Effects of the oestrous cycle and exogenous ovarian steroids on metabolism of beta-phenylethylamine in rat lung.

1 Metabolism of [14C]-beta-phenylethylamine (PEN), a substrate for monoamine oxidase-B (MAO-B), was measured in lung homogenates and in perfused lungs during the 4 day oestrous cycle of the rat. 2 Metabolism in vitro was high during met-oestrus and di-oestrus and low during pro-oestrus and oestrus; this variation in activity correlated with changes in Vmax of the enzyme without changes in Km. 3 PEN metabolism in lung homogenates was also altered by treatment of rats with 17 beta-oestradiol but not by progesterone treatment. 4 Metabolism of [14C]-PEN in perfused lungs was the same during either pro-oestrus or met-oestrus. Uptake of [14C]-PEN in perfused lung measured directly was also the same at these two stages. 5 These results demonstrate that in lungs MAO-B activity was affected by endogenous changes in steroid level but that such changes in enzymic activity were not reflected in the metabolic properties of whole lung.

Ontogenesis of uptake and deamination of 5-hydroxytryptamine, dopamine and beta-phenylethylamine in isolated perfused lung and lung homogenates from rats.

1. Uptake of 5-hydroxytryptamine (5-HT) and beta-phenylethylamine (PEA) was studied in perfused lung from male rats between 10 and 70 days old. 2. Monoamine oxidase (MAO) activity towards 5-HT, PEA and dopamine was studied in homogenate preparations of lung from rats aged between 5 and 80 days. 3. Uptake of 5-HT (10 microM) decreased throughout the age range studied but uptake of PEA (50 microM) increased for the first 30 days and beyond this age it decreased. Metabolites formed for both amines reflected the changes in uptake. 4. MAO activity deaminating 5-HT is well developed by day 10 and reaches its maximum by day 40. For dopamine and PEA, MAO activity remained low until day 20, and the developed rapidly, reaching a maximum by day 40 for dopamine; activity towards PEA did not reach a maximum by day 80. 5. These results show that uptake and MAO activity changes with age and thus the lung responds like other tissues. 6. These results also demonstrate the independent development of uptake and MAO activity towards 5-HT, PEA and dopamine.

Rate-limiting step in the metabolism of polar and non-polar monoamines in lung and liver.

1 Uptake of the non-hydroxylated amines, [14C]-tryptamine and [14C]-benzylamine in rat lung, infused through the pulmonary circulation, was not saturable over the concentration range 2.5-1,000 microM. 2 The kinetic constants for deamination of a variety of hydroxylated and non-hydroxylated monoamines in liver, perfused via the portal circulation, with monoamine oxidase activity in homogenates of liver were similar. 3 In lung, uptake of both [14C]-tryptamine and [14C]-benzylamine was inhibited by the monoamine oxidase inhibitor deprenyl and competition occurred between tryptamine, benzylamine and beta-phenylethylamine for uptake. 4 These results indicate that tryptamine and benzylamine metabolism in lung is not limited by uptake, unlike that of the hydroxylated amines 5-hydroxytryptamine and noradrenaline and that uptake resembles that of beta-phenylethylamine in lung. 5 the selectivity of the lung in handling monoamines is not shown by the liver, suggesting that lung has a specific role in clearing certain biogenic monoamines.

Effect of hyperoxia on uptake and metabolism of 5-hydroxytryptamine and beta-phenylethylamine in rat lung: a sex difference.

1 The uptake of 5-hydroxytryptamine (5-HT) and beta-phenylethylamine (PEA) and their deamination by monoamine oxidase (MAO) were studied in perfused lung from male and female rats exposed to 100% O(2) at 1 ATA for up to 60 h.2 The uptake and metabolism of 5-HT in lungs from both male and female rats was not changed by exposure to O(2).3 The uptake and metabolism of PEA by lungs from male rats was unchanged. Uptake of PEA by lungs from female rats was inhibited 20% and 62% after 37 h and 50 h exposure respectively.4 MAO activity, both in vitro and in perfused lung, was increased towards PEA after 35 h of hyperoxia.5 Metabolism of PEA in perfused lung, measured over 30 min, was inhibited 52% after 50 h of O(2) hyperoxia.6 These results show that exposure to high concentrations of O(2) damages lung, resulting in inhibition of uptake of PEA and consequently in inhibition of metabolism of PEA.7 These results also indicate that, in lung from female rats, MAO-type B is more susceptible to changes in O(2) tension than MAO type A.

Binding of oestradiol, progesterone and prolactin in rat lung.

Binding of [3H]oestradiol and of [3H]progesterone in the cytosol from lungs of adult male rats was suppressible, dependent on incubation time and on protein concentration and was protein in nature. Suppressible binding of oestradiol consisted of a high affinity site (dissociation constant (Kd), 1 x 10(-9) +/- 0.2 x 10(-9) mol/l; maximum number of binding sites (Bmax), 0.7 +/- 0.2 pmol/mg protein) and a lower affinity site (Kd, 2.4 x 10(-8) +/- 0.6 x 10(-8) mol/l; Bmax, 6.3 +/- 0.4 pmol/mg protein) and showed evidence of positive co-operation. Suppressible binding of progesterone consisted of a single site with a Kd of 6 x 10(-9) +/- 1 x 10(-9) mol/l and Bmax of 44.5 +/- 8 fmol/mg protein. Binding of 125I-labelled ovine prolactin was found in homogenates of fetal lung (20 days of gestation) but not of adult lung (80 days of age). Treatment of adult rats with ovine prolactin was associated with an increase in the number of binding sites of high affinity for 125I-labelled ovine prolactin but these sites showed an altered specificity. This 'up-regulation' of the prolactin binding may provide a mechanism by which prolactin stimulates surfactant production in lung. These results, together with the known effects of these hormones on certain lung functions, provide further evidence that lung is a target organ for oestradiol, progesterone and prolactin.

Sensitization does not alter phasic and tonic responses to carbachol in isolated epithelium-free guinea-pig trachea.

An alteration in the handling of Ca2+ has been proposed as the pathogenic mechanism underlying the airway smooth muscle hyperresponsiveness of asthma. The present study tested the hypothesis that the altered responsiveness of receptor operated contraction to carbachol in allergic asthma results from a change in the phasic or tonic components. Using a kinetic approach, the phasic and tonic responses to 10 microM carbachol were quantitated in isolated epithelium-free trachea 21 days after guinea-pigs were sensitized with ovalbumin and aluminum hydroxide (as adjuvant) to generate preferentially IgE-like antibodies. Sensitization was confirmed by challenge of the isolated trachea with ovalbumin. The steady-state and kinetic characteristics of the phasic and tonic responses were the same from sensitized animals and animals treated with saline and aluminum hydroxide (control) and before and after challenge of the trachea from both groups of animals. The present results demonstrate that immunologic sensitization and challenge do not appear to elicit a defect in the phasic or tonic responses of receptor mediated contractions in airway smooth muscle and suggest there is no alteration in the handling of Ca2+ in smooth muscle from sensitized and challenged guinea-pig trachea.

Clinical potency of calcium channel blockers in asthma may be related to their inhibition of receptor-mediated phasic responses in vitro.

The calcium channel blockers (CCB) have been clinically effective in exercise-induced asthma. The completeness of protection with the CCB might be related specifically to inhibition of Ca2+ influx or release. To examine this hypothesis, the rank order of potency of inhibition of the CCB, nicardipine, diltiazem and verapamil on the steady-state and kinetic parameters of the phasic and tonic responses to the muscarinic receptor agonist carbachol (10 microM) and KCl (40 mM) in the intact isolated guinea-pig trachea was determined. The Ca2+ channel agonist Bay K 8644 was also examined for its effects on intracellular Ca2+. Nicardipine abolished the KCl response at both 0.1 microM and 1 microM concentrations. The amplitude of the KCl response was inhibited equally by 1 microM diltiazem (61% inhibition) and 1 microM verapamil (68% inhibition). The rate constant of onset of the KCl response was similarly inhibited 60% by diltiazem and 66% by verapamil. Nicardipine abolished the carbachol phasic response at the 1 microM concentration. The amplitude of the phasic response was inhibited equally by 0.1 microM nicardipine (61.3% inhibition), 1 microM diltiazem (64.5% inhibition) and 1 microM verapamil (71% inhibition). The rate constant of decay of the phasic response was inhibited equally by 0.1 microM nicardipine (43% inhibition) and 1 microM diltiazem (29% inhibition). The rate constant of onset of the phasic response was unaffected by nicardipine, diltiazem and verapamil. Only 1 microM nicardipine inhibited the amplitude and rate constant of onset of the tonic response. The only effect of Bay K 8644 (1 microM) was to increase the phasic response amplitude. The CCB demonstrate a similar order of potency for inhibition of the phasic responses and clinical efficacy of the CCB in exercise-induced asthma (nicardipine > verapamil > diltiazem).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of oestrous cycle and exogenous ovarian steroids on 5-hydroxytryptamine metabolism in rat lung.

1. The metabolism of 5-hydroxytryptamine (5-HT) in homogenates of lung and in perfused isolated lungs, was measured, using [14C]5-HT, over the 4-day oestrous cycle in rats. 2. In lung homogenates, 5-HT metabolism (reflecting monoamine oxidase (MAO) activity) was greater during met-oestrus than during the rest of the cycle. 3. In homogenates, the apparent Km for MAO activity was lowest during met-oestrus whereas the, Vmax value was highest during di-oestrus. 4. In perfused lungs, 5-HT metabolism and uptake was greater during pro-oestrus than during met-oestrus. 5. In lung homogenates prepared from female rats treated with 17 beta-oestradiol or progesterone for 8 days, the metabolism of 5-HT was increased. 6. We conclude that although the lung is not usually considered a target organ for ovarian hormones, its metabolic activity is affected by changes in the levels of endogenous or exogenous ovarian steroids, an effect comparable with those already observed in uterus, ovary and other more commonly accepted target organs.

Effect of volatile anesthetics on endogenous tryptophan, 5-hydroxytryptophan and 5-hydroxytryptamine in rat lung.

Volatile anesthetics inhibit the pulmonary inactivation of 5-hydroxytryptamine (5-HT) possibly via an effect on endogenous lung 5-HT. The consequent higher systemic arterial 5-HT concentrations may predispose the heart to dysrhythmias. The direct effect of the anesthetics on endogenous 5-HT, its metabolites, and precursors in the isolated ventilated perfused rat lung was determined by high-pressure liquid chromatography. Halothane (0.45, 1.4, and 2.3 minimum alveolar concentration (MAC] and 35% nitrous oxide (N2O) increased lung 5-HT (11, 70, 94, and 54% respectively). The effect of 0.45 MAC halothane and 35% N2O on 5-HT was synergistic. Isoflurane (2.9 MAC) had no effect on lung 5-HT. The lung concentration of tryptophan (TRP) was increased 51% by 2.9 MAC isoflurane, but the rate of efflux of TRP from the lung was unchanged. There was no effect of the anesthetics on 5-hydroxytryptophan (5-HTP). The ratio of 5-HT:5-HTP was significantly increased by 2.3 MAC halothane and 0.5 MAC halothane +35% N2O. The 5-HTP:TRP ratio was unchanged. The metabolite of 5-HT, 5-hydroxyindole acetic acid (5-HIAA), was not always detected. The results suggest that the increase in lung 5-HT by halothane reflects an increase in 5-HTP decarboxylase activity and that halothane and isoflurane exert selective effects on lung 5-HT synthesis. The results do not support the hypothesis that lung 5-HT controls the inactivation of 5-HT in the pulmonary circulation.

Metabolism of 5-hydroxytryptophan in the isolated perfused rat lung.

The metabolism of 5-hydroxytryptophan (5-HTP) and tryptophan (TRP) in a single pass across the pulmonary circulation was studied in the isolated ventilated perfused rat lung and by high pressure liquid chromatography. The metabolism of 5-HTP was dependent on the rate of lung perfusion and the duration of infusion of 5-HTP, and was a saturable process with an apparent Km of 1.8 mM and Vmax of 0.14 mumol/g/3 min. The indoles found in the lung were 5-HTP, 5-hydroxytryptamine (5-HT) and 5-hydroxyindole acetic acid (5-HIAA); only 5-HIAA was detected in the lung effluent. The efflux of 5-HTP from the lung had two exponential components with half-lives of 0.15 and 3.65 min. After an infusion of 3H-5-HTP, the radiolabel that accumulated in lung was located mainly in the soluble fraction. An infusion of TRP resulted in the synthesis of 5-HTP, 5-HT and 5-HIAA in the lung, and 5-HTP was detected in the lung effluent. The results suggest that 5-HT can be synthesized in the intact lung from circulating TRP and 5-HTP. Since the rate of lung metabolism is low and no 5-HT is released into the lung effluent, the contribution of the lung to circulating levels of 5-HT is likely to be insignificant. Synthesis of 5-HT in intact lung suggests an intrapulmonary role for 5-HT.

Kinetic characterization of 5-hydroxytryptamine receptor desensitization in isolated guinea-pig trachea and rabbit aorta.

Desensitization of the contractile response mediated by the 5-hydroxytryptamine2 (5-HT2) receptor in the isolated guinea-pig trachea and rabbit aorta is a time-dependent process and therefore it has been characterized by an apparent rate constant obtained from a kinetic analysis. Under similar conditions, desensitization of the response in the trachea is 7-fold faster than in the aorta. Desensitization is homologous and reversible and is not affected by inhibition of neuronal and extraneuronal uptake, monoamine oxidase activity, alpha 1 adrenergic, cholinergic muscarinic or histamine H1 receptors. Desensitization does not depend on removal of epithelium from the trachea or endothelium and adventitia from the aorta or on the release of a stable relaxant factor. It is also not affected by the removal of extracellular Ca++, which is needed for tonic contraction. The dependence of desensitization on agonist concentration, number of receptors and the intrinsic activity of the agonist was determined. The observed values of the rate constants for desensitization and of the peak tension (T peak) in trachea show a saturable dependence on the concentration of 5-HT, indicating that occupancy of the 5-HT2 receptor is needed for desensitization. The less efficacious agonists, N-methyl serotonin, dimethyltryptamine, quipazine, 5-methoxytryptamine, 5-methyltryptamine, 5-methoxy dimethyltryptamine, 4-hydroxytryptamine and bufotenine induce significantly slower desensitization than 5-HT. A 25 to 75% reduction in 5-HT2 receptor number by alkylation had no effect on the observed rate constants for desensitization.(ABSTRACT TRUNCATED AT 250 WORDS)

Halothane- and enflurane-induced inhibition of phasic responses to carbachol in isolated guinea pig trachea.

Volatile anesthetics produce bronchodilatation by a combination of effects on the autonomic nervous system as well as direct effects on the airway smooth muscle. We sought to further explain the direct effects on airway smooth muscle by studying the effect of volatile anesthetics on the phasic and tonic components of the contractile response. The effects of 1.2%, 2.3%, and 4.3% halothane and 1.6%, 3.2%, 4.9%, and 8.7% enflurane on the phasic and tonic components of the contractile response to 10 microM carbachol and 40 mM KCl in isolated guinea pig trachea were determined using steady-state and kinetic analyses. No direct effect of either anesthetic on resting tension was observed. The peak tensions of the phasic responses to 10 microM carbachol were significantly inhibited by all concentrations of halothane and enflurane. The inhibition by enflurane was concentration-dependent and eliminated the phasic response at 8.7%. The peak tensions of the tonic responses to 10 microM carbachol were unaffected. The peak tensions of the responses to 40 mM KCl were unaffected by halothane but were significantly inhibited by 4.9% enflurane. The decay rate of the phasic response was unaffected by the anesthetics. The onset rate of the tonic response was significantly inhibited only by 8.7% enflurane, whereas halothane was without effect. The EC50 concentration for carbachol, as determined by cumulative concentration response curves, was increased by both 8.7% enflurane and 4.3% halothane. The results support the hypothesis that halothane and enflurane inhibit that portion of the contractile response to carbachol dependent on the release of intracellular Ca2+.(ABSTRACT TRUNCATED AT 250 WORDS)

Phasic responses to carbachol in isolated guinea pig trachea are augmented by cooling and inhibited by nifedipine.

Steady-state and kinetic approaches were used to study the effects of cooling to 25 degrees C and nifedipine (1 microM) on the phasic and tonic responses to 0.3 and 10 microM carbachol (Carb) in isolated segments of epithelium-free guinea pig trachea. Cooling and nifedipine had no effect on the steady-state tensions of the contractile responses to Carb in Krebs-bicarbonate buffer, but did inhibit the response to 40 mM KCl. Cooling increased the tensions of the phasic responses to both concentrations of Carb in Ca(++)-depleted, ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid-containing buffer, whereas the rate constant of decay of the phasic response (kdecay) was decreased only at 0.3 microM Carb. Nifedipine inhibited the tension and increased the kdecay of the phasic response to 10 microM Carb at both 37 and 25 degrees C. The phasic response to 0.3 microM Carb was essentially completely inhibited by nifedipine. The tension of the tonic response to Carb was unaffected by cooling and was only inhibited by nifedipine at 0.3 microM Carb at 37 degrees C. Both cooling and nifedipine, at 37 degrees C, decreased the rate constant of onset of the tonic response (kon) to Carb. The predominant effect of cooling to augment the phasic component of the contractile response to Carb, whereas it does not provide the mechanism to explain the enhanced bronchoconstrictor response of airways to cold, does suggest that Ca++ mobilization is altered by cold and that this may contribute to the enhanced bronchoconstrictor response.(ABSTRACT TRUNCATED AT 250 WORDS)

Inactivation of monoamines by the lung.

Inactivation of monoamines was the first pharmacokinetic property of lung to be identified. For some amines inactivation in perfused lung but not in liver is limited by uptake. Inactivation of beta-phenylethylamine in lung is limited not by uptake but by the activity of intracellular monoamine oxidase. The uptake of this amine is like that of basic drugs such as amphetamine and propranolol. Changes in the external environment of the lung (exposure in vivo to gaseous anaesthetics or to high concentrations of oxygen, greater than 95%) change amine inactivation processes in lung. The oestrous cycle and administration of ovarian steroids also affect the inactivation of 5-hydroxytryptamine and phenylethylamine. Monoamine oxidase activity in lung homogenates is highest at met- and lowest at pro-oestrus for both substrates. However, in perfused lung, inactivation is better correlated with variations in uptake. Therefore the main effect of the oestrous cycle is on uptake rather than on enzymic activity. Uptake processes are thus crucial determinants of monoamine activation in lung and the pharmacokinetic properties of lung are capable of responding to changes in its internal or external environment.

Modulation of rate at which serotonin-induced contraction decays in guinea pig trachea.

Stimulation of the type 2 serotonin (5-HT2) receptor in guinea pig trachea with 5-HT results in a contraction that decays in the continued presence of 5-HT. The decay of the 5-HT contraction has been proposed to be dependent on 5-HT2 receptor activation and to reflect desensitization of the receptor. The characteristics of the decay of the 5-HT contraction may also be dependent on other properties of the tissue. The effects of modulation of biochemical pathways implicated in airway smooth muscle contraction on the 5-HT contraction in isolated guinea pig trachea were determined with the use of a kinetic approach we developed previously. Decay of the 5-HT contraction was inhibited by cooling, increased by forskolin, 3-isobutylmethyl-1-xanthine, and 8-bromoadenosine 3',5'-cyclic monophosphate, and unaffected by staurosporine, H-7, H-8, phorbol 12,13-dibutyrate, and by inhibitors of the three major pathways of arachidonic metabolism. The results suggest that decay of the 5-HT contraction in guinea pig trachea is dependent on both the receptor and the biochemical state of the tissue.

The effect of lumbar epidural and general anesthesia on plasma catecholamines and hemodynamics during abdominal aortic aneurysm repair.

Twenty-four patients undergoing abdominal aortic aneurysm (AAA) repair were studied to compare the effects of lumbar epidural anesthesia (LEA) and general anesthesia (GA) on plasma catecholamine levels and hemodynamics before and during infrarenal aortic cross-clamping. Patients received either a high dose of opioid anesthetic (GA group, n = 12), or lumbar epidural anesthesia to T4 sensory level with a light general anesthetic (LEA group, n = 12). Systemic vascular resistance (SVR) and norepinephrine (NE) and epinephrine (E) levels were measured before anesthetic induction (before epidural activation in the LEA group, and before general anesthesia induction in the GA group), 15 min before cross-clamping, and 1,5, and 10 min after cross-clamping. There was a large (P < 0.05) increase in NE and E in the GA group by 15 min before aortic cross-clamping, but NE and E levels in the LEA group did not increase. The GA group had significantly higher levels of NE and E than the LEA group 15 min before cross-clamping and also after clamping. NE levels in the LEA group increased after cross-clamping, and NE levels in the GA group remained constant. E levels remained stable in both groups after cross-clamping. After clamping, SVR increased in both groups, but the increase occurred after 1 min in the GA group and took 5 min to become significant in the LEA group. There was no significant correlation between changes in NE or E and changes in SVR in either group. This study shows that epidural anesthesia to T4 prevents NE and E increases in response to abdominal surgery.(ABSTRACT TRUNCATED AT 250 WORDS)