Mechanical properties of human bronchial smooth muscle in vitro.

The in vitro mechanical properties of smooth muscle strips from 10 human main stem bronchi obtained immediately after pneumonectomy were evaluated. Maximal active isometric and isotonic responses were obtained at varying lengths by use of electrical field stimulation (EFS). At the length (Lmax) producing maximal force (Pmax), resting tension was very high (60.0 +/- 8.8% Pmax). Maximal fractional muscle shortening was 25.0 +/- 9.0% at a length of 75% Lmax, whereas less shortening occurred at Lmax (12.2 +/- 2.7%). The addition of increasing elastic loads produced an exponential decrease in the shortening and velocity of shortening but increased tension generation of muscle strips stimulated by EFS. Morphometric analysis revealed that muscle accounted for 8.7 +/- 1.5% of the total cross-sectional tissue area. Evaluation of two human tracheal smooth muscle preparations revealed mechanics similar to the bronchial preparations. Passive tension at Lmax was 10-fold greater and maximal active shortening was threefold less than that previously demonstrated for porcine trachealis by us of the same apparatus. We attribute the limited shortening of human bronchial and tracheal smooth muscle to the larger load presumably provided by a connective tissue parallel elastic component within the evaluated tissues, which must be overcome for shortening to occur. We suggest that a decrease in airway wall elastance could increase smooth muscle shortening, leading to excessive responses to contractile agonists, as seen in airway hyperresponsiveness.     

Different ontogeny of rate of force generation and shortening velocity in guinea pig trachealis.

Juveniles of many species, including humans, display greater airway responsiveness than do adults. This may involve changes in airway smooth muscle function. In the present work we studied force production and shortening velocity in trachealis from 1-wk-old (1 wk), 3-wk-old (3 wk), and 3-mo-old (adult) guinea pigs. Strips were electrically stimulated (60 Hz, 18 V) at their optimal length (l(o)) to obtain maximum active stress (P(o)) and rate of stress generation. Then, force-velocity curves were elicited at 2.5 s from the onset of the stimulus. By applying a recently developed modification of Hill's equation for airway smooth muscle, the maximum shortening velocity at zero load (V(o)) and the value alpha. gamma/beta, an index of internal resistance to shortening (Rsi), were calculated (alpha, beta, and gamma are the constants of the equation). P(o) increased little with maturation, whereas the rate of stress generation increased significantly (0.40 +/- 0.03, 0.45 +/- 0.03, 0. 51 +/- 0.03 P(o)/s for 1 wk, 3 wk, and adult animals). V(o) slightly increased early with maturation to decrease significantly later (1. 79 +/- 0.67, 2.45 +/- 0.92, and 0.55 +/- 0.09 l(o)/s for 1 wk, 3 wk, and adult animals), whereas the Rsi showed an opposite trend (14.98 +/- 5.19, 8.99 +/- 3.01, and 32.07 +/- 5.54 mN. mm(-2). l(o)(-1). s for 1 wk, 3 wk, and adult animals). This early increase of force generation in combination with late increase of Rsi may explain the changes of V(o) with age. An elevated V(o) may contribute to the incidence of airway hyperresponsiveness in healthy juveniles.     

Relaxation of guinea pig trachealis during electrical field stimulation increases with age.

Our laboratory has previously shown that maturation of airway smooth muscle (ASM) contractility may play a role in the airway hyperresponsiveness displayed by juveniles of many species, including humans (Chitano P, Wang J, Cox CM, Stephens NL, and Murphy TM. J Appl Physiol 88: 1338-1345, 2000). ASM relaxation, which could also contribute to airway hyperresponsiveness, has neither been described nor quantified during maturation. Therefore, we studied ASM relaxation during and after electrical field stimulation (EFS) in tracheal strips from 1-wk-old, 3-wk-old, and 3-mo-old guinea pigs. Strips were stimulated (60 Hz, 18 V) at their optimal length for 15, 20, and 25 s, with and without the cyclooxygenase inhibitor indomethacin. To evaluate the role of the epithelium, deepithelialized strips from adult animals were also studied. New indexes were developed to quantify relaxation during EFS. We measured the time course of tension relaxation and its maximum rate (RTR) during the EFS, as well as the residual tension at the end of the EFS (TCT(end)). After EFS, we measured the maximum RTR and the time needed to reduce to half the TCT(end). Relaxation during the EFS significantly increased with age. Indomethacin reduced this age difference by increasing relaxation in strips from younger animals. By contrast, removal of the epithelium in adult strips decreased relaxation. Relaxation after EFS decreased with age and was not affected by indomethacin. In adult strips, it was further reduced by epithelium removal. Our results show that during EFS 1) airway smooth muscle relaxation increases with age, 2) cyclooxygenase metabolites oppose relaxation in younger animals, and 3) epithelium removal inhibits relaxation. We suggest that a reduced ASM relaxing ability during stimulation may be involved in juvenile airway hyperresponsiveness.     

Active and passive components in the length-dependent stiffness of tracheal smooth muscle during isotonic shortening.

Contraction of smooth muscle tissue involves interactions between active and passive structures within the cells and in the extracellular matrix. This study focused on a defined mechanical behavior (shortening-dependent stiffness) of canine tracheal smooth muscle tissues to evaluate active and passive contributions to tissue behavior. Two approaches were used. In one, mechanical measurements were made over a range of temperatures to identify those functions whose temperature sensitivity (Q(10)) identified them as either active or passive. Isotonic shortening velocity and rate of isometric force development had high Q(10) values (2.54 and 2.13, respectively); isometric stiffness showed Q(10) values near unity. The shape of the curve relating stiffness to isotonic shortening lengths was unchanged by temperature. In the other approach, muscle contractility was reduced by applying a sudden shortening step during the rise of isometric tension. Control contractions began with the muscle at the stepped length so that properties were measured over comparable length ranges. Under isometric conditions, redeveloped isometric force was reduced, but the ratio between force and stiffness did not change. Under isotonic conditions beginning during force redevelopment at the stepped length, initial shortening velocity and the extent of shortening were reduced, whereas the rate of relaxation was increased. The shape of the curve relating stiffness to isotonic shortening lengths was unchanged, despite the step-induced changes in muscle contractility. Both sets of findings were analyzed in the context of a quasi-structural model describing the shortening-dependent stiffness of lightly loaded tracheal muscle strips.     

Three paradigms of airway smooth muscle hyperresponsiveness in young guinea pigs

Evidence for contributions of airway smooth muscle (ASM) to the hyperresponsiveness of newborn and juvenile airways continues to accumulate. In our laboratory, 3 novel paradigms of hyperresponsiveness of newborn and young ASM have recently emerged using a guinea pig model of maturation in 3 age groups: 1 week (newborn), 3 weeks (juvenile), and 2-3 months (adult). The first paradigm includes evidence for a natural decline after newborn and juvenile life of the velocity of ASM shortening associated with a decrease in regulatory myosin light chain phosphorylation and a parallel decline in the content of myosin light chain kinase. Associated with the decrease in ASM shortening with age is an increase in the internal resistance to shortening. Dynamic stiffness is shown to relate inversely to the expression of myosin light chain kinase. This suggests that developmental changes in shortening relate inversely to the stiffness of the ASM early in shortening, suggesting a dynamic role for the cytoskeleton in facilitating and opposing ASM shortening. This relationship can be approximated as (dP/dt)max approximately (dP/dL)passive x (dL/dt)max (the maximal rate of increase of active stress generation approximately to the passive stiffness x the maximal shortening velocity). The second paradigm demonstrates that newborn ASM, unlike that in adults, does not relax during prolonged electric field stimulation. The impaired relaxation is related to changes in prostanoid synthesis and acetylcholinesterase function. The third paradigm demonstrates that, whereas oscillatory strain serves to transiently relax adult ASM, in newborns it induces (after the initial relaxation) a sustained potentiation of active stress. This is related to developmental changes in the prostanoid release. Together, these paradigms demonstrate that ASM contributes by multiple mechanisms to the natural hyperresponsiveness of newborn and juvenile airways. Future studies will elaborate the mechanisms and extend these paradigms to ASM hyperresponsiveness following sensitization in early life.     

Mechanisms of airway smooth muscle relaxation during maturation

Greater airway responsiveness in healthy juveniles is considered a factor in the higher asthma prevalence at a young age compared with adults. We have developed a guinea pig maturational model that utilizes tracheal strips from 1-week-, 3-week-, and 3-month-old guinea pigs to study the role of airway smooth muscle (ASM) in juvenile airway hyperresponsiveness. Because a reduced ability of ASM to spontaneously relax may contribute to airway hyperresponsiveness by maintaining bronchospasm and thus high airway resistance, we have employed this model to study ASM spontaneous relaxation during electrical field stimulation (EFS). Since relaxation during EFS had been neither described nor quantified during maturation, we developed new indices that allowed an appropriate comparison of the relaxing response from strips of different age animals. Using these indices we found that, whereas strips from adult animals relax to a level of tension similar to that found in the absence of stimulation, this ability to spontaneously relax is essentially absent in trachealis from infant animals. These results confirmed that maturation of ASM relaxation may play a role in juvenile airway hyperresponsiveness and that our maturational model is suitable to study the mechanisms regulating spontaneous relaxation in physiological conditions. We investigated the role of prostanoids in ASM relaxation and showed that cyclooxygenase inhibition increases relaxation in infant ASM to levels similar to adults. These results suggest that prostanoids regulate the ability of ASM to spontaneously relax, i.e., they reduce relaxation. We have produced preliminary data suggesting a maturational change in the level of prostanoids. Moreover, the possible action of acetylcholinesterase on maturation of ASM relaxation is discussed here on the basis of a preliminary study. We suggest that impairment of ASM relaxation likely contributes to increased airway responsiveness.     

Adaptation to chronic length change in explanted airway smooth muscle.

It has been shown that airway smooth muscle in vitro is able to maintain active force over a large length range by adaptation in the absence of periodic stimulations at 4 degrees C (Wang L, Paré PD, and Seow CY. J Appl Physiol 90: 734-740, 2001). In this study, we show that such adaptation also takes place at body temperature and that long-term adaptation results in irreversible functional change in the muscle that could lead to airway hyperresponsiveness. Rabbit tracheal muscle explants were passively maintained at shortened and in situ length for 3 and 7-8 days in culture media; the length-tension relationship was then examined. The length associated with maximal force generation decreased by 10.5 +/- 3.8% (SE) after 3 days and 37.7 +/- 8.5% after 7 or 8 days of passive shortening. At day 3, the left shift in the length-tension curve due to adaptation at short lengths was reversible by readapting the muscle at a longer length. The shift was, however, not completely reversible after 7 days. The results suggest that long-term adaptation of airway smooth muscle could lead to increased muscle stiffness and force-generating ability at short lengths. Under in vivo condition, this could translate into resistance to stretch-induced relaxation and excessive airway narrowing.     

Pulmonary vascular smooth muscle: biochemical and mechanical developmental changes.

To evaluate the developmental changes in pulmonary vascular smooth muscle contractile protein content, mechanical properties, and their contribution to the high resistance characteristic of the fetal and immediate neonatal period, we studied pulmonary vessels of fetal, newborn, and adult sheep, as well as newborn and adult pigs. Strips of the second- through fifth-generation vessels were dissected, and their content of tissue total smooth muscle cell protein, myosin, and actin-to-myosin ratio were measured; the mechanical properties of the second-generation vascular strips were also studied. For all ages the smooth muscle protein and myosin content of the second-generation vessels were significantly greater than for the lower pulmonary vascular orders (P less than 0.05). The myosin content in fetal sheep (0.77 +/- 0.03 micrograms/mg wet tissue) was similar to that of the newborn (0.79 +/- 0.04) and adult (0.86 +/- 0.05). However, the smooth muscle protein content (7.94 +/- 0.21 micrograms/mg wet tissue) and the actin-to-myosin ratio of the pulmonary vascular tissue of the fetus (1.00 +/- 0.04) were lower (P less than 0.01) in the fetal than in the newborn (9.16 +/- 0.26 and 1.60 +/- 0.12) and adult (9.38 +/- 0.3 and 1.60 +/- 0.11, respectively). No differences were observed for these parameters between the newborn and adult pig. Stress (16.5 +/- 1.7 mN/mm2) and the maximum shortening capacity (13.0 +/- 1.5% of optimal length) in the newborn pulmonary vascular strips were significantly greater than for the fetus (6.8 +/- 1.4 and 5.9 +/- 1.0, respectively) but similar to those of the adult sheep.(ABSTRACT TRUNCATED AT 250 WORDS)     

Perfused bronchial segment and bronchial strip: narrowing vs. isometric force by mediators

When bronchial segments were perfused with Krebs solution at a constant pressure (5-6 cmH2O), the resistance rose exponentially with increasing concentrations of either carbachol or histamine in the lumen. The pressure-flow relationship was linear. Histamine and carbachol caused 43 and 47% muscle shortening, respectively, and produced the same maximum effect (Emax) because they both stopped perfusion. In bronchial strips the maximum isometric force or isotonic shortening to carbachol was more than twice that of histamine and the responses showed a plateau. There were no significant differences in sensitivities [negative log of the concentration producing half-maximal response (EC50)] to either carbachol or histamine in the strips (isotonic and isometric) and the segments perfused at constant pressure. When airway segments were perfused at a constant flow, however, responses plateaued and the sensitivities to carbachol and histamine were reduced more than tenfold compared with the strips [4.71 +/- 0.20 and 6.22 +/- 0.08 (SE) for carbachol in segments and isometric strips, respectively, and 3.92 +/- 0.13 and 4.94 +/- 0.11 (SE) for histamine]. We conclude that when segments are perfused at a constant pressure, airway closure occurs before maximal pharmacological activation, as seen in airway strips.     

Tracheal smooth muscle responses to substance P and neurokinin A in the piglet.

The tachykinins substance P (SP) and neurokinin A (NKA) have been shown to induce airway smooth muscle contraction in mature animals, and the enzyme neutral endopeptidase (NEP) modulates this effect. We evaluated maturation of SP- and NKA-induced tracheal smooth muscle contraction and modulation of their effects by NEP in anesthetized, paralyzed, and artificially ventilated piglets less than 4 days, 2-3 wk, and 10 wk of age. Tracheal smooth muscle tension was measured in vivo from an open tracheal segment by use of a force transducer. Intravenous SP caused a dose-dependent increase in tracheal tension in all three age groups; however, the response in less than 4-day-old piglets was significantly weaker than in 2- to 3- and 10-wk-old piglets. NKA caused a dose-dependent increase in tracheal tension only in 2- to 3- and 10-wk-old piglets. The response of tracheal tension to NKA was weaker than the response to SP in all age groups. Atropine (2 mg/kg) significantly diminished the responses of tracheal tension to SP and NKA, indicating a cholinergic contribution to these responses at all ages. Intravenous thiorphan, a known NEP inhibitor, potentiated the effects of SP only in 2- to 3- and 10-wk-old piglets and did not affect the response of tracheal tension to NKA at any age. Biochemical analyses demonstrated a significant increase in tracheal NEP activity in comparably aged piglets over the first 10 wk of life.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bronchial smooth muscle mechanics of a canine model of allergic airway hyperresponsiveness.

Although we have reported that tracheal smooth muscle from sensitized dogs shows altered mechanical properties, we did not know, because of technical difficulties with the preparation, whether similar changes occur in the properties of sensitized central bronchial smooth muscle (BSM), the site at which the acute asthmatic response is believed to develop. We have now succeeded in developing a cartilage-free BSM preparation that retains optimal mechanical properties. Such strips were obtained from mongrel dogs that had been sensitized to ragweed pollen. Controls were littermates injected with adjuvant alone. Length-tension relationships were obtained for both control and sensitized BSM strips (CBSM and SBSM, respectively). The maximal active stresses were the same (P greater than 0.05) when normalized to muscle fraction in total tissue cross-sectional area [6.2 +/- 0.6 x 10(4) and 5.9 +/- 0.6 x 10(4) (SE) for SBSM and CBSM, respectively]. This suggests that optimal tension is an insensitive indicator of bronchial hyperresponsiveness and that isotonic studies might be more revealing. The maximal shortening velocity (Vo) for SBSM at 2 s [0.35 +/- 0.017 (SE) lo/s, where lo signifies optimal muscle length], in the course of a 10-s contraction, was significantly greater (P less than 0.05) than Vo measured for CBSM (0.27 +/- 0.015 lo/s). However, Vo did not differ at the 8-s point of contraction. The sensitized group demonstrated a statistically significantly greater maximal shortening capacity (0.67 +/- 0.04 lo) than the control group (0.51 +/- 0.04 lo). At 2 s of contraction, 80% of maximal SBSM shortening had been completed and was significantly greater than for CBSM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Maturation of respiratory reflex responses in the piglet

Stimulation of chemo-, irritant, and pulmonary C-fiber receptors reflexly constricts airway smooth muscle and alters ventilation in mature animals. These reflex responses of airway smooth muscle have, however, not been clearly characterized during early development. In this study we compared the maturation of reflex pathways regulating airway smooth muscle tone and ventilation in anesthetized, paralyzed, and artificially ventilated 2- to 3- and 10-wk-old piglets. Tracheal smooth muscle tension was measured from an open tracheal segment by use of a force transducer, and phrenic nerve activity was measured from a proximal cut end of the phrenic nerve. Inhalation of 7% CO2 caused a transient increase in tracheal tension in both age groups, whereas hypoxia caused no airway smooth muscle response in either group. The phrenic responses to 7% CO2 and 12% O2 were comparable in both age groups. Lung deflation and capsaicin (20 micrograms/kg iv) administration did not alter tracheal tension in the younger piglets but caused tracheal tension to increase by 87 +/- 28 and 31 +/- 10%, respectively, in the older animals (both P less than 0.05). In contrast, phrenic response to both stimuli was comparable between ages: deflation increased phrenic activity while capsaicin induced neural apnea. Laryngeal stimulation did not increase tracheal tension but induced neural apnea in both age groups. These data demonstrate that between 2 and 10 wk of life, piglets exhibit developmental changes in the reflex responses of airway smooth muscle situated in the larger airways in response to irritant and C-fiber but not chemoreceptor stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mechanical properties of human tracheal cartilage.

Biomechanical changes in airway cartilage could influence the mechanics of maximal expiratory flow and cough and the degree of shortening of activated airway smooth muscle. We examined the tensile stiffness of small samples of human tracheal cartilage rings in specimens obtained at autopsy from 10 individuals who ranged in age from 17 to 81 yr. The tensile properties of the cartilage were compared with its content of water (%water), glycosaminoglycans (chondroitin sulfate equivalents, mg/mg dry wt), and hydroxyproline content (mg hydroxyproline/mg dry weight). The average values for tensile stiffness ranged between 1 and 15 MPa and increased significantly with increasing age [tensile stiffness = 0.19 x (age in yr) + 2.02; r = 0.83, P less than 0.05]. The outermost layer of cartilage was the most stiff in all individuals, and the deeper layers were progressively less stiff. Water content and hydroxyproline content both decreased with increasing age. Thus tensile stiffness correlated inversely with water content and hydroxyproline content [tensile stiffness = -0.83 x (%water) + 16.4; r = 0.82, P less than .05 and tensile stiffness = -342 x (hydroxyproline content) + 25; r = 0.87, P less than 0.05]. Total tissue content of glycosaminoglycans did not change with age, although changes in glycosaminoglycan type and proteoglycan structure with increasing age have been described. We conclude that there are age-related changes in the biomechanical properties and biochemical composition of airway cartilage that could influence airway dynamics.     

Decreased phosphatase activity, increased Ca2+ sensitivity, and myosin light chain phosphorylation in urinary bladder smooth muscle of newborn mice

Developmental changes in the regulation of smooth muscle contraction were examined in urinary bladder smooth muscle from mice. Maximal active stress was lower in newborn tissue compared with adult, and it was correlated with a lower content of actin and myosin. Sensitivity to extracellular Ca2+ during high-K+ contraction, was higher in newborn compared with 3-wk-old and adult bladder strips. Concentrations at half maximal tension (EC50) were 0.57 +/- 0.01, 1.14 +/- 0.12, and 1.31 +/- 0.08 mM. Force of the newborn tissue was inhibited by approximately 45% by the nonmuscle myosin inhibitor Blebbistatin, whereas adult tissue was not affected. The calcium sensitivity in newborn tissue was not affected by Blebbistatin, suggesting that nonmuscle myosin is not a primary cause for increased calcium sensitivity. The relation between intracellular [Ca2+] and force was shifted toward lower [Ca2+] in the newborn bladders. This increased Ca2+ sensitivity was also found in permeabilized muscles (EC50: 6.10 +/- 0.07, 5.77 +/- 0.08, and 5.55 +/- 0.02 pCa units, in newborn, 3-wk-old, and adult tissues). It was associated with an increased myosin light chain phosphorylation and a decreased rate of dephosphorylation. No difference was observed in the myosin light chain phosphorylation rate, whereas the rate of myosin light chain phosphatase-induced relaxation was about twofold slower in the newborn tissue. The decreased rate was associated with a lower expression of the phosphatase regulatory subunit MYPT-1 in newborn tissue. The results show that myosin light chain phosphatase activity can be developmentally regulated in mammalian urinary bladders. The resultant alterations in Ca2+ sensitivity may be of importance for the nervous and myogenic control of the newborn bladders.     

Could an increase in airway smooth muscle shortening velocity cause airway hyperresponsiveness?

Airway hyperresponsiveness (AHR) is a characteristic feature of asthma. It has been proposed that an increase in the shortening velocity of airway smooth muscle (ASM) could contribute to AHR. To address this possibility, we tested whether an increase in the isotonic shortening velocity of ASM is associated with an increase in the rate and total amount of shortening when ASM is subjected to an oscillating load, as occurs during breathing. Experiments were performed in vitro using 27 rat tracheal ASM strips supramaximally stimulated with methacholine. Isotonic velocity at 20% isometric force (Fiso) was measured, and then the load on the muscle was varied sinusoidally (0.33 ± 0.25 Fiso, 1.2 Hz) for 20 min, while muscle length was measured. A large amplitude oscillation was applied every 4 min to simulate a deep breath. We found that: 1) ASM strips with a higher isotonic velocity shortened more quickly during the force oscillations, both initially (P < 0.001) and after the simulated deep breaths (P = 0.002); 2) ASM strips with a higher isotonic velocity exhibited a greater total shortening during the force oscillation protocol (P < 0.005); and 3) the effect of an increase in isotonic velocity was at least comparable in magnitude to the effect of a proportional increase in ASM force-generating capacity. A cross-bridge model showed that an increase in the total amount of shortening with increased isotonic velocity could be explained by a change in either the cycling rate of phosphorylated cross bridges or the rate of myosin light chain phosphorylation. We conclude that, if asthma involves an increase in ASM velocity, this could be an important factor in the associated AHR.     

Echocardiographic assessment of cardiac function in diabetic db/db and transgenic db/db-hGLUT4 mice

Control db/+ and diabetic db/db mice at 6 and 12 wk of age were subjected to echocardiography to determine whether contractile function was reduced in vivo and restored in transgenic db/db-human glucose transporter 4 (hGLUT4) mice (12 wk old) in which cardiac metabolism has been normalized. Systolic function was unchanged in 6-wk-old db/db mice, but fractional shortening and velocity of circumferential fiber shortening were reduced in 12-wk-old db/db mice (43.8 +/- 2.1% and 8.3 +/- 0.5 circs/s, respectively) relative to db/+ control mice (59.5 +/- 2.3% and 11.8 +/- 0.4 circs/s, respectively). Doppler flow measurements were unchanged in 6-wk-old db/db mice. The ratio of E and A transmitral flows was reduced from 3.56 +/- 0.29 in db/+ mice to 2.40 +/- 0.20 in 12-wk-old db/db mice, indicating diastolic dysfunction. Thus a diabetic cardiomyopathy with systolic and diastolic dysfunction was evident in 12-wk-old diabetic db/db mice. Cardiac function was normalized in transgenic db/db-hGLUT4 mice, indicating that altered cardiac metabolism can produce contractile dysfunction in diabetic db/db hearts.     

Length oscillation induces force potentiation in infant guinea pig airway smooth muscle

Deep inspiration counteracts bronchospasm in normal subjects but triggers further bronchoconstriction in hyperresponsive airways. Although the exact mechanisms for this contrary response by normal and hyperresponsive airways are unclear, it has been suggested that the phenomenon is related to changes in force-generating ability of airway smooth muscle after mechanical oscillation. It is known that healthy immature airways of both humans and animals exhibit hyperresponsiveness. We hypothesize that the profile of active force generation after mechanical oscillation changes with maturation and that this change contributes to the expression of airway hyperresponsiveness in juveniles. We examined the effect of an acute sinusoidal length oscillation on the force-generating ability of tracheal smooth muscle from 1 wk, 3 wk, and 2- to 3-mo-old guinea pigs. We found that the length oscillation produced 15-20% initial reduction in active force equally in all age groups. This was followed by a force recovery profile that displayed striking maturation-specific features. Unique to tracheal strips from 1-wk-old animals, active force potentiated beyond the maximal force generated before oscillation. We also found that actin polymerization was required in force recovery and that prostanoids contributed to the maturation-specific force potentiation in immature airway smooth muscle. Our results suggest a potentiated mechanosensitive contractile property of hyperresponsive airway smooth muscle. This can account for further bronchoconstriction triggered by deep inspiration in hyperresponsive airways.     

Expression of airway contractile properties and acetylcholinesterase activity in swine

We studied the effect of maturation on contractile properties of tracheal smooth muscle from seventeen 2-wk-old swine (2ws) and fifteen 10-wk-old swine (10ws) in situ and in vitro. The response to parasympathetic stimulation was studied in situ in isometrically fixed segments. Contraction was elicited at lower frequencies [half-maximal response to electrical stimulation (ES50) = 6.7 +/- 0.05 Hz] in 2ws than in 10ws (ES50 = 9.1 +/- 0.4 Hz; P less than 0.01). Despite substantial differences in morphometrically normalized cross-sectional area in 2ws (0.012 +/- 0.003 cm2) and 10ws (0.028 +/- 0.001 cm2; P less than 0.01), maximal active tension elicited by parasympathetic stimulation was similar (12.4 +/- 3.2 g/cm in 2ws vs. 13.3 +/- 2.3 g/cm in 10ws; P = NS). In separate in vitro studies in 25 tracheal smooth muscle strips from 10 swine, concentration-response curves generated with potassium-substituted Krebs solution (KCl) were similar in 2ws and 10ws. In 58 other strips (10 swine), maximal active force elicited with acetylcholine (ACh) in 2ws was significantly greater than for 10ws (P less than 0.001). Removal of the epithelium had no effect. However, cholinesterase inhibition with 10(-7) M physostigmine augmented the response to ACh in 10ws (P less than 0.02) but not 2ws. We demonstrate increased force generation and sensitivity to vagal stimulation in 2ws vs. 10ws, which corresponds to increased reactivity to ACh in vitro. The relative hyperresponsiveness in 2ws is specific for cholinergic response and is attenuated at least in part by maturation of the activity of acetylcholinesterase enzyme.     

Effect of theophylline on the velocity of diaphragmatic muscle shortening

The present study examined the effect of theophylline on the shortening velocity of submaximally activated diaphragmatic muscle (i.e., muscles were activated by the use of a level of stimulation, 50 Hz, within the range of phrenic neural firing frequencies achieved during breathing, whereas maximum activation is achieved at 300 Hz). Experiments were performed in vitro on strips of diaphragmatic muscle obtained from 21 Syrian hamsters. Muscle shortening velocity was assessed during isotonic contractions against a range of afterloads, and Hill's characteristic equation was used to calculate velocity at zero load. In addition, unloaded shortening velocity was also measured by the slack test, i.e., from the time required for muscles to take up slack after a sudden reduction in muscle length. Theophylline (160 mg/l) increased the velocity of muscle shortening against a wide range of external loads (0-14 N/cm2) and increased the extrapolated unloaded velocity of shortening from 6.4 +/- 0.9 to 7.9 +/- 1.1 (SE) lengths/s (P less than 0.01). Theophylline reduced the time required to take up slack for any given step change in muscle length, increasing the unloaded velocity of shortening assessed by the slack test from 7.6 +/- 0.9 to 9.3 +/- 1.1 lengths/s (P less than 0.002). The effect of theophylline on diaphragmatic shortening velocity was evident at concentrations as low as 40 mg/l and increased progressively as theophylline concentrations were increased to 320 mg/l. Theophylline increased the shortening velocity of fatigued as well as fresh muscles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Increased in vitro airway responsiveness in sheep following repeated exposure to antigen in vivo

We have studied the effect of repeated in vivo antigen exposure on in vitro airway responsiveness in sensitized sheep. Fourteen sheep underwent five biweekly exposures to aerosolized Ascaris suum antigen or saline. Following this exposure regimen, the animals were killed and tracheal smooth muscle and lung parenchymal strips were prepared for in vitro studies of isometric contraction in response to histamine, methacholine, prostaglandin F2 alpha, and a thromboxane A2 analogue. No alteration in tracheal smooth muscle responsiveness was observed between saline- and antigen-exposed tissue. In contrast, by use of lung parenchymal strips as an index of peripheral airway responsiveness, significant increases in responsiveness to histamine and a thromboxane A2 analogue (10(-6) and 10(-5) M) were observed in antigen-exposed tissue compared with saline controls. These results demonstrate that repeated antigen exposure in vivo selectively increase the responsiveness of peripheral lung smooth muscle to certain chemical mediators of anaphylaxis.