M. Oratz

Adenyl cyclase activity and cyclic AMP in acute cardiac overload: A method for measuring cyclic AMP production based on ATP specific activity☆

Abstract As early as 20 min after onset of acute hemodynamic overload in the perfused guinea pig heart, nuclear RNA polymerase activity is increased followed by increased synthesis of microsome protein and ultimately myosin. In order to evaluate the mechanism triggering the above sequence, the effect of overload on adenyl cyclase activity was studied. In addition, a method was devised to measure directly the formed cyclic 3′5′ AMP using [ 3 H]adenosine as a precursor. After 10 min, a point midway between onset of stress and first appearance of increased nuclear polymerase activity, ATP concentration, [ 3 H]ATP specific activity, and 3 H-cyclic AMP were the same in control and overloaded left ventricles. However, adenyl cyclase activity was significantly increased in the particulate fraction obtained from overloaded left ventricles. It is interesting to suggest a possible role of adenyl cyclase in the mechanism triggering increased protein synthesis in overload.

Protein synthesis in prolonged cardiac arrest

Abstract Cardiac protein synthesis has been found to increase as pressure stress is applied both in vivo and in vitro, but the relationship between contraction, per se, and protein synthesis has not been characterized. A cardiac preparation with right ventricular loading and controlled coronary flow was used to examine the possibility of such a relation, in vitro. In both aerobically perfused contracting and high K+ arrested hearts, ATP, creatine phosphate, potassium and glycogen levels were maintained, and lactate production was at low levels. Incorporation of [14C]lysine into right and left ventricular protein was the same in contracting and aerobically perfused arrested hearts after 3 h of perfusion. In contrast, anoxia induced arrest decreased incorporation of lysine into protein with a more profound drop in [14C]lysine incorporation in the left ventricle. Alterations of protein synthesis in 3 h of anoxia were associated with a fall in ATP (greater in the left ventricle than in the right ventricle), creatine phosphate, glycogen and potassium with a sharp rise in lactate production. These changes in anoxia were minimized when anoxic hearts were concomitantly perfused with high K+ (16 mEq/l). The data indicate that cardiac arrest associated with metabolic integrity in this preparation produced little change in protein synthesis and suggest that a baseline rate of protein synthesis occurs in such tissue which is not dependent on contraction, per se.

ALBUMIN-OSMOTIC FUNCTION†

Publisher Summary This chapter presents albumin–osmotic function. The term oncotic pressure is synonymous with colloid osmotic pressure. Also measurements of oncotic pressure in plasma indicate a better correlation with albumin content than with total protein content. The function of oncotically active molecules is to retain water in the blood in the capillary beds despite the hydrostatic pressure that is trying to force this water out. Oncotic pressure is quite constant and varies from 240–465 mm H 2 O. The variations in mean values are primarily because of the techniques of measurement. However, in individual the oncotic pressure is quite stable. There is some diurnal variation, but this was found to be a postural effect and probably related to the change in blood volume upon arising. Moreover, the loss in water from the capillary beds to extravascular areas could be prevented by the presence of an osmotically active particle in the blood that would not be rapidly filtered through the capillary walls. The protein, albumin, is ideally suited for this osmotic function. Of the plasma proteins it has a relatively low molecular weight and its large net negative charge at blood pH makes this molecule osmotically more effective than an equal weight of the other plasma proteins.

Ethanol and Cardiac Protein Synthesis

Acute exposure of the heart to ethanol does not appear to alter the rate of young guinea pig cardiac protein synthesis when assayed in vitro. In contrast, the primary metabolite of ethanol, acetaldehyde, markedly diminishes synthesis despite its chronotropic and inotropic effects. On the other hand, after 11-13 weeks of ethanol-drinking during growth and maturation, the synthetic capacity of the working right ventricle was decreased when measured in vitro with normal perfusate. Assay of synthesis of the contractile proteins myosin heavy and light chains, actin and tropomyosin suggests a change in synthesis or pool size of actin reflected in an alteration of relative synthesis of this protein compared to that of heavy chains. The relative synthesis of the other proteins remained at control levels. When hearts from ethanol-drinking and matched control animals were perfused under conditions of severe ischemia, there was a profound fall in protein synthesis in all hearts, and ethanol did not enhance the inhibition of synthesis. However, the hearts from ethanol-drinking animals showed a more marked and significant impairment of maintaining ejection pressure with a marked increase in coronary resistance as the perfusion progressed. It is postulated that some impairment of protein metabolism may occur during prolonged ethanol exposure, which may influence the cardiac response of another induced stress, e.g., ischemia.

PROTEIN SYNTHESIS IN THE HEPATOCYTE

The level of serum albumin has often been relied upon t o reflect the status of the liver in health and disease. It is well known that, while the concentration of serum albumin is decreased in alcohol-induced cirrhosis, data are available indicating that in many cases the exchangeable albumin pool is not depressed;’ further, when alcohol is removed and adequate nutrition supplied, the albuminsynthesizing potential is restored.* The question whether the removal of alcohol or the institution of adequate nutrition was responsible for the liver’s recovery has been the subject of much debate. In vivo the level of albumin is the net result of synthesis, degradation, and distribution, and in the whole animal albumin synthesis is influenced by the interplay of the nutritional status of the animal, e n ~ i r o n m e n t , ~ hormones, oncotic equilibrium,“ and toxins, as well as the state of health. Thus, in vivo, it is difficult t o study the effect of one isolated factor in albumin synthesis, and for this reason the isolated perfused liver was employed. The advantages are obvious: (a) the perfusion medium can be altered in a particular manner with any substance with the knowledge that the substance perfusing the liver has not been altered by prior passage through any tissue: (b) the effects of pretreatment of the inact animal on the liver’s ability to synthesize albumin can be studied by removal of the liver after pretreatment; and (c) subcellular systems can be isolated following perfusion and correlative evidence of altered microstructure with altered liver function may be obtained. In the present study the isolated perfused liver was used t o study the effects of altered nutrition, in the absence and presence of ethyl alcohol, on the liver’s ability t o synthesize albumin as well as the correlative effect on the endoplasmic reticulum-bound polysome, the organelle responsible for the synthesis of albumin. The results indicated that a short term fast of 24 hours was sufficient t o decrease albumin synthesis to one-half that found in livers from fed donors. The decrease in albumin synthesis was coincident with a disaggregation of the bound polysome. However, this effect could be readily reversed by the presence

Alcoholic Cardiomyopathy: Studies of Protein Metabolism

It is now well known that cardiomyopathy may follow prolonged ingestion of ethanol in man. This myopathy appears to be unrelated to the associated liver dysfunction and damage. Studies with animal models have been difficult because of different diets, species of animal studied, and the duration of administration of ethanol. The results have also been inconsistent. However, what has emerged in the last decade is the general finding that although the heart may behave normally under “control” conditions, when an additional stress, either chemical or hemodynamic, is applied, the heart from the ethanol exposed animal reacts abnormally. As examples, little impairment of cardiac contractile action has been noted in alcohol exposed hearts unless there is the added stress of increased afterload (1,2). Similarly, both fast electrical pacing (3) and ischemia (4) produce a greater impairment of contractile action in the ethanol exposed heart than in the matched control. Further, in vitro studies with hearts from ethanol exposed animals show a decresed contractile response to increases in calcium when compared to the control (5).

Alcohol Effects on Albumin Synthesis

Alcoholism and alcohol-induced organ damage are two separate entities — the former, representing an abnormal syndrome, may show organ involvement early or late, while the latter may occur without true addiction. Either the effects of alcohol or its metabolite acetaldehyde or the resultant effects of complete alcohol metabolism on protein synthesis are probably organ specific, modified by genetic predisposition, altered by nutritional status, hormone inbalance, absorptive capabilities, and the presence or absence of prior alcohol-induced or associated organ damage. To try to isolate the acute and the chronic effects of ethanol exposure on a specific protein synthetic pathway in vivo is not a simple task. The picture is further complicated by the complex nature of the normal protein synthetic and cellular transport systems. It is only necessary to consider how many potential sites in the assembly line process of protein synthesis may be altered by alcohol to see how complex a scheme we are facing (Mueckler and Pitot 1981, 1982; Bantle et al. 1980a; Krieg et al. 1980; Prehn et al. 1981; Thomas et al. 1981; Emr et al. 1980; Walter and Blobel 1981; Rothman 1981; Dorling et al. 1975; Algranati and Sabatini 1979; Schreir et al. 1977; Blobel and Dobberstein 1975; Weigand et al. 1982; Peters et al. 1971; Redman and Cherian 1972; Glauman and Ericsson 1970). Following the final posttranscriptional modification, the appropriate messenger RNA (mRNA) becomes available for the translation process. In its most simplified form this process requires the presence of initiation factors which promote the formation of the ribosomal complex, a complex of the two ribosome subunits and the particular mRNA.

Cardiac Protein Synthesis in Stress: Overload, Ethanol and Anoxia

Protein synthesis is of major importance in maintenance of cellular structure and viability. In the heart, proteins include those with structural, contractile and enzymatic function. Since changes in cardiac size, gross structure and function have long been known to occur with different stresses, the assay of protein synthesis has become of increasing importance, and the present paper will examine alterations of such synthesis in specific stresses as hemodynamic overload, anoxia and exposure to ethanol or its metabolites.

The Influence of Ethanol on Albumin Metabolism

The mechanisms responsible for the production of protein within the liver cells are the subject of continued detailed investigations and the myriad of steps located between the initial turning on of the assembly line to the final extrusion of the completed molecule offer innumerable areas for regulation or modification and thus control. In attempting to outline the effects of any particular stress on the synthetic mechanism it will ultimately be necessary to delineate the steps as they proceed in the formation of the protein. Further, it will be necessary to use as specific an isolated system as possible so that the interplay of in vivo factors tending to dampen the effects of one stress or another can be eliminated.