Renal viability evaluated by the multiprobe assembly: a unique tool for the assessment of renal ischemic injury.

BACKGROUND: One of the major causes of transplanted organs' dysfunction is ischemia-reperfusion injury, where mitochondrial dysfunction is the primary contributor to cell damage. Mitochondrial NADH fluorescence reliably describes intracellular oxygen deficiency and mitochondrial function. Therefore, its monitoring at the tissue level, together with other physiological parameters, can serve to evaluate tissue vitality. METHODS: The multiprobe assembly (MPA) enabled the assessment of renal blood flow (RBF) using laser Doppler flowmetry, mitochondrial NADH redox state using the fluorometric technique, and ionic homeostasis using specific mini-electrodes (K(+) and H(+)). The MPA was utilized in two rat groups in which ischemia was induced for a period of 25-30 min (group 1) or for 60 min (group 2), and RBF and NADH were also monitored in a group of rats that underwent a complete kidney ischemia 24 h before the monitoring - a well-known model of acute renal failure. RESULTS: During ischemia, the RBF was completely abolished, NADH and extracellular potassium levels increased, and extracellular pH decreased. Immediately after the reperfusion, full recovery was observed; however, in the rats undergoing 60-min ischemia followed by 24-hour reperfusion, the tissue hemodynamic and mitochondrial functions were significantly impaired. CONCLUSION: This study demonstrates the advantage of using the MPA for real-time evaluation of kidney physiological state, which may serve as a practical instrument for the evaluation of graft viability during transplantation procedures.

Brain oxygen balance under various experimental pathophysiologycal conditions.

Normally, brain tissue copes with negative oxygen balance by increasing cerebral blood flow (CBF). We examined the effects of increasing oxygen demand, by inducing spreading depression (SD) under various oxygen balance states, on brain O2 balance. The Tissue Vitality Monitoring System was used, which enables real time simultaneous in vivo monitoring of CBF, mitochondrial NADH and tissue HbO2 from the same region of the cerebral cortex. SD was induced during normoxia, hypoxia, hyperoxia, ischemia, and in normal and ischemic brain after systemic epinephrine administration. Under normoxia, hyperoxia and ischemia & epinephrine, the compensation of energy demand induced by SD, was carried out by increasing CBF. The higher oxygen delivery under hyperoxia and epinephrine did not change the pattern of recovery from SD as compared to normoxia, whereas in the ischemic and hypoxic brain, the recovery from SD was prolonged, indicating a lake in oxygen delivery. Epinephrine infusion in the ischemic rat, decreased oxyhemoglobin utilization during SD, indicating that tissue oxygen balance improves even under higher oxygen demand induced by SD.

Effects of anesthesia on brain mitochondrial function, blood flow, ionic and electrical activity monitored in vivo.

Thiopental, a well-known barbiturate, is often used in patients who are at high risk of developing cerebral ischemia, especially during brain surgery. Although barbiturates are known to affect a variety of processes in the cerebral cortex, including oxygen consumption by the mitochondria, the interrelation between mitochondrial function and anesthetics has not been investigated in detail under in vivo conditions. The aim of this study was to examine the effects of thiopental on brain functions in normoxia and under partial or complete ischemia. The use of the multiparametric monitoring system permitted simultaneous measurements of microcirculatory blood flow, NADH fluorescence, tissue reflectance, and ionic and electrical activities of the cerebral cortex. Thiopental caused a significant, dose-dependent decrease in blood flow and a significant decrease in extracellular levels of potassium, with no significant changes in NADH levels in normoxic and ischemic rats. Following complete ischemia (death), the increase in the reflectance was significantly smaller in the anesthetized normoxic group versus the awake normoxic group. The time until the secondary increase in reflectance, seen in death, was significantly shorter in the anesthetized ischemic group. In conclusion, it seems that the protective effect of thiopental occurs only under partial ischemia and not under complete ischemia.

Effect of hyperbaric oxygenation on brain hemodynamics, hemoglobin oxygenation and mitochondrial NADH.

To determine the HbO(2) oxygenation level at the microcirculation, we used the hyperbaric chamber. The effects of hyperbaric oxygenation (HBO) were tested on vitality parameters in the brain at various pressures. Microcirculatory hemoglobin oxygen saturation (HbO(2)), cerebral blood flow (CBF) and mitochondrial NADH redox state were assessed in the brain of awake restrained rats using a fiber optic probe. The hypothesis was that HBO may lead to maximal level in microcirculatory HbO(2) due to the amount of the dissolved O(2) to provide the O(2) consumed by the brain, and therefore no O(2) will be dissociated from the HbO(2). Awake rats were exposed progressively to 15 min normobaric hyperoxia, 100% O(2) (NH) and to 90 min hyperbaric hyperoxia (HH) from 1.75 to 6.0 absolute atmospheres (ATA). NH and HH gradually decreased the blood volume measured by tissue reflectance and NADH but increased HbO(2) in relation to pO(2) in the chamber up to a nearly maximum effect at 2.5 ATA. Two possible approximations were found to describe the relationship between NADH and HbO(2): linear or logarithmic. These findings show that the increase in brain microcirculatory HbO(2) is due to an increase in O(2) supply by dissolved O(2), reaching a maximum at 2.5 ATA. NADH is oxidized (decreased signal) in parallel to the HbO(2) increase, showing maximal tissue oxygenation and cellular mitochondrial NADH oxidation at 2.5 ATA. In conclusion, in the normoxic brain, the level of microcirculatory HbO(2) is about 50% as compared to the maximal level recorded at 2.5 ATA and the minimal level measured during anoxia.

Effects of anesthesia on the responses to cortical spreading depression in the rat brain in vivo.

OBJECTIVES: The aim of this study was to evaluate the effect of cortical spreading depression (CSD) on the metabolic, hemodynamic, electrical and ionic properties during anesthesia as compared with the awake state. METHODS: The mitochondrial NADH redox state, reflected light, direct current (DC) potential, electrocorticography (ECoG), cerebral blood flow (CBF) and volume (CBV), and extracellular K(+) concentrations ([K(+)](e)), were measured continuously and simultaneously in real time using two unique monitoring systems that evaluate brain function. Three consecutive CSD waves were initiated using a KCl solution in both awake and anesthetized rats. RESULTS AND DISCUSSION: CSD caused typical amplitude changes: biphasic waves in reflectance, oxidation cycles in NADH, an increase in CBF, CBV and in [K(+)](e), a negative shift in DC potential and depression in ECoG. Anesthesia by equithesin decreased significantly the baseline levels of CBF and [K(+)](e), showing a reduction in oxygen supply and demand. After anesthesia, CSD significantly decreased [K(+)](e) and NADH oxidation cycles, indicating a reduction in oxygen demand and in oxygen balance, respectively. Furthermore, anesthesia reduced CSD wave frequencies by slowing the recovery period, showing a decline in energy production during brain activation, or by changing electrophysiological properties of the tissue. No changes were found in the propagation rate and in the initiation period of CSD, which may indicate that equithesin does not block CSD initiation. In addition, we found that the whole cerebral cortex reacts homogenously to CSD and that equithesin may reduce oxygen demand and energy production, which may have a protective effect on the brain exposed to pathophysiological conditions.

Can the "brain-sparing effect" be detected in a small-animal model?

BACKGROUND: Under O(2) imbalance in the body, blood redistribution occurs between more vital organs and less vital organs. This response is defined as the "brain-sparing effect". The study's aim was to develop a new rat model for simultaneous real-time monitoring of tissue viability in a highly vital organ, the brain, and a less vital organ, the small intestine, under various metabolic perturbations and emergency-like situations. MATERIAL/METHODS: The cerebral cortex and intestinal serosa were exposed in anesthetized rats and a multi-site multi-parametric (MSMP) monitoring system was connected to both. Tissue blood flow (TBF) was monitored using laser Doppler flowmetry and mitochondrial function by NADH fluorometry. The perturbations performed were anoxia (30 sec) and 20 minutes of hypoxia, hypercapnia, or hyperoxia. RESULTS: Under oxygen deficiency, cerebral blood flow (CBF) increased (315+/-53% in anoxia and 140+/-12% in hypoxia), whereas intestinal blood flow decreased (60+/-11% in anoxia and 56+/-13% in hypoxia). Mitochondrial NADH significantly increased in both organs (119+/-2.8% and 151+/-14% in the brain and intestine, respectively). Under hyperoxia, NADH was oxidized in both organs (up to 9% change). Hypercapnia led to an increase in CBF (143+/-11%) and oxidation of mitochondrial NADH (by 10%), with no significant changes in the intestine. CONCLUSIONS: The two organs respond significantly differently to lack of O(2) by activating the sympathetic nervous system. Monitoring less vital organs may indicate an early response to emergency situations. Therefore, a less vital organ could be used as a surrogate organ to be monitored in order to spare the brain.

Effect of hypothermia on brain multi-parametric activities in normoxic and partially ischemic rats.

Hypothermia, as well as anesthesia, are known to protect the brain against ischemia, hypoxia and other pathological damages. One of the mechanisms of this improvement could be by lowering brain function, and thereby lowering oxygen demand. We examined the effect of hypothermia on brain function and blood supply in awake and anesthetized rats and studied the interaction between partial ischemia and the responses to hypothermia. The brain function multiprobe (BFM) used enabled simultaneous measurements of cerebral blood flow (CBF), mitochondrial NADH redox state, extracellular K(+) concentration, DC potential and ECoG from the cerebral cortex in rats whose brain temperature was lowered by 5 degrees C. Hypothermia was induced in awake, anesthetized and brain ischemic-anesthetized rats. In anesthetized and ischemic-anesthetized rats, the time required for lowering the brain temperature by 5 degrees C was five times less than in the normal awake animals. No significant changes in CBF and NADH levels were found in response to hypothermia in the awake animals. In contrast, a significant decrease in extracellular K(+) concentration was recorded under hypothermia, probably due to the lower rate of depolarization. Hypothermia in anesthetized and in ischemic-anesthetized rats did not significantly affect the levels of mitochondrial NADH, CBF and extracellular K(+). Hypothermia under ischemia was expected to be more effective.