Low- and high-blood flow regions in the normal pig heart are equally vulnerable to ischaemia during partial coronary stenosis.

Myocardial perfusion is heterogeneous, even in the normal heart. It is unknown whether the resting normal blood flow level predicts the severity of mismatch between local blood flow and metabolism during acute ischaemia. In the present study local blood flow (measured with radioactively labelled microspheres) and metabolic indicators of ischaemia [tissue contents of lactate and inosine (INO), a breakdown product of adenosine triphosphate (ATP)] were determined in 84-102 simultaneously frozen samples (approximately 0.9 g) of normal (n = 7) and partially ischaemic (n = 4) porcine left ventricles. Ischaemia was induced for 20 min by partially occluding the left anterior descending artery to reduce perfusion pressure from 107 +/- 17 mm Hg to 39 +/- 10 mm Hg (mean +/- SD). Flow reduction in the ischaemic region was strongly variable, both within the subepicardium (range 6-66%, average 34%) and the subendocardium (range 33-84%, average 57%), indicating redistribution of blood flow inside transmural layers in addition to the well-known preferential decrease in subendocardial perfusion. The relative flow reduction during stenosis was not dependent on normal local perfusion level (Spearman rank correlation coefficient -0.002, P = 0.99). Samples with low or high myocardial blood flows before stenosis showed similar increases in lactate content and INO/ATP content ratio, as long as the percentage blood flow reduction was the same. It is concluded that regions with low and high resting flows in the normally perfused heart are equally susceptible to metabolism-perfusion mismatch resulting from coronary stenosis.

Influence of fluid resuscitation on renal function in bacteremic and endotoxemic rats.

PURPOSE: Fluid resuscitation, which is the most important primary therapy in sepsis, is not always able to prevent acute renal failure. In this study, we investigated in two different rat models of distributive shock whether fluid resuscitation would increase renal plasma flow (RPF) and subsequently glomerular filtration rate (GFR). MATERIALS AND METHODS: In pentobarbital anesthetized wistar rats Haemaccel (Behring Pharma, Hoechst, the Netherlands) infusion (1.2 mL/100 g/h for 3 hours) was started immediately during either bacteremia (bolus of living Escherichia coli bacteria, 10(9) or endotoxemia (1 hour infusion of E. coli endotoxin, 8 mg/kg), as well as in time-matched healthy controls. RESULTS: After 3 hours, this treatment had increased RPF (clearance of 131I-hippurate) above normal in control (+67%) and bacteremic rats (+75%), whereas in endotoxemic animals, the significantly decreased RPF was normalized. On the other hand, in bacteremic animals, the lowered GFR (clearance of creatinine; x44%) was normalized, whereas in endotoxemic animals GFR remained depressed (x30%). The lack of improvement in GFR during endotoxemia was also indicated by a profound fall in urine flow, which by contrast steadily increased in control and bacteremic rats owing to volume loading. In both shocked groups, the decreased renal oxygen delivery was normalized, but the higher renal oxygen consumption than expected on the basis of the work needed for sodium reabsorption was not influenced by Haemaccel treatment, despite the fact that it caused this work load to rise in bacteremic but not in endotoxemic rats. In both shock models, renal cortical adenosine triphosphate content did not differ from healthy controls and was not influenced by volume loading. CONCLUSIONS: In conclusion, our study suggests that a decrease in GFR caused by live bacteria in the circulation may benefit from fluid resuscitation, while during endotoxemia this therapy could not prevent acute renal failure.

Renal function and oxygen consumption during bacteraemia and endotoxaemia in rats.

BACKGROUND: The hypothesis that renal failure during septic shock may occur as a result of hypoxia-related cell dysfunction was investigated in two rat models of distributive shock. METHODS: Pentobarbitone-anaesthetized rats received either a bolus (1 ml) of living Escherichia coli bacteria (hospital-acquired strain, 1 x 10(9) CFU/ml; BA-group, n = 7), or a 1-h infusion of endotoxin (E. coli O127.B8: 8 mg/kg; ET-group, n = 7), or saline to serve as time matched controls (C-group, n = 7). RESULTS: Urine flow in the BA- and ET-group reached a nadir at 1 h, but thereafter increased and reached values higher than control at 3 h. At this time point, renal oxygen delivery had decreased, in the BA-group mainly due to a fall in arterial oxygen content and in the ET-group to a fall in renal plasma flow (clearance of 131I-hippurate). However, renal oxygen extraction had significantly increased, by 31% in the BA and by 59% in the ET group, while renal oxygen consumption remained the same. Net tubular sodium reabsorption had decreased by 55% in the BA and by 25% in the ET group, due to a fall in glomerular filtration rate (clearance of creatinine). Hence, an excess oxygen consumption was found which was caused neither by an increased renal glucose release nor by the presence of an increased number of leukocytes stuck in the glomeruli. Renal tubular cells showed normal morphology. An indication that proximal tubular function in the BA and ET group remained largely intact were normal ATP levels, absence of urinary glucose, and a normal fractional excretion of sodium. However, since urine flow had increased in shocked rats at 3 h, water appeared selectively lost. CONCLUSIONS: Our data indicate that in rat models of septic shock renal failure is not caused by cortical hypoxia or a shortage of cellular energy supply.

Determination of dimethylamine in biological samples by high-performance liquid chromatography.

A reversed-phase HPLC method for the quantification of dimethylamine in serum and urine is presented. Dimethylamine (DMA) is converted into a stable fluorescent product by precolumn derivatization with fluorenylmethylchloroformate. The DMA derivative is resolved from derivatives of other amines and amino acids by gradient elution with a total run-time of 15 min. The lower limit of determination in biological samples is 0.1 micromol/l. Recoveries from spiked serum samples were 99-107%. Within- and between-run precision were better than 6%. Concentrations of DMA in serum from normal human subjects (n=8) and from continuous ambulatory peritoneal dialysis patients (n=15) were 3.3+/-1.5 and 29.1+/-12.1 micromol/l, respectively.

Local mitochondrial enzyme activity correlates with myocardial blood flow at basal workloads.

To study whether heterogeneous myocardial blood flow relates to the local oxidative capacity of cardiac muscle, local blood flow at resting cardiac workloads and the activity of the mitochondrial enzyme succinate dehydrogenase (SDH) were determined in small regions of the left ventricle of seven anaesthetized, mechanically ventilated, open-chest pigs (25-35 kg). Following injection of radioactive microspheres (15 microns phi) into the left atrium, the heart was rapidly excised and cut into five transverse slices, which were simultaneously freeze-clamped between two aluminum blocks precooled at -80 degrees C. The left ventricle was then subdivided into 84 samples of about 0.9 g. Myocardial blood flow was 0.88 +/- 0.34 ml/min/g wet weight (ww), and SDH activity 1.46 +/- 0.33 mumol/min/g ww (mean +/- S.D., n = 7). Local data were normalized to their respective mean values in each pig, and then pooled. Local blood flow ranged from 0.32 to 1.63 of the mean, and blood flow heterogeneity characterized by the coefficient of variation (CV = S.D./mean) was 18.4%. Normalized local SDH activity ranged from 0.16 to 1.94, with a CV of 21.8%, significantly exceeding measurement error (CV = 4.5%). Local blood flows and SDH activities did not vary among transmural sublayers of the left ventricle, but variation within each sublayer was considerable. In six of the seven pigs, local blood flow correlated (P < 0.05) with SDH activity, with correlation coefficients (r) ranging from 0.26 to 0.54 (for pooled data: r = 0.27, P < 0.0001). When expressed per gram dry weight, heterogeneity of SDH activity increased (P < 0.05), and here also local blood flow correlated with SDH activity in all pigs (for pooled data: r = 0.45, P < 0.0001). Hence, heterogeneity of mitochondrial capacity within cardiac muscle partly explains the heterogeneity of myocardial blood flow, even though myocardial perfusion at rest was studied in relation with a maximal enzyme rate. The low correlation coefficient clearly indicates that at resting workloads other factors also play a role.

Simultaneous determination of creatine compounds and adenine nucleotides in myocardial tissue by high-performance liquid chromatography.

A rapid high-performance liquid chromatography method was developed for the determination of creatine phosphate, creatine, adenine nucleotides, and related compounds in myocardial tissue. Analysis was performed by reversed-phase chromatography on a C18 column containing 3-microns particles, employing gradient elution and uv detection at 210 nm. Separation was achieved in less than 5 min. Total analysis time, including equilibration of the column after return of the gradient to starting conditions, was 8 min. The high reproducibility and short analysis time make this method suitable for the routine analysis of large series of samples.

Rapid determination of glutamine in biological samples by high-performance liquid chromatography.

A high-performance liquid chromatographic method for the determination of glutamine in biological samples is presented. Glutamine was derivatized with ortho-phthalaldehyde reagent containing 3-mercaptopropionic acid and separated by reversed phase chromatography on a C18 column containing 3-microns particles. No interference from other amino acids was observed. The assay was linear over a range from 1 to 2,000 mumol/l. Analytical recovery of plasma samples spiked with glutamine was 98.6 +/- 3.8%. Within- and between-batch imprecision were 1.5% and 2.2%, respectively. The derivatization step was fully automated. Total analysis time, including derivatization and chromatography, amounted to 6 min. The method can be used for the determination of glutamine in plasma, urine, cerebrospinal fluid and tissue homogenates.