Inhibitory effects of 4,4′-diisothiocyano stilbene-2,2′-disulfonic acid (DIDS) in the response of isolated hepatocytes to phalloidin

Summary The influence of pretreatment with monooxygenase inducers on total irreversible binding of metabolically activated [³H]-benzo(a)pyrene to cellular DNA and the formation of benzo(a)pyrene metabolite-deoxyribonucleoside adducts after cytochrome P-448 induction was studied in perfused rat lungs. Pretreatment with the cytochrome P-448 inducer -naphthoflavone increasing binding by a factor of 23. In lungs of induced animals, 0.45 pmoles of benzo(a)pyrene equivalents were bound per mg DNA. Binding to RNA and to protein was also considerably induced by -naphthoflavone. Phenobarbital treatment did not significantly increase binding to cellular macromolecules of rat lung. Analysis of hydrolyzed DNA of lungs from -naphthoflavone-treated rats by Sephadex LH 20 chromatography revealed the formation of at least two nucleoside adducts with metabolically activated benzo(a)pyrene one of which is probably due to modification of the DNA with a benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide and the other to modification of DNA with secondary metabolites of benzo(a)pyrene phenols.This study was supported by the Deutsche Forschungsgemeinschaft, Bonn-Bad Godesberg.In part subject of the doctoral thesis of Erik Klaus, Fachbereich Biologie, University of Mainz.     

The mechanism of the drinking response to some hypotensive drugs: Activation of the renin-angiotensin system by direct or reflex-mediated stimulation of β-receptors

Summary The increase in water intake and plasma renin concentration (PRC) in response to the administration of the -receptor blocker phentolamine (10 mg/kg) were simultaneously diminished or abolished by ganglionic blockade with pempidine (10 mg/kg) or camphidonium (10 mg/kg) as well as by pretreatment with reserpine or 6-OH-dopamine. In contrast, the effects of the -mimetic agent, isoprenaline (250 g/kg), on both parameter remained unchanged. The (–)-isomer of propranolol (200 g/kg) selectively blocked the dipsogenic and PRC-raising effects of isoprenaline and phentolamine, thus demonstrating that a specific -receptor blockade antagonizes the drinking and increase in PRC induced by the drugs used.From these results it is concluded that the stimulation fo certain -receptors causes an increase in PRC. The -receptors may be stimulated either directly by -mimetic agents such as isoprenaline or indirectly by a reflex-mediated activation of the sympathetic nervous system brought about by hypotensive, drugs such as phentolamine.The activation of the renin-angiotensin system triggers off the drinking response.Some of the results were presented to the Deutsche Pharmakologische Gesellschaft (Meyer and Hertting, 1971).     

Serum glycoside concentrations after single or repeated intravenous doses ofβ-methyl-digoxin and digoxin

Summary The aim of the present investigation was to estimate the ratio of the intravenous doses of-methyl-digoxin and digoxin required to produce identical serum glycoside concentrations in man.20 patients on intravenous maintenance therapy were changed from-methyl-digoxin to the identical dose of digoxin or vice versa. Each drug was given for 7 days. Serum concentrations 13% higher were found during administration of-methyl-digoxin. Assuming a half life of 60 h after with drawal, the dose of digoxin producing the same minimum serum concentration was estimated to be 1.16 times higher than that of-methyl-digoxin.18 healthy volunteers received 0.4 mg -methyldigoxin, and 23 the same dose of digoxin, as an intravenous infusion over 2 h. The serum concentrations and urinary glycoside excretion were measured over a period of 32 hrs. During the first hour after the infusion the serum concentration of digoxin declined more rapidly than that of-methyl-digoxin. Thereafter, the ratio of the serum concentrations did not change appreciably up to the end of the investigation. The area under the serum concentration/time curve was about 13% greater for-methyl-digoxin than for digoxin; this difference was not significant.The average renal clearance was 96±9 ml for-methyl-digoxin, 151±13 ml for digoxin. Since the total body clearance of digoxin is only about 1.16 times higher than that of-methyl-digoxin, the lower renal clearance of-methyl-digoxin must partly be compensated by higher extrarenal clearance.From the ratios of the areas under the serum concentration/time curves after single doses of -methyldigoxin and digoxin, and the minimum serum concentrations during maintenance therapy, it was concluded that the dose of digoxin to produce the same average serum concentrations would be about 1.15 times higher than that of-methyl-digoxin. In comparison with the large variations in individual dosage of digoxin and-methyl-digoxin, this difference is too small to be of practical importance.     

α-adrenoceptors mediating the positive inotropic effect of phenylephrine in the right ventricular muscle of the monkey (Macaca fuscata)

Summary The property of adrenoceptors mediating the positive inotropic effect (PIE) in the ventricular muscle of the Japanese monkey (Macaca fuscata) was investigated by the use of phenylephrine (PE) and adrenoceptor antagonists. The intrinsic activity (0.6) and the pD₂-value (5.41) for PE were comparable to those in other mammalian species. The -adrenoceptor antagonist, pindolol (3×10^–8 mol/l) shifted only the upper part of the concentration-response curve (CRC) for PE to the right; the pD₂-value for PE was not significantly affected by pindolol. On the other hand, phentolamine (10^–6 mol/l) shifted the lower part of the CRC for PE more than the upper part. In the presence of both pindolol and phentolamine the curve was shifted to the right in a parallel manner. The time required for twitch relaxation was negatively correlated to the degree of PIE of PE in the presence of phentolamine but not pindolol. These results indicate that both - and -adrenoceptors mediate the positive inotropic action in the ventricular muscle of the Japanese monkey and that in contrast to the action via -adrenoceptors the action via -adrenoceptors is not accompanied by the relaxant effect.     

A dose-ranging study to evaluate the β1-adrenoceptor selectivity of bisoprolol

Summary A dose-ranging study was performed to compare the ₁-adrenoceptor selectivity of bisoprolol with that of atenolol and nadolol. Seven normal subjects (mean age 26 y) were given single oral doses of bisoprolol 5 mg (B5), 10 mg (B10), 20 mg (B20); atenolol 50 mg (A50), 100 mg (A100); nadolol 40 mg (N40); and placebo (PL), in a single blind randomised cross-over design. ₂-adrenoceptor responses were assessed by attenuation of finger tremor and cardiovascular responses to graded isoprenaline infusions. Dose-response curves were constructed, and doses of isoprenaline required to increase finger tremor by 100% (IT₁₀₀), heart rate by 25 beats/min (IH₂₅), SBP by 25 mm Hg (IS₂₅), cardiac output by 35% (IC₃₅), and decrease DBP by 10 mm Hg (ID₁₀), after each treatment were calculated. These indices were compared with placebo response and expressed as dose-ratios. Exercise heart rate (EHR) was used to assess ₁-adrenoceptor blockade.There were dose-related increases in plasma concentrations of bisoprolol and atenolol. Reduction of EHR was significantly less with B5 (16.8%) in comparison with all other treatments: B10 21.9%, B20 23.1%; A50 22.5%, A100 22.6%; N40 22.9%. There were small but significant reductions in isoprenaline-induced tachycardia with bisoprolol and atenolol, although mean dose-ratios were considerably less in comparison with N40 (IH₂₅ dose-ratios): B5 2.55, B10 3.18, B20 3.93, A50 2.91, A100 4.89, N40 17.23. There were similar patterns for the other isoprenaline responses.These results show that conventional doses of bisoprolol (10 mg) and atenolol (50 mg) produced equal antagonism of ₁ and ₂-adrenoceptors, and therefore possess equal degrees of ₁-adrenoceptor selectivity. Increasing doses of bisoprolol and atenolol were associated with partial loss of selective ₁-adrenoceptor blockade, although antagonism of ₂-adrenoceptors was significantly less compared with the effects of nadolol.     

Enzymic control of irreversible binding of metabolically activated benzo(a)pyrene in perfused rat liver by monooxygenase activity

Addition of [³H]-benzo(a)pyrene to the perfusion medium of isolated rat livers results in irreversible binding of radioactivity to DNA, RNA and protein. Binding to DNA accounted for about 0.1% of the total radioactivity which was bound in livers from animals treated with oil or saline and was increased by a factor of 3–5 after pretreatment of the animals with -naphthoflavone or with phenobarbital. When the inhibitiors of monooygenase activity, -naphthoflavone or metyrapone, were present in the perfusion medium, irreversible binding was reduced in livers from both -naphthoflavone- and phenobarbital-pretreated animals, irrespective of the inhibitor used.In livers from animals treated with oil or saline protein and a RNA fraction containing tightly associated protein were able to bind [³H]-benzo(a)pyrene metabolites to about the same extent but after induction by pretreatment with -naphthoflavone binding to the RNA fraction was enhanced to a much higher extent than binding to the protein fraction. Pretreatment with phenobarbital did not result in an increased irreversible binding to RNA and protein.A considerable amount of 15–25% of the total radioactivity added to the perfusion medium was excreted into the bile after treatment of the animals with the tested inducers of monooxygenase activity compared to an excretion of 3% in animals treated with oil or saline.The results indicate that nucleic acid and protein adduct formation in the liver is controlled by the action of the cytochrome P-450-dependent monooxygenases.In part subject of the doctoral thesis of Erik Klaus, Fachbereich Biologie, University of Mainz     

Plasma nicotine,plasma β-endorphin and mood states during periods of chronic smoking,abstinence and nicotine replacement

Nicotine is known to release neuroendocrine substances which may subsequently reinforce smoking behavior by improving mood states. The purpose of this study was to examine changes in plasma-endorphin and mood states during periods of chronic smoking, abstinence from smoking, and abstinence while chewing nicotine gum. A modified A-B-A-C design was used. Normal male volunteers were randomly assigned to an experimental or control group. Over a 12-day protocol, experimental subjects smoked ad libitum for 2 days, were abstinent for 4 days, resumed smoking for 2 days, and then chewed nicotine gum for the final 4 days. Control subjects smoked ad libitum throughout the entire protocol. Results indicated that changes in plasma-endorphin levels were not related to changes in the four smoking conditions. Plasma nicotine and mood states were related, such that dysphoric moods increased during abstinence from smoking in comparison to the control group. To investigate further the relationships between nicotine,-endorphin and reinforcement for smoking, it may be necessary to characterize endogenous opioid peptide release in the central nervous system during smoking.     

Nasal Absorption Enhancement of 17β-Estradiol by Dimethyl-β-Cyclodextrin in Rabbits and Rats

A new formulation for nasal administration containing 17-estradiol (E₂) with dimethyl--cyclodextrin (DMC) as a solubilizer and absorption enhancer is described. Nasal administration of this E₂-DMC formulation gave a significantly higher E₂ absorption than an E₂ suspension in both rabbits and rats. Relative to an intravenous injection of the E₂-DMC formulation, absolute bioavailabilities of 94.6 and 67.2% were calculated for the nasal E₂-DMC formulation in rabbits and rats, respectively. Differences in bioavailability may have resulted from differences in experimental animal conditions. The effects on human nasal ciliary activity of the E₂-DMC formulation were studied with an in vitro method. The formulation was found to exert only a minor effect on ciliary beat frequency. Thus, nasal delivery of E₂, using a cyclodextrin inclusion formulation, may have potential for clinical application, e.g., in the therapy of postmenopausal disorders.     

Urinary trehalase activity as an indicator of kidney injury due to environmental cadmium exposure

One hundred and seventy-eight subjects, patients with Itai-itai disease and their family members, aged 12–87 years living in a cadmium (Cd)-polluted area in the Jinzu River basin (Cd-exposed group) and 176 controls (control group) were examined. In the Cd-exposed group urinary trehalase increased with increasing age, urinary 2-microglobulin (2-m) and retinol-binding protein. Although urinary cadmium was higher in the Cd-exposed group, no particular correlation was found between urinary trehalase and urinary cadmium. Seventeen men and 11 women showed raised urinary trehalase activities despite normal values of urinary 2-m (<300 g/g.creatinine), suggesting that urinary trehalase increases earlier than urinary 2-m. In 19 patients with Itai-itai disease included in the Cd-exposed group, urinary trehalase decreased with decreasing reciprocal of serum creatinine, suggesting that urinary trehalase decreases in the most advanced cases of chronic cadmium nephropathy due to reduced tubular cell mass.     

Naloxone effects on β-endorphin,cortisol, prolactin,growth hormone,HVA and MHPG in plasma of normal volunteers

8 mg of naloxone were administered IV to 14 normal volunteers in a placebo-controlled, double-blind experiment. Plasma levels of -endorphin, cortisol, prolactin, growth hormone, HVA and MHPG were determined before and 45 min after administration. Naloxone elicited significant increases in cortisol and MHPG but did not change plasma levels of the other compounds. In an additional experiment on two subjects, 20 mg of naloxone caused elevations of -endorphin as well as of cortisol. This parallel increase indicates that the linkage between the secretion of -endorphin and ACTH/cortisol may be dose-dependent. The increase in MHPG is in agreement with the hypothesized association of noradrenergic hyperactivity and opiate withdrawal.     

Protection Afforded by Maltosyl-β-cyclodextrin Against α-Chymotrypsin-Catalyzed Hydrolysis of a Luteinizing Hormone-Releasing Hormone Agonist, Buserelin Acetate

Purpose. The present study addresses how maltosyl--cyclodextrin (G₂--CyD) impacts upon the -chymotrypsin-catalyzed hydrolysis of buserelin acetate, an agonist of luteinizing hormone-releasing hormone with emphasis upon the direct effect of G₂--CyD on the activity of the protease. Methods. Kinetic and solubility studies were performed in isotonic phosphate buffer (pH 7.4) at 25°C and 37°C. The interaction of -chymotrypsin with G₂--CyD in the buffer solution was examined by differential scanning calorimetry. Results. G₂--CyD decelerated the -chymotrypsin-catalyzed hydrolysis of buserelin acetate to give the 1–3 tripeptide and the 4–9 hexapeptide fragments. This deceleration can be explained solely by a nonproductive encounter between a complex of the substrate with G₂--CyD and the protease at relatively low CyD concentrations, while the direct inhibitory effect of G₂--CyD on the proteolytic activity made a considerable contribution to the overall deceleration of the hydrolysis at higher CyD concentrations. Calorimetric studies indicate the presence of intermediate states in the thermal unfolding of -chymotrypsin, simultaneously accompanied by the autolysis. By contrast, a two-state thermal unfolding of -chymotrypsin was observed in the presence of G₂--CyD, suggesting reduced proteolytic activity upon binding to G₂--CyD. Conclusions. These results suggest that G₂--CyD at higher concentrations inhibits the proteolytic action of -chymotrypsin through direct interaction with the protease, as well as through the formation of a non-productive complex with the substrate.     

Unlabeled hemoglobin adducts of 4,4′-methylenebis (2-chloroaniline) in rats and guinea pigs

The capacity of N-oxidized metabolites of 4,4-methylenebis(2-chloroaniline) (MBOCA) to form hemoglobin (Hb) adducts was determined in vitro, and the formation of Hb adducts following in vivo administration of MBOCA was assessed with or without prior induction of cytochrome P-450 enzymes with phenobarbital or -naphthoflavone. Hb adduct formation was determined by electron-capture GLC of MBOCA as the heptafluorobutyryl derivative following mild acid hydrolysis of protein-bound MBOCA. The method was confirmed by gas chromatography-mass spectrometry with selected ion monitoring. N-hydroxy- and mononitroso-MBOCA, but not MBOCA itself, formed adducts to rat and human Hb in vitro in a dose-related manner. Binding was inhibited by cysteine and glutathione but not oxidized glutathione or methionine. Intravenous administration of as little as 0.04 mol/kg N-hydroxy-MBOCA to rats resulted in measurable formation of MBOCA-Hb adducts (0.9 ng/50 mg Hb). Intraperitoneal administration of 0.5–50 mg/kg MBOCA to rats, and subcutaneous administration of 5–500 mg/kg MBOCA to rats and 4–100 mg/kg to guinea pigs resulted in dose-related formation of Hb adducts. MBOCA-Hb remained elevated in blood for greater than 10 weeks following a single subcutaneous dose in guinea pigs. Pretreatment of rats with phenobarbital induced microsomal benzphetamine N-demethylase (BND) activity and resulted in a small increase in in vitro N- andortho- hydroxylation of MBOCA, but did not increase in vivo Hb adducts. Pretreatment of rats with -naphthoflavone induced microsomal aryl hydrocarbon hydroxylase as well as ethoxyresorufin-O-deethylase, and increased in vitro N- but notortho-hydroxylation of MBOCA. -Naphthoflavone pretreatment increased the formation of MBOCA-Hb adducts when rats were dosed with MBOCA at 100 and 500 mg/kg, but not 20 mg/kg subcutaneously.     

Enzyme-inducing drug combinations and their effects on liver microsomal enzyme activity in man

Summary The effect of 2 different drug combinations on liver microsomal activity was investigated in healthy volunteers by administering antipyrine 1200 mg and phenobarbitone 100 mg, or the same dose of antipyrine with rifampicin 600 mg daily for 14 days. The effect of rifampicin 1200 mg given for only 8 days was also studied. Before and after each drug regimen, estimates were made of the total body clearance of antipyrine, -glutamyl-transferase (-GT) and urinary excretion of 6--hydroxycortisol as in vivo parameters of liver microsomal enzyme activity. Following combined antipyrine and phenobarbitone administration, the antipyrine clearance was increased by 80%, after antipyrine with rifampicin by 128%, and after rifampicin alone by 104%. 6--hydroxycortisol, corrected for 17-hydroxycorticosteroids, increased from 2.6% to 8% following antipyrine plus phenobarbitone, from 4.4% to 27.9% following antipyrine plus rifampicin, and from 5.4% to 29.7% after rifampicin given alone. Based on previous studies, antipyrine given with phenobarbitone produced slightly more induction than phenobarbitone given alone. Following antipyrine 1200 mg with rifampicin 600 mg for 14 days a significantly greater increase in antipyrine clearance and 6--hydroxycortisol excretion was observed than when either drug was given alone.     

Both β1- and β2-adrenoceptors mediate catecholamine-evoked arrhythmias in isolated human right atrium

Summary The involvement of ₁- and ₂-adrenoceptors in catecholamine-evoked arrhythmias was investigated in isolated human right atrial appendages obtained from 22 patients chronically treated with blockers (usually ₁-selective) and 9 patients not treated with blockers. A simple experimental model that assesses the incidence of arrhythmic contractions as a function of heart rate (pacing) is introduced. ₁-adrenoceptors were activated by (–)-noradrenaline during ₂-adrenoceptor blockade with 50 nmol/l ICI 118551. ₂-adrenoceptors were activated by (–)-adrenaline during ₁-adrenoceptor blockade with 300 nmol/l CGP 20712A. Both (–)noradrenaline and (–)-adrenaline caused arrhythmic contractions whose incidence was greater at low than at high pacing rates. CGP 20712A (300 nmol/l) blocked the (–)-noradrenaline-evoked contractions in 1/1 atrial strip from 1/1 patient not treated with a blocker and 17/17 atrial strips from 15/15 patients chronically treated with blockers. ICI 118551 (50 nmol/l) blocked the (–)-adrenaline-evoked contractions in 3/4 atrial strips from 3/4 patients not treated with blockers and 17/20 atrial strips from 15/18 patients chronically treated with blockers. The incidence of arrhythmic contractions evoked by both (–)-noradrenaline and (–)-adrenaline was higher in chronically blocked patients than in non blocked patients. We conclude that both ₁- and ₂-adrenoceptors mediate atrial arrhythmias and that the generation of these arrhythmias is facilitated by chronic ₁-adrenoceptor blockade. Correspondence to: A. J. Kaumann at the Clinical Pharmacology Unit, University of Cambridge, as above     

In kitten ventricular myocardium,the inotropic potency of an agonist is determined by both its intrinsic activity for the adenylyl cyclase and its affinity for the β-adrenoceptors

Summary Three -adrenoceptor-related parameters were measured for 23 agonists on kitten ventricular myocardium: inotropic potencies (EC₅₀'s) on papillary muscles, intrinsic activities for stimulation of the adenylyl cyclase in membrane particles, and affinities for -adrenoceptors labelled with ³H-(–)-propranolol in membrane particles.For catecholamines a dissociation between inotropic potencies and -adrenoceptor affinities was observed. Inotropic potencies are 1.3–1.8 orders of magnitude higher than affinities. This dissociation between inotropic activation and binding is similar for (+)- and (–)-enantiomers of catecholamines, despite the higher affinities of the latter. The dissociation is considerably smaller for dopamine and dobutamine, which suggests a role of the -OH group in the phenomenon. The effect is also smaller with resorcinols than with catechols; it decreases further, or is not observed at all, when one of the ring hydroxyl groups is replaced or absent. With most agonists the degree of dissociation appears to be directly proportional to the degree of adenylyl cyclase stimulation. About 1/4 of maximum adenylyl cyclase stimulation suffices to activate the inotropic processes fully. It is suggested that both the magnitude of the dissociation and the degree of adenylyl cyclase activation are indicators of the degree of change induced by agonist in the conformation of the -adrenoceptor.The production of marked inotropic effects by noradrenaline and adrenaline with low -adrenoceptor occupancy and little adenylyl cyclase stimulation may permit the regulation of heart function. Conditions of desensitization that reduce maximum activation of the adenylyl cyclase may not necessarily reduce maximum inotropic effects of these catecholamines.The agonist (±)-hydroxybenzylpindolol caused positive inotropic effects only at high -adrenoceptor occupancy. This is consistent with the idea that sufficient conformational change occurs only near receptor saturation.     

Haemophilus influenzae-induced decreases in lung β-adrenoceptor function and number coincide with decreases in spleen noradrenaline

Summary In the guinea-pig intraperitoneal administration of the Gram-negative bacterium Haemophilus infuenzae induces a decrease of -adrenoceptor number and results in impairment of -adrenoceptor function in the peripheral and central airways, respectively. In the present study, the time-course of these events was studied and compared with changes in catecholamines in plasma, in organs involved in immunoregulation (spleen, thymus), and in the heart and the lung. The number of -adrenoceptor binding sites in peripheral lung tissue and -adrenoceptor function in isolated tracheal spirals were significantly decreased 3 and 4 days after administration of H. infuenzae (24–33%). No significant changes were observed at day 1 and day 8. The effects on tracheal -adrenergic receptor function were characterized by a decrease of maximal relaxation only, whereas EC₅₀-values were not affected. These data are indicative of an effect on the functional coupling of the receptors to the biochemical events leading to smooth muscle relaxation. No changes were observed in catecholamine concentrations in the lung, heart, and the thymus after H. infuenzae-treatment. Plasma noradrenaline, though, was significantly increased at day 1 after H. infuenzae. At day 8 plasma noradrenaline had returned to control levels. Interestingly, the effect on spleen noradrenaline was opposite to the effect seen in plasma. A significant decrease in spleen noradrenaline was observed after H. infuenzae at days 1, 3, and 8, with a maximum of 42% at day 1. It is suggested that the decrease in spleen noradrenaline may have a causal relationship with the changes in lung -adrenoceptors. Send offprint requests to F. Engels     

Regional distribution of β1- and β2-adrenoceptors in the failing and nonfailing human heart

Summary Total -adrenoceptor density and ₁- and ₂-subtype distribution in right and left atria and in different ventricular regions from 14 failing and seven nonfailing human hearts have been compared. End-stage heart failure was due to idiopathic dilated cardiomyopathy (n=8) or ischaemic cardiomyopathy (n=6).In nonfailing hearts the total -adrenoceptor density was similar in the right and left atria and in all the ventricular regions studied (about 70 to 80 fmol/mg protein). The ₁:₂-adrenoceptor ratio in both nonfailing atria was similar (about 70:30%) and was significantly smaller than in the different regions of both ventricles (about 80:20%). The ₁-subtype density was similar in nonfailing atria and ventricles (about 55 fmol/mg protein). The ₂-subtype density was significantly higher in the right and left atrium (about 25 fmol/mg protein) than in both ventricles (about 15 fmol/mg protein).In patients with end-stage heart failure due to idiopathic dilated cardiomyopathy or ischaemic cardiomyopathy the total -adrenoceptor density was reduced by 50–60% in all regions. On the other hand, the ₁- and ₂-subtype distribution differed with the cause of heart failure. In patients with idiopathic dilated cardiomyopathy, the ₁-adrenoceptor density was lower in all regions, but the ₂-adrenoceptor density was not significantly reduced. In patients with ischaemic cardiomyopathy both ₁- and ₂-adrenoceptors were reduced in all regions.It is concluded that downregulation of -adrenoceptors in patients with end-stage idiopathic dilated cardiomyopathy or ischaemic cardiomyopathy occurs uniformly throughout the heart. The results support the hypothesis that changes in -adrenoceptor subtypes may be related to the cause of heart failure.     

The influence of hyperthyroidism on β-adrenoceptor-mediated relaxation of isolated small mesenteric arteries

We investigated the influence of hyperthyroidism on relaxant responses of small mesenteric resistance arteries to -adrenoceptor agonists and to compounds stimulating the corresponding second-messenger system. Hyperthyroidism was induced by feeding rats for 28 days with 5 mg/kg L-thyroxine (T4)-containing rat chow. This treatment produced a stable hyperthyroid state, as indicated by several biochemical/metabolic and haemodynamic parameters. Preparations of small mesenteric arteries were mounted in an isometric wire myograph. Subsequently, concentration-effect curves were determined for isoproterenol, noradrenaline and salbutamol as well as for forskolin, dibutyryl-cAMP and theophylline. We also determined concentration-effect curves to the -adrenoceptor agonists in the presence of ICI 118,551 and CGP 20712A (i.e., in the presence of a selective ₂- and ₁-adrenoceptor antagonist, respectively). Apparent pA₂-values were calculated to determine which -adrenoceptor subtype causes vasodilation. These experiments indicate that -adrenoceptor-mediated vasodilation involves both ₁- and ₂-adrenoceptors in mesenteric resistance vessels of both hyperthyroid and control rats. In our experiments hyperthyroidism has a sensitizing influence on vascular responses induced by the -adrenoceptor agonist isoproterenol and the selective ₂-adrenoceptor agonist salbutamol. Sensitization to isoproterenol was abolished in the presence of ICI 118,551, whereas it was emphasized in the presence of CGP 20712A. Although this was not fully supported by the results obtained with noradrenaline, these results indicate that the sensitization to -adrenoceptor agonists is probably limited to the ₂-adrenoceptor/G-protein complex and not associated with alterations of the corresponding second messenger system.     

Effects of polychlorinated terphenyls and paraffins on rat liver microsomal cytochrome P-450 and in vitro metabolic activities

The polychlorinated terphenyl Aroclor 5460 and the polychlorinated paraffins Witaclor 171 P and Witaclor 149 increased to different degrees the total microsomal concentration of cytochrome P-450 in the rat liver after intraperitoneal injection of 0.3, 1.0, and 1.0 g · kg^–1 body weight, respectively, each day for four days. The multiple forms of cytochrome P-450 were affected differently with an induction of RLvMc P-450₅₀ and RLvMc P-450₅₄ by all chemicals, and an additional induction of RLvMc P-450₅₅ by the polychlorinated terphenyl. The rat liver weights were extensively increased after treatment with the polychlorinated paraffins. Alterations were found in the in vitro metabolism of biphenyl, benzo(a)pyrene and the steroid hormones, 4-androstene-3,17-dione and 5-androstane-3,17-diol, after exposure to all chemicals. Changes in the in vitro formation of benzo(a)pyrene metabolites were found to correlate with changes in the multiple forms of cytochrome P-450. The present study demonstrate that only limited information can be obtained from alterations in the total concentration of cytochrome P-450 and show the importance of studying changes in the multiple forms and the metabolism of different substrates. Our results further indicate that exposure to any of the investigated polychlorinated chemicals may alter the biological effects of other environmental contaminants, drugs and endogenous substances which are metabolized by the cytochrome P-450 enzyme system.     

Differential Effects of Sulfate and Sulfobutyl Ether of β-Cyclodextrin on Erythrocyte Membranes in Vitro

The hemolytic activity of -cyclodextrin (-CyD) on rabbit erythrocytes was reduced by the introduction of negatively-charged groups onto the hydroxyls of -CyD; the membrane disrupting abilities decreased in the order of -CyD > 2-hydroxypropyl--CyD (HP--CyD) > sulfobutyl--CyD (SB--CyD) >> -CyD sulfate (S--CyD). Under pre-hemolytic concentrations, both -CyD and SB--CyD induced shape changes of membrane invagination on the erythrocytes. In sharp contrast, S--CyD showed biphasic effect on the shape of the erythrocytes; i.e. the crenation at relatively low concentrations and the invagination at higher concentrations. The S--CyD-induced membrane crenation arose from a direct action on the membranes rather than cell metabolism-mediated effects. Unlike -CyD, S--CyD was found to bind to the erythrocytes and may be confined to the outer surface of the membrane bilayer, which may expand the exterior layer relative to the cytoplasmic half, thereby inducing the cells to crenate. On the other hand, the membrane invagination mediated by the three - CyDs was initiated by extracting specific membrane lipids from the cells, depending upon their inclusion abilities, subsequently leading to the lysis of the cells. These results indicate that SB--CyD and S--CyD interact with the erythrocyte membranes in a differential manner and possess lower membrane disrupting abilities than the parent -CyD and HP--CyD.