Comparison of amounts of collagen and elastin in pleura and parenchyma of dog lung

Pressure-volume characteristics of the lung have been thought to be due primarily to the properties of the network of alveolar septa. However, Hajji et al. (J. Appl. Physiol.: Respirat . Environ. Exercise Physiol. 47: 175-181, 1979) attributed a substantial role to the visceral pleura. Seeking a structural explanation for this result, we compared the relative amounts of collagen fibrils and elastin fibers in the visceral pleura and alveolar parenchyma using stereological measurements in five canine lobes. We found about one-fifth as much collagen and one-tenth as much elastin in the pleura as in the alveolar parenchyma. This structural result confirms the functional conclusions of Hajji et al. We argue that such a substantial structure is not needed for protection against overinflation but may have to do with stabilization of lobe shape or handling of frictional forces.     

The epithelial Na(+) channel (ENaC) is related to the hypertonicity-induced Na(+) conductance in rat hepatocytes

The epithelial Na(+) channel (ENaC) is composed of the subunits alpha, beta, and gamma [Canessa et al., Nature 367 (1994) 463-467] and typically exhibits a high affinity to amiloride [Canessa et al., Nature 361 (1993) 467-470]. When expressed in Xenopus oocytes, conflicting results were reported concerning the osmo-sensitivity of the channel [Ji et al., Am. J. Physiol. 275 (1998) C1182-C1190; Hawayda and Subramanyam, J. Gen. Physiol. 112 (1998) 97-111; Rossier, J. Gen. Physiol. 112 (1998) 95-96]. Rat hepatocytes were the first system in which amiloride-sensitive sodium currents in response to hypertonic stress were reported [Wehner et al., J. Gen. Physiol. 105 (1995) 507-535; Wehner et al., Physiologist 40 (1997) A-4]. Moreover, all three ENaC subunits are expressed in these cells [B?hmer et al., Cell. Physiol. Biochem. 10 (2000) 187-194]. Here, we injected specific antisense oligonucleotides directed against alpha-rENaC into single rat hepatocytes in confluent primary culture and found an inhibition of hypertonicity-induced Na(+) currents by 70%. This is the first direct evidence for a role of the ENaC in cell volume regulation.     

Modeling of respiratory system impedances in dogs

Mechanical impedances between 4 and 64 Hz of the respiratory system in dogs have been reported (A.C. Jackson et al. J. Appl. Physiol. 57: 34-39, 1984) previously by this laboratory. It was observed that resistance (the real part of impedance) decreased slightly with frequency between 4 and 22 Hz then increased considerably with frequency above 22 Hz. In the current study, these impedance data were analyzed using nonlinear regression analysis incorporating several different lumped linear element models. The five-element model of Eyles and Pimmel (IEEE Trans. Biomed. Eng. 28: 313-317, 1981) could only fit data where resistance decreased with frequency. However, when the model was applied to these data the returned parameter estimates were not physiologically realistic. Over the entire frequency range, a significantly improved fit was obtained with the six-element model of DuBois et al. (J. Appl. Physiol. 8: 587-594, 1956), since it could follow the predominate frequency-dependent characteristic that was the increase in resistance. The resulting parameter estimates suggested that the shunt compliance represents alveolar gas compressibility, the central branch represents airways, and the peripheral branch represents lung and chest wall tissues. This six-element model could not fit, with the same set of parameter values, both the frequency-dependent decrease in Rrs and the frequency-dependent increase in resistance. A nine-element model recently proposed by Peslin et al. (J. Appl. Physiol. 39: 523-534, 1975) was capable of fitting both the frequency-dependent decrease and the frequency-dependent increase in resistance. However, the data only between 4 and 64 Hz was not sufficient to consistently determine unique values for all nine parameters.     

Inhomogeneity during deflation of excised canine lungs. I. Alveolar pressures

Factors both intrinsic and extrinsic to the lung may cause inhomogeneity of alveolar pressures during deflation. Wilson et al. (J. Appl. Physiol. 59: 1924-1928, 1985) predicted that any such inhomogeneity would be limited by interdependence of regional expiratory flows. To test this hypothesis and to explore how the pleural pressure gradient might affect inhomogeneity of alveolar pressures, we deflated at submaximal flows excised canine lobes that first were suspended in air and then were immersed in foams that simulated the vertical gradient of pleural pressure. Interregional inhomogeneity of regional transpulmonary pressures was measured with use of an alveolar capsule technique. Flow-dependent inhomogeneity of alveolar pressures was present, with differences in alveolar pressure quickly relaxing to a constant limiting value at each flow. Foam immersion increased inhomogeneity at a given flow. We conclude that factors intrinsic to the lung cause significant inhomogeneity of alveolar pressures at submaximal expiratory flows and that this inhomogeneity is enhanced by the extrinsic gradient of pleural pressure. These observations are consistent with the interdependence of flow proposed by Wilson et al.     

Alveolar pressure in fluid-filled occluded lung segments during permeability edema

In a model of increased hydrostatic pressure pulmonary edema Parker et al. (J. Appl. Physiol.: Respirat. Environ. Exercise Physiol. 44: 267-276, 1978) demonstrated that alveolar pressure in occluded fluid-filled lung segments was determined primarily by interstitial fluid pressure. Alveolar pressure was subatmospheric at base line and rose with time as hydrostatic pressure was increased and pulmonary edema developed. To further test the hypothesis that fluid-filled alveolar pressure is determined by interstitial pressure we produced permeability pulmonary edema-constant hydrostatic pressure. After intravenous injection of oleic acid in dogs (0.01 mg/kg) the alveolar pressure rose from -6.85 +/- 0.8 to +4.60 +/- 2.28 Torr (P less than 0.001) after 1 h and +6.68 +/- 2.67 Torr (P less than 0.01) after 3 h. This rise in alveolar fluid pressure coincided with the onset of pulmonary edema. Our experiments demonstrate that during permeability pulmonary edema with constant capillary hydrostatic pressures, as with hemodynamic edema, alveolar pressure of fluid-filled segments seems to be determined by interstitial pressures.     

Low-frequency respiratory system resistance in the normal dog during mechanical ventilation

The low-frequency resistances of the respiratory system, lung, and chest wall were investigated in four anesthetized paralyzed dogs mechanically ventilated at various frequencies between 0.08 and 0.83 Hz. The resistances were calculated by three different methods: 1) as the real part of the complex impedance obtained from regular ventilation data, 2) as the effective resistance of a two-compartment model fitted to the same data, and 3) as the resistance of a single-compartment model fitted to data obtained during sinusoidal ventilation at various frequencies. Alveolar pressures were measured by a closed-chest alveolar capsule technique and afforded a direct measure of airways resistance. All three resistance estimates were very similar and decreased markedly with frequency between 0 and 1 Hz. The real part of lung impedance at the higher frequencies investigated (around 5 Hz) closely approximated airways resistance, as predicted by the eight-parameter viscoelastic model of respiratory mechanics proposed by Bates et al. (J. Appl. Physiol. 67:2276-2285, 1989).     

A model evaluation of estimates of breath-to-breath alveolar gas exchange

A mathematical model has been implemented for evaluation of methods for estimating breath-to-breath alveolar gas exchange during exercise in humans. This model includes a homogeneous alveolar gas exchange compartment, a dead space compartment, and tissue spaces for CO2 (alveolar and dead space). The dead space compartment includes a mixing portion surrounded by tissue and an unmixed (slug flow) portion which is partitioned between anatomical and apparatus contributions. A random sinusoidal flow pattern generates a breath-to-breath variation in pulmonary stores. The Auchincloss algorithm for estimating alveolar gas exchange (Auchincloss et al., J. Appl. Physiol. 21: 810-818, 1966) was applied to the model, and the results were compared with the simulated gas exchange. This comparison indicates that a compensation for changes in pulmonary stores must include factors for alveolar gas concentration change as well as alveolar volume change and thus implies the use of end-tidal measurements. Although this algorithm yields reasonable estimates of breath-to-breath alveolar gas exchange, it does not yield a "true" indirect measurement because of inherent error in the estimation of a homogeneous alveolar gas concentration at the end of expiration.     

Evidence for Na+–glucose cotransporter in type I alveolar epithelium

Functional evidence of Na⁺–glucose cotransport in rat lung has been provided by Basset et al. (J. Physiol. 384:325–345, 1987). By autoradiography [³H]phloridzin binding has been found confined to alveolar epithelial type II cells in mouse and rabbit lungs (Boyd, J. Physiol. 422: 44P, 1990). In this research we checked by immunofluorescence whether Na⁺–glucose cotransporter (SGLT1) is also expressed in alveolar type I cells. Lungs of anesthetized rats and lambs were fixed by paraformaldehyde, perfused in pulmonary artery, or instilled into a bronchus, respectively. Tissue blocks embedded in paraffin or frozen were sectioned. Two specific anti-SGLT1 antibodies for rat recognizing aminoacid sequence 402–420, and 546–596 were used in both species. Bound primary antibody was detected by secondary antibody conjugated to fluorescein isothiocianate or Texas red, respectively. In some sections cellular nuclei were also stained. In rats alveolar type I cells were identified by fluorescent Erythrina cristagalli lectin. Sections were examined by confocal laser-scanning microscope. Both in rats and lambs alveolar epithelium was stained by either antibody; no labeling occurred in negative controls. Hence, SGLT1 appears to be also expressed in alveolar type I cells. This is functionally relevant because type I cells provide 95–97% of alveolar surface, and SGLT1, besides contributing to removal of lung liquid under some circumstances, keeps low glucose concentration in lining liquid, which is useful to prevent lung infection.     

20-hydroxyeicosatetraenoic acid (20-HETE): structural determinants for renal vasoconstriction

The effects of natural and synthetic eicosanoids on the diameter of rat interlobular arteries studied in vitro were compared to that of the potent, endogenous vasoconstrictor 20-HETE. Vasoconstrictor activity was optimum for chain lengths of 20-22 carbons with at least one olefin or epoxide between located between C(13)-C(15) and an oxygen substituent at C(20)-C(22). The presence of delta (Zou et al. Am. J. Physiol. 1996, 270, R228; Gebremedhin, D. et al. Am. J. Physiol. 1998, 507, 771)-, delta (Carroll et al. Am. J. Physiol. 1996, 271, R863; Vazquez et al. Life Sci. 1995, 56, 1455)-, or delta (Imig et al. Hypertension 2000, 35, 307; Lopez et al. Amer. J. Physiol. 2001, 281, F420)-olefins had no influence on the vasoconstrictor response whereas the introduction of a C(7)-thiomethylene enhanced potency. A sulfonamide or alcohol, but not a lactone, could replace the C(1)-carboxylate. These data were used to construct a putative binding domain map of the 20-HETE receptor consisting of: (i) a comparatively open, hydrophilic binding site accommodating the C(1)-functionality; (ii) a hydrophobic trough spanning the olefins; (iii) a shallow pocket containing a critical pi-pi binding site in the vicinity of the pi (Ito et al. Am. J. Physiol. 1998, 274, F395; Quigley, R.; Baum, M.; Reddy, K. M.; Griener, J. C.; Falck, J. R. Am. J. Physiol. 2000, 278, F949)-olefin; and (iv) an oxyphilic binding site proximate to the omega-terminus.     

A model of the single atrial cell: relation between calcium current and calcium release

The hypothesis that calcium release from the sarcoplasmic reticulum in cardiac muscle is induced by rises in free cytosolic calcium (Fabiato 1983, Am. J. Physiol 245) allows the possibility that the release could be at least partly regenerative. There would then be a non-linear relation between calcium current and calcium release. We have investigated this possibility in a single-cell version of the rabbit-atrial model developed by Hilgemann & Noble (1987, Proc. R. Soc. Lond. B 230). The model predicts different voltage ranges of activation for calcium-dependent processes (like the sodium-calcium exchange current, contraction or Fura-2 signals) and the calcium current, in agreement with the experimental results obtained by Earm et al. (1990, Proc. R. Soc. Lond. B 240) on exchange current tails, Cannell et al. (1987, Science, Wash. 238) by using Fura-2 signals, and Fedida et al. (1987, J. Physiol., Lond. 385) and Talo et al. (1988, Biology of isolated adult cardiac myocytes) by using contraction. However, when the Fura-2 concentration is sufficiently high (greater than 200 microM) the activation ranges become very similar as the buffering properties of Fura-2 are sufficient to remove the regenerative effect. It is therefore important to allow for the buffering properties of calcium indicators when investigating the correlation between calcium current and calcium release.     

Multiple Forms and Distribution of Calcium/Calmodulin-Stimulated Protein Kinase II in Brain

In recent years, the enzyme Ca²⁺/calmodulin-stimulated protein kinase II¹ (CaM-PK II) as attracted a great deal of interest. CaM-PK II is the most abundant calmodulin-stimulated protein kinase in brain, where it is particularly enriched in neurons (Ouimet et al., 1984; Erondu and Kennedy, 1985; Lin et al., 1987; Scholz et al., 1988). Neuronal CaM-PK II has been suggested to be involved in several phenomena associated with synaptic plasticity (Lisman and Goldring, 1988; Kelly, 1992), including long-term potentiation (Malinow et al., 1988; Malenka et al.,1989), neurotransmission (Nichols et al., 1990; Siekevitz, 1991), and learning (for review, see Rostas, 1991). This enzyme has also been postulated to be selectively vulnerable in several pathological condition, including epilepsy/kindling (Bronstein et al.,1990; Wu et al., 1990), cerebral ischemia (Taft et al., 1988), and organophosphorus toxicity (Abou-Donia and Lapadula, 1990).     

Connexons and cell adhesion: a romantic phase

Recent evidence indicates, that gap junction forming proteins do not only contribute to intercellular communication (Kanno and Saffitz in Cardiovasc Pathol 10:169-177, 2001; Saez et al. in Physiol Rev 83:1359-1400, 2003), ion homeostasis and volume control (Goldberg et al. in J Biol Chem 277:36725-36730, 2002; Saez et al. in Physiol Rev 83:1359-1400, 2003). They also serve biological functions in a mechanical sense, supporting adherent connections between neighbouring cells of epithelial and non-epithelial tissues (Clair et al. in Exp Cell Res 314:1250-1265, 2008; Shaw et al. in Cell 128:547-560, 2007), where they stabilize migratory pathways in the developing central nervous system (Elias et al. in Nature 448:901-907, 2007; Malatesta et al. in Development 127:5253-5263, 2000; Noctor et al. in Nature 409:714-720, 2001; Rakic in Brain Res 33:471-476, 1971; J Comp Neurol 145:61-83 1972; Science 241:170-176, 1988), or mediate polarized movements and directionality of neural crest cells during organogenesis (Kirby and Waldo in Circ Res 77:211-215, 1995; Xu et al. in Development 133:3629-3639, 2006). Since, most data describing adhesive properties of gap junctions delt with connexin 43 (Cx43) (Beardslee et al. in Circ Res 83:629-635, 1998), we will focus our brief review on this isoform.     

Inhomogeneity during deflation of excised canine lungs. II. Alveolar volumes

We have previously demonstrated appreciable inhomogeneity of alveolar pressures measured by a capsule technique in excised canine lobes deflated at submaximal flows (J. Appl. Physiol. 65: 1757-1765, 1988). We further analyzed the results of these experiments by estimating alveolar volumes (VA) and regional flows from regional transpulmonary pressures, assuming that regional pressure-volume relationships were homogeneous. Deflation at submaximal flows of lungs suspended in air caused significant flow-dependent inhomogeneity of VA that increased as lung volume decreased. Immersion of lungs in stable foams that simulated the gradient of pleural pressure modified the pattern of emptying, but not always to a gravity-dependent sequence. Limitation of regional expiratory flow was often asynchronous during both air suspension and foam immersion. There was no evidence of a common regional flow-volume curve. Submaximal deflation is a complex heterogeneous process, with the interregional pattern of emptying determined by the interaction of factors that are both intrinsic and extrinsic to the lungs.     

Geometric hysteresis of alveolated ductal architecture

Low Reynolds number airflow in the pulmonary acinus and aerosol particle kinetics therein are significantly conditioned by the nature of the tidal motion of alveolar duct geometry. At least two components of the ductal structure are known to exhibit stress-strain hysteresis: smooth muscle within the alveolar entrance rings, and surfactant at the air-tissue interface. We hypothesize that the geometric hysteresis of the alveolar duct is largely determined by the interaction of the amount of smooth muscle and connective tissue in ductal rings, septal tissue properties, and surface tension-surface area characteristics of surfactant. To test this hypothesis, we have extended the well-known structural model of the alveolar duct by Wilson and Bachofen (1982, "A Model for Mechanical Structure of the Alveolar Duct," J. Appl. Physiol. 52(4), pp. 1064-1070) by adding realistic elastic and hysteretic properties of (1) the alveolar entrance ring, (2) septal tissue, and (3) surfactant. With realistic values for tissue and surface properties, we conclude that: (1) there is a significant, and underappreciated, amount of geometric hysteresis in alveolar ductal architecture; and (2) the contribution of smooth muscle and surfactant to geometric hysteresis are of opposite senses, tending toward cancellation. Quantitatively, the geometric hysteresis found experimentally by Miki et al. (1993, "Geometric Hysteresis in Pulmonary Surface-to-Volume Ratio during Tidal Breathing," J. Appl. Physiol. 75(4), pp. 1630-1636) is consistent with little or no smooth muscle tone in anesthetized rabbits in control conditions, and with substantial smooth muscle activation following methacholine challenge. The observed local hysteretic boundary motion of the acinar duct would result in irreversible acinar flow fields, which might be important mechanistic contributors to aerosol mixing and deposition deep in the lung.     

Error due to dead-space admixture in single-breath method for estimation of pulmonary blood flow

The single-breath method of Kim et al. (J. Appl. Physiol. 21: 1338-1344, 1966) for the estimation of pulmonary blood flow is based on a single-alveolus lung model for which an analytical relationship has been established between the kinetic behavior of the alveolar O2 and CO2 tensions and the pulmonary blood flow. The analysis is based on the assumption that the dead-space contribution to the expirate is negligible after expiration of a predefined volume. We have examined the influence of this assumption on the estimation of pulmonary blood flow by computer simulation in a lung model that incorporates deadspace contribution to the expirate. Data on the fractional contribution of the dead space to the expired gas were obtained from Tsunoda et al.'s study (J. Appl. Physiol. 32: 644-649, 1972) on the emptying pattern of normal adult lungs. The results show that failure to take account of the dead-space contribution can cause an underestimation in the pulmonary blood flow of greater than 30%. The error can be reduced by ignoring the first part of the expiration but only at the cost of an increase in the sensitivity of the single-breath method to measurement noise. This property of the system is demonstrated experimentally. The error due to dead-space admixture depends on the total volume of dead-space gas, the measurement noise, the pulmonary blood flow, and the emptying characteristics of the dead-space compartment during expiration. In normal subjects it is possible to optimize the experimental design so that the systematic error is less than 5% and the coefficient of variation is less than 10%.(ABSTRACT TRUNCATED AT 250 WORDS)     

The significance of reutilization of surfactant phosphatidylcholine

To assess the magnitude of reutilization of surfactant phosphatidylcholine, 68 3-day-old rabbits were injected intratracheally with a trace dose of [3H]choline-labeled surfactant mixed with [14C]palmitate-labeled synthetic dipalmitoylphosphatidylcholine. After timed kills we measured the total phosphatidylcholine associated counts/min in whole lung and alveolar wash and the specific activities of phosphatidylcholine in the alveolar wash, lamellar bodies, and microsomes isolated from the lung of each rabbit. Using a modification of the compartment analysis of Skinner et al. (Skinner, S. M., Clark, R. E., Baker, N., and Shipley, R. A. (1959) Am. J. Physiol. 196, 238-244), we found that surfactant phosphatidylcholine was reutilized with greater than 90% efficiency. The turnover time of the alveolar wash phosphatidylcholine was estimated to be 10.1 h and 9.3 h as measured by the 3H and 14C labels, respectively. From the ratios of alveolar wash-associated natural to synthetic phosphatidylcholine specific activities and from similar ratios obtained in 30 additional rabbits using [14C]choline-labeled natural surfactant and [3H]choline-labeled dipalmitoylphosphatidylcholine, we showed that phosphatidylcholine was reutilized intact rather than as component parts. Within 6 h of injection, the synthetic dipalmitoylphosphatidylcholine functioned metabolically as that administered in the form of natural surfactant.     

Pulmonary alveolar macrophages express a polyamine transport system

Polyamine transport is an important mechanism by which cells regulate their intracellular polyamine content. It is well established that the lung has a high capacity for polyamine transport, and recently the polyamine putrescine has been shown to be selectively accumulated into the type II pneumocyte of rabbit lung slices (Saunders et al.: Lab. Invest., 95:380-386, 1988). In addition, it has been suggested that there may be more than one polyamine transport system in lung tissue (Byers et al.: Am. J. Physiol., 252:C663-C669, 1987). In the present study, we have examined whether there are differences in the distribution of putrescine and spermidine uptake activities in isolated rabbit lung cells. We report that pulmonary alveolar macrophages have a greater rate of uptake of both putrescine and spermidine than the total lung cell population. Kinetic analysis of the polyamine uptake system present in macrophages showed putrescine uptake consisted of a saturable (Km = 2.1 microM) and nonsaturable component whilst spermidine uptake consisted of both a high- and a low-capacity saturable component (Km = 0.16 microM and 1.97 microM, respectively). The rate of polyamine transport was similar to those reported for many proliferative or tumor cell-lines and appears to be greater than any other major lung cell type. Inhibition studies of the transport of polyamines into pulmonary alveolar macrophages suggested that the uptake of both putrescine and spermidine was mediated by the same system, which could not be described by simple Michaelis-Menten kinetics. The transport appears to be reversible due to significant efflux. This is the first study to describe the presence of multiple polyamine transport systems in pulmonary alveolar macrophages.     

Breath-by-breath measurement of the volume displaced by diaphragm motion.

To develop an accurate method to measure the volume displaced by diaphragm motion (DeltaVdi) breath by breath, we compared DeltaVdi measured by a previously evaluated biplanar radiographic method (Singh B, Eastwood PR, and Finucane KE. J Appl Physiol 91: 1913-1923, 2001) at several lung volumes during vital capacity inspirations in 10 healthy and nine hyperinflated subjects with 1) DeltaVdi measured from the same chest X-rays by two previously described uniplanar methods (Petroll WM, Knight H, and Rochester DF. J Appl Physiol 69: 2175-2182, 1990; Verschakelen JA, Deschepper K, and Demendts M. J Appl Physiol 72: 1536-1540, 1992) and a proposed method that considered actual cross-sectional shape of the rib cage and spinal volume (DeltaVdi(S)); and 2) DeltaVdi(S) measured by lateral fluoroscopy in the same 10 healthy subjects. Relative to biplanar DeltaVdi, DeltaVdi(S) values from lateral chest X-rays and fluoroscopy were not different, whereas DeltaVdi values of Petroll et al. and Verschakelen et al. were increased by (means +/- SD) 1.98 +/- 1.59 and 1.16 +/- 0.82 liters, respectively (both P < 0.001). During quiet breathing, DeltaVdi(S) by lateral fluoroscopy was 66 +/- 16% of tidal volume and similar to that between functional residual capacity and one-half inspiratory capacity by the biplanar radiographic method. We conclude that accurate breath-by-breath measurements of DeltaVdi can be made by using lateral fluoroscopy.     

Thoracic gas volume in rabbits by low-frequency ambient pressure changes

R. Peslin et al. measured thoracic gas volume (TGV) in adults using a new method employing low-frequency ambient pressure changes (APC) (J. Appl. Physiol. 62: 359-363, 1987). We extended that methodology and then tested the hypothesis that this technique was applicable to small mammals. TGV [at functional residual capacity (FRC)] by APC and by conventional Boyle's law was compared in 12 rabbits. The rabbits were anesthetized, tracheostomized, intubated, and placed in a pressure plethysmograph. Although in the method of Peslin et al. box pressure was oscillated at a single frequency, in our extension box pressure was oscillated simultaneously at two frequencies (0.1 and 0.2 Hz). Flow at the airway opening consisted of rapid events due to spontaneous breathing, superposed on slower events due to the alveolar gas compression. The slower events were analyzed to yield alveolar gas compliance and, by Boyle's law, FRC. FRC by APC was highly correlated to FRC by conventional plethysmography (r = 0.85). Compared with the methodology of Peslin et al., our extension relaxes a key limitation and yields systematically higher estimates of FRC. We conclude that this method is applicable to small mammals, despite an inherently more compliant chest wall, and that the methodological extension improves the estimate of FRC.