Characterization of L-arginine uptake by plasma membrane vesicles isolated from cultured pulmonary artery endothelial cells

We investigated the mechanisms of [3H]-L-arginine transport via System Y+ using plasma membrane vesicles derived from cultured pulmonary artery endothelial cells. [3H]-L-arginine uptake into plasma membrane vesicles was Na-independent, sensitive to trans-stimulation, unaffected by proton-conducting ionophores, and selectively inhibited by cationic amino acids. Kinetic experiments performed over a wide range of substrate concentrations revealed only one population of L-arginine transporters with Km = 130 microM. To elucidate the driving force for L-arginine transport, we measured [3H]-L-arginine uptake by plasma membrane vesicles at different transmembrane ion gradients. Plasma membrane vesicles accumulated [3H]-L-arginine only when a membrane potential was imposed across the vesicles, and the velocity of uptake was linearly related to the magnitude of the created membrane potential. The presence of potassium ions inside the vesicles was not essential for uptake of L-arginine into vesicles, but it was essential for trans-stimulation of L-arginine transport. [3H]-L-arginine accumulated in plasma membrane vesicles can be released by agents that dissipate transmembrane potassium gradients (e.g. saponin, gramicidin, and nigericin). Diazoxide and pinacidil, activators of K(+)-channels, had no significant effect on [3H]-L-arginine uptake, whereas tetraethylammonium chloride, 4-aminopyridine, and glibenclamide, inhibitors of K(+)-channels, caused decreases in [3H]-L-arginine transport by plasma membrane vesicles. This study demonstrates for the first time a specific role for potassium ions in the mechanism of L-arginine transport, particularly in the phenomenon of trans-stimulation.     

Amlodipine dynamic effects and myocardial pharmacokinetics in the isolated and perfused guinea-pig heart

Myocardial dynamic effects and pharmacokinetics of amlodipine were studied in the isolated retrogradely perfused and spontaneously beating guinea-pig heart. Pharmacokinetic analysis of drug accumulation showed one-compartment characteristics with an half-life of 76 min. Whereas disposition exhibited two-compartment characteristics with phasic half-lives of 25 and 174 min., respectively. Myocardial drug accumulation was increased by 600 times at steady-state compared to the perfusion liquid. Dynamic effect parameters were studied during increasing amlodipine concentrations from 0.16 to 220 nM. Dynamic steady-states developed within 20 min. Coronary flow-rate increased with an Emax of 119% and an EC50 of 1.2 x 10(-8) M. Amlodipine produced inhibitory effects on contraction amplitude and velocities of contraction and relaxation. Observed Emax-values and curve-fitted EC50-values were: 97, 97 and 94% and 1.10(-8), 7.7 x 10(-9) and 2.1 x 10(-8) M, respectively. Heart frequency was not changed. Oxygen consumption increased markedly to a maximum of 44% at 3 x 10(-8) M amlodipine and then decreased to nearly initial values. The frequency-corrected QT-interval decreased to a maximal extent of 20% at the three highest concentrations. Myocardial efficiency expressed as the ratio of contraction velocity times frequency to oxygen consumption exhibited a progressive decline to about 2% of initial values. The PQ-interval was not changed and the QRS-interval showed only a small but significant decrease at the highest amlodipine concentration. No arrythmogenic effects were observed. The study demonstrated a very slow accumulation and disposition of amlodipine in the guinea-pig heart with a steady-state myocardial drug concentrating accumulation of 600 times. Marked increase in coronary flow-rate and oxygen consumption accompanied by a progressive negative inotropic effect were observed.     

The effects of some cholinesterase reactivators on contractility of the isolated guinea-pig heart atria

The effect of eight monoquaternary and bisquaternary pyridine aldoxime cholinesterase reactivators was tested on isolated guinea-pig heart atria. 2. Acetylcholine and methylfurthretonium in concentrations ranging from 10(-7) M to 10(-5) M have negative inotropic effects in the electrically stimulated atria and negative chronotropic effects in the spontaneously beating atria. 3. In the presence of higher concentration of cholinesterase reactivators alone, the parameters of heart muscle contractility are significantly altered. 4. Cumulative dose-response curves of methylfurthretonium in the presence of reactivators in the range of concentrations from 10(-5) M to 10(-3) M are shifted parallelly to higher concentrations of the agonist.     

Effects of a lipoxygenase inhibitor on digoxin-induced cardiac arrhythmias in the isolated perfused guinea-pig heart

1. The effects of a lipoxygenase inhibitor, BW A4C, on digoxin-induced arrhythmias and cardiac dynamics (contractile force, perfusion pressure, heart rate) were investigated in Langendorff-perfused isolated guinea-pig hearts. In the control group, arrhythmias were induced by 25 micrograms/ml digoxin at a perfusion rate of 0.5 ml/min. In the treated groups, BW A4C (1 and 0.3 microM) perfused continuously from 15 min prior to digoxin until cardiac arrest occurred. Digoxin exposure (microgram/g wet weight of heart) for the occurrence of arrhythmias and cardiac arrest were the parameters evaluated to assess cardiotoxicity. 2. Digoxin caused a marked increase in leukotriene B4 release in the coronary effluent, and was collected during tachyarrhythmias. BW A4C markedly inhibited the digoxin-induced elevation of LTB4. 3. BW A4C (1 and 0.3 microM) did not prevent the onset of ventricular fibrillation and ventricular tachycardia despite a slight delay in the occurrence of ventricular fibrillation and cardiac arrest at the 0.3 microM concentration. 4. Contractile force increased significantly after digoxin infusion which was concomitant with the time of onset of arrhythmias. In the presence of BW A4C, the contractile force increased, but not significantly. Perfusion pressure increased initially after digoxin infusion in the absence and the presence of BW A4C, but not significantly. 5. These findings show that the lipoxygenase inhibitor lacked any protective action on digoxin-induced arrhythmias despite its effective suppression of digoxin-induced elevation of LTB4 in coronary effluent.     

Electromechanical effects of newly synthetized propafenone derivatives on isolated guinea-pig heart muscle preparations

AIMS: To clarify the significance of apoptosis in the progression of uterine cervical neoplasias, including cervical intraepithelial neoplasia (CIN), microinvasive carcinoma (MIC), and invasive squamous cell carcinoma (ISCC) categories, in relation to cell proliferation and human papilloma virus (HPV) infection. METHODS: Forty six cases of CIN I/II, 75 of CIN III, 16 of MIC, and 44 of ISCC were examined using formalin fixed and paraffin wax embedded samples. The TdT mediated dUTP-biotin nick end labelling (TUNEL) method for detection of apoptotic cells was performed along with Ki-67 immunohistochemistry. Presence of HPV-DNA was confirmed by PCR-RFLP assay. RESULTS: Apoptotic labelling indices, calculated after counting positive nuclei among at least 2000 nuclei, showed significant positive correlation with histological malignant grading in CIN and tumour cell invasion into stroma. In contrast, similar Ki-67 labelling index values were found in CIN, MIC, and ISCC. Although HPV-DNA was detected in 35/46 CIN I/II (76.1%), 53/74CIN III (71.6%), 9/16 MIC (56.3%), and 36/44 ISCC (81.8%), there was no apparent relation with the apoptotic labelling indices. CONCLUSIONS: Apoptosis in cervical neoplasias may be closely related to tumour cell differentiation and progression. It also seems unlikely that HPV itself is directly related to pathways regulating apoptosis.     

Bradykinin mediates myocardial ischaemic preconditioning against free radical injury in guinea-pig isolated heart

1. Myocardial ischaemic preconditioning (IP) against free radical injury and its possible mediator(s) was investigated in a Langendorff-perfused guinea-pig heart. 2. 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) was used for triggering free radical injury in cardiac tissue. It reduced left ventricular developed pressure (LVDP), +/- dp/dtmax, heart rate (HR) and coronary flow (CF) and increased thiobarbituric acid-reactive substances (TBARS) in cardiac tissue. 3. Ischaemic preconditioning (5 min global ischaemia and 5 min reperfusion) exerted cardioprotection against DPPH-induced functional impairment, with significant improvement in LVDP, +/- dp/dtmax, HR and CF. The formation of TBARS in cardiac tissue was reduced. Blockade of bradykinin (BK) B2 receptors with icatibant (HOE 140) abolished the cardio-protective effects of IP. 4. Bradykinin (10(-7) mol/L) perfusion for 10 min protected the heart against free radical injury. The cardioprotection induced by BK was reversed by HOE 140. 5. Pretreatment with IP and BK results in cardiac protection against free radical injury through the activation of B2 receptors. Endogenously generated BK may mediate IP in the guinea-pig heart.     

Uptake and metabolism of tryptophan by the isolated perfused guinea-pig mammary gland

Tryptophan uptake by the isolated perfused lactating guinea-pig mammary gland was 46.5+/-4.6 mug/h per g. Results of absorption studies and the use of [methylene-14C]tryptophan suggest that tryptophan is one of the group of amino acids that are transferred almost quantitatively from blood plasma to milk protein.     

Muscarinic contraction in isolated guinea-pig trachea and antagonism by noradrenaline

In contrast to other muscarinic agonists, WAL 2014 FU does not induce bronchospasm in laboratory animals. The present investigation was intended to test the hypothesis that this is due to a particular susceptibility of the drug's effect to antagonism by catecholamines, as a result of partial M3-agonism. The tonic activity of the muscarinic agonists, aceclidine, arecoline, carbachol, McN-A-343, RS 86, thiopilocarpine and WAL 2014 FU, was tested in groups of isolated tracheal muscle of the guinea-pig. Susceptibility to functional antagonism by beta-adrenoceptor stimulation was measured by the displacement of the concentration-force curves by 3 microM noradrenaline. Evaluation of the concentration-force relationship revealed differences in potency and intrinsic activity (carbachol-100%) ranging from 114% for arecoline to 36% for thiopilocarpine (WAL 2014 FU-63%). The catecholamine increased the concentration of agonist which induced 5% of the maximum effect achievable (EC05) values fivefold (carbachol) to more than 4,680 fold (thiopilocarpine) (WAL 2014 FU: 2,860 fold). Regression analysis between the intrinsic activity of the seven compounds and the antagonistic effect of noradrenaline revealed a significant correlation (Spearman correlation coefficient (r[s])=-0.79; p=0.036). Inhibition of the effects of endogenous catecholamines by beta-adrenolysis with 50 microM toliprolol increased the maximal contraction induced by 1 mM WAL 2014 FU, but did not affect maximal contraction induced by 30 microM arecoline. Pretreatment with 0.3-1.0 mM dibutyrylcyclic adenosine monophosphate (DBcAMP) shifted the concentration-response curves of arecoline, WAL 2014 FU and thiopilocarpine in a similar manner to noradrenaline. The results exclude an important contribution of adenylate cyclase-coupled M2-receptors to the susceptibility of tracheal contraction by muscarinic agonists to functional antagonism by noradrenaline, but emphasize the importance of intrinsic activity at the M3-receptors. The pronounced susceptibility of WAL 2014 FU-induced contraction to functional antagonism by beta-adrenoceptor activation provides an explanation for the failure of the drug to induce bronchospasm in vivo.     

Excitatory motor and electrical effects produced by tachykinins in the human and guinea-pig isolated ureter and guinea-pig renal pelvis

1. In isolated tissue experiments, neurokinin A (NKA) produced concentration-dependent contraction of human and guinea-pig ureter (pD2 = 6.7 and 7.2, respectively); an effect greatly reduced (>80% inhibition) by the tachykinin NK2 receptor-selective antagonist MEN 11420 (0.1 microM). The tachykinin NK1 and NK3 receptor agonists septide and senktide, respectively, were ineffective. 2. Electrical field stimulation (EFS) of the guinea-pig isolated renal pelvis produced an inotropic response blocked by MEN 11420 (0.01-1 microM). In the same preparation MEN 11420 (0.1 microM) blocked (apparent pK(B) = 8.2) the potentiation of spontaneous motor activity produced by the NK2 receptor-selective agonist [betaAla8]NKA(4-10). 3. In sucrose-gap experiments, EFS evoked action potentials (APs) accompanied by phasic contractions of human and guinea-pig ureter, which were unaffected by tetrodotoxin or MEN 11420 (3 microM), but were blocked by nifedipine (1-10 microM). NKA (1-3 microM) produced a slow membrane depolarization with superimposed APs and a tonic contraction with superimposed phasic contractions. NKA prolonged the duration of EFS-evoked APs and potentiated the accompanying contractions. MEN 11420 completely prevented the responses to NKA in both the human and guinea-pig ureter. 4. Nifedipine (1-10 microM) suppressed the NKA-evoked APs and phasic contractions in both human and guinea-pig ureter, and slightly reduced the membrane depolarization induced by NKA. A tonic-type contraction of the human ureter in response to NKA persisted in the presence of nifedipine. 5. In conclusion, tachykinins produce smooth muscle excitation in both human and guinea-pig ureter by stimulating receptors of the NK2 type only. NK2 receptor activation depolarizes the membrane to trigger the firing of APs from latent pacemakers.     

The release of a coronary vasodilator metabolite from the guinea-pig isolated perfused heart stimulated by catecholamines, histamine and electrical pacing and by exposure to anoxia

The results of surveys and inquiries to identify autistic children, carried out in England and Wales, the U.S.A. and Denmark, are compared. Three studies, in each of which either a total population of children or a wide range of handicapped children was screened, using case-note inspection and interviews, all estimated the prevalence of the autistic syndrome to be between four and five children per 10,000 aged under 15 years. Inquiries that counted diagnosed cases only or that relied upon local authority records produced much lower prevalence rates for the autistic syndrome. The reasons for this are examined, and the implications for prevalence studies of handicapping conditions are discussed.     

L-arginine uptake and metabolism by lung macrophages and neutrophils following intratracheal instillation of silica in vivo

Cold air inhalation and exercise-induced bronchoconstriction (EIB) have both been used as measures of bronchial responsiveness. Both stimuli are often combined in the Nordic climate. The main objective of the present study was to investigate the climatic influence of cold temperatures upon exercise-induced asthma. The secondary aims were: (a) to assess metacholine bronchial hyper-responsiveness and EIB in children with bronchial asthma (n = 32; mean age 10.8 years) compared to children with other chronic lung diseases (CLD) (n = 26, mean age 10.1 years); and (b) to assess the influence of cold air inhalation upon EIB in the two groups of children. Methods used were: (a) the metacholine concentration causing a reduction in FEV1 of 20% (PC20-M), (b) maximum FEV1 fall (delta FEV1) after submaximal treadmill run (EIB test); and (c) delta FEV1 after submaximal treadmill run while inhaling cold (-20 degrees C) dry air (CA-EIB test). Geometric mean PC20-M did not differ significantly between the asthma children (1.28 mg ml-1) and the CLD children (2.90 mg ml-1). In the asthma children, mean delta FEV1 after EIB test was 12.8% vs 21.8% after adding cold air (P < 0.0001), compared to 5.2 and 7.4%, respectively (P = 0.03), in the CLD group. Maximum sensitivity and specificity for the EIB test were 69.8% at a fall in FEV1 of 6.8%; for the CA-EIB test, 72% at a fall in FEV1 of 10.2%; and for metacholine provocation, 56% at a PC20-M of 1.5 mg ml-1. In conclusion, children with bronchial asthma are substantially more sensitive to cold air than children with CLD, and EIB is markedly increased by cold air inhalation in asthmatic children, maintaining the specificity of the EIB test and increasing the sensitivity. The low sensitivity of the EIB test is probably influenced by the use of inhaled steroids. Metacholine inhalation test has less specificity and sensitivity in discriminating asthma from other chronic lung diseases.     

Characterization of the calcium-activated chloride channel in isolated guinea-pig hepatocytes

Macroscopic and unitary currents through Ca(2+)-activated Cl- channels were examined in enzymatically isolated guinea-pig hepatocytes using whole-cell, excised outside-out and inside-out configurations of the patch-clamp technique. When K+ conductances were blocked and the intracellular Ca2+ concentration ([Ca2+]i) was set at 1 microM (pCa = 6), membrane currents were observed under whole-cell voltage-clamp conditions. The reversal potential of the current shifted by approximately 60 mV per 10-fold change in the external Cl- concentration. In addition, the current did not appear when Cl- was omitted from the internal and external solutions, indicating that the current was Cl- selective. The current was activated by increasing [Ca2+]i and was inactivated in Ca(2+)-free, 5 mM EGTA internal solution (pCa > 9). The current was inhibited by bath application of 9-anthracenecarboxylic acid (9-AC) and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) in a voltage-dependent manner. In single channel recordings from outside-out patches, unitary current activity was observed, whose averaged slope conductance was 7.4 +/- 0.5 pS (n = 18). The single channel activity responded to extracellular Cl- changes as expected for a Cl- channel current. The open time distribution was best described by a single exponential function with mean open lifetime of 97.6 +/- 10.4 ms (n = 11), while at least two exponentials were required to fit the closed time distributions with a time constant for the fast component of 21.5 +/- 2.8 ms (n = 11) and that for the slow component of 411.9 +/- 52.0 ms (n = 11). In excised inside-out patch recordings, channel open probability was sensitive to [Ca2+]i. The relationship between [Ca2+]i and channel activity was fitted by the Hill equation with a Hill coefficient of 3.4 and the half-maximal activation was 0.48 microM. These results suggest that guinea-pig hepatocytes possess Ca(2+)-activated Cl- channels.     

The uptake of calcium by isolated chromaffin granules of the adrenal medulla

11 beta-Hydroxysteroid dehydrogenase (11 beta-HSD) catalyses the interconversion of biologically active cortisol to inactive cortisone in man, and corticosterone to 11-dehydrocorticosterone in rodents. As such, this enzyme has been shown to confer aldosterone-selectivity on the mineralocorticoid receptor and to modulate cortisol/corticosterone access to the glucocorticoid receptor (GR). Two kinetically distinct isoforms of this enzyme have been characterized in both rodents and man; a low-affinity NADP(H)-dependent enzyme (11 beta-HSD1) which predominantly acts as an oxoreductase and, more recently, a high-affinity NAD-dependent uni-directional dehydrogenase (11 beta-HSD2). In this study we have analysed the expression of both 11 beta-HSD1 and 11 beta-HSD2 isoforms in rat adrenal cortex and medulla and have investigated their possible roles with respect to glucocorticoid-regulated enzymes mediating catecholamine biosynthesis in adrenal medullary chromaffin cells. Using a rat 11 beta-HSD1 probe and a recently cloned in-house mouse 11 beta-HSD2 cDNA probe, Northern blot analyses revealed expression of mRNA species encoding both 11 beta-HSD1 (1.4 kb) and 11 beta-HSD2 (1.9 kb) in the whole adrenal. Consistent with this, 11 beta-dehydrogenase activity (pmol 11-dehydrocorticosterone formed/mg protein per h, mean +/- S.E.M.) in adrenal homogenates, when incubated with 50 nM corticosterone in the presence of 200 microM NAD, was 97.0 +/- 9.0 and with 500 nM corticosterone in the presence of 200 microM NADP, was 98.0 +/- 1.4. 11-Oxoreductase activity (pmol corticosterone formed/mg protein per h) with 500 nM 11-dehydrocorticosterone in the presence of 200 microM NADPH, was 187.7 +/- 31.2. In situ hybridization studies of rat adrenal cortex and medulla using 35 S-labelled antisense 11 beta-HSD1 cRNA probe revealed specific localization of 11 beta-HSD1 mRNA expression predominantly to cells at the corticomedullary junction, most likely within the inner cortex. In contrast, 11 beta-HSD2 mRNA was more abundant in cortex versus medulla, and was more uniformly distributed over the adrenal gland. Negligible staining was detected using control sense probes. Ingestion of the 11 beta-HSD inhibitor, glycyrrhizic acid (> 100 mg/kg body weight per day for 4 days) resulted in significant inhibition of adrenal NADP-dependent (98.0 +/- 1.4 vs 42.5 +/- 0.4) and NAD-dependent (97.0 +/- 9.0 vs 73.2 +/- 6.7) 11 beta-dehydrogenase activity and 11-oxoreductase activity (187.7 +/- 31.2 vs 67.7 +/- 15.3). However, while levels of 11 beta-HSD1 mRNA were similarly reduced (0.85 +/- 0.07 vs 0.50 +/- 0.05 arbitrary units), those for 11 beta-HSD2 remained unchanged (0.44 +/- 0.03 vs 0.38 +/- 0.01). Levels of mRNA encoding the glucocorticoid-dependent enzyme phenylethanolamine N-methyltransferase which catalyses the conversion of noradrenaline to adrenaline, were also significantly reduced in those rats given glycyrrhizic acid (1.12 +/- 0.04 vs 0.78 +/- 0.04), while those for the glucocorticoid-independent enzyme tyrosine hydroxylase (1.9 kb), which catalyses the conversion of tyrosine to DOPA, were unchanged (0.64 +/- 0.04 vs 0.61 +/- 0.04). In conclusion, the rat adrenal gland expresses both 11 beta-HSD1 and 11 beta-HSD2 isoforms. 11 beta-HSD1 gene expression is localized to the adrenal cortico-medullary junction, where it is ideally placed to regulate the supply of cortex-derived corticosterone to the medullary chromaffin cells. This, together with our in vivo studies, suggests that 11 beta-HSD1 may play an important role with respect to adrenocorticosteroid regulation of adrenaline biosynthesis. The role of 11 beta-HSD2 in the adrenal remains to be elucidated.     

Mechanism of valproic acid uptake by isolated rat brain microvessels

In an effort to characterize putative transport systems of valproic acid (VPA) at the blood-brain barrier, the effects of various substrates and inhibitors of known anion transporters on the equilibrium vessel-to-medium concentration (vessel/medium) ratio of VPA were investigated using isolated rat brain microvessels. The equilibrium vessel/medium ratio of VPA was decreased by the presence of high millimolar concentration of unlabeled VPA, indicating that a saturable transport system was involved in VPA transport from medium to microvessels. Short-chain monocarboxylates such as propionic acid, pyruvic acid, and L-lactic acid did not alter the vessel/medium ratio, whereas medium-chain fatty acids and unsaturated metabolites of VPA significantly inhibited the net transport of VPA. Dicarboxylates, tricarboxylate, and p-aminohippuric acid did not affect VPA accumulation in the brain microvessels. Several anionic drugs including salicylic acid, penicillin G, cefazolin, and probenecid significantly reduced the vessel/medium ratio of VPA. In addition, disulfonate inhibitors of inorganic anion exchangers, SH-group modifying reagent, and metabolic inhibitor showed remarkable inhibitory effects on the net transport of VPA between brain microvessels and medium. These results suggest that VPA may be actively transported through the antiluminal membrane via a carrier-mediated system shared by other anionic drugs.     

Mechanism of uptake of technetium-tetrofosmin. II: Uptake into isolated adult rat heart mitochondria

BACKGROUND: 99mTc-labeled tetrofosmin is a new myocardial imaging agent that gives stable heart uptake. However, little is known about the mechanism of uptake in heart tissue. The aim of this study was to assess the factors responsible for the uptake and retention of 99mTc-labeled tetrofosmin in isolated heart mitochondria. METHODS AND RESULTS: Mitochondria were isolated from adult rat heart tissue with competent metabolic function (i.e., respiratory control ratio of 3 and adenosine diphosphate/oxygen ratio of 2) for succinate oxidation. Intramitochondrial volume measured by the distribution of 3H-water and 14C-sucrose was 1.16 +/- 0.23 microliters/mg protein (mean +/- SD). In the isolated mitochondria, uptake could be demonstrated within 30 seconds of addition of oxidative substrate, but adenosine triphosphate alone did not stimulate marked uptake. Uptake was proportional to the amount of mitochondrial protein over a range of 0.2 to 3 mg protein but independent of Tc-labeled tetrofosmin concentration over a range of 0.4 to 200 pmol/L (0.1 to 50 microCi/ml). The presence of Tc-labeled tetrofosmin had no effect on the oxidative capability of the mitochondria. Use of the mitochondrial uncoupler 2,4-dinitrophenol caused release of 92% of radioactivity. Addition of Ca2+ to the mitochondria to partially depolarize the membrane resulted in partial release of activity. Application of the Nernst equation to the uptake data gave rise to a value of -163 mV for the mitochondrial membrane potential. CONCLUSION: It was concluded that the accumulation of 99mTc-labeled tetrofosmin by the mitochondria is related to their ability to transduce metabolic energy into electronegative membrane potential.     

Flufenamic and tolfenamic acids and lemakalim relax guinea-pig isolated trachea by different mechanisms

The effects of K+ channel inhibitors on the relaxations induced by flufenamic and tolfenamic acids and lemakalim were examined in guinea-pig isolated trachea precontracted with prostaglandin F2alpha (PGF2alpha, 1 microM). Flufenamic and tolfenamic acids (0.1-33 microM) and lemakalim (0.01-33 microM) relaxed guinea-pig trachea in a concentration-dependent manner. Tetraethylammonium (0.5-2 mM), a nonspecific inhibitor of K+ channels, inhibited the relaxations induced by flufenamic and tolfenamic acids without affecting lemakalim-induced relaxation. Charybdotoxin (ChTX, 33-100 nM), an inhibitor of the large Ca2+-activated K+ channels (BK(Ca)), also inhibited the relaxations induced by flufenamic and tolfenamic acids without affecting lemakalim-induced relaxation. Glipizide (3.3-33 microM), an inhibitor of the ATP-sensitive K+ channels (K(ATP)) inhibited lemakalim-induced relaxation without affecting those induced by flufenamic and tolfenamic acids. Our results indicate that the relaxations of guinea-pig isolated trachea by flufenamic and tolfenamic acids are due to activation of BK(Ca). The relaxant mechanism of flufenamic and tolfenamic acids thus differs from that of lemakalim, an activator of K(ATP).     

Effect of bodyweight on the rate of glucose uptake by isolated rat muscles

The uptake of glucose by isolated extensor digitorum longus muscles was measured in rats of 78-350 g bodyweight. The rate of uptake per unit weight of muscle fell as the weight of the animal increased. It is concluded that in metabolic studies with isolated rat skelatal muscles, only muscles from weight-matched rats should be compared.