Changes of respiratory chain activity in mitochondrial and synaptosomal fractions isolated from the gerbil brain after graded ischaemia

In this study we have examined (1) the integrated function of the mitochondrial respiratory chain by polarographic measurements and (2) the activities of the respiratory chain complexes I, II-III, and IV as well as the ATP synthase (complex V) in free mitochondria and synaptosomes isolated from gerbil brain, after a 30-min period of graded cerebral ischaemia. These data have been correlated with cerebral blood flow (CBF) values as measured by the hydrogen clearance technique. Integrated functioning of the mitochondrial respiratory chain, using both NAD-linked and FAD-linked substrates, was initially affected at CBF values of approximately 35 ml 100 g-1 min-1, and declined further as the CBF was reduced. The individual mitochondrial respiratory chain complexes, however, showed differences in sensitivity to graded cerebral ischaemia. Complex I activities decreased sharply at blood flows below approximately 30 ml 100 g-1 min-1 (mitochondria and synaptosomes) and complex II-III activities decreased at blood flows below 20 ml 100 g-1 min-1 (mitochondria) and 35-30 ml 100 g-1 min-1 (synaptosomes). Activities declined further as CBF was reduced below these levels. Complex V activity was significantly affected only when the blood flow was reduced below 15-10 ml 100 g-1 min-1 (mitochondria and synaptosomes). In contrast, complex IV activity was unaffected by graded cerebral ischaemia, even at very low CBF levels.     

Tolerance to ischaemia and ischaemic preconditioning in neonatal rat heart

Although there is much information on ischaemic preconditioning in the adult myocardium, this phenomenon has not yet been investigated in neonatal hearts. To examine the early development of cardiac tolerance to ischaemia and the possible protective effects of preconditioning, rat hearts isolated on days 1, 4 and 7 of postnatal life were perfused (Langendorff) with Krebs-Henseleit solution at constant pressure, temperature (37 degrees C) and rate (200 beats/min). Developed force (DF) of contraction was measured by an isometric force transducer, and analysed using an on-line computer. Hearts were exposed to 40 or 60 min of global ischaemia followed by 30 min of reperfusion. Preconditioning was induced by three 3-min periods of global ischaemia, each separated by 5-min periods of reperfusion. Developmental changes in expression of protein kinase C (PKC) isoforms, and their activation following preconditioning, were estimated using Western blot analysis. Recovery of contractile function during reperfusion decreased from day 1 (48 +/- 2%) to day 4 (42 +/- 1%) and day 7 (33 +/- 2%). Preconditioning failed to improve ischaemic tolerance on day 1 (46 +/- 2%) and on day 4 (43 +/- 3%), but pronounced effect was observed on day 7 (40 +/- 2%). Prolonging the period of sustained ischaemia from 40 to 60 min on day 1 did not lead to a better recovery of contractile function in preconditioned hearts. PKC isoforms alpha, delta, epsilon and zeta were expressed in the ventricular myocardium during the first week of life, but there was no evidence of translocation following preconditioning on day 7. It may be assumed that the decreasing tolerance of the heart to ischaemia during early postnatal life is counteracted by the development of an endogenous protection.     

Depressed myofilament co-operativity associated with post-cardioplegic myocardial depression

The mechanism underlying myocardial depression after procedures involving cardioplegia are unknown. We tested the hypothesis that such depression was associated with altered myofilament interactions, using isolated hearts perfused with warm (37 degreesC), oxygenated (95% O2/5% CO2) Krebs-Ringer's bicarbonate (KRB) buffer. A latex balloon was inserted into the left ventricle (LV) to monitor LV function. All hearts underwent a 30-min equilibration period. One group of hearts (CPL+RPR) were arrested with St Thomas #2 cardioplegic solution (4 degreesC; 3 ml followed by 1 ml every 15 min) for 120 min, followed by reperfusion with warm, oxygenated KRB. A second group underwent cardioplegic arrest with no reperfusion (CPL). A third group underwent 60 min of warm, oxygenated perfusion with KRB beyond the equilibration period (60 MIN). The last group only underwent the equilibration period (EQUIL). LV function was assessed at the end of equilibration, and at 30 and 60 min of reperfusion (or 30 and 60 min additional perfusion in the 60 MIN group). All hearts were frozen at the end of the temporal protocol for each group, and stored at -70 degreesC for later measurement of Ca2+-stimulated Mg2+ ATPase activity after isolation of myofibrils. CPL+RPR hearts demonstrated significant depression of systolic pressure and elevation diastolic pressure at fixed volumes, compared to baseline and 60 MIN group values. There were no significant changes in the amount of constituent myofilament proteins, as assessed by densinometric analyses of Western blots. There were also no changes in the minimal or maximal ATPase activities, nor in the pCa50, indicating no effect of cardioplegic arrest on myofilament sensitivity to calcium. However, all hearts that underwent cardioplegic arrest were found to have significantly lower Hill coefficients (1.85+/-0.09 and 1.85+/-0.13 v 2.31+/-0.13 and 2.34+/-0. 14 in CPL+RPR and CPL v 60 MIN and EQUIL hearts, respectively), suggesting decreased co-operativity of the actomyosin interaction. Such a decrease in co-operativity would contribute to both the systolic and diastolic alterations associated with myocardial depression after cardioplegic arrest. These changes were associated with the cardioplegic event, and appeared to be independent of reperfusion.     

Sucralfate attenuates gastric mucosal lesions and increased vascular permeability induced by ischaemia and reperfusion in rats

Recent evidence suggests that oxygen-derived free radicals are involved in mediating gastric microvascular and parenchymal cell injuries induced by ischaemia and reperfusion. Therefore, the effect of the locally acting anti-ulcer drug, sucralfate, was studied on ischaemia and reperfusion (e.g. induced gastric lesions, intraluminal bleeding, changes in vascular permeability and non-protein sulfhydryl levels in the rat stomach). Allopurinol was used as a known standard antioxidant drug. Rats were subjected to 30 min of gastric ischaemia in the presence of 100 mmol/L hydrochloric acid and reperfusion periods of 15, 30 or 60 min duration. The gastric lesions were assessed microscopically under an inverted microscope. The vascular permeability was quantified by measuring the extravasated Evans blue in the stomach. There were significantly greater numbers of gastric lesions, intraluminal bleeding and leakage of Evans blue during all reperfusion periods as compared with those of ischaemia, with maximum effects occurring at 60 min following reperfusion. Pretreatment with sucralfate (31.25-250 mg/kg, p.o.) or allopurinol (12.5-50 mg/kg, i.p.) 30 min before the procedure, dose-dependently reduced the gastric lesions, intraluminal bleeding, and decreased the vascular permeability induced by ischaemia and reperfusion. Furthermore, sucralfate dose-dependently reverses the ischaemia and reperfusion-induced depletion of mucosal non-protein sulfhydryl levels and inhibited the superoxide radical production in both cell-free xanthine-xanthine oxidase and in the stimulated polymorphonuclear cellular systems. These results suggest that the protection produced by sucralfate against gastric injury may be due to its antioxidant effects.     

Optimal depth and duration of mild hypothermia in a focal model of transient cerebral ischemia: effects on neurologic outcome, infarct size, apoptosis, and inflammation

BACKGROUND and PURPOSE: Mild hypothermia is possibly the single most effective method of cerebroprotection developed to date. However, many questions regarding mild hypothermia remain to be addressed before its potential implementation in the treatment of human stroke. Here we report the results of 2 studies designed to determine the optimal depth and duration of mild hypothermia in focal stroke and its effects on infarct size, neurological outcome, programmed cell death, and inflammation. METHODS: Rats underwent a 2-hour occlusion of the left middle cerebral artery. In the first study (I) animals were kept (intraischemically) at either 37 degreesC (n=8), 33 degreesC (n=8), or 30 degreesC (n=8). Study II consisted of 4 groups: (1) controls (37 degreesC, n=10), (2) 30 minutes of hypothermia started at ischemic onset (33 degreesC, n=9), (3)1 hour (33 degreesC, n=8), and (4) 2 hours (33 degreesC, n=8). Brain temperature was measured by a thermocouple probe placed in the contralateral cortex. After suture removal, all animals were rewarmed and reperfused for 22 hours (I) or 70 hours (II). RESULTS: Mild hypothermia to 33 degreesC or 30 degreesC was neuroprotective (17+/-7% and 27+/-6%, respectively) relative to controls (53+/-8%, P<0.02), but 33 degreesC was better tolerated and recovery from anesthesia was faster. The neurological score of hypothermic animals was significantly better than that of controls (I & II) at both 24 and 72 hours postischemia except for the 30-minute group (II), which showed no improvement. In Study II, 2 hours of hypothermia reduced injury by 59%, 1 hour reduced injury by 84% whereas 30 minutes did not reduce injury. Normalized for infarct size, 2 hours of mild hypothermia decreased neutrophil accumulation by 57% whereas both 1 hour and 30 minutes had no effect. At 72 hours, 1 and 2 hours of mild hypothermia decreased transferase dUTP nick-end labeling (TUNEL) staining by 78% and 99%, respectively, and 30 minutes of hypothermia had no effect. CONCLUSIONS: Intraischemic mild hypothermia must be maintained for 1 to 2 hours to obtain optimal neuroprotection against ischemic cell death due to necrosis and apoptosis.     

The role of the house fly (Musca domestica) in the dissemination of hookworm

The effect of cooling rate, warming rate, and duration of phase transition upon survival of frozen canine kidneys was investigated. In the present study, 11 kidneys out of 14 rapidly cooled (2--4degreesC/min) to --22degreesC and thawed (70--110degreesC/min) were viable following contralateral nephrectomy. The serum creatinine and BUN levels rose to a maximum of 8.4 and 30 mg%, respectively, on the eighth day post-contralateral nephrectomy. Average survival time was 10 days; however, two of the dogs in this group were allowed to survive, one for 3 months and one for over 2 years. Eight kidneys out of 16 slowly cooled (0.25-1.0degreesC/min) and either rapidly or slowly warmed (20-30degreesC/min) had function to produce small amounts of urine; however, they did not survive more than 5 days after contralateral nephrectomy. Cooling rates of 0.1 and 10degreesC/min were too harmful to the kidney to have renal function after reimplantation. The minimum renal cell damage as assessed by LDH and GOT in the post-freeze perfusate was found in the 2-4degreesC/min cooling rate following rapid warming (70degrees-110degreesC/min). Correlation of the duration of phase transition time to renal cell damage was linear for LDH and GOT (r=0.93). This result suggests that the duration of phase transition time also is an important factor during the freezing process, affecting post-thaw survival of canine kidneys.     

Post-mortem changes in the rabbit retina. A study by light microscopy

The course of post-mortem changes in rabbit retina has been followed. Short post-mortem periods are accompanied by degenerative changes limited mainly to the visual cells and retinal pigment epithelium. Long post-mortem periods are associated with degenerative changes throughout the retina. Retinal tissue maintained at room temperature was less affected than that kept at body temperature (37degreesC). Post-mortem changes are similar to those observed following periods of pressure induced ischaemia and it is thought that the mechanical effects of pressure on retinal tissue are minimal at the level of resolution afforded by light microscopy.     

Conversion of temperature-sensitive to -resistant gene expression due to mutations in the promoter region of the melibiose operon in Escherichia coli

The melibiose utilization system of Escherichia coli W3133, a derivative of K12, is nonfunctional between 37 and 42 degreesC. The reason for this temperature sensitivity was thought to be that the melibiose transporter (MelB) of W3133 cells was temperature-sensitive. A mutant W3133-2 has been isolated as a temperature-resistant strain that can utilize melibiose between 37 and 42 degreesC. However, we found that the melibiose transporter of the W3133-2 was still temperature-sensitive. Half-life activities of the melibiose transporter at 37 degreesC (or 40 degreesC) in both E. coli W3133 and W3133-2 were exactly the same. Furthermore, we found that the nucleotide sequence of coding region of the melB structural gene (the second gene of the melibiose operon) of W3133-2 was exactly the same as that of W3133. Activity of alpha-galactosidase (product of the first gene, melA, of the melibiose operon) of W3133 cells grown at 40 degreesC was very low, although that of W3133-2 cells grown at 40 degreesC was high. These observations suggested that expression of the melibiose operon in W3133 is also temperature-sensitive. In fact, we found that the expression in W3133 cells was temperature-sensitive, while that in W3133-2 cells was temperature-resistant, by analyzing mRNA levels using the Northern blot method. Furthermore, we identified mutations in the promoter region of the melibiose operon of W3133-2 that resulted in the elongation of an 18 nucleotide inverted repeat sequence to a 28-nucleotide repeat sequence present immediately upstream of the -35 region. This may stabilize a possible stem structure due to the inverted repeat at 37-42 degreesC.     

Leukotrienes D4 and E4 produced in myocardium impair coronary flow and ventricular function after two hours of global ischaemia in rat heart

OBJECTIVE: Leukotrienes D4 and E4 are potent coronary vasoconstrictors and myocardial depressants. The aim was to investigate the contribution of myocardial leukotrienes to impairment of coronary flow and recovery of contractile function in rat hearts subjected to 2 h of global ischaemia. METHODS: Rat hearts were mounted on a working Langendorff apparatus and perfused with oxygenated Krebs-Henseleit solution at 37 degrees C for 30 min. Hearts were then arrested with either standard potassium crystalloid cardioplegic solution (n = 6), or with cardioplegic solution containing the leukotriene D4, E4 receptor antagonist Ly171883 (n = 6). Arrested hearts were maintained at 15 degrees C for 2 h, then rewarmed to 37 degrees C during 30 min working reperfusion. Coronary effluent was analysed by radioimmunoassay for leukotriene C4, D4, E4, and F4 levels. Immediately prior to cardiac arrest, and again after 30 min reperfusion, coronary flow, and aortic outflow and pressure were measured. RESULTS: Postischaemic leukotriene levels were increased compared to preischaemic levels in both groups [pooled measurements: 133.3 (SD 136.4) v 20.7(17.8) pg.0.1 ml-1, p < 0.05]. Postischaemic coronary vascular resistance was increased by 80% in controls (p < 0.001) compared to 19% (p = NS) in treated hearts. In addition, functional recovery was significantly greater in treated hearts compared to controls [82(3)% v 53(3)% for coronary flow; 79(3)% v 50(2)% for cardiac output; 82(4)% v 54(3)% for stroke work]. CONCLUSIONS: Leukotrienes are endogenously produced by the heart, and this production is significantly increased after global ischaemia and reperfusion. Reversal of significantly increased coronary vascular resistance coupled with improved functional recovery in hearts treated with LY171883 demonstrates an important contribution of endogenously produced leukotrienes to coronary vascular impairment and functional stunning of the globally ischaemic, reperfused heart.     

In vivo free radical production after cross-clamping and reperfusion of the renal artery in the rabbit

Postischaemic reperfusion injury is often attributed to the generation of oxygenated free radicals which may subsequently promote lipid peroxidation in cell membranes. Electron paramagnetic resonance spectroscopy in association with the spin trap molecule alpha-phenyl-N-tert-butyl-nitrone allowed direct confirmation of lipid free radical production after renal ischaemia-reperfusion in an in vivo rabbit model. A 60-min period of ischaemia followed by reperfusion caused free radical production twofold greater than after 15 min of ischaemia. Glutathione and alpha-tocopherol have been measured in renal tissue, as indirect markers of lipid peroxidation. After 15 min of ischaemia followed by 10 min of reperfusion, the mean(s.e.m.) glutathione content of the ischaemic kidney was slightly but significantly reduced by 11.9(2.5)% (P < 0.003). The content of alpha-tocopherol was unchanged. However, 10 min of reperfusion following 60 min of ischaemia led to significant decrease in mean(s.e.m.) content of both glutathione (30.4(3.7)%) (2.23(0.2) versus 3.14(0.18) mumol/g wet tissue, P < 0.001) and alpha-tocopherol (46.1(7.8)%) (0.57(0.10) versus 1.09(0.14) micrograms/g wet tissue, P < 0.001) when compared to the control kidney. Under these experimental conditions, desferrioxamine (15 mg/kg administered intravenously before inducing ischaemia), a drug known to limit free radical production, significantly limited the decrease of alpha-tocopherol to 20.8(6.4)% (0.83(0.08) versus 1.05(0.04) micrograms/g wet tissue, P < 0.05), but did not prevent glutathione consumption in the reperfused kidney.     

Homeobox genes in hematopoiesis and leukemogenesis

The effect of cold and warm intermittent antegrade blood cardioplegia, on the intracellular concentration of taurine in the ischaemic/reperfused heart of patients undergoing aortic valve surgery, was investigated. Intracellular taurine was measured in ventricular biopsies taken before institution of cardiopulmonary bypass, at the end of 30 min of ischaemic arrest and 20 min after reperfusion. There was no significant change in the intracellular concentration of taurine in ventricular biopsies taken after the period of myocardial ischaemia in the two groups of patients (from 10.1 +/- 1.0 to 9.6 +/- 0.9 mumol/g wet weight for cold and from 9.3 +/- 1.3 to 10.0 +/- 1.3 mumol/g wet weight for warm cardioplegia, respectively). Upon reperfusion however, there was a fall in taurine in both groups but was only significant (P < 0.05) in the group receiving cold blood cardioplegia (6.9 +/- 0.8 mumol/g wet weight after cold blood cardioplegia versus 8.0 +/- 0.8 mumol/g wet weight following warm blood cardioplegia). Like taurine, there were no significant changes in the intracellular concentration of ATP after ischaemia in the two groups of patients (from 3.2 +/- 0.32 to 2.95 +/- 0.43 mumol/g wet weight for cold and from 2.75 +/- 0.17 to 2.62 +/- 0.21 mumol/g wet weight for warm cardioplegia, respectively). However upon reperfusion there was a significant fall in ATP in both groups with the extent of the fall being less in the group receiving warm cardioplegia (1.79 +/- 0.19 mumol/g wet weight for cold and 1.98 +/- 0.27 mumol/g wet weight for warm cardioplegia, respectively). This work shows that reperfusion following ischaemic arrest with warm cardioplegia reduces the fall in tissue taurine seen after arrest with cold cardioplegia. Accumulation of intracellular sodium provoked by hypothermia and a fall in ATP, may be responsible for the fall in taurine by way of activating the sodium/taurine symport to efflux taurine.     

Myocardial protective effect of trilinolein: an antioxidant isolated from the medicinal plant Panax pseudoginseng

In a previous study we demonstrated that trilinolein, a natural plant triacylglycerol, is a novel myocardial protective agent in vivo. The mechanism probably involves an antioxidant effect. This work investigated the mechanism of myocardial protection of trilinolein to determine if inhibition of calcium influx and alteration of activity of superoxide dismutase are involved. In isolated cardiomyocytes, pretreatment with trilinolein at a low concentration of 10(-9) M effectively reduced 45Ca2+ influx stimulated by hypoxia/normoxia by 34%. In isolated perfused rat heart subjected to 60 min global hypoxemia without reperfusion, pretreatment with 10(-7) M trilinolein for 15 min reduced infarct size by 37%. Assay of superoxide dismutase-mRNA by Northern blot analysis in in vivo rat heart subjected to 30 min ischaemia and 10 min reperfusion showed pretreatment with 10(-7) M trilinolein had a synergistic action with antioxidant systems preventing the rise in superoxide dismutase-mRNA. These results reconfirm the myocardial protection of trilinolein and suggest it may be related to antioxidant activity and inhibition of 45Ca2+ influx.     

Glucagon-like peptide 1(7-36) amide''s central inhibition of feeding and peripheral inhibition of drinking are abolished by neonatal monosodium glutamate treatment

BACKGROUND AND AIMS: Injuries caused by ischaemia and ischaemia/reperfusion in the small intestine have been widely accepted as resulting in necrosis. The aim of this study was to ascertain whether apoptosis also occurs. METHODS: Intestinal epithelium from rats subjected to ischaemia (15-90 minutes) and ischaemia/reperfusion (15 minutes ischaemia followed by 15-75 minutes of reperfusion) was studied using histological, immunohistochemical, and molecular biological methods as well as FACS. RESULTS: Mucosal injury was induced by both ischaemia and ischaemia/reperfusion. Detachment of epithelial cells from the villous stroma was an early morphological change indicating mucosal injury. More than 80% of the detached cells exhibited characteristic morphological features of apoptosis (condensation of chromatin and nuclear fragmentation). The remainder demonstrated necrotic features. The apoptotic cells eventually underwent spontaneous degeneration with membrane rupture, a process morphologically identical to necrosis. DNA fragmentation was also confirmed by immunohistochemical methods and agarose gel electrophoresis. CONCLUSION: Apoptosis is a major mode of cell death in the destruction of rat small intestinal epithelial cells induced by ischaemia and ischaemia/reperfusion injury. Disruption of epithelial cell-matrix interactions ("anoikis") may play an important part in induction of apoptosis in detached enterocytes.     

New model of renal warm ischaemia-reperfusion injury for comparative functional, morphological and pathophysiological studies

BACKGROUND: Renal warm ischaemia-reperfusion injury is pertinent to vascular and transplant surgery. While established models provide functional and morphological data the authors wanted to be able to correlate this with the underlying pathophysiology at any chosen time point, thus allowing future interventional effects on reperfusion injury to be evaluated. METHODS: In a rodent model bilateral renal warm ischaemia (15-60 min) and then reperfusion (20 or 80 min) before nephrectomy allowed for analysis of early reperfusion pathophysiology. The remaining kidney provided functional data (glomerular filtration rate (GFR)) at days 2 and 7 before nephrectomy for late analysis and morphology using a new grading system. RESULTS: Acceptable survival rate (ten of 12 animals) was seen with up to 45 min of warm ischaemia. Renal function was impaired at day 2 following 30-60 min of warm ischaemia (P< 0.01) and day 7 in the 45- and 60-min groups (P < 0.05 and P < 0.01 respectively). Strong correlation existed between duration of ischaemia and GFR at day 2 (r2=0.88) and day 7 (r2=0.95). Histological damage in the cortical tubules was evident in the 45- and 60-min groups (P< 0.01). CONCLUSION: This new model allowed comparative functional, morphological and pathophysiological studies while minimizing the number of animals required. Overall 45 min of warm ischaemia gave significant, recoverable injury and is recommended for investigating renal reperfusion injury.     

RNA cleavage without hydrolysis. Splitting the catalytic activities of binase with Asn101 and Thr101 mutations

We have developed a model of ischaemia-reperfusion injury in C57BL/6 mice involving ischaemia for 0.5 to 2.5 h with an elastic tourniquet on one hind limb and reperfusion for 24 h, analogous to a well-established model of ischaemia-reperfusion injury in the rat. Viability was assessed in tissue homogenates of the gastrocnemius muscles from the affected and contralateral control limb by a triphenyl tetrazolium chloride dye reaction, measuring the activity of the oxidative mitochondrial enzymes. After 1.5 h ischaemia and 24 h reperfusion, viability in the ischaemic-reperfused limb was 13%, with the control muscle regarded as 100% viable. Significant improvements in viability to 86% (P < 0.05) and 56% (P < 0.05) were achieved, with administration 30 min prior to tourniquet release, of the nitric oxide (NO) synthase inhibitor nitro-L-arginine methyl ester (L-NAME, 30 mg/kg) and the anti-inflammatory glucocorticoid dexamethasone (2.5 mg/kg) respectively, with similar findings in the rat tourniquet model.     

Effects of ischaemia and reperfusion on vasoactive neuropeptide levels in the canine infrarenal aortic revascularization model

Infrarenal aortic cross-clamping is associated with remote vascular events, including myocardial infarction and renal insufficiency. The purpose of this study was to determine whether hindlimb ischaemia and reperfusion associated with infrarenal aortic cross-clamping results in the production of vasoactive regulatory neuropeptides. A canine model of infrarenal aortic cross-clamping was used for the study. Serial blood samples were drawn, prior to, at the time of, and serially following placement of the clamp and subsequent release of the clamp and reperfusion. Ischaemia resulted in increased mean(s.e.m.) plasma levels of neuropeptide Y (NPY) (initial 10.0(1.8) pmol/l versus ischaemia 24.7(2.3) pmol/l, P < 0.001) and vasoactive intestinal polypeptide (VIP) (initial 2.53(0.5) pmol/l versus ischaemia, 7.3(1.3) pmol/l, P < 0.05). Reperfusion produced three-fold elevation of VIP (initial 2.5(0.5) pmol/l versus reperfusion 9.6(1.5) pmol/l, P < 0.001), two-fold elevation in the plasma levels of endothelin-1 (initial 1.3(0.1) pmol/l versus reperfusion maximum 2.5(0.3) pmol/l, P < 0.01) and NPY (initial 10.0(0.8) pmol/l versus reperfusion maximum 23.9(2.3) pmol/l, P < 0.001). Ischaemia and reperfusion did not alter calcitonin gene-related peptide (CGRP) (a potent vasodilator) levels. Endothelin-1 (ET-1) plasma levels were also increased following haemorrhagic shock (initial 1.3(0.1) pmol/l versus exsanguination 3.4(0.4) pmol/l, P < 0.001), but not during ischaemia (initial 1.3(0.1) pmol/l versus ischaemia maximum 1.7(0.2) pmol/l, P = 0.7). It was concluded that vasoactive regulatory peptides are released following ischaemia, reperfusion and shock in the canine infrarenal aortic revascularization model and, therefore could contribute to remote vascular events observed with infrarenal aortic cross-clamping.     

Intrathecal neostigmine, but not sympathectomy, relieves mechanical allodynia in a rat model of neuropathic pain

OBJECTIVE: In myocardial ischaemia, slow conducting capsaicin-sensitive C-fibres are activated. Apart from the mediation of pain, activation of these fibres causes release of various peptides, such as calcitonin gene-related peptide (CGRP), which is a potent vasodilator. The aim of this study was to investigate the role of CGRP in the context of myocardial ischaemia in vivo. METHODS: The left anterior descending coronary artery (LAD) was occluded during 45 min in 27 anaesthetised open-chest pigs. LAD flow, mean arterial pressure (MAP), heart rate, peak dP/dt, arterial and coronary venous concentration of CGRP was measured prior to ischaemia, and during 4 h of reperfusion. The extent of myocardial infarction was measured using staining with triphenyl tetrazolium chloride. RESULTS: Retroinfusion of CGRP (100 micrograms) into the ischaemic myocardium was associated with a more pronounced hyperaemia, and systemic hypotension, during early reperfusion. The infarct size in relation to the area at risk was not affected by CGRP or the CGRP antagonist CGRP(8-37), and averaged 67 +/- 3%. There were no changes in plasma CGRP levels during ischaemia or reperfusion. CONCLUSION: Exogenously administered CGRP can cause systemic hypotension and augments postischaemic coronary flow. In this model, no cardioprotective effect of CGRP could be proven.     

Moderate exercise increases postexercise thresholds for vasoconstriction and shivering

The purpose of this study was to evaluate the effect of exercise on the subsequent postexercise thresholds for vasoconstriction and shivering. On two separate days, with six subjects (3 women), a whole body water-perfused suit slowly decreased mean skin temperature (approximately 7.0 degreesC/h) until thresholds for vasoconstriction and shivering were clearly established. Subjects were then rewarmed by increasing water temperature until both esophageal and mean skin temperatures returned to near-baseline values. Subjects either performed 15 min of cycle ergometry (65% maximal O2 consumption) followed by 30 min of recovery (Exercise) or remained seated with no exercise for 45 min (Control). Subjects were then cooled again. We mathematically compensated for changes in skin temperatures by using the established linear cutaneous contribution of skin to the control of vasoconstriction and shivering (20%). The calculated core temperature threshold (at a designated skin temperature of 30.0 degreesC) for vasoconstriction increased significantly from 36.64 +/- 0.20 to 36.89 +/- 0.22 degreesC postexercise (P < 0.01). Similarly, the shivering threshold increased from 35.73 +/- 0.13 to 36.13 +/- 0.12 degreesC postexercise (P < 0.01). In contrast, sequential measurements, without exercise, demonstrate a time-dependent decrease in both the vasoconstriction (0.10 degreesC) and shivering (0.12 degreesC) thresholds. These data indicate that exercise has a prolonged effect by increasing the postexercise thresholds for both cold thermoregulatory responses.     

Arginine vasopressin release inhibitor RU51599 attenuates brain oedema following transient forebrain ischaemia in rats

RU51599 is an arginine vasopressin (AVP) release inhibitor and a selective kappa opioid agonist which has a pure water diuresis effect without associated electrolyte excretion. The effect of RU51599 on brain oedema following transient forebrain ischaemia in rats was examined. Under microscopy, the visible vertebral arteries at the second vertebra could be easily electrocauterized and completely cut by microscissors to yield complete cessation of circulation of both vertebral arteries. Transient forebrain ischaemia was induced by this improved highly reproducible technique of four-vessel occlusion model. Forty-three male Wistar rats were separated into six groups; saline-treated (1 ml/kg) normal rats (n = 10), RU51599-treated (1 mg/kg) normal rats (n = 4), saline-treated (1 ml/kg) rats with complete occlusion of both vertebral arteries (n = 5), RU51599-treated (1 mg/kg) rats with complete occlusion of both vertebral arteries (n = 5), saline-treated (1 ml/kg) rats with both complete occlusion of both vertebral arteries and carotid occlusion bilaterally during 45 minutes followed by 60 minutes of reperfusion (n = 11), RU51599-treated (1 mg/kg) rats with both complete occlusion of both vertebral arteries and carotid occlusion bilaterally during 45 minutes followed by 60 minutes of reperfusion (n = 8). The brain water content was determined by the dry-wet weight method. Cerebral blood flow was monitored during ischaemia and reperfusion was performed by laser Doppler flowmetry to make sure to obtain reversible forebrain ischaemia. Effects of RU51599 on concentration of glutamate released from the hippocampal CA1 of rats subjected to 5 minutes four-vessel occlusion and 60 minutes of reperfusion were also investigated by the microdialysis method. This modified four-vessel occlusion method produced reversible forebrain ischaemia with a high level of success. Bilateral carotid occlusion followed by 60 minutes reperfusion caused a significant increase in brain water content (P < 0.01), which was significantly attenuated by RU51599 (P < 0.01). These findings indicate that the AVP-release inhibitor RU51599 reduced brain oedema following transient forebrain ischaemia in rats.     

Low-dose midazolam attenuates predatory odor avoidance in rats

Based on the premise that optimal drug delivery might improve the efficacy of locoregional treatment for solid tumors, the authors set up an experimental model for isolation perfusion in surgical specimens from patients resected for carcinoma of the colon. Ten surgical specimens were cannulated, washed internally and externally with saline solution, promptly cooled to 4 degreesC, connected to a circuit, and perfused with Krebs-Henselait modified solution, concentrated red blood cells, albumin, desamethasone, glucose, and heparin for 60 minutes at a target temperature of 37 degreesC. Organ temperature, flow rate, perfusion pressure, and metabolic and functional parameters were checked at 5, 20, and 60 minutes of perfusion. A paraphysiologic perfusion procedure was achieved. Mean values (and ranges) were as follows: temperature 37 degreesC (35. 1-39.6 degreesC); flow rate 10.2 (5.6-17.9) ml/min/100 g; arterial pressure 96 (42-154) mmHg; arterial pH 7.3 (7.1-7.5); arterial PO2 183 (78-304) mmHg; arterial PCO2 36 (31-46) mmHg. No important signs of tissue damage were found at histology. Autonomous or stimulated peristalsis (or both) was present throughout the experiment. Mean O2 extraction was 7.9 ml/min/100 g (range 3.1-11.0). Mean glucose consumption was 229 mg/100 g (range 174-252). The model worked well and appears promising, particularly for future use in various pharmacokinetic and pharmacodynamic studies of antiblastic agents.