Isolated Rat Atria Research Articles

Bicuculline actions on isolated rat atria, mouse vas-deferens and guinea-pig ileum are unrelated to GABA A receptor blockade

1. 1. Some new pharmacological activities of bicuculline were found in isolated rat atria, mouse vas deferens and guinea-pig ileum. 2. 2. In isolated rat atria bicuculline (10–300 μM) induced potent positive inotropic and negative chronotropic effects which were not antagonized by propranolol (1 μM), 6-hydroxydopamine pretreatment (50 mg/kg i.v. twice), ranitidine (3 μM) or atropine (1 μM). Bicuculline (10–300 μM) potentiated electrically evoked contractions in mouse vas deferens and inhibited them (30–500 μM) in guinea-pig ileum. It was inactive on unstimulated mouse vas deferens. 3. 3. The above effects were completely reproduced by the bicuculline related-substance, β-hydrastine, but not by the bicuculline N-methyl derivative, bicuculline methiodide (BMI), on the isolated rat atria. BMI inhibited instead of potentiating the mouse vas deferens twitches and potentiated instead of inhibiting the guinea-pig ileum twitches. 4. 4. Picrotoxin, the other classic non-competitive GABA A antagonist, was completely devoid of the effects reported for bicuculline. 5. 5. We concluded that, on the three preparations studied, bicuculline possesses some effects which are unrelated to its GABA A receptor blocking activity.

Aging: Effects on chronotropic actions of muscarinic agonists in isolated rat atria

Negative chronotropic effects of acetylcholine and carbachol were compared in right atria isolated from adult and aged Fischer 344 rats. Preparations from aged rats were more sensitive to the action of acetylcholine; however, there was no difference in responsiveness to carbachol or to acetylcholine after pretreatment with diisopropylfluorophosphate, an irreversible cholinesterase inhibitor. These data suggest that the enhanced sensitivity to the direct chronotropic action of acetylcholine is the result of aging-related reductions in acetylcholinestrase activity.

A New Approach to the Direct Detection of Free Radicals in the Intact Myocardium

A new method for the direct ESR detection of free radicals in rat myocardial tissue is described. Isolated rat atria are continuously monitored for heart rate and contractile force; at the end of the experimental period the beating organs are inserted into quartz ESR tubes and immediately frozen in liquid nitrogen. Spectra obtained from these preparations show the presence of very weak radical signals. When ESR spectra are recorded on samples obtained from pools of rat atria pulverized under liquid nitrogen, the radical lines are markedly stronger than those observed for intact organs; contaminating metals are also frequently detected. These findings indicate that crushing or grinding procedures carried out under liquid nitrogen produce artifactual ESR active species. The new method described in the present paper does not involve mechanical interventions and therefore should yield reliable artifact-free results.

Experimental studies on the acute cardiovascular toxicity of formalin and its antidotal treatment

Rats anesthetized with pentobarbital and ventilated artificially were infused with 0.01 ml formalin (= 0.12 mmol formaldehyde)/kg.min. They exhibited a sharp decline of arterial blood pressure, heart rate and peripheral resistance and a slower one of cardiac output and died after 59.9 +/- 6.0 min of infusion. Sinus bradycardia and, in some cases, AV-arrhythmia occurred in the ECG. The additional infusion with cysteine attenuated the cardiovascular failure and more than doubled the survival time of formalin-infused rats. Infusion of N-acetylcysteine or correction of formalin-induced metabolic acidosis with sodium bicarbonate, on the other hand, did not exert antidotal activity. On isolated rat atria in vitro, formalin decreased the rate and the contractility and cysteine antagonized these effects of formalin. In conclusion, the severe and often lethal incidents observed following the therapeutic administration of formalin are due to the cardiovascular-depressive activity of formaldehyde and may be antagonized by cysteine.

Effect of Glutathione and N-Acetylcysteine on in Vitro and in VIVO Cardiac Toxicity of Doxorubicin

The effects of two sulfhydryl compounds, glutathione (GSH) and N-acetylcysteine (NAC), on the cardiotoxicity of doxorubicin (DXR) were tested on in vitro and in vivo models. DXR was administered to rats as 4 weekly i.v. doses of 3 mg/kg. GSH (1.5 mmoles/kg), given i.v. 10 min before and 1 hr after DXR, was found to prevent the development of the delayed cardiotoxic effects of DXR, as assessed by electrocardiographic and mechanical parameters, as well as by histological examination of left ventricular preparations. In contrast, equimolar oral doses of NAC (1 hr before and 2 hrs after DXR) were found to be ineffective. Both GSH and NAC prevented the negative inotropic effect produced by DXR on isolated rat atria. A good correlation exists between the cardioprotective effects of the two agents and their ability to enhance the non-protein sulfhydryl group content of the myocardium. Differences observed in vivo between GSH and NAC might be accounted for by pharmacokinetic factors.

Cardiotoxicity and antitumor activity of a copper(II)-doxorubicin chelate

The cardiotoxic and cytotoxic effects of the Cu(II)-doxorubicin (DXR) complex [Cu(DXR)]n are compared with those of the parent drug. It is shown that 10(-4) M [Cu(DXR)]n has no depressant effects on isolated rat atria, in contrast with an equimolar concentration of the parent drug. No differences were found between the cytotoxic activities of the Cu(II) complex and free DXR on B16 melanoma and HeLa cells. A reduced penetration of the polymeric [Cu(DXR)]n into the myocardial cells as compared with the free drug was invoked to account for the absence of cardiotoxicity of the DXR complex. On the other hand, the observation that copper-complexation does not affect the cytotoxicity of the drug suggests that extracellular as well as intracellular mechanisms may be involved in the development of its antitumor activity.

Alinidine antagonizes the myocardial effects of adenosine

The bradycardiac drug alinidine (2.8–182.3 μM) antagonized the negative inotropic effect of adenosine in isolated left rat atria in a non-competitive fashion. A 50% reduction of the maximal adenosine effect was achieved with 64.6 μM alinidine. The shortening of the action potential duration and the suppression of slow action potentials in guinea pig atria by 1–10 μM adenosine could be abolished by 5.7 μM alinidine. Adenosine (1 mM) exerted a negative inotropic effect on isolated strips from the right ventricle of rat heart exposed to 0.36 μM isoproterenol, an effect which was fully antagonized by 91.1 μM alinidine. Addition of 1 mM adenosine to spontaneously beating isolated rat atria induced pronounced bradycardia. Under these conditions, the bradycardiac agent alinidine increased the contractile rate concentration dependently (11.4–182.3 μM), while the chemically unrelated bradycardiac compound UL-FS 49 did not accelerate the beat frequency. As alinidine antagonized all the cardiac actions of adenosine that we tested, it may be concluded that the drug interferes with A 1-receptor-mediated effects.

Alinidine reverses the descending staircase of isolated rat atria by an antimuscarinic action

Contractile force of isolated atria from most mammalian species increases with the rate of electrical stimulation, resulting in an ascending staircase. In contrast, in the rat, contractile force decreases with increasing rate of stimulation (descending staircase). The bradycardic and antianginal drug alinidine (5.7-91.2 mumol/l) reversed the descending staircase to ascending by a positive inotropic effect at higher stimulation rates. Maximal positive inotropy was obtained with 45.6 mumol/l, a concentration which also caused maximal bradycardia in spontaneously beating atria. Concentrations of 1 mumol/l of the antimuscarinic compounds atropine as well as the quaternary salt ipratropium bromide also reversed the descending staircase of rat atria. Addition of alinidine did not cause any further increase in force of contraction under these conditions. Addition of 1 mumol/l physostigmine to isolated left atria from guinea pigs for blockade of acetylcholinesterase decreased contractility at all stimulation rates, but did not change the ascending character of the staircase. Alinidine antagonized the negative inotropic effect of physostigmine. The known antimuscarinic action of alinidine was quantified in electrically driven (0.25 Hz) left rat atria by antagonism of the negative inotropic effect of oxotremorine (0.01-10 mumol/l). Alinidine acted as a strictly competitive antagonist with a pA2 of 5.82. In isolated papillary muscle from guinea pigs, pretreated with reserpine for depletion of catecholamines, carbachol (0.1-3000 mumol/l) exerted positive inotropic effects. Alinidine antagonized also this effect in a competitive fashion with a pA2 value of 5.58.(ABSTRACT TRUNCATED AT 250 WORDS)

Alpha-adrenoceptor-mediated chronotropic response: influence of temperature and basal spontaneous rate

The chronotropic response to phenylephrine of isolated rat atria was determined at 37 degrees C and 30 degrees C. At both temperatures similar maximal increases in rate were obtained (91 +/- 4 beats/min and 93 +/- 8 beats/min, respectively). Beta-adrenoceptor blockade (10(-6) M propranolol) abolished the chronotropic effect of phenylephrine at 37 degrees C whereas at 30 degrees C a propranolol-resistant chronotropic effect was detected. The propranolol-resistant chronotropic effect of phenylephrine was sensitive to prazosin blockade (10(-6) M) indicating alpha adrenoceptors involvement in the response. At 37 degrees C a propranolol-resistant, prazosin-sensitive effect of phenylephrine could be obtained if basal rate was lowered (by cholinergic stimulation or acidosis) or if extracellular calcium was increased over 1.35 mM. When the chronotropic effect of phenylephrine was compared at similar basal rates, it was greater at 37 degrees C than at 30 degrees C. These results make unlikely the possibility of increases in sensitivity or in density of alpha-adrenoceptors at low temperature. The fact that alpha-adrenoceptor mediated chronotropic response is enhanced at low basal frequencies makes this mechanism a potential safety factor during bradycardia.

Endothelin stimulates release of atrial natriuretic peptides in vitro and in vivo

The effect of endothelin (END) on the release of atrial natriuretic peptides (ANP) was studied in isolated rat atria and in conscious rats. END stimulates the ANP release in vitro in a dose-dependent manner. An increase in ANP plasma levels and cyclic GMP plasma levels was also observed in conscious rats after injection of END. When a monoclonal antibody directed against ANP was injected together with END the increase in cyclic GMP was completely blocked. From this study it is concluded that END is a potent secretagogue for ANP both in vitro and in vivo.

Dynorphin stimulates the release of ANP from isolated rat atria

La dynorphine, peptide opioide endogene, augmente la concentration plasmatique en peptide natriuretique atrial. In vitro, la mesure de la liberation d'ANP a partir d'un atrium de rat isole confirme l'activite stimulatrice directe de la dynorphine sur la secretion

Effects of thyroid status on atrial natriuretic peptide release from isolated rat atria

The effect of thyroid hormone on circulating levels of atrial natriuretic peptide (ANP) was studied in experimental hyperthyroid and hypothyroid rats. Plasma ANP was 102 +/- 5 pg/ml in euthyroid rats, 82 +/- 4 pg/ml in hypothyroid rats, and 138 +/- 11 pg/ml in hyperthyroid rats. We have also measured immunoreactive ANP in the atria of euthyroid, hypothyroid, and hyperthyroid rats. ANP content and concentration in the atria were lower (546 +/- 32 pg/mg tissue) in hyperthyroid rats than in hypothyroid rats (802 +/- 74 pg/mg tissue). Right atrium from euthyroid, hypothyroid, and hyperthyroid rats was superfused with a modified Langendorff preparation. Spontaneous release of ANP was significantly higher from the hyperthyroid rats (20 +/- 2 pg.min-1.mg-1) than from the hypothyroid rats (5.6 +/- 0.5 pg.min-1.mg-1). ANP release from the euthyroid rats was 9.3 +/- 1.2 pg.min-1.mg-1. These results indicate hyperthyroidism causes an increase in ANP secretion and a decreased release occurs during hypothyroidism.

The Effects of 4-Pentenoic and Pentanoic Acids on the Hypoxic Rat Atria

When exposed to hypoxia, the isolated rat atria released lactate into the bathing medium and underwent a rise in resting tension and a decline of the contractions frequency. In some of them, it also occurred a complete cessation of the pacemaker activity. Atria from 24-h fasted rats, when compared to those from fed ones, exhibited a lower lactate output, a higher rise in resting tension, a faster decay of the contraction frequency and an increased occurrence of atrial arrest. In both the fed and fasted rats atria, some triacylglycerol lipolysis remained throughout the hypoxic incubation. Addition of 2 mM 4-pentenoic acid abolished the lipolytic activity and reduced lactate output in both groups of atria. In the fed rats atria it also accelerated the decrease of the pacemaker frequency. Pentanoic acid reduced lactate output in both groups of atria and in those from fed rats it did not alter lipolysis but increased the rise in resting tension, the decline of the pacemaker frequency and the occurrence of atrial arrest. Present data indicate that although 4-pentenoic acid inhibits fatty acid oxidation and endogenous lipolysis, it was not able to reduce the noxious effects of hypoxia. Since the effects of 4-pentenoic acid were rather similar to those of fasting and pentanoic acid, they might be ascribed to the accumulation of its own oxidative metabolites which could be detrimental for the hypoxic atria.

Effects of calcium and ouabain on the release of atrial natriuretic factor.

In order to examine the role of calcium in the secretory process of atrial natriuretic factor (ANF), we studied the effects of hypercalcaemia and ouabain on the plasma concentration of immunoreactive ANF, and the effect of calcium on immunoreactive ANF release from isolated rat atria. Anaesthetized dogs were treated with CaCl2 infusion, ouabain or phenylephrine injection. With CaCl2 infusion, serum calcium and plasma immunoreactive ANF respectively increased to three and four times their basal levels. Ouabain increased plasma immunoreactive ANF to two and a half times the initial level. Neither CaCl2 nor ouabain produced any effect on right atrial pressure and heart rate, but they both significantly increased arterial pressure. Phenylephrine caused a greater increase in arterial pressure than both CaCl2 and ouabain. However, there was no significant increase in plasma immunoreactive ANF. Moreover, calcium stimulated the release of immunoreactive ANF from isolated rat atria. These results suggest that the calcium may play a key role in the secretory process of ANF.

Free creatine available to the creatine phosphate energy shuttle in isolated rat atria.

To measure the actual percentage of intracellular free creatine participating in the process of energy transport, the incorporation of [1-14C]creatine into the "free" creatine and phosphocreatine (PCr) pools in spontaneously beating isolated rat atria, under various conditions, was examined. The atria were subjected to three consecutive periods, control, anoxia, and postanoxic recovery, in medium containing tracers of [1-14C]creatine. The tissue content and specific activity of creatine and PCr were determined at the end of each period. The higher specific activity found for tissue PCr (1.87 times) than creatine, independent of the percentage of total intracellular creatine that was present as free creatine, provides evidence for the existence of two separate pools of free creatine. Analysis of the data shows that in the normal oxygenated state approximately equal to 9% of the total intracellular creatine is actually free to participate in the process of energy transport (shuttle pool). About 36% of the total creatine is bound to unknown intracellular components and the rest exists as PCr. The creatine that was taken up and the creatine that was released from the breakdown of PCr have much greater access to the site of phosphorylation than the rest of the intracellular creatine. A sharp increase in the specific activity of residual PCr on prolongation of anoxic time was also observed. This provides evidence for a nonhomogeneous pool of PCr, for the most recently formed (radioactive) PCr appeared to be hydrolyzed last.

Calcitonin gene-related peptide(CGRP) stimulates the release of atrial natriuretic peptide(ANP) from isolated rat atria

The effect of calcitonin gene-related peptide(CGRP) on the release of atrial natriuretic peptide(ANP) was studied in spontaneously beating, isolated rat atria. CGRP stimulated the ANP release in a dose-dependent manner. When the atria were incubated with a combination of phentolamine, propranolol, and atropine, these antagonists blocked neither the rise in ANP release nor the positive chronotropic and inotropic effects of CGRP. Therefore, we conclude that CGRP stimulates ANP release as well as cardiac contractility independently of adrenergic and cholinergic receptors.

Effects of piperine on calcitonin gene-related peptide (CGRP)-containing nerves in the isolated rat atria

Piperine, a major pungent agent of black peppers, induced both positive chronotropic and inotropic responses in the isolated atria of rats. These responses were not blocked by various antagonists but exhibited rapid tachyphylaxis. After exposure to piperine in vitro, endogenous calcitonin gene-related peptide (CGRP) was profoundly depleted from intracardiac nerves. These results suggest that piperine releases endogenous CGRP from intracardiac non-adrenergic non-cholinergic nerves and that released CGRP exerts positive chronotropic and inotropic effects.

Class III Antiarrhythrnic Action Linked with Positive Inotropy: Effects of the d‐ and 1‐Isomer of Sotalol on Isolated Rat Atria at Threshold and Suprathreshold Stimulation

The beta-adrenoceptor blocker sotalol has been shown to possess class III antiarrhythmic action. The present study was designed to test the hypothesis that class III antiarrhythmic action and positive inotropy may be linked. Since d,l-sotalol has previously been reported to have variable inotropic effect, we studied direct effects of the d- and l-stereoisomer of sotalol on refractoriness and inotropy of isolated rat atria at different strengths of electrical stimulation. Both the d- and l-stereoisomer of sotalol (7.5 x 10(-5) M) increased the effective refractory period, and to the same extent. However, d-sotalol increased isometric contractile force by 10%, while l-sotalol had no significant effect, when the atria were stimulated at close to threshold values (10 +/- 1 mA). Probably due to release of noradrenaline within the myocardium, contractile force increased when the atria were stimulated at suprathreshold values (40 mA). At suprathreshold stimulation, however, d-sotalol induced a 10% decrease and l-sotalol a 20% decrease, in contractile force. In conclusion, the class III antiarrhythmic action of sotalol is linked with positive inotropy. In the presence of neurotransmitter release, negative inotropic effect of d- and in particular l-sotalol, may occur due to beta-adrenoceptor blocking activity.

Effects of magnesium depletion on myocardial high-energy phosphates and contractility

The effect of prolonged magnesium depletion on contractility, phosphorylating activity, and organic phosphates of spontaneously beating isolated rat atria was studied. Rats were fed a Mg-deficient diet for 8 weeks, during which serum Mg fell from 1.85 ± 0.02 to 0.52 ± 0.10 mg/dl. Atrial contractile activity was measured for 1 hr and at the end of this period tissue samples were taken for the determination of the phosphorylated intermediates. Mg depletion was associated with (a) reduced intracellular inorganic phosphorus and adenine nucleotides; (b) elevated creatine phosphate; (c) reduction in contractile force (CF) with no change in atrial beat rate (BR). There were no significant differences in the activities of creatine phosphokinase and adenylate kinase in control and Mg-depleted rat atrial homogenates determined in the presence of 5 m m MgCl 2. Addition of various concentrations of MgCl 2 to the medium resulted in an immediate reduction in both CF and BR of normal and Mg-depleted rat atria. Intraperitoneal administration of MgCl 2 to Mg-depleted rats resulted in complete recovery of CF of isolated atria. This improvement in CF occurred without changes in the levels of inorganic phosphate and adenine nucleotides. The reduced intracellular level of high-energy phosphate or inorganic phosphate cannot therefore be responsible for the impaired contractility seen in Mg-depleted heart muscle. On the other hand, the fact that the creatine phosphate levels were higher in magnesium depletion suggests that myofibrillar utilization of creatine phosphate is more impaired than production, analogous to phenomena seen in postanoxic recovery.

Interleukin 2 stimulates heart contractility in the presence of exogenous arachidonate or the calcium ionophore A 23187

An increase in the isometric developed tension (IDT) of isolated rat atria was observed shortly after the addition of human interleukin 2 (IL-2) to the organ preparation with subthreshold concentrations of either arachidonate (AA, 1.98 × 10 −6 M) or the calcium ionophore A 23187 (1.9 × 10 −6 M). Both natural purified IL-2 (nIL-2) and yeast recombinant IL-2 (rIL-2) were active in this experimental system. It was determined that this lymphokine was active at 2 × 10 −11 M, considering as a reference the specific activity of rIL-2. Anti-IL-2 monoclonal antibody (anti-IL-2 MAb) abolished this reaction. Inhibition of atrial phospholipase C activity by nitrocarboxyphenyl N,N-diphenylcarbamate (NCDC, 5 × 10 −6 M) prevented the development of the inotropic positive effect of IL-2 in the presence of either AA or A 23187. The synthetic diacylglyceride 1-oleoyl, 2-acetyl-glycerol (OAG) replaced the IL-2 as stimulatory signal but NCDC had no effect on the reaction. The results suggest that IL-2 can alter the physiologic behaviour of the heart and that its mechanism of action is probably similar to the one proposed for other IL-2 targets (IL-2 receptor-positive T lymphocytes, T cell lines).