Effects of chronic run training on Na+-dependent Ca2+ efflux from rat left ventricular myocytes

The effects of endurance run training on Na+-dependent Ca2+ regulation in rat left ventricular myocytes were examined. Myocytes were isolated from sedentary and trained rats and loaded with fura 2. Contractile dynamics and fluorescence ratio transients were recorded during electrical pacing at 0.5 Hz, 2 mM extracellular Ca2+ concentration, and 29 degreesC. Resting and peak cytosolic Ca2+ concentration ([Ca2+]c) did not change with exercise training. However, resting and peak [Ca2+]c increased significantly in both groups during 5 min of continuous pacing, although diastolic [Ca2+]c in the trained group was less susceptible to this elevation of intracellular Ca2+. Run training also significantly reduced the rate of [Ca2+]c decay during relaxation. Myocytes were then exposed to 10 mM caffeine in the absence of external Na+ or Ca2+ to trigger sarcoplasmic reticular Ca2+ release and to suppress cellular Ca2+ efflux. This maneuver elicited an elevated steady-state [Ca2+]c. External Na+ was then added, and the rate of [Ca2+]c clearance was determined. Run training significantly reduced the rate of Na+-dependent clearance of [Ca2+]c during the caffeine-induced contractures. These data demonstrate that the removal of cytosolic Ca2+ was depressed with exercise training under these experimental conditions and may be specifically reflective of a training-induced decrease in the rate of cytosolic Ca2+ removal via Na+/Ca2+ exchange and/or in the amount of Ca2+ moved across the sarcolemma during a contraction.     

An evaluation of the allergic contact dermatitis potential of colloidal grain suspensions

OBJECTIVE: To assess the efficacy of Kangke Injection in treating viral myocarditis. METHODS: Kangke Injection is the effective ingredient extracted from Radix Sophora flavescens (RSF). Seventy-six cases of virus myocarditis suffering from the continuous positive Coxsackie B virus ribose nucleic acid-polymerase chain reaction (CBVRNA-PCR) in blood, their peak value in blood was determined by high performance liquid chromatography (HPLC) and compared with 50 cases treated by glucose-insulin-potassium chloride (GIK). RESULTS: The clearance rate of CBVRNA-PCR and RSF was dose-dependent. The effective rate of RSF on palpitation, chest distress, dispnea was 96.02%, and that of arrhythmia was 100%, all of them were better than those of control. After RSF therapy, the parameters of heart function of ejection fraction (EF), stroke volume (SV), cardiac output (CO), and cardiac index (CI) elevated significantly (P < 0.01), left ventricular mass (LVM) and left ventricular mass index (LVMI) were decreased statistically significantly, while after 5 month therapy, the anti-Coxsackie group B virus neutralizing antibodies of RSF group was returned to normal titer, natural killer (NK) cell activity elevated, P < 0.01. CONCLUSION: RSF was an effective substance for regressing the "Pathologic status" of viral myocarditis.     

Acid-base and electrolyte effects of shortening steeplechase in a three-day-event

This study was designed to characterise the acid-base and electrolyte effects of shortening the distance required during steeplechase (Phase B) in the face of hot and humid weather conditions during a treadmill-simulated Speed and Endurance test. Eight conditioned Thoroughbred horses underwent 3 randomised permutations of a standardised exercise test on a high speed treadmill. Each test consisted of trotting at 3.7 m/s for 10 min (Phase A); galloping at 11 m/s (Phase B) for 4 (cool laboratory conditions), 3 (hot and humid), or 2 (hot and humid) min; trotting at 3.7 m/s for 30 min (Phase C); and walking at 1.8 m/s for 10 min (Phase X). The treadmill slope was 4% for trotting and galloping and 0% for walking. Cool versus hot and humid conditions were 20 degrees C and 50-60% relative humidity vs. 26-28 degrees C and 80-85% relative humidity, respectively. Pulmonary artery blood samples were obtained at rest prior to exercise (Rest); at the end of Phases A (A10) and B (B2-4); at 10 (C10), 20 (C20) and 30 (C30) min through Phase C; and at 5 min into Phase X (X5). Additional samples for lactate (LA) and glucose (GLC) analysis were obtained 5 min into Phase C (C5) and at the end of Phase X (X10). Samples were analysed for packed cell volume (PCV), haemoglobin (HB), total plasma protein (TP), sodium (Na), potassium (K), chloride (Cl), anion gap (AG), plasma glucose (GLC) and lactate (LA), pH, PCO2, bicarbonate (HCO3) and base excess (BE). Shortening steeplechase distance by 50% under hot and humid conditions (2 min B) resulted in a consistent return to control measurements (4 min B) only for plasma LA. Changes in PCV, HB, TP, K and Cl were related more to the longer galloping distance in the 4 min B trials than to hot vs. cold laboratory conditions. Alternatively, changes in LA, GLC, pH, PCO2 and AG were more related to hot and humid laboratory conditions than they were to galloping distance. These latter variables, when combined with physical measures such as core temperature, bodyweight loss, point of fatigue on Phase C and recovery heart rates may serve as the best monitors of positive responses in future studies of proposed modifications to Phase C, rather than those variables which were more distance than weather-related.     

The role of an alpha subtype M2-M3 His in regulating inhibition of GABAA receptor current by zinc and other divalent cations

Sensitivity of GABAA receptors (GABARs) to inhibition by zinc and other divalent cations is influenced by the alpha subunit subtype composition of the receptor. For example, alpha6beta3gamma2L receptors are more sensitive to inhibition by zinc than alpha1beta3gamma2L receptors. We examined the role of a His residue located in the M2-M3 extracellular domain (rat alpha6 H273) in the enhanced zinc sensitivity conferred by the alpha6 subtype. The alpha1 subtype contains an Asn (N274) residue in the equivalent location. GABA-activated whole-cell currents were obtained from L929 fibroblasts after transient transfection with expression vectors containing GABAA receptor cDNAs. Mutation of alpha1 (alpha1(N274H)) or alpha6 (alpha6(H273N)) subtypes did not alter the GABA EC50 of alphabeta3gamma2L receptors. alpha1(N274H)beta3gamma2L receptor currents were as sensitive to zinc as alpha6beta3gamma2L receptor currents, although alpha6(H273N)beta3gamma2L receptor currents had the reduced zinc sensitivity of alpha1beta3gamma2L receptor currents. We also examined the activity of other inhibitory divalent cations with varying alpha subtype dependence: nickel, cadmium, and copper. alpha6beta3gamma2L receptor currents were more sensitive to nickel, equally sensitive to cadmium, and less sensitive to copper than alpha1beta3gamma2L receptor currents. Studies with alpha1 and alpha6 chimeric subunits indicated that the structural dependencies of the activity of some of these cations were different from zinc. Compared with alpha6beta3gamma2L receptor currents, alpha6(H273N)beta3gamma2L receptor currents had reduced sensitivity to cadmium and nickel, but the sensitivity to copper was unchanged. Compared with alpha1beta3gamma2L receptor currents, alpha1(N274H)beta3gamma2L receptor currents had increased sensitivity to nickel, but the sensitivity to cadmium and copper was unchanged. These findings indicate that H273 of the alpha6 subtype plays an important role in determining the sensitivity of recombinant GABARs to the divalent cations zinc, cadmium, and nickel, but not to copper. Our results also suggest that the extracellular N-terminal domain of the alpha1 subunit contributes to a regulatory site(s) for divalent cations, conferring high sensitivity to inhibition by copper and cadmium.     

Action of prostanoids on the emetic reflex of Suncus murinus (the house musk shrew)

Several prostanoids were investigated for a potential to induce emesis in Suncus murinus. The TP receptor agonist 11alpha,9alpha-epoxymethano-15S-hydroxyprosta-5Z,13E-dienoic acid (U46619) induced emesis at doses as low as 3 microg/kg, i.p. but the DP receptor agonist 5-(6-Carboxyhexyl)-1-(3-cyclohexyl-3-hydroxypropyl) hydantoin (BW245C) was approximately 1000 times less potent. The emetic action of U46619 (300 microg/kg, i.p.) was antagonized significantly by the TP receptor antagonist, vapiprost (P<0.05). EP (prostaglandin E(2), 17-phenyl-omega-trinor prostaglandin E(2), misoprostol and sulprostone), FP (prostaglandin F(2alpha) and fluprostenol) and IP (iloprost and cicaprost) receptor agonists failed to induce consistent emesis at doses up to 300-1000 microg/kg, i.p. Fluprostenol reduced nicotine (5 mg/kg, s.c.)-but not copper sulphate (120 mg/kg, intragastric)-induced emesis; the other inconsistently emetic prostanoids were inactive to modify drug-induced emesis. The results indicate an involvement of TP and possibly DP and FP receptors in the emetic reflex of S. murinus.     

Inhibition of sickle beta-chain (betaS)-dependent polymerization by nonhuman alpha-chains. A superinhibitory mouse-horse chimeric alpha-chain

Horse alpha-chain inhibits sickle beta-chain-dependent polymerization; however, its inhibitory potential is not as high as that of mouse alpha-chain. Horse alpha-(1-30) and alpha-(31-141) segments make, respectively, minor and major contributions to the inhibitory potential of horse alpha-chain. The sum of the inhibitory potential of the two segments does not account for the inhibitory potential of the full-length horse alpha-chain. Although the polymerization inhibitory potential of horse alpha-chain is lower than mouse alpha-chain, the inhibitory potential of horse alpha-(31-141) is comparable to that of mouse alpha-(31-141). When mouse alpha-(1-30) is stitched to horse alpha-(31-141), the product is a chimeric alpha-chain with an inhibitory potential greater than mouse alpha-chain. In contrast, the stitching of horse alpha-(1-30) with mouse alpha-(31-141) had no additional inhibitory potential. Molecular modeling studies of HbS containing the mouse-horse chimeric alpha-chain indicate altered side-chain interactions at the alpha1beta1 interface when compared with HbS. In addition, the AB/GH corner perturbations facilitate a different stereochemistry for the interaction of the epsilon-amino group of Lys-16(alpha) with the beta-carboxyl group of Asp-116(alpha), resulting in a decrease in the accessibility of the side chain of Lys-16(alpha) to the solvent. Based on molecular modeling, we speculate that these perturbations by themselves, or in synergy with the altered conformational aspects of the alpha1beta1 interactions, represent the molecular basis of the superinhibitory potential of the mouse-horse chimeric alpha-chains.     

Role of active site tyrosine residues in catalysis by human glutathione reductase

Tyr114 and Tyr197 are highly conserved residues in the active site of human glutathione reductase, Tyr114 in the glutathione disulfide (GSSG) binding site and Tyr197 in the NADPH site. Mutation of either residue has profound effects on catalysis. Y197S and Y114L have 17% and 14% the activity of the wild-type enzyme, respectively. Mutation of Tyr197, in the NADPH site, leads to a decrease in Km for GSSG, and mutation of Tyr114, in the GSSG site, leads to a decrease in Km for NADPH. This behavior is predicted for enzymes operating by a ping-pong mechanism where both half-reactions partially limit turnover. Titration of the wild-type enzyme or Y114L with NADPH proceeds in two phases, Eox to EH2 and EH2 to EH2-NADPH. In contrast, Y197S reacts monophasically, showing that excess NADPH fails to enhance the absorbance of the thiolate-FAD charge-transfer complex, the predominant EH2 form of glutathione reductase. The reductive half-reactions of the wild-type enzyme and of Y114L are similar; FAD reduction is fast (approximately 500 s-1 at 4 degreesC) and thiolate-FAD charge-transfer complex formation has a rate of 100 s-1. In Y197S, these rates are only 78 and 5 s-1, respectively. The oxidative half-reaction, the rate of reoxidation of EH2 by GSSG, of the wild-type enzyme is approximately 4-fold faster than that of Y114L. These results are consistent with Tyr197 serving as a gate in the binding of NADPH, and they indicate that Tyr114 assists the acid catalyst His467'.