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PubMed1996

Osmotic tolerance limits and properties of murine spermatozoa.

Osmotic tolerance of spermatozoa is a critical determinant of functional survival after cryopreservation. This study first tested the hypothesis that mouse spermatozoa behave as linear osmometers, using an electronic particle counter to measure the change in sperm volume in response to anisosmotic solutions. The resulting Boyle-van't Hoff plot was linear (r2 = 0.99) from 75 to 1200 mOsmolal and indicates that 60.7% of the total cell volume is osmotically inactive. Next, mouse sperm tolerance to osmotic stress was determined by assessment of plasma membrane integrity, mitochondrial viability, and motility. Each functional endpoint was measured after exposure to anisosmotic solutions and again after return to isosmolality. The dual fluorescent stains-carboxyfluorescein diacetate with propidium iodide and Rhodamine 123 with propidium iodide-were used to determine membrane integrity and functional mitochondria, respectively. Motility was measured by video microscopy in the range of 1-2400 mOsmolal and was further analyzed from 140 to 600 mOsmolal using computer-assisted semen analysis. The data indicate that motility is substantially more sensitive to osmotic stress than either mitochondrial viability or membrane integrity and that mouse spermatozoa should be maintained within 76-124% of their isosmotic volume during cryopreservation in order to maintain > 80% of pretreatment motility.

Animals↗
PubMed1997

Biochemical characterization of sperm agglutination antigen-1, a human sperm surface antigen implicated in gamete interactions.

The anti-sperm monoclonal antibody (mAb) S19 was previously demonstrated to agglutinate human spermatozoa, inhibit sperm penetration of cervical mucus, and inhibit sperm-zona pellucida binding. These results implicated the cognate S19 antigen, designated sperm agglutination antigen-1 (SAGA-1), in gamete interactions and identified SAGA-1 as an attractive candidate for immunocontraceptive development. In the present study, evaluation of sperm agglutination with video microscopy showed that the S19 mAb rapidly and completely agglutinated human spermatozoa in a "tangled" pattern of agglutination. One- and two-dimensional immunoblot analyses identified SAGA-1 as a highly acidic, polymorphic sperm protein with an apparent molecular mass of 15-25 kDa and an isoelectric point of 2.5-3.0. Periodate treatment abolished this immunoreactivity, demonstrating that the S19 mAb reacted with a carbohydrate epitope and indicating that SAGA-1 is a glycoprotein. Absence of S19 immunoreactivity in postvasectomy seminal fluid implicated the testis, epididymis, and/or proximal vas deferens in the expression of SAGA-1. In solubility and phase partitioning assays, SAGA-1 was extracted from spermatozoa in Triton X-114 and exhibited the hydrophobic characteristics of integral and glycosylphosphatidyl inositol-anchored membrane proteins. These results identify SAGA-1 as a hydrophobic, highly acidic sperm glycoprotein that is localized on the entire sperm surface and has potential significance as a target for antibodies that inhibit sperm function and gamete interactions.

Antibodies, Monoclonal↗
PubMed1999

Statoliths motions in gravity-perceiving plant cells: does actomyosin counteract gravity?

Statocytes from plant root caps are characterized by a polar arrangement of cell organelles and sedimented statoliths. Cortical microtubules and actin microfilaments contribute to development and maintenance of this polarity, whereas the lack of endoplasmic microtubules and prominent bundles of actin microfilaments probably facilitates sedimentation of statoliths. High-resolution video microscopy shows permanent motion of statoliths even when sedimented. After immunofluorescence microscopy using antibodies against actin and myosin II the most prominent labeling was observed at and around sedimented statoliths. Experiments under microgravity have demonstrated that the positioning of statoliths depends on the external gravitational force and on internal forces, probably exerted by the actomyosin complex, and that transformation of the gravistimulus evidently occurs in close vicinity to the statoliths. These results suggest that graviperception occurs dynamically within the cytoplasm via small-distance sedimentation rather than statically at the lowermost site of sedimentation. It is hypothesized that root cap cells are comparing randomized motions with oriented motions of statoliths and thereby perceiving gravity.

Actomyosin↗
PubMed1988

Irreversible injury in anoxic hepatocytes precipitated by an abrupt increase in plasma membrane permeability.

Using low-light digitized video microscopy, the onset, progression, and reversibility of anoxic injury were assessed in single hepatocytes isolated from fasted rats. Cell-surface bleb formation occurred in three stages over 1-3 h after anoxia. Stage I was characterized by formation of numerous small blebs. In stage II, small blebs enlarged by coalescence and fusion to form a few large terminal blebs. Near the end of stage II, cells began to swell rapidly, ending with the apparent breakdown of one of the terminal blebs. Breakdown of the bleb membrane initiated stage III of injury and was coincident with a rapid increase of nonspecific permeability to organic cationic and anionic molecules. On reoxygenation, stages I and II were fully reversible, and plasma membrane blebs were resorbed completely within 6 min of reoxygenation without loss of viability. Stage III, however, was not reversible, and no morphological changes occurred on reoxygenation. The results indicate that onset of cell death owing to anoxia is a rapid event initiated by a sudden increase of nonspecific plasma membrane permeability caused by rupture of a terminal bleb. Anoxic injury is reversible until this event occurs.

Animals↗
PubMed1994

Induced redistribution of cell surface receptors by alternating current electric fields.

The molecular mechanisms that underlie the biological effects of low frequency sinusoidal electric fields may involve induced changes in the physical state of charged cell surface receptors. We have used intensified fluorescence video microscopy to study the redistribution of cell surface receptors, including transferrin receptors (TFR) and low density lipoprotein receptors (LDL-R), in response to externally applied alternating current electric fields in the 3 to 23 V/cm range (peak to peak). Redistribution of both TFR and LDL-R was prominent at frequencies of 1 and 10 Hz but negligible at frequencies of 60 and 120 Hz. Application of a 1 Hz, 23 V/cm field for 15 min caused a twofold change in local TFR surface density, whereas application of a 60 Hz, 23 V/cm field resulted in no significant TFR redistribution. The extent of TFR redistribution induced by a 1 Hz field changed by only 20% over the field strength range from 3.5 to 23 V/cm. AC field-induced cell surface receptor migration did not consistently follow electric field lines, suggesting that mechanisms more complex than classical electrophoresis and electroosmosis mediate receptor redistribution. Joule heating and plasma membrane calcium channel activation were shown not to be involved in the mechanism of receptor redistribution. Applied external electric fields may reorganize cytoskeletal and plasma membrane structures, providing pathways for cell surface receptors to migrate anharmonically.

Calcium Channels↗
PubMed2004

Inhalation of carbon monoxide prevents liver injury and inflammation following hind limb ischemia/reperfusion.

The induction of heme oxygenase (HO), the rate limiting enzyme in the conversion of heme into carbon monoxide (CO) and biliverdin, limits liver injury following remote trauma such as hind limb ischemia/reperfusion (I/R). Using intravital video microscopy, we tested the hypothesis that inhaled CO (250 ppm) would mimic HO-derived liver protection. Hind limb I/R significantly decreased sinusoidal diameter and volumetric flow, increased leukocyte accumulation within sinusoids, increased leukocyte rolling and adhesion within postsinusoidal venules, and significantly increased hepatocyte injury compared with naive animals. Inhalation of CO alone did not alter any microcirculatory or inflammatory parameters. Inhalation of CO following I/R restored volumetric flow, decreased stationary leukocytes within sinusoids, decreased leukocyte rolling and adhesion within postsinusoidal venules, and significantly reduced hepatocellular injury following hind limb I/R. HO inhibition did not alter microcirculatory parameters in naive mice, but did increase inflammation, as well as increase hepatocyte injury following hind limb I/R. Inhalation of CO during HO inhibition significantly reduced such microcirculatory deficits, hepatic inflammation, and injury in response to hind limb I/R. In conclusion, these results suggest that HO-derived hepatic protection is mediated by CO, and inhalation of low concentrations of CO may represent a novel therapeutic approach to prevent remote organ injury during systemic inflammatory response syndrome, or SIRS.

Administration, Inhalation↗
PubMed2006

Increased migration and metastatic potential of tumor cells expressing aquaporin water channels.

Aquaporin (AQP) water channels are expressed in high-grade tumor cells of different tissue origins. Based on the involvement of AQPs in angiogenesis and cell migration, we tested whether AQP expression in tumor cells might enhance their migration and metastatic potential. Transfection of B16F10 and 4T1 tumor cells with AQP1 did not affect their appearance, size, growth, or substrate adherence but increased their plasma membrane osmotic water permeability by 5- to 10-fold. In vitro analysis of cell migration by transwell assay, wound healing and video microscopy showed a 2- to 3-fold accelerated migration of the AQP1-expressing tumor cells compared to control cells. In mice, AQP1 expression increased tumor cell extravasation by >1.5-fold as quantified by counting tumor cells in lung at 6 h after tail vein injection of a mixture of fluorescently tagged AQP1-expressing and control tumor cells. AQP1 expression also increased by 3-fold the number of lung metastases 14 days after tail vein injection of tumor cells, with alveolar wall infiltration seen with AQP1-expressing tumor cells. Our results provide evidence for AQP-facilitated tumor cell migration and spread, suggesting a novel function for AQP expression in high-grade tumors. AQP inhibition may thus reduce the metastatic potential of some tumors.

Animals↗
PubMed1996

Effect of volatile anesthetics on hydrogen peroxide-induced injury in aortic and pulmonary arterial endothelial cells.

BACKGROUND: Oxidant damage to endothelial cells occurs during inflammation and reperfusion after ischemia, mediated in part by endogenously produced hydrogen peroxide (H2O2). Previous studies have established a role for increased cytosolic calcium in the mechanism of endothelial oxidant injury, and have suggested that volatile anesthetics may exacerbate oxidant injury in pulmonary endothelium. However, the effect of volatile anesthetics on oxidant injury to systemic arterial endothelial cells, and their effect on oxidant-related changes in cytosolic calcium homeostasis, have not been reported previously. METHODS: Primary cultures of human aortic and pulmonary arterial endothelial cells were studied. The rate of cell death after H2O2 exposure was determined in cell suspension by propidium iodide fluorimetry and lactate dehydrogenase release. The final extent of cell death 24 h after H2O2 exposure was determined in monolayer cultures by methyl thiazolyl tetrazolium reduction. Cytosolic calcium and cell death were determined in single cells using fura-2 and propidium iodide imaging with digitized, multiparameter, fluorescent video microscopy. RESULTS: In aortic endothelial cells, clinical concentrations of halothane (1.0%) and isoflurane (1.5%) decreased both the rate of cell death and the final extent of cell death after H2O2 exposure, with halothane being more protective. Supraclinical concentrations of halothane (2.7%) and isoflurane (4.0%) were less protective. In pulmonary arterial endothelial cells, halothane and isoflurane had essentially no effect on H2O2-mediated cell death. The protective effect of anesthetic in aortic endothelial cells was not due to an enhanced removal of H2O2 by endogenous enzymes. Hydrogen peroxide exposure caused a large increase in cytosolic calcium well before cell death, and this was moderated by anesthetic treatment. CONCLUSIONS: The effect of volatile anesthetics on oxidant injury to endothelial cells may differ between cells derived from systemic and pulmonary vascular beds. Halothane, and to a lesser extent, isoflurane, protects against oxidant injury in aortic endothelial cells. The mechanism of protection may involve modulation of the interaction of H2O2 with vital cellular constituents, and/or amelioration of the toxic increase in cytosolic calcium that follows such interaction.

Anesthetics, Inhalation↗
PubMed1996

Direct vasomotor effects of isoflurane in subepicardial resistance vessels from collateral-dependent and normal coronary circulation of pigs.

BACKGROUND: The purpose of the current investigation was to determine and compare the direct effect of isoflurane on normal resistance coronary arteries and similarly sized coronary arteries in collateral-dependent (CD) circulation. A differential vasomotor effect of isoflurane might contribute to flow redistribution, either adverse or favorable, between normal and CD regions. METHODS: The authors used a swine model of chronic coronary occlusion. Six weeks after ameroid occluder placement around the left circumflex artery, the heart was removed. Myocardial blood flow, measured in vivo before the removal, was lower during rapid atrial pacing in the CD region than in the normally perfused region. These in vivo studies established the existence and location of CD circulation. Subepicardial microvessels, measuring approximately 100 microns, were dissected from both the CD region and the normal region. Either with or without preconstriction of the vessels with the thromboxane analogue U46619 1 microM, direct vasomotor effect of isoflurane was examined in vitro by video microscopy. In addition, vasomotor responses to endothelin-1, the endothelium-dependent dilator adenosine 5' diphosphate, and the endothelium-independent dilator sodium nitroprusside were compared between CD and normal resistance vessels. RESULTS: Isoflurane caused greater concentration-dependent constriction of the normal vessels than of the CD vessels (P < 0.05); this constrictive effect was masked by preconstriction of the vessels. Vasodilation of ADP was less in CD vessels than in normal vessels (P < 0.001), but vasodilation to sodium nitroprusside was not significantly different (P = 0.45). Vasoconstriction to endothelin-1 was greater in CD vessels than in normal vessels (P < 0.01). CONCLUSIONS: The vasoconstrictive effect of isoflurane normally observed in coronary resistance vessels is impaired in resistance vessels that supply a CD area. The basis of this may be related to endothelial dysfunction.

Anesthetics, Inhalation↗
PubMed1997

Leukocyte-endothelium interaction in the rat mesenteric microcirculation during halothane or sevoflurane anesthesia .

BACKGROUND: The effects of inhalational anesthetics on the microcirculation, including leukocyte dynamics, remain to be clarified. The authors investigated halothane and sevoflurane anesthesia to determine if these agents evoked leukocyte adhesion through endothelial cell-dependent mechanisms involving such adhesion molecules. METHODS: Rats were anesthetized with halothane or sevoflurane in 100% oxygen and the lungs were mechanically ventilated. Leukocyte behavior in mesenteric venules was recorded through intravital video microscopy under monitoring microvascular hemodynamics. To examine the mechanisms for leukocyte rolling and adhesion, these studies were repeated after animals were pretreated with a monoclonal antibody against P-selectin (MAb PB1.3) or against intracellular adhesion molecule-1 (ICAM-1; MAb 1A29): P-selectin required for rolling of circulating leukocytes and ICAM-1 for firm adhesive interactions with leukocyte integrins. RESULTS: Under baseline anesthetic conditions (1 minimum alveolar concentration [MAC]), venular wall shear rates, an index of the disperse force on marginating leukocytes, in the sevoflurane-treated rats were about two times higher than those with halothane. At 2 MAC, halothane caused a marked arteriolar constriction and decreasing shear rates concurrent with an increasing density of venular leukocyte adhesion. Sevoflurane at 2 MAC induced leukocyte rolling and adhesion, which were attenuated by PB1.3 and 1A29, without alterations in the wall shear rates. Halothane-induced leukocyte adhesion was not prevented by PB1.3 but it was by 1A29. CONCLUSIONS: Halothane or sevoflurane anesthesia induces venular leukocyte rolling and adhesion: P-selectin upregulation plays a crucial role in leukocyte rolling and adhesion during sevoflurane anesthesia, whereas low-flow perfusion is likely to evoke ICAM-1-dependent leukocyte adhesion during halothane anesthesia.

Anesthetics, Inhalation↗
PubMed1993

The fast axonal transport in hippocampal neurones is acutely enhanced by db-cAMP.

It has been observed that neurones have a certain capacity for the upregulation of fast axonal transport, for instance during nerve regeneration or reactive sprouting. However, the molecular regulation of this transport system is largely unknown. We show here by quantitative video-microscopy of endogenous organelles that application of 1 mM db-cAMP increases the velocity of fast axonal transport maximally by 32% within 60 minutes in neonatal hippocampal cells. At the same time, the jump length of the saltatory motions remains largely unchanged. The data suggest that activation of protein kinase A plays a role in the immediate upregulation of axonal transport.

Animals↗
PubMed1997

The RGD-containing domain of exogenous HIV-1 Tat inhibits the engulfment of apoptotic bodies by dendritic cells.

OBJECTIVE: HIV-1 Tat can be released by infected cells and exert various extracellular functions on bystander cells, possibly contributing to immunodeficiency. In order to investigate whether exogenous Tat can affect antigen presentation, the effects of synthetic Tat on the function of dendritic cells displaying antigen presenting cell phenotype were studied. DESIGN: Cultured dendritic cells were challenged with apoptotic bodies and monitored for cell engulfment and free intracellular calcium ([Ca2+]i) increase. The effect of synthetic HIV-1 Tat and its RGD-containing domain (peptide 65-80) or basic domain (peptide 46-60) on both functions was investigated. METHODS: Dendritic cells were obtained by culture of monocytes with granulocyte-macrophage colony-stimulating factor. Apoptosis was induced in Jurkat cells by sub-lethal irradiation. Engulfment of radiolabelled apoptotic bodies by dendritic cells was obtained by a 45 min co-incubation at 37 degrees C. Non-ingested apoptotic bodies were removed and cell-associated radioactivity evaluated in a gamma-counter after cell lysis. Single cell analysis of calcium fluxes was performed by video-microscopy and ratio-imaging, after cell staining with the fluorescent calcium chelator FURA-2. RESULTS: Apoptotic bodies were engulfed by dendritic cells: this process was accompanied by [Ca2+]i rise. Synthetic HIV-1 Tat inhibited both apoptotic body engulfment and [Ca2+]i increase. The same inhibition was obtained with the RGD-containing domain (peptide 65-80), but not with the basic domain (peptide 46-60) of Tat, suggesting the involvement of an integrin. This integrin is likely to be alpha v beta 3, since RGD-containing peptides from vitronectin, but not from fibronectin, inhibited apoptotic body engulfment. Furthermore, both HIV-1 Tat and its 65-80 peptide blocked [Ca2+]i increase due to beta 3-integrin cross-linking. CONCLUSIONS: Our results support a role for HIV-1 Tat in decreasing the function of dendritic cells, possibly impairing antigen presentation.

Antigen Presentation↗
PubMedDate not supplied

Endothelial cell seeding with rotation of a ventricular blood sac.

Successful establishment of a durable endothelial cell (EC) monolayer inside a ventricular blood sac requires homogeneous coverage of the entire luminal surface with attached cells. For this purpose, a new device was developed that slowly rotates a fully assembled cardiac prosthesis with three degrees of freedom. We seeded ECs derived from human adipose tissue at a density of approximately 3.5 x 10(4) cells/cm2 onto the surfaces of polyurethane-made blood sacs and "ersatz" bladders (consisting of T-25 tissue culture flasks). The kinetics of cell attachment, spreading, and proliferation were determined using video microscopy combined with image analysis and cell viability assays. After 60 min of seeding at 5-10 rotations/hr, the plating efficiency inside the blood sacs was 35.7 +/- 11%, with cell viability remaining approximately 90 +/- 5%. After 3 hr, when the plating efficiency reached a plateau (approximately 70%), the rotation was stopped and the ECs were allowed to spread and proliferate under static conditions. Within 48 hr, the entire luminal surface was evenly covered by a confluent EC monolayer. Our long-term studies show that with a proper feeding schedule, such an EC monolayer can be maintained intact in vitro for more than 2 weeks.

Adipose Tissue↗
PubMedDate not supplied

Evaluation of a new heparin bonding process in prolonged extracorporeal membrane oxygenation.

A novel heparin bonding method has been developed for in situ surface modification using ozone oxidation, and evaluated in vitro and in vivo during prolonged extracorporeal membrane oxygenation (ECMO) experiments. The ECMO system consisted of a Capiox hollow fiber membrane oxygenator (MO; Terumo Corp., Tokyo, Japan) with an integral heat exchanger, a Capiox centrifugal pump (CP), and an extracorporeal circuit. The blood contacting surfaces of the system were completely modified using the heparin bonding process, and evaluated in a chronic sheep model for extended period of time, ranging from 96 to 168 hr, under minimal systemic heparinization. The heparin bonded surface was able to maintain high levels of heparin bioactivity, and showed improved blood compatibility in in vitro epifluorescent video microscopy experiments by suppressing platelet adhesion/activation and complement activation. For the ECMO experiments, extracorporeal blood flow was maintained at 3 L/min and the activated clotting time was maintained at 150 sec. There was no significant change in the gas transfer capability or mean pressure drop across the MO over experimental times of up to 168 hr. Platelet count and other coagulation parameters remained stable within the physiologic range throughout the experiment. There were no detectable thrombi in the CP, tubing, or connectors in the test circuit. Local thrombus formation was noted in a stagnant area of the MO, although this did not interfere with its function or lead to any significant embolization. Based on these results, the heparin bonded ECMO system appears to be a safe and effective device for prolonged extracorporeal circulation under minimal systemic heparinization.

Animals↗
PubMedDate not supplied

Bacterial adhesion on polyurethane surfaces conditioned with thrombus components.

Thrombosis and infection are two major complications associated with cardiovascular devices such as ventricular assist devices, total artificial hearts, vascular grafts, and catheters. When blood contacts an artificial biomaterial, protein deposition occurs, as do activation of the blood coagulation cascade, platelet adhesion, activation, and aggregation, all of which lead to thrombus formation. An increased incidence of bacterial infection also has been seen clinically with indwelling biomaterial devices. Some evidence suggests a possible association between thrombosis and infection, in that adherent bacteria may provide a nidus for thrombus formation, or adherent thrombi composed of platelets and fibrin may form sheltered sites for bacterial adhesion. In the current study, the authors examined Staphylococcus aureus adhesion to sulfonated, aminated, and phosphonated polyurethane surfaces that had been pre-adsorbed with solutions of increasing complexity, in an effort to approach and simulate clot formation on the surface. These solutions included various combinations of fibrinogen, albumin, plasma, thrombin, and isolated platelets. Bacterial adhesion was observed in a radial flow chamber mounted on the motorized stage of a video microscopy system, with image processing software used to perform automated data collection and image analysis. Scanning electron microscopy also was used to visualize cross-linked fibrin and bacterial adhesion on these surfaces. Bacterial adhesion was found to be lowest on the phosphonated polyurethane. The presence of fibrin or isolated platelets significantly increased bacterial adhesion compared to surfaces pre-adsorbed with albumin.

Bacterial Adhesion↗
PubMedDate not supplied

Improved blood compatibility of DLC coated polymeric material.

There is currently an increasing interest in the use of DLC (diamond like carbon) films in biomedical applications. These investigations making use of DLC in the biomedical area indicate its attractive properties. In this study, we succeeded in depositing DLC on polymer substrates and found the best conditions and method for this application. We evaluated the blood compatibility of polycarbonate substrates coated by DLC (PC-DLC) under different conditions by using epifluorescent video microscopy (EVM) combined with a parallel plate flow chamber. Segmented polyurethane (SPU), which has been used to fabricate medical devices including an artificial heart, and proven to have acceptable blood compatibility, was compared with polycarbonate substrates coated with DLC film. The EVM system measured platelet adhesion on the surface of the DLC, by using whole human blood containing Mepacrine labeled platelets perfuse at a wall shear rate of 100 s(-1) at 1 min intervals for a period of 20 min. PC-DLC demonstrated that Tecoflex showed higher complement activation than PC-DLC. There were significant differences between the PC-DLC substrates. On the basis of these results, it is recommended for use as a coating material in implantable blood contacting devices such as artificial hearts, pacemakers, and other devices. This DLC seems to be a promising candidate for biomaterials applications and merits further investigation.

Carbon↗
PubMed1995

Ketamine attenuates endotoxin-induced leukocyte adherence in rat mesenteric venules.

OBJECTIVES: To determine the influence of ketamine on endotoxin-induced leukocyte adherence and venular microhemodynamics. DESIGN: Randomized, controlled trial. SETTING: Experimental laboratory. SUBJECTS: Thirty male Wistar rats. INTERVENTIONS: The rats were pretreated with ketamine (10 mg/kg iv) or 0.9% saline, and both groups were given endotoxin (Escherichia coli lipopolysaccharide; 5 mg/kg iv). The control group received two doses of 0.9% saline. MEASUREMENTS AND MAIN RESULTS: The rates of leukocyte adherence and changes in microhemodynamics were monitored in rat mesenteric venules, using in vivo video microscopy. The number of adherent leukocytes was determined on-line in 10-min intervals from 60 mins before until 2 hrs after endotoxin administration. Venular diameters, red blood cell velocity, volumetric blood flow, and the venular wall shear rate were monitored before and at 10, 30, and 60 mins after endotoxin exposure. A 6.3-fold increase in the number of adherent leukocytes was observed 10 mins after administration of endotoxin when compared with control animals (5.87 +/- 0.69 vs. 0.93 +/- 0.21 adherent cells/100 microns; p < .001). This increase remained unchanged for 120 mins. In ketamine-pretreated rats, a 2.6-fold increase in leukocyte adherence occurred during the first 20 mins after endotoxin exposure (2.40 +/- 0.46 vs. 0.93 +/- 0.21 adherent cells/100 microns; p < .01). However, no difference in the number of adherent leukocytes between ketamine-pretreated and control animals was found after this 20-min period. In animals of the control group, no increase in leukocyte adherence occurred during the entire observation time. Diameters of mesenteric venules did not change after endotoxin exposure in any of the groups. Red blood cell velocity and venular blood flow in the endotoxin-treated groups decreased 10 mins after the injection of endotoxin when compared with controls, but these values did not show any difference when they were compared between ketamine and saline-pretreated animals. Similarly, venular wall shear rate in the endotoxin-treated groups decreased 10 and 30 mins after injection of endotoxin. However, no significant difference occurred between ketamine and saline-pretreated animals. CONCLUSIONS: Pretreatment with ketamine attenuates endotoxin-induced leukocyte adherence by a shear rate-independent mechanism, suggesting reduced expression of adhesion molecules. These results indicate that ketamine exerts an anti-inflammatory effect, which might be beneficial in septic patients.

Animals↗
PubMed1998

Neither dopamine nor dobutamine corrects mesenteric blood flow depression caused by positive end-expiratory pressure in a rat model of acute lung injury.

OBJECTIVE: To determine if either dopamine or dobutamine would counteract the deleterious effect that positive end-expiratory pressure (PEEP) has on cardiac output and mesenteric blood flow in a rat model of acute lung injury. DESIGN: Prospective, randomized, controlled trial in a clinically relevant model of acute lung injury. SETTING: Microcirculation research laboratory. SUBJECTS: Male Sprague-Dawley rats. INTERVENTIONS: The animals were anesthetized with pentobarbital (30 mg/kg) by intraperitoneal injection. They underwent tracheostomy, jugular and femoral vein cannulation, femoral artery cannulation, carotid artery thermistor placement, and bowel preparation for in vivo video microscopy. Acute lung injury was created by administering 0.1 N hydrochloric acid (1 mL/kg) via the tracheostomy. Dopamine or dobutamine (2.5 or 12.5 microg/kg/min), followed by two intravenous fluid boluses, was administered to rats ventilated with 5, 10, 15, and 20 cm H2O of PEEP. MEASUREMENTS AND MAIN RESULTS: Mean arterial pressure, thermodilution cardiac output, mesenteric arteriolar diameter, and red blood cell velocity were measured and mesenteric blood flow was calculated. Cardiac output was depressed in rats exposed to 20 cm H2O of PEEP by 32+/-2%. The corresponding values for cardiac output depression at 20 cm H2O of PEEP in rats receiving 2.5 and 12.5 microg/kg/min of dopamine and 2.5 and 12.5 microg/kg/min of dobutamine were 31+/-1%, 21+/-1%, 29+/-0%, and 24+/-2%, respectively. Mesenteric blood flow was depressed in rats ventilated with 20 cm H2O of PEEP by 74+/-3%, while the corresponding values in rats exposed to 20 cm H2O of PEEP and receiving 2.5 or 12.5 microg/kg/min of dopamine or 2.5 or 12.5 microg/kg/min of dobutamine were 86+/-3%, 77+/-3%, 73+/-3%, and 66+/-3%, respectively. Fluid boluses did not correct the deficits in cardiac output or mesenteric blood flow caused by the combination of acute lung injury and PEEP. CONCLUSIONS: The higher doses of dopamine and dobutamine partially, but insignificantly, corrected the cardiac output depression caused by PEEP in a model of acute lung injury. Neither dose of dopamine nor dobutamine was able to improve PEEP-induced mesenteric blood flow depression.

Animals↗