How To Take Creatine

Creatine Practical Guide.

Creatine: A practical guide evolved from the thousands of questions asked by professional and amateur athletes from around the globe. Learn How To Most Effectively Combine Exercise, Nutrition And Smart Creatine Use For Explosive Muscle Growth And Improved Overall Health. Here is just a small sampling of the many questions addressed by this e-book How long can I keep creatine on the shelf? Will I lose muscle after I stop supplementing? Not all creatine brands recommend the same amount. What gives? Is mixing creatine with protein powder a bad idea? Why do so many creatine brands contain so much dextrose? Is loading really necessary? Im currently taking Accutane for nodular acne. Is it safe for me to supplement? Will creatine stunt my growth? Im training twice as much these days and Im still not making any gains! Why? If creatine isnt a steroid, then how come it gave me a positive doping result? Will creatine shrink my package?! Read more here...

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Cellular Systems Of Creatine Biosynthesis And Use In The Brain And Retina

Differential Cellular Expression of the Creatine Biosynthesis Enzyme, GAMT, and Creatine Kinases, uCK-Mi and CK-B, in the Adult Mouse Brain The cellular system of creatine biosynthesis and its function in the adult mouse brain have been investigated by immunohistochemical localization studies for GAMT, ubiquitous mitochondrial creatine kinase (uCK-Mi) and brain-type cytoplasmic creatine kinase (CK-B) (Tachikawa et al., 2004). GAMT is highly expressed in oligodendrocytes (Figure 4A, arrow) and olfactory ensheathing glia and moderately in astrocytes (Figure 4A, arrowhead), whereas the level of GAMT is very low in neurons and microglia and below the detection threshold in capillaries. The ability to synthesize creatine from glycine has been confirmed for astrocytes when 13C glycine was added to the culture medium, radioactive creatine was detected in cell extracts of astrocyte-rich primary cultures (Dringen et al., 1998). It remains unknown whether oligodendrocytes and olfactory...

In Vivo Magnetic Resonance Spectroscopy Of Transgenic Mice With Altered Expression Of Guanidinoacetate

This chapter reviews results of in vivo MRS studies on transgenic mice with alterations in the expression of the enzymes creatine kinase and guanidinoacetate methyltrans-ferase. The particular metabolic consequences of these enzyme deficiencies in skeletal muscle, brain, heart and liver are addressed. An overview is given of metabolite levels determined by in vivo MRS in skeletal muscle and brain of wild-type and transgenic mice. MRS studies on mice lacking guanidinoacetate methyltransferase have demonstrated metabolic changes comparable to those found in the deficiency of this enzyme in humans, which are (partly) reversible upon creatine feeding. Apart from being a model for a creatine deficiency syndrome, these mice are also of interest to study fundamental aspects of the biological role of creatine. MRS studies on transgenic mice lacking creatine kinase isoenzymes have contributed significantly to the view that the creatine kinase reaction together with other enzymatic steps...

By Guanidinoacetate Methyltransferase And Creatine Kinase

Creatine is a small metabolite that fulfills a central role in energy metabolism. The final step in its biosynthesis is controlled by the enzyme guanidinoacetate methyltransferase (GAMT), which catalyzes the methylation of guanidinoacetate to form Cr. In mammals, this biosynthesis is assumed to take place mainly in the pancreas and liver. Subsequently, Cr is exported into the blood and taken up by tissues requiring Cr, such as muscle and brain (Wyss and Kaddurah-Daouk, 2000). Intake of creatine-containing food may also increase the blood level of creatine, and tissues such as muscle and brain may have the enzymatic machinery to synthesize creatine themselves. Tissues taking up creatine are generally characterized as having high and fluctuating energy demands. In these tissues, Cr is reversibly phosphorylated to phospho-creatine (PCr) in the near-equilibrium reaction Cr + ATP PCr + ADP + H+, which is catalyzed by the enzyme creatine kinase (CK). This reaction is thought to be important...

Cerebral Creatine Deficiency Syndromes Clinical Aspects Treatment And Pathophysiology

Abstract Cerebral creatine deficiency syndromes (CCDSs) are a group of inborn errors of creatine metabolism comprising two autosomal recessive disorders that affect the biosynthesis of creatine - i.e. arginine glycine amidinotransferase deficiency (AGAT MIM 602360) and guanidinoacetate methyltransferase deficiency (GAMT MIM 601240) - and an X-linked defect that affects the creatine transporter, SLC6A8 deficiency (SLC6A8 MIM 300036). The biochemical hallmarks of these disorders include cerebral creatine deficiency as detected in vivo by 1H magnetic resonance spectroscopy (MRS) of the brain, and specific disturbances in metabolites of creatine metabolism in body fluids. In urine and plasma, abnormal guanidinoacetic acid (GAA) levels are found in AGAT deficiency (reduced GAA) and in GAMT deficiency (increased GAA). In urine of males with SLC6A8 deficiency, an increased creatine creatinine ratio is detected. The common clinical presentation in CCDS includes mental retardation, expressive...

Presymptomatic Treatment Of Creatine Biosynthesis Defects

Abstract Recent observations in two patients, one with AGAT deficiency (AGAT-D) and one with GAMT deficiency (GAMT-D), both diagnosed already at birth, provide first evidence for important therapeutic effects of pre-symptomatic treatment with creatine (Cr) supplementation in AGAT-D and Cr supplementation plus guanidinoacetate lowering strategies in GAMT-D. Although long-term data are lacking, the results suggest that complete prevention of neurological sequelae in early treated patients could be feasible (Battini et al, 2006 Schulze et al, 2006)

Clinical Utility Of Creatine In Neuromuscular Disorders

Some of the earliest interest in the role of creatine and muscular dystrophy emerged in the 1950's (Benedict et al., 1955 Milhorat, 1953). A review on biochemical aspects of muscular dystrophy summarized the available data showing that there was a marked increase in creatine excretion (Benedict et al., 1955), to the extent that some investigators were suggesting that muscular dystrophy could be due to disturbance of creatine metabolism (which we now know is not correct). The hypothesis that the increased urinary creatine was due to an inability of the dystrophic muscle to transport creatine (Benedict et al., 1955) is likely to be correct (Tarnopolsky et al., 2003 Tarnopolsky and Parise, 1999). A re-emergence of the interest in the clinical utility of CrM in patients with neuromuscular disorders emerged following the increased interest in creatine metabolism in the early 1990's. Our group reported the results of an open study (N 21) followed by a single-blinded study looking at...

Genetic And Secondary Disorders Of Creatine Metabolism

Inborn errors of creatine metabolism in humans may provide better insight into the pathogenic mechanisms associated with these diseases. Mutations in two enzymes responsible for creatine biosynthesis, L-arginine glycine amidino-transferase (AGAT) and guanidinoacetate methyltransferase (GAMT), as well as mutations in a sodium- and chloride-dependent creatine transporter (SLC6A8), have been identified. In patients with GAMT and AGAT mutations, clinical symptoms include mental and motor retardation, extra-pyramidal symptoms, and seizures (see chapter 8 Stockler et al., 2007 Item et al., 2001 Stromberger et al., 2003). The gene encoding SLC6A8 is located on the X-chromosome, thereby usually affecting males, who show more severe mental retardation and absence of speech than affected females (Mancini et al., 2004). While not always consistent, plasma and CSF levels of creatine are usually low in these patients with variable, but usually elevated creatinine levels in urine (Verhoeven et al.,...

Historical Aspects Of Creatine Kinase And Creatine

Functions of Creatine Kinase and Phosphocreatine in Muscle Contraction As already mentioned, CK was considered a strictly soluble metabolic enzyme just for trivial buffering of cellular ATP levels according to its equilibrium constant (Meyer et al., 1984). The facts that (i) CK existed as several tissue-specific and developmentally regulated cytosolic isoforms (Eppenberger et al., 1964, 1967), that (ii) a CK isoform was identified that is located within mitochondria (Jacobs et al., 1964 Jacobus and Lehninger, 1973), and that (iii) a small but significant fraction of soluble muscle-type MM-CK was shown to bind specifically to the sarcomeric M-band of skeletal muscle (Turner et al., 1973 Wallimann, 1975), to the sarcoplasmic reticulum (SR) (Rossi et al., 1990), as well as to the plasma membrane (Saks and Kupriyanov, 1982 for a recent review, see Wallimann et al., 2007), were met with some resistance by the muscle research community. It was said that a soluble enzyme cannot be...

Creatine Supplementation In Neurodegenerative Disorders

As noted above, creatine administration has several potential neuroprotective effects including buffering of intracellular energy reserves, stabilizing intracellular calcium, reducing extracellular glutamate, inhibiting activation of the mitochondrial permeability transition, and acting as an anti-oxidant. The salubrious neuroprotective effects of creatine have been widely reported in experimental models of neurological diseases, particularly in neurotoxin and transgenic models of HD (Andreassen et al., 2001a Balestrino et al., 1999 Brewer and Wallimann, 2000 Brustovetsky et al., 2001 Carter et al., 1995 Dedeoglu et al., 2003 Ferrante et al., 2000 Hausmann et al., 2002 Ikeda et al., 2000 Klivenyi et al. 2003, 2004a,b Malcon et al., 2000 Matthews et al., 1998, 1999 Royes et al., 2003 Ryu et al., 2005 Shear et al., 2000 Sullivan et al., 2000 Zhu et al., 2004). Both creatine and phosphocre-atine prevented death of cultured striatal and hippocampal neurons when exposed to 3-NP...

Conclusion Of Creatine Kinase

High Creatine Kinase

Our recent research provides novel molecular-anatomical evidence that (i) CRT at the BBB and iBRB is involved in regulating the creatine concentration in the brain and retina and that (ii) local creatine is preferentially synthesized in the glial cells in the brain and retina. These findings provide important information that will increase our understanding of the mechanism of creatine supply and creatine use in the brain and retina and of creatine supplementation in patients with creatine deficiency syndromes. In particular, the BBB and iBRB play essential roles in maintaining energy homeostasis not only in terms of supplying energy sources (glucose and lactate), but also for supplying an energy 'buffer' (creatine). Although oral creatine supplementation is reported to be an effective treatment for AGAT- and GAMT-deficient patients (Stockler et al., 1994, 1996), high doses of creatine given over a long period of time only partially replenish brain creatine pools. In this regard, the...

Modulation and regulation of creatine transport

Little effect on creatine uptake rates has been observed with PCr, creatinine, ornithine, glycine, glutamic acid, histidine, alanine, arginine, leucine, methionine or cysteine (Fitch, 1988 Loike et al., 1988 M ller and Hamprecht, 1989). The sodium-dependent uptake of creatine in culture is sensitive to extracellular creatine concentration (Loike et al., 1988). Myoblasts maintained for 24 h in a medium containing 1 mM creatine exhibited one-third of the uptake activity of cells bathed for the same duration in a medium lacking creatine. The downregulation was slowed by inhibitors of protein synthesis. From this observation one can conclude that extracellular creatine regulates the induction of the expression of a protein that decreases the sodium-dependent uptake activity only after the creatine enters the cell. No alteration in the efflux rate for creatine was observed in studies of the effects of changes in extracellular creatine concentrations (Loike et al., 1988). If creatine was...

Clinical Utility Of Creatine In Neurometabolic Disorders

There have been a number of small case series and case reports on the effects of creatine supplementation in mitochondrial cytopathies. A case report of an 18 year old with MELAS syndrome taking creatine at 12 g d for 12 days followed by 5g d for 28 months showed a definite improvement in MRI and MR spectroscopy values with an increase in brain creatine as well as marked behavior and cognitive improvements. There appeared to be a deterioration of renal function with an impairment of renal creatinine clearance in this patient who had pre-existing nephropathy (Barisic et al., 2002). A small open study in four patients with mitochondrial cytopathy (Kearns Sayre syndrome, NARP and MELAS) evaluated the effects of CrM supplementation (0.1-0.35g kg day) (Komura et al., 2003). This was actually a retrospective study for nine months to almost five years and found an improvement in cycle ergometry performance of about 12 during steady-state exercise with no significant effects on peak power Our...

Pleiotropic Effects Of Creatine

Capacity for intracellular ATP and ADP concentrations, one could explain at least part of the Cr-dependent protection of cells from various stressors, especially the neuroprotective effects of Cr (see chapters 8-11 Stockler et al., 2007 Schulze and Battini, 2007 Tarnopolsky, 2007 Klein and Ferrante, 2007), by the classical beneficial effects of Cr in improving the cellular energy state. Cr supplementation mediates remarkable neuroprotection in experimental animal models of amyotrophic lateral sclerosis (ALS), Huntington's disease (HD), Parkinson's disease, and traumatic brain and spinal cord injury (see chapters 10 and 11 herein Tarnopolsky, 2007 Klein and Ferrante, 2007). Prophylactic Cr administration also mediates neuroprotection in cerebral ischemia in mice (Adcock et al., 2002 Prass et al., 2007 Zhu et al., 2004). Transgenic mice expressing high levels of BB-CK in liver cells, which normally express only very low levels of this enzyme, show a high degree of resistance to tumour...

Early Evolution Of The Creatine Kinase Gene Family And The Capacity For Creatine Biosynthesis And Membrane Transport

Abstract The creatine kinase (CK) phosphocreatine (PCr) energy buffering system is widespread in animal groups. Recent genomic sequencing and experimental results support the view that the capacity for creatine biosynthesis and membrane transport may have evolved quite early, perhaps coincident with CK. Conventional wisdom would suggest that CK evolved from an ancestral protein most similar to the CK homologue, arginine kinase. This early CK gene subsequently diverged into the cytoplasmic, mitochondrial and flagellar CK gene families. It is now clear that both the mitochondrial and cytoplasmic-flagellar genes were present prior to the divergence of sponges from the multi-cellular animal (metazoan) lineage, possibly as long as a billion years ago. Sponges constitute the most ancient, extant metazoan group. It is likely that the primary function of the CK-PCr system in these primitive animals was to mitigate reaction-diffusion constraints in highly polarized cells such as spermatozoa...

The Neuroprotective Role Of Creatine

Abstract Significant progress has been made in identifying neuroprotective agents and their translation to patients with neurological disorders. While the direct causative pathways of neurodegeneration remain unclear, they are under great clinical and experimental investigation. There are a number of interrelated pathogenic mechanisms triggering molecular events that lead to neuronal death. One putative mechanism reported to play a prominent role in the pathogenesis of neurological diseases is impaired energy metabolism. If reduced energy stores play a role in neuronal loss, then therapeutic strategies that buffer intracellular energy levels may prevent or impede the neurodegenerative process. Recent studies suggest that impaired energy production promotes neurological disease onset and progression. Sustained ATP levels are critical to cellular homeostasis and may have both direct and indirect influence on pathogenic mechanisms associated with neurological disorders. Creatine is a...

Creatine uptake

Most studies on creatine transport have focused on creatine influx and several tissues have been examined (Ku and Passow, 1980 Loike et al., 1986a M ller and Hamprecht, 1989 Odoom et al., 1993). This saturable active transport is highly specific for creatine, sodium dependent, concentrative and sensitive to metabolic inhibitors (Fitch et al., 1968b Fitch and Chevli, 1980 Ku and Passow, 1980 Loike et al., 1986a, 1988 M ller and Hamprecht, 1989 Bennett et al., 1991). Two sodium molecules are transported for every creatine molecule and the Km for sodium is 55 mm (M ller and Hamprecht, 1989) thus favouring Cr transport. The uptake of creatine appears to be little affected by extracellular pH over the range pH 6.9-7.9 (Syllm-Rapoport et al., 1980). The electrogenic nature of creatine transport is important in the mechanism of uptake of the molecule. The transporter exploits the negative membrane potential as well as the inwardly directed sodium gradient in order to maximize creatine...

Creatine Kinase

Creatine phosphate serves as a cellular storage species of high-energy phosphate that is readily available for the formation of ATP from ADP by the action of creatine kinase (EC 2.7.3.2). Creatine phosphate is a high-energy phosphate (AG' -10 kcal mol-1 for hydrolysis) by virtue of being a phosphoramidate. The reversibility of the reaction of creatine kinase (eq. 10-5), allows creatine phosphate to be formed whenever the concentration of ATP is high, and it makes creatine phosphate an efficient reserve of high-energy phosphate to generate ATP under conditions of high-energy demand. The action of creatine kinase requires a divalent metal ion, either Mg2+ or Mn2+ in complex with ADP. The more prevalent Mg2+ is the likely activating metal ion in cells. Structurally and stereochemically distinct complexes of divalent metal ions with ADP or ATP exist in solution, and species that bind to a particular enzyme can be determined in various ways. For example, the following two structures of...

Gajja S Salomons1 And Markus Wyss2

To make a project a success, you typically need to have three things a good idea, an enthusiastic team, and a committed 'customer' (in this case, the Springer publishing company). When J. Robin Harris, the Series Editor of Subcellular Biochemistry, first approached us with the idea of a book on Creatine and Creatine Kinase in Health and Disease, we were both surprised and delighted. Next, we were impressed by the dedicated participation of many of the key players in creatine and creatine kinase research from all over the world. In fact, all authors we asked to participate accepted the invitation, and have submitted high-quality manuscripts in a timely manner. Consequently, the book is up to date, and both editors and authors are confident that this book will become an important contribution to expand the knowledge and funding of creatine and creatine kinase research. Finally, the continuous support of the Springer publishing company and, in particular, of Mike van den Bosch, Marie...

Historical Introduction To Phosphagen Kinases

Lohmann is credited with the discovery of creatine kinase (CK) in 1934. He believed that the reaction of adenosine monophosphate (AMP) with two phosphocreatine (PCr) molecules gave adenosine triphosphate (ATP) and creatine (Cr) and that the observed orthophosphate (P,) released from PCr in contracting muscle was due to ATP hydrolysis Creatine kinase was first purified by Kuby et al. (1954) from rabbit muscle and its properties reported by Kuby and Noltman (1963). Creatine and Creatine Phosphate In general, CK is characteristic of vertebrates and arginine kinase is characteristic of invertebrates. However, although arginine kinase has never been reported from vertebrates, CK is found in various invertebrate phyla (e.g. Porifera, Annelida, Echinodermata and Chordata) (Watts, 1971). In most cases a single species has one phosphagen but there are examples of more than one kinase occurring in some species of annelids and echinoderms. Phosphagens and their kinases are not restricted to...

Ck Isoform Divergence Occurred At The Dawn Of The Radiation Of The Metazoa

Creatine kinases actually constitute a superfamily of three isoform gene families each targeted to different intracellular compartments - cytoplasmic (CytCK), mitochondrial (MtCK) and flagellar (FlgCK) CKs. The former two gene families code for proteins that exist as oligomers - CytCKs are dimers while typical MtCKs exist as both dimers and octamers, with the latter predominating under physiological conditions (Wallimann et al., 1992 Wyss et al., 1992). FlgCKs are contiguous trimers consisting of three fused, complete CK domains in a single polypeptide chain and most likely resulted from a gene duplication fusion event followed by unequal crossing over (Wothe et al., 1990). CytCKs, octameric MtCKs and FlgCKs evolved before the divergence of the protostome (most invertebrates such as arthropods, mollusks, worms) and deuterostome (echinoderms, protochordates, vertebrates) lineages, at least 650 million years ago (Ellington et al., 1998 Suzuki et al., 2004). and are widely distributed in...

Expression And Function Of Agat Gamt And Ct1 In The Mammalian Brain

Abstract In mammals, creatine is taken up from the diet and can be synthesized endogenously by a two-step mechanism involving the enzymes arginine glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT). Creatine (Cr) is taken up by cells through a specific transporter, CT1. While the major part of endogenous synthesis of Cr is thought to occur in kidney, pancreas and liver, the brain widely expresses AGAT, GAMT and CT1 , both during development and in adulthood. The adult central nervous system (CNS) has a limited capacity to take up Cr from periphery, and seems to rely more on its endogenous Cr synthesis. In contrast, the embryonic CNS might be more dependent on Cr supply from periphery than on endogenous synthesis. This review will focus on the expression and function of AGAT, GAMT and CT1 in the mammalian CNS, both during development and in adulthood. Emphasis will also be placed on their specific roles in the different cell types of the brain, to analyze...

Catalytic and substrate binding domain

Kinetic studies suggest separate but synergistic binding of MgATP and creatine. Recently, using site-directed mutagenesis, Lin et al., (1994) and others (Furter et al., 1993) tried evaluating further the role of cysteine-283. In Lin et al.'s study of mitochondrial CK expressed by the Escherichia coli system,

Regulation Of The

The prime function of the CRT is for the cellular accumulation of creatine which, in turn, is crucial for the maintenance of ATP in tissues with high energy demands like brain and muscle. Also, creatine is neuroprotective in many animal models of neurological diseases (Wyss and Schulze, 2002). Consequently, the regulation of the CRT is an important topic. It is essential to know whether creatine supplementation down-regulates the transporter. Alternatively, if we can learn how to up-regulate the transporter, then it may be possible to enhance the uptake of creatine into the brain, facilitating its use as a neuroprotective agent. Efforts have been made to understand how the CRT is regulated in relation to tissue creatine levels, particularly for muscle. However, deficiencies in the specificity of some of the available CRT antibodies (discussed in section 5.2 above) have so far limited our understanding of the regulation of CRT activity at the cellular level. Understanding of the...

Study Design Issues In Clinical Trials

The ultimate goal of a therapeutic intervention is to cure the disease. Given that a cure is not possible for essentially every neuromuscular and neurometabolic disorder, the goal is control of symptoms and functional improvement. Consequently, to determine the efficacy of a therapeutic intervention, the ultimate evidence of efficacy is an improvement in quality of life and functional capacity as well as a reduction in morbidity and mortality. An evaluation of the effect of an intervention on these clinical outcomes usually requires very large numbers of subjects followed for a long period of time to detect significant differences as is possible with common conditions such as cancer, heart disease and diabetes. Most of the neuromuscular and neurometabolic disorders are relatively rare and the recruitment of hundreds to thousands of patients is not feasible. Another issue is that of the cost of running such trials which can run into millions of dollars for clinical outcome evaluations....

The Normal Heart Resting Metabolism

Human and animal cardiac spectra are composed of peaks derived from phosphates at different chemical shifts. On reading the spectrum from left to right, 2,3 diphosphoglycerate (2,3 DPG), inorganic phosphate (P,), phospho-diester (PDE), creatine phosphate (PCr) and three peaks for ATP can be identified. Blood, heart and skeletal muscle contain P PDE and ATP. Creatine phosphate is present in skeletal and cardiac muscle and absent from blood. The presence of a prominent 2,3 DPG peak indicates that the sample volume has a significant amount of blood present. Figure 9.1. Cardiac spectrum from a control acquired using the phase-modulated rotating frame imaging technique (PMRFI) (2,3 DPG+P,, 2,3 diphosphoglycerate and inorganic phosphate PDE, phosphodiester PCr, creatine phosphate ATP, adenosine triphosphate, y, a and phosphates ppm, frequency in parts per million). Figure 9.1. Cardiac spectrum from a control acquired using the phase-modulated rotating frame imaging technique (PMRFI) (2,3...

Substrate Specificity And Transport Mechanism Of The

Creatine Kinase Mechanism Cysteine

The CRT is highly specific for creatine. Neither creatinine nor phosphocreatine are substrates. The key features for substrate specificity are a carboxyl group and a guanidino group, separated by no more than 2-3 carbon atoms (Guimbal and Kilimann, 1993 Moller and Hamprecht, 1989 Saltarelli et al., 1996 Sora et al., 1994). A benzyl ester derivative of creatine which no longer contains a free carboxyl group is not a substrate for the transporter although it can be taken up by brain slices in a non-transporter dependent manner (Lunardi et al., 2006). j8-Guanidinopropionate is a better competitive inhibitor than guanidinoacetate or 4-guanidinobutyrate. 2-Amino-3-guandinopropionate competes more efficiently with creatine for rabbit than electric ray CRT, suggesting slightly altered binding sites in different species (Guimbal and Kilimann, 1993, 1994). Arginine, citrulline, carnitine, GABA and choline are all without a significant effect on creatine transport. Many studies have shown that...

Role Of Phosphagens In Clinic

Recent work from the laboratory of Eric Hultman in Stockholm has demonstrated that, in man, maximum sprinting can only be supported by using both glycogen and phosphocreatine simultaneously as fuels. This has led to considerable interest in providing creatine for athletes (see Newsholme et al., 1994 and Chapter 13). These results suggest that, in insect flight muscles, rat heart and possibly hearts from other animals, phosphagen kinase may not play a role in the transfer of energy-rich phosphate from mitochondria to myofibrils, at least under conditions of maximum rates of ATP turnover. It is possible that the role of PCr Cr in the energy shuttle process depends upon the distances between the sites of ATP production and ATP utilization (see Chapter 5). With large distances as in spermatozoa, the creatine system functions as an important energy shuttle system (Juretschke and Kamp, 1990). Creatine phosphate inhibits phosphofructokinase, a regulatory enzyme in glycolysis (Uyeda and...

Mitochondrial Encephalopathies

Metabolic activity within the central nervous system is relatively high, making brain tissue especially vulnerable to mitochondrial respiratory chain abnormalities. Mitochondrial encephalopathies are multi-system diseases characterized by biochemical and genetic mitochondrial defects with differing clinical symptomatologies and brain area involvements. Mitochondrial encephalopathies are age-related with respect to disease onset, occurring mostly in infants and young children, but also in adulthood in some patients (Barisic et al., 2002 Komura et al., 2003 Kremer et al., 1993 Lofberg et al., 2001 Scaglia and Northrop, 2006 Tarnopolsky et al., 1998 Walter et al., 2002). Included in this group of mitochondrial dysfunction disorders are lactic acidosis with stroke-like episodes (MELAS) Leigh syndrome myoclonus epilepsy and ragged-red-fibers (MERRF) Kearns-Sayre syndrome (KSS) neuropathy, ataxia, and retinitis pigmentosa (NARP) mitochondrial neurogastrointestinal encephalopathy (MNGIE)...

Cardiac and skeletal muscle

Myocardial gene expression of CK is developmentally regulated (Lamers et al., 1989 Hasselbaink et al., 1990). In the mouse BB-CK alone is expressed during prenatal development, until the last week, when MM-CK increases dramatically whilst the BB-CK isoform decreases concomitantly. Associated with the co-expression of M-CK and B-CK gene peripartum, and during the first 3 weeks postnatally, there is up to a maximum of 35 MB-CK activity (Hall and DeLuca, 1975 Ingwall, 1991). The mitochondrial isoform does not get expressed until several days postnatally (Hall and DeLuca, 1975 Ingwall, 1991). In several diseased states with altered cardiac function, alteration of CK gene expression also occurs. In pressure overload or volume overload induced cardiac hypertrophy of both humans (Jansson et al., 1987 Ingwall, 1991) and experimental animals (Meerson and Javich, 1982 Younes et al., 1984 Schuyler and Yarbrough, 1990 Field et al., 1992), alteration of the CK isozyme expression develops. In the...

Subcellular localization domain

A specific domain exists for sarcoplasmic reticulum localization or association with other sarcoplasmic reticulum proteins has not been established. Recently, Rojo et al. (1991) showed association of the CK to model membrane preparation. The mitochondrial CK interacts with a phospholipid, cardiolipin of the inner mitochondrial membrane (Ottaway, 1967 Schlame and Augustin, 1985 Cheneval and Carafoli, 1988), and with a model phospholipid membrane (Rojo et al., 1991). Further delineation of the subcellular localization domain is important to the understanding of the creatine phosphate shuttle and awaits analysis by site-directed mutagenesis through domain-deletion or domain-switching experiments.

Valdur Saks12 Tuuli Kaambre2 Rita Guzun1 Tiia Anmann2 Peeter Sikk2 Uwe Schlattner13 Theo Wallimann3 Mayis Aliev4 And

Abstract In this review, we summarize the main structural and functional data on the role of the phosphocreatine (PCr) - creatine kinase (CK) pathway for compartmentalized energy transfer in cardiac cells. Mitochondrial creatine kinase, MtCK, fixed by cardiolipin molecules in the vicinity of the adenine nucleotide translocator, is a key enzyme in this pathway. Direct transfer of ATP and ADP between these proteins has been revealed both in experimental studies on the kinetics of the regulation of mitochondrial respiration and by mathematical modelling as a main mechanism of functional coupling of PCr production to oxidative phosphorylation. In cells in vivo or in permeabilized cells in situ, this coupling is reinforced by limited permeability of the outer membrane of the mitochondria for adenine nucleotides due to the contacts with cytoskeletal proteins. Due to these mechanisms, at least 80 of total energy is exported from mitochondria by PCr molecules. Mathematical modelling of...

Pathophysiological Considerations

Present evidence suggests that each of the CCDSs has potential effects on several aspects of creatine physiology, including the intermediates of creatine biosynthesis and their biochemical reactions, the transfer of creatine from biosynthesis to usage compartments, intracellular creatine phosphorylation cycles, and all cellular functions normally supported by creatine in developing and mature brain. 6.1. Disrupted Creatine Transport The concept of an organ-specific transfer of creatine from sites of biosynthesis in liver, pancreas and kidney to sites of usage in brain and muscle must be refined, at least with regard to the brain. Immunohistochemical and functional studies have established the presence of AGAT and GAMT in all types of brain cells in vivo (Braissant et al., 2001) and the capacity of cultured astrocytes to synthesise creatine (Dringen et al., 1998). This suggests a dual origin of cerebral creatine from intracerebral biosynthesis and from transport across the blood-brain...

Transgenic animal model

Over-expression of B-CK by transgenic mice has been reported (Brosnan et al., 1993). Under the control of a skeletal muscle actin promoter, the transgene was expressed and produced MB-CK (22-32 ), which is normally absent in adult skeletal muscle. Over-expression of CK, however, did not alter ATP, creatine phosphate, or creatine levels, supporting the generally held belief that CK-catalysed skeletal muscle reactions are normally in equilibrium. A transgenic model with liver-specific expression of B-CK has also been reported (Brosnan et al., 1990). The hepatocytes of the liver do not normally express any detectable CK.

Molecular Characterization Of The

Molecular cloning showed the CRT to be a member of solute carrier family 6, a large family of membrane transporters that mediate the translocation of a range of solutes across plasma membranes, through the co-transport of sodium and chloride down their electrochemical gradients (Chen et al., 2004). The -y-aminobutryic acid (GABA) and norepinephrine transporters (Guastella et al., 1990 Paczkowski et al., 1999) were the first to be discovered, followed rapidly by identification of transporters for dopamine and serotonin, accounting for reference to this family as Na+ Cl- -dependent neurotransmitter transporters or neurotransmitter sodium symporters (NSS) (Saier, 1999). Subsequently, homologous transporters for solutes other than neurotransmitters were discovered, e.g. for taurine, betaine and creatine. The human genome encodes twenty SLC6 transporters, a large sub-branch of which are Na+-dependent amino acid transporters (Broer, 2006). A role in amino acid transport is likely to account...

Tissue Distribution Of The

Localization of the CRT protein identifies sites of creatine uptake and thus has functional significance. Unfortunately, studies of the localization of the CRT protein have been scarce due to the limited availability and specificity of CRT antibodies. Several antibodies are available that appear to give reliable identification of the CRT by immunocytochemistry (Acosta et al., 2005 Murphy et al., 2001 Ohtsuki et al., 2002 Peral et al., 2002). However, the development of antibodies with adequate specificity and sensitivity to detect the CRT in tissue extracts by Western blotting has been more problematic. The CRT does not appear to be an abundant component of plasma membranes, even in tissues containing high levels of creatine and phosphocreatine. Also, depending on conditions and possibly different glycosylation states, the CRT may run with different apparent molecular masses on SDS polyacrylamide gels. The CRT may also aggregate once solubilized from membranes. Antibodies to the CRT,...

Michael A Conway Ronald Ouwerkerk Bheeshma Rajagopalan and George K Radda

Creatine phosphate changes in tachycardia and rhythm disorders 150 11. Changes in creatine phosphate the effects of drug therapy and diagnostic potential 152 Key words creatine phosphate, magnetic resonance spectroscopy, myocardium aortic stenosis, aortic incompetence, hypertrophy, heart failure

Biochemical Pathology And Laboratory Diagnosis

CCDSs are characterized by an almost complete lack of creatine in the brain, which is shown in vivo by brain H MRS. It should be noted that in females with a heterozygous mutation in the SLC6A8 gene, a milder reduction in cerebral creatine is usually detected. However, cerebral creatine can vary in heterozygous females from being absent to normal levels, depending on the skewing in X-inactivation. Interestingly, in CCDS patients, creatine deficiency is much less severe in muscle than in brain. Although creatine levels in skeletal muscle have not been investigated extensively in CCDS patients and insights are based on case studies only, creatine content of skeletal muscle was measured in vivo by proton MRS and in vitro in a muscle biopsy of a SLC6A8-deficient patient (Pyne-Geithman et al., 2004). This case report revealed normal creatine levels, suggesting that endogenous creatine biosynthesis is able to maintain proper creatine levels, and or that another creatine transporter is able...

Jaliashvilf Eugeny A Konorev2 Sergi A Kryzkanovsky2 and Ettore Strumia1

Creatine and Creatine Phosphate transplantation (Hearse, 1980 Semenovsky et al., 1987). One agent which shows promise in this respect is creatine phosphate (Neoton). Creatine phosphate (PCr) was discovered by Eggleton and Eggleton in 1927 (see Ennor and Morrison, 1958, for review), and the discovery predated that of ATP. It has taken more than 50 years of intensive study to understand various aspects of the physiological function of this molecule. It is now accepted that PCr has at least a dual role in cells with high energy turnover such as in the heart, skeletal muscle, brain, retina, spermatozoa and nerve endings (Saks et al., 1978 Bessman, 1985 Jacobus, 1985 Wallimann et al., 1989). The intracellular PCr content is high and ranges from 2 to 30 mM (Dawson, 1983 Gard et al., 1985). Evidence to date suggests that it acts as an energy (ATP) buffer (Mommaerts, 1969) and also as an intracellular energy carrier (Saks et al., 1978 Bessman, 1985 Jacobus, 1985 Wallimann et al., 1989). The...

Theodore J Lampidis Yufang Shi and Luigisilvestro1

Protection by creatine phosphate against Adriamycin 117 Key words creatine phosphate, muscle, cardiac, hypoxia, arrhythmia, adriamycin, phosphocreatine, phosphoarginine, cardiotoxicity Creatine and Creatine Phosphate Copyright 1996 Academic Press Limited With the ability to maintain functionally active cardiac cells in culture for prolonged periods, we began to develop this system as an in vitro model to study cardiopathogenesis produced in patients treated with the potent anti-tumour agent Adriamycin (ADM). ADM-induced cardiomyopathy is a cumulative dose-limiting toxicity which prevents a more efficacious use of this drug as an antitumour agent. We have previously shown that ADM and a number of other anthracycline analogues which are positively charged (at physiological pH) are accumulated preferentially in cardiac muscle cells as opposed to co-cultured cardiac fibroblasts (Lampidis et al., 1981). It has also been observed with compounds other than the anthracyclines (xanthene dyes)...

Oxidative Phosphorylation

Schematic representation of creatine-stimulated respiration at the inner mitochondrial membrane of a saponin-skinned cardiac fibre. Excess creatine (Cr) shifts the local Mi-CK reaction in favour of an increase in local ADP. ADP is then transported via the adenine nucleotide translocase into the mitochondrial matrix to stimulate respiration. IMM, inner mitochondrial membrane OMM, outer mitochondrial membrane SL, sarcolemma mit-CK, mitochondrial creatine kinase bound to the IMM. causes an increased sensitivity of mitochondrial respiratory control to changes in ADP. Through this mechanism Cr stimulates respiration (Veksler et al., 1988, 1991 Saks et al., 1991a, 1993). Creatine shifts the local Mi-CK equilibrium (Eqn 1, page 52) towards the production of PCr and ADP (ADP in turn stimulates respiration). Thus creatine and Mi-CK can lower the Km, or increase the sensitivity, for respiratory control of oxidative phosphorylation with respect to ADP (Fig. 4.3). For further details...

Traumatic Brain And Spinal Cord Injury

The primary cause of neuronal death is the physical damage of impact, the secondary effects of trauma are less apparent and are associated with edema, ischemia, inflammation, altered calcium homeostasis, and impaired energy metabolism related to mitochondrial dysfunction and ATP loss (Sullivan et al., 1998, 1999 Taoka and Okajima, 1998 Xiong et al., 1997, 1998). ATP loss may also be the result of vascular damage and consequent hypoxia. As such, enhanced neuronal survival in traumatic injuries may be dependent upon adequate ATP supplies. As discussed above, creatine may provide enhanced neuroprotection after these cellular events. There have been efforts to demonstrate the efficacy of creatine in experimental models of TBI and SCI (Hausmann et al., 2002 Rabchevsky et al., 2003 Scheff and Dhillon, 2004). Surgically-induced cortical contusions representing the sequelae of human closed-head injury were performed in mice. These resulted in severe behavioral deficits, cortical tissue loss,...

Metabolism

Clinical Experience with Creatine Phosphate Therapy 185 Paola Fault tto and Ettore Sirumia 12. Creatine Phosphate Added to St Thomas' Cardioplegia 199 David J. Chambers 13. Uses of Creatine Phosphate and Creatine Supplementation for the Athlete 217 Joseph F. Clark 2. Experimental creatine supplementation 218 3. Oral creatine supplementation 219 5. Creatine phosphate 221 14. Creatine and Creatine Phosphate Future Perspectives 227 Michael A. Conwy and Joseph F. Clark Assay for Creatine and Creatine Phosphate 233

Acknowledgements

Creatine and Creatine Phosphate Scientific and Clinical Perspectives was compiled with the support of a large number of contributors who undertook to complete their manuscripts within a very short period of time. We are very grateful to them for their enthusiastic support.

Shuttle

One of the central tenets regarding the regulation of mitochondrial oxidative metabolism is that ADP produced in the cytoplasm diffuses back into the mitochondria to stimulate the production of more ATP (i.e. respiratory control). Alternatively Jacobus and Lehninger (1973) proposed that ADP produced by high metabolic activity is immediately rephosphorylated to ATP through the action of creatine kinase (CK), using the phosphate from creatine phosphate (PCr). The respiratory signal to the mitochondria, in this Creatine and Creatine Phosphate hypothesis, is creatine. The creatine, produced at sites of high metabolic activity, diffuses back into the mitochondria to be rephosphorylated to PCr through the action of a different CK isoenzyme, the mitochondrial CK. The location of mitochondrial CK facilitates this reaction since it is bound to the outer surface of the inner mitochondrial membrane. Such a cycle, in which creatine and PCr serve as the diffusible intermediates connecting sites of...

Intestinal epithelia

Cance of an intestinal epithelial cell creatine phosphate shuttle. In vitro studies support the shuttle hypothesis and demonstrate that, at the brush border, locally CK generated ATP with PCr as substrate, is used selectively for myosin filament shortening at the same location (Gordon and Keller, 1992).

Agat Deficiency

AGAT deficiency affects the first enzyme in creatine biosynthesis. This disorder was first described in two Italian sisters with unspecific developmental and speech delay, and occasional (fever-induced) seizures in one of them. In vivo H MRS of the brain revealed creatine deficiency, but opposite to GAMT deficiency, GAA levels were low in body fluids. This combination pointed to a defect in GAA biosynthesis, and AGAT deficiency was finally confirmed by deficient enzyme activity and identification of a homozygous nonsense mutation in the AGAT gene (Item et al., 2001). The AGAT gene has been mapped to chromosome 19p13.3, thus confirming that AGAT deficiency is an autosomal recessive disorder. Supplementation with creatine monohydrate resulted in almost complete normalisation of cerebral creatine levels and a catch-up in psychomental development (Bianchi et al., 2000). Although, after 6 years of treatment, the 13- and 11-year-old sisters still have a moderate intellectual deficit, there...

Valdura Saks

Key words adenine nucleotides, cardiomyocytes, compartmentation, creatine kinase, diffusion, free energy exchange, mitochondria, oxidative phosphorylation, regulation The creatine kinase (CK) isoenzymes function, in vivo, coupled to intracellular structures (mitochondria, myofibrils and cellular membranes). They operate at equilibrium in the cytoplasm as part of the intracellular creatine phosphate (PCr) pathway, or phosphocreatine circuit for energy channelling (Saks et al., 1978, 1991a Bessman and Geiger, 1981 Bessman and Carpenter, 1985 Jacobus, 1985 Wallimen et al., 1992 Wegmann et al., 1992 Wyss et al., 1992 Aliev and Saks, 1993). Chapters 1,2 and 4 provided an introduction to the CK system. We will now see how compartmentalization of CK enzymes are involved in cardiac energy metabolism. Creatine and Creatine Phosphate Figure 5.1. (A) The dependence of the rate of respiration of the skinned cardiac fibres on the external ADP concentration in the absence (curve 1) and in the...

Ischemic Stroke

The maintenance of ion gradients across the neuronal cell membrane involves a significant degree of metabolic energy provided by ATP. Within minutes of reduced or lack of blood flow, a cascade of events occur resulting in failure of sodium potassium pumps, influx of extracellular calcium ions, and subsequent excitotoxicity that ultimately results in neuronal death. Neurons most severely affected by hypoxic injury die rapidly by necrosis, while those neurons that are exposed to a lesser degree of hypoxia in the penumbral zone succumb via apoptosis (Tatsumi et al., 2003). Creatine supplementation may result in improved ability of the neuron to withstand ischemia-mediated energetic deficiency. Creatine supplementation has been reported to be neuroprotective in an experimental model of anoxia in neonatal mice (Wilken et al., 2000). After 30 minutes of anoxia, both ATP and PCr concentrations were significantly higher in creatine-treated pups than unsupplemented controls, suggesting that...

GPA feeding

In an attempt to develop an animal model that might identify biochemical changes and pathology associated with creatine and PCr depletion, Chevli and Fitch fed rats an analogue of creatine 3-guanidinopropionic acid ( 3-GPA) (Fitch et al., 1968b, 1974 Chevli and Fitch, 1979). This agent has been used to compete with creatine for entry into the skeletal muscle cells (1 of their diet w w) as a means to study creatine transport (Fitch et al., 1968b). Further work by Shields and Whitehair (1973) showed that rats receiving diets containing 3-GPA had abnormal creatine metabolism, with chronically reduced levels of muscle creatine combined with increased levels of excreted creatinine. Fitch et al. (1974) demonstrated that 3-GPA feeding resulted in a decrease in muscle PCr with concomitant replacement by the phosphorylated analogue, 3-GPA-P. Muscles from animals fed 3-GPA were found to have decreased 3-GPA-P during hypoxic contraction indicating that, like PCr, 3-GPA-P could serve with CK in a...

Future Directions

The current data appear to support the concept that CrM administration over a period of several months can increase muscle strength and enhance body composition in patients with muscular dystrophy. It will be important to determine whether these objective markers of muscle strength translate into functional improvements and, ultimately, a reduction in clinical severity markers (such as the time to wheelchair dependence in boys with Duchenne muscular dystrophy). Clearly, these clinical outcome trials will require a multi-center approach. The potential for creatine to attenuate some of the side effects of corticosteroid treatment appears encouraging from preliminary data however, this needs to be carefully evaluated in a number of neuromuscular conditions where corticosteroid use is common (dystrophinopathy, polymyositis, myasthenia gravis). There does not appear to be a benefit from CrM supplementation in patients with inherited neuropathies at least over the short term however,...

Photoreceptors

Is converted to PCr, which then diffuses into the outer segments to regenerate ATP for visual cycle events (i.e. regeneration of cGMP and phosphorylation reactions). The creatine phosphate shuttle then serves to supply energy from the inner to the outer segments and nerve terminals for the rapid regeneration of ATP at the point of energy consumption. These results have been confirmed and extended by studies of bovine retina (Hemmer et al., 1993). Using specific antisera and light and electron microscopy, two distinct isoforms of CK (BB-CK and mitochondrial CK) were shown to be separately compartmentalized in the outer and inner segments of the photoreceptors. Furthermore, the ATP-generating potential of the CK system was sufficiently large to regenerate the entire ATP pool during a photic cycle (Hemmer et al., 1993).

Future Applications

There is ample ultrastructural, biochemical as well as physiological experimental evidence for the presence of the creatine phosphate shuttle pathway. It is also clear that during organ development and in certain disease states, the alteration of CK isoform expression occurs. The question, therefore, is no longer whether there is a functional role for CK in cell metabolism, but to what extent the compartmentalization of CK isozymes is important in the homeostasis of the specialized cells and in response to environmental cues. This question can be investigated by several of the newly developed recombinant DNA technologies, including gene targeting and dominant negative modulation.

Summary

All cells use ATP as the immediate energy source. Cellular ATP is generated by oxidative metabolism and by glycolysis. Many specialized cells in an organism require fast and constant energy utilization to maintain specialized cellular function. However, most cellular ATPases utilize ATP and therefore rapid accumulation of end-product ADP and AMP will significantly alter the equilibrium kinetics around the cellular ATPase it may be myosin ATPase, sarcolemmal ion-stimulated ATPase, SR calcium ATPase, or other regulatory ATPases. Compartmentalized isoforms of CK provide an adaptive mechanism which can rephosphorylate ADP to ATP, using creatine phosphate, thus maintaining the free ADP concentration at an appropriately low level. Another compartmentalized mitochondrial CK at the site of mitochondrial ATP generation catalyses the phosphorylation of creatine to creatine phosphate, thus completing the two components of a creatine phosphate shuttle pathway. It is now evident that many cells...

CNS neurons

Neurons of the central and peripheral nervous system are morphologically polarized cells, being highly specialized for the reception and transmission of (synaptic) information at the axon and dendrites, respectively. We have found that B-CK and mitochondrial creatine kinase are co-expressed in the same Golgi Type I (large) neurons, and not by the vast majority of central nervous system (CNS) interneurons and glia (Friedman and Roberts, 1994). Furthermore, these two isoforms of CK are expressed at morphologically distinct parts of the cell. The mitochondrial isoform of CK is localized primarily in the cell body of all Golgi Type I neurons. In contrast, B-CK mainly localizes within the cell processes, and to a lesser extent within the cell body. The distinct spatial arrangement of CK isoforms occurs throughout the brain including cerebral cortex, hippocampus, cerebellum and brainstem. Thus, the creatine phosphate shuttle of brain neurons serves to transfer metabolic energy from...

Epilepsy

Creatine, as an intracellular facilitator of high-energy phosphate transport and as an anti-oxidant, makes it a likely therapeutic candidate in epilepsy. There have been many recent spectroscopy studies examining alterations in creatine levels in vitro as well as in vivo in animal models of epilepsy and in humans. In vitro studies showed that stimulated hippocampal neurons fired faster and had faster recovery rates when phosphocreatine (PCr) NAA levels were higher (Williamson et al., 2005). In in vivo trials, creatine administration has been shown to ameliorate hypoxic seizures in both rabbit pups and rats (Holtzman et al., 1998, 1999). Rabbit pups were given subcutaneous creatine injections three days before inducing hypoxia. Early seizures were prevented and late seizures were significantly reduced by 60 . Additionally, brain levels of creatine, as detected by NMR spectroscopy, initially doubled as compared to controls and eventually reached adult levels (Holtzman et al., 1999)....

Research Outlook

Multidisciplinary approaches to creatine research will lead to new insights into the functional relationships between the BBB and iBRB, glia, and neurons (photoreceptor cells) for brain and retinal energy homeostasis and will offer improved therapeutic strategies for brain and retinal disorders. Genetic manipulation of BBB and iBRB function in mice or rats, e.g. by BBB-, iBRB- or glia-selective gene transfer and silencing, will be a future key method. Using such techniques will be a particularly important issue to investigate the relationships between creatine transport dysfunction at the BBB and iBRB and mental retardation, speech problems, language delay, and epilepsy. We recently produced a transgenic rat harboring the mouse Tie2 promoter enhancer linked green fluorescent protein (GFP) gene (Ohtsuki et al., 2005). The mouse Tie2 Oral administration of creatine is a potentially promising treatment for neurode-generative diseases owing to the neuroprotective effects found in animal...

Prospects

CCDSs are potentially treatable disorders. However, progress in understanding efficacy of treatment and development of new treatment strategies has been delayed due to the rareness of the single disorders. As with other rare inborn errors of metabolism, worldwide networks and orphan disease registries are needed to facilitate progress in understanding the natural history and in the development of strategies for treatment and prevention. Creatine deficiency syndromes should be investigated using these tools in the future. Development of treatment strategies depends on the understanding of pathophysiology. Our understanding of pathophysiology and pathobiochemistry in CCDSs is still incomplete. The pathobiochemical actions and pathophysiological consequences of accumulation of GAA, GAA's role in brain function, and pharmacological inhibition of GAA's action in the brain need to be understood fundamentally in order to develop more effective treatment strategies for GAMT deficiency. The...

Conclusion

Neuroprotective compounds targeting identified pathologic mechanisms of disease have the potential to delay the onset and slow the progression of neurological diseases. A large number of studies in both patients and experimental model systems have validated high-energy phosphate deficiency as a promising therapeutic target in neurological diseases. Compounds such as creatine buffer neuronal energy demands and are attractive candidates for targeting this important disease mechanism. Creatine has some advantages over other similar-acting compounds, which include more straightforward bioavailability and the potential to serve as an in vivo biomarker of premanifest and manifest disease. Creatine is available for human use and represents an immediate candidate as a neuroprotective agent for large-scale clinical trials in neurological diseases. Given the safety and tolera-bility of creatine, it may be especially well suited for long-term use in neurological disorders, providing prophylactic...

David L Christie

Abstract Creatine and phosphocreatine provide an intracellular, high-energy phosphate buffering system, essential to maintain ATP levels in tissues with high energy demands. A specific plasma membrane creatine transporter (CRT) is required for the cellular uptake of creatine. This transporter is related to the -aminobutyric acid (GAT) and norepinephrine (NET) transporters and is part of a large gene family of Na+- and Cl--dependent neurotransmitter transporters, now known as solute carrier family 6 (SLC6). CRT is essential for normal brain function as mutations in the CRT gene (SLC6A8) result in X-linked mental retardation, associated with the almost complete lack of creatine in the brain, severe speech and language delay, epilepsy, and autistic behaviour. Insight into the structure and function of the CRT has come from studies of creatine transport by tissues and cells, in vitro studies of CRT mutations, identification of mutations associated with CRT deficiency, and from the recent...

Mark A Tarnopolsky

Abstract Many of the neuromuscular (e.g., muscular dystrophy) and neurometabolic (e.g., mitochondrial cytopathies) disorders share similar final common pathways of cellular dysfunction that may be favorably influenced by creatine monohydrate (CrM) supplementation. Studies using the mdx model of Duchenne muscular dystrophy have found evidence of enhanced mitochondrial function, reduced intra-cellular calcium and improved performance with CrM supplementation. Clinical trials in patients with Duchenne and Becker's muscular dystrophy have shown improved function, fat-free mass, and some evidence of improved bone health with CrM supplementation. In contrast, the improvements in function in myotonic dystrophy and inherited neuropathies (e.g., Charcot-Marie-Tooth) have not been significant. Some studies in patients with mitochondrial cytopathies have shown improved muscle endurance and body composition, yet other studies did not find significant improvements in patients with mitochondrial...

Excretory epithelia

Creatine kinase immunoreactivity is most intense in the inner stripe of the outer medulla of the kidney. Both mitochondrial and BB-CK isoforms are specifically localized to the distal nephron, especially in the thick ascending limb. The pattern of renal expression of CK correlates with the region of greatest oxygen consumption, ATP utilization and sodium transport (Friedman and Perryman, 1991). In the very specialized sodium excretory (rectal) gland of the shark, there are also high levels of expression of multiple CK isoforms and PCr (Epstein et al., 1981 Friedman and Roberts, 1992). Stimulation of this excretory epithelial gland with cAMP causes PCr levels to rapidly decrease with no change in the levels of ATP (Epstein et al., 1983). Both the mitochondrial CK and BB-CK are expressed at high levels, equivalent to that measured in mammalian myocardium. Localization of the two isoforms is restricted to the basal but not the apical aspect of the tubules, consistent with the energy...

Concluding Remarks

As described in this chapter, MRS provides unique contributions to the understanding of creatine-related bioenergetics and metabolism in tissues of transgenic animals. Typical properties of MRS, such as its non-destructive and non-invasive character, which enables dynamic and longitudinal studies, are major assets as compared to more invasive approaches in investigations of tissues in which rapid changes in bioenergetics and metabolism may occur. Because of its relatively crude spatial resolution, the use of results from MRS examinations in the interpretation of processes at the (sub)cellular level may not be trivial and will require careful consideration of morphological, physiological, and biochemical knowledge of the tissue being examined, or dedicated experimental approaches (e.g., Wiseman and Kushmerick, 1995). Cellular heterogeneity in tissues may be an issue when the tissue composition changes due to the mutation introduced. Phenotypic consequences may also not be 100...

Joseph F Clark

Key words creatine, creatine kinase, creatine phosphate, 3-guanidinopropi-onic acid, mutation, NMR Many functions have been assigned to the creatine phosphate (PCr) creatine kinase (CK) system and some of these are described in Chapters 2, 4, 5 and 7. Little work has been performed, however, to determine how essential the CK system is in the normal function of the entire organism. Basically, there are two ways in which one can evaluate the requirement for a functioning CK system in vivo. The first method involves altering the intracellular substrates in order to short-circuit the system. This has been done using various creatine analogues such as 3-guanidinopropionic acid ( 3-GPA), cyclocreatine and 3-guanidinobuteric acid (Zweier et al., 1991 Clark et al., 1994 Boehm et al., 1995). Creatine analogues act as competitive CK inhibitors owing to their different kinetics and affinities (Wallimann et al., 1992 Clark et al., 1994). Another technique for examining the CK system in vivo is to...

Ppm 150

Selected region of 13C MR spectra of adult hindlimb muscle after administration of 13C-4 labeled creatine. M-CK ScCKmit-- -- double knockout animals show uptake and phosphorylation of 13C-labeled creatine (right) although PCr tCr ratios are significantly lower than in wild-type animals (left). Adapted from in't Zandt et al. (2003) and reprinted with permission

The Book

The appearance in print of the present volume of the Subcellular Biochemistry Series entitled Creatine and Creatine Kinase in Health and Disease, edited by Gajja S. Salomons and Markus Wyss, seems entirely timely. The importance and physiological significance of creatine kinase (CK) as well as the pleiotropic effects of creatine (Cr) and phosphocreatine (PCr) in health and disease have been largely underappreciated historically. Based on new discoveries in recent years, however, they are currently attracting much interest and even experience center stage attention, for instance with the recently announced large clinical Cr study with Parkinson's patients in the USA (Couzin, 2007). The comprehensive earlier review on Creatine and Creatinine Metabolism by Markus Wyss and Kaddurah-Daouk (2000), as well as the most recent review on Cr by John and Margaret Brosnan (2007), give a pre-taste of this new and exciting era of CK- and Cr-related research to come.

Characterization of Translation

Augment the host system (e.g. initiation or elongation factors) have not yet been identified. Each viral protein species appears to be encoded by a unique mRNA. Cells infected with HCMV do not show the generalized early shutoff of cell protein synthesis that occurs in HSV-1 infected cells. There are, nevertheless, very early changes in the metabolism of HCMV-infected cells that include stimulated transcription of the genes for heat shock protein 70, ornithine decarboxylase, thymidine kinase, and creatine kinase decreased transcription of the fibronectin gene and changes similar to those induced by the G protein signaling pathway, such as decreased intracellular Ca2+ stores and increased levels of intracellular cAMP.

Neuroleptic Malignant Syndrome

Neuroleptic malignant syndrome is usually thought of as a syndrome of fever and catatonic rigidity, with increased serum creatine phosphokinase levels in the setting of neuroleptic administration. The etiology of this disorder is obscure, and lack of consistent case definitions has hampered fuller study of this syndrome. The initial description of neuroleptic malignant syndrome referred to it as pallor and hyperthermia with isolated primary symptoms of fever, akinesia or stupor with hypertonicity, and pulmonary congestion. Since then, a number of diagnostic criteria have been proposed as summarized at length by Adityanjee, Mathews, and Aderibigbe in their 1999 review. In essence, the syndromic definitions stress the presence of rigidity, fever, and autonomic changes in the presence of neuroleptic exposure. Alteration in sensorium is also a common feature of several of these criteria.

Pathological features

Depending on the results of the laboratory tests, those patients meeting the criteria above are classified into groups listed below. The following studies are suggested complete blood count, erythrocyte sedimentation rate, chemistry profile, creatine kinase, antinuclear antibody, thyroid functions, serum and urine immunoglobulin studies (to include either immunofixation electrophoresis or immunoelectrophoresis, and HIV and hepatitis serology. The list of laboratory studies is not comprehensive. For instance, in certain clinical circumstances, other studies may be indicated, such as phytanic acid, long-chain fatty acids, porphyrins, urine heavy metals, a-lipoprotein, p-lipoprotein, glucose tolerance test, imaging studies of the central nervous system, and lymph node or bone marrow biopsy. Classification of CIDP

Subcellular Systems For Studying Pglycoprotein

Plasma membrane vesicles have also proved useful in studies of Pgp-mediated drug transport. Most vesicle preparations consist of a mixture of right-side-out and inside-out vesicles,54 and if they are well sealed, the latter population can transport drug from the external medium into the vesicle lumen when provided with ATP. When using a vesicle system where other membrane-bound ATPases are present, it is often necessary to add an ATP-regenerating system, such as creatine kinase and creatine phosphate, to prevent rapid depletion of ATP in the external solution. Substrate uptake into the vesicle interior can be measured in one of two ways. If drug is available in radioactive form (e.g., 3H colchicine, 3H vinblastine, 125I peptide), it is added to the vesicle preparation at time zero, together with ATP and a regenerating system, and vesicles are removed at various times (typically, ranging up to 30 minutes) and collected by rapid filtration.54 Drug uptake into the vesicles increases with...

Statistical Approaches for Data Analysis

Duchenne muscular dystrophy (DMD) on the basis of serum creatine kinase (CK) measurements, we developed a procedure to combine independent serial measurements of CK on individuals into a single index which then was combined with family history information to yield the probability that the individual being tested was a carrier of the DMD gene (299). In order to efficiently distinguish female carriers of hemophilia from healthy normal individuals, logistic coefficients were derived from the ratio of measurements of factor VIII activity (which is defective in hemophilia A) and von Willebrand factor (to which factor VIII binds) into a single index which was combined with family history information to yield the probability that the individual being tested was a carrier of hemophilia A (300).

Hormonal Interactions During Exercise

For the few seconds of the 100-yard dash, endogenous ATP reserves in muscle, creatine phosphate and glycogen, are the chief sources of energy. For short-term maximal effort, energy must be released from fuel before circulatory adjustments can provide the required oxygen. Breakdown of glycogen to lactate provides the needed ATP and is activated in part through intrinsic biochemical mechanisms that activate glycogen phos-phorylase and phosphofructokinase. For example, calcium released from the sarcoplasmic reticulum in response to neural stimulation not only triggers muscle contraction but also activates glycogen phosphorylase. These intrinsic mechanisms are reinforced by epinephr-ine and norepinephrine released from the adrenal medullae and sympathetic nerve endings in response to central activation of the sympathetic nervous system.

Clinical Description Dm1

Ninety percent of DM1 patients present at adulthood with delayed muscle maturation, distal muscle weakness, wasting, myotonia, cataracts, cardiac abnormalities, smooth muscle dysfunction, insulin resistance, daytime sleepiness, testicular atrophy (low reproductive fitness), ''difficult'' personality, neuropsychiatric disturbances, and frontal balding. 1 Ten percent of the patients present at infancy with hypotonia (floppy infant), oromotor dysfunction, tent-shaped mouth, feeding and respiratory insufficiency (diaphragmatic hypoplasia), arthrogryposis, and mental retardation in those who survive until adulthood (congenital DM). 1 All manifestations show a progressive course. Usually, creatine kinase is elevated. Muscle biopsy shows type 1 predominance, centrally located nuclei, severe fiber atrophy with nuclear clumps, hypertrophic and angulated fibers, and occasionally, necrotic fibers, fibrosis, or fat deposits. Cardiac involvement comprises conduction defects (mostly HV...

Clinical Findings

Clinical symptoms usually appear in adulthood and are characterized by exercise intolerance with muscle cramps that can be accompanied by attacks of myoglobinuria. Serum creatine kinase is usually elevated and increases after exercise. Avoidance of strenuous exercise can prevent major episodes of rhabdomyolysis.

If You Take Oral Medicines

In most cases you can take your usual medicines the day before the procedure, but none on the day of the procedure, and then restart them when you start eating. If you are on metformin and you need a procedure where you get a special x-ray with contrast dye, you may be asked to stop your metformin for a couple of days until a serum creatine confirms that your kidneys are not affected by the contrast dye. When you stop metformin, your glucose levels may run high, and your doctor may ask you to take tolbutamide, repaglinide, or nateglinide, or even a little bit of insulin, to control the glucose levels until you can restart the metformin.

Animal Models of Motor Neuron Disease

SOD1 transgenic rodent models most accurately recapitulate the major clinical and histopathological hallmarks of ALS and thus have been the standard model in which to evaluate novel drugs (Ripps et al. 1995 Lee et al. 1996). Further, these animals carry a genetic mutation demonstrated in a group of individuals with familial ALS. However, studies which appear to alleviate ALS in rodents do not always translate into successful human therapeutics. For example, creatine, a nitrogenous organic acid shown to improve mitochondrial function, was found to extend survival in SOD1 mice (Klivenyi et al. 1999) but had no beneficial effect in human trials (Groeneveld et al. 2003). Results should hence be interpreted with caution, taking into consideration both the faster rate of degeneration of motor neurons in high expressing SOD1 mice and the relative uncommonness of SOD1 mutations in ALS patients (1-2 ).

Neuromuscular and Cutaneous Syndromes

Polyneuropathy, polymyositis, and myopathy are all known to occur with RCC. The myopathy is characterized by involvement of proximal muscles and association of elevated serum levels of creatine kinase and aldolase (Solon etal. 1994 Evans etal. 1990). In each condition, there are reports of resolution after nephrecto-my. With the development of metastatic disease, poly-myositis and polyneuromyopathy have recurred.

Oxidative And Nitrosative Induced Cell Death In Diabetic Neuropathy

NADH coupled with failure of the Mt creatine phosphate pump to regenerate ATP from ADP also results in disruption of the Mt electron transfer chain, and depletion of ATP (56). Depletion of ATP is associated with apoptosis in vitro (20), and similar changes are seen in chronic neuropathy in diabetic animals (57). Oxidative stress can be prevented by inhibitors of the Mt electron transport chain, for example, Mt complex II or III inhibitors, for example, TTFA or myxothiazole (20,58). Failure to prevent oxidative stress results in Mt DNA damage. DNA damage might be repaired by base excision repair, however, if not repaired there is furthermore disruption of the electron transport chain and production of further ROS. This vicious cycle of ROS production and Mt DNA damage ultimately leads to energy depletion in the cell and apoptosis (29,59).

Figure

Most enzyme biosensors use either membrane-based enzyme entrapment, or the enzyme is covalently bound to the inside of a nylon tube. The stability of the electrode is dependent on the stability of the enzyme, which is partially dependent on the method of immobilization. Many enzyme electrodes are available to detect, for example, glucose, urea, creatine, and pyruvate in clinical samples. However, only the glucose biosensor has been widely commercialized.

Clinical Diagnosis

Serum creatine kinase (CK) is markedly increased. This marked elevation of serum CK is the most important hallmark for the diagnosis of BMD, but the level of elevation of serum CK is not so high compared to DMD. During the asymptomatic period, elevation of serum CK is the sole sign for BMD. Some BMD patients are identified accidentally because of elevations of AST or ALT which are commonly examined for liver function as serum CK elevation is accompanied with elevations of AST and ALT.

Eukaryotic Genes

The existence of introns allows for the creation of multiple proteins from one gene, by the use or exclusion of different exons. Such alternative splicing gives rise to protein isoforms, highly similar but slightly different proteins, with functions that vary as well. Isoforms are typically tissue-specific. For example, the muscle enzyme creatine kinase exists in one form in the heart, and another form in the skeletal muscles (such as the biceps), which have different ends formed through use of different exons. Even though it codes for two or more proteins, most scientists call such a DNA sequence a single gene.

Muscular System

The ATP for muscle contractions can come from four sources aerobic oxidation, or glycolysis with fermentation to lactic acid, creatine phosphorylation, and free ATP. Under resting conditions, muscle cells produce ATP from the oxidation of fatty acids via aerobic respiration. Because it requires oxygen, this mechanism can provide energy only as fast as the required amount of oxygen can be provided to the muscle by the bloodstream. Although this is most efficient, aerobic respiration does not provide ATP fast enough for continuous contractions such as those during athletic performances. It can, however, support moderate continuous activity such as hiking or, literally, aerobic exercise. For very brief, explosive activity, such as a single leap or a throw, the muscle cell has two immediate sources of ATP. One is the small amount of free ATP always present in cells. This can support about 2 seconds of contractions. The second immediate source is a high-energy compound called creatine...

Autonomic Ganglia

Prominent physiological and less dramatic pathological changes are present in auto-nomic ganglia of DAN. There is a significant reduction in blood flow in autonomic ganglia such as superior cervical ganglion (32,33). This reduction by about 50 is present as early as 1 week and is persistent over 24 weeks (4). Glucose uptake was reduced to 30 of control values in superior cervical ganglion in rats with DAN. a-Lipoic acid supplementation had no effect on glucose uptake in normal nerves at any dose, but reversed the deficit in DAN, with a threshold between 10 and 25 mg kg. ATP, creatine phosphate, and lactate were measured in sciatic nerve and superior cervical ganglion. a-Lipoic acid prevented the reduction in autonomic ganglion creatine phosphate (34).

In Diabetes

As mentioned above there is circumstantial evidence that nitrosative stress and peroxynitrite formation importantly contribute to the pathogenesis of diabetic cardiomyopathy both in animals and humans. We have tested a novel metalloporphyrin peroxynitrite decomposition catalyst, FP15, in murine models of diabetic cardiovascular complications (92). We hypothesized that neutralization ofperoxynitrite with FP15 would ameliorate the development of cardiovascular dysfunction in a STZ-induced murine model of diabetes. To ensure that the animals received the FP15 treatment at a time when islet cell destruction was already complete and hyperglycemia has stabilized the treatment was initiated 6 weeks after the injection of STZ. Although FP15 did not affect blood glucose levels, it provided a marked protection against the loss of endothelium-depen-dent relaxant ability of the blood vessels (Fig. 1A) and improved the depression of both diastolic (Fig. 1B) and systolic function of the heart (92)....

Multicenter Trial

As indicated by Ebbeling and Clarkson (1989), muscle soreness and elevations in serum creatine kinase (CK) can be used as a biochemical marker for skeletal muscle injury. Recently a study was conducted to evaluate the hypothesis that exposure of skeletal muscle to exercise-induced stress in combination with prior administration of a study drug (e.g., drug A) will produce a greater increase in CK than the effects of exercise alone. This

Heart Disease

The ability of PA to chelate iron and possibly reduce its free radical generating potential and subsequent lipid peroxidative damage Figure 14.3(C) may also protect the heart from ischemic and reperfusion injury 113 . Rao et al. 113 intravenously injected rats with saline or PA at levels up to 15 mg per 100 g of BW. Shortly after, the hearts were excised, and reperfusion injury was induced in vitro. The higher levels of injected PA resulted in lessened reperfusion injury by significantly reducing creatine kinase release, decreasing lipid peroxidation

Who to screen

More frequent screening is indicated in patients with known risk factors such as the presence of a goiter, a history of type 1 diabetes mellitus, previous thyroid surgery or radiation, pernicious anemia, premature gray hair, vitiligo, or any previous history or family history of thyroid disease. Additionally, in any patient with an unexplained laboratory abnormality such as hypercholesterolemia, hyponatremia, anemia, hypercalcemia, or creatine kinase elevation serum TSH levels should be determined (11). Patients with new-onset atrial fibrillation should also be tested, since approximately 15 will be thyrotoxic (13,14). Patients should be evaluated for thyroid dysfunction before starting therapy with iodine-containing medications and then periodically while taking those medications.

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