Reversible carnitine palmitoyltransferase inhibitors with broad chemical diversity as potential antidiabetic agents.

A series of carnitine related compounds of general formula XCH(2)CHZRCH(2)Y were evaluated as CPT I inhibitors in intact rat liver (L-CPT I) and heart mitochondria (M-CPT I). Derivative 27 (ZR = -HNSO(2)R, R = C(12), X = trimethylammonium, Y = carboxylate, (R) form) showed the highest activity (IC(50) = 0.7 microM) along with a good selectivity (M-CPT I/L-CPTI IC(50) ratio = 4.86). Diabetic db/db mice treated orally with 27 showed a significant reduction of serum glucose levels.

Seminal carnitine and acetylcarnitine content and carnitine acetyltransferase activity in young Maremmano stallions.

The reproductive characteristics and seminal carnitine and acetylcarnitine content as well as carnitine acetyltransferase activity of young Maremmano stallions (n=25) are reported. The stallions were subjected to semen collection in November and January; in each trial two ejaculates were collected 1h apart. The total motile morphologically normal spermatozoa (TMMNS) and the progressively motile spermatozoa at collection and during storage at +4 degrees C were evaluated. Seminal L-carnitine (LC), acetylcarnitine (AC), pyruvate and lactate were measured using spectrophotometric methods, whereas carnitine acetyltransferase activity was measured by radioenzymatic methods. Since there were no major significant differences in seminal and biochemical characteristics between the November and January trials, data were also pooled for the first and second ejaculates. Significant differences (P<0.001) were observed between the first and second ejaculates for sperm count (0.249+/-0.025 versus 0.133+/-0.014x10(9)/ml), total number spermatozoa by ejaculate (12.81+/-1.23 versus 6.36+/-0.77x10(9)), progressively motile spermatozoa (48.6+/-3.0 versus 52.6+/-3.0%) and TMMNS (3.35+/-0.50 versus 2.02+/-0.37x10(9)). In the raw semen the LC and AC were significantly higher in the first ejaculate than in the second (P<0.001), whereas, pyruvate and pyruvate/lactate ratio were higher in the second ejaculate (P<0.05). Seminal plasma AC and LC concentrations resulted higher in the first ejaculate (P<0.001). The pyruvate/lactate ratio was higher in the second ejaculate (P<0.05). Both raw semen and seminal plasma LC and AC concentrations were positively correlated with spermatozoa concentration (P<0.01); in raw semen AC was also correlated to TMMNS (P<0.01). Lactate levels of raw semen was correlated to progressively motile spermatozoa after storage (P<0.01). In the second ejaculate, significant correlations were also observed among AC/LC ratio in raw semen and progressively motile spermatozoa after 48 and 72h of refrigeration. Furthermore, AC levels were correlated to lactate concentration. The positive correlation between LC, AC and spermatozoa concentration, and between AC and TMMNS indicated carnitine as potential semen quality marker. Moreover, the correlation between AC/LC ratio and progressive spermatozoa motility after refrigeration, suggests that carnitine may contribute towards improving the maintenance of spermatozoa viability during in vitro storage.

L-carnitine and acetyl-L-carnitine in human nerves from normal and diabetic subjects.

Marked reduction in the contents of L-carnitine and acetyl-L-carnitine has been reported in peripheral nerves of rats with experimental diabetes. Since these substances have been claimed to improve a number of signs and symptoms of peripheral neuropathy in controlled clinical trials, this study was aimed at assessing whether nerves from diabetic subjects would also reveal similar decrease in the concentration of L-carnitine and acetyl-L-caritine. To this end, these substances were measured in nerves obtained from 11 patients with diabetic neuropathy, 13 patients with ischemic non-diabetic neuropathy, and 12 normal controls. Nerves from patients with either diabetic neuropathy and ischemic non-diabetic neuropathy showed levels of both carnitines lower than those from normal controls. However, differences among the three groups were not statistically significant, indicating that a reduction in these amino acids probably represents only a co-factor in the development of the variegated clinical picture of human diabetic neuropathy.

Carnitine and derivatives in the central nervous system of chick embryo.

1. Carnitine contents and the activity of carnitine acetyltransferase in the egg, in the embryo, and in different brain areas of central nervous system in chick embryo were determined in the course of development. 2. The egg showed low levels of free carnitine and acetylcarnitine. 3. In the whole embryo, at first stages of development, long chain acylcarnitine and acetylcarnitine were the best represented classes of carnitines. 4. In the brain regions acetylcarnitine levels, high at the first days, showed a continual decrease during development. 5. The activity of carnitine acetyltransferase increased and was totally related to development.

Tissue lipid accumulation by L-aminocarnitine, an inhibitor of carnitine-palmitoyltransferase-2. Studies in intact rats and isolated mitochondria.

Tissues of fasted animals treated with L-aminocarnitine (L-3-amino-4-trimethylaminobutyrate) showed an accumulation of long-chain acylcarnitines and triacylglycerols. Blood levels of free fatty acids, long-chain acylcarnitines and triacylglycerol-rich lipoproteins were found to be increased, whereas glucose was reduced. The liver mitochondria isolated from rats treated with L-aminocarnitine utilized both pyruvate and succinate normally, but were not able to oxidize palmitoylcarnitine. In vitro oxidation of palmitoylcarnitine by liver mitochondria was inhibited by L-aminocarnitine in a concentration-dependent fashion in contrast to succinate and pyruvate oxidation which was not modified. L-aminocarnitine proved to be a potent and selective inhibitor (IC50 = 805 nM) of the carnitine palmitoyltransferase isoenzyme, located on the inner side of the mitochondrial inner membrane (CPT2). The activity of the carnitine palmitoyltransferase isoenzyme located on the mitochondrial outer membrane inhibitable by malonyl-CoA (IC50 = 19 microM), was not inhibited by 0.8 microM L-aminocarnitine. Both in vitro and in vivo effects of L-aminocarnitine suggest that the substance has a specific and potent inhibitory action on CPT2. Its in vivo inhibition results in a dramatic increase of long-chain acylcarnitines in various organs, that it is why this increase can be considered a very good marker of CPT2 inhibition. A markedly altered lipid metabolism was observed.

Levels of carnitines in brain and other tissues of rats of different ages: effect of acetyl-L-carnitine administration.

Male Sprague-Dawley rats, aged 2, 5, 16, 20 and 30 months and normally fed, were used for determination of carnitines in the brain, serum, heart, tibial muscle, liver and urine. With respect to 5-month-old animals, those aged 30 months exhibited a statistically significant decrement of total carnitine levels in the brain, serum, heart and tibial muscle, accompanied by a dramatic increment in the liver. This suggests impaired net transport of carnitines from the liver to the blood in old age. Urinary excretion was similar in the two age groups. One group received from 5 months on daily 75 mg/kg acetyl-L-carnitine in drinking water. At 20 months, the treated animals showed levels of brain, heart and serum carnitines similar to those of 5-month-old animals. The recovery of brain, heart and serum carnitines in the old animals treated with acetyl-L-carnitine indicates that intestinal absorption and tissue uptake remain sufficiently efficient in the course of aging. The lower level of brain lipofuscins due to acetyl-L-carnitine treatment may be related to the effect of the compound on acetylcholine metabolism.

L-carnitine effect on plasma lipoproteins of hyperlipidemic fat-loaded rats.

The effect of oral L-carnitine administration to rats fed olive oil has been studied. Carnitine significantly decreased triglyceride, cholesterol and phospholipid levels. Particularly, the levels of chylomicron and very low density lipoproteins in the blood were lowered. Low density lipoprotein levels were not affected, and high density lipoproteins were found to be decreased by 20%. Because carnitine did not change the composition of chylomicron and very low density lipoproteins fraction or affect the gastrointestinal triglyceride residue (about 1/3 of the original load), an effect of carnitine on hepatic fatty acid handling is most likely. The lowering of plasma free fatty acid levels by carnitine administration is in favor of an effect of carnitine on fatty acid handling. The effect on the liver is illustrated by the study of acetoacetate formation in in vitro perfused livers from previously olive oil loaded +/- carnitine-treated rats. Carnitine pretreatment stimulated ketogenesis. It is speculated that carnitine administration, by promoting beta-oxidation, lowers the production of very low density lipoproteins. This may be accomplished partly by an increase in the hepatic level of fatty acid binding protein, which also has been observed.

The effect of exogenous L-carnitine on biochemical parameters in serum and in heart of the hyperlipidaemic rat.

In previous experiments we have demonstrated that L-carnitine administration is capable of reducing olive oil-induced lipidaemia in the rat. In the present study we determined the effect of L-carnitine on the levels of (acyl)carnitines in heart and serum in addition to its effect on serum levels of lipids and ketone bodies after olive oil gavage feeding. L-carnitine was found to reduce the level of myocardial long-chain acylcarnitine which was increased by the olive oil treatment. It also increased the levels of carnitine and acid soluble acylarnitines in both heart and serum. L-carnitine administration caused a clearcut decrease of olive oil-induced lipidaemia and ketonaemia. These effects of added L-carnitine strongly suggest that the stimulation of the beta-oxidation in the mitochondria (at the expense of extra mitochondrial triglycerceride synthesis) is suboptimal after fat loading.