Synthesis of [Tl4Se16]: a novel tetranuclear Tl polyselenide

(Ph₄P)₄[Tl₄Se₁₆] was prepared hydrothermally in a sealed pyrex tube by the reaction of TlCl, K₂Se₄ and Ph₄PCl in a 1:1:1 molar ratio at 110 °C for one day. The red crystals were obtained in 50% yield. Crystals of (Ph₄P)₄[Tl₄Se₁₆]: triclinic P (No. 2), Z=1, a=12.054(9), b=19.450(10), c=11.799(6) Å, α=104.63(4), β=98.86(6), γ=101.99(6)° and V=2555(3) ų at 23 °C, 2θ_max=40.0°, μ=120.7 cm⁻¹, D_calc=2.23. The structure was solved by direct methods. Number of data collected: 5206. Number of unique data having F_o²>3σ(F_o²): 1723. Final R=0.075 and R_w=0.089. [Tl₄Se₁₆]⁴⁻ consists of four, almost already linearly arranged, tetrahedral thallium centers which are coordinated by two chelating Se₄²⁻, two bridging Se₂²⁻ and four bridging Se²⁻ ligands. [Tl₄Se₁₆]⁴⁻ sits on an inversion center and possesses a central {Tl₂Se₂}²⁺ planar core. The Tl(1)–Tl(1)′ distance in this core is 3.583(6) Å. These two thallium atoms are then each linked to two cyclic Tl(Se₄) fragments via bridging Se₂²⁻ and Se²⁻ ligands forming Tl₂Se(Se₂) five-membered rings.     

Direct oxidation of boronates by peroxynitrite: Mechanism and implications in fluorescence imaging of peroxynitrite

In this study, we show that boronates, a class of synthetic organic compounds, react rapidly and stoichiometrically with peroxynitrite (ONOO⁻) to form stable hydroxy derivatives as major products. Using a stopped-flow kinetic technique, we measured the second-order rate constants for the reaction with ONOO⁻, hypochlorous acid (HOCl), and hydrogen peroxide (H₂O₂) and found that ONOO⁻ reacts with 4-acetylphenylboronic acid nearly a million times (k = 1.6 × 10⁶ M^− 1 s^− 1) faster than does H₂O₂ (k = 2.2 M^− 1 s^− 1) and over 200 times faster than does HOCl (k = 6.2 × 10³ M^− 1 s^− 1). Nitric oxide and superoxide together, but not alone, oxidized boronates to the same phenolic products. Similar reaction profiles were obtained with other boronates. Results from this study may be helpful in developing a novel class of fluorescent probes for the detection and imaging of ONOO⁻ formed in cellular and cell-free systems.     

Gill (Na+,K+)-ATPase from the blue crab Callinectes danae: modulation of K+-phosphatase activity by potassium and ammonium ions

The kinetic properties of a microsomal gill (Na⁺,K⁺)-ATPase from the blue crab Callinectes danae were analyzed using the substrate p-nitrophenylphosphate. The (Na⁺,K⁺)-ATPase hydrolyzed PNPP obeying cooperative kinetics (n=1.5) at a rate of V=125.4±7.5 U mg⁻¹ with K_0.5=1.2±0.1 mmol l⁻¹; stimulation by potassium (V=121.0±6.1 U mg⁻¹; K_0.5=2.1±0.1 mmol l⁻¹) and magnesium ions (V=125.3±6.3 U mg⁻¹; K_0.5=1.0±0.1 mmol l⁻¹) was cooperative. Ammonium ions also stimulated the enzyme through site–site interactions (n_H=2.7) to a rate of V=126.1±4.8 U mg⁻¹ with K_0.5=13.7±0.5 mmol l⁻¹. However, K⁺-phosphatase activity was not stimulated further by K⁺ plus NH₄⁺ ions. Sodium ions (K_I=36.7±1.7 mmol l⁻¹), ouabain (K_I=830.3±42.5 μmol l⁻¹) and orthovanadate (K_I=34.0±1.4 nmol l⁻¹) completely inhibited K⁺-phosphatase activity. The competitive inhibition by ATP (K_I=57.2±2.6 μmol l⁻¹) of PNPPase activity suggests that both substrates are hydrolyzed at the same site on the enzyme. These data reveal that the K⁺-phosphatase activity corresponds strictly to a (Na⁺,K⁺)-ATPase in C. danae gill tissue. This is the first known kinetic characterization of K⁺-phosphatase activity in the portunid crab C. danae and should provide a useful tool for comparative studies.     

Oxygen and carbon dioxide transport by the haemocyanin of an amphibious crab,Holthuisana transversa

Summary The oxygen and carbon dioxide transporting properties of the haemolymph from an amphibious Australian crab,Holthuisana transversa were investigated. Within the temperature range 15 to 35°C increasing temperature markedly decreased oxygen affinity (H=–54 kJ·mol^–1). The Bohr effect was small at all temperatures with a mean value of –0.13. Over the temperature range 15–35°C there was a significant increase in the cooperativity of oxygen binding. Changing the concentration of Ca,l-lactate or haemocyanin in the haemolymph could elicit no significant change in either O₂ affinity or cooperativity of O₂ binding. There was no evidence in support of a specific effect of CO₂ on oxygen affinity of either non-dialysed or dialysed haemolymph.The amount of CO₂ that could be carried byH. transversa haemolymph was significantly reduced by increased temperature (approx. 14 to 12.5 mmol·l^–1 CO₂). Comparisons of oxygenated and deoxygenated haemolymph at a fixed pH were unable to demonstrate the presence of a significant Haldane effect. Combining data from oxygenated and deoxygenated haemolymph the buffer value was calculated to be in the range –6.2 to –8.5 mmol·l^–1 HCO ₃ ^– ·pH unit^–1.The insensitivity ofH. transversa haemocyanin function to all modulating influences except temperature is discussed with respect to the ecology of this crab.     

Metabolic rate, maximum metabolism, and advantages of torpor in the fat mouse Steatomys pratensis natalensis Roberts, 1921 (Dendeomurinae)

1. The fat mouse Steatomys pratensis natalensis (mean body mass 37.4±0.43 (se)) has a low euthermic body temperature T_b=30.1–33.8 °C and a low basal metabolic rate (BMR)=0.50 ml O₂ g⁻¹ h⁻¹.

2. Below an ambient temperature (T_a)=15 °C, the mice were hypothermic.

3. The lowest survivable T_a=10 °C.

4. Torpor is efficient in conserving energy between T_a=15–30 °C, below T_a=15 °C, the mice arouse.

5. Euthermic and torpid mice were hyperthermic at T_a=35 °C.

6. Thermal conductance was 0.159 ml O₂ g⁻¹ h⁻¹ °C⁻¹, 98.8% of the expected value.

7. Non-shivering thermogenesis (NST) was 2.196 ml O² g⁻¹ h⁻¹ (3.69×BMR).

8. Maximal oxygen consumption, however, was 3.83 ml O₂ g⁻¹ h⁻¹ (6.44×BMR), indicating that other methods of heat production are additive.

9. Because fat mice conserve energy by torpor only between T_a=15–30 °C, we suggest that torpor may be a more important mechanism for surviving food shortages than for surviving cold weather.

Effects of thiosulphate on haemocyanin oxygen affinity in the isopod Saduria entomon (L.) and the brown shrimp Crangon crangon (L.)

The sulphide-tolerant brackish water isopod Saduria entomon has haemocyanin with a high affinity for oxygen. Sulphide is oxidized in the hepatopancreas to thiosulphate. Oxygen needed for this oxidation is transported bound to haemocyanin. Thiosulphate was found to improve haemocyanin oxygen affinity in S. entomon. The modulating effect was time and concentration dependent. The effect was greater at higher pH, i.e. the Bohr effect was greater (−1.82) compared to that seen without thiosulphate (−1.36). S. entomon exposed to simultaneous severe hypoxia and sulphide had a low haemolymph pH and the partial pressure of oxygen required to reach 50% saturation was even lower than that measured in vitro. This is ascribed to the combined effects of thiosulphate and lactate as well as unknown co-factors. Thiosulphate had no effect at all in the sulphide and hypoxia non-tolerant natantian Crangon crangon. It is suggested that thiosulphate can be important as a haemocyanin modulator for crustaceans digging in sulphide-rich sediments or constantly exposed to sulphide. Accepted: 11 August 1999     

Long-term exposure of the freshwater shrimp Macrobrachium olfersii to elevated salinity: Effects on gill (Na,K)-ATPase α-subunit expression and K-phosphatase activity

The kinetic properties of a microsomal gill (Na⁺,K⁺)-ATPase from the freshwater shrimp, Macrobrachium olfersii, acclimated to 21‰ salinity for 10 days were investigated using the substrate p-nitrophenylphosphate. The enzyme hydrolyzed this substrate obeying cooperative kinetics at a rate of 123.6 ± 4.9 U mg^− 1 and K_0.5 = 1.31 ± 0.05 mmol L^− 1. Stimulation of K⁺-phosphatase activity by magnesium (V_max = 125.3 ± 7.5 U mg^− 1; K_0.5 = 2.09 ± 0.06 mmol L^− 1), potassium (V_max = 134.2 ± 6.7 U mg^− 1; K_0.5 = 1.33 ± 0.06 mmol L^− 1) and ammonium ions (V_max = 130.1 ± 5.9 U mg^− 1; K_0.5 = 11.4 ± 0.5 mmol L^− 1) was also cooperative. While orthovanadate abolished p-nitrophenylphosphatase activity, ouabain inhibition reached 80% (K_I = 304.9 ± 18.3 μmol L^− 1). The kinetic parameters estimated differ significantly from those for freshwater-acclimated shrimps, suggesting expression of different isoenzymes during salinity adaptation. Despite the ≈2-fold reduction in K⁺-phosphatase specific activity, Western blotting analysis revealed similar α-subunit expression in gill tissue from shrimps acclimated to 21‰ salinity or fresh water, although expression of phosphate-hydrolyzing enzymes other than (Na⁺,K⁺)-ATPase was stimulated by high salinity acclimation.     

Synthesis and crystal structure of a complex between lanthanum nitrate and a bicyclic polyether

Reaction between lanthanum nitrate hexahydrate and a macrobicyclic polyether in ethanol has yielded a product of overall stoichiometry 3:2. The ligand, 21R, 26S, 29R, 34S-21, 22, 23, 24, 25, 26, 29, 30, 31,32,33, 34-dodecahydro-1,4,7,14,17,20,28,35-octaoxa (2^3,29syn2^18,34syn) (7.7) orthocyclophane, L, provides 8 oxygen donor atoms. Crystals were obtained from MeOH/EtOH (50/50). Crystal structure determination on 9590 observations, R=0.059, has shown the triclinic unit cell, a = 26.562(6), b = 13.486(3), c = 12.154(3) Å, α=63.9(1), β = 100.0(1), γ = 102.0(1)° space group P , V = 3806 Å to contain, as the asymmetric unit, two complex cations (La(NO₃)₂L)⁺ and one complex anion (La- (NO₃)₅MeOH)²⁻. The lanthanum is 11-coordinated in the anion and one of the cations, in which there is one bidentate and one monodentate nitrate anion and 12-coordinated in the other cation. For the monodentate nitrate La---O = 2.448(9) Å, all other nitrate ions are bidentate (La---O = 2.594(9)−2.743(10) Å). Most La---O bonds are shorter in the 11 than in the 12-coordinated cation. There are large differences in La---O bond lengths according to the nature of the carbon atoms to which the oxygen is attached. The methanol molecule forms a hydrogen bond to one oxygen atom of the monodentate nitrate group.     

The X-ray structure analysis of bis-2,2′,N,N′-bipyridyl ketone cobalt(III) nitrate dihydrate

An X-ray structural analysis of bis-2,2′,N,N′-bipyridyl ketone cobalt(III) nitrate dihydrate, CoC₂₂H₂₀N₄O₄⁺· NO₃⁻·2H₂O,M_r=559.38 g/mol, P , a=8.862(2), b=16.195(3), c=8.772(2) Å, α=103.54(2), β=95.74(3), γ=105.07°, V=1164.4(4) ų, Z=2, D_x=1.595 g/cm³, Mo Kα radiation (λ=0.71073 Å), μ=7.8 cm⁻¹ and R=0.079, revealed a Co(III) cation in a slightly distorted octahedral environment. The structure reveals that the ligand di-2-pyridyl ketone (dpk) has undergone a hydration reaction across the ketone double bond and one of the hydrate oxygen atoms coordinated to the metal forming a tridentate chelate. This new Co(dpk-hydrate)₂⁺ complex displays the least distorted geometry yet reported for either 1:1 or 1:2 (metal:ligand) complexes. A geometry optimization using the INDO model Hamiltonian as implemented in the program ZINDO was performed on the title complex with the Co³⁺ modeled as a singlet. The result of the computation corroborates the geometry of the title complex as that expected for Co³⁺.     

Thiopurine methyl transferase activity: new extraction conditions for high-performance liquid chromatographic assay

A new liquid–liquid extraction is described for thiopurine methyl transferase (TPMT, EC 2.1.1.67) activity determination: the use of a pH 9.5 NH₄Cl buffer solution, before adding the solvent mixture, allows more rapid extraction, avoiding a centrifugation step, and reduces the global cost of analysis. After the extraction step, 6-methylmercaptopurine, synthesised during the enzymatic reaction, is determined by a liquid chromatographic assay. Analytical performance of the assay was tested on spiked erythrocyte lysates. The linear concentration range was 5–250 ng ml ⁻¹ (r≥0.997, slope=1.497, intercept=−0.367). The recoveries were 82.8, 89.9 and 82.2% for 75, 125 and 225 ng ml⁻¹, respectively. The coefficients of variation were ≤6.1% for within-day assay (n=6) and ≤9.5% for between-day assay precision (n=6; 14 days). TPMT activity was determined in a French adult Caucasian population (n=70). The results ranged from 7.8 to 27.8 nmol h⁻¹ ml⁻¹ packed red blood cells and the frequency distribution histogram is similar to that previously published.     

Comparison of oxygen mass transfer coefficient in simple and extractive fermentation systems

Aeration and agitation are important variables to ensure effective oxygen transfer rate during aerobic bioprocesses; therefore, the knowledge of the volumetric mass transfer coefficient (k_La) is required. In view of selecting the optimum oxygen requirements for extractive fermentation in aqueous two-phase system (ATPS), the k_La values in a typical ATPS medium were compared in this work with those in distilled water and in a simple fermentation medium, in the absence of biomass. Aeration and agitation were selected as the independent variables using a 2² full factorial design. Both variables showed statistically significant effects on k_La, and the highest values of this parameter in both media for simple fermentation (241 s⁻¹) and extractive fermentation with ATPS (70.3 s⁻¹) were observed at the highest levels of aeration (5 vvm) and agitation (1200 rpm). The k_La values were then used to establish mathematical correlations of this response as a function of the process variables. The exponents of the power number (N³D²) and superficial gas velocity (V_s) determined in distilled water (α = 0.39 and β = 0.47, respectively) were in reasonable agreement with the ones reported in the literature for several aqueous systems and close to those determined for a simple fermentation medium (α = 0.38 and β = 0.41). On the other hand, as expected by the increased viscosity in the presence of polyethylene glycol, their values were remarkably higher in a typical medium for extractive fermentation (α = 0.50 and β = 1.0). A reasonable agreement was found between the experimental data of k_La for the three selected systems and the values predicted by the theoretical models, under a wide range of operational conditions.     

Removal of municipal solid waste COD and NH4–N by phyto-reduction: A laboratory–scale comparison of terrestrial and aquatic species at different organic loads

Laboratory scale tests on phytodepuration of raw and pre-treated leachate from municipal sanitary waste were carried out with four vegetable aquatic and terrestrial species at different organic loads. We used the terrestrial species Stenotaphrum secundatum and the free-floating aquatic species Lemna minor, Eichhornia crassipes and Myriophyllum verticellatum to purify leachate from municipal solid waste. The organic load characterized by COD varied from 2–30 g m⁻² day⁻¹. Blanks using tap water served as controls. Duration of the experiments varied from 9–90 days. Maximum concentrations in the experiments were 1600 mg l⁻¹ COD and 300 mg l⁻¹ NH₄–N for S. secundatum. Best results in terms of COD, BOD, and ammonia removal were obtained for raw leachate with COD=2 g m⁻² day⁻¹ in free water surface (FWS) wetlands, and with 2 and 5 g m⁻² day⁻¹ in subsurface flow (SSF) wetlands. Results show that for pretreated leachate (labeled c) low in BOD and NH₄–N, the aquatic species showed low removal and stress even at the lowest load of COD=2 g m⁻² day⁻¹. We cannot say if this is due to the pretreatment itself or the chemical or microbial composition of this leachate. The Stenotaphrum system operated well with this load of leachate c. For untreated leachate (type a and b) the removal and plant growing conditions seemed good at COD=2 g m⁻² day⁻¹. For S. secundatum a load of COD=5 g m⁻² day⁻¹ operated well. All loads above COD=5 g m⁻² day⁻¹ caused low removal and stress, and the green parts of the plants disappeared. Oxygen was, however, consumed throughout the experimental period. For pretreated leachate (type c), the removal of COD were low (−24 to 17%) but good for NH₄–N (52–91%). This leachate also experienced high ammonia removal from the beginning of the experiments, probably due to existing consortia of nitrifying bacteria in it. Statistical analysis shows that the S. secundatum and L. minor systems maintained higher oxygen levels than the M. verticellatum and E. crassipes systems, when operated with tap water. For Lemna minor, this may be due to a better capacity for transporting oxygen into the water. With leachate all S. secundatum systems have higher oxygen levels than the aquatic systems, basically because the water content of the soil has been kept well below saturation. S. secundatum shows a significantly lower removal of COD than did the aquatic systems at a loading of COD=2 g m⁻² day⁻¹ of raw leachate. There is no significant difference between the systems in the removal of NH₄–N at a loading of COD=2 g m⁻² day⁻¹ of both types of leachate. E. crassipes has a lower removal of NH₄–N than M. verticellatum and S. secundatum at a loading of 5 g m⁻² day⁻¹ of COD of both types of leachate. In our experiments, it appears that the amount of free ammonia explains the toxicity of the leachate to the plants. This, however, does not exclude other possible toxic factors.     

The effects of intensity of exercise on excess postexercise oxygen consumption and energy expenditure in moderately trained men and women

This experiment investigated the effects of intensity of exercise on excess postexercise oxygen consumption (EPOC) in eight trained men and eight women. Three exercise intensities were employed 40%, 50%, and 70% of the predetermined maximal oxygen consumption (VO_2max). All ventilation measured was undertaken with a standard, calibrated, open circuit spirometry system. No differences in the 40%, 50% and 70% VO_2max trials were observed among resting levels of oxygen consumption (V0₂) for either the men or the women. The men had significantly higher resting VO₂ values being 0.31 (SEM 0.01) 1·min^–1 than did the women, 0.26 (SEM 0.01) 1·min^–1 (P < 0.05). The results indicated that there were highly significant EPOC for both the men and the women during the 3-h postexercise period when compared with resting levels and that these were dependent upon the exercise intensity employed. The duration of EPOC differed between the men and the women but increased with exercise intensity: for the men 40% – 31.2 min; 50% – 42.1 min; and 70% – 47.6 min and for the women, 40% – 26.9 min; 50% – 35.6 min; and 70% – 39.1 min. The highest EPOC, in terms of both time and energy utilised was at 70% VO_2max. The regression equation for the men, where y=O₂ in litres, and x=exercise intensity as a percentage of maximum was y=0.380x + 1.9 (r ²=0.968) and for the women is y=0.374x–0.857 (r ²=0.825). These findings would indicate that the men and the women had to exercise at the same percentage of their VO_2max to achieve the maximal benefits in terms of energy expenditure and hence body mass loss. However, it was shown that a significant EPOC can be achieved at moderate to low exercise intensities but without the same body mass loss and energy expenditure.     

Surface-enhanced Raman spectroscopy combined with atomic force microscopy for ultrasensitive detection of thrombin

Indole-3-acetic acid (IAA) amide conjugates play an important role in balancing levels of free IAA in plant cells. The GH3 family of proteins conjugates free IAA with various amino acids. For example, auxin levels modulate expression of the Oryza sativa (rice) GH3-8 protein, which acts to prevent IAA accumulation by coupling the hormone to aspartate. To examine the kinetic properties of the enzyme, we developed a liquid chromatography–tandem mass spectrometry (LC–MS/MS) assay system. Bacterially expressed OsGH3-8 was purified to homogeneity and used to establish the assay system. Monitoring of the reaction confirms the reaction product as IAA–Asp and demonstrates that production of the conjugate increases proportionally with both time and enzyme amount. Steady-state kinetic analysis using the LC–MS/MS-based assay yields the following parameters: V/E_t^IAA = 20.3 min⁻¹, K_m^IAA = 123 μM, V/E_t^ATP = 14.1 min⁻¹, K_m^ATP = 50 μM, V/E_t^Asp = 28.8 min⁻¹, K_m^Asp = 1580 μM. This is the first assignment of kinetic values for any IAA–amido synthetase from plants. Compared with previously described LC- and thin-layer chromatography (TLC)-based assays, this LC–MS/MS method provides a robust and sensitive means for performing direct kinetic studies on a range of IAA-conjugating enzymes.     

Nitrogen utilization by the raphidophyte Heterosigma akashiwo: Growth and uptake kinetics in laboratory cultures

The nitrogen uptake and growth capabilities of the potentially harmful, raphidophycean flagellate Heterosigma akashiwo (Hada) Sournia were examined in unialgal batch cultures (strain CCMP 1912). Growth rates as a function of three nitrogen substrates (ammonium, nitrate and urea) were determined at saturating and sub-saturating photosynthetic photon flux densities (PPFDs). At saturating PPFD (110 μE m⁻² s⁻¹), the growth rate of H. akashiwo was slightly greater for cells grown on NH₄⁺ (0.89 d⁻¹) compared to cells grown on NO₃⁻ or urea, which had identical growth rates (0.82 d⁻¹). At sub-saturating PPFD (40 μE m⁻² s⁻¹), both urea- and NH₄⁺-grown cells grew faster than NO₃⁻-grown cells (0.61, 0.57 and 0.46 d⁻¹, respectively). The N uptake kinetic parameters were investigated using exponentially growing batch cultures of H. akashiwo and the ¹⁵N-tracer technique. Maximum specific uptake rates (V_max) for unialgal cultures grown at 15 °C and saturating PPFD (110 μE m⁻² s⁻¹) were 28.0, 18.0 and 2.89 × 10⁻³ h⁻¹ for NH₄⁺, NO₃⁻ and urea, respectively. The traditional measure of nutrient affinity—the half saturation constants (K_s) were similar for NH₄⁺ and NO₃⁻ (1.44 and 1.47 μg-at N L⁻¹), but substantially lower for urea (0.42 μg-at N L⁻¹). Whereas the α parameter (α = V_max/K_s), which is considered a more robust indicator for substrate affinity when substrate concentrations are low (<K_s), were 19.4, 12.2 and 6.88 × 10⁻³ h⁻¹/(μg-at N L⁻¹) for NH₄⁺, NO₃⁻ and urea, respectively. These laboratory results demonstrate that at both saturating and sub-saturating N concentrations, N uptake preference follows the order: NH₄⁺ > NO₃⁻ > urea, and suggests that natural blooms of H. akashiwo may be initiated or maintained by any of the three nitrogen substrates examined.     

Structure and properties of a novel OH bridged dimetal helical complex with 6,6-dimethyl-2,2′:6′,2″:6″,2:6,2-quinquepyridine ligand

A new monohelical OH⁻ bridged dinuclear complex [Zn₂(dmqpy)(OOCCH₃)₂(μ-OH)][ClO₄] · 0.5EtOH, where dmqpy is 6,6-dimethyl-2,2′:6′,2″:6″,2:6,2-quinquepyridine, has been synthesized and characterized by X-ray crystallography: monoclinic, space group P2₁/c, a=13.670(1), b=14.751(1), c=16.782(1) Å, β=96.59(1)°, U=3361.7(4) ų, Z=4, R=0.0601. Two Zn(II) ions are in different coordination modes, one is five-coordinate with a N₃O₂ donor set and the other is N₂O₂ four-coordinate with a distorted tetrahedral geometry, and the zinc ions are bridged by a hydroxyl group. The presence of the OH⁻ bridge is further confirmed by electrospray mass and infrared spectroscopies. The solution properties of the complex were investigated by ¹H NMR spectroscopy. The results of NMR indicate that the complex has higher symmetry in solution than in the solid state.     

Comparisons of the release of vasodilator substances from left and right cardiac chambers of the isolated perfused rabbit heart: Implications for intraventricular thrombus formation

Prostacyclin (PgI₂) and endothelium-derived nitric oxide (EDNO) are produced by the arterial and venous endothelium. In addition to their vasodilator action on vascular smooth muscle, both act together to inhibit platelet aggregation and promote platelet disaggregation. EDNO also inhibits platelet adhesion to the endothelium. EDNO and PgI₂ have been shown to be released from the cultured endocardial cells. In this study, we examined the release of vasoactive substances from the intact endocardium by using isolated rabbit hearts perfused with physiological salt solution (95% O₂/5% CO₂, T = 37 °C). The right and left cardiac chambers were perfused through separate constant-flow perfusion loops (physiological salt solution, 8 ml min⁻¹). Effluent from left and right cardiac, separately, was bioassayed on canine coronary artery smooth muscle, which had been contracted with prostaglandin F_{2α_}(2 × 10⁻⁶ M) and no change in tension was exhibit. However, addition of calcium ionophore A23187 (10⁻⁶ M) to the cardiac chambers’ perfusion line induced vasodilation of the bioassay coronary ring, 61.4 ± 7.4% versus 70.49 ± 6.1% of initial prostaglandin F_2α contraction for the left and right cardiac chambers perfusate, respectively (mean ± SEM, n = 10, p > 0.05). Production of vasodilator was blocked totally in the left heart but, only partially blocked in the right heart by adding indomethacin (10⁻⁵ M) to the perfusate, respectively, 95.2 ± 2.2% versus 41.5 ± 4.8% (mean ± SEM, n = 10, p < 0.05). 6-Keto prostaglandin F_1α, measured in the endocardial superfusion effluent was also higher for the left cardiac chambers than for the right at the time of stimulation with the A23187, respectively, 25385.88 ± 5495 pg/ml (n = 8) versus 13,132.45 ± 1839.82 pg/ml (n = 8), (p < 0.05). These results showed that cyclooxygenase pathway plays major role in generating vasoactive substances for the left cardiac chamber endocardium; while it is not the main pathway for the right ventricular endocardium at which EDNO and PgI₂ could act together and potentiate their antithrombogenic activities in isolated perfused rabbit heart. This may be an explanation for the intraventricular thrombus mostly seen in left ventricle rather than in right ventricle as a complication of myocardial infarction.     

Active NaCl absorption across split lamellae of posterior gills of Chinese crabs (Eriocheir sinensis) adapted to different salinities

Split lamellae of posterior gills of Eriocheir sinensis adapted to fresh water, brackish waters (9 or 18‰) or seawater (36‰) were mounted in Ussing chambers, and transepithelial short-circuit currents and conductances were measured with salines, containing approximately in vivo-like NaCl concentrations. Active sodium and chloride absorption (I_Na and I_Cl), the transcellular conductances and the leak conductance were identified with external amiloride and/or DIDS. Split gill lamellae of crabs adapted to fresh water displayed similar magnitudes of I_Na and I_Cl with 10 mmol l⁻¹ NaCl in the external medium (internally haemolymph-like NaCl saline). Augmenting external NaCl (50 mmol l⁻¹) resulted in an increase of I_Cl, whereas I_Na decreased. Split gill lamellae of crabs adapted to brackish waters (external NaCl of 125 and 225 mmol l⁻¹, respectively) showed lower currents than preparations of freshwater crabs (50 mmol l⁻¹ external NaCl). With split gill lamellae of seawater crabs no currents were detected (450 mmol l⁻¹ NaCl on both sides). The transcellular conductances showed similar changes as the currents. The leak conductance of split gill lamellae of crabs adapted to fresh or brackish waters was low (0.3–0.8 mS cm⁻²), whereas it was much higher (7 mS cm⁻²) with preparations of seawater crabs.     

3-(1′,1′-Dimethylbutyl)-1-deoxy-Δ-THC and related compounds: synthesis of selective ligands for the CB2 receptor

The synthesis and pharmacology of 15 1-deoxy-Δ⁸-THC analogues, several of which have high affinity for the CB₂ receptor, are described. The deoxy cannabinoids include 1-deoxy-11-hydroxy-Δ⁸-THC (5), 1-deoxy-Δ⁸-THC (6), 1-deoxy-3-butyl-Δ⁸-THC (7), 1-deoxy-3-hexyl-Δ⁸-THC (8) and a series of 3-(1′,1′-dimethylalkyl)-1-deoxy-Δ⁸-THC analogues (2, n=0–4, 6, 7, where n=the number of carbon atoms in the side chain−2). Three derivatives (17–19) of deoxynabilone (16) were also prepared. The affinities of each compound for the CB₁ and CB₂ receptors were determined employing previously described procedures. Five of the 3-(1′,1′-dimethylalkyl)-1-deoxy-Δ⁸-THC analogues (2, n=1–5) have high affinity (K_i=<20 nM) for the CB₂ receptor. Four of them (2, n=1–4) also have little affinity for the CB₁ receptor (K_i=>295 nM). 3-(1′,1′-Dimethylbutyl)-1-deoxy-Δ⁸-THC (2, n=2) has very high affinity for the CB₂ receptor (K_i=3.4±1.0 nM) and little affinity for the CB₁ receptor (K_i=677±132 nM).

Scheme 3. (a) (C₆H₅)₃PCH₃⁺ Br⁻, n-BuLi/THF, 65°C; (b) LiAlH₄/THF, 25°C; (c) KBH(sec-Bu)₃/THF, −78 to 25°C then H₂O₂/NaOH.     

Erythrocytic ATP release in the presence of modified cell-free hemoglobin

The red blood cell (RBC) has been proposed as an O₂ sensor through a direct link between the desaturation of intracellular hemoglobin (Hb) and ATP release, leading to vasodilation. We hypothesized that the addition of cell-free Hb to the extracellular space provides a supplementary O₂ source that reduces RBC desaturation and, consequently, ATP release. In this study, the saturation of RBC suspensions was lowered by additions of deoxygenated hemoglobin-based oxygen carrier (HBOC) and then assayed for extracellular ATP. When an acellular human Hb intramolecularly cross-linked between α subunits (ααHb, p50 = 33 mmHg) was added to the red cell suspension, ATP production was significantly less than that in the presence of a lower p50 HBOC (Hb cross-linked between β subunits, ββHb, p50 = 8 mmHg). These results provide a potential mechanism for the O₂ affinity of HBOCs to interfere with a vasodilatory signal.