Parameters of Cell Death and Proliferation of Prostate Cancer Cells with Altered Expression of Myosin 1C Isoforms.

Myosin 1C is a monomeric myosin motor with a truncated tail domain. Such motors are referred as slow "tension sensors." Three isoforms of myosin 1C differ in short N-termed amino acid sequences, the functional differences between isoforms have not been elucidated. Myosin 1C isoform A was described as a diagnostic marker for prostate cancer, but its role in tumor transformation remains unknown. Based on data on the functions of myosin 1C, we hypothesized the potential role of myosin 1C isoforms in maintaining the tumor phenotype of prostate cancer cells. In our work, we showed that a decrease in the expression level of myosin 1C isoform C leads to an increase in the proliferative activity of prostate tumor cells.

[Modifications to the solution for reperfusion of the ischemic heart]. / Modifikatsii rastvora dlia reperfuzii ishemizirovannogo serdtsa.

Isolated working guinea pig hearts were subjected to 30-min total normothermic ischemia and 30-min reperfusion with modified Krebs solution to which various agents were added. The presence of glutamate in the solution resulted in better recovery of cardiac pump function and higher myocardial energy phosphate levels. Addition of phrelone, a calmodulin inhibitor, or adenosine and ribose did not improve pump function. The similar functional recovery was observed after addition of taurine instead of glutamate and their combination exerted even higher functional recovery while further addition of phosphocreatine was not so effective.

[Formation of products of anaerobic metabolism in the ischemic myocardium]. / Obrazovanie produktov anaérobnogo obmena v ishemicheskom miokarde.

The effect of ischemia on the formation of products of anaerobic metabolism and their release into the cardiac effluent in isolated perfused guinea pig hearts was studied. During 30 min normothermal ischemia, the myocardial ATP and phosphocreatine levels decreased to 34% and 15% of the initial values, respectively. The net alanine formation in ischemia was approximately a stoichiometric glutamate decrease; the increase in the tissue malate content corresponded to the aspartate----oxaloacetate----malate anaplerotic flux, the succinate production being commensurable to alpha-ketoglutaric acid formation in the alanine aminotransferase reaction. Using 1H-NMR, it was shown that the release of trace amounts of lactate, alanine, succinate, creatine and pyruvate into cardiac effluents occurred during the first 5 minutes of reperfusion. The rate of metabolite release decreased in the following order: lactate much greater than alanine greater than succinate greater than creatine. By the 30th minute of reperfusion, the decrease in the tissue levels of these metabolites to preischemic values was accompanied by the recovery of ATP and phosphocreatine to 65% and 90% of the initial levels, respectively. The data obtained suggest that the formation and release of alanine, creatine or succinate as well as lactate from ischemic myocardium may testify to significant disturbances in energy metabolism of the myocardium.

[Mechanisms for improving the energy metabolism and function of the hypoxic myocardium with amino acids]. / Mekhanizmy uluchsheniia énergeticheskogo obmena i funktsii gipoksicheskogo miokarda aminokislotami.

The relationship between metabolism of the main myocardial amino acids, glutamate, aspartate and alanine, and energy state of hypoxic myocardium, was studied. Depression of cardiac contractile function during asphyxia in rats was accompanied by a decrease in glutamate mitochondrial and tissue contents and an increase in the tissue alanine and mitochondrial aspartate. The reduction of mitochondrial glutamate in asphyxia was related to losses of intramitochondrial ATP and state 3 respiration with glutamate and malate. Using NMR technique, exogenous glutamate and oxaloacetate were shown to increase succinate formation coupled with ATP and CTP production in the rat heart mitochondria in absence of aeration. These data suggest that glutamate and products of its transamination decrease the contraction of hypoxic myocardium stimulating anaerobic energy formation in the tricarboxylic acid cycle.

The relationship between the cardiac contractile function, adenine nucleotides and amino acids of cardiac tissue and mitochondria at acute respiratory hypoxia.

The effect of asphyxia and subsequent resumption of respiration on the content of adenine nucleotides and some amino acids in heart tissue and mitochondria, as well as respiration of heart mitochondria was studied in rats. The depression of cardiac contractile function during asphyxia showed a better correlation with losses in mitochondrial adenine nucleotides (ATP + ADP + AMP) than those in cardiac tissue. The decrease in the heart work index was accompanied by a decrease in state 3 respiration with glutamate and malate as well as uncoupled respiration with these substrates. This did not occur with succinate. Nonphosphorylating (state 4) respiratory rates and ADP/O ratios were slightly affected by asphyxia, when respiratory substrates of both types were used. The decreased level of glutamic acid in the tissue and mitochondria of asphyxic hearts was simultaneously observed with a significant increase of alanine in cardiac tissue and of aspartic acid in the mitochondria. The losses of intramitochondrial ATP and respiratory activity with NAD-dependent substrates during asphyxia were associated with a reduction of glutamic acid level in mitochondria. The recovery of cardiac function during resumption of respiration was related to the restoration of mitochondrial respiration supported by glutamate and malate, as well as to the restoration of mitochondrial adenine nucleotides and glutamic acid. The results suggest that the depression of cardiac function caused by acute respiratory hypoxia may be attributed to impairment of electron transport, particularly in complex I of the respiratory chain and changes in metabolism of glutamic acid.

[Relation between glutamate and adenine nucleotide levels of heart mitochondria during hypoxia]. / Vzaimosviaz' mezhdu soderzhaniem glutamata i adeninnukleotidov v mitokhondriiakh serdtsa pri gipoksii.

The effect of acute respiratory hypoxia in rats on mitochondrial respiration, adenine nucleotides and some amino acids of the heart was studied. The decrease in the total (ATP + ADP + AMP) and exchangeable (ATP + ADP) adenine nucleotide pool of the mitochondria was accompanied by a pronounced loss of state 3 respiration with glutamate plus malate and a slight decrease with succinate plus rothenone. The uncoupled respiration of mitochondria with glutamate and malate was decreased in the same degree as in the absence of 2,4-dinitrophenol. State 4 respiration with substrates of both types was unaffected by hypoxia. These data point to a hypoxia-induced impairment of complex I of the respiratory chain. The decrease of tissue and mitochondrial glutamate was accompanied by the elevation of alanine content in the heart and an increase in intramitochondrial aspartate. The ADP-stimulated respiration of mitochondria was correlated with mitochondrial glutamate and ATP as well as with exchangeable adenine nucleotide pools during hypoxia. The experimental results suggest that mitochondrial dysfunction induced by hypoxia may also be attributed to the low level of mitochondrial glutamate.

[Formation of the intermediate products of the tricarboxylic acid cycle and ammonia from free amino acids in anoxic heart muscle]. / Obrazovanie promezhutochnykh produktov tsikla trikarbonovykh kislot i ammiaka iz svobodnykh aminokislot myshtsy serdtsa pri anoksii.

Glutamate and aspartate showed the highest rate of catabolism in oxygenated isolated rat heart with the formation of glutamine, asparagine and alanine. Under anoxia, the catabolism of branch chained amino acids and that of lysine, proline, arginine and methionine was inhibited. However, glutamate and aspartate catabolized at a higher rate as compared with oxygenation. Alanine was the product of their excessive degradation. During oxygenation, 70% of ammonia were produced via deamination of amino acids. Under anaerobic conditions the participation of amino acids in ammoniagenesis decreased to 4%; the principal source of ammonia was the adenine nucleotide pool. The total pool of the tricarboxylic acid cycle intermediates increased 2.5-fold due to accumulation of succinate. The data obtained suggest that the constant influx of intermediates into the cycle from amino acids is supported by coupled transamination of glutamate and aspartate. This leads to the formation of ATP and GTP in the tricarboxylic acid cycle during blocking of aerobic energy production.

Adenine nucleotides, glutamate and respiratory function of heart mitochondria during acute hypoxia.

The effect of acute hypoxia on adenine nucleotides, glutamate, aspartate, alanine and respiration of heart mitochondria was studied in rats. The losses of intramitochondrial adenine nucleotides (ATP+ADP+AMP) during hypoxia were related to depression of state 3 respiration supported by glutamate and malate, as well as decrease in uncoupled respiration. Hypoxia had less prominent effect on succinate-dependent state 3 respiration. Non-phosphorylating (state 4) respiratory rates and ADP/O ratios were slightly affected by oxygen deprivation. Glutamate fall in tissue and mitochondria of hypoxic hearts was concomitant with significant increase in tissue alanine and mitochondrial aspartate. The losses of intramitochondrial ATP and respiratory activity with NAD-dependent substrates during hypoxia were related to a decrease in mitochondrial glutamate. The results suggest that hypoxia-induced impairment of complex I of respiratory chain and a loss of glutamate from the matrix may limit energy-producing capacity of heart mitochondria.

The role of amino acid catabolism in the formation of the tricarboxylic acid cycle intermediates and ammonia in anoxic rat heart.

Amino acid catabolism, the tricarboxylic acid cycle intermediates and ammonia formation were studied in isolated perfused rat heart under anoxia. The total net anaplerosis due to amino acid degradation in anoxia was equal to that in oxygenation (6.29 and 6.09 mumol/g dry weight per h, respectively) as a result of the increased transamination of glutamic and aspartic acids. During anoxic perfusion, the rate of catabolism of glutamic and aspartic acids was 1.5-times higher than in normoxia, while depletion of branched-chain amino acids, lysine, proline, arginine and methionine, was inhibited. Alanine was the product of excessive degradation of glutamic and aspartic acids. Under anaerobic conditions, in spite of inhibition of amino acid deamination, ammonia formation was increased 2.7-fold as compared to oxygenation. The principal amount of ammonia (96%) was produced at degradation of adenine nucleotides. A 2.5-fold increase in the pool of the tricarboxylic acid cycle intermediates under anoxia was associated mainly with accumulation of succinate. The data suggest that the coupling of alanine- and aspartate amino transferases is a mechanism controlling the tricarboxylic acid cycle pool size in anoxic heart.

On the mechanism of enhanced ATP formation in hypoxic myocardium caused by glutamic acid.

The effect of glutamic acid on the cardiac contractile function and sources of anaerobic ATP formation in hypoxic myocardium was studied in isovolumic rat hearts. The presence of glutamic acid (5 mM) in the perfusate significantly diminished an increment in diastolic pressure caused by 60 min hypoxia, and facilitated its complete recovery during 30 min reoxygenation. This effect was combined with the maintenance of a higher ATP level during hypoxia and reoxygenation. The total content of lactate in the heart-perfusate system rose exactly as during hypoxia without glutamic acid, while pyruvate content decreased due to increased alanine formation. Restoration of tissue content of glutamate and aspartate in the presence of exogenous glutamic acid was accompanied by a more than 2-fold increase in succinate formation, the end-product of the Krebs' cycle under anaerobic conditions. The products of glutamic acid transamination with oxaloacetic acid, aspartic and alpha-ketoglutaric acids (5mM each), induced the same functional and metabolic alterations as glutamic acid. Amino-oxyacetic acid, a tramsaminase inhibitor, eliminated the effects caused by glutamic acid. Moreover, the inhibition of transamination was accompanied by a decreased succinate and alanine synthesis as well as insignificantly increased lactate formation compared to hypoxia without additives. The results suggest that the beneficial effect of glutamic acid is due to the activation of anaerobic ATP formation in the mitochondria rather than stimulation of glycolysis.

[Effect of glutamic acid on the anaerobic formation of ATP in the heart muscle]. / Vliianie glutaminovoi kisloty na anaérobnoe obrazovanie ATP v myshtse serdtsa.

The effect of exogenous glutamic acid on nitrogenous and energetic metabolism of isolated perfused heart of the rat was studied under anoxia. Addition of 5 mM glutamic acid to the perfusate significantly increased the ATP level in anoxic heart. Perfusion of anoxic heart with 5 mM glutamic acid recovered the glutamate and aspartate tissue content and caused augmented production of alanine and succinate, while lactate formation did not change. In the presence of glutamic acid, the glutamine and asparagine contents in the heart-perfusate system markedly increased; however, the ammonia content did not reduce significantly. The effect on glutamic acid was reproduced by its transamination products, 5 mM aspartic and 5 mM alpha-ketoglutaric acids, and was totally eliminated by 2 mM aminooxyacetic acid, an inhibitor of transaminases. These data suggest that the glutamate-induced protective effect of ATP is probably related to the stimulation of substrate phosphorylation in mitochondria, resulting in succinate synthesis that is coupled with glutamate transamination.

Protective effect of glutamic acid on cardiac function and metabolism during cardioplegia and reperfusion.

The effect of glutamic acid added to cardioplegic solution containing 20 mM K+ on the cardiac function and metabolism was studied in isolated working rat hearts. 30-min cardiac arrest resulted in profound fall in creatine phosphate and ATP content, by four- and two-fold, respectively, as well as in four-fold rise in AMP content. Simultaneously, during cardioplegia a decline in tissue glutamate and aspartate content and an increase in tissue ammonia and alanine content were found. After reperfusion, an incomplete restoration of ATP, AMP, and creatine phosphate content were observed; the cardiac output recovered only to 39 percent of the initial value. An addition of glutamic acid to cardioplegic solution was associated with significantly less decline in the content of high-energy phosphates and less prominent rise in AMP content during cardioplegia. It also prevented the decline in tissue aspartate content and caused a lesser ammonia accumulation in myocardial tissue due to the activation of glutamine synthesis. In spite of this the tissue ammonia level remained elevated. Reperfusion with Krebs-Henseleit buffer resulted in the recovery of cardiac output to 75% of the initial value as well as better restoration of high-energy phosphate content. The addition of glutamic acid in the perfusate during reperfusion led to further improvement of ATP and creatine phosphate content. It is suggested that an addition of glutamic acid may have beneficial effect in open heart surgery.

Effect of exogenous amino acids on the contractility and nitrogenous metabolism of anoxic heart.

The effect of exogenous glutamic acid and arginine on the contractility of isolated perfused rat heart and on the metabolism of some nitrogenous compounds was studied. Sixty-minute anoxic perfusion (95% N2 + 5% CO2) led to a fall in developed isovolumic pressure and an elevation in diastolic pressure, to an increase in the production of alanine, glutamine, and ammonia, and to a decrease in the tissue content of aspartate and glutamate. The total pool of free amino acids and taurine under these conditions remained unchanged. Subsequent 40-min reoxygenation partially restored the contractile function. Addition of 3.5 mM glutamic acid or 5 mM arginine into the perfusate before anoxia resulted in a higher level of developed pressure and a lower level of diastolic pressure during anoxia and almost complete recovery of cardiac function after subsequent reoxygenation. Both amino acids had no effect on ammonia formation by the anoxic heart but enhanced its binding in myocardial tissue via formation of glutamine and urea. It is suggested that the exogenous amino acid effect on anoxic heart is mediated by activation of substrate phosphorylation rather than the ability to bind tissue ammonia.

Effect of glutamic and aspartic acids on adenine nucleotides, nitrogenous compounds and contractile function during underperfusion of isolated rat heart.

The experiments were carried out to find out whether exogenous glutamic or aspartic acid could diminish changes in the cardiac contractile function and high-energy phosphate content caused by underperfusion of isolated isovolumic rat heart. After 40 min of reduced coronary flow (from 10 to 3 ml/min) there was an almost four-fold fall in the developed pressure, and more than three-fold rise in the diastolic pressure as well as a profound fall in creatine phosphate (CP) and ATP content. Glutamic (68 mM) or aspartic (75 mM) acids were added to the perfusate after 10 min of underperfusion when the developed pressure had declined almost to the same level as was observed after 40 min and the content of CP was reduced more than two-fold. Glutamic acid completely prevented the rise in the diastolic pressure and significantly increased the CP content, as compared to its level observed before addition of glutamate, but glutamic acid did not change the developed pressure. As a result, the CP and ATP contents were three- and two-fold higher, respectively, after addition of glutamic acid as compared to control underperfused hearts. Similar, but slightly less prominent effects were observed when aspartic acid was added instead of glutamic acid. These results suggest that high concentrations of glutamic and aspartic acids can exert beneficial effects on ischemic heart muscle.

[Effect of exogenous amino acids on myocardial contraction and metabolism of nitro-compounds in the myocardium during anoxia]. / Vliianie ékzogennykh aminokislot na sokratitel'nuiu funktsiiu i metabolizm azotistykh soedinenií serrdtsa pri anoksii.

Anoxic perfusion of an isolated rat heart resulted in contractility disorder, increased production of ammonia, alanine, glutamine and lowered levels of glutaminic and asparaginic acids, while the total pool of free amino acids and taurine remained unchanged. Subsequent reoxygenation partly recovered cardiac contractility. When 3.5 mM glutaminic acid or 5 mM arginine was added to the perfusate, the anoxic contracture was less pronounced, and the heart maintained a higher pressure than would be common for anoxia, while reoxygenation resulted in virtually complete recovery of contractility. Both amino acids did not basically affect ammonia synthesis, but enhanced its binding in heart tissue, i. e. glutamine and urea synthesis, the reactions requiring increased ATP spending. The findings suggest that the mechanism of exogenous amino acids' action in anoxic conditions is based on substrate phosphorylation rather than their ability to bind ammonia.

[Effect of glutamic and aspartic acids on heart metabolism and function in hypoperfusion]. / Vliianie glutaminovoi i asparaginovoi kislot na metabolizm i funktsiiu serdtsa pri gipoperfuzii.

The effects of glutamic and aspartic acids were studied during hypoperfusion of the rat isolated heart. The ischemic contracture that develops during hypoperfusion was prevented by glutamic acid. This effect was accompanied by preservation of higher tissue levels of ATP and CP, elimination of glutamate and aspartate deficiency, intensification of ammonia binding through the synthesis of glutamine, asparagine and urea by the myocardium. Nevertheless the level of free ammonia in the tissue remained fairly high. Aspartic acid had a similar but less pronounced effect on heart function and metabolism. The mechanism of contractile function preservation by amino acids appears to be connected with activation of oxidative and substrate phosphorylation in mitochondria rather than with the reduced level of free ammonia.

[Coronary perfusion rate as a factor determining the degree of heart contractile function decrease in an energy formation disorder]. / Skorost' koronarnoi perfuzii kak faktor, opredeliaiushchii stepen' umen'sheniia sokratitel'noi funktsii serdtsa pri narushenii énergoobrazovaniia.

Experiments on the isolated hearts of guinea pigs have shown that administration of dinitrophenol in the concentrations of 0.01-0.10 mmol elicited, depending on the dose given, a decrease in pressure of a latex balloon, placed into the cavity of the left ventricle and an increase in the displacement of the SI segment on the ECG, recorded from the subendocardial layer of the apex. It is established that the rate of the coronary flow, limiting washing out of K+ and other metabolites determines the degree of depression of the contractile function with energy formation disorders and also its restoration during reoxygenation.

Influence of adaptation to altitude hypoxia on the behavior of rats in a conflict situation.

Adaptation to altitude hypoxia under conditions of conflict between thirst and defensive motivation was studied. The subjects adapted to altitude hypoxia made three times more attempts to drink from a water bowl despite receiving an intense shock. Adaptation to altitude hypoxia increased the stability of an initiated drinking reaction over intense shock. In adapted animals that began to drink, the drinking response changed to defensive behavior with twice the shock intensity as in control animals. In the absence of conflict, there was no difference in the satisfaction of the thirst and the defensive motivation in animals adapted to hypoxia and in controls. Hence, the effect of adaptation to hypoxia consists in making the animals more capable of inhibiting the responses to defensive motivation under conditions where it is necessary to satisfy another vital motivation.

[Influence of adaptation to altitude hypoxia and subsequent de-adaptation on the retention of a temporary connection]. / Vliianie adaptatsii k vysotnoi gipoksii i posleduiushchei deadaptatsii na sokhranenie vremennoi sviazi.

Adaptation to simulated altitude hypoxia was followed by a significant increase of the degree of retention of active avoidance of the conditioned reflex. Twenty four hours after the reflex elaboration the adapted rats required only half of conditioned and unconditioned stimulus combinations in the reflex retention test than control ones. This effect was reproduced 1 and 5 days after the cessation of hypoxic training. The obtained increase in the degree of the conditioned reflex retention disappeared during deadaptation 10 and 27 days after the cessation of hypoxic training.