Antimycin A: stimulation of cell division and protein synthesis in Tetrahymena pyriformis.

Addition of antimycin A to a culture of Tetrahymena pyriformis caused an increase in cell division and protein synthesis in this ciliated protozoan. The antimycin effect is a function of the time of exposure to the antibiotic as well as of the age of the culture. A large accumulation of endoplasmic reticulum, reflecting increased protein synthesis, was visualized by electron microscopy in cells stimulated by the antimycin A.

Properties of carnitine transport in rat kidney cortex slices.

The properties of carnitine transport were studied in rat kidney cortex slices. Tissue:medium concentration gradients of 7.9 for L-[methyl-14C]carnitine were attained after 60-min incubation at 37 degrees C in 40 microM substrate. L- and D-carnitine uptake showed saturability. The concentration curves appeared to consist of (1) a high-affinity component, and (2) a lower affinity site. When corrected for the latter components, the estimated Km for L-carnitine was 90 microM and V = 22 nmol/min per ml intracellular fluid; for D-carnitine, Km = 166 microM and V = 15 nmol/min per ml intracellular fluid. The system was stereospecific for L-carnitine. The uptake of L-carnitine was inhibited by (1) D-carnitine, gamma-butyrobetaine, and (2) acetyl-L-carnitine. gamma-Butyrobetaine and acetyl-L-carnitine were competitive inhibitors of L-carnitine uptake. Carnitine transport was not significantly reduced by choline, betaine, lysine or gamma-aminobutyric acid. Carnitine uptake was inhibited by 2,4-dinitrophenol, carbonyl cyanide m-chlorophenyl-hydrazone, N2 atmosphere, KCN, N -ethylmaleimide, low temperature (4 degrees C) and ouabain. Complete replacement of Na+ in the medium by Li+ reduced L- and D-carnitine uptake by 75 and 60%, respectively. Complete replacement of K+ or Ca2+ in the medium also significantly reduces carnitien uptake. Two roles for the carnitine transport system in kidney are proposed: (1) a renal tubule reabsorption system for the steady-state maintenance of plasma carnitine; and (2) maintenance of normal carnitine levels in kidney cells, which is required for fatty acid oxidation.

Inhibition of the adenine nucleotide translocator by matrix-localized palmityl-CoA in rat heart mitochondria.

The activity of the adenine nucleotide translocator in rat heart mitochondria was quantitatively determined by the rate of [14C]ATP transport at 2 degrees C using the carboxyatractyloside inhibitor-stop technique. Linear uptake was obtained for 15 s, and with differing protein concentrations. The effect of matrix long-chain acyl-CoA esters upon the adenine nucleotide translocator activity was determined in these mitochondria. Incubation with palmitylcarnitine produced an increase in matrix long-chain acyl-CoA esters and decreased the velocity of [14C]ATP transport. Mitochondria isolated in the presence of KCN showed elevated levels of long-chain acyl-CoA esters, decreased transportable nucleotides, and very low adenine nucleotide translocator activity. Addition of potassium ferricyanide to these mitochondria caused a reduction in matrix acyl-CoA esters and partially restored adenine nucleotide translocator activity. Potassium ferricyanide also lowered matrix acyl-CoA in freshly isolated mitochondria and increased [14C]ATP transport. These findings show that the level of long-chain acyl-CoA esters within the mitochondrial matrix affects adenine nucleotide translocator activity and regulates mitochondrial activity.

Effect of dehydroepiandrosterone sulfate on carnitine acetyl transferase activity and L-carnitine levels in oophorectomized rats.

Alteration in energy metabolism of postmenopausal women might be related to the reduction of dehydroepiandrosterone sulfate (DHEAS). DHEA and DHEAS decline with age, leveling at their nadir near menopause. DHEA and DHEAS modulate fatty acid metabolism by regulating carnitine acyltransferases and CoA. The purpose of this study was to determine whether dietary supplementation with DHEAS would also increase tissue L-carnitine levels, carnitine acetyltransferase (CAT) activity and mitochondrial respiration in oophorectomized rats. Plasma L-carnitine levels rose following oophorectomy in all groups (P < 0.0001). Supplementation with DHEAS was not associated with further elevation of plasma L-carnitine levels, but with increased hepatic total and free L-carnitine (P = 0.021 and P < 0.0001, respectively) and cardiac total L-carnitine concentrations (P = 0.045). In addition, DHEAS supplementation increased both hepatic and cardiac CAT activities (P < 0.0001 and P = 0.05 respectively). CAT activity positively correlated with the total and free carnitine levels in both liver and heart (r = 0.764, r = 0.785 and r = 0.700, r = 0.519, respectively). Liver mitochondrial respiratory control ratio, ADP:O ratio and oxygen uptake were similar in both control and supplemented groups. These results demonstrate that in oophorectomized rats, dietary DHEAS supplementation increases the liver and heart L-carnitine levels and CAT activities. In conclusion, DHEAS may modulate L-carnitine level and CAT activity in estrogen deficient rats. The potential role of DHEAS in the regulation of fatty acid oxidation in postmenopausal women is worthy of investigation.

Protection of the ischaemic myocardium by L-propionylcarnitine: effects on the recovery of cardiac output after ischaemia and reperfusion, carnitine transport, and fatty acid oxidation.

The effects of L-propionylcarnitine on the recovery of cardiac contractile performance after global ischaemia and reperfusion were studied in isolated perfused rat hearts. The addition of either 5.5 or 11 mmol X litre-1 L-propionylcarnitine significantly improved the recovery of cardiac output, left ventricular pressure, and dP/dt after 90 min of ischaemia and 15 min of reperfusion. Myocardial adenosine triphosphate and creatine phosphate concentrations were significantly higher in the L-propionylcarnitine treated hearts than in controls, but the concentrations of long chain acyl carnitine and coenzyme A were unaffected. The protecting effects of L-propionylcarnitine were compared with those of L-carnitine and L-acetylcarnitine. A 11 mmol X litre-1 dose of L-propionylcarnitine and L-acetylcarnitine significantly improved the recovery of cardiac output after 90 min of ischaemia and 15 min of reperfusion, but L-carnitine did not. L-Propionylcarnitine was the most protective agent. The effects of these derivatives on L-3H-carnitine transport and 14C-palmitate oxidation were also measured. All of these derivatives competitively inhibited L-3H-carnitine transport in isolated cardiac myocytes, but L-propionylcarnitine was the most potent. Carnitine and L-propionylcarnitine stimulated palmitate oxidation in the homogenate, whereas L-acetylcarnitine inhibited it. In myocytes only L-propionylcarnitine affected palmitate oxidation. These data show that L-propionylcarnitine protects the ischaemic myocardium. Its protection is greater than that for L-carnitine or L-acetylcarnitine, and the difference in effectiveness may relate to the rate of transport into the cells and the effects on fatty acid utilisation.

L-carnitine improvement of cardiac function is associated with a stimulation in glucose but not fatty acid metabolism in carnitine-deficient hearts.

OBJECTIVES: Increasing myocardial carnitine content can improve heart function in patients with carnitine deficiency. We were interested in determining the effects of L-carnitine on cardiac function and substrate metabolism in a rat model of carnitine deficiency. METHODS: Carnitine deficiency was induced in male Sprague-Dawley rats by supplementing the drinking water with 20 mM sodium pivalate. Control animals received an equimolar concentration of sodium bicarbonate. Following treatment, cardiac function and myocardial substrate utilization were determined in isolated working hearts perfused with glucose and relevant levels of fatty acids. To increase tissue levels of carnitine, hearts were perfused with 5 mM L-carnitine for a period of 60 min. RESULTS: Hearts from sodium pivalate-treated animals demonstrated a 60% reduction in total heart carnitine content, depressions in cardiac function and rates of palmitate oxidation, and elevated rates of glycolysis compared to control hearts. Treatment with L-carnitine increased total carnitine content and reversed the depression in cardiac function seen in carnitine-deficient hearts. However, this was not associated with any improvement in palmitate oxidation. Rates of glycolysis and glucose oxidation, on the other hand, were increased with L-carnitine. CONCLUSIONS: Our findings indicate that acute L-carnitine treatment is of benefit to cardiac function in this model of secondary carnitine deficiency by increasing overall glucose utilization rather than normalizing fatty acid metabolism.

Malonyl CoA inhibition of carnitine palmityltransferase in rat heart mitochondria.

The effects of malonyl CoA on carnitine palmityltransferase I (CPT-I), fatty acid-supported state 3 respiration, and carnitine reversal of palmityl CoA inhibition of state 3 respiration and of the adenine nucleotide translocator, were studied in isolated rat heart mitochondria. Malonyl CoA was a potent competitive inhibitor of CPT-I with an I50 of 0.8 microM. Fasting did not affect CPT-I activity or the I50 value of malonyl CoA. Malonyl CoA inhibited fatty acid-supported respiration and prevented carnitine from reversing the inhibition of the adenine nucleotide translocator by palmityl CoA. These findings suggest that malonyl CoA may affect fatty acid oxidation in the heart.

The effect of enteral carnitine administration in humans.

We previously determined that the L-carnitine uptake by human duodenal tissue occurs by both active (KT 558 mumol/L) and passive mechanisms. The effects of enteral carnitine was studied in humans. A hamburger meal (345 mumol total carnitine) induced peak jejunal fluid free (unesterified) and short-chain acylcarnitine concentrations (SCAC) of 209 and 130 mumol/L, respectively. Plasma carnitine concentrations and the percent renal reabsorption remained unchanged. By contrast, a pharmacologic dose of free carnitine (25,298 mumol) raised peak intraluminal free and SCAC to 20,660 and 4204 mumol/L. Plasma total carnitine concentrations doubled to 93 mumol/L, and the percent renal reabsorption of free and SCAC declined to 76% and 52%, respectively. In triple-lumen perfusions, 200 mumol carnitine/L was absorbed at 484 nmol.min-1.30 cm-1 jejunum, a rate sufficient for prandial but not pharmacologic assimilation. Our findings indicate that absorption of physiologic and pharmacologic amounts of carnitine occurs predominantly by active transport and passive diffusion, respectively.

Nearly fatal muscle carnitine deficiency with full recovery after replacement therapy.

A 23-year-old woman became quadriplegic and respirator-dependent after 18 years of weakness and rhabdomyolysis. Her muscle tissue and that of a deceased sister contained lipid-laden fibers. Treatment with D,L-carnitine 4 grams per day was followed by a dramatic improvement within 10 days. Muscle function was normal at 8 months and has remained so during 3 subsequent years of L-carnitine 3 grams per day. Pretreatment muscle biopsy had documented low levels of free carnitine and short-chain acylcarnitine compounds. Carnitine palmityltransferase was slightly elevated. The asymptomatic parents had low-normal muscle carnitine levels, slight increase in muscle fiber lipid droplets, osmiophilic lipid-laden Schwann's cell vacuoles, and myelin lamellae with different periodicities.

Fatal rhabdomyolysis following influenza infection in a girl with familial carnitine palmityl transferase deficiency.

Severe rhabdomyolysis following an influenza B infection developed in a previously well 13-year-old girl. There was no history of trauma. Her course was complicated by episodes of severe hyperkalemia, hypocalcemia, hyperphosphatemia, and myoglobinuria. Renal failure, hypertension, and life-threatening arrhythmias developed; she died. Muscle biopsy revealed that this girl had carnitine palmityl transferase deficiency. An asymptomatic sister was demonstrated to have the same disorder. Although carnitine palmityl transferase deficiency is usually associated with mild bouts of rhabdomyolysis that become apparent only in adulthood, severe forms of this disorder may be seen in children. Life-threatening rhabdomyolysis and myoglobinuria may follow any infection associated with decreased intake. If carnitine palmityl transferase deficiency is diagnosed in a proband, other siblings should be evaluated so that proper preventative measures can be undertaken to help prevent the development of symptoms in susceptible individuals who have not been recognized to have the disease.

Defective myocardial carnitine metabolism in congestive heart failure secondary to dilated cardiomyopathy and to coronary, hypertensive and valvular heart diseases.

Reduced myocardial carnitine concentrations in the explanted heart and elevated plasma levels have been found in patients undergoing heart transplant for end-stage congestive heart failure (CHF). To evaluate a possible loss of myocardial carnitine in less severe stages of CHF, total myocardial carnitine levels were compared in right ventricular endomyocardial biopsies from 28 patients with mild, moderate and severe dilated cardiomyopathy, 8 patients with CHF of different origin and 13 normal control subjects. If possible, free myocardial carnitine and free and total plasma carnitine were also determined. For the first time, myocardial carnitine levels have been measured in endomyocardial biopsies from 13 normal human hearts (control values: 9.9 +/- 0.8 nmol/mg noncollagen protein). In comparison with these control values, total myocardial carnitine was significantly reduced in patients with dilated cardiomyopathy (6.1 +/- 0.5 nmol/mg noncollagen protein, p less than 0.0001), and CHF of other origins (6.6 +/- 1.1 nmol/mg noncollagen protein, p less than 0.02). Free myocardial carnitine concentrations in dilated cardiomyopathy (4.6 +/- 0.4 nmol/mg noncollagen protein) and CHF of different origin (4.4 +/- 0.5 nmol/mg noncollagen protein) were also significantly different from control values (control values: 9.7 +/- 0.7 nmol/mg noncollagen protein, p less than 0.0001 and p less than 0.005 for both groups). The loss of free and total myocardial carnitine was comparable in dilated cardiomyopathy and CHF due to other diseases. In contrast, plasma free and total carnitine levels in the CHF patients were significantly elevated (67 +/- 5.5 mumol/liter, control values 41 +/- 3.7 mumol/liter, p less than 0.005). Alterations in myocardial carnitine metabolism represent nonspecific biochemical markers in CHF with yet unknown consequences for myocardial function.

Improved pacing tolerance of the ischemic human myocardium after administration of carnitine.

The possibility that DL-carnitine has a protective effect during myocardial ischemia was evaluated by performing two rapid coronary sinus pacing studies 15 minutes apart in 21 patients with coronary artery disease. Eleven patients received DL-carnitine (20 or 40 mg/kg) before the second pacing study. The treated group had a significant increase in mean heart rate (12.5 beats/min, P less than 0.001), pressure-rate product (1,912 units, P less than 0.01) and pacing duration (3.2 minutes, P less than 0.001) after the administration of carnitine. The treated group also had improvements in percent myocardial lactate extraction (8.8 percent increase, P less than 0.001) and left ventricular end-diastolic pressure (a decrease of 5.3 mm Hg, P less than 0.05). There was significantly less S-T segment depression during the second pacing period in both the untreated and treated groups. The results of this study suggest that in ischemic human hearts with reasonably well preserved left ventricular function, DL-carnitine may improve the tolerance for stress associated with an increase in heart rate and pressure-rate product.

Improvement of myocardial function in diabetic rats after treatment with L-carnitine.

The effects of L-carnitine administration on the severity of diabetes were investigated. Serum glucose, free fatty acids (FFA), triglycerides, and ketones from diabetic and normal rats injected for 2 weeks with 3 g/kg/d of either L-carnitine or saline were assayed. Hearts were analyzed for carnitine and long-chain acyl coenzyme A. L-carnitine treatment to diabetic rats significantly reduced serum glucose, FFA, triglycerides, and ketones. In nondiabetic rats, carnitine increased serum ketones while FFA and triglycerides were decreased. L-carnitine treatment to diabetic rats prevented a decrease in myocardial total carnitine content. Long-chain acyl carnitine increased while long-chain acyl coenzyme A decreased. In another experiment, L-carnitine administration (750 mg/kg/d for 14 days) significantly improved the recovery of cardiac output after 60, 90, and 120 minutes of ischemia in diabetic perfused hearts. These results suggest that L-carnitine therapy may reduce the severity of diabetes mellitus and improve myocardial performance.

Fatty acid oxidation and cardiac function in the sodium pivalate model of secondary carnitine deficiency.

Carnitine-deficiency syndromes are often associated with alterations in lipid metabolism and cardiac function. The present study was designed to determine whether this is also seen in an experimental model of carnitine deficiency. Carnitine deficiency was induced in male Sprague-Dawley rats supplemented with sodium pivalate for 26 to 28 weeks. This treatment resulted in nearly a 60% depletion of myocardial total carnitine content as compared with control hearts. When isolated working hearts from these animals were perfused with 5.5 mmol/L glucose and 1.2 mmol/L palmitate and subjected to incremental increases in left-atrial filling pressures, cardiac function remained dramatically depressed. The effects of carnitine deficiency on glucose and palmitate utilization were also assessed in hearts perfused at increased workload conditions. At this workload, function was depressed in carnitine-deficient hearts, as were rates of 1.2-mmol/L [U-14C]-palmitate oxidation, when compared with control hearts (544 +/- 37 vs 882 +/- 87 nmol/g dry weight.min, P < .05). However, glucose oxidation rates from 5.5 mmol/L [U-14C]-glucose were slightly increased in carnitine-deficient hearts. To determine whether the depressed fatty acid oxidation rates were a result of reduced mechanical function in carnitine-deficient hearts, the workload of hearts was reduced. Under these conditions, mechanical function was similar among control and carnitine-deficient hearts. Palmitate oxidation rates were also similar in these hearts (526 +/- 69 v 404 +/- 47 nmol/g dry weight.min for control and carnitine-deficient hearts, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Protection of mitochondrial and heart function by amino acids after ischemia and cardioplegia.

The effects of amino acids in protecting against ischemic/reperfusion injury were tested in two experimental models: the isolated perfused rat heart subjected to 21 min of zero flow ischemia (37 degrees) followed by 40 min of reperfusion and the isolated perfused rabbit heart subjected to 300 min of cardioplegic arrest (29 degrees) followed by 60 min of reperfusion. In both cases, the addition of amino acids to the perfusion medium significantly improved the recovery of cardiac contractile function. The protective effects of amino acids were associated with a preservation of mitochondrial respiratory activity. These findings suggest that amino acids by replenishing mitochondrial matrix levels of critical TCA cycle substrates, such as malate, stimulate mitochondrial respiration and thereby enhance the recovery of heart function.

Carnitine protection against adriamycin-induced cardiomyopathy in rats.

The effects of chronic adriamycin toxicity on myocardial carnitine content and contractile function were studied in rats, along with potential protective effects of L-carnitine administration. Cardiomyopathy was induced over a 6- to 7-week period by weekly intravenous injections of adriamycin, 2 mg/kg. In vivo myocardial tissue levels of carnitine were not significantly changed by adriamycin, but plasma levels were elevated. Cardiac output was depressed in isolated perfused hearts from adriamycin-treated rats perfused with 11 mM glucose. In a second experiment, 4-week-old male rats were divided into four groups: saline-treated control, L-carnitine-treated control, saline-treated adriamycin, and L-carnitine-treated adriamycin. L-Carnitine was given intraperitoneally each day at a dose of 500 mg/kg. Myocardial histology and ultrastructure were analyzed. Cardiac performance was determined in hearts perfused with 1.2 mM palmitate and 5.5 mM glucose. Hearts from saline-treated adriamycin rats showed histopathological changes and a significantly diminished cardiac output at various preloads when compared to saline-treated controls. Daily intraperitoneal L-carnitine reduced histopathological alterations and improved cardiac performance.

Sodium pivalate reduces cardiac carnitine content and increases glucose oxidation without affecting cardiac functional capacity.

This study determined how selected functional, metabolic, and contractile properties were impacted by sodium pivalate, a compound which creates a secondary carnitine deficiency. Young male rats received either sodium pivalate (20 mM, PIV) or sodium bicarbonate (20 mM, CONTR) in their drinking water. After 11-12 weeks cardiac function and glucose oxidation rates were measured in isolated, perfused working heart preparations. Hearts were also analyzed for carnitine content, activities of hexokinase (HK), citrate synthase (CS), and B-hydroxyacyl CoA dehydrogenase (HOAD), and myosin isoenzyme distribution. Sodium pivalate treatment significantly reduced cardiac carnitine content and increased glucose oxidation but did not alter cardiac functional capacity. HK activity was increased in the PIV group (p < 0.05), and HOAD activity decreased (p < 0.05). CS activity and myosin isoform distribution (VI > 85%) remained unchanged. These results demonstrate that pivalate treatment of this duration and the accompanying carnitine deficiency shift cardiac substrate utilization without compromising cardiac functional capacity.

Myocardial L-carnitine deficiency in a family of dogs with dilated cardiomyopathy.

Dilated cardiomyopathy in a family of dogs was found to be associated with decreased myocardial L-carnitine concentrations, when compared with those in control dogs. In 2 affected dogs, treatment with high doses of L-carnitine was associated with increased myocardial L-carnitine concentration and greatly improved health and myocardial function. Withdrawal of L-carnitine supplementation from these dogs resulted in development of myocardial dysfunction and clinical signs of dilated cardiomyopathy.