Sexually dimorphic phenotype of arteriolar responsiveness to shear stress in soluble epoxide hydrolase-knockout mice

We hypothesized that potentiating the bioavailability of endothelial epoxyeicosatrienoic acids (EETs) via deletion of the gene for soluble epoxide hydrolase (sEH), or downregulation of sEH expression, enhances flow/shear stress-induced dilator responses (FID) of arterioles. With the use of male (M) and female (F) wild-type (WT) and sEH-knockout (KO) mice, isolated gracilis muscle arterioles were cannulated and pressurized at 80 mmHg. Basal tone and increases in diameter of arterioles as a function of perfusate flow (5, 10, 15, 20, and 25 μl/min) were recorded. The magnitude of FID was significantly smaller and associated with a greater arteriolar tone in M-WT than F-WT mice, revealing a sex difference in FID. This sex difference was abolished by deletion of the sEH gene, as evidenced by an enhanced FID in M-KO mice to a level comparable with those observed in F-KO and F-WT mice. These three groups of mice coincidentally exhibited an increased endothelial sensitivity to shear stress (smaller WSS50) and were hypotensive. Endothelial EETs participated in the mediation of enhanced FID in M-KO, F-KO, and F-WT mice, without effects on FID of M-WT mice. Protein expression of sEH was downregulated by approximately fourfold in vessels of F-WT compared with M-WT mice, paralleled with greater vascular EET levels that were statistically comparable with those observed in both male and female sEH-KO mice. In conclusion, sex-different regulation of sEH accounts for sex differences in flow-mediated dilation of microvessels in gonadally intact mice.

Glomerular tubular balance is suppressed in adenosine type 1 receptor-deficient mice

Glomerular tubular balance maintains a stable fractional solute and fluid reabsorption in the proximal tubule over a range of glomerular filtration rates. The mediators of this process are unknown. We tested the hypothesis that adenosine, produced in proximal tubule cells acting on adenosine type 1 receptors (A1-AR) promotes Na+ and fluid uptake and mediates glomerular tubular balance. Absolute proximal fluid reabsorption ( Jv) was measured by in vivo microperfusion in A1-AR knockout and wild-type mice during perfusion of the closed proximal tubule at 2–10 nl/min. Jv increased with perfusate flow from 2–4 nl/min in both strains, but the fractional increase was lower in A1-AR−/− mice (A1-AR+/+: 114% vs. A1-AR−/−: 38%; P < 0.001), suggesting reduced glomerular tubular balance (GTB). At higher perfusion rates, Jv increased modestly in both strains, indicating less GTB at higher flow. The physiological effects of reduced GTB in A1-AR−/− mice were assessed from the response to an acute volume load (1 ml/2 min). Na+ excretion and urine flow increased 76 and 73% more in A1-AR−/− mice than A1-AR+/+ over the following 30 min, accompanied by a higher proximal tubule flow (A1-AR−/−: 6.9 ± 0.9 vs. A1-AR+/+: 5.2 ± 0.6 nl/min; P < 0.05). The expression of the sodium-hydrogen exchanger 3 and sodium phosphate cotransporter-2 were similar between strains. In conclusion, GTB is dependent on adenosine acting on type 1 receptors in the proximal tubule. This may contribute to acute changes in Na+ and fluid reabsorption.

Vaporized Perfluorohexane Attenuates Ventilator-induced Lung Injury in Isolated, Perfused Rabbit Lungs

Background The authors tested the hypothesis that administration of vaporized perfluorohexane may attenuate ventilator-induced lung injury. Methods In isolated, perfused rabbit lungs, airway pressure-versus-time curves were recorded. At baseline, peak inspiratory pressure and positive end-expiratory pressure of mechanically ventilated lungs were set to obtain straight pressure-versus-time curves in both the lower and upper ranges, which are associated with less collapse and overdistension, respectively. After that, peak inspiratory pressure and positive end-expiratory pressure were set at 30 cm H2O and 0, respectively, and animals were randomly assigned to one of two groups: (1) simultaneous administration of 14% perfluorohexane vapor in room air (n = 7) and (2) control group-ventilation with room air (n = 7). After 20 min of cycling collapse and overdistension, tidal volume and positive end-expiratory pressure were set back to baseline levels, administration of perfluorohexane in the therapy group was stopped, and mechanical ventilation was continued for up to 60 min. Lung weight, mean pulmonary artery pressure, and concentration of thromboxane B2 in the perfusate were measured. In addition, the distribution of pulmonary perfusate flow was assessed by using fluorescent-labeled microspheres. Results Significantly higher peak inspiratory values developed in control lungs than in lungs treated with perfluorohexane. In addition, upper ranges of pressure-versus-time curves were closer to straight lines in the perfluorohexane group. Lung weight, mean pulmonary arterial pressure, and release of thromboxane B2 were significantly higher in controls than in perfluorohexane-treated lungs. Also, redistribution of pulmonary perfusate flow from caudal to cranial zones was less important in the treatment group. Conclusion The authors conclude that the administration of perfluorohexane vapor attenuates the development of ventilator-induced lung injury in isolated, perfused rabbit lungs.

Development of an in situ perfused kidney preparation for elasmobranch fish: action of arginine vasotocin

Acclimation of the European lesser-spotted dogfish Scyliorhinus canicula to reduced environmental salinity [85–70% seawater (SW)] induced a significant diuresis in addition to a significant decrease in plasma osmolality in vivo. The threshold for this diuresis was determined to be 85% SW. Therefore, S. canicula acclimated to 85% SW was selected for further study as a diuretic model in the development of an in situ perfused kidney preparation. The renal role of arginine vasotocin (AVT) in the in situ perfused trunk preparation was investigated. In SW, perfusion of 10−9 and 10−10 M AVT resulted in a glomerular antidiuresis and decreases in tubular transport maxima for glucose and perfusate flow. In 85% SW, 10−10 M AVT had no significant effect on these renal parameters with the exception of transport maxima for glucose and perfusate flow. Tubular parameters remained unchanged by either 10−9 or 10−10 M AVT. The results demonstrate that the perfused kidney preparation was a viable tool for the investigation of renal parameters in elasmobranch fish and that AVT induced a glomerular antidiuresis.

Flow-mediated release of nitric oxide in isolated, perfused rabbit lungs

The effects of changing perfusate flow on lung nitric oxide (NO) production and pulmonary arterial pressure (Ppa) were tested during normoxia and hypoxia and after N G-monomethyl-l-arginine (l-NMMA) treatment during normoxia in both blood- and buffer-perfused rabbit lungs. Exhaled NO (eNO) was unaltered by changing perfusate flow in blood-perfused lungs. In buffer-perfused lungs, bolus injections of ACh into the pulmonary artery evoked a transient increase in eNO from 67 ± 3 (SE) to 83 ± 7 parts/billion with decrease in Ppa, whereas perfusate NO metabolites (pNOx) remained unchanged. Stepwise increments in flow from 25 to 150 ml/min caused corresponding stepwise elevations in eNO production (46 ± 2 to 73 ± 3 nl/min) without changes in pNOx during normoxia. Despite a reduction in the baseline level of eNO, flow-dependent increases in eNO were still observed during hypoxia.l-NMMA caused declines in both eNO and pNOx with a rise in Ppa. Pulmonary vascular conductance progressively increased with increasing flow during normoxia and hypoxia. However,l-NMMA blocked the flow-dependent increase in conductance over the range of 50–150 ml/min of flow. In the more physiological conditions of blood perfusion, eNO does not reflect endothelial NO production. However, from the buffer perfusion study, we suggest that endothelial NO production secondary to increasing flow, may contribute to capillary recruitment and/or shear stress-induced vasodilation.

Flow Monitoring in Microdialysis Systems for Continuous Sampling

Continuous in vivo sampling of medically relevant substances using the microdialysis method requires the most exact possible knowledge about the involved perfusate flow. As this flow is typically in the range of few microliters per hour a flow sensor with a resolution of less than 1 microliter per hour is required. This paper will present such a sensor based on a calorimetrical measurement. The described principle of operation allows not only the flow measurement itself but also an additional detection of the involved fluids thermal characteristics, allowing the compensation of important interferents in this and similar micro fluidic applications, e.g. gas enclosures.

Effects of adenosine receptor antagonists on the responses to contrast media in the isolated rat kidney

Contrast media can induce both a decrease in renal blood flow and a reduction in glomerular filtration rate (GFR) when administered to both experimental animals and humans. In the present study we have examined the role of adenosine in mediating these effects using the isolated perfused rat kidney. Kidneys were perfused with a 6.7%-(w/v)-albumin-based perfusate supplemented with glucose and amino acids (n = 6 per group). They were exposed to diatrizoate [20 mg of iodine (mgI)/ml; osmolality 1650 mOsm/kg of water] or iotrolan (20 mgI/ml; osmolality 320 mOsm/kg of water) in the presence or absence of theophylline (10.8 µg/ml), or to diatrizoate in the presence or absence of a specific adenosine A1 receptor antagonist (KW-3902; 2 µg/ml) or a specific A2 receptor antagonist (KF17837; 6 µg/ml). Diatrizoate (n = 6) produced a fall in GFR from 0.65±0.04 to 0.42±0.03 ml·min-1·g-1 (P < 0.05); renal perfusate flow (RPF) also declined, from 36.5±3.8 to 22.0±3.2 ml·min-1·g-1 (P < 0.05). Iotrolan (n = 6) produced a fall in GFR from 0.64±0.02 to 0.48±0.04 ml·min-1·g-1 (P < 0.05) and in RPF from 33.3±3.8 to 24.0±3.0 ml·min-1·g-1 (P < 0.05). Theophylline (10.8 µg/ml) prevented the fall in GFR caused by either diatrizoate (baseline, 0.63±0.05 ml·min-1·g-1; diatrizoate+theophylline, 0.60±0.04 ml·min-1·g-1) or iotrolan (basline, 0.64±0.04 ml·min-1·g-1; iotrolan+theophylline, 0.67±0.05 ml·min-1·g-1), but did not affect the decreases in RPF caused by either agent. KW-3902 (2 µg/ml) also prevented the fall in GFR produced by diatrizoate (baseline, 0.66±0.05 ml·min-1·g-1; diatrizoate+KW-3902, 0.61±0.05 ml·min-1·g-1), while the fall in RPF remained unaffected. KF17837 (6 µg/ml) had no effect on the decreases in either GFR or RPF induced by diatrizoate (n = 6 per group). The results suggest a role for adenosine acting at the A1 receptor in mediating the decrease in GFR induced by contrast media. This effect is independent of a change in renal vascular resistance, and possibly secondary to mesangial cell contraction causing a decrease in the ultrafiltration coefficient.

Flow reduces the amplitude and increases the frequency of lymphatic vasomotion: role of endothelial prostanoids

Fluid dynamic forces have substantial effects on the movement of lymph and activity of lymph vessels. The effect of increases in intraluminal flow on spontaneous pumping activity of isolated collecting lymphatics has not yet been characterized in a condition in which the intraluminal pressure is constant. Thus, in afferent lymph microvessels isolated from rat iliac lymph nodes, changes in maximum (Dmax) and minimum (Dmin) diameter to increases in perfusate flow were investigated in the presence of a constant perfusion pressure of 6 cmH2O. Intraluminal flow was elicited by increases in the difference between outflow and inflow pressures (Pdiff, from 0 to 6 cmH2O). Diameters were measured by videomicroscopy. In response to increases in perfusate flow, Dmax and Dmin of lymphatics decreased from 157.5 ± 6.1 to 90.9 ± 5.6 μm and from 91.9 ± 5.3 to 66.3 ± 3.6 μm, respectively, whereas vasomotion frequency increased from 18.0 ± 0.7 min−1 to 23.4 ± 1.1 min−1 (at Pdiff of 4 cmH2O). Removal of extracellular Ca2+ abolished spontaneous diameter oscillations; under these conditions the passive diameter of lymphatics was 216.0 ± 7.1 μm and did not change in response to increases in perfusion. In the absence of endothelium, flow-induced changes in Dmax, Dmin, and oscillation frequency were eliminated. N ω-nitro-l-arginine methyl ester, an inhibitor of nitric oxide synthase, did not affect flow-induced changes in diameter of lymphatics. In contrast, indomethacin, an inhibitor of prostaglandin synthesis, or SQ-29,548, a PGH2/thromboxane A2(PGH2/TxA2) receptor blocker, inhibited the perfusion-induced reduction of Dmax and Dmin of lymphatics and also the increase in the frequency of vasomotion. These findings suggest that the sensitivity of lymphatic endothelium to increases in intraluminal flow could provide an important local intrinsic mechanism for the control of lymphatic resistance by release of constrictor prostanoids PGH2/TxA2.

Microdialysis of skeletal muscle at rest

Techniques in human skeletal muscle research are by necessity predominantly 'descriptive'.Microdialysis has raised high expectations that it could meet the demand for a method that allows 'mechanistic' investigations to be performed in human skeletal muscle. In the present review, some views are given on how well the initial expectations on the use of the microdialysis technique in skeletal muscle have been fulfilled, and the areas in which additional work is needed in order to validate microdialysis as an important metabolic technique in this tissue. The microdialysis catheter has been equated to an artificial blood vessel, which is introduced into the tissue. By means of this 'vessel' the concentrations of compounds in the interstitial space can be monitored. The concentration of substances in the collected samples is dependent on the rate of perfusate flow. When perfusate flow is slow enough to allow complete equilibration between interstitial and perfusate fluids, the concentration in the perfusate is maximal and identical to the interstitial concentration. Microdialysis data may be influenced by changes in blood flow, especially in instances where the tissue diffusivity limits the recovery in vivo, i.e. when recovery in vitro is 100 %, whereas the recovery in vivo is less than 100 %. Microdialysis data indicate that a significant arterial-interstitial glucose concentration gradient exists in skeletal muscle but not in adipose tissue at rest. While the concentrations of glucose and lactate in the dialysate from skeletal muscle are close to the expected values, the glycerol values obtained for muscle are still puzzling. Ethanol added to the perfusate will be cleared by the tissue at a rate that is determined by the nutritive blood flow (the microdialysis ethanol technique). It is concluded that microdialysis of skeletal muscle has become an important technique for mechanistic studies in human metabolism and nutrition.