The involvement of the polyamines binding sites at the NMDA receptor in creatine-induced spatial learning enhancement.

Achievements made over the last years have highlighted the important role of creatine in health and disease. However, studies of its effect on cognition function have been limited. In the present study, we investigated the effect of creatine on early consolidation of the spatial learning in rats. Statistical analysis showed that intrahippocampal administration of creatine (2.5 and 7.5 nmol/hippocampus) (post-training) decreased the latency for scape and mean number of errors in Barnes maze test. The involvement of polyamine binding site at NMDA receptor in creatine-induced spatial learning enhancement was investigated by co-administration of arcaine (0.02 nmol/hippocampus) or spermidine (0.02 nmol/hippocampus) with creatine (2.5 nmol/hippocampus) (post-training). Statistical analysis revealed that creatine-induced spatial learning enhancement was reverted by co-administration of arcaine (0.02 nmol/hippocampus) and intensified by spermidine (0.02 nmol/hippocampus). These results provide evidence that creatine not only seem to be involved in energy metabolism but may also play an important role in early consolidation of spatial learning in hippocampus which participation of polyamines binding site at the NMDA receptor.

Pharmacokinetics of creatine.

Research has demonstrated that creatine supplementation has some therapeutic benefit with respect to muscle function and more recently neurological function. Despite the growing body of literature on the pharmacologic effect of creatine, very little is known about the disposition of creatine after supraphysiologic doses. The movement of creatine throughout the body is governed by transport processes which impact the absorption of creatine from the intestine, clearance of creatine from the kidney, and access of creatine to target tissues. With repeated doses of creatine, it appears that the clearance of creatine decreases mainly due to the saturation of skeletal muscle stores. Insulin and insulin-stimulating foods appear to enhance muscle uptake of creatine but at the same time, high carbohydrate meals may slow the absorption of creatine from the intestine. Little is known about creatine disposition in special populations including the elderly and patients with neuromuscular disease. Knowledge of creatine disposition in these clinically relevant populations can help remove some of the guess work of dose selection during clinical trials.

Study of hypothalamic metabolism in cluster headache by proton MR spectroscopy.

The authors used 1H-MRS to investigate hypothalamic metabolism in 26 patients with cluster headache (CH) and 12 healthy subjects. Hypothalamic N-acetylaspartate/creatine was reduced in patients with CH vs controls (p < 0.01). Dividing the patients into episodic CH outside- and in-cluster periods and chronic CH, the hypothalamic N-acetylaspartate/creatine in all three subgroups of patients was reduced. The reduction of the neuronal marker N-acetylaspartate is consistent with hypothalamic neuronal dysfunction in patients with CH.

Anti-inflammatory activity of creatine supplementation in endothelial cells in vitro.

1 Creatine (CR) supplementation augments muscle strength in skeletal muscle cells by increasing intracellular energy pools. However, the effect of CR supplementation on endothelial cells remains to be clarified. 2 In this study, we investigated whether CR supplementation had any anti-inflammatory activity against human pulmonary endothelial cells in culture. 3 We confirmed that supplementation with 0.5 mM CR significantly increased both intracellular CR and phosphocreatine (PC) through a CR transporter while keeping intracellular ATP levels constant independent of CR supplementation and a CR transporter antagonist. 4 In the assay system of endothelial permeability, supplementation with 5 mM CR significantly suppressed the endothelial permeability induced by serotonin and H(2)O(2). 5 In cell adhesion experiments, supplementation with 5 mM CR significantly suppressed neutrophil adhesion to endothelial cells. 6 In the measurement of adhesion molecules, CR supplementation with more than 0.5 mM CR significantly inhibited the expressions of ICAM-1 and E-selectin on endothelial cells, and the inhibition was significantly suppressed by an adenosine A(2A) receptor antagonist. 7 The present study suggests that CR supplementation has anti-inflammatory activities against endothelial cells.

Muscle creatine uptake and creatine transporter expression in response to creatine supplementation and depletion.

The total creatine pool size [Cr(total); creatine (Cr) + phosphocreatine (PCr)] is crucial for optimal energy utilization in skeletal muscle, especially at the onset of exercise and during intense contractions. The Cr(total) likely is controlled by long-term modulation of Cr uptake via the sodium-dependent Cr transporter (CrT). To test this hypothesis, adult male Sprague-Dawley rats were fed 1% Cr, their muscle Cr(total) was reduced by approximately 85% [1% beta-guanidinoproprionic acid (beta-GPA)], or their muscle Cr(total) was repleted (1% Cr after beta-GPA depletion). Cr uptake was assessed by skeletal muscle (14)C-Cr accumulation to Cr and PCr by using hindlimb perfusion, and CrT protein content was assessed by Western blot. Cr uptake rate decreased with dietary Cr supplementation in the white gastrocnemius (WG; 45%) only. Depletion of muscle Cr(total) to approximately 15% of normal increased Cr uptake in the soleus (21%) and red gastrocnemius (22%), corresponding to 70-150% increases in muscle CrT content. In contrast, the inherently lower Cr uptake rate in the WG was unchanged with depletion of muscle Cr(total) even though CrT band density was increased by 230%. Thus there was no direct relationship between apparent muscle CrT abundance and Cr uptake rates. However, Cr uptake rates scaled inversely with decreases in muscle Cr(total) in the high-oxidative muscle types but not in the WG. This implies that factors controlling Cr uptake are different among fiber types. These observations may help explain the influence of initial muscle Cr(total), time dependency, and variations in muscle Cr(total) accumulation during Cr supplementation.

Effect of creatine manipulation on fast-twitch skeletal muscle of the mouse.

1. The effect of short-term, reversible alteration of muscle total creatine content (Crtot) on force development was sought in fast-twitch extensor digitorum longus (EDL) muscles of female mice. 2. Three in vivo interventions were investigated: 1% creatine-supplementation, treatment with the creatine-uptake inhibitor beta-guanidino propionic acid (beta-GPA; 1%) or beta-GPA treatment followed by creatine supplementation for 5 days. 3. The Crtot of isolated muscles, determined using reverse-phase high-performance liquid chromatography, was 133 +/- 38 mmol/kg dry in 11 EDL control muscles and was not significantly affected by dietary creatine-supplementation (152 +/- 25 mmol/kg dry; n = 8). Significant creatine depletion was observed in the beta-GPA-fed group (65 +/- 6 mmol/kg dry; n = 9) and this was reversed by 5 days of creatine supplementation (133 +/- 21 mmol/kg dry; n = 10). 4. Creatine depletion did not affect maximum tetanic stress. However, when muscle creatine was restored by creatine supplementation, a substantial increase in tetanic stress was observed. Significant correlations were observed between Crtot and maximum tetanic stress (r = 0.56) and relaxation rate (r = 0.43). The enhancement of force was not due to effects of creatine on muscle fibre type because neither mechanical tests of fibre characteristics nor the fibre types of the muscles were affected. 5. We conclude that, in muscles that contain large numbers of fast-twitch fibres, maximum tetanic stress is determined, in part, by muscle creatine stores.

Supplemental creatine may decrease serum homocysteine and abolish the homocysteine 'gender gap' by suppressing endogenous creatine synthesis.

Creatine synthesis is responsible for the large majority of the methyl group transfers in normal hepatic metabolism, and is significantly greater in men than in women. Since methyl group depletion impairs the efficiency of homocysteine disposal, it is believed that the comparatively high rate of creatine synthesis in men is primarily responsible for the 'gender gap' in homocysteine levels. Adequate supplemental intakes of creatine can suppress creatine synthesis by inhibiting expression of the enzyme arginine-glycine transamidase. Thus, it is proposed that creatine supplementation may represent a practical strategy for decreasing plasma homocysteine levels.

Cyclosporin A inhibits creatine uptake by altering surface expression of the creatine transporter.

The immunosuppressive drug cyclosporin A (CsA) inhibited the hCRT-1 cDNA-induced creatine uptake in Xenopus oocytes and the endogenous creatine uptake in cultured C(2)C(12) muscle cells in a dose- and time-dependent manner. FK506, another potent immunosuppressant, was unable to mimic the effect of CsA suggesting that the inhibitory effect of CsA was specific. To delineate the mechanism underlying, we investigated the effect of CsA on the K(m) and V(max) of creatine transport and also on the cell surface distribution of the creatine transporter. Although CsA treatment did not affect the K(m) (20-24 microm) for creatine, it significantly decreased the V(max) of creatine uptake in both oocytes and muscle cells. CsA treatment reduced the cell surface expression level of the creatine transporter in the muscle cells by approximately 60% without significantly altering its total expression level, and the reduction in the cell surface expression paralleled the decrease in creatine uptake. Taken together, our results suggest that CsA inhibited creatine uptake by altering the surface abundance of the creatine transporter. We propose that CsA impairs the targeting of the creatine transporter by inhibiting the function of an associated cyclophilin, resulting in an apparent loss in surface expression of the creatine transporter. Our results also suggest that prolonged exposure to CsA may result in chronically creatine-depleted muscle, which may be a cause for the development of CsA-associated clinical myopathies in organ transplant patients.

Caffeine counteracts the ergogenic action of muscle creatine loading.

This study aimed to compare the effects of oral creatine (Cr) supplementation with creatine supplementation in combination with caffeine (Cr+C) on muscle phosphocreatine (PCr) level and performance in healthy male volunteers (n = 9). Before and after 6 days of placebo, Cr (0.5 g x kg-1 x day-1), or Cr (0.5 g x kg-1 x day-1) + C (5 mg x kg-1 x day-1) supplementation, 31P-nuclear magnetic resonance spectroscopy of the gastrocnemius muscle and a maximal intermittent exercise fatigue test of the knee extensors on an isokinetic dynamometer were performed. The exercise consisted of three consecutive maximal isometric contractions and three interval series of 90, 80, and 50 maximal voluntary contractions performed with a rest interval of 2 min between the series. Muscle ATP concentration remained constant over the three experimental conditions. Cr and Cr+C increased (P < 0.05) muscle PCr concentration by 4-6%. Dynamic torque production, however, was increased by 10-23% (P < 0.05) by Cr but was not changed by Cr+C. Torque improvement during Cr was most prominent immediately after the 2-min rest between the exercise bouts. The data show that Cr supplementation elevates muscle PCr concentration and markedly improves performance during intense intermittent exercise. This ergogenic effect, however, is completely eliminated by caffeine intake.

Preincubation with creatine enhances levels of creatine phosphate and prevents anoxic damage in rat hippocampal slices.

We have investigated the relationship between energy metabolism, NMDA-receptor antagonism, and anoxic damage in vitro. Anoxic damage was assessed by measuring protein synthesis, defined as the incorporation of [14C]lysine into perchloric acid-insoluble tissue extracts. The concentrations of energy metabolites were measured by ion-exchange HPLC. Anoxia caused an inhibition of protein synthesis, a reduction in phosphocreatine and adenosine triphosphate, and extensive neuronal damage. The reduction of protein synthesis depended on the duration of anoxia and the time allowed for recovery. Preincubation with the creatine dose-dependently (0.03-3 mmol/L) increased baseline levels of phosphocreatine, reduced the anoxia-induced decline in phosphocreatine and adenosine triphosphate, prevented the impairment of protein synthesis, and reduced neuronal death. Incubation with (R,S)-3-guanidinobutyric acid, a synthetic analogue of creatine that cannot be phosphorylated, did not prevent the anoxia-induced impairment of protein synthesis and did not enhance the levels of phosphocreatine and adenosine triphosphate. Incubation with a combination of both creatine and the noncompetitive NMDA antagonist MK-801 provided complete protection. These results indicate that energy status is a major factor controlling anoxic damage in the rat hippocampal slice.

Age dependence of myosin heavy chain transitions induced by creatine depletion in rat skeletal muscle.

This study was designed to test the hypothesis that myosin heavy chain (MHC) plasticity resulting from creatine depletion is an age-dependent process. At weaning (age 28 days), rat pups were placed on either standard rat chow (normal diet juvenile group) or the same chow supplemented with 1% wt/wt of the creatine analogue beta-guanidinopropionic acid [creatine depletion juvenile (CDJ) group]. Two groups of adult rats (age approximately 8 wk) were placed on the same diet regimens [normal diet adult and creatine depletion adult (CDA) groups]. After 40 days (CDJ and normal diet juvenile groups) and 60 days (CDA and normal diet adult groups), animals were killed and several skeletal muscles were removed for analysis of creatine content or MHC distribution. In the CDJ group, creatine depletion (78%) was accompanied by significant shifts toward expression of slower MHC isoforms in two slow and three fast skeletal muscles. In contrast, creatine depletion in adult animals did not result in similar shifts toward slow MHC isoform expression in either muscle type. The results of this study indicate that there is a differential effect of creatine depletion on MHC transitions that appears to be age dependent. These results strongly suggest that investigators contemplating experimental designs involving the use of the creatine analogue beta-guanidinopropionic acid should consider the age of animals to be used.

Bioenergetic consequences of cardiac phosphocreatine depletion induced by creatine analogue feeding.

To further evaluate the bioenergetic role of phosphocreatine, we assessed several parameters in normal and depleted rat hearts. Rats were fed (8 weeks) a diet containing either 1% beta-guanidinoproprionic acid or 2% beta-guanidinobutyric acid (beta-GBA), resulting in an 80% phosphocreatine depletion compared to controls. Left ventricular pressure-volume curves were obtained to determine contractile function. At any volume, the developed pressure in depleted hearts was lower than in controls. At the plateau, the rate-pressure product was between 37-45% lower: 34,000 (beta-GBA), 30,174 (beta-guanidinoproprionic acid) versus 54,400 (control). 31P NMR spectroscopy on beta-GBA-treated hearts obtained the [ATP] and [phosphocreatine], which with saturation transfer estimated the rates of creatine kinase and ATP production. In depleted hearts, the rate constant for ATP synthesis from phosphocreatine was increased 33%. However, the flux was 72% lower. ATP production from ADP and Pi were similar under normal conditions, in spite of higher rates of oxygen consumption in the depleted hearts. The addition of 50 mM creatine to control perfusate had no effect on function or high energy phosphates. In contrast, a 28% increase in function and a 52% increase in [phosphocreatine] was seen in beta-GBA hearts. There was a marked increase in free [ADP] in beta-GBA hearts, resulting in a lower estimated ATP phosphorylation potential. Overall, the results suggest that phosphocreatine may play an important function by optimizing the thermodynamics of cardiac high energy phosphate utilization.

[Cardiac relaxation and distensibility in energy deficit]. / Rasslabimost' i rastiazhimost' serdtsa pri energodefitsite.

Changes in the cardiac pump function, contractility, relaxation and distensibility in progressive metabolic blockade induced with dinitrophenol, were compared in the guinea pig isolated heart. A fall in relaxation and distensibility occurred first and was more obvious than a fall in contractility or the cardiac output. The same relationships were observed in isolated hearts of rats with myocardial phosphocreatine deficiency. The fall in relaxation and distensibility seem to be the necessary sign of acute or chronic myocardial energy deficiency.

Biochemical and ultrastructural changes in skeletal muscle induced by a creatine antagonist.

To evaluate the essentiality of creatine and phosphocreatine for the maintenance of the ultrastructure of skeletal muscle, chicks were fed a creatine antagonist, beta-guanidinobutyric acid (beta-GBA), as 2% of a Chow diet. Chicks fed beta-GBA exhibited growth retardation and weakness, and they accumulated large amounts of a monosubstituted guanidino compound, presumably beta-GBA, in their skeletal muscles. After 2 wk, there was a 74% decrease in the uptake of [14C]-1-creatine into pectoralis muscles of chicks fed beta-GBA. After 2 wk there as a significant decrease in phosphocreatine concentrations in pectoralis muscles from 20.1 +/- 2.8 mumoles per g wet weight (mean +/- S.D.) for 8 control chicks to 16.5 +/- 2.5 for 7 chicks fed beta-GBA. Selected fibers of the pectoralis and gastrocnemius muscles of chicks fed beta-GBA exhibited ultrastructural abnormalities including loss of thick and thin filaments, disruption of the Z band, dilated mitochondria, and dilated and displaced sarcoplasmic reticulum. The pectoralis muscles of chicks given 6% creatine in addition to 2% beta-GBA in the diet accumulated little beta-GBA, maintained normal phosphocreatine concentrations, and exhibited no significant ultrastructural abnormalities. These findings are the first experimental evidence that high concentrations of phosphocreatine are essential for the maintenance of the ultrastructural integrity of skeletal muscle.

Mechanism of thyroid-induced creatinuria in rat, with special reference to creatine synthesis in liver and creatine loss from skeletal muscle.

Mechanism of thyroid-induced creatinuria was investigated in rats. The rats were injected with triiodothyronine (T3) (100 mug/100g, sc) at 9.00 hr. Oxygen consumption increased 12 hr after T3 injection, reaching peak value 48 hr after the injection and decreasing to the pre-injection level at 96 hr. Urinary creatine excretion also increased during the first 10 hr after the injection, approaching maximal value 34-48 hr after the injection, and decreasing to the pre-injection level 72-82 hr after the injection. Throughout the experimental period, urinary creatinine decreased with time after the injection, although the difference between the groups was not significant. A decreased creatine tolerance was also observed after T3 injection. Beta-Guanidinopropionic acid (beta-GPA), a competitive inhibitor of creatine transport into skeletal muscle, as well as partial hepatectomy, caasing inhibition of creatine synthesis, were without effect on the difference in urinary creatine excretion between T3-treated and control animals. T3 increased the plasma creatine level both in bilateral nephrectomized and in bilateral nephrectomized plus beta-GPA administrated groups. These results suggest that increased creatine loss from skeletal muscle in addition to decreased creatine uptake of skeletal muscle rather than increased hepatic synthesis of creatine play an important role in T3-induced creatinuria.