Node Preparations Research Articles

Effect of ischaemia on overdrive suppression in isolated, blood-perfused atrial preparations of dogs.

The magnitude of overdrive suppression of the SA nodal pacemaker activity was markedly influenced by its ischaemia which was produced by occlusion of the perfusion tubing close to the sinus node artery in the in-situ and isolated canine sino-atrial node preparations. In all isolated preparations, postdrive suppression was significantly augmented after occlusion of the perfusion system for at least 120 seconds. It is concluded that total ischaemia depresses sino-atrial node function in the atropinized canine heart and that postdrive suppression is a good index of such impaired function.

Effects of atrio-ventricular conduction of calcium-antagonistic coronary vasodilators, local anaesthetics and quinidine injected into the posterior and the anterior septal artery of the atrio-ventricular node preparation of the dog.

The effects on atrio-ventricular (A-V) conduction and blood flow of calcium-antagonists (verapamil, nifedipine and diltiazem), local anaesthetics (procaine and lidocaine) and quinidine were investigated in the isolated, cross-circulated A-V node preparation of the dog. The drugs were injected individually into the posterior septal artery (PSA) through which the upper part of the A-V node is mainly perfused or into the anterior septal artery (ASA) through which the lower part of the node and the more distal conduction system are perfused. Single injections into the PSA of nifedipine (0.3-10 mug), verapamil (1-30 mug), diltiazem (1-30 mug), quinidine (30-300 mug), lidocaine (100 mug-1 mg) and procaine (300 mug-3 mg) produced a dose-related increase in the A-V conduction time and with higher doses of these drugs a second or third degree block of A-V conduction occurred. Nifedipine (0.3-30 mug) and verapamil (1-100 mug) injected into the ASA scarcely affected A-V conduction. Quinidine (30 mug-1 mg) and lidocaine (100 mug-3 mg) injected into the ASA prolonged the A-V conduction time in a dose-related manner, although the effects were less prominent than those produced upon injection into the PSA. High doses of quinidine (3 mg) and lidocaine (3-10 mg) injected into the ASA altered the shape of ventricular bipolar electrograms and prolonged the time interval between an electrogram of the right bundle branch and that of the ventricle. The results are consistent with the hypothesis that in excitation of A-V nodal cells a slow calcium current rather than a fast sodium current plays an important role and that in the His-Purkinje-ventricular system the fast sodium current is predominant. Single injections of the 6 drugs into the PSA produced a dose-related increase in blood flow through the PSA. All drugs but nifedipine increased the blood flow in almost the same dose range that caused impairment of A-V conduction. Nifedipine was 10 times more potent in increasing the blood flow than in impairing A-V conduction.

Modification by manganese ions and verapamil of the responses of the atrioventricular node to norepinephrine

In isolated and cross-circulated canine A–V node preparations single injections of Mn 2+ (1–30 μmol) and verapamil (0.3–10 nmol) into the posterior septal artery (PSA) caused a dose-dependent increase in the A–V conduction time and, in large doses, produced A–V conduction block. The A–V conduction time increased by Mn 2+ and verapamil is restored by 1-norepinephrine (1-NE) injected into the PSA. The A–V nodal tachycardia induced by 1-NE is suppressed by Mn 2+ but not by verapamil. These indicate the following: (1) the slow Ca 2+ channel plays an important role in A–V conduction; (2) the positive dromotropic response of the A–V node to 1-NE may have some relations to the Ca 2+ channel; (3) the A–V nodal tachycardia induced by 1-NE may be mediated by the Ca 2+ channel. Possible causes which produced differences between Mn 2+ and verapamil in the effects in antagonizing the 1-NE-induced A–V nodal tachycardia are discussed.

Effects of fusaric acid and its derivative on the cardiovascular system

The cardiovascular effects of fusaric acid, a dopamine-beta-hydroxylase (DBH) inhibitor and a hypotensive agent, and 5-(4'-chlorobutyl) picolinic acid, one of the most potent DBH inhibitors in fusaric acid derivatives, were investigated in anesthetized dogs. Fusaric acid (10-30 mg/kg) given intravenously caused immediately and dose-dependently a fall in blood pressure, an increase in heart rate, a stimulation of respiration, a marked increase in the rate of superior mesenteric arterial flow, and a decrease in the rate of femoral arterial flow. Fusaric acid (0.3-3 mg) given close-arterially caused a dose-dependent decrease in the sinus rate and in the developed tension in isolated blood-perfused sinoatrial node and papillary muscle preparations, respectively. For the superior mesenteric, renal, and femoral circulations which were perfused with blood, a close-arterial injection of fusaric acid (10-30 mg) increased in the rates of flow in a dose-dependent manner. 5-(4'-Chlorobutyl) picolinic acid showed similar responses as fusaric acid quantitatively or qualitatively. These results indicate that hypotension induced rapidly after intravenous administration of fusaric acid or 5-(4'-chlorobutyl) picolinic acid is not due to the enzyme inhibition, but rather to the direct depression of cardiac function and decrease in peripheral vascular resistance.

COMPARISON OF NEWLY SYNTHESIZED β-ADRENERGIC BLOCKERS, OPC 1085 AND SQ 11725, WITH PINDOLOL AND PROPRANOLOL IN THE BLOOD-PERFUSED CANINE SA NODE AND PAPILLARY MUSCLE PREPARATIONS

COMPARISON OF NEWLY SYNTHESIZED β-ADRENERGIC BLOCKERS, OPC 1085 AND SQ 11725, WITH PINDOLOL AND PROPRANOLOL IN THE BLOOD-PERFUSED CANINE SA NODE AND PAPILLARY MUSCLE PREPARATIONS

Effects of dopamine on sinus rate and ventricular contractile force of the dog heart in vitro and in vivo.

1 Experiments were carried out on dog isolated papillary muscle and sinus node preparations perfused with arterial blood from a donor dog. The chronotropic and inotropic effects of dopamine were analysed by using reserpine, desmethylimipramine (DMI), cocaine and phenoxybenzamine, and the relative chronotropic and inotropic effects defined, the results being compared with those for noradrenaline (NA). 2 The effects of dopamine administered intra-arterially into the isolated preparations were reduced by pretreatment of animals with reserpine both in the papillary muscle and sinus node. The chronotropic effects, however, were affected less by pretreatment with reserpine than were the inotropic effects. 3 Desmethylimipramine (DMI) reduced the inotropic and chronotropic effects of dopamine, and enhanced the effects of NA and nerve stimulation; the chronotropic effects of the amines were less affected than the inotropic effects. 4 Cocaine enhanced considerably the inotropic and chronotropic effects of NA, and decreased the inotropic but not the chronotropic effect of dopamine. 5 Phenoxybenzamine enhanced the inotropic effects of dopamine, NA and nerve stimulation, but did not affect the chronotropic effects of the amines. 6 When dopamine (1 to 300 mug/kg) was administered intravenously to the donor dog, it increased preferentially the contractile force of the ventricular myocardium with a comparatively small change of the sinus rate in the isolated preparations as well as in the heart in vivo. NA (0.1 to 10 mug/kg) caused effects similar to those of dopamine. The maximal inotropic responses to these catecholamines were reached with lower doses than the chronotropic ones. 7 It is concluded that both the positive inotropic and the positive chronotropic responses to dopamine are mediated partially by a direct and partially by an indirect stimulant effect on beta-adrenoceptors in the dog heart. The present results suggest that the difference in activity of dopamine and NA between the ventricular myocardium and the sinus node may be ascribed to the unequal innervation with adrenergic nerve fibres of the atrium and the ventricle (Furnival, Linden & Snow, 1971). The sinus node which is densely innervated by adrenergic nerve fibres may inactivate noradrenaline and dopamine more effectively than the ventricular myocardium through the uptake into the nerve and thereby be less sensitive to the exogenous catecholamines.

Effects of neurotransmitters injected into the posterior and the anterior septal artery on the automaticity of the atrioventricular junctional area of the dog heart.

The effects of neurotransmitters injected into the posterior and the anterior septal artery on the automaticity of the atrioventricular (AV) junctional area were examined in the excised, blood-perfused canine AV node preparation. An AV rhythm of 57.4 +/- 2.8 beats/min (N = 28) developed after the destruction of the sinoatrial node. l-Norepinephrine injected into the anterior septal artery increased the frequency of AV rhythm, but l-norepinephrine injected into the posterior septal artery caused a pacemaker shift from the anterior septal artery area to the posterior septal artery area. Acetylcholine injected into the posterior septal artery blocked retrograde conduction but did not decrease the frequency of AV rhythm; however, acetylcholine injected into the anterior septal artery decreased AV rhythm but did not affect retrograde conduction. Destruction of the posterior septal artery area failed to change AV rhythm. The surgically separated anterior septal artery preparation had the same rate as did the AV node preparation. The posterior septal artery preparation had a definitely lower rate and responded to smaller doses of l-norepinephrine than did the AV node preparation. The results of the present study indicate that AV rhythm in the AV node originates in the area supplied by the anterior septal artery and that the area supplied by the posterior septal artery, which has extremely low automaticity, is highly responsive to l-norepinephrine, resulting in nodal tachycardia.

Effects of nifedipine, a potent calcium-antagonistic coronary vasodilator, on atrioventricular conduction and blood flow in the isolated atrioventricular node preparation of the dog

The effects of the calcium-antagonist nifedipine on atrioventricular (A-V) conduction and blood flow were investigated in comparison with those of verapamil by the use of the isolated, arterially blood-perfused A-V node preparation of the dog. Single injections of 0.3--30 mug of nifedipine and verapamil into the A-V node artery produced a dose-related increase in the A-V conduction time and at 30 mug the two drugs caused second degree block of A-V conduction. The results are compatible with the hypothesis that a slow calcium channel plays an important role in excitation of A-V nodal cells. The rate of blood flow through the A-V node artery was about 10 times more sensitive to nifedipine than was the A-V conduction time and increased in a dose-related manner. In contrast, an increase in blood flow rate by verapamil occurred in almost the same dose range as did impairment of A-V conduction. This indicates that the action of nifedipine is more pronounced on coronary smooth muscle cells than on the A-V nodal cells.

Effect of apomorphine on the canine SA node.

A constant pressure perfusion of the sinus node artery was performed in 10 in situ and in 3 isolated SA node preparations. The selective administration of apomorphine caused a negative chronotropic effect which was not blocked by treatment with atropine. The sinus deceleration was usually produced by apomorphine, 10-300 mug, while sinus irregularity and finally atrioventricular nodal rhythm in the in situ preparation or sinus arrest in the isolated one resulted from doses above 1 mg. In the in situ preparation at a higher dose from 300 mug to 1 mg, a slight sinus acceleration was occasionally induced following sinus deceleration; In this time, a fall in blood pressure was usually observed; This acceleration response to apomorphine was inhibited by propranolol. From these results, it is concluded that apomorphine induces direct depressive action on the SA node and, in extremely high doses, a slight sinus acceleration may be induced by reflex mechanism.

Adenylate cyclase activity in lymphoid organs during regional and systemic graft-versus-host reactions

In order to gain further information concerning the pathogenesis of graft-versus-host disease, the activity of adenylate cyclase (EC 4.6.1.1) was studied in lymphoid and non-lymphoidal organs of animals undergoing graft-versus-host reaction. A marked increase in the adenylate cyclase activity was observed in the washed cell particles isolated from lymph nodes and spleen of animals undergoing regional and systemic graft-versus-host reactions, respectively. Isoproterenol was found to stimulate the enzyme in all the control and experimental preparations; however, NaF failed to stimulate the enzyme in preparations obtained from lymph nodes of rats undergoing regional graft-versus-host reaction. The elevated adenylate cyclase activity in lymph node preparations from rats undergoing graft-versus-host reaction was found to be associated with a decrease in K m value without any significant change in the value for V. The adenylate cyclase activities in the non-lymphoidal organs such as heart, kidney and brain from animals undergoing graft-versus-host reaction were not significantly different from the control values. It is suggested that the observed activation of adenylate cyclase during graft-versus-host reaction may be a direct consequence of cell-cell interaction.

Effect of desmethylimipramine on the canine SA node.

Direct perfusion of the sinus node artery at constant pressure of 100 mmHg was performed in eleven canine hearts in situ. The injection of desmethylimipramine (DMI) into the sinus node artery caused an initial short deceleration followed by markedly long-lasting acceleration of the heart rate. The initial deceleration was not influenced by treatment with atropine. The secondary acceleration was inhibited by propranolol, and, furthermore, suppressed by tetrodotoxin. After the treatment with propranolol or tetrodotoxin, DMI produced only a negative chronotropic effect dose-relatedly. It was difficult to observe the blocking effect of DMI on tyramine action, because DMI alone induced marked sinus acceleration. On the other hand, tyramine action was not influenced by tetrodotoxin. Therefore, the use of both DMI and tetrodotoxin is beneficial to observe blocking effect of DMI on tyramine-like action in the in situ SA node preparation.

Intracellular stimulation and recording from single cardiac cells in the rabbit atrium

The mechanism of pacemaker action in an isolated rabbit sinoatrial node preparation was studied by means of intracellular stimulation and recording through a single-barrelled microelectrode. This technique permitted not only stimulation of individual cells of the various types found in and around the pacemaker tissue but also the recording of nonpropagated activity in cells stimulated or in those in the immediate vicinity. It was found that: 1) The myocardial atrial fiber has a threshold of the order of 10 −6 A. when stimulated intracellulary with a 15 msec duration pulse. This threshold was constant throughout diastole, except during its very early phase. The resulting response was of an all-or-none type and propagated over the tissue. 2) Thresholds for latent pacemaker cells were different from cell to cell; within a cell, it also varied slightly with the degree of diastolic depolarization attained. The response initiated by intracellular stimulation was also of an all-or-none type and, in most cases, it was propagated to the atrium. When a response was induced at an early phase in diastole, a conduction failure was sometimes observed. When such a nonpropagated response occurred in a latent pacemaker cell, the duration of diastole was increased. 3) Most pacemaker cells tested did not respond to intracellular stimulation. In a few cells, however, a graded response proportional to stimulus intensity, was obtained. Such response did not propagate to the atrium. These graded responses differed from the local responses which occur in other types of cell. It was concluded that a synchronization of pacemaker cell activity is required for effective pacemaker action.

Positive chronotropic effect, of ouabain in the excised and blood-perfused canine SA node preparation of the dog

The effect of ouabain on the sinus rate was investigated in excised and blood-perfused canine SA node preparations which had 101±3 regular beats/min (mean±S.E., N=46) and 3.2±0.3 ml/min of blood flow through the sinus node artery. When ouabain was rapidly injected in a wide range of doses from 0.3 to 30 μg into the sinus node artery, it caused a dose-dependent acceleration of the regular sinus rate without any latency. The positive chronotropic response to ouabain was not blocked by tetrodotoxin, β-adrenoceptor blocking agents such as propranolol and alprenolol, or pretreatment with reserpine. These observations suggest that the ouabain-induced acceleration is not mediated through adrenergic mechanism. Although a decrease of sinus rate by 7.3±2.3% (mean±S.E., N=6) of the control level was induced by MnCl2 infusion at a rate of 30 or 60 μg/min, the acceleration response to ouabain was selectively blocked by MnCl2 while that to noradrenaline was not modified. These results suggest that ouabain causes an increase of influx of Ca ions across the membrane of the SA nodal pacemaker cell, resulting in the acceleration response, and that this increase of Ca ion influx is probably linked with potassium and/or sodium permeability in the slow diastolic depolarization.

Selective stimulation of the parasympathetic preganglionic nerve fibres in the excised and blood-perfused SA node preparation of the dog.

Experiments were conducted on the canine SA node preparation cross-circulated with the arterial blood from the donor dog through the cannulated sinus node artery. The sinus rate of 98±4 beats/min persisted with a regular rhythm over 10 h. Subthreshold electrical stimulation to the myocardium was applied to various portions of the right atrium. Stimulation of the caval margin caused a simple deceleration of the sinus rate, and a portion slightly anterior to the mid-point between the two caval ostia was most sensitive. Stimulation at the portion where the sinoatrial node was supposed to be caused far less deceleration, while that at the atrial free wall failed to affect sinus rhythm. The negative chronotropic response to subthreshold electrical stimulation was abolished either by injection of hexamethonium or pempidine, or by that of tetrodotoxin or atropine into the sinus node artery, while physostigmine enhanced it. These results indicate that the deceleration response induced by subthreshold electrical stimulation is due to excitation of preganglionic fibres of the parasympathetic nerve. On the other hand, the propranolol-sensitive acceleration did not occur until myocardial excitation was induced by a stronger intensity of stimulation. This suggests that the sympathetic postganglionic fibres have a higher electrical threshold than the parasympathetic preganglionic fibres. These results contradict previous observations in excised atria perfused with physiological salt solution in which hypoxia of the peripheral neurone cannot be avoided.

The electrophysiologic effects of low and high digoxin concentrations on isolated mammalian cardiac tissue: reversal by digoxin-specific antibody.

The effects of digoxin on electrophysiologic properties were evaluated in isolated perfused cardiac tissue. In canine Purkinje fiber (PF)-ventricular muscle (VM) preparations, control measurements, using microelectrode technique, were made of: resting potential (RP), action potential (AP) amplitude, rate of rise, overshoot, duration (APD), membrane responsiveness, conduction velocity (CV), and refractory period. The preparation was then exposed to 1 x 10(-7) M digoxin and repeat measurements were carried out every 15 min. At slow (30/min) rates of stimulation APD initially prolonged then markedly shortened. With more rapid stimulation (75 and 120/min) no initial APD prolongation was observed. When stimulated at 75/min, RP and AP rate of rise, amplitude, and CV remained near control values for 60-75 min then rapidly decreased until electrical inexcitability (110+/-15 min). At that time fibers were perfused with serum containing digoxin-specific antibody (DSA) or one of a group of test solutions. In the preparations exposed to DSA, membrane characteristics improved by 15 min, and by 60 min approximated control values. No beneficial effect was seen with the various test solutions. DSA also reversed digoxin-induced enhanced phase 4 depolarization in PF. Effective (ERP) and Functional (FRP) refractory periods of rabbit atrioventricular (AV) node preparations were measured in the control state. The tissue was then exposed to 1 x 10(-7) M digoxin and refractory period measurements repeated. At a time when AV conduction prolonged by 20%, associated with marked prolongation of ERP and FRP, DSA or various test solutions were perfused. The prolongation in ERP, FRP, and AV conduction time rapidly returned to normal only in the DSA perfused tissue. It is concluded that DSA has the ability to reverse pronounced toxic electrophysiological effects of digoxin in in vitro cardiac tissue.

Competence of thoracic duct cells in the transfer of adjuvant disease and delayed hypersensitivity. Evidence that mycobacterial components are required for the successful transfer of the disease

Transfer of rat adjuvant disease into syngeneic recipients was achieved with lymph node and spleen cells from adjuvant injected donors, but not with washed thoracic duct (TD) cells, despite the fact that these were capable of transferring delayed hypersensitivity (DH) to PPD. Addition of mycobacteria enabled the TD cells to transfer disease. Injection of adjuvant caused accelerated development of disease in the recipients of sensitized TD cells. Since addition of mycobacterial components, probably already present in lymph node and spleen cell preparations, was necessary to transfer disease with sensitized TD cells, it appears unnecessary to invoke an autoimmune or viral etiology for the disease. It is concluded that adjuvant disease results from prolonged DH type reactions to mycobacterial components deposited in the sites of inflammation. TD cells drained late were more efficient than those drained early after cannulation in the transfer of DH to PPD. This was probably due to changes in the proportions of different cell populations present in the TD lymph.

The isolated and cross-circulated AV node preparation of the dog.

The isolated and cross-circulated AV node preparation of the dog.

Lymph Node Cells Carrying Receptor Sites Detected by Diffusion Dialysis Techniques

Abstract Hartley strain guinea pigs were immunized with bovine γ globulin (BGG) using various techniques in order to produce humoral antibodies, delayed hypersensitivity or both humoral antibodies and hypersensitivity to BGG. A diffusion dialysis technique was utilized to demonstrate cell binding of antigen (125I-BGG). Cells contained in regional lymph node preparations from guinea pigs with delayed hypersensitivity to BGG demonstrated high degrees of cell binding of 125I-BGG. Cytophilic antibodies to BGG were also produced in some guinea pigs. The results of these studies fail to demonstrate a relationship between cell binding of antigen associated with the development of delayed hypersensitivity and the presence of cytophilic antibodies.

Negative dromotropic response to phenylephrine blocked by phenoxybenzamine.

In most canine excised and cross-circulated A-V node preparations phenylephrine induced a monophasic long-lasting positive dromotropic effect, and in some a triphasic response, i.e., positive-negativepositive dromotropic one. The negative dromotropic response was abolished by phenoxybenzamine and also blocked by l-hyoscyamine. Then, the triphasic response was converted to a monophasic positive one which was blocked by 1-alprenolol.

Early stimulation of potassium uptake in lymphocytes treated with PHA

even narrower region (density 1.0662 to 1.0687) of high radioactivity. By contrast, the distribution of 14C-labeled (“short-lived”) cells in all the other organs was such that there were relatively more of these cells in the low density range of the gradients than there were total cells in that range. This was particularly pronounced in the lymph node preparation and the bone marrow preparation. Enrichment for “long-lived” cells was seen only in certain portions of the gradient which contained the cells from spleen and in the low-density region of the lymph node gradient. A relatively large fraction of the total cells from lymph nodes (density I .062 to 1.0662) were poor both in “short-” and “long-lived” cells. A similar, but smaller isotopically inert population was also seen in the gradient containing spleen cells (density 1.0650 to 1.0685). In view of the method of labeling which was employed, it is not possible to determine if these represent “very long-lived cells” (i.e., cells which did not divide even once in a period of a whole month) or “intermediate cells” (i.e., cells which divided after the end of the 3H labeling period but too slowly to incorporate 14C). The former alternative seems more likely particularly in view of the very large range in specific activities for either isotope which occurred in a single organ and especially in the spleen (fig. 1). Our results thus suggest that the categorization of lymphoid cells into the “short-lived” and the “long-lived” categories is a gross oversimplification. They are consistent with the interpretation that there are a multitude of subclasses of lymphoid cells, having discretely different life expectancies in the range from a few days or less to well over 1 month. The relative abundance of cells having these different half-lives in the different organs presumably explains the overall variation in longevity of cells from organ to organ which