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R W Mitchell

Publications and source records attributed to R W Mitchell.

At least 37 records · Page 2Linked to original sources
PubMed1994

Immune sensitization augments epithelium-dependent spontaneous tone in guinea pig trachealis.

We examined epithelial modulation of tracheal smooth muscle (TSM) responsiveness in vitro from guinea pigs receiving active immune sensitization in vivo. Initially, guinea pigs were either ovalbumin sensitized (by aerosol) or sham sensitized with normal saline; TSM responsiveness was assessed isometrically as active tension (AT) after equilibration by electrical field stimulation in vitro. For epithelium-intact (Epi+) tissues, sensitization caused an increase in baseline active spontaneous tone (1.89 +/- 0.20 g AT) vs. sham-sensitized tissues (1.18 +/- 0.28 g AT; P = 0.02). Spontaneous tone in sensitized TSM in which the epithelium was removed (Epi-) (1.01 +/- 0.14 g AT) was substantially less than from Epi+ tissues (P = 0.01) and did not differ from sham-sensitized epithelium-denuded tissues (0.82 +/- 0.24 g AT; P > 0.05). Indomethacin caused a reduction in spontaneous tone to comparable magnitude for all treatment paradigms. Immune sensitization caused physiological reduction in the ability to relax in response to isoproterenol; the concentration of isoproterenol eliciting 50% relaxation of spontaneous tone was 7.10 +/- 0.13 (-log M) for TSM from sensitized guinea pigs compared with 8.20 +/- 0.27 (-log M) for sham-sensitized tissues (P = 0.006). However, after precontraction with exogenous acetylcholine, relaxation caused by isoproterenol was not affected by either indomethacin or epithelial removal. Muscarinic responsiveness to acetylcholine was augmented by immune sensitization; however, the increase in response to acetylcholine was attenuated by epithelium removal or cyclooxygenase blockade.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1994

Passive sensitization of human bronchi augments smooth muscle shortening velocity and capacity.

We assessed whether incubation with human serum from atopic individuals containing high concentrations of immunoglobulin E (IgE) causes augmentation of maximal contraction of human bronchial smooth muscle from non-atopic subjects in vitro. Bronchi were obtained from eight patients undergoing lung resection, and force-velocity relationships were determined for eight pairs of epithelium-intact bronchial rings of generations 6-7 using an electromagnetic lever system, which allowed isotonic shortening when load-clamps [from 0 to maximal isometric force (P0)] were applied at specific times after onset of contraction. Contractions were elicited by supramaximal electrical field stimulation (50 Hz, 10 s train duration, 25 V). Optimal length (Lo) for each tissue was determined during equilibration. After resection, tissues were sensitized passively with human sera containing high titers (> 1,000 U/ml) of IgE by incubation for 16 h at 20 degrees C. Maximal shortening velocity (Vmax) was increased for passively sensitized bronchi [0.1150 +/- 0.0240 1/2 circumferences/s (1/2Cir/s)] compared with sham-sensitized bronchi [0.0731 +/- 0.0152 1/2Cir/s, P = 0.038]. Similarly, maximal shortening (delta Lmax) was augmented in sensitized bronchial rings (11.27 +/- 1.80 %Lo) compared with sham-sensitized tissues (8.19 +/- 1.39 %Lo, P = 0.012). However, P0 did not differ between sensitized (122.5 +/- 24.4 mN/cm2) compared with sham-sensitized tissues (138.4 +/- 32.1 mN/cm2, P = 0.642). Our data are the first demonstration that Vmax and delta Lmax are augmented in sensitized but not challenged human bronchial rings after passive sensitization using human serum containing high concentrations of IgE.

Bronchi↗
PubMed1994

Exposure of immature rats to hyperoxia increases tracheal smooth muscle stress generation in vitro.

Recently, we demonstrated that chronic exposure to hyperoxia causes in vivo airway muscarinic receptor hyperresponsiveness in the developing rat [Am. J. Physiol. 262 (Lung Cell. Mol. Physiol. 6): L263-L269, 1992]. To test whether airway cholinergic hyperresponsiveness might result from intrinsic alterations in smooth muscle contractility, we measured the effect of in vivo hyperoxia on the contractile force elicited by acetylcholine (ACh) of isometrically mounted tracheal rings in vitro. Tracheal rings were obtained from 3-wk-old rats exposed to air or to > 95% O2 for 8 days. Muscarinic responses were determined by measuring the force elicited by exposure to increasing concentrations of ACh. Responses were normalized to the morphometrically determined tracheal smooth muscle cross-sectional area in a plane perpendicular to the axis of force generation. In vivo O2 exposure significantly increased maximal ACh-induced stress generation (response to 10(-3) M ACh: air, 15.92 +/- 1.37 g/mm2; O2, 21.78 +/- 1.52 g/mm2; P = 0.010). The ACh-induced stress generation of cylinders from hyperoxic rats was substantially reduced by both epithelial removal and treatment with the cyclooxygenase inhibitor indomethacin. We conclude that in vivo hyperoxic exposure increases tracheal smooth muscle contractile function in vitro and that epithelium-derived prostaglandin(s) contributes to the observed increase in maximal contractile responsiveness.

Animals↗
PubMed1993

The 3' terminal region of a gene encoding a cysteine-rich surface protein in Giardia duodenalis.

DNA derived from chromosome band 3 of the cloned Giardia duodenalis line, WB-1B was used to construct a cloned library in E. coli. One of these clones, C3/23, has been identified as the 3' coding region of a G. duodenalis cysteine-rich variable surface protein (CRVSP) gene by homology with other published CRVSPs and also contains 720 bp of the 3' flanking region. The sequence of C3/23, was derived from genomic DNA independently of cDNA, or expression copies of the CRVSP genes. The 3' flanking region is not homologous to the 3' untranslated regions of published CRVSPs which probably reflects its genomic origin. Subclones of C3/23 were used to show that the 3' flanking region was conserved in all strains examined in this study and was repeated many times in the genome. The 3' flanking repeats were located on three chromosome bands and were not always associated with the coding sequence of C3/23 which was represented, although not equally, on all chromosome bands. The highly conserved nature of the 3' flanking region and its multiple representation in the genome emphasize the probable role of this sequence in the localization or regulation of expression of the CRVSPs in G. duodenalis.

Amino Acid Sequence↗
PubMed1993

Familiarity and the rarity of deception: two theories and their relevance to play between dogs (Canis familiaris) and humans (Homo sapiens).

One theory of the relation between familiarity and the frequency of deception predicts that familiarity leads to the rarity of deception and another, that familiarity increases deception. We examined which theory applied to play by comparing familiar and unfamiliar partners during play between dogs (Canis familiaris) and humans (Homo sapiens). Deceptions by humans were based on directionality of movement and petting the dog and on the projects show object and throw object, which are specialized for play. Likewise, deceptions by dogs were based on directionality of movement and the project retrieve object (an analogue to show object). Deceptions based on directionality and petting were rare among familiars (and unfamiliars), whereas those based on show object, throw object, and retrieve object were more frequent. The findings suggest that, in play at least, deception may occur frequently.

Adolescent↗
PubMed1993

Effect of immune sensitization on stimulated ACh release from trachealis muscle in vitro.

We assessed the effect of immune sensitization on acetylcholine (ACh) release from parasympathetic nerve terminals in tracheal smooth muscle (TSM) strips from ragweed-sensitized (RWS) and sham-sensitized, littermate control (LMC) dogs. Strips of TSM were tethered to force transducers at optimal length in perfusion chambers containing [3H]choline and a fixed volume of physiological perfusate. Tissues were equilibrated for 1 h by electrical field stimulation (EFS) every 5 min to facilitate uptake of label into parasympathetic nerves as ACh. Fresh perfusate (containing 3 x 10(-8) M physostigmine) was collected at 5-min intervals for 1 h, and a rate coefficient of [3H]ACh release was determined. Tissues were exposed to agonists in the seventh collection period, and the increase in label release (ratio change where < or = 1.00 = baseline) and force production were determined. Ragweed antigen challenge stimulated [3H]ACh release and contraction in RWS but not LMC tissues. [3H]ACh release was 1.93 +/- 0.22 x baseline in RWS vs. 0.92 +/- 0.02 in control tissues (P < 0.01); contraction was 31.2 +/- 9.5% of that elicited by EFS (% EFS) in RWS vs. 0% EFS in LMC tissues (P < 0.01). Strips of TSM from RWS but not LMC dogs demonstrated concentration-dependent, augmented release of ACh caused by histamine. After 10(-4) M histamine, [3H]ACh release in RWS was 1.94 +/- 0.37 x baseline vs. 1.05 +/- 0.06 for LMC tissues (P < 0.05); histamine also caused greater contraction in RWS (106.5 +/- 5.9% EFS) vs. LMC (86.5 +/- 5.6% EFS; P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1993

Effect of airway inflammation on smooth muscle shortening and contractility in guinea pig trachealis.

We studied the effect of either 1) immunogenic inflammation caused by aerosolized ovalbumin or 2) neurogenic inflammation caused by aerosolized capsaicin in vivo on guinea pig tracheal smooth muscle (TSM) contractility in vitro. Force-velocity relationships were determined for nine epithelium-intact TSM strips from ovalbumin-sensitized (OAS) vs. seven sham-sensitized controls and TSM strips for seven animals treated with capsaicin aerosol (Cap-Aer) vs. eight sham controls. Muscle strips were tethered to an electromagnetic lever system, which allowed isotonic shortening when load clamps [from 0 to maximal isometric force (Po)] were applied at specific times after onset of contraction. Contractions were elicited by supramaximal electrical field stimulation (60 Hz, 10-s duration, 18 V). Optimal length for each muscle was determined during equilibration. Maximal shortening velocity (Vmax) was increased in TSM from OAS (1.72 +/- 0.46 mm/s) compared with sham-sensitized animals (0.90 +/- 0.15 mm/s, P < 0.05); Vmax for TSM from Cap-Aer (0.88 +/- 0.11 mm/s) was not different from control TSM (1.13 +/- 0.08 mm/s, P = NS). Similarly, maximal shortening (delta max) was augmented in TSM from OAS (1.01 +/- 0.15 mm) compared with sham-sensitized animals (0.72 +/- 0.14 mm, P < 0.05); delta max for TSM from Cap-Aer animals (0.65 +/- 0.11 mm) was not different from saline aerosol controls (0.71 +/- 0.15 mm, P = NS). We demonstrate Vmax and delta max are augmented in TSM after ovalbumin sensitization; in contrast, neurogenic inflammation caused by capsaicin has no effect on isolated TSM contractility in vitro. These data suggest that airway hyperresponsiveness in vivo that occurs in association with immunogenic or neurogenic inflammation may result from different effects of these types of inflammation on airway smooth muscle.

Aerosols↗
PubMed1993

Pertussis toxin augments beta-adrenergic relaxation of muscarinic contraction in canine trachealis.

We studied the effect of pertussis toxin (PT) and partial muscarinic antagonism using pirenzepine (PIR) on beta-adrenergic relaxation of muscarinic contraction in 188 tracheal smooth muscle (TSM) preparations from 25 dogs in vitro. Strips of TSM were incubated for 4 h at 37 degrees C in Krebs-Henseleit (K-H) perfusate with or without 10 micrograms/ml of PT. In tissues contracted to target tension (TT; 50% of maximal response to 127 mM potassium-substituted K-H [KCl]) with acetylcholine (ACh), pretreatment with PT decreased the concentration of isoproterenol (ISO) causing 30% relaxation from TT (RC30) from 1.3 +/- 0.8 x 10(-7) M (control) to 2.8 +/- 0.7 x 10(-8) M (p = 0.013). Pretreatment with PT also augmented the maximal relaxation elicited by 10(-5) M ISO. In separate studies, strips of TSM were contracted with ACh; pretreatment with 10(-7) M PIR decreased the concentration of ISO causing 50% relaxation (RC50) from 3.4 +/- 0.6 x 10(-7) to 9.6 +/- 1.5 x 10(-8) M (p = 0.042). Pretreatment with PIR did not affect relaxation elicited by ISO for strips contracted equivalently with KCl. In addition, PIR increased both the potency and efficacy of ISO in relaxing muscarinic contraction in sham-incubated strips of TSM but had no effect after incubation with PT. Neither PT nor PIR affected beta-adrenergic relaxation of TSM contracted with KCl. Our data demonstrate that beta-adrenergic receptor relaxation of muscarinic contraction is augmented by (1) incubation with PT and (2) partial blockade of muscarinic receptors.

Acetylcholine↗
PubMed1993

Physiologic significance of epithelial removal on guinea pig tracheal smooth muscle response to acetylcholine and serotonin.

We studied the modulatory effect of airway epithelium on guinea pig tracheal smooth muscle (TSM) contraction. Isometric force was measured in vivo before and after removal of the tracheal epithelium. In parallel studies, TSM contraction was also measured isometrically in epithelium-intact and epithelium-denuded TSM strips in vitro. Epithelial removal in vivo did not alter the contractile response of TSM to acetylcholine (ACh) or serotonin. In nine guinea pigs, active tension (AT) caused by 3 x 10(-7) mol/kg of intravenous ACh was 0.74 +/- 0.14 g force per longitudinal length of the segment (g/cm) in the presence of epithelium versus 0.89 +/- 0.16 g/cm after removal of airway epithelium (confirmed histologically) (p NS). The threshold response to ACh was also unchanged (-8.0 +/- 0.3 log mol/kg control versus -8.3 +/- 0.3 log mol/kg after epithelial removal, p NS). In six guinea pigs, the AT caused by 3 x 10(-8) mol/kg of intravenous serotonin was 1.92 +/- 0.63 g/cm with an intact epithelium versus 2.15 +/- 0.70 g/cm after epithelial removal in vivo (p NS). Epithelial removal in vitro increased the sensitivity of TSM contraction to ACh when the data were expressed as the percentage maximal response to ACh. The concentration of ACh causing 50% of the maximal response (EC50) was -5.74 +/- 0.25 log M in eight epithelium-intact TSM strips versus -6.37 +/- 0.16 log M after epithelial removal in controls (n = 8) (p = 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1993

Heterogeneity of tracheobronchial lymphatic smooth muscle responses to histamine and 5-hydroxytryptamine.

We assessed the responsiveness of tracheobronchial lymphatic smooth muscle to mediators of inflammation to determine whether homogeneous responses to histamine and 5-hydroxytryptamine (5-HT) are demonstrated among species typically used in studies of lymph vessels. Fresh porcine and bovine tracheobronchial lymph vessels were suspended from force-displacement transducers in baths containing oxygenated Krebs solution. Concentration-response curves were generated by cumulative addition of histamine (10(-7) to 10(-3) M) or 5-HT (10(-7) to 3 x 10(-4) M). Active tension (AT) was expressed in milligrams and as a percentage of initial vessel ring response to 65mM KCl. Histamine elicited concentration-dependent contraction, yielding maximum AT in porcine rings of 1116 +/- 127 mg (n = 39; 129.1 +/- 10.5% of KCl response) and in bovine rings of 733 +/- 106 mg (n = 20; 65.8 +/- 12.9%; P = 0.0005 for percent responses). PD2 values (negative log10 of the concentration at half-maximum effect) were 4.49 +/- 0.08 and 4.82 +/- 0.08; (P = 0.0034). 5-HT elicited concentration-dependent contraction, yielding maximum AT of 560 +/- 50 mg in porcine rings (n = 15; 97.2 +/- 9.7%) and 2892 +/- 454 mg in bovine rings (n = 27; 159.0 +/- 29%; P < 0.0001 for percent responses). PD2 values were 6.25 +/- 0.05 and 5.28 +/- 0.04 (P < 0.0001). The data demonstrate a role for inflammatory mediators in the modulation of tracheobronchial lymphatic smooth muscle tone that is species- and mediator-specific, and support the potential for paracrine regulation of tracheobronchial lymph flow.

Animals↗
PubMed1993

Length-tension characteristics of bovine tracheobronchial lymphatic smooth muscle.

Current information regarding the physiology of lymphatic smooth muscle is derived from experiments on mesenteric and thoracic duct lymph trunks. We hypothesized that tracheobronchial lymphatics share many of the same properties possessed by the mesenteric lymphatics, and examined the passive and active length-tension characteristics of the two. Fresh isolated lymph vessel rings were prepared from bovine mesenteric and tracheobronchial lymphatic collectors, mounted in organ baths, and connected to force-displacement transducers. Isometric contractions were induced by exposure to 65mM KCl-substituted perfusate after intermittent ring length changes. Active tension was calculated. Optimal vessel length was greater in tracheobronchial vessel rings, averaging 4.9 +/- 0.4mm vs 2.8 +/- 0.3mm in mesenteric rings (p < 0.001). Optimal resting tension and ATmax were similar for both truncal types, measuring 738 +/- 95mg and 2379 +/- 289mg in tracheobronchial vessel rings, and 625 +/- 108mg and 2501 +/- 320mg in mesenteric vessel rings, respectively. Stress developed at L(o) (optimal length) was similar for tracheobronchial (35.4 +/- 4.3mN mm-2) and mesenteric (26 +/- 4.3mN mm-2) lymphatics (P = N.S.). The data demonstrate that tracheobronchial lymph vessels are similar to mesenteric lymph vessels in their ability to generate significant stress, and suggest that these lymphatics participate in the regulation of lymph flow.

Animals↗
PubMed1992

Physiological mechanisms mediating enhanced force generation during development and immune sensitization.

We examined the development of acetylcholinesterase (AChase) activity and tracheal smooth muscle (TSM) contraction elicited by acetylcholine (ACh) in a swine model of maturation and a dog model of allergic bronchospasm. Strips of TSM were tethered isometrically at optimal length and responses were expressed as a percentage of the maximum to KCl-substituted perfusate (% KCl). Maximal contraction (ATmax) to ACh in 2-week-old swine (168 +/- 8% KCl) was greater than in 10-week-old swine (142 +/- 2% KCl; p less than 0.02). The AChase inhibitor, physostigmine, augmented ACh-elicited ATmax in 10-week-old (27% increase; p less than 0.01) but not in 2-week-old swine (2% increase; p is NS) and caused a greater increase in sensitivity to muscarinic activation in 2 versus 10 week-old swine (p less than 0.02), thus demonstrating increased contraction of TSM in 2 versus 10-week-old swine, which results at least in part from reduced AChase activity in immature animals. In another study, TSM from ragweed-sensitized dogs demonstrated augmented efficacy to ACh-elicited contraction (180 +/- 6% KCl) compared with TSM from sham-sensitized, littermate controls (163 +/- 4% KCl; p less than 0.05). In the presence of physostigmine, ATmax was not different between ragweed-sensitized and control TSM.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1992

Ontogeny of shortening velocity in porcine trachealis.

We examined the effect of maturation on force-velocity (F-V) parameters in porcine tracheal smooth muscle (TSM) to determine the relationship between maximal isometric contractile force (Po) and maximal velocity of shortening (Vmax). Strips of TSM excised from 1-day-old neonatal swine (neo; n = 8), 2-wk-old swine (2ws; n = 7), and 10-wk-old swine (10ws; n = 7) were tethered to an electromagnetic lever system for F-V analysis of contractility. TSM strips were activated by electrical field stimulation at optimal resting tension, voltage, and length (Lo) so that maximal reproducible contractile force (Po) was elicited. Velocities were measured at the early phase of isometric contraction (3.1 +/- 0.1 s for neo, 2.9 +/- 0.1 s for 2ws, and 3.1 +/- 0.1 s for 10ws; P = NS). Shortening velocity increased progressively with maturation; Vmax was 0.164 +/- 0.011 Lo/s for neo, 0.194 +/- 0.013 Lo/s for 2ws (P less than 0.05 vs. neo), and 0.260 +/- 0.024 Lo/s for 10ws (P less than 0.01 vs. neo; P less than 0.05 vs. 2ws). Maximal isometric force generation increased substantially during the first 2 wk of postnatal life and thereafter returned to neonatal levels; Po was 71.5 +/- 2.1 mN/mm2 for neo, 95.4 +/- 7.0 mN/mm2 for 2ws, and 74.7 +/- 6.2 mN/mm2 for 10ws (P less than 0.05, 2ws vs. neo and 10ws). In separate studies, we also determined whether differences in Vmax occurred during the normal cycling phase of the cross bridge (3 s) or during the slowly cycling phase of the latch bridge (8 s) in tissue from 12 additional animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1991

Effect of in vitro preconditioning on tracheal smooth muscle responsiveness.

We evaluated the effect of preconditioning of the isometric contractile response of canine tracheal smooth muscle (TSM) in vitro. Strips of epithelium-free TSM (n = 90) were excised from 16 anesthetized dogs and fixed isometrically in tissue perfusion chambers. Experiments were performed using methods previously reported in which the following parameters were investigated: 1) quiescent equilibration time in the perfusion chamber (0-120 min); 2) effect of repeated exchange of perfusate; 3) method of determining the optimal resting length (Lmax) for presetting of resting tension (RT); 4) effect of precontraction during the equilibration phase on the contractile response to agonists administered subsequently; and 5) method of determining RT on the response to muscarinic stimulation. When other variables were uniform, neither equilibration time nor perfusate exchange affected potency or efficacy of the response generated subsequently to acetylcholine (ACh). However, both potency (range of EC50: -5.71 +/- 0.14 log M to -6.52 +/- 0.24 log M; P less than 0.02) and efficacy (range of maximal active tension: 1,143 +/- 268 g/cm2 to 2,878 +/- 151 g/cm2; P less than 0.001) of ACh were altered substantially as a result of the method used to estimate Lmax. Repeated precontraction by electrical field stimulation or with 127 mM KCl did not alter potency or efficacy of contraction elicited by ACh. Maximal active tension generated with 10(-3) M ACh was 2,878 +/- 151 g/cm2 after 12-15 tetanizing field stimulations and 2,696 +/- 198 g/cm2 after 5-7 contractions with 127 mM KCl (P = NS).(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1991

Effect of maturational changes in myosin content and morphometry on airway smooth muscle contraction.

We studied the relationship of airway morphometry, the content of myosin heavy-chain and isoform stoichiometry, and the distribution of bronchoconstrictor responses in the airways of maturing swine. Lungs were excised in 2-wk-old farm swine (2ws; n = 13) and 10-wk-old swine (10ws; n = 13), and tracheal smooth muscle strips and bronchial rings from generations 2-5 were fixed for in vitro isometric measurement of force generation. Split samples were placed in formaldehyde solution or glutaraldehyde for light- or electron-microscopic morphometry or frozen for analysis of tissue myosin content. The rank order of force generation elicited by both receptor- and nonreceptor-dependent mechanisms for both 2ws and 10ws was generation 4 greater than 3 greater than or equal to 2. For all matched airway generations, contractile force was 25-100% greater in 2ws than 10ws. Differences in force generation were not related to morphometric differences in smooth muscle mass content among airways. The relative cross-sectional area of smooth muscle derived by computerized morphometry was 5.5-7% for each airway generation and did not change with age. Electron-microscopic morphometry demonstrated comparable myocyte content within muscle bundles for all airways in both age groups. In generation 4 airways, myocyte size in 2ws (27.3 +/- 0.8 nuclei/2,500 microns2) hypertrophied approximately 15% in 10ws (20.4 +/- 0.6 nuclei/2,500 microns2; P less than 0.01). Tissue content of myosin measured by computerized laser densitometry of gel electrophoresis of homogenates was greater in trachea from 2ws than 10ws (135 +/- 10 vs. 90 +/- 4 micrograms/g tissue; P less than 0.01); homology of 200- and 205-kDa isoforms was confirmed by Western blot against polyclonal myosin antibody and Cleveland digest analysis of each band. Differences in contractile forces between generations in 2ws and 10ws were not correlated to functional myosin isoform content. We demonstrate a maturational downregulation of contractile forces in maturing swine. This response is independent of smooth muscle receptor distribution and is not related to morphological changes in airways muscle mass, cellularity, changes in content of nonmyocyte tissues, or tissue content of functional myosin isoform.

Aging↗
PubMed1991

Ontogenic expression of acetylcholinesterase activity in trachealis of young swine.

Previous investigations have demonstrated that cholinergic contraction of porcine tracheal smooth muscle (TSM) decreases between the second and tenth weeks of life. In this investigation, we hypothesized that the greater contractile response to acetylcholine (ACh) in TSM of 2-wk-old swine (2ws) vs. 10-wk-old swine (10ws) was the result of a relative decrease in activity of acetylcholinesterase (AChase). To examine this hypothesis, we assessed AChase activity directly in homogenates of TSM from eight 2ws and seven 10ws using a newly adapted method that measures the rate of cleavage of acetylthiocholine; enzyme activity was expressed as absorbance units per minute per milligram protein. The AChase from tissues of both age groups saturated at approximately 3 mM substrate. However, maximal AChase activity (Vmax) was significantly greater in 10ws than 2ws. Eadie-Hofstee analysis of enzyme kinetics revealed similar Michaelis-Menten constants for 2ws and 10ws. The concentration of physostigmine (PS), an inhibitor of cholinesterase, that elicited half-maximal inhibition of AChase activity also was similar for 2ws and 10ws. In separate studies, contraction of TSM strips was assessed in vitro at optimal resting length and expressed as a function of maximal force generation to potassium chloride. Strips of TSM from 2ws contracted with greater force than those of 10ws. After pretreatment with 10(-8) M PS, contractile forces were similar in 2ws and 10ws. We conclude that AChase activity measured directly in muscle homogenates is significantly reduced in TSM of 2ws vs. 10ws and that this may result in augmented contraction to ACh under conditions of zero-order kinetics.

Acetylcholine↗
PubMed1991

Extracellular Ca2+ mobilization in potential-dependent contraction of trachealis of maturing swine.

We studied the effect of maturation on potassium-induced parasympathetic activation and Ca2+ entry in tracheal smooth muscle (TSM) from fifteen 2-wk-old (2ws) and sixteen 10-wk-old (10ws) male domestic farm swine. Atropine (10(-7) M) caused inhibition of the maximal contraction elicited by potassium to 50.3 +/- 2.6% maximum of control response (P less than 0.001) in TSM from 2ws but had no significant effect in TSM from 10ws (94.6 +/- 4.2% maximum; P = NS vs. control). Verapamil (10(-7) M) plus 10(-7) M atropine reduced contraction elicited by potassium in both 2ws (23.7 +/- 5.8% maximum; P less than 0.001 vs. control) and 10ws (50.6 +/- 6.3% maximum; P less than 0.001 vs. control, P less than 0.05 vs. 2ws); 10(-6)M verapamil caused greater than 95% blockade of contraction caused by potassium in both 2ws and 10ws. In separate studies, atropine-treated strips were equilibrated with extracellular Ca2+ concentrations ([Ca2+]o) ranging from normal (1X [Ca2+]o) to four times normal (4x [Ca2+]o). Increasing [Ca2+]o increased maximal contractile response in atropine-treated TSM strips from 68.7 +/- 3.8% maximum for 1x [Ca2+]o to 100.8 +/- 4.8% maximum for 4x [Ca2+]o (P less than 0.001) in 2ws. Neither atropine nor [Ca2+]o affected maximal responses of TSM in 10ws (103.5 +/- 3.0% maximum for 1x [Ca2+]o; P = NS vs. control). However, in the presence of atropine and verapamil, 4x [Ca2+]o augmented KCl-elicited contraction of TSM from both 2ws (46.9 +/- 6.3% maximum; P less than 0.01 vs. control) and 10ws (78.6 +/- 2.3% maximum; P less than 0.005 vs. control).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗