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1 Bristol Heart Institute, University of Bristol, Bristol Royal Infirmary, Bristol BS2 8HW, UK2 Department of Biochemistry, School of Medical Sciences, University Walk, Bristol BS8 1TD, UK
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Abstract |
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(Received 5 January 2004;
accepted after revision 3 February 2004;
first published online 6 February 2004)
Corresponding author N. King: Bristol Heart Institute, Bristol Royal Infirmary, Bristol BS2 8HW, UK. Email: n.king{at}bris.ac.uk
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Introduction |
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Exposure to ischaemia–reperfusion has been shown to induce a fall in the endogenous myocardial aspartate concentration in both experimental models (Pisarenko et al. 1995) and patients (Suleiman et al. 1997). Exogenous aspartate has been implicated in myocardial protection (Pisarenko, 1996), but this remains controversial (Buckberg, 1996). One possible reason for the controversy could be the poor understanding of aspartate transport in the heart. The likelihood that exogenous aspartate will be taken up into heart cells in order to improve metabolism and function will be dependent upon the characteristics of aspartate transport across the cardiac sarcolemma.
Recently we characterized aspartate transport in normal rat heart using sarcolemmal vesicles and isolated cardiac myocytes (King et al. 2001). This revealed a high affinity sodium-dependent transport system, which was inhibited by L- but not D-glutamate. These characteristics, consistent with the Xag– transport system, were accompanied by expression in rat heart cells and vesicles (King et al. 2001) and human heart vesicles (N. King unpublished observations) of the system Xag– transporter, EAAT3.
All of the above work has been carried out using the normal heart, whereas very little is known about the adult hypertrophied heart in terms of aspartate transport or in relation to the ability of aspartate to impart myocardial protection. In truth, there are few studies investigating myocardial protection techniques during cardiac surgery in patients with ventricular hypertrophy (Anderson et al. 1995; Jin et al. 1995; Calafiore et al. 1996; Dorman et al. 1997). This is mainly due to the fact that techniques used to protect hearts with ischaemic disease are uncritically extended to hypertrophied hearts. Hypertrophied hearts are metabolically different from ischaemically diseased heart (Suleiman et al. 1998; Ascione et al. 2002), which will have implications for the efficacy of cardioplegic techniques.
The aim of this study was twofold. First we tested the hypothesis that aspartate transporter expression was different in the hypertrophied compared to the normal heart. Secondly, we hypothesized that the expression and characteristics of aspartate transport in the hypertrophied heart would influence the effectiveness of aspartate in protecting the hypertrophied heart from an ischaemic insult.
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Methods |
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L-[14C]Aspartate and Rainbow molecular weight markers were obtained from Amersham (Little Chalfont, Buckinghamshire, UK); collagenase Type I, was from Worthington Biochemical Corporation (Lakewood, NJ, USA); anti-EAAT3 monoclonal antibody was from Chemicon (Temecula, CA, USA); horseradish peroxidase (HRP)-conjugated anti-rabbit or -mouse IgG was from Dako A/S (Denmark). All other chemicals were from either Sigma or BDH and were of analytical grade.
Source of normal and hypertrophied hearts
Hearts were obtained from two strains of male rats, namely the spontaneous hypertensive rat (SHR) and their corresponding normotensive control, the Wistar Kyoto (WKY) (Doggrell & Brown, 1998; Atlante et al. 1996). The rats were age matched at 18 ± 3 weeks old, which was chosen to coincide with reports documenting the development of myocardial hypertrophy in the SHR (Atlante et al. 1996; Doggrell & Brown, 1998). The rats were also weight matched at 354.7 ± 3.8 g WKY versus 349.2 ± 3.1 g SHR (n= 69, all values are means ±S.E.M.). Cardiac hypertrophy in the SHR was confirmed by the significantly greater ventricle weight (1389 ± 28 mg SHR versus 1151 ± 29 mg WKY, P > 0.001, t test, n= 69) and significantly greater ventricular mass index (0.599 ± 0.03 WKY versus 0.738 ± 0.03 SHR, P < 0.02, t test, n= 69), with these values consistent with previous reports about WKY and SHR (Doggrell & Brown, 1998; Atlante et al. 1996).
All rats were killed humanely by cervical dislocation and the hearts processed for the preparation of cardiac sarcolemmal vesicles or for Langendorff perfusion for either cell isolation or protection studies.
Preparation of cardiac sarcolemmal vesicles
Homogenization and differential centrifugation were used to prepare cardiac sarcolemmal vesicles from normal and hypertrophied hearts as previously described (King et al. 2001). Marker enzyme assays were used to assess the purity of the completed vesicle samples. The enzymes measured were Na+,K+-ATPase (sarcolemmal membrane), Ca2+,(K+)-ATPase (sarcoplasmic reticulum) and Mg2+-ATPase (other intracellular membranes) (King et al. 2001). All of the vesicles were enriched in Na+,K+-ATPase activity (21 ± 8-fold normal versus 17 ± 2-fold hypertrophied, n= 6) without any enrichment in Ca2+,(K+)-ATPase activity (1 ± 0.1-fold in both preparations, n= 6) or Mg2+-ATPase activity (1.4 ± 0.2-fold normal versus 1.3 ± 0.1-fold hypertrophied, n= 6). There were no significant differences in the enrichment and recovery of marker enzymes in vesicles prepared from normal versus hypertrophied hearts. A comparison was made of the protein concentration in the completed vesicle samples. This was performed as previously described (King et al. 2001) with all the vesicles whether from hypertrophied or normal hearts suspended in 0.75 ml. There were no significant differences between the protein concentrations of the normal (4 ± 1 mg ml–1, n= 6) compared to the hypertrophied (3.4 ± 0.3 mg ml–1, n= 6) vesicle samples. The protein concentration of whole normal and hypertrophied hearts was not determined.
Measurement of L-[14C]aspartate uptake into cardiac sarcolemmal vesicles
The initial rate at 1 s of 0.001–0.3 mmol l–1L-[14C]aspartate uptake was carried out by rapid filtration at room temperature as previously described (King et al. 2001). The internal solution of the vesicles was (mmol l–1): 100 potassium gluconate, 100 mannitol, 10 Hepes/Tris (pH 7.4). The transport solution was (mmol l–1): 100 NaCl or 100 choline-Cl, 100 mannitol, 10 Hepes/Tris (pH 7.4) and 0.001–0.3 L-[14C]aspartate. Results are presented as the sodium-dependent rate of L-[14C]aspartate uptake, calculated as the uptake in the NaCl containing solution minus the uptake in the choline-Cl containing solution.
Preparation of isolated cardiac myocytes
A combination of enzyme digestion and mechanical dispersion was used to isolate cardiac ventricular myocytes from normal and hypertrophied hearts (King et al. 2001). The viability and morphology of the isolated cells was examined by light microscopy and Trypan Blue exclusion. This indicated that > 80% of the myocytes from normal and hypertrophied hearts were rod shaped and able to exclude Trypan Blue.
Western blotting of EAAT3 and glutamate dehydrogenase
Expression of EAAT3 and glutamate dehydrogenase (GDH) proteins in normal and hypertrophied samples were investigated by Western blotting (King et al. 2001). EAAT3 protein expression was investigated in cardiac sarcolemmal vesicles and isolated cardiac myocytes. The anti-GDH antibody was manufactured in the University of Bristol following an identical procedure to that previously described (Nicholson & McGivan, 1996), except that the initial peptide was designed from the C terminal sequence of GDH. GDH expression was investigated using the same isolated cardiac myocyte samples that were used to investigate EAAT3 protein expression. In order to achieve this, the blots were stripped using a kit from ECL and then re-probed.
Every sample was loaded onto the gel containing precisely the same amount of protein, namely 50 µg. Band density on the resultant X-ray films was visualized and quantified on a Bio-Rad scanning densitometer (Bio-Rad Laboratories, Hemel Hempstead, UK) with incorporation into Molecular Analyst software (version 2.1 for Macintosh, Bio-Red). The results were expressed as OD units mm2 with the values for EAAT3 being adjusted for the internal control (i.e. subtraction of the density measured for the respective normal and hypertrophied glutamate dehydrogenase samples).
RT-PCR of system Xag-transporters
Total RNA was isolated from hearts and brains of SHR and WKY rats using TRI Reagent (Sigma) in accordance with the manufacturer's instructions. Complimentary DNA was synthesized using an Ambion kit (Ambion Inc. Cat. no. 1710) as instructed. cDNAs were stored at –20°C until use.
PCR was carried out in a reaction volume of 50 µl containing cDNA, dNTPs, polymerase buffer (Sigma), primers, and Taq DNA polymerase (Sigma). Amplification of EAAT2-4 was performed with initial denaturation at 94°C for 3 min followed by 27 cycles comprising 30 s at 94°C, 45 s at 50°C, 30 s at 72°C followed by a 7-min extension at 72°C. For EAAT1, amplification was performed as follows: initial denaturation at 94°C for 3 min, then 30 cycles of 30 s at 94°C, 30 s at 60°C, and 60 s at 72°C with a final extension at 72°C for 7 min. Amplified products were subjected to electrophoresis in 1% agarose gels, and visualized by ultraviolet illumination in the presence of ethidium bromide (0.1 mg ml–1).
These PCR conditions and the primer sequences were obtained from literature reports (Suchak et al. 2003; Velasco et al. 2003). In brief, the specific oligonucleotide primers were for EAAT1: sense 5'-CTACTCACCGTCAGCGCTGT-3' and antisense 5'-AGCACAAATCTGGTGATGCG-3' (expected product size 1019 bases) (Velasco et al. 2003); EAAT2: sense 5'-ATGTCTTCGTGCATTCGGTGTTGGG-3' and antisense 5'-AGCCGTGGCACCATCTTCATAGC-3' (expected product size 326 bases); EAAT3: sense 5'-CCCAAACGCATCACCCAGAAC G-3' and antisense 5'-TCGGCAACCCTTCCAGTTACATTCC-3' (expected product size 374 bases); EAAT4: sense 5'-GCTTGTGCCATGAGTGACTTATAGG-3' and antisense 5'-CGTGTCCTGAGGGATTTCTTCG-3' (expected product size 674 bases) (Suchak et al. 2003).
Langendorff heart perfusion
The use of the Langendorff preparation in this lab has been previously described (Javadov et al. 2000). In brief, the perfusing solution was Krebs solution containing (mmol l–1): 118 NaCl, 25 NaHCO3, 4.8 KCl, 1.2 KH2PO4, 1.2 MgSO4, 11 glucose and 1.2 CaCl2. This was delivered at 11 ml min–1; maintained at 37°C; and aerated with 95%O2–5% CO2 (pH 7.4). A water-filled balloon connected to a pressure transducer was inserted into the left ventricle to monitor cardiac performance.
After an initial equilibration period of 30 min, the pump was switched off and the heart immersed in solution at 37°C to induce global normothermic ischaemia. Reperfusion was initiated by restarting the pump and was continued for 60 min. Initially, an ischaemic period of 40 min was used for all hearts, but in later experiments, the ischaemic period for normal hearts was increased to 70 min in order to match the level of functional recovery and reperfusion injury between the control normal and hypertrophied hearts (see Results). Where used, 0.5 mmol l–1 aspartate was added to the perfusate 10 min into the equilibration period and was present throughout ischaemia and reperfusion.
The effect of ischaemia and reperfusion on membrane integrity was assessed from the release of lactate dehydrogenase (LDH) into the effluent. This was measured with a Cell Cytotoxicity kit (LDH) from Roche using known quantities of lactate dehydrogenase (Sigma) as a standard.
Collection and analysis of intracellular metabolites
ATP, lactate and free amino acid concentrations were measured in biopsy samples taken from normal and hypertrophied hearts. Samples were obtained from freshly isolated hearts after a brief perfusion with Krebs solution to flush out the blood. Tissue samples were also obtained from a different set of hearts at the beginning (0 min) and end of ischaemia (40 min for hypertrophied or 70 min for normal) with or without aspartate addition to the perfusate.
ATP in neutralized extract was separated and quantified by a high performance liquid chromatography technique based on previous reports (Imura et al. 2001). Amino acids were determined according to the Waters Pico-Tag method as reported previously (Imura et al. 2001), whilst lactate was measured using a kit from Sigma.
Data analysis
Data are presented as means ±S.E.M. comparisons between datasets were made using either the student's t test or ANOVA with an appropriate post test. This was performed using INSTAT (Graphical Software, Inc.) on an IBM compatible computer. P < 0.05 was considered significant.
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Results |
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EAAT3 protein was expressed in cardiac sarcolemmal vesicles (Fig. 1A) and cardiac myocytes (Fig. 1B) isolated from normal and hypertrophied hearts. In the vesicles, the mean density of the bands from hypertrophied hearts was 8.5 ± 0.5 optical density (OD) units mm2, which was significantly greater than the 2.4 ± 0.5 OD units mm2 measured for normal hearts (P < 0.05, t test, n= 5). In isolated cardiac myocytes, the mean density of the bands from hypertrophied samples was 1.8 ± 0.3 OD units mm2, significantly greater than the 0.1 ± 0.4 OD units mm2 in the normal samples (P < 0.01, t test, n= 6).
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Effect of hypertrophy on expression of mRNA for the system Xag– transporters
The possibility that other system Xag– transporters were expressed in normal and hypertrophied hearts was investigated by reverse transcription-polymerase chain reaction (RT-PCR). Figure 2 shows the results of experiments using specific primers directed against EAAT1-4. Of these four transporters, EAAT1 and EAAT3 were detected with bands of a similar intensity in both normal and hypertrophied hearts, whilst EAAT2 and EAAT4 were not detected in any heart samples. In control experiments using samples of brain mRNA isolated from the normal and Spontaneously hypertensive rats all four transporters were detected (not shown).
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The kinetics of sodium dependent L-[14C]aspartate uptake were measured in the same vesicle populations from normal and hypertrophied hearts that had been used to investigate EAAT3 expression. These results are shown in Fig. 3. The affinity (Km) of L-[14C]aspartate transport was similar in the two groups, whilst the maximal velocity (Vmax) of L-[14C]aspartate uptake for vesicles from hypertrophied hearts was significantly higher than for vesicles from normal hearts (n= 6, t test, P < 0.001).
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The effect of 40 min global normothermic ischaemia on the function and recovery of normal compared to hypertrophied hearts is shown in Table 1. Under basal conditions, hypertrophied hearts had a significantly greater left ventricular developed pressure (LVDP) and higher rate pressure product (RPP; heart rate x LVDP). During ischaemia, the hypertrophied hearts tended to enter rigor more swiftly than the normal hearts, although this did not quite reach significance. Rigor contracture was significantly greater in hypertrophied hearts compared to normal hearts. The most dramatic differences occurred during reperfusion, when in every parameter investigated the hypertrophied hearts were significantly worse in comparison to the normal hearts and in comparison to the basal values.
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After 40 min ischaemia, the functional recovery and membrane integrity of normal hearts were clearly superior to those of hypertrophied hearts. Therefore, in order to properly assess the effects of aspartate addition on normal compared to hypertrophied hearts, the hearts were matched as to the level of functional recovery and reperfusion injury evident during reperfusion. This was achieved by increasing the ischaemic period for the normal hearts to 70 min. Table 2 shows how under these conditions, the performance and outcome of the normal and hypertrophied hearts were affected by the addition of 0.5 mmol l–1 aspartate to the perfusate.
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The effect of aspartate addition upon reperfusion injury in the normal and hypertrophied hearts was also assessed from LDH loss as shown in Fig. 4B. The total LDH release from hypertrophied hearts with aspartate was significantly less than control hypertrophied hearts and both groups of normal hearts.
Changes to the amino acid pool in normal and hypertrophied hearts
Table 3 shows the concentration of aspartate, glutamate, glutamine and alanine in freshly isolated normal and hypertrophied serum and hearts. There were no significant differences in the tissue, serum or tissue–serum gradient between the normal and hypertrophied groups.
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At the beginning of ischaemia, the ATP concentration was similar in all hearts with 11.1 ± 1.8 and 13.4 ± 1.9 nmol (g protein)–1 in normal hearts with and without aspartate compared to 14.9 ± 1.4 and 11.8 ± 1 nmol (g protein)–1 in hypertrophied hearts with and without aspartate. At this time the alanine/glutamate ratio was also comparable in the four groups of hearts with 0.4 ± 0.1 in both groups of normal hearts and with 0.3 ± 0.1 in both groups of hypertrophied hearts. The lactate concentration at the beginning of ischaemia in each of the four groups was not significantly different from zero. Come the end of ischaemia, the ATP concentration was higher, the lactate concentration lower and the alanine/glutamate ratio smaller in the hypertrophied hearts with aspartate compared to any of the other groups (Fig. 5).
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Discussion |
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Hypertrophy increases expression of EAAT3 protein and the rate of aspartate uptake
Expression of the aspartate transporter EAAT3 protein in cardiac sarcolemmal vesicles and cardiomyocytes (Fig. 1A and B) from hypertrophied hearts was greater than in samples from control hearts. This increase did not represent a global change in protein expression in hypertrophy, since the level of expression of glutamate dehydrogenase was similar in cells from hypertrophied and normal hearts (Fig. 1C). In fact, these results reflect a general pattern of selected change either up or down to different metabolic enzymes during the progression/regression of cardiac hypertrophy (Rossi & Lortet, 2001).
The increased EAAT3 expression in vesicles from hypertrophied hearts compared to normal hearts was accompanied by a significantly faster rate (Vmax) of aspartate transport in the same vesicle samples (Fig. 3). The absence of any significant difference in the Km of the two groups suggests that the same transporter facilitates aspartate uptake in normal and hypertrophied vesicles. The uptake characteristics in all the samples comprising high affinity, low capacity and sodium dependence (not shown) are entirely consistent with EAAT3 expression (Palacín et al. 1998; King et al. 2001).
EAAT3 belongs to a family of five transport proteins called the excitatory amino acid transporters (EAAT) (Palacín et al. 1998). mRNAs for two family members, EAAT1 and EAAT3, were expressed in the normal and hypertrophied hearts (Fig. 2). The absence of any obvious differences in band intensity suggests firstly that the mRNA level might be similar in the normal compared to the hypertrophied groups and secondly that the up-regulation of EAAT3 may occur at the protein level. However, because the effect of hypertrophy upon the expression of EAAT1 protein has yet to be determined, the possibility that EAAT1 could contribute to the up-regulation of aspartate transport and the improvement in protection of the hypertrophied heart cannot be ruled out. In a recent comprehensive review about amino acid transporters and signalling it was reported that there is currently no direct evidence to show that substrate binding to EAAT proteins controls direct interactions of these transporters with other proteins or the activity of transporter-associated signalling molecules (Hyde et al. 2003).
Does increased EAAT3 expression and activity improve protection with aspartate in hypertrophied hearts?
In the absence of any interventions hypertrophied hearts were more susceptible to 40 min ischaemia followed by reperfusion than normal hearts (Table 1 and Fig. 4). Although the vulnerability to ischaemia–reperfusion and changes to metabolites of hypertrophied hearts compared to normal hearts varies with different models and causes of hypertrophy (Allard et al. 1994; Ji et al. 1994), our results are qualitatively similar to other observations using the SHR/WKY model of hypertrophy (Snoeckx et al. 1986; Anderson et al. 1987).
After matching the recovery of the normal and hypertrophied hearts, perfusion with 0.5 mmol l–1 aspartate significantly improved the performance of the hypertrophied hearts without affecting normal hearts. This was demonstrated by a better functional recovery (LVDP and RPP, Table 2); reduced ischaemic stress (ATP and lactate, Fig. 5); and less reperfusion injury (LDH release, Fig. 4B) in the hypertrophied hearts perfused with aspartate compared to any of the other hearts.
One explanation for the improved performance of hypertrophied hearts would be that it resulted from aspartate loading of the hypertrophied cardiomyocytes stimulated by their higher level of EAAT3 expression and faster rate of aspartate transport compared to normal hearts. This raises the question as to whether other EAAT3 substrates would also protect hypertrophied hearts, although it is unlikely that poorly metabolized substrates such as D-aspartate would have any potential. Indeed in experiments using L-
-aminoadipic acid (Palacín et al. 1998), a poorly metabolized substrate of EAAT1-3, there was no effect either good or bad on cardiac function in normal or hypertrophied hearts (N. King, unpublished observations). On the other hand, there is strong circumstantial evidence to suggest that glutamate may protect hypertrophied hearts. Indeed, throughout the current experiment the two amino acids which were at a significantly greater level and were preserved better in hypertrophied hearts perfused with aspartate compared to any other group of hearts were glutamate and glutamine. The loss of these amino acids from normal hearts during baseline perfusion is consistent with previous findings in guinea-pig hearts (Suleiman & Chapman, 1993) and can possibly be explained by the large outward gradient, which is present for these amino acids during perfusion with Krebs solution.
Glutamate and glutamine have been implicated in myocardial protection in the normal heart (Pisarenko et al. 1995; Pisarenko, 1996), although their role in the hypertrophied heart has not been investigated. Both these amino acids can be formed via aspartate firstly through the actions of glutamate/aspartate aminotransferase (Pisarenko, 1996) to produce glutamate and then glutamine synthetase to yield glutamine (Abcouwer et al. 1995). A possible benefit of having higher levels of these amino acids at the beginning of reperfusion could be related to an improved ability to counteract ionic imbalances. During ischaemia sodium accumulates in the cardiomyocytes, which then contributes to the calcium overload that occurs upon reperfusion (Suleiman et al. 2001). This harmful chain of events could potentially be alleviated in hypertrophied hearts perfused with aspartate through the extrusion of sodium with either glutamine or glutamate. This could be accomplished via a reversal of the sodium-dependent glutamine (Rennie et al. 1996) and sodium-dependent glutamate (Dinkelborg et al. 1995) transporters that have been characterized in heart.
The choice to use 0.5 mmol l–1 aspartate was based on the low Km of EAAT3 as seen in the transport characteristics of aspartate in the vesicles (Fig. 3). This concentration is lower than that used by other investigators where typically hearts have been perfused with 3–13 mmol l–1 aspartate often in combination with either glutamate (Engelman et al. 1991; Morita et al. 1995) or -ketoglutarate (Pisarenko et al. 1995). These differences in addition to species differences and other variations in exact experimental protocol, may explain why perfusion with 0.5 mmol l–1 aspartate did not aid the normal hearts (Table 2).
In conclusion changes to aspartate transporter expression and activity in the hypertrophied heart positively influence the efficiacy of aspartate as a protective agent, which suggests that the use of aspartate as an additive to cardioplegic solution could provide significant protection to the hypertrophied heart.
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References |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Allard MF, Schönekess BO, Henning SL, English DR & Lopaschuk GD (1994). Contribution of oxidative metabolism and glycolysis to ATP production in hypertrophied hearts. Am J Physiol 267, H742–H750.[Medline]
Anderson WA, Berrizbeitia LD, Ilkowski DA, Cha R, Gu J, Fernandez J, Laub GW, Adkins MS, Chen C & McGrath LB (1995). Normothermic retrograde cardioplegia is effective in patients with left ventricular hypertrophy – a prospective randomised study. J Cardio Thorac Surg 36, 17–24.
Anderson PG, Bishop SP & Digerness SB (1987). Transmural progression of morphological changes during ischemic contracture and reperfusion in the normal and hypertrophied rat heart. Am J Pathol 129, 152–167.[Abstract]
Ascione R, Caputo M, Gomes WJ, Lotto AA, Bryan AJ, Angelini GD & Suleiman M-S (2002). Ischaemia-reperfusion in hypertrophic hearts of patients undergoing aortic surgery using cold or warm blood cardioplegia. Eur J Cardio Thoracic Surg 21, 440–446.[CrossRef]
Atlante A, Seccia TM, Marra E, Minervini GM, Vulpis V, Pirrelli A & Passarella S (1996). Carrier-mediated transport controls hydroxyproline catabolism in heart mitochondria from spontaneously hypertensive rats. FEBS Letts 396, 279–284.[CrossRef][Medline]
Buckberg GD (1996). Invited editorial on ‘effects of glutamate and aspartate on myocardial substrate oxidation during potassium arrest’. J Thorac Cardiovasc Surg 112, 1661–1663.
Calafiore AM, Teodori G, Bosco G, Di Giammarco G, Vitolla G, Fino C & Contini M (1996). Intermittent antegrade warm blood cardioplegia in aortic valve replacement. J Cardiac Surg 11, 348–354.[Medline]
Dinkelborg LM, Kinne RKH & Grishaber MK (1995). Characterisation and pH dependence of l-glutamate transport in sarcolemmal vesicles from rat hearts. Am J Physiol 268, H194–H201.[Medline]
Doggrell SA & Brown L (1998). Rat models of hypertension, cardiac hypertrophy and failure. Cardiovasc Res 39, 89–105.
Dorman BH, Hebbar L, Clair MJ, Hinton RB, Roy RC & Spinale FG (1997). Potassium channel opener augmented cardioplegia-protection of myocyte contractility with chronic left ventricular dysfunction. Circulation 96, 253–259.
Engelman RM, Rousou JA, Flack JE, Iyengar Y, Kimura Y & Das DK (1991). Reduction of infarct size by systemic amino acid supplementation during reperfusion. J Thorac Cardiovasc Surg 101, 855–859.[Abstract]
Hyde R, Taylor PM & Hundal HS (2003). Amino acid transporters: roles in amino acid sensing and signalling in animal cells. Biochem J 373, 1–18.[CrossRef][Medline]
Imura H, Caputo M, Parry A, Pawade A, Angelini GD & Suleiman M-S (2001). Age-dependent and hypoxia-related differences in myocardial protection during pediatric open heart surgery. Circulation 103, 1551–1556.
Javadov SA, Lim KHH, Kerr PM, Suleiman M-S, Angelini GD & Halestrap AP (2000). Protection of hearts from reperfusion injury by propofol is associated with inhibition of the mitochondrial permeability transition. Cardiovasc Res 45, 360–369.
Ji LL, Fu RG, Mitchell EW, Griffiths M, Waldrop TG & Swartz HM (1994). Cardiac-hypertrophy alters myocardial response to ischaemia and reperfusion in-vivo. Acta Physiol Scand 151, 279–290.[Medline]
Jin XY, Gibson DG & Pepper JR (1995). Early changes in regional and global left ventricular function after aortic valve replacement – comparison of crystalloid, cold blood and warm blood cardioplegia. Circulation 92, 155–162.
King N, Williams H, McGivan JD & Suleiman M-S (2001). Characteristics of l-aspartate transport and expression of EAAC-1 in sarcolemmal vesicles and isolated cells from rat heart. Cardiovasc Res 52, 84–94.
Morita K, Ihnken K, Buckberg GD, Matheis G, Sherman MP & Young HH (1995). Studies of hypoxemic/reoxygenation injury: without aortic clamping. J Thorac Cardiovasc Surg 110, 1228–1234.[CrossRef][Medline]
Nicholson B & McGivan JD (1996). Induction of high affinity glutamate transport activity by amino acid deprivation in renal epithelial cells does not involve an increase in the amount of transporter protein. J Biol Chem 21, 12159–12164.
Palacín M, Estévez R, Bertran J & Zorzano A (1998). Molecular biology of mammalian plasma membrane amino acid transporters. Physiol Rev 78, 969–1054.
Peuhkurinen KJ (1984). Regulation of the tricarboxylic acid cycle pool size in heart muscle. J Mol Cell Cardiol 16, 487–495.[CrossRef][Medline]
Pisarenko OI (1996). Mechanisms of myocardial protection by amino acids: facts and hypotheses. Clin Exptl Pharmacol 23, 627–633.[CrossRef]
Pisarenko OI, Studneva IM, Shulzhenko VS, Korchazhkina OV & Kapelko VI (1995). Substrate accessability to cytosolic aspartate aminotransferase improves posthypoxic recovery of isolated rat heart. Biochem Mol Med 55, 138–148.[CrossRef][Medline]
Rennie MJ, Ahmed A, Khogali SEO, Low SY, Hundal HS & Taylor PM (1996). Glutamine metabolism and transport in skeletal muscle and heart and their clinical relevance. J Nutr 126, S1142–S1149.
Rossi A & Lortet S (2001). Energy metabolism patterns in mammalian myocardium adapted to chronic physiopathological conditions. Cardiovasc Res 52, 84–94.
Safer B (1975). The metabolic significance of the malate-aspartate cycle in heart. Circ Res 37, 527–533.
Snoeckx LHEH, Van der Vusse GJ, Coumans WA, Willemsen PHM, Van der Nagel T & Reneman RS (1986). Myocardial function in normal and spontaneously hypertensive rats during reperfusion after a period of global ischaemia. Cardiovasc Res 20, 67–75.
Suchak SK, Baloyianni NV, Perkinton MS, Williams RJ, Meldrum BS & Rattray M (2003). The ‘glial’ glutamate transporter, EAAT2 (Glt-1) accounts for high affinity glutamate uptake into adult rodent nerve endings. J Neurochem 84, 522–532.[CrossRef][Medline]
Suleiman M-S, Ascione R, Angelini GD, Bryan AJ & Gomes WJ (1998). Metabolic differences between hearts with ischaemic disease and hearts with left ventricular hypertrophy. J Mol Cell Cardiol 30, 2519–2523.[CrossRef][Medline]
Suleiman M-S & Chapman RA (1993). Changes in the principal free intracellular amino-acids in the Langendorf perfused guinea-pig heart during arrest with calcium-free or high potassium media. Cardiovasc Res 27, 1810–1814.
Suleiman M-S, Dihmis M, Caputo M, Angelini GD & Bryan AJ (1997). Changes in myocardial concentration of glutamate and aspartate during coronary artery surgery. Am J Physiol 272, H1063–H1069.[Medline]
Suleiman M-S, Halestrap AP & Griffiths EJ (2001). Mitochondria: a target for myocardial protection. Pharmacol Therapeut 89, 29–46.[CrossRef][Medline]
Velasco I, Velasco-Velázquez MA, Salazar P & Lajud N (2003). Influence of serum-free medium on the expression of glutamate transporters and the susceptibility to glutamate toxicity in cultured neurons. J Neurosci Res 71, 811–818.[CrossRef][Medline]
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P < 0.05 versus control normal hearts (both tests ANOVA with Tukey–Kramer post test). Open bars control normal hearts; cross-hatched bars normal hearts + aspartate; filled bars control hypertrophied hearts and horizontal-lined bars hypertrophied hearts + aspartate. ‘–’ control; ‘+’ with aspartate. Each data point represents the mean of 10 hearts.