Walmor C. De Mello

Angiotensin II and the Heart: On the Intracrine Renin-Angiotensin System

-The active end product of the renin-angiotensin system, angiotensin II (Ang II), through the activation of specific Ang II receptors, regulates cardiac contractility, cell coupling, and impulse propagation and is involved in cardiac remodeling, growth, and apoptosis. We review these subjects, as well as the second messengers that are involved, and the synthesis of Ang II in the heart under normal and pathological conditions. Finally, we discuss the possibility that there is an intracrine renin-angiotensin system in the heart that plays a role in the control of cell communication and inward Ca(2+) current.

Intracellular Angiotensin II Regulates the Inward Calcium Current in Cardiac Myocytes

Abstract —The influence of intracellular administration of angiotensin II (Ang II) on the inward calcium current (I Ca ) was investigated in single myocytes isolated from adult rat ventricle. Comparative studies were also made in ventricular cells of Golden hamsters. The I Ca was measured in single cells using the whole-cell voltage clamp configuration. The results indicated that Ang II (10 −8 mmol/L) dialyzed into the rat myocytes reduced the peak I Ca by 35±5.5% (n=20; P −7 mmol/L) added to the bath did not suppress the effects of Ang II, indicating that the peptide is acting intracellularly. Moreover, the intracellular dialysis of losartan (10 −6 mmol/L) or [Sar 1 Val 5 Ala 8 ] Ang II (10 −6 mmol/L) did not change the effect of Ang II. Stimulation of I Ca by exogenous cAMP or inhibition of protein kinase C did not alter the effect of Ang II on I Ca . Zaprinast (100 μmol/L), an inhibitor of cGMP phosphodiesterase, when added to the bath solution increased appreciably the effect of Ang II on I Ca ( P −8 mmol/L) increased I Ca by 36±2.4% (n=20; P >0.05). The effect of the peptide was not altered by the intracellular administration of losartan (10 −6 mmol/L), by [Sar 1 Val 5 Ala 8 ] Ang II (10 −6 mmol/L), or by the inhibitor of protein kinase A. The inhibition of protein kinase C, however, prevented the effect of Ang II I Ca in the hamster myocytes. The results particularly suggest that the activation of the cardiac renin-angiotensin system regulates I Ca and myocardial contractility, an effect that varies with the species.

Renin-Angiotensin System and Cell Communication in the Failing Heart

Abstract The influence of heart failure on the process of cell communication was investigated in cell pairs isolated from the ventricle of cardiomyopathic hamsters (11 months old) and the results compared with age-matched normal hamsters. The gap junctional conductance (gj) was measured with two voltage-clamp amplifiers. The results showed two major populations of cell pairs with respect to gj values: one with very low values (0.8 to 2.5 nS) and the other with higher values (7 to 35 nS). In normal hamsters, the most frequent gj values were in the range of 40 to 100 nS. Angiotensin II (Ang II, 1 μg/mL) caused cell uncoupling in myopathic myocytes with low gj but reduced gj by 53±6.6% (±SE) in cell pairs with higher gj values (7 to 35 nS). The effect of Ang II on gj of myopathic cell pairs was suppressed by losartan (10−7 mol/L). In cardiomyopathic cell pairs with low gj (0.8 to 2.5 nS), enalapril (1 μg/mL) caused an appreciable increase in gj (219±20.3%), whereas in cell pairs with higher gj (7 to 35 nS), the gj increment was smaller (80±10.8%) but still larger than that seen in controls (33±5.4%). Intracellular dialysis of Ang I (10−8 mol/L) abolished cell communication in myopathic cell pairs with low gj (0.8 to 2.5 nS) and reduced gj by 66±1.7% in the other pairs (7 to 35 nS). The effect of Ang I on gj was greatly reduced by enalaprilat (10−9 mol/L) added to the cytosol. Dialysis of Ang II (10−8 mol/L) into the myopathic cell reduced gj by 48±4.2%, an effect abolished by losartan (10−8 mol/L). The results indicate that the decline in gj seen in the ventricle of cardiomyopathic hamsters is in part due to activation of the cardiac renin-angiotensin system.

Cell-to-cell communication in heart and other tissues

The establishment of cell communities requires the development of a particular ability of the functional units to recognize one another. Recognition and further adhesion are initial steps in the process of organ formation (Grobst~4n, 1954 and Spiegel, 1954). During the embryogenesis cells move randomly until they recognize the final target. Chemotactic pathways established in the form of large or small molecules seems to be involved in this process (Wolpert, 1969 and Wolpert et al., 1974). In fetal no, cortex the migration of neurons probably occurs by contact guidance (Rakic, 1974) provided by the extracellular structures or neighboring cells. At the beginning of this century, Wilson (1907) and Holtfreter (1939) showed that dispersed cells are able to reassociate, a process involving components of the cell surface. Studies carried out on slime molds (Barondes and Rosen, 1976) indicated that species specific aggregation is related to the appearance of lectin-like macromolecules on the surface of the cells. The role of surface giycoproteins to cell recognition and adhesion has been supported by studies in sponges (Humphreys, 1963 and Gessner and Ginsberg, 1964). In 1947 Weiss proposed that lock and key molecules located at the surface of apposing cells might promote ~ o n ff they are complementary. A major advance came through the development of procedures for cell aggregation by rotation (Moscona, 1961) and the use of enzymic methods which made it possible to approach cell aggregation quantitatively. Interestingly, these studies show that reassociation of trypsin-dissociated embryonic cells into complex aggregates follows a reconstruction of their original pattern (Moscona, 1957).

Modulation of junctional permeability.

Cell-to-cell coupling through low-resistance junctions represents a very old mechanism of intercellular communication. In sponges and medusae, for instance, without nervous tissue, the epithelia receive external stimuli and convert them into electrical pulses that are conducted in all directions through low-resistance junctions (Mackie, 1964).

Influence of Intracellular Renin on Heart Cell Communication

Abstract The influence of intracellular renin and angiotensinogen on the control of cell-to-cell communication in heart muscle was investigated in cell pairs isolated from adult rat ventricle. Junctional conductance was measured with two separated voltage-clamp circuits. Intracellular dialysis of renin (0.2 pmol/L) caused a decrease in junctional conductance of 29±3.8% (±SEM, P −9 mol/L) dialyzed into the cell caused an appreciable reduction in the effect of renin. The intracellular administration of renin (0.2 pmol/L) plus angiotensinogen (0.4 pmol/L) produced a faster and stronger fall in junctional conductance (84.3±1.35%, P

Angiotensin (1-7) re-establishes impulse conduction in cardiac muscle during ischaemia-reperfusion. The role of the sodium pump

INTRODUCTION The effect of angiotensin (1-7) (Ang 1-7) on membrane potential and excitability of rat heart muscle under ischaemia/reperfusion was investigated. MATERIALS AND METHODS The hearts of adult rats were removed under deep anaesthesia and perfused using the Langendorff method. After 40 minutes of global no-flow ischaemia, the heart was reperfused for five minutes and the right ventricle was dissected out and transferred to a transparent chamber, through which normal oxygenated Krebs solution flowed continuously (37 degrees C). Measurements of membrane potential were performed using an intracellular microelectrode connected to a high impedance preamplifier. The muscle was stimulated with rectangular current pulses (3 ms duration; 0.6 Hz) generated by an electronic stimulator and isolation unit. To study the influence of Ang (1-7) on sodium pump current, isolated myocytes were voltage-clamped at -40 mV and the current generated by the pump was recorded before and after the administration of Ang (1-7) (10-8 M) to the bath. RESULTS Ang (1-7) (10-8 M) hyperpolarised the ischaemic heart fibre and re-established impulse propagation. The increment of resting potential was related to the activation of the sodium pump. Indeed, Ang (1-7) (10-8 M) enhanced the transient outward current generated by an electrogenic sodium pump. Both effects of Ang (1-7) on membrane potential and pump current were abolished by ouabain (10-7 M). The cardiac refractoriness was also increased by Ang (1-7) (10-8 M). CONCLUSIONS Ang (1-7) activates the sodium pump, hyperpolarises the heart cell and re-establishes the impulse conduction during ischaemia/reperfusion. These effects of Ang (1-7), and the increment of cardiac refractoriness, provide an explanation for the reduced incidence of arrhythmias during ischaemia/reperfusion in the presence of Ang (1-7).

On the local cardiac renin angiotensin system. Basic and clinical implications

In the present review we reevaluated the experimental and clinical evidence that there is a local renin angiotensin system in the heart as well as the presence of a functional intracrine component which is activated during pathological conditions like heart failure and hypertension. The implications of these findings for cardiology were discussed. The novel finding that cell swelling impairs cell coupling and impulse propagation through activation of ionic channels with consequent generation of cardiac arrhythmias and the evidence that AT1 receptors are mechanosensors able to alter the heart function independently of Ang II were discussed. Particular attention was given to the role of salt loading on the activation of a local cardiac renin angiotensin and its consequences.

Beneficial versus harmful effects of Angiotensin (1-7) on impulse propagation and cardiac arrhythmias in the failing heart

Introduction. The presence of Angiotensin (1-7) (Ang 1-7) and ACE 2 in the ventricle of cardiomyopathic hamsters as well as the influence of Ang (1-7) on membrane potential, impulse propagation and cardiac excitability were investigated. Methods. Histology and immunochemistry were used to demonstrate the presence of Ang (1-7) and ACE 2 in the ventricle of cardiomyopathic hamsters. Measurements of transmembrane potentials, conduction velocity and refractoriness were made using conventional intracellular microelectrodes. The influence of Ang (1-7) on sodium pump current was investigated in voltageclamped myocytes isolated from the ventricle. Results. The results indicated the presence of Ang (1-7) and ACE 2 in myocytes of cardiomyopathic hamsters. Moreover, Ang (1-7) (10-8 M) hyperpolarised the heart cell, increased the conduction velocity, and I reduced transiently the action potential duration. The cardiac refractoriness was also increased by the heptapeptide, an effect in part reduced by an inhibitor of mas receptor. These findings indicate that Ang (1-7) has important antiarrhythmic properties. However, the beneficial effects of Ang (1-7) are dose-dependent because at higher concentration (10-7 M) the heptapeptide elicited an appreciable increase of action potential duration and early-after depolarisations. Since losartan (10-7 M) did not counteract this effect of the high dose of the heptapeptide, it is possible to conclude that activation of AT1-receptors is not involved in this effect of Ang (1-7).To investigate the mechanism of the hyperpolarising action of Ang (1-7) the influence of the heptapeptide on the sodium potassium pump current was studied in myocytes isolated from the ventricle of cardiomyopathic hamsters. The peak pump current density was measured under voltage clamp using the whole cell configuration. The results indicated that Ang (1-7) (10—8 M) enhanced the electrogenic sodium pump, an effect suppressed by ouabain (10—7 M). Conclusions. Ang (1-7) has beneficial effects on the failing heart by activating the sodium pump, hyperpolarising the cell membrane and increasing the conduction velocity. These effects as well as the increment of refractoriness indicate that Ang (1-7) has antiarrhythmic properties. At higher concentrations (10—7 M), however, the heptapeptide induced early-after depolarisations which leads to the conclusion that an optimal generation of Ang (1-7) must be achieved to permit a protective role of Ang (1-7) on cardiac arrhythmias.

Cell Coupling and Impulse Propagation in the Failing Heart

Cell Coupling and Impulse Propagation. Cell coupling and impulse propagation were investigated in the ventricle of cardiomyopathic hamsters at an advanced stage of heart failure. An appreciable decline in junctional conductance was found, a phenomenon in part related to activation of the plasma and cardiac renin‐angiotensin systems. Decreased expression of conucxin43 or an alteration of junctional proteins also might be implicated in the decreased cell coupling. Morphologic abnormalities such as fibrosis, necrosis, and rupture of cell contacts contribute to the decline of conduction velocity or to the blockade of impulse propagation in some areas of the ventricle, creating the conditions for anisotropic conduction and cardiac arrhythmias. The decrease in membrane potential found in myopathic cells is related in part to depression of Na‐KATPase activity, and the lack of action of beta‐adrenergic agonists on junctional conductance is explained by down‐regulation of beta receptors and an abnormality of adenyl cyclase.