Hydroxylation of salicylate by the in vitro diaphragm: evidence for hydroxyl radical production during fatigue

There is increasing evidence that oxygen-derived free radicals produced during strenuous work by the diaphragm may contribute to diaphragm fatigue and/or injury. However, the precise identity of these oxygen radicals remains unknown, inasmuch as oxygen free radicals are extremely short lived and their detection in biologic systems is quite difficult. There is recent evidence that the salicylate-trapping method may be a useful means of monitoring tissue production of hydroxyl radical (.OH). This method is predicated on the fact that salicylate's phenolic ring can be attacked by .OH at the 3 or 5 position to yield 2,3- or 2,5-dihydroxybenzoic acid (DHB). These metabolites are stable and can be identified by high-performance liquid chromatography (HPLC) coupled with electrochemical or ultraviolet detection. To test the hypothesis that hydroxylated salicylates are produced during diaphragm fatigue, we exposed in vitro rat diaphragm strips to a physiological saline solution containing 2.0 mM sodium salicylate for approximately 15 min. The solution was then removed, and the strips were fatigued (20 Hz, 200-ms train duration, 1 train/s) via phrenic nerve stimulation for 30 s-10 min. The diaphragm strips were subsequently homogenized, and the homogenate was analyzed by HPLC coupled with ultraviolet detection. Levels of 2,3-DHB were significantly higher in fatigued than in control nonfatigued strips. There was also a significant correlation between the amount of 2,3-DHB in the fatigued muscle and the accumulated tension-time product developed during fatigue. 2,5-DHB was not consistently identified in control or experimental strips.(ABSTRACT TRUNCATED AT 250 WORDS)     

Preconceptional education

Although studies have examined the susceptibility and pattern of injury induced by infusion of free radical-generating solutions into a number of vital organs, no such investigation has been performed for the diaphragm. The purpose of the present study was to examine the susceptibility of the diaphragm to damage by a free radical-generating solution (iron-ADP complexes). Studies were performed using an in situ canine diaphragmatic strip preparation in which the phrenic artery supplying the strip was cannulated and perfused with blood from the ipsilateral femoral artery. Four groups of studies were performed: (1) a group in which saline was infused into the arterial supply of the diaphragm for 15 min; (2) a group in which a solution of iron-ADP was infused; (3) a group in which both iron-ADP and superoxide dismutase (SOD), a free radical scavenger, were infused; and (4) a group given iron-ADP and denatured SOD. Strip tension and blood flow were monitored during electrically induced diaphragmatic contractions for 15 min before intraphrenic infusions, during the period of infusions, and for 90 min after cessation of infusions. We found that diaphragm tension did not change over time in saline-treated control animals but fell significantly in animals in which iron-ADP was infused. The effects of iron-ADP were largely prevented by concomitant administration of active SOD, but not by denatured SOD. On average, at 90 min after cessation of infusions, tension had fallen to 82 +/- 6, 41 +/- 8, 63 +/- 4, and 28 +/- 9% of its initial value in saline, iron-ADP, iron-ADP/SOD, and iron-ADP/denatured SOD groups, respectively (p < 0.001 for comparison of the four groups, with saline and iron-ADP/SOD groups different from the other two groups). Diaphragm blood flow did not change significantly in any group. These data suggest that free radical-mediated diaphragmatic injury can result in a marked reduction in diaphragm contractility.     

Influence of hypoxia and hypoxia/hypercapnia upon brain and blood peroxidative and glutathione status in normal weight and growth-restricted newborn piglets

Glutathione (reduced (GSH) and oxidized (GSSG)), lipid peroxidation products (TBAR) and in vitro production of reactive oxygen species (ROS, by means of stimulated lipid peroxidation, H2O2 formation and amplified chemiluminescence (CL) in 9000 xg brain supernatants) were studied in the cerebellum (C) and temporoparietal area (TP) of the brain of normal weight (NW) and spontaneously intra-uterine growth-restricted newborn piglets (IUGR) after 1 hour hypoxia (fractional inspired oxygen concentration (FiO2) 8%), and in combination with 10% CO2, followed by 3 hours recovery (FiO2 30%). The strong GSH depletion accompanied by an increased concentration of GSSG and TBAR, more distinct in IUGR, is the most important result in the brain after hypoxia and reoxygenation. Hypercapnia-related acidosis seems to protect the brain of IUGR from hypoxia/reoxygenation induced injury by reducing GSH depletion as well as GSSG and TBAR increases. But stimulated lipid peroxidation and H2O2 formation in 9000 xg supernatants of C and TP were found to be higher in acidosis and hypercapnia. Decreased or unchanged amplified CL, demonstrating lower in vitro production of ROS, cannot explain the GSH depletion after hypoxia and reoxygenation. The scarce changes in erythrocyte GSH and GSSG as well as plasma TBAR concentrations did not reflect the findings in the brain. Nevertheless, the changes in the brain support the hypothesis that oxidative stress plays a role in neuronal damage after hypoxic stress, but the brain of IUGR did not reveal a special response to moderate hypoxia.     

The determination of T-lymphocyte antigenic markers in the serum of patients with atopic dermatitis

PURPOSE: We tested the role of lipid peroxidation in the demyelination and white matter necrosis associated with radiation injury of the central nervous system. METHODS AND MATERIALS: We irradiated the cervical spinal cords of female F344 rats (23 Gy) and assayed for the accumulation of the peroxidation byproducts malondialdehyde and hydroxyeicosatetraenoic acids, and for the consumption of the endogenous free radical scavengers vitamins E and C. We further tested the role of lipid peroxidation in radiation injury of the central nervous system by determining the sensitivity of the cervical spinal cord to radiation in rats on diets containing deficient, normal, and supplemental levels of the antioxidant vitamin E. Rats were placed on these diets at 4 weeks of age and irradiated (18.5-21.5 Gy) 16 weeks later. RESULTS: During the 5 months between irradiation and the onset of paralysis, no accumulation of peroxidation byproducts or consumption of endogenous scavengers was seen in the cervical spinal cords of the irradiated rats. The cervical spinal cords of some of the rats placed on the diets with deficient, normal, and supplemental levels of vitamin E were analyzed at the time of irradiation and contained 197 +/- 57, 501 +/- 19, and 717 +/- 35 pmol vitamin E/mg protein, respectively. Despite the statistical differences in these levels, the radiation sensitivity of the cervical spinal cord (ED50 for white matter necrosis) in rats receiving the three diets was not different (20.4, 20.7, and 20.6 Gy). CONCLUSION: These data do not support a role for free radical-induced lipid peroxidation in the white matter damage seen in radiation injury of the central nervous system.     

Human respiratory muscles: sensations, reflexes and fatiguability

1. Given the importance of the ventilatory 'pump' muscles, it would not be surprising if they were endowed with both sensory and motor specializations. The present review focuses on some unexpected properties of the respiratory muscle system in human subjects. 2. Although changes in blood gas tension were long held not to influence sensation directly, studies in subjects who are completely paralysed show that increases in arterial CO2 levels elicit strong sensations of respiratory discomfort. 3. Stretch reflexes in human limb muscles contain a monosynaptic spinal excitation and a long-latency excitation. However, inspiratory muscles show an initial inhibition when tested with brief airway occlusions during inspiration. This inhibition does not depend critically on input from pulmonary or upper airway receptors. 4. Human inspiratory muscles (including the diaphragm) have been considered to fatigue during inspiratory resistive loading. However, recent studies using phrenic nerve stimulation to test the force produced by the diaphragm show that carbon dioxide retention (hypoventilation) and voluntary cessation of loading occur before the muscles become overtly fatigued.     

Post-exercise depression of motor evoked potentials as a function of exercise duration

Post-exercise facilitation and post-exercise depression are phenomena described in motor evoked potentials (MEPs) elicited to transcranial magnetic stimulation. Brief, non-fatiguing muscle activation produces post-exercise facilitation, and prolonged fatiguing muscle activation produces post-exercise depression. We studied 12 normal subjects to determine whether post-exercise depression occurs before fatigue is reached. We recorded MEPs from the resting extensor carpi radialis muscle after increasing the duration of isometric wrist extension, at 50% of maximum voluntary contraction, until the muscle fatigued. Fatigue was defined as the inability to maintain that force. The mean exercise duration before the muscle fatigued was 130 s, and post-exercise depression occurred only beyond 90 s of exercise. We conclude that post-exercise depression is detectable only after prolonged muscle activation.     

Renal disease. II. Urinary tract infection in women

The paper is devoted to the investigation of the lipid peroxidation level according to measurements of the accumulation level of malon dialdehyde in fragments of the liver tissue of newborn and adult pigs, depending on the cooling rate and cryoprotectants used. The conclusion has been made on the basis of the data obtained. That the lipid peroxidation level in the liver tissue fragments is decreased with the increase of cooling rates. That is activated the least of all with the use of average cooling rate (100 degrees C/min) with dimethylsulphoxide (DMSO) as cryoprotectant in concentrations of 10, 20% and rapid cooling rate (80,000 degrees C/min) with polyethylenglycol-1500 (PEG-1500) and DMSO in analogous concentrations. These cryopreservation data have shown the lowest of the lipid peroxidation level in the liver tissue fragments when comparing one with native tissue (before freezing).     

Glutathone and glutathione ethyl ester supplementation of mice alter glutathione homeostasis during exercise

The present study examined the effect of glutathione (GSH) and glutathione ethyl ester (GSH-E) supplementation on GSH homeostasis and exercise-induced oxidative stress. Male Swiss-Webster mice were randomly divided into 4 groups: starved for 24 h and injected with GSH or GSH-E (6 mmol/kg body wt, i.p.) 1 h before exercise, starved for 24 h and injected with saline (S); and having free access to food and injected with saline (C). Half of each group of mice was killed either after an acute bout of exhaustive swimming (E) or after rest (R). Plasma GSH concentration was 100-160% (P < 0.05) higher in GSH mice vs. C or S mice at rest, whereas GSH-E injection had no effect. Plasma GSH was not affected by exercise in C or S mice, but was 44 and 34% lower (P < 0.05) in E vs. R mice with GSH or GSH-E injection, respectively. S, GSH- and GSH-E-treated mice had significantly lower liver GSH concentration and the GSH:glutathione disulfide (GSSG) ratio than C mice. Hepatic and renal GSH and the GSH:GSSG ratio were significantly lower in E vs. R mice in all groups. GSH-E-treated mice had a significantly smaller exercise-induced decrease in GSH vs. C, S, and GSH-treated mice and no difference in the GSH:GSSG ratio in the kidney. Activities of gamma-glutamylcysteine synthetase and gamma-glutamyltranspeptidase in the liver and kidney were not affected by either GSH treatment or exercise. GSH concentration and the GSH:GSSG ratio in quadriceps muscle were not different among C, S and GSH-treated mice, but significantly lower in GSH-E-treated mice (P < 0.05). Hepatic malondialdehyde (MDA) content was greater in exercised mice in all but GSH-E-treated groups. GSH and GSH-E increased MDA levels in the kidney of E vs. R mice, but attenuated exercise-induced lipid peroxidation in muscle. Swim endurance time was approximately 2 h longer in GSH (351 +/- 22 min) and GSH-E (348 +/- 27) than S mice (237 +/- 17). We conclude that 1) acute GSH and GSH-E supplementation at the given doses does not increase tissue GSH content or redox status; 2) both GSH and GSH-E improve endurance performance and prevent muscle lipid peroxidation during prolonged exercise; and 3) while both compounds may impose a metabolic and oxidative stress to the kidney, this side effect is smaller with GSH-E supplementation.     

Cellular adaptations in the diaphragm in chronic obstructive pulmonary disease

BACKGROUND: In patients with severe chronic obstructive pulmonary disease, the diaphragm undergoes physiologic adaptations characterized by an increase in energy expenditure and relative resistance to fatigue. We hypothesized that these physiologic characteristics would be associated with structural adaptations consisting of an increased proportion of less-fatigable slow-twitch muscle fibers and slow isoforms of myofibrillar proteins. METHODS: We obtained biopsy specimens of the diaphragm from 6 patients with severe chronic obstructive pulmonary disease (mean [+/-SE] forced expiratory volume in one second, 33+/-4 percent of the predicted value; residual volume, 259+/-25 percent of the predicted value) and 10 control subjects. The proportions of the various isoforms of myosin heavy chains, myosin light chains, troponin, and tropomyosin were determined by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis. We also used immunocytochemical techniques to determine the proportions of the various types of muscle fibers. RESULTS: The diaphragm-biopsy specimens from the patients had higher percentages of slow myosin heavy chain I (64+/-3 vs. 45+/-2 percent, P<0.001), and lower percentages of fast myosin heavy chains IIa (29+/-3 vs. 39+/-2 percent, P=0.01) and IIb (8+/-1 vs. 17+/-1 percent, P<0.001) than the diaphragms of the controls. Similar differences were noted when immunohistochemical techniques were used to compare the percentages of these fiber types in the two groups. In addition, the patients had higher percentages of the slow isoforms of myosin light chains, troponins, and tropomyosin, whereas the controls had higher percentages of the fast isoforms of these proteins. CONCLUSIONS: Severe chronic obstructive pulmonary disease increases the slow-twitch characteristics of the muscle fibers in the diaphragm, an adaptation that increases resistance to fatigue.     

Synergistic behavior of inspiratory muscles after diaphragmatic fatigue in the newborn

We studied diaphragmatic and intercostal muscle activity and the pattern of motion of rib cage and abdomen after diaphragmatic muscle fatigue in 15 newborn infants (birth wt 1,251 +/- 424 g, mean +/- SD). Rib cage and abdominal motion were monitored with magnetometers and intercostal and diaphragmatic electromyograms (EMG's) with surface electrodes. Twelve infants showed a total of 66 episodes of muscle fatigue identified by EMG frequency spectrum analysis. Two patterns of responses to fatigue were observed. In the first case, five infants consistently recruited their intercostal muscles; this was followed by a normalization of the diaphragmatic frequency spectrum. In these infants, recruitment of intercostal muscles successfully prevented any clinical deterioration. In the second, seven infants showed no change in their intercostal muscle activity, and diaphragmatic fatigue was followed by apnea. We conclude that in newborn infants the synergistic behavior of the diaphragm and intercostal muscles can maximize the performance of these muscles and, in some infants, seems to prevent development of apnea.     

Increased compliance in diaphragm muscle of the cardiomyopathic Syrian hamster

We investigated the hypothesis that diaphragm compliance was abnormal in cardiomyopathic Syrian hamsters (CSH), an experimental model of myopathy. The passive elastic properties of isolated diaphragm muscles were analyzed at both the muscle and sarcomere levels. We used the following passive exponential relationship between stress (sigma) and strain (epsilon): sigma = (Eo/beta) (ebetaepsilon - 1), where Eo is the initial elastic modulus and beta is the stiffness constant. Immunocytochemistry procedures were used to analyze the distribution of two key elastic components of muscle, extracellular collagen and intracellular titin elastic components, as well as the extracellular matrix glycoprotein laminin. Muscle and sarcomere values of beta were nearly twofold lower in CSH (8.7 +/- 1.9 and 8.3 +/- 1.4, respectively) than in control animals (19.7 +/- 1.7 and 16.8 +/- 2.1, respectively) (P < 0.01 for each). Compared with controls, Eo was higher in CSH. Sarcomere slack length was significantly longer in CSH than in control animals (2.1 +/- 0.1 vs. 1.9 +/- 0.1 micrometer, P < 0.05). The surface area of collagen I was significantly larger in CSH (17.4 +/- 1.8%) than in control animals (12.4 +/- 0.7%, P < 0.05). There was no change in the distribution of titin or laminin labelings between the groups. These results demonstrate increased diaphragm compliance in cardiomyopathic hamsters. The increase in CSH diaphragm compliance was observed despite an increase in the surface area of collagen and was not associated with an abnormal distribution of titin or laminin.     

Effect of N-acetylcysteine on human diaphragm strength and fatigability

Free radical injury is believed to be important in diaphragm dysfunction. N-Acetylcysteine (NAC) is a potent free radical scavenger shown in animal models to attenuate diaphragm fatigue; however, its effects on human diaphragm function are unknown. We assessed diaphragm function by electrophrenic twitch stimulation (PdiT) and twitch occlusion (to yield Pdimax) in four healthy subjects 35 +/- 3 yr of age (mean +/- SD). We intravenously administered NAC (150 mg/kg in 250 ml D5W) or placebo (CON) (250 ml D5W) in a randomized manner after subjects were premedicated with antihistamines. There were no significant side effects with the infusion. After infusion, we measured baseline Pdimax and PdiT at FRC. Diaphragm fatigue was then induced by subjects breathing through an inspiratory resistive load. Pdimax and PdiT were then measured at 15 to 30 min and 1, 2, 3, 4, and 20-25 h after fatigue. Times to fatigue were 13 +/- 4 min (CON) and 21 +/- 6 min (NAC) (p = 0.04). At 15 min after fatigue, PdiT was reduced to 40% (CON) compared with 30% (NAC) initial PdiT value (p = 0.05). Other twitch characteristics (maximal rate of relaxation and maximal contraction rate) were reduced to a greater degree after placebo compared with NAC. There were no significant differences in the rate of recovery between CON and NAC. Pdimax at 30 min after fatigue was significantly greater with NAC; however, at 1 h after fatigue, Pdimax for CON and NAC were not different, suggesting similar rates of recovery in high-frequency fatigue. These data suggest that NAC may attenuate low-frequency human diaphragm fatigue.     

Effects of fenoterol on inspiratory effort sensation and fatigue during inspiratory threshold loading

We studied the effects of a single dose of fenoterol on the relationship between inspiratory effort sensation (IES) and inspiratory muscle fatigue induced by inspiratory threshold loading in healthy subjects. The magnitude of the threshold was 60% of maximal static inspiratory mouth pressure (PI,mmax) at functional residual capacity, and the duty cycle was 0.5. Subjects continued the threshold loaded breathing until the target mouth pressure could no longer be maintained (endurance time). The intensity of the IES was scored with a modified Borg scale. Either fenoterol (5 mg) or a placebo was given orally 2 h before loading in a randomized double-blind crossover protocol. The endurance time with fenoterol (34.4 +/- 8.6 min) was longer than that with the placebo (22.2 +/- 7.1 min; P < 0.05). The ratio of high- to low-frequency power of the diaphragmatic electromyogram (EMGdi) decreased during loading; the decrease was less with fenoterol (P < 0.05). The EMGdi also decreased with loading; the decrease was greater on fenoterol treatment (P < 0.01). The PI,mmax and maximal transdiaphragmatic pressure (Pdi) were similarly decreased after loading on either treatment. The intensity of the IES rose with time during loading in both groups but was lower with fenoterol than with the placebo (P < 0.05). The ratio of Pdi to integrated activity of the EMGdi increased with fenoterol (P < 0.05). Fenoterol treatment increased both superimposed Pdi twitch and Pdi twitch of relaxed diaphragm and decreased the value of (1-superimposed Pdi twitch/Pdi twitch of relaxed diaphragm). Thus we conclude that in normal subjects fenoterol reduces diaphragmatic fatigue and decreases the motor command to the diaphragm, resulting in a decrease in IES during inspiratory threshold loading and a prolongation of endurance.     

Intestinal permeability and inflammation in patients on NSAIDs

PURPOSE: The purpose of this study was to determine whether high frequency fatigue was present in the diaphragm after intense whole body endurance exercise. METHODS: We used bilateral phrenic nerve stimulation (BPNS) before and during recovery from whole body exercise to detect fatigue in the diaphragm. To detect high frequency fatigue we used paired stimuli at 10, 20, 50, 70, and 100 Hz frequency and determined the transdiaphragmatic pressure (Pdi) response to the second stimulation (T2). RESULTS: The subjects (N = 10) exercised at 93.3 +/- 2.3% of their VO2max for 9.9 +/- 0.5 min. The Pdi response to "twitch" and 10 Hz "tetanic" stimulation was decreased immediately after exercise versus pre-exercise values (-23.4 +/- 3.3%). The T2 amplitude was substantially reduced at all frequencies immediately after exercise (-28.0%), but by 30 min into recovery the T2 amplitude at 70 and 100 Hz was not different from pre-exercise values. In contrast, at 10 and 20 Hz the T2 response was still significantly reduced. CONCLUSIONS: We interpret these data to mean that high frequency fatigue as well as low frequency fatigue were present in the diaphragm after intense whole body endurance exercise.     

Time-level relationship for lipid peroxidation and the protective effect of alpha-tocopherol in experimental mild and severe brain injury

OBJECTIVE: Oxygen free radical-mediated lipid peroxidation has been proposed to be one of the major mechanisms of secondary damage in traumatic brain injury. The first purpose of this study was to establish the time-level relationship for lipid peroxidation in injured brain tissue. The second purpose was to examine the protective effect of alpha-tocopherol against lipid peroxidation. METHODS: For this study, 65 guinea pigs in five groups were studied. Five of the animals were identified as a control group, and the remaining 60 animals were divided equally into four groups (Groups A, B, C, and D). Mild injury (200 g x cm) (Groups A and C) and severe injury (1000 g x cm) (Groups B and D) were produced by the method of Feeney et al. Alpha-tocopherol (100 mg/kg) was administered intraperitoneally before brain injury in Groups C and D. Five animals from each group were killed immediately after trauma, five after 1 hour, and the remaining five animals after 36 hours. Lipid peroxidation in traumatized brain tissues was assessed using the thiobarbituric acid method. RESULTS: In all groups with traumatic brain injuries, levels of malondialdehyde, a lipid peroxidation product, were higher than in the control group. The amount of lipid peroxidation was increased by the severity of the trauma. Alpha-tocopherol significantly suppressed the rise in lipid peroxide levels in traumatized brain tissues. CONCLUSION: This study demonstrates that lipid peroxidation is increased by the severity of trauma and that alpha-tocopherol has a protective effect against oxygen free radical-mediated lipid peroxidation in mild and severe brain injury.     

A convenient and inexpensive chemo-radiosensitivity assay for lung cancer cells using Terasaki''s microplate

Pharyngeal dilator muscles are critical for maintaining upper airway patency in the neonatal period. The present study examined in vitro the contractile properties of a pharyngeal dilator muscle, the sternohyoid, in 1-7-day-old piglets (n = 24). Isometric contraction and half-relaxation times were 36.7 +/- 1.1 and 30.9 +/- 1.2 msec, respectively. Twitch potentiation ('staircase phenomenon') and post-tetanic potentiation were noted following repetitive stimulation. During prolonged repetitive stimulation with a standard (40 Hz) fatigue test, muscle force declined gradually over time, with loss of half of the initial force occurring over 138 +/- 11 sec, and a 2-min fatigue index (ratio of force at 2 min to initial force) of 0.52 +/- 0.03. An additional 10 piglets were studied at ages of 14-20 days. Muscle from older piglets had comparable isometric twitch kinetics as that of younger animals. However, sternohyoid muscle from the older piglets had worse endurance than muscle from the younger animals, as indicated by a shorter time required for force to decrease by half (86 +/- 10 sec, P < 0.01) and a lower 2-min fatigue index (0.36 +/- 0.03, P < 0.01). These data indicate that for the sternohyoid muscle of the newborn piglet (a) physiological properties are consistent with moderate to fast contraction with good endurance, (b) force potentiates during repetitive twitch stimulation and following a brief period of tetanic stimulation, and (c) there is worsening of endurance but no change in isometric twitch kinetics with increasing age during the first weeks of life.     

Effect of dimethyl sulfoxide pretreatment on activities of lipid peroxide formation, superoxide dismutase and glutathione peroxidase in the mouse liver after whole-body irradiation

We investigated the effects of dimethyl sulfoxide (DMSO) on radiation damage in the mouse. DMSO (i.p. 0.11 g/mouse) administered 30 min before exposure protected the mice from the gamma-whole body irradiation: the 30 days lethality was significantly decreased from 44% to 16% (P < 0.05). The contents of thiobarbituric acid reactive substances(TBA-RS) in the mouse liver increased linearly between days 2 and 10 after 9 Gy gamma ray irradiation. The TBA-RS contents in the liver on days 2 to 10 after irradiation were reduced by DMSO pretreatment. The irradiation decreased superoxide dismutase (SOD) activity in the liver on day 10. Decrease in SOD activity was prevented by DMSO pretreatment. In the electron microscopic study, the mitochondria in the irradiated mouse liver were swollen, but we could observe no change after DMSO pretreatment. The results suggest that DMSO has radioprotective effects, probably due to inhibition of lipid peroxidation.     

Regional, developmental, and cell cycle-dependent differences in mu, delta, and kappa-opioid receptor expression among cultured mouse astrocytes

Of all skeletal muscles examined in the rat, the spinotrapezius (S) and diaphragm (D) have the closest fiber-type composition. However, their oxidative capacities differ by two- to threefold. We have developed an intravital microscopy preparation to study diaphragm microcirculation in vivo. Using this preparation and the standard spinotrapezius model first described by S. D. Gray (1973, Microvasc. Res. 5, 395-400), we tested the hypothesis that pronounced microcirculatory differences would exist between these two muscles as a function of their disparate oxidative capacities. The lineal density of all capillaries in the spinotrapezius was 33.6 +/- 1.5 compared to 65.1 +/- 3.3 capillaries/mm in the diaphragm (P < 0.001). In the diaphragm compared with the spinotrapezius muscle, a significantly (P < 0.05) greater proportion of capillary countercurrent flow (D, 29 +/- 6% vs 8 +/- 6%) existed. Within both muscles, there was a similar proportion of capillaries supporting red blood cell (RBC) flow (S, 89 +/- 7% vs D, 92 +/- 2%). However, the diaphragm supported significantly (P < 0.001) greater intracapillary RBC velocities (D, 302 +/- 11 vs S, 226 +/- 9 micron/s) and fluxes (D, 33.4 +/- 1.1 vs S, 19.2 +/- 2.1 cells/s) compared with the spinotrapezius. Capillary "tube" hematocrit was greater (P = 0.01) in the diaphragm (0.32 +/- 0.02) than in the spinotrapezius (0.22 +/- 0.03) muscle. These data demonstrate that microcirculatory flow characteristics in resting muscle can be regulated independent of muscle fiber-type composition and may be related to muscle oxidative capacity.     

Posttreatment with eicosatetraynoic acid decreases lung edema in guinea pigs exposed to phosgene: the role of leukotrienes

Acetylenic acids such as 5,8,11,14-eicosatetraynoic acid (ETYA), have been shown to be effective in preventing pulmonary edema formation (PEF). In phosgene-exposed guinea pigs, we examined the effects of ETYA on PEF, measured as real time lung weight gain (lwg). Pulmonary artery pressure (Ppa), airway pressure (Paw), perfusate leukotrienes (LT) C4/D4/E4/B4, and lung tissue lipid peroxidation (TBARS) were measured using the isolated, buffer-perfused lung model. Guinea pigs were challenged to 175 mg/m3 (44 ppm) phosgene for 10 minutes giving a concentration x time product of 1750 mg.min/m3 (437 ppm.min). Five minutes after removal from the exposure chamber, guinea pigs were treated, i.p., with 200 microL of 100 microM ETYA. 200 microL of 50 microM ETYA was added to the perfusate every 40 minutes, beginning at 60 minutes after start of exposure (t = 0). There were four groups in this study: air-treated, phosgene-exposed, ETYA-posttreated + phosgene, and ETYA-posttreated + air ETYA-posttreated + phosgene guinea pigs had significantly lower Ppa (P = .006), Paw (P = .009), and lwg (P = .016) compared with phosgene-exposed animals. Phosgene exposure reduced LTB4 compared with air-treated controls (P = .09). ETYA-posttreatment + phosgene had significantly increased perfusate LTB4 (P = .0006) compared with phosgene exposure only group. Total perfusate, LTC4 + LTD4 + LTE4, was not different between phosgene-exposed, air-treated or ETYA-posttreatment + phosgene over time. Posttreatment with ETYA significantly lowered TBARS formation, 206 +/- 13 versus 285 +/- 23 nmol/mg protein (P = .016), compared with phosgene-exposed lungs. Paradoxically, ETYA posttreatment decreased PEF and lipid peroxidation, but increased sulfidopeptide LT release from the lung during perfusion. We conclude that LTC4/D4/E4, and B4, may play different roles than previously thought for PEF in the isolated perfused lung model.     

Salbutamol enhances isotonic contractile properties of rat diaphragm muscle

The effects of the beta2-adrenoceptor agonist salbutamol (Slb) on isometric and isotonic contractile properties of the rat diaphragm muscle (Diamus) were examined. A loading dose of 25 microg/kg Slb was administered intracardially before Diamus excision to ensure adequate diffusion. Studies were then performed with 0.05 microM Slb in the in vitro tissue chamber. cAMP levels were determined by radioimmunoassay. Compared with controls (Ctl), cAMP levels were elevated after Slb treatment. In Slb-treated rats, isometric twitch and maximum tetanic force were increased by approximately 40 and approximately 20%, respectively. Maximum shortening velocity increased by approximately 15% after Slb treatment, and maximum power output increased by approximately 25%. During repeated isotonic activation, the rate of fatigue was faster in the Slb-treated Diamus, but both Slb-treated and Ctl Diamus fatigued to the same maximum power output. Still, endurance time during repetitive isotonic contractions was approximately 10% shorter in the Slb-treated Diamus. These results are consistent with the hypothesis that beta-adrenoceptor stimulation by Slb enhances Diamus contractility and that these effects of Slb are likely mediated, at least in part, by elevated cAMP.