The effect of alpha 2-adrenoceptor agonists on the acid secretory responses of rat isolated gastric mucosa to electrical field stimulation.

The effects of clonidine, UK-14,304, noradrenaline, para-aminoclonidine and phenylephrine were examined on the acid secretory response of the rat isolated gastric mucosa preparation to electrical field stimulation. Clonidine, UK-14,304, noradrenaline and para-aminoclonidine but not phenylephrine (10 microM) reduced the response of the gastric mucosa stimulated at 2.5 Hz; gastric mucosae stimulated at higher frequencies were insensitive to the action of these alpha 2-adrenoceptor agonists. The inhibitory effect of the selective alpha 2-adrenoceptor agonist UK-14,304 was antagonized by idazoxan but not by prazosin. These findings indicate that clonidine and other alpha 2-adrenoceptor agents inhibit the acid secretory response of the rat gastric mucosa to electrical field stimulation by an action at alpha 2-adrenoceptors, which are probably located on cholinergic nerve terminals.     

Role of acetylcholine in electrical stimulation-induced arrhythmia in rat isolated atria.

In this study, we used the spontaneously beating, isolated rat right atrium as an in vitro model to study arrhythmogenic effects of electrical stimulation. A tetrapolar platinum electrode was used for stimulation and recording of atrial electrical activity at 36.5 degrees C (spontaneous rate, 4.9+/-0.3 Hz). A flutter-like pattern of arrhythmia was reproducibly induced by application of stimulus trains (250 pulses, 66.7 Hz). Arrhythmia was characterized by regular and very short cycle length (40-70 ms), each episode lasting from 3 s to >5 min. In control conditions, application of one to five pulse trains was sufficient to induce arrhythmia. However, atropine (but not propranolol) completely blocked arrhythmia induction (10-15 consecutive trains were ineffective). The ability of electrical stimulation to evoke arrhythmia was restored after atropine washout. A milder stimulation protocol (30 pulses, 50 Hz), which was unable to evoke arrhythmia in control conditions, was fully effective in the presence of 1 microM acetylcholine (ACh). Furthermore, a similar flutter-like pattern could be induced in isolated left atria in the presence of ACh. Our results point out an arrhythmogenic effect of neurally released ACh in the isolated right atrium on atrial electrical stimulation.     

Influence of chronic volume overload-induced atrial remodeling on electrophysiological responses to cholinergic receptor stimulation in the isolated rat atria

Whereas molecular mechanisms of atrial fibrillation (AF) have been widely investigated, there is limited information regarding interrelation between chronic volume overload and parasympathetic nervous system in the pathophysiology of AF. In this study, we investigated the influence of abdominal aorto-venocaval shunt (AVS)-induced atrial remodeling on electrophysiological responses to cholinergic receptor stimulation in the isolated rat atria. Interstitial fibrosis, cardiomyocyte hypertrophy and atrial enlargement, known as structural arrhythmogenic substrates for AF, took place after one month of AVS operation. Carbachol at 0.1 and 1 μM shortened the effective refractory period, acting as functional arrhythmogenic substrates, but increased the conduction velocity both in the atria of the sham-operated and AVS rats. The extents of the electrophysiological responses to carbachol in the atria of the AVS rat were greater than those in the sham-operated ones. Also, the higher inducibility and longer duration of carbachol-mediated AF were detected in the AVS atria than those in the sham-operated ones. These results showed that chronic volume overload-induced atrial remodeling promoted electrophysiological responses to cholinergic receptor stimulation in the isolated atria of rats, suggesting possible synergistic actions between structural arrhythmogenic substrate in the remodeled atria and functional arrhythmogenic substrates modulated by parasympathetic nerve activity.     

Mechanical responses of rat isolated uterine horns to transmural stimulation.

1 Transmural stimulation of rat isolated uterine horns at low pulse width produced contractions. These were antagonized by hyoscine or tetrodotoxin and potentiated by physostigmine. 2 In the presence of hyoscine, and during bradykinin-induced contractions, transmural stimulation produced inhibition. This inhibition was antagonized by guanethidine, propranolol or tetrodotoxin. 3 Hexamethonium or pempidine did not affect responses to transmural stimulation. 4 It is suggested that transmural stimulation is a method of exciting cholinergic motor and noradrenergic inhibitory postganglionic neurones to the rat myometrium.     

Diminution by benzodiazepines of the chronotropic responses to noradrenaline in rat isolated atria

The effects of various benzodiazepines on chronotropic responses were assayed in spontaneously beating rat isolated atria. The increases in atrial rate obtained from concentration-response curves to noradrenaline were reduced dose dependently by both the peripheral agonist, Ro 5-4864 5 and 10 microM, and the mixed agonist, diazepam 5, 10 and 50 microM, but not by the central benzodiazepine agonist, clonazepam 10 and 30 microM. The inhibitory effects of the benzodiazepines on the atrial responses to noradrenaline were not counteracted by either the peripheral benzodiazepine antagonist, PK 11195 10 microM, or the central benzodiazepine antagonist, Ro 15-1788 10 and 100 microM. Both 10 microM Ro 5-4864 and 10 microM diazepam also reduced the increases in atrial rate produced by either the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine, or the adenylate cyclase activator, forskolin. On the contrary, diazepam and Ro 5-4864 did not modify the chronotropic responses of the atria either to direct exposure to CaCl2 or to the calcium agonist, BAY K 8644. The increases in the intracellular levels of cAMP induced by noradrenaline were not modified by Ro 5-4864 and were even increased by 50% in the presence of diazepam. It is concluded that benzodiazepines probably reduce the chronotropic responses to noradrenaline in rat isolated atria through the interaction with the cAMP-linked chain of events that follows the activation of beta-adrenoceptors.     

Thermogenic responses to adrenoceptor agonists and brown fat adrenoceptors in overfed rats

Rats fed a cafeteria diet to produce hyperphagia showed increases in the maximal thermogenic responses (rise in oxygen consumption) to isoprenaline (mixed beta-agonist), prenalterol (beta 1-selective agonist) and clenbuterol (beta 2-agonist), and left-shifts in the dose-response curves to the latter two. The maximal response to phenylephrine (alpha-agonist) was similar for control and cafeteria rats. Ligand binding studies revealed increases in beta-adrenoceptor density of 33-38% in brown fat cells and isolated membranes from cafeteria-fed rats, but a 30% reduction in beta-receptors in heart membranes. Cold-adaptation caused a 22% reduction in beta-receptor density in brown fat membranes, but no change in heart. The ratio of beta 1/beta 2-receptors in brown fat was reduced from 59/45 in control to 47/54 in cafeteria-fed rats, but was not significantly altered in heart (58/44) or in brown fat from cold-adapted animals (64/30). alpha-Adrenoceptor density was increased above control values by 69 and 25% in brown adipose tissue from cafeteria and cold-adapted rats, respectively.     

Involvement of prostaglandins in the inhibitory response of adrenoceptor agonists in the rat isolated uterus

1. The possible involvement of intramurally generated prostaglandins in the responses produced by noradrenaline (NA), adrenaline (ADR) and salbutamol (SAL) has been investigated in the rat isolated uterus. 2. NA, ADR and SAL produced concentration-related inhibition of acetylcholine-induced tone in preparations from the proestrus, oestrus, metoestrus and dioestrus phases of the oestrous cycle, and from ovariectomized animals. 3. The cyclo-oxygenase inhibitor, flurbiprofen (FBF), enhanced uterine inhibitory response to all three agonists under the different hormonal conditions. 4. The dual inhibitor of cyclo-oxygenase and lipoxygenase, 3-amino-l-(m-trifluoromethyl)-phenyl-2-pyrazolone (BW 755C) also potentiated uterine response to NA, ADR and SAL, but the effect produced by BW 755C was similar to that achieved in the presence of FBF. 5. In contrast, the inhibition elicited by histamine and papaverine were unaffected by FBF treatment. 6. The above results provide pharmacological evidence that adrenoceptor agonists may influence prostaglandin production in the rat uterus both in the presence, and absence of the ovarian hormones.     

The influence of oxidative stress on various inotropic responses in isolated rat left atria

The effects of free radicals, generated by electrolysis of a physiological salt solution, on various inotropic responses to drugs in isolated rat left atria were studied. Evidence for the generation of hydroxyl radicals was obtained from an appropriate fluorimetric assay. The amount of free radicals produced by electrolysis of the medium proved current-dependent. Exposure of isolated rat left atria to the medium which had been subjected to electrolysis caused a current-dependent decrease in contractile force. Oxidative stress, as a result of the electrolysis of the medium, caused altered inotropic responses to extra cellular Ca²⁺ (pD₂ control group: 2.62 ± 0.06 vs. 2.44 ± 0.07 electrolysis group), sodium withdrawal (rise in contractile force control group: 1.73 ± 0.19 mN vs. 0.48 ± 0.21 mN electrolysis group) and lowering of stimulation frequency. The response to isoprenaline was diminished in atria subjected to oxidative stress and led to a rightward shift of the concentration response curves (pD₂ control group: 7.56 ± 0.10 vs. 6.77 ± 0.11 electrolysis group). In addition, the inotropic responses to forskolin (pD₂ control group: 6.17 ± 0.12 vs. < 4.5 electrolysis group) and dibutyryl cAMP (rise in contractile force caused by 1 × 10^–5 M db-cAMP in control group: 2.15 ± 0.01 mN vs. 1.21 ± 0.10 mN electrolysis group) proved blunted as well. Measurement of the adenylyl cyclase activity revealed that free radicals attenuated the basal (by 11.1%) and forskolin stimulated (155.0 ± 5.1 vs. 48.0 ± 1.8 pmol cAMP/mg prot./min for control and electrolysis group respectively) activity of the adenylyl cyclase. DMSO, a well known hydroxyl radical scavenger, was able to abolish the free radical-induced decrease in the response to isoprenaline. Surprisingly, addition of α-adrenoceptor agonists to atria subjected to electrolysis-generated free radicals led to a rapid decrease in contractile force. DMSO was unable to counteract the negative intropic effect of methoxamine in atria subjected to oxidative stress. This negative inotropic response to α-adrenoceptor agonists in atria subjected to electrolysed medium is unlikely to be the direct result of phospholipase C or protein kinase C activation. Angiotensin II (which stimulates PLC as well) did not reduce contractile force and chelerythrine (a PKC inhibitor) was unable to counteract the negative inotropic effect of the adrenoceptor agonists. In addition, the negative inotropic effect of methoxamine proved insensitive to 10^–6 M phentolamine and 10^–5 M doxazosin, which indicates an α-adrenoceptor independent mechanism. From this study we conclude that free radicals alter responses to various inotropic stimuli. These alterations may be the result of injured contractile elements, transporter molecules and molecules involved in signal transduction. Addition of α-adrenoceptor agonists after oxidative stress leads to a α-adrenoceptor, PLC and PKC independent decrease in contractile force. Received: 17 June 1996 / Accepted: 6 November 1996     

Sertraline inhibits the contractile responses to noradrenaline,KCI and electrical field stimulation of rat isolated vas deferens

1 The aim of this study was to investigate the effect of sertraline, a selective serotonin re-uptake inhibitor, on contractile responses to noradrenaline (NA), KCI, serotonin (5-HT) and electrical field stimulation of rat isolated vas deferens. 2 Pre-treatment with 10^-4 M sertraline showed inhibitory effects on responses to NA, KCI, 5-HT and electrical field stimulation, while pre-treatment with 10 ^-6 and 10^-5 M sertraline caused potentiation of responses to NA (10^-7 and 10^-6 M). 3 A voltage-dependent calcium channel activator, Bay K 8644, restored the inhibited responses when sertraline was washed out of the organ bath, although restoration could not be seen when sertraline was not removed. 4 The inhibition of the contractile responses by sertraline pre-treatment may be via a mechanism through calcium channels which is additional to the selective serotonin re-uptake inhibitory effect of sertraline.     

Sertraline inhibits the contractile responses to noradrenaline, KCl and electrical field stimulation of rat isolated vas deferens

1. The aim of this study was to investigate the effect of sertraline, a selective serotonin re-uptake inhibitor, on contractile responses to noradrenaline (NA), KCl, serotonin (5-HT) and electrical field stimulation of rat isolated vas deferens. 2. Pre-treatment with 10(-4) M sertraline showed inhibitory effects on responses to NA, KCl, 5-HT and electrical field stimulation, while pre-treatment with 10(-6) and 10(-5) M sertraline caused potentiation of responses to NA (10(-7) and 10(-6) M). 3. A voltage-dependent calcium channel activator, Bay K 8644, restored the inhibited responses when sertraline was washed out of the organ bath, although restoration could not be seen when sertraline was not removed. 4. The inhibition of the contractile responses by sertraline pre-treatment may be via a mechanism through calcium channels which is additional to the selective serotonin re-uptake inhibitory effect of sertraline.     

Uptake of 5-hydroxytryptamine in rat isolated atria.

1. The mode of 5-hydroxytryptamine (5-HT) uptake by the rat isolated atria was studied and compared to noradrenaline (NA) uptake. 2. Rat isolated atria were incubated with 14C-5-HT (46 microM) or 3H-NA 0.4 microM). After washing, the radioactivity fixed in atria was counted and the total NA, 5-HT and 5-hydroxyindol-3-acetic acid (5-HIAA) atria contents were measured by HPLC. 3. 14C-5-HT uptake was reduced in atria from 6-hydroxydopamine (100 mg/kg, i.p., 48 hr before experiments) or reserpine (2.5 mg/kg, i.p., 24 and 48 hr before experiments) pretreated rats. 4. The incubation of atria with 5-HT (50 microM) at the same time as 3H-NA reduced the 3H-NA value fixed. 5. Addition of desipramine (1 microM) or hydrocortisone (150 microM) before the incubation of atria with 14C-5-HT was without any effect on 14C-5-HT uptake. In contrast, fluvoxamine (1 microM) or indalpine (5 microM) caused a slight inhibition. 6. These data indicate that 5-HT is taken into the NA storage vesicles within the atria sympathetic nerves. This uptake does not use the NA carrier and involves partly the 5-HT carrier. An extraneuronal accumulation was noticed and a part of it is intracellular.     

Actions of mescaline on isolated rat atria.

Mescaline, in concentrations of 5 x 10(-4) and 1 x 10(-3) M, produced negative chronotropic and positive inotropic responses in isolated, spontaneously beating rat atria. In tissues driven at a constant rate, the inotropic response was diminished greatly, indicating that the increment in the force of contraction was secondary to the reduction in rate. These chronotropic and inotropic responses were not altered consistently by pretreatment with the histamine antagonists chlorpheniramine and metiamide.     

Calcium antagonists inhibit positive chronotropic responses to alpha 1-adrenoceptor activation in rat isolated atria

Positive chronotropic responses of rat isolated atria to phenylephrine were reduced by propranolol (0.3 microM) and the residual response was further depressed by the selective alpha 1-adrenoceptor antagonist prazosin (1 nM) but not yohimbine (10 nM), confirming that a component of the response to phenylephrine was due to activation of alpha 1-adrenoceptors. When beta-adrenoceptors were blocked by propranolol, the positive chronotropic response to phenylephrine was enhanced by increasing the calcium concentration and by the calcium channel activator Bay K 8644 (0.1 microM), whereas the response was decreased by lowering the calcium concentration and by the calcium antagonists verapamil (10 nM), nifedipine (10 nM) and diltiazem (100 nM). In the presence of prazosin, when phenylephrine acts only on beta-adrenoceptors, calcium antagonists had no effect on the response. In rat isolated aortic strips in a calcium-free, high K+ (40 mM) solution, verapamil (10 nM), nifedipine (10 nM) and diltiazem (100 nM) shifted the calcium-induced contraction curves to the right, but prazosin (10 nM) had no effect, indicating that it is not a calcium antagonist. The calcium antagonists in the concentrations stated above had no effect on phenylephrine-induced contractions of rat aortic strips in normal Krebs-Henseleit solution, indicating that they did not block alpha 1-adrenoceptors in these concentrations. Taken together, these data suggest that the positive chronotropic effect of phenylephrine resulting from activation of alpha 1-adrenoceptors involves an increased influx of calcium through channels that are sensitive to organic calcium antagonists.     

Influence of desipramine on the contractile responses to noradrenaline, clonidine and to electrical field stimulation in the rat isolated seminal vesicle

The effect of desipramine (DMI) on the contractile responses to noradrenaline (NA), clonidine and to electrical field stimulation (FS), was studied in the rat isolated seminal vesicle. The aim was to analyse the mechanism of action of DMI in this preparation and compare the results obtained with those already published for other similar preparations, e.g. the guinea-pig seminal vesicle. The results showed that DMI, which on its own had little effect on contractility of rat seminal vesicle, significantly (P less than 0.001) enhanced the Na-induced contractions (by 30-50%), whereas it markedly reduced (by 40%) the contractions produced by clonidine. DMI also enhanced the FS-induced contractions (by about 50%) in the same preparation. The enhancement of the NA-induced responses by DMI can be explained in terms of inhibition of the NA uptake by the preparation, whereas the reduction in clonidine response may be due to blockade of adrenoceptors in the rat seminal vesicle. Other possibilities and mechanisms are also discussed in this and other types of smooth muscle preparations.     

Relationship of blood pressure to the responsiveness of an isolated human artery to selected agonists and to electrical stimulation

Intrinsic vascular responsiveness to norepinephrine, transmural electrical stimulation, 5-hydroxy-tryptamine, and vasopressin was studied in isolated helical cut strips of cystic artery (downstream branch of hepatic artery) from 120 subjects and related to blood pressure. Blood pressure, thickness of the tunica media, and passive elastic properties of arterial strips were each significantly correlated with age. With the exception of blood pressure in the female subjects, it is doubtful that these relationships are of major biologic significance. Nevertheless, in subgroup formation, care was taken to control for age. Hypertension was arbitrarily defined in three different ways as: (a) two diastolic pressure measurements greater than or equal to 90 mm Hg (HT90); (b) two diastolic pressure measurements greater than or equal to 95 mm Hg (HT95); or (c) treatment for hypertension instituted by a physician 6 months to 2 years after arteries were studied (HTQ). In arteries of hypertensive female subjects, responsiveness to norepinephrine (and possibly to 5-hydroxytryptamine) was increased significantly over the first half of the dose-response curve, particularly in the arteries of HT95 and HTQ subjects. Responses to transmural electrical stimulation and vasopressin were not consistently different. Such differences were not seen in arteries of male subjects where, if anything, responsiveness to norepinephrine (but not 5-hydroxytryptamine) was decreased. The present observations were made in the absence of any substantive difference in arterial dimensions (e.g., cross-section area) or in the maximal response to norepinephrine. The data support the notion that, at least for female subjects, alteration in intrinsic vascular responsiveness may play a role in the pathogenesis of human essential hypertension.