fluorescence resonance energy transfer (FRET). The presence of AChE triggers the hydrolysis of the substrate acetylcholine (ATCh) to enzymatic thiocholine (TCh), which reduces MnO2 nanowires to Mn2+ and releases the attached H1 strands into the solution. The H1 strands hybridize with H2 strands through the strand displacement reaction. Meanwhile, the activated DNAzyme cleaves the RNA nucleotide of the DNA-F signal probe to release FAM. The fluorescence of FAM at 518 nm is thus recovered, corresponding to a decrement of NR emission at 632 nm owing to the blocking of FRET. The fluorescence ratio of F518/F632 serves as a signal readout for the AChE assay within 5 × 10-4-10 U mL-1, with a limit of detection (LOD) of 2.7 × 10-4 U mL-1. The feasibility of this method was demonstrated by the measurement of AChE activity in human blood, which reveals its promising potential in clinical assays.

The production of nitric oxide in the coeliac ganglion modulates the effect of cholinergic neurotransmission on the rat ovary during the preovulatory period

The aim of the present work was to investigate whether the nitric oxide produced by the nitric oxide/nitric oxide synthase (NO/NOS) system present in the coeliac ganglion modulates the effects of cholinergic innervation on oxidative status, steroidogenesis and apoptotic mechanisms that take place in the rat ovary during the first proestrous. An ex vivo Coeliac Ganglion- Superior Ovarian Nerve- Ovary (CG-SON-O) system was used. Cholinergic stimulation of the CG was achieved by 10−6 M Acetylcholine (Ach). Furthermore, 400 μM Aminoguanidine (AG) – an inhibitor of inducible-NOS was added in the CG compartment in absence and presence of Ach. It was found that Ach in the CG compartment promotes apoptosis in ovarian tissue, probably due to the oxidative stress generated. AG in the CG compartment decreases the release of NO and progesterone, and increases the release of estradiol from the ovary. The CG co-treatment with Ach and AG counteracts the effects of the ganglionic cholinergic agonist on ovarian oxidative stress, increases hormone production and decreases Fas mRNA expression. These results suggest that NO is an endogenous modulator of cholinergic neurotransmission in CG, with implication in ovarian steroidogenesis and the apoptotic mechanisms that take place in the ovary during the preovulatory period in rats.

The effect of hydroalcoholic extract of Achillea eriophora DC. on blood pressure of anaesthetized male rat.

Achillea eriophora (Asteraceae) is a medicinal plant commonly used in Iran. This study was performed to determine the cardiovascular effects of hydroethanolic extract of A. eriophora (HEAE) and the underlying mechanisms in anaesthetized rats. The acute effects of intravenous (i.v.) administration of different doses of HEAE (40, 50, 60, 80 mg/kg), and its probable interaction with cholinergic and nitrergic systems were investigated in the presence of ACh and NOS blocker (L-NAME) as well as ethanol (HEAE solvent in sham group). Intravenous administration of different doses of HEAE induced hypotension. HEAE (60 mg/kg) significantly reduced mean arterial blood pressure (MAP), systolic arterial blood pressure (SBP) and diastolic arterial blood pressure (DBP) compared to control rats that treated with ethanol only, but no change in heart rate (HR) was seen in both groups. The results showed significant decrease in MAP, SBP, DBP and increase of HR in the presence of HEAE plus ACh (10 µg/kg) compared to when ACh was injected alone. Finally i.v. administration of HEAE, significantly reduced MAP and DBP in L-NAME (5 mg/kg) treated animals, while bradycardic responses to L-NAME were not significantly changed by HEAE. It can be concluded that Achillea eriophora induced hypotensive effect via lowering total peripheral resistance and cardiac output that may be synergist with cholinergic and independent of nitrergic system.

Oxidative stress and altered steroidogenesis in the ovary by cholinergic stimulation of coeliac ganglion in the first proestrous in rats. Implication of nitric oxide

An ex-vivo Coeliac Ganglion-Superior Ovarian Nerve-Ovary (CG–SON–O) system from virgin rats in the first proestrous was used to test whether cholinergic stimulation of CG affects oxidative status and steroidogenesis in the ovary. The CG and the O were placed in separate buffered-compartments, connected by the SON, and the CG was stimulated by acetylcholine (Ach). To test a possible role of nitric oxide (NO) in the ovarian response to cholinergic stimulation of CG, aminoguanidine (AG) – an inhibitor of inducible-NO synthase was added to the O compartment. After 180 min incubation, the oxidative status was assessed in O whereas nitrite and steroidogenesis were assessed at 30, 120 and 180 min. Ach in CG decreased the total antioxidant capacity, but increased NO production and protein carbonization in O. Ach stimulation of CG increased estradiol, but decreased progesterone release in O by reducing the mRNAs related to their synthesis and degradation. The addition of AG to the O compartment caused an opposite effect, which was more pronounced in the presence of Ach in the CG compartment than in its absence. These results show that the stimulation of the extrinsic-cholinergic innervation of the O increases the concentration of NO, causes oxidative stress and modulates steroidogenesis in the first rat proestrous.

Regulation of nerve-evoked contractions of rabbit vas deferens by acetylcholine.

Stimulation of intramural nerves in the vas deferens of many species yields a classical biphasic contraction comprised of an initial fast component, mediated by P2X receptors and a second slower component, mediated by α1-adrenoceptors. It is also recognized that sympathetic nerve-mediated contractions of the vas deferens can be modulated by acetylcholine (Ach), however there is considerable disagreement in the literature regarding the precise contribution of cholinergic nerves to contraction of the vas deferens. In this study we examined the effect of cholinergic modulators on electric field stimulation (EFS)-evoked contractions of rabbit vas deferens and on cytosolic Ca2+ levels in isolated vas deferens smooth muscle cells (VDSMC). The sustained component of EFS-evoked contractions was inhibited by atropine and by the selective M3R antagonist, 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (4-DAMP). EFS-evoked contractions were potentiated by Ach, carbachol (Cch), and neostigmine. The sustained phase of the EFS-evoked contraction was inhibited by prazosin, an α1-adrenoceptor antagonist and guanethidine, an inhibitor of noradrenaline release, even in the continued presence of Ach, Cch or neostigmine. The soluble guanylate cyclase (sGC) inhibitor, 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one enhanced the amplitude of EFS-evoked contractions and reduced the inhibitory effects of 4-DAMP. Isolated VDSMC displayed spontaneous Ca2+ oscillations, but did not respond to Cch. However, the α1-adrenoceptor agonist, phenylephrine, evoked a Ca2+ transient and contracted the cells. These data suggest that EFS-evoked contractions of the rabbit vas deferens are potentiated by activation of M3 receptors and reduced by activation of a sGC-dependent inhibitory pathway.

Experimental evidence and mathematical modeling of thermal effects on human colonic smooth muscle contractility.

It has been shown, in animal models, that gastrointestinal tract (GIT) motility is influenced by temperature; nevertheless, the basic mechanism governing thermal GIT smooth muscle responses has not been fully investigated. Studies based on physiologically tuned mathematical models have predicted that thermal inhomogeneity may induce an electrochemical destabilization of peristaltic activity. In the present study, the effect of thermal cooling on human colonic muscle strip (HCMS) contractility was studied. HCMSs were obtained from disease-free margins of resected segments for cancer. After removal of the mucosa and serosa layers, strips were mounted in separate chambers. After 30 min, spontaneous contractions developed, which were measured using force displacement transducers. Temperature was changed every hour (37, 34, and 31°C). The effect of cooling was analyzed on mean contractile activity, oscillation amplitude, frequency, and contraction to ACh (10(-5) M). At 37°C, HCMSs developed a stable phasic contraction (~0.02 Hz) with a significant ACh-elicited mean contractile response (31% and 22% compared with baseline in the circular and longitudinal axis, respectively). At a lower bath temperature, higher mean contractile amplitude was observed, and it increased in the presence of ACh (78% and 43% higher than the basal tone in the circular and longitudinal axis, respectively, at 31°C). A simplified thermochemomechanical model was tuned on experimental data characterizing the stress state coupling the intracellular Ca(2+) concentration to tissue temperature. In conclusion, acute thermal cooling affects colonic muscular function. Further studies are needed to establish the exact mechanisms involved to better understand clinical consequences of hypothermia on intestinal contractile activity.

Colorimetric and fluorometric assays for acetylcholinesterase and its inhibitors screening based on a fluorescein derivate

A fluorescein-based sensor was developed for the AChE activity assay and the inhibitor screening. The sensor provided the dual assay methods for the screening of AChE activity in the presence or absence of inhibitor. The colorimetric and fluorometric assays were based on the following processes: (1) owing to the hydrolysis of acetylthiocholine in the presence of AChE, the fluorescein-based probe can rapidly induce 1,4-addition of the hydrolysis product thiocholine to α,β-unsaturated ketone in the compound 1, resulting in strong fluorescence and absorption changes; (2) in the presence of the corresponding inhibitor, the fluorescence enhancement or the absorption change would be inhibited in that the formation of thiocholine was hindered.

Anticholinesterase Activity of Phenolic Acids and their Derivatives

The ability of 36 phenolic acids and their derivatives to inhibit acetyl- and butyrylcholinesterase was studied. The most efficient acetylcholine inhibitors were: carnosic acid = gentisic acid > 3-hydroxy-4-methoxycinnamic acid = ethyl ferulate = ethyl vanillate = nordihydroguaiaretic acid > ethyl 4-hydroxybenzoate = methyl ferulate. The order of effectiveness towards butyrylcholinesterase was: carnosic acid > nordihydroguaiaretic acid = ethyl ferulate > salicylic acid > gentisic acid > rosmarinic acid = caftaric acid > homogentisic acid. The inhibitory activity was dependent on the number/position of OH or/and OCH3 groups attached to a phenol ring. It can be speculated that OCH3 substitution in the phenol ring can promote a higher antibutyrylcholinesterase activity (although not statistically confirmed at p < 0.05). The presence of a CH=CH-COOH group had a highly favourable effect on the antiacetylcholinesterase activity compared with a CH2-CH2-COOH or a COOH group. Methyl and ethyl esters were more potent inhibitors than the corresponding free acids. The molecular weight of the compounds (in the range of M = 154.12 - 474 g/mol) played a minor role in this context.

The Effect of Achromobacter xylosidans in Adult Cystic Fibrosis Lung Transplant Recipients

Purpose Acromobacter xylosidans is a multidrug resistant gram-negative rod that has been recently described to colonize or infect the airways of 10-29% of cystic fibrosis (CF) patients. We describe our single center experience of CF lung transplant recipients colonized with this bacterium prior to transplantation. Methods and Materials CF patients who were transplanted between 1995-2011 at our institution were reviewed. Patients with pre-transplant A. xylosidans colonization (ACH) were compared to those who did not harbor this pathogen (non-ACH). Variables collected included: time to positive ACH culture post-transplant, duration of intravenous (IV) antibiotics, number of inpatient days, and survival. Duration of IV antibiotics and the number of inpatient days were analyzed by a Mann-Whitney U test, while survival was compared using Kaplan-Meier analysis. Results 48 of 474 patients had a CF diagnosis; 10 (21%) were colonized with ACH prior to transplantation. All 10 grew the organism after transplantation at a median of 125 days (3-1055 days) as detected by bronchoalveolar lavage performed during surveillance bronchoscopy. The ACH group was hospitalized a greater number of days after transplantation ( Table 1 ) and required significantly longer courses of IV antibiotics compared to the non-ACH group (p=0.048). ACH patients also showed a trend towards worse survival (p = 0.085) with separation between the groups noted three years after transplantation. Conclusions Longer post-operative hospital stays, a prolonged antibiotic course, and the possibility of poor mid-term survival when compared to non-ACH-CF transplant recipients, suggests a negative impact by the presence of ACH in CF transplant recipients. ACH (n=10) Non-ACH (n=38) P value Inpatient days, median (range) 87 (37-171) 50 (12-250) 0.084 Weeks of IV antibiotics, median (range) 12 (3-42) 7.5 (2-25) 0.049

High-Speed 2-Dimentional Observation of Stepwise Motions in Single nAChR and AChBP using Diffracted X-Ray Tracking (DXT)

The nicotinic acetylcholine receptor (nAChR) is a pentameric ligand-gated ion channel in the central and the peripheral nervous system. The structure of nAChR has been determined by Miyazawa et al in 2003. There are also similar proteins to nAChR such as GluCl and protein from some bacteria. Based on the structural analysis of these proteins, two models were proposed to explain the gating mechanism of nAChR. One is Rotation and the other is Tilt. In this study, we directly observed single molecular motion of nAChR and AChBP using diffracted X-ray tracking (DXT). DXT is a method to detect single molecular motions.There are two important characteristics of the technique. First, both tilt and rotation can be observed. Second, the spacial and time resolutions of DXT are extremely high. In fact, theoretically, 100 pm scale measurements are possible and time resolution of the technique is now sub micro seconds.We applied DXT to AChBP and nAChR in the time scale of 100μs which is assumed channel opening time from the study of patch cramp. We observed the motion of AChBP and nAChR in the presence and absence of ACh. In the presence of ACh, both tilting and rotational motions are activated. Thus, we think combination of tilt and rotate is essential for the gating mechanism. In addition to the data, we observed interesting trajectories. They have three stepwise motions, tilt, rotate and tilt again. These spots can not only explain our data but also support both tilt and rotate models.

Desensitization of nicotinic acetylcholine receptors in central nervous system neurons of the stick insect ( Carausius morosus) by imidacloprid and sulfoximine insecticides

Imidacloprid, sulfoxaflor and two experimental sulfoximine insecticides caused generally depressive symptoms in stick insects, characterized by stillness and weakness, while also variably inducing postural changes such as persistent ovipositor opening, leg flexion or extension and abdomen bending that could indicate excitation of certain neural circuits. We examined the same compounds on nicotinic acetylcholine receptors in stick insect neurons, which have previously been shown to desensitize in the presence of ACh. Brief U-tube application of 10 −4 M solutions of insecticides for 1 s evoked currents that were much smaller than ACh-evoked currents, and depressed subsequent ACh-evoked currents for several minutes, indicating that the compounds are low-efficacy partial agonists that potently desensitize the receptors. Much lower concentrations of insecticides applied in the bath for longer periods did not activate currents, but inhibited ACh-evoked currents via desensitization of the receptors. Previously described fast- and slowly-desensitizing nACh currents, I ACh1 and I ACh2 respectively, were each found to consist of two components with differing sensitivities to the insecticides. Imidacloprid applied in the bath desensitized high-sensitivity components, I ACh1H and I ACh2H with IC 50s of 0.18 and 0.13 pM, respectively. It desensitized the low-sensitivity slowly desensitizing component, I ACh2L, with an IC 50 of 2.6 nM, while a component of the fast-desensitizing current, I ACh1L, was least sensitive, with an IC 50 of 81 nM I ACh1L appeared to be insensitive to the three sulfoximines tested, whereas all three sulfoximines potently desensitized I ACh1H and both slowly desensitizing components, with IC 50s between 2 and 7 nM. We conclude that selective desensitization of certain nAChR subtypes can account for the insecticidal actions of imidacloprid and sulfoximines in stick insects.

Glutamine 57 at the Complementary Binding Site Face Is a Key Determinant of Morantel Selectivity for α7 Nicotinic Receptors

Nicotinic receptors (AChRs) play key roles in synaptic transmission. We explored activation of neuronal α7 and mammalian muscle AChRs by morantel and oxantel. Our results revealed a novel action of morantel as a high efficacy and more potent agonist than ACh of α7 receptors. The EC50 for activation by morantel of both α7 and α7-5HT3A receptors is 7-fold lower than that determined for ACh. The minimum morantel concentration required to activate α7-5HT3A channels is 6-fold lower than that of ACh, and activation episodes are more prolonged than in the presence of ACh. By contrast, oxantel is a weak agonist of α7 and α7-5HT3A, and both drugs are very low efficacy agonists of muscle AChRs. The replacement of Gln57 in α7 by glycine, which is found in the equivalent position of the muscle AChR, decreases the efficacy for activation and turns morantel into a partial agonist. The reverse mutation in the muscle AChR (ϵG57Q) increases 7-fold the efficacy of morantel. The mutations do not affect activation by ACh or oxantel, indicating that this position is selective for morantel. In silico studies show that the tetrahydropyrimidinyl group, common to both drugs, is close to Trp149 of the principal face of the binding site, whereas the other cyclic group is proximal to Gln57 of the complementary face in morantel but not in oxantel. Thus, position 57 at the complementary face is a key determinant of the high selectivity of morantel for α7. These results provide new information for further progress in drug design.

Modulation of erythrocyte deformability by PKC activity

The interactions between membrane, peripheral and cytoskeleton proteins are responsible for the maintenance of erythrocyte deformability (EEI) and some of these interactions are modulated by PKC activity. Protein band 3 of the erythrocyte membrane is phosphorylated by phosphotyrosine kinases (PTK) and dephosphorylated by phosphotyrosine phosphatase (PTP). It was previously described by us a signal transduction mechanism that describes a possible pathway connecting an erythrocyte external membrane protein, acetylcholinesterase (AChE), with protein band 3. So how does PKC activity modulate EEI when protein band 3 is phosphorylated or dephosphorylated in absence or presence of AChE effectors?To answer this we used phorbol 12-myristate 13-acetate (PMA) as an activator and chelerythrine chloride as inhibitor of PKC and also band 3 modulators of band 3 phosphorylation degree, in presence and absence of AChE effectors in order to measure in whole blood samples EEI. Our results showed that erythrocyte deformability was significantly (i) decreased by inhibition of PKC, in absence and presence of AChE inhibitor velnacrine (ii) increased with PMA in absence and presence of ACh and (iii) decreased in presence of calpeptin in absence and presence of either chelerythrine or PMA. These results establish dependence between cytoskeleton proteins, PKC activity, band 3 phosphorylation degrees and EEI. Better understanding of those proteins interactions on transduction mechanisms might trigger possible targets for drug action that would modulate EEI.

1H nuclear magnetic resonance relaxation investigation of huperzine E binding to acetylcholinesterase

In order to search for better acetylcholinesterase (AchE) inhibitors, the binding properties of AchE with huperizine E, which is a derivative of huperzine A, were investigated with 1H nuclear magnetic resonance (1H NMR) method. The nonselective, selective and double-selective spin-lattice relaxation rates of some protons in huperzine E were acquired in the absence and presence of AchE at a concentration ratio of [ligand]/[protein] = 1: 0.005. The enhancements of selective relaxation rates of these protons were obvious after adding AchE. The molecular motional correlation times of two pairs of protons, H-1a/H-1b and H-2/H-3, in the bound state at T = 298 K were 11.7 and 9.46 ns respectively, while they were 27.7 and 35.2 ps in the free state. All of these show that huperzine E has high binding affinity with AchE.

Essential oil of Pterodon polygalaeflorus inhibits electromechanical coupling on rat isolated trachea

The present work studied the effects of the essential oil of Pterodon polygalaeflorus (EOPP), a plant used to treat bronchitis and amigdalytis, on rat airway smooth muscle in vitro. In Ca 2+-containing medium, EOPP (100–1300 μg/ml) inhibited preferentially high KCl- than 5-HT-induced muscle contractions in a concentration-dependent fashion, but did affect neither basal muscle tension nor ACh-induced contractions. In preparations maintained in either 60 mM K + or 10 μM ACh in Ca 2+-free medium, EOPP (100, 600 and 1300 μg/ml) inhibited maximum contractile response induced by cumulative Ca 2+ addition (0.1–20 mM). Verapamil (10, 30 and 100 μg/ml), a Ca 2+ channel blocker, also inhibited Ca 2+-induced concentration–effect curve in presence of ACh in Ca 2+-free medium, whilst it was ineffective to decrease cholinergic contractions in Ca 2+-containing medium. In presence of 150 mM K + in Ca 2+-containing medium, EOPP (1300 μg/ml) did not reversed ACh-induced contractions. In contrast, under similar conditions, EOPP almost fully relaxed cholinergic contractions of tracheal smooth muscle in Ba 2+-containing medium. In medium containing 10 mM tetraethylammonium and 2 mM Ba 2+ instead of Ca 2+, both EOPP (1300 μg/ml) and verapamil (∼5 μg/ml) significantly decreased ACh-induced contractions. Thus, in rat isolated trachea, EOPP induces inhibitor effects on contractions preferentially triggered by an electromechanical coupling mode.

Angiotensin II-induced modulation of endothelium-dependent relaxation in rabbit mesenteric resistance arteries

The role of local endogenous angiotensin II (Ang II) in endothelial function in resistance arteries was investigated using rabbit mesenteric resistance arteries. First, the presence of immunoreactive Ang II together with Ang II type-1 receptor (AT1R) and angiotensin converting enzyme (ACE) was confirmed in these arteries. In endothelium-intact strips, the AT1R-blocker olmesartan (1 μm) and the ACE-inhibitor temocaprilat (1 μm) each enhanced the ACh (0.03 μm)-induced relaxation during the contraction induced by noradrenaline (NA, 10 μm). Similar effects were obtained using CV-11974 (another AT1R blocker) and enalaprilat (another ACE inhibitor). The nitric-oxide-synthase inhibitor NG-nitro-l-arginine (l-NNA) abolished the above effect of olmesartan. In endothelium-denuded strips, olmesartan enhanced the relaxation induced by the NO donor NOC-7 (10 nm). Olmesartan had no effect on cGMP production (1) in endothelium-intact strips (in the absence or presence of ACh) or (2) in endothelium-denuded strips (in the absence or presence of NOC-7). In β-escin-skinned strips, 8-bromoguanosine 3′,5′ cyclic monophosphate (8-Br-cGMP, 0.01–1 μm) concentration dependently inhibited the contractions induced (a) by 0.3 μm Ca2+ in the presence of NA+GTP and (b) by 0.2 μm Ca2++GTPγS. Olmesartan significantly enhanced, while Ang II (0.1 nm) significantly inhibited, the 8-Br-cGMP-induced relaxation. We propose the novel hypothesis that in these arteries, Ang II localized within smooth muscle cells activates AT1Rs and inhibits ACh-induced, endothelium-dependent relaxation at least partly by inhibiting the action of cGMP on these cells.