Inhibition Of Extraneuronal Uptake Research Articles

Chronic Mild Stress–Induced Depression-Like Symptoms in Rats and Abnormalities in Catecholamine Uptake in Small Arteries

Major depression and cardiovascular diseases have a strong comorbidity; however, the reason for this is unknown. In the chronic mild stress (CMS) model of depression, only a fraction of rats develop a major feature of depression-anhedonia-like behavior, whereas other rats are stress resilient. Previous studies suggested that CMS rats also have increased total peripheral vascular resistance. On the basis of CMS-induced changes of sucrose intake, a reliable measure for anhedonia, rats were divided into "resilient" and "anhedonic" groups. An interaction between hedonic status and vascular function was studied after 4 and 8 weeks of CMS exposure in vitro in wire myograph on saphenous arteries and mesenteric small arteries (MSAs) from these rats. When comparing the different experimental rat groups, arterial sensitivities to noradrenaline (NA) were similar under control conditions, but in the presence of the neuronal reuptake inhibitor cocaine, arteries from anhedonic rats were more sensitive to NA. No change in perivascular innervation was found, but elevated expression of neuronal NA transporter was detected. Inhibition of extraneuronal uptake with corticosterone (1 μM) suggests that this transport is diminished in MSAs after CMS. The corticosterone-sensitive transporter organic cation cotransporter 2 was shown to be reduced in MSAs after CMS. No CMS-induced changes in the corticosterone-sensitive transport were found in saphenous arteries. Our results indicate that CMS-induced depression-like symptoms in rats are associated with changes in catecholamine uptake pathways in the vascular wall, which potentially modulates the effect of sympathetic innervation of resistance arteries.

Influence of inhibition of extraneuronal uptake and of O-methylation on the hyperglycaemia caused by sympathomimetic amines in depancreatized rats.

This study aimed at testing whether the O-methylating system (extraneuronal uptake + O-methylation) modulates, in-vivo, beta-adrenoceptor-mediated responses. The influences of U-0521 (3,4-dihydroxymethylpropiophenone, an inhibitor of catechol-O-methyl transferase (COMT)) and hydrocortisone (an inhibitor of extraneuronal uptake) on the hyperglycaemia evoked by isoprenaline and adrenaline were compared. Both inhibitors enhanced the increase of the plasma glucose level induced by either isoprenaline (0.36 nmol kg-1 min-1) or adrenaline (0.55 nmol kg-1 min-1). The enhancement caused by U-0521 developed faster than that caused by hydrocortisone, but was of the same magnitude. This is the first report of supersensitivity to sympathomimetic amines caused by inhibition of either COMT or extraneuronal uptake in-vivo and for a response not involving smooth muscle cells.

Effect of ω-Conotoxin GVIA on Noradrenaline Release from Postganglionic Sympathetic Neurones in Rabbit Aorta

The aim of the present work was to examine the effect of the selective N-type calcium blocking agent omega-conotoxin GVIA on stimulation-evoked release of noradrenaline from sympathetic nerves in rabbit isolated aorta with regard to stimulation frequency, extracellular Ca2+ concentration, and transmitter uptake. Rings of rabbit isolated aorta were preloaded with (-)-3H-noradrenaline and the fractional 3H-overflow evoked by electrical-field stimulation was determined by liquid scintillation spectrometry. Omega-conotoxin GVIA (3 x 10(-10)-3 x 10(-8) M) did not alter the spontaneous 3H-outflow. Omega-conotoxin GVIA (3 x 10(-10)-3 x 10(-8) M) caused a slowly developing reduction of stimulation-evoked 3H-overflow at 1 and 30 Hz. The Emax for the omega-conotoxin-induced inhibition was less (70%) at 30 Hz than that (96%) seen at 1 Hz. Short-term incubation with omega-conotoxin GVIA caused a subsequent steady-state inhibition. The inhibitory action of omega-conotoxin GVIA (3 x 10(-10)-3 x 10(-9) M) was inversely related to the extracellular Ca2+ concentration (6.5 x 10(-4)-2.7 x 10(-3) M). Cocaine (3 x 10(-5) M) plus corticosterone (4 x 10(-5) M), neuronal and extraneuronal uptake inhibitors, respectively, did not alter the inhibitory effect of omega-conotoxin GVIA (3 x 10(-9) M) on 3H-overflow evoked by stimulation at a frequency of either 1 or 30 Hz. It is concluded that omega-conotoxin GVIA acts on prejunctional N-type calcium channels to inhibit stimulation-evoked noradrenaline release from sympathetic neurone terminals in rabbit aorta. At a high frequency, another subtype calcium channel may possibly be involved. The action of omega-conotoxin GVIA is independent of neuronal and extraneuronal uptake mechanisms for noradrenaline, but dependent on the amount of Ca2+ to be transported across the neurilemma from the extracellular space into the neurone.

The influence of animal age on beta-adrenoceptor density and function in tracheal airway smooth muscle.

We examined the influence of animal age on the functional response of guinea-pig (0-156 weeks) and rat (4-136 weeks) isolated tracheal tissue to beta-adrenoceptor agonists. In addition, the binding density and affinity of [125I]iodocyanopindolol ([125I]CYP) binding to tracheal tissue was examined with respect to animal age. Significant age-related changes in isoprenaline potency were observed in tracheal ring preparations taken from animals during the early maturation phase of animal growth in the guinea-pig and rat. In addition, in rat isolated tracheal tissue, age-related decreases in fenoterol potency were observed during senescence, but not maturation. The changes in the functional responsiveness of tracheal tissue were not reflected by changes in the binding density or affinity for [125I]cyanopindolol ([125I]CYP) of beta-adrenoceptors, or in changes in specific autoradiographic grain density over smooth muscle tissue. In both guinea-pig and rat, no significant age-related changes in the influence of catechol-O-methyl transferase (COMT) or of extraneuronal uptake inhibition were detected. This study has demonstrated significant age-related changes in the responsiveness of guinea-pig and rat isolated tracheal tissue to beta-adrenoceptor agonists that were not related to changes in the density or affinity of the beta-adrenoceptor population or in the activity of COMT or extraneuronal uptake. The possibility of age-related changes in receptor-signal transduction coupling should be explored.

Extraneuronal uptake inhibitor U-0521 decreases contractile responses in rat vas deferens

1. 1. The influence of catechol-O-methyltransferase inhibitor U-0521 on isotonic contraction of isolated rat vas deferens was examined to determine optimal concentration and nonspecific effects. 2. 2. Maximum responses to (-)-epinephrine were increased at 0.4 μM and 1 μM concentrations of U-0521. Epinephrine responses were progressively decreased in the presence of higher concentrations (10 μM, 30 μM and 100 μM) of U-0521. 3. 3. The response to the nonadrenergic agonist neurokinin A was similarly depressed in the presence of 100 μM U-0521. 4. 4. U-0521 not only inhibits COMT, at concentrations above 1 μM it nonspecifically depresses contraction of the rat vas deferens by both adrenergic and nonadrenergic agonists.

Disprocynium24, a novel inhibitor of the extraneuronal monoamine transporter, has potent effects on the inactivation of circulating noradrenaline and adrenaline in conscious rat.

The role of extraneuronal uptake in terminating the actions of catecholamines has been difficult to evaluate in vivo, largely because of lack of suitable inhibitors. The compound, 1,1'-diisopropyl-2,4'-cyanine iodide or disprocynium24 (D24), is a novel inhibitor of extraneuronal uptake with a high degree of potency in vitro. This study examined the actions of D24 on the inactivation and metabolism of circulating noradrenaline and adrenaline in conscious rats. Animals received i.v. infusions of 3H-labelled noradrenaline and adrenaline, and their extraneuronal O-methylated metabolites, normetanephrine and metanephrine. Plasma concentrations of endogeneous and 3H-labelled catecholamines and metanephrines were measured before and after D24. D24 caused large increases in plasma concentrations of noradrenaline and adrenaline, effects due to both decreases in their plasma clearances and increases in their rates of release into plasma. Plasma concentrations of normetanephrine and metanephrine also increased due to their decreased clearance from plasma. Increased release of normetanephrine into plasma did not contribute to increased plasma concentrations of normetanephrine. In fact, the contribution of extraneuronal O-methylation to noradrenaline clearance decreased substantially after D24. The data indicate that D24 is a potent inhibitor of the extraneuronal catecholamine transporter in vivo and that this process contributes importantly to the removal of circulating catecholamines and their O-methylated amine metabolites. Increased release of noradrenaline into plasma may reflect an increase in the proportion of transmitter that escapes from sites of release into the circulation. However, increased adrenaline release indicates that the drug also causes sympathoadrenal activation.

Uptake mechanisms of meta-[ 123I]iodobenzylguanidine in isolated rat heart

In order to clarify the uptake and retention mechanisms of radioiodinated meta-iodobenzylguanidine (MIBG) in heart, the kinetics of no-carrier-added [ 123I]MIBG were studied in the isolated working rat heart in interaction with pharmacologic agents. The tracer was administered in the perfusate as a 10-min pulse, followed by a 90-min washout period. Kinetic analysis of the externally monitored time-activity curves of control hearts showed avid uptake ( K i = 4.4 ± 0.7 mL/min/g), and monoexponential clearance ( k o = 0.0056 ± 0.0017 l/min), indicating a distribution volume ( V d = K i k o ) of 834 ± 214 mL/g. Blocking experiments ( n = 41) were performed with neuronal uptake (uptake-1) inhibitor desipramine (DMI; 50–100 nM) and the extraneuronal uptake (uptake-2) inhibitor N-(9-fluorenyl)- N-methyl-β-chloroethylamine (SKF550; 0.4–0.8 μM). Uptake rate was 27% reduced ( P < 0.05) by 50 nM DMI but not significantly affected by 0.4 μM SKF550. Distribution volume was 88% reduced ( P < 0.0005) by 50 nM DMI and 28% reduced ( P < 0.05) by 0.4 μM SKF550. In DMI-blocked hearts, uptake rate was dramatically decreased (−80%, P < 0.0005) by SKF550 (0.4 μM), indicating uptake-2 transport contributed predominantly to the extraneuronal uptake of the tracer. The slow uptake rate seen with concomitant inhibition of uptake-1 and uptake-2 was further decreased by addition of unlabeled MIBG (1–10 μM) in a concentration-dependent manner, yet unaffected by addition of the vesicular uptake inhibitor Ro 4–1284 (1 μM). Thus, the uptake rate of [ 123I]MIBG is primarily dependent on uptake-1 and uptake-2 activity. Other possible mechanisms of uptake such as passive diffusion in association with intracellular binding are significant only in conditions where uptake-1 and uptake-2 mechanisms are largely inhibited.

Post-natal development modulates the rat seminal vesicle sensitivity to sympathomimetic agonists.

The post-natal development-related changes in the sensitivity to sympathomimetic agonists were studied in the seminal vesicle of 40-, 60- and 120-day old rats by determining the pD2 values for adrenaline, phenylephrine and methoxamine. The age-related changes in the neuronal and extraneuronal uptake of sympathomimetic agonists were also determined. The seminal vesicle sensitivity to adrenaline, phenylephrine and methoxamine increased in sexually mature rats (60- and 120-day old) (concentration-effect curves shifted to the left and pD2 values increased) when compared with immature rats (40-day old). Cocaine, a neuronal uptake inhibitor, induced supersensitivity to adrenaline at all ages, but did not affect the sensitivity to methoxamine. The ratio between the EC50 values for adrenaline in the presence and in the absence of cocaine was reduced in tissues of mature animals. This reduction suggests a loss of function of the neuronal uptake system with sexual development. In the presence of cocaine the pD2 value for adrenaline was increased during sexual development. Thus, the age-related changes in the adrenaline sensitivity were observed even when neuronal uptake was not operative, suggesting age-related changes at the post-junctional level, which was confirmed by the increase in the seminal vesicle sensitivity to methoxamine. In the presence of the extraneuronal uptake inhibitor, corticosterone, the sensitivity to adrenaline did not change and the age-related changes persisted.(ABSTRACT TRUNCATED AT 250 WORDS)

The influence of the density of adrenergic innervation on the extracellular steady-state concentration gradient for 3H-noradrenaline.

After inhibition of extraneuronal uptake by corticosterone, isolated right atria and lengthwise halved vasa deferentia of the rat were incubated with 0.2 mumol/l 3H-noradrenaline for 60 min, washed out for 100 min and then prepared for autoradiography. The autoradiographic images were digitized, and silver grain density was determined as a function of the distance from the surface. Silver grain density declined towards the centre of the tissue; the decline was monophasic exponential and significantly steeper in the vas deferens (0.016 microns-1) than in the less densely innervated right atrium (0.011 microns-1). Silver grain density at the surface of the tissue was higher in vas deferens than in right atrium. The results show that the extracellular steady-state concentration gradient for 3H-noradrenaline (generated by uptake1 during the incubation with this amine) largely depends on the density of the adrenergic innervation.

Uptake of norepinephrine in an isolated artery from normotensive humans.

Previous studies have suggested that catecholamine uptake may play a role in vascular responsiveness in hypertension. The current study was undertaken to characterize amine uptake and effects of its inhibition in an isolated human artery from normotensive subjects and to provide a basis for future study in hypertensive individuals. Accumulation of tritiated norepinephrine into the artery of normotensive subjects was time-dependent and 16.9-fold more than the incubation media concentration (1 microM) of amine after 60 minutes. There was a lesser accumulation of tritiated normetanephrine (3.1-fold), and it was not increased over time. Increasing the concentration of norepinephrine to 30 microM did not significantly change the proportional accumulation. Inhibition of neuronal uptake with cocaine (10 microM) reduced the average accumulation of both concentrations of tritiated norepinephrine to 3.9-fold (p less than 0.001). Inhibition of extraneuronal uptake with corticosterone alone (10 microM) had no significant effect on average accumulation of norepinephrine, and where combined with cocaine, there was no further effect of corticosterone. Neither cocaine nor corticosterone had any effects on accumulation of normetanephrine. In spite of elimination of approximately 75% of the uptake of norepinephrine, cocaine had very little potentiating effect on mechanical responses to exogenous norepinephrine and neurally released transmitter. Thus, norepinephrine uptake in human cystic artery is characteristic of neuronal uptake, but cocaine treatment has only a very modest potentiating effect on responsiveness to endogenous norepinephrine and no significant effect on responsiveness to exogenous amine.

Subsensitivity to β-adrenoceptor agonists in pacemakers isolated from senescent rats

1. 1. The effects of aging on the responsiveness of the isolated pacemaker of the rat to the chronotropic effect of norepinephrine, isoproterenol and soterenol were studied. 2. 2. Pacemakers isolated from senescent rats (22–24 months old) showed subsensitivity to norepinephrine, isoproterenol and soterenol, when compared with pacemakers isolated from young rats (3–4 months old). The maximum response to the partial agonist soterenol was reduced. 3. 3. Determination of the pA 2 value of metoprolol in pacemakers isolated from senescent and young rats showed that the chronotropic response is mediated by a homogeneous β 1-adrenoceptor population. 4. 4. Inhibition of extraneuronal uptake did not potentiate the chronotropic effect of isoproterenol in pacemakers isolated from senescent rats. Addition of cocaine shifted the concentration-effect curve for norepinephrine only 2.2-fold to the left in senescent rats. 5. 5. It is concluded that, during aging, impairment of the extraneuronal and neuronal uptake mechanisms has an important role in the control of chronotropic responsiveness to catecholamines.

Characterization of norepinephrine accumulation by a crude synaptosomal-mitochondrial fraction isolated from rat heart

Norepinephrine (NE) uptake into a heart synaptosomal-mitochondrial fraction was assessed under conditions where neuronal uptake (type 1) was linear with respect to both time and protein concentration. The NE accumulation process was sensitive to incubation temperature, sodium ion concentration and medium osmolality. Furthermore, NE uptake was attenuated by the neuronal uptake inhibitor desmethylimipramine (DMI) in a concentration dependent manner; the IC 50 value was approximately 10 nM and maximum inhibition was obtained at 100 nM. In contrast, the extraneuronal uptake inhibitor, metanephrine did not significantly attenuate NE uptake. Kinetic analysis demonstrated that the DMI sensitive NE accumulation is saturable with a K M of approximately 400 nM and that NE uptake occurs via a single uptake process. This demonstration of neuronal type NE uptake by a synaptosomal-mitochondrial fraction constitutes a successful demonstration of the preparation of a rat heart subcellular fraction containing functional synaptosomes.

β1‐Adrenoceptors mediate smooth muscle relaxation in mouse isolated trachea

1. The relaxant effects to the beta-adrenoceptor agonists isoprenaline, adrenaline, noradrenaline, RO363, procaterol and fenoterol were investigated in carbachol-contracted mouse isolated tracheal preparations. 2. The order of potencies for those beta-adrenoceptor agonists that induced full relaxation of carbachol-contracted mouse tracheal preparations was isoprenaline greater than RO363 greater than noradrenaline = adrenaline greater than fenoterol. The EC50 value of isoprenaline for relaxation was 46 nM. The beta 1-adrenoceptor-selective agonist, RO363 was ten times more potent than the beta 2-adrenoceptor-selective agonist, fenoterol. The highly beta 2-adrenoceptor-selective agonist procaterol was a partial relaxant and induced only 28 +/- 4% relaxation. 3. Relaxations induced by noradrenaline and isoprenaline were not significantly affected by the neuronal uptake inhibitor, cocaine (10 microM) or by the extraneuronal uptake inhibitor, deoxycorticosterone acetate (25 microM) respectively. The alpha-adrenoceptor agonist methoxamine induced no observable elevation of mouse tracheal smooth muscle tone. 4. Schild plots for the beta-adrenoceptor antagonists, atenolol and betaxolol (beta 1-adrenoceptor-selective) and ICI 118,551 (beta 2-adrenoceptor-selective) were linear, with slope values approaching unity. Mean pA2 values derived for atenolol, betaxolol and ICI 118,551 for antagonism of beta-adrenoceptor-mediated relaxation were 7.1, 8.4 and 7.2, respectively. These data were independent of the use of isoprenaline or noradrenaline as the agonist. 5. These findings indicate that beta-adrenoceptor-mediated relaxations of mouse isolated trachea occur predominantly through activation of beta 1-adrenoceptors.

The activity of the neuronal and extraneuronal catecholamine-metabolizing enzymes of the perfused rat heart.

In a comparative study the neuronal and extraneuronal metabolism of several 3H-catecholamines (all of which were tritiated in the C-7 position of the side chain only) was determined in isolated rat hearts perfused at a concentration of the 3H-amines of 50 nmol/l. While the neuronal MAO activity was determined after inhibition of extraneuronal uptake (100 mumol/l OMI) and COMT (10 mumol/l U-0521), the extraneuronal MAO activity was estimated after inhibition of neuronal uptake (30 mumol/l cocaine) and COMT. The extraneuronal COMT activity was determined under conditions of inhibition of both neuronal uptake and MAO (pretreatment with pargyline). Hearts were perfused with the 3H-catecholamines until the rate of appearance of the various 3H-metabolites in the venous effluent has reached a steady state. From these rates (vst-st) and the steady-state content of the unchanged 3H-catecholamines in the tissue (Si), the rate constants (Vmax/Km) for the unsaturated intracellular enzymes COMT (kCOMT) and MAO (kMAO) were calculated. The kCOMT values for all four catecholamines, (-)-noradrenaline, dopamine, (-)-adrenaline and (+/-)-isoprenaline exhibit a range from 0.24 to 0.78 min-1; the metabolism of the catecholamines by the COMT differs: (-)-noradrenaline = dopamine less than (-)-adrenaline less than (+/-)-isoprenaline. The extraneuronal MAO activity was low for all three catecholamines, (-)-adrenaline, (-)-noradrenaline and dopamine (range of kMAO from 0.05 to 0.28 min-1) and declined in the order.(ABSTRACT TRUNCATED AT 250 WORDS)

Characteristics of the cocaine-sensitive accumulation and O-methylation of 3H-(-)-noradrenaline by rabbit endometrium.

1. The extraneuronal uptake and O-methylation of 2,5,6 3H-(-)-noradrenaline was studied in segments of uterine endometrium from rabbits pretreated with 17 beta-oestradiol and progesterone. 2. The uptake of 3H-noradrenaline was measured in MAO- and COMT-inhibited tissues and obeyed Michaelis-Menten kinetics with an apparent Km of 78 mumol/l and a Vmax of 5.4 nmol/g min. Uptake was inhibited by low Na+, and by potential substrates in the order dopamine greater than (-)adrenaline greater than (-)isoprenaline = 5-hydroxytryptamine. 3. Following uptake at 1.2 mumol/l, efflux of 3H-noradrenaline was slow and appeared to be from two compartments, of which the first (I) had a t1/2 of 53 min and a capacity of 1.8 nmol/g. The presence of the second compartment (II) was inferred from the tissue content of 3H after 60 min of efflux, which was 3-4 times greater than predicted if the 3H was present in compartment I only. Following incubation with 3H-noradrenaline in the presence of cocaine 30 mumol/l the 3H efflux was rapid and the combined capacities of compartments I and II were greatly decreased. 4. 3H-NMN formation, measured in MAO-inhibited tissues, obeyed Michaelis-Menten kinetics with a half-saturating outside concentration of 12 mumol/l and a Vmax of 0.9 nmol/g X min. The formation was inhibited by the neuronal uptake inhibitors, desipramine 3 mumol/l and metaraminol 100 mumol/l (each by 80%), but was unaffected by the extraneuronal uptake inhibitor, NMN 100 mumol/l, and by oxytetracycline 100 mumol/l and methoxamine 10 mumol/l. 5.(ABSTRACT TRUNCATED AT 250 WORDS)

A role for corticosterone in the in vivo regulation of the extraneuronal uptake of [3H]-isoprenaline into rat atria.

The role of corticosterone, a potent in vitro inhibitor of extraneuronal uptake, in the regulation of the extraneuronal accumulation of [3H]-isoprenaline into rat atria was examined. Procedures which increased plasma corticosterone levels resulted in a decrease in the corticosterone-sensitive component of extraneuronal accumulation of [3H]-isoprenaline (64% after reserpine treatment and 25% after chronic cold exposure). Procedures which decreased levels of plasma corticosterone resulted in an increase in the corticosterone-sensitive component of extraneuronal accumulation of [3H]-isoprenaline (approximately 20% after adrenalectomy and 32% after hypophysectomy). This increase was partially prevented by the in vivo administration of dexamethasone (40 micrograms kg-1). There was a strong inverse correlation between the plasma corticosterone concentration and the level of extraneuronal uptake into atria (P less than 0.01). Corticosterone appears to play a major role in the regulation of extraneuronal uptake into tissues of the rat.

The influence of oestrogen and oestrogen metabolites on the sensitivity of the isolated rabbit aorta to catecholamines

In the present study the influence of oestradiol, catechol oestrogens, and O-methylated oestrogens was determined on the contractile responses of the isolated rabbit aorta to (-)-adrenaline. Oestradiol (40 mumol/l), 2-hydroxyoestradiol (2OHE2) (20 mumol/l), and 2-methoxyoestradiol (2MeOE2) (20 mumol/l) all sensitized the rabbit aorta to contractile responses to (-)-adrenaline. Similarly, the 2-hydroxy and 2-methoxy derivatives of oestrone and oestriol also sensitized the aorta to (-)-adrenaline-induced contractions. The largest degree of sensitization was seen in the presence of the 2-methoxysteroids. Oestradiol and 2OHE2 did not increase responses of the aorta to (-)-noradrenaline, while slight potentiation of contraction was seen in the presence of 2MeOE3. The potentiating effect of 2OHE2 on contractile responses to (-)-adrenaline was abolished by prior treatment of the tissue with a COMT inhibitor (U-0521, 55 mumol/l). Conversely, pretreatment of the tissue with 2OHE2 prevented the augmented aortic contraction to (-)-adrenaline usually seen after inhibition of COMT. The non-additive nature of the sensitization seen after combined treatment with 2OHE2 and U-0521 was qualitatively similar to that seen following combined exposure to maximally effective concentrations of U-0521 and an inhibitor of extraneuronal uptake (hydrocortisone 100 mumol/l). Oestradiol and 2MeOE2 reduced the formation of both the 3H-O-methylated, 3H-deaminated and the 3H-O-methylated deaminated metabolites of 3H-(-)-adrenaline (0.15 mumol/l) during exposure of the aorta to the tritiated catecholamine.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of extraneuronal uptake inhibitors on the positive chronotropic response to isoprenaline and on the accumulation of isoprenaline in perfused rat heart after inhibition of catechol-O-methyl transferase.

Experiments were carried out in isolated perfused rat hearts. The presence of tropolone (100 mumol/l), an inhibitor of catechol-O-methyl transferase (COMT), significantly potentiated the positive chronotropic response to isoprenaline (0.1, 1, 3 and 10 nmol/l). Two uptake2 inhibitors, 3-O-methylisoprenaline (100 mumol/l) and normetanephrine (100 mumol/l), induced a positive chronotropic response, but corticosterone (100 mumol/l) and hydrocortisone (100 mumol/l) had no such effect. 3-O-methylisoprenaline (100 mumol/l) and normetanephrine (100 mumol/l) enhanced the positive chronotropic response to isoprenaline (0.1, 1, 3 and 10 nmol/l). Corticosterone (100 mumol/l) potentiated the positive chronotropic response to isoprenaline (0.1 and 1 nmol/l). Hydrocortisone (30 mumol/l) enhanced the response to 0.1 nmol/l isoprenaline but did not affect the positive chronotropic responses to 1, 3 or 10 nmol/l isoprenaline. The addition of uptake2 inhibitors (3-O-methylisoprenaline, 100 mumol/l; normetanephrine, 100 mumol/l; corticosterone, 100 mumol/l) to the perfusion medium significantly reduced the positive chronotropic response to the perfusion with isoprenaline (3 nmol/l) and tropolone (100 mumol/l). The accumulation of 3H-isoprenaline in the heart perfused with 3H-isoprenaline (1, 10 and 100 nmol/l) was significantly increased by the presence of tropolone (100 mumol/l): the accumulation for 1, 10 and 100 nmol/l of 3H-isoprenaline was 5.07, 47.0 and 500 pmol/g, respectively. The high accumulation observed during perfusion with 3H-isoprenaline (3 nmol/l) and tropolone (100 mumol/l) was significantly decreased by the addition of an uptake2 inhibitor, 3-O-methylisoprenaline (100 mumol/l), normetanephrine (100 mumol/l) or corticosterone (100 mumol/l), but not by hydrocortisone (30 mumol/l).(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of epithelium removal on the sensitivity of guinea‐pig isolated trachealis to bronchodilator drugs

Mechanical removal of the epithelium increased the sensitivity of tracheal strips to isoprenaline, sodium nitroprusside, and to adenosine (only in the presence of inhibitors of its uptake and metabolism). Epithelium removal was without effect on sensitivity to salbutamol or papaverine. Preincubation of tracheal strips with an inhibitor of extraneuronal uptake, corticosterone (50 microM), had no effect on tissue sensitivity to either salbutamol or papaverine. However, the steroid both increased sensitivity to isoprenaline, and abolished the effect of epithelium removal on sensitivity to this catecholamine. These results suggest that in the guinea-pig, the tracheal epithelium is a major source of extraneuronal uptake for catecholamines. Furthermore, the increase in trachealis sensitivity to isoprenaline following epithelium removal is probably due to loss of these sites of extraneuronal uptake. The fact that sensitivity to salbutamol, papaverine and adenosine (in the absence of metabolic inhibitors) was not increased by denuding the epithelium indicates that loss of a diffusion barrier to drugs is not the mechanism of increased sensitivity. Adenosine (and possibly nitroprusside) may cause the epithelium to release a smooth muscle excitatory factor. Thus, removal of the epithelium attenuates this excitatory influence and enhances smooth muscle responsiveness to adenosine. These results provide further evidence that the epithelium has an important role in modulating the sensitivity of guinea-pig trachealis to drugs.

Differential effects of ketamine isomers on neuronal and extraneuronal catecholamine uptake mechanisms.

Contractile responses of isolated rabbit aortic strips to epinephrine and norepinephrine were potentiated in a dose-related manner by (+) ketamine but not by (-) ketamine (1.1 X 10(-5) M - 3.7 X 10(-4) M). Potentiation was blocked completely by pretreatment with the extraneuronal uptake inhibitor cortisol (83-138 microM) but was unaffected by the neuronal uptake inhibitor cocaine (29 microM). Responses of the rat anococcygeus muscle to these catecholamines were potentiated by both isomers, with (+) ketamine being more potent than its optical antipode. These effects were blocked completely in tissues from 6-hydroxydopamine sympathectomized animals. Results suggest that inhibition of extraneuronal uptake of catecholamines by racemic ketamine is due solely to an action of the (+) isomer, whereas both isomers appear capable of inhibiting neuronal uptake.