Compensating for a stressful pregnancy? Glucocorticoid treatment during gravidity reduces metabolic rate in female fence lizards post-parturition.

Reproduction is a critical part of an animal's life history, but one which incurs significant costs to survival and future reproductive potential. These physiological consequences are likely to be influenced by context - for example, if an individual is subject to environmental stressors, physiological and behavioral changes associated with reproduction may be altered. Glucocorticoids, hormones produced as part of the physiological response to stressors, may alter how reproduction affects female physiology and behavior, and therefore the outcomes of reproductive trade-offs. Glucocorticoids prioritize immediate survival over reproduction, for example through changes in immune function, metabolic rate, and foraging, which may reduce energy expenditure or increase energy gain. However, we previously found that female eastern fence lizards (Sceloporus undulatus) experiencing elevated glucocorticoid levels during gestation were nevertheless able to maintain reproductive output and body condition. Here we investigate compensatory mechanisms by which eastern fence lizard females may maintain reproduction under experimental increases in a glucocorticoid, corticosterone (CORT). We found that, although CORT-treated females had similar immune function and behavior, they had reduced metabolic rates 3-5 days post-parturition compared to control females. Given that CORT-treated females spent a similar time basking and had equal food intake compared to control females, we suggest that the reduced metabolic rate is a mechanism by which CORT-treated females maintain their energy balance and reduce the energetic costs of gestation during periods of stress. This study suggests that physiological responses to reproduction may be context-dependent and could act to minimize costs of reproduction in situations where CORT is elevated (such as during periods of environmental stress).

Glucocorticoids do not influence a secondary sexual trait or its behavioral expression in eastern fence lizards.

Secondary sexual traits and associated behaviors can be influenced by environmental factors such as exposure to stressors. Such effects may be mediated by the physiological stress response, which is typified by the release of glucocorticoid hormones. The effects of glucocorticoids on sexual traits such as plumage and display coloration have most commonly been studied in isolation rather than in conjunction with other pertinent aspects of signalling, such as behavior and habitat use, though these have substantial potential to alter signal perception. Here we test the effects of corticosterone (CORT), a common glucocorticoid, on a secondary sexual trait (badge coloration) in male eastern fence lizards (Sceloporus undulatus), and behaviors associated with its expression. We show that neither baseline nor experimentally manipulated CORT levels were associated with badge coloration. Further, elevation of CORT levels in the field did not alter signalling or associated territorial behaviors. There was a trend for CORT-treatment to influence perch height selection, which may influence signal perception. We suggest that future studies investigating the effects of environmental stressors and associated physiological changes on secondary sexual traits should consider behaviors and ecology relevant to signal perception in order to best understand the influence of stressors in nature.

Survival and reproductive costs of repeated acute glucocorticoid elevations in a captive, wild animal.

Organisms are continuously encountering both predictable and unpredictable ecological stressors within their environment. The activation of the hypothalamic-pituitaryadrenal (stress) axis is a fundamental process allowing animals to cope with and respond to such encounters. A main consequence of HPA axis activation is the release of glucocorticoid hormones. Although short-term glucocorticoid elevations lead to changes in physiological and behavioral processes that are often adaptive, our understanding of fitness consequences of repeated acute elevations in glucocorticoid hormones over a longer time period is largely lacking. This is of particular current importance as animals are facing a significant increase in exposure to stressors including those associated with human-induced rapid environmental change. Here, we test fitness-relevant consequences of repeated exposure to glucocorticoids in the absence of natural challenges, by treating wild-caught gravid female eastern fence lizards (Sceloporus undulatus) with a daily transdermal dose of a glucocorticoid hormone until laying. This treatment causes an increase in plasma glucocorticoids that mimics the natural response lizards have when they encounter a stressor in the wild, without confounding effects associated with the encounter itself. This treatment reduced females' reproductive success (hatching success) and survival. Further, glucocorticoid-induced reductions in reproductive success were greater when females had experienced higher temperatures the previous winter. This demonstrates the potential significant consequences of repeated exposure to acute elevations in glucocorticoid hormones. Additionally, the costs of repeated glucocorticoid elevation may be further exaggerated by an individual's previous experience, such as the potential compounding effects of winter warming increasing animals' vulnerability to increased glucocorticoid levels during spring breeding.

Serum and sputum neurotrophin levels in chronic persistent cough.

BACKGROUND: Neurotrophins (NTs) are a family of growth factors, including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin3 (NT-3) that are involved in inflammation. Serum and induced sputum NT levels are increased in asthma and in cough because of idiopathic pulmonary fibrosis, respectively. Neurogenic inflammation is implicated in the pathogenesis of chronic cough in individuals with normal chest radiography, but the role of NTs in this condition is unknown. OBJECTIVE: To assess if NT levels are elevated in the serum and airways in subjects with chronic persistent cough. METHODS: Eighty-one subjects with chronic cough persistent for over 1 year; with normal chest radiography and spirometry were included. Thirty healthy subjects were controls. Serum NGF, BDNF and NT-3 were measured by enzyme immunoassay. In a subset, NGF was measured in induced sputum. Sputum cell counts and allergen-specific serum IgE were measured and all patients received specific sequential treatment trials to achieve a final diagnosis for the cough. RESULTS: There was no significant difference either in the levels of serum or sputum NTs in chronic cough subjects compared with controls or between the most common causes of cough: post-nasal drip syndrome, gastro-oesophageal reflux disease, asthma and bronchiectasis. The median (inter-quartile range) for sputum NGF (pg/mL) was 516 (296-772) in healthy controls and 580 (312-880) in subjects with chronic cough (P=0.284). There was no correlation between NT levels and sputum cell counts. Sputum NGF levels correlated with duration of cough (r=0.34, P=0.002). CONCLUSION: NTs are not elevated in induced sputum or serum of subjects with chronic persistent cough. This implies that NTs do not have a central role in perpetuating airway inflammation in chronic persistent cough.

Interleukin-18 levels in induced sputum are reduced in asthmatic and normal smokers.

BACKGROUND: IL-18 is a cytokine which is known to have an important role in the development of a Th1 lymphocyte response. As such, it may have a regulatory role in asthma by modifying Th2 lymphocyte responses. Cigarette smoking may amplify the airway inflammation associated with asthma. OBJECTIVE: This study investigated if IL-18 could be detected in induced sputum from asthmatics and normal subjects and if smoking altered IL-18 levels. METHODS: Induced sputum was obtained from asthmatic (31 smokers, 35 non-smokers) and normal (20 smokers, 20 non-smokers) subjects. All smokers had a smoking history of > or =15 pack years. IL-18 levels in sputum supernatant were measured by ELISA. IL-18 mRNA expression and cellular localization were assessed by quantitative PCR and immunocytochemistry, respectively. RESULTS: Smoking was associated with a significant reduction in IL-18 levels (median (interquartile range) - smokers 20 (0-102) pg/mL vs. non-smokers 358 (50-876) pg/mL, P<0.001). This was more pronounced in asthmatics (smokers, 47 (40-64) pg/mL vs. non-smokers, 530 (30-1484) pg/mL; P<0.001) than in normal subjects (smokers, 25 (0-78) pg/mL vs. non-smokers, 247 (50-656) pg/mL; P<0.01). Within each of the smoking and non-smoking groups there was no significant difference in IL-18 levels between asthmatic and normal subjects. There was no correlation between sputum IL-18 levels and any specific cell type in the sputum samples nor serum IgE levels. IL-18 mRNA expression was reduced in asthmatic smokers compared with non-smokers. IL-18 production was localized to sputum macrophages by immunocytochemistry. CONCLUSIONS: IL-18 is detectable in induced sputum samples from both asthmatic and normal subjects. Cigarette smoking significantly reduces sputum IL-18 levels. This effect is more pronounced in asthmatics than in normal subjects.

Influence of cigarette smoking on inhaled corticosteroid treatment in mild asthma.

BACKGROUND: Although inhaled corticosteroids have an established role in the treatment of asthma, studies have tended to concentrate on non-smokers and little is known about the possible effect of cigarette smoking on the efficacy of treatment with inhaled steroids in asthma. A study was undertaken to investigate the effect of active cigarette smoking on responses to treatment with inhaled corticosteroids in patients with mild asthma. METHODS: The effect of treatment with inhaled fluticasone propionate (1000 microg daily) or placebo for 3 weeks was studied in a double blind, prospective, randomised, placebo controlled study of 38 steroid naïve adult asthmatic patients (21 non-smokers). Efficacy was assessed using morning and evening peak expiratory flow (PEF) readings, spirometric parameters, bronchial hyperreactivity, and sputum eosinophil counts. Comparison was made between responses to treatment in non-smoking and smoking asthmatic patients. RESULTS: There was a significantly greater increase in mean morning PEF in non-smokers than in smokers following inhaled fluticasone (27 l/min v -5 l/min). Non-smokers had a statistically significant increase in mean morning PEF (27 l/min), mean forced expiratory volume in 1 second (0.17 l), and geometric mean PC20 (2.6 doubling doses), and a significant decrease in the proportion of sputum eosinophils (-1.75%) after fluticasone compared with placebo. No significant changes were observed in the smoking asthmatic patients for any of these parameters. CONCLUSIONS: Active cigarette smoking impairs the efficacy of short term inhaled corticosteroid treatment in mild asthma. This finding has important implications for the management of patients with mild asthma who smoke.

High resolution computed tomographic assessment of airway wall thickness in chronic asthma: reproducibility and relationship with lung function and severity.

BACKGROUND: In some patients chronic asthma results in irreversible airflow obstruction. High resolution computed tomography (HRCT) has been advocated for assessing the structural changes in the asthmatic lung and permits investigation of the relationships between airway wall thickening and clinical parameters in this condition. METHODS: High resolution CT scanning was performed in 49 optimally controlled asthmatic patients and measurements of total airway and lumen diameter were made by two independent radiologists using electronic callipers. Wall area as % total airway cross sectional area (WA%) and wall thickness to airway diameter ratio (T/D) were calculated for all airways clearly visualised with a transverse diameter of more than 1.5 mm, with a mean value derived for each patient. Intra- and inter-observer variability was assessed for scope of agreement in a subgroup of patients. Measurements were related to optimum forced expiratory volume in 1 second (FEV1), forced mid expiratory flow, carbon monoxide gas transfer, two scores of asthma severity, airway inflammation as assessed with induced sputum, and exhaled nitric oxide. RESULTS: Neither observer produced a statistically significant difference between measurements performed on two occasions but there was a significant difference between observers (limits of agreement -2.6 to 6.8 for WA%, p<0.0001). However, mean WA% measured on two occasions differed by no more than 5.4% (limits of agreement -4.0 to 5.4; mean (SD) 0.7 (2.4)). Statistically significant positive associations were observed between both WA% and T/D ratio and asthma severity (r(S)=0.29 and 0.30, respectively, for ATS score), and an inverse association with gas transfer coefficient was observed (r(S)=-0.43 for WA% and r(S)=-0.41 for T/D). No association was identified with FEV1 or airway inflammation. CONCLUSIONS: The airway wall is thickened in more severe asthma and is associated with gas transfer coefficient. This thickening does not relate directly to irreversible airflow obstruction as measured with FEV1.

Smoking and airway inflammation in patients with mild asthma.

STUDY OBJECTIVES: Cigarette smoking is common in asthmatic patients, and we investigated the impact of cigarette smoking on airway inflammation in asthma. DESIGN: Single-center observational study of airway inflammation in asthmatic and healthy smokers and nonsmokers. SETTING: Asthma research unit in a university hospital. PATIENTS OR PARTICIPANTS: Sixty-seven asthmatic and 30 nonasthmatic subjects classified as smokers or nonsmokers. Asthmatics had chronic, stable asthma and were not receiving inhaled or oral steroids at the time of the study. INTERVENTIONS: We examined induced-sputum cell counts and levels of interleukin (IL)-8 and eosinophilic cationic protein (ECP). Bronchial hyperreactivity was assessed using methacholine challenge. MEASUREMENTS AND RESULTS: Asthmatic smokers had higher total sputum cell counts than nonsmoking asthmatics and both smoking and nonsmoking healthy subjects. Smoking was associated with sputum neutrophilia in both asthmatics and nonasthmatics (median, 47% and 41%, respectively) compared with nonsmokers (median, 23% and 22%, respectively), and sputum IL-8 was increased in smokers compared with nonsmokers, both in subjects with asthma (median, 945 pg/mL vs 660 pg/mL, respectively) and in healthy subjects (median, 1,310 pg/mL vs 561 pg/mL, respectively). Sputum eosinophils and ECP levels were higher in both nonsmoking and smoking asthmatics than in healthy nonsmokers. In smoking asthmatics, lung function (FEV(1) percent predicted) was negatively related to both sputum IL-8 (r = - 0.52) and sputum neutrophil proportion (r = - 0.38), and sputum IL-8 correlated positively with smoking pack-years (r = 0.57) and percent neutrophil count (r = 0.51). CONCLUSIONS: In addition to the eosinophilic airway inflammation observed in patients with asthma, smoking induces neutrophilic airway inflammation; a relationship is apparent between smoking history, airway inflammation, and lung function in smoking asthmatics.

Conserved cysteine and tryptophan residues of the endothelin-converting enzyme-1 CXAW motif are critical for protein maturation and enzyme activity.

The neprilysin (NEP)/endothelin-converting enzyme (ECE) family of metalloproteases contains a highly conserved carboxyl-terminal tetrapeptide sequence, CXAW, where "C" is cysteine, "X" is a polar amino acid, "A" is an aliphatic residue, and "W" is tryptophan. Although this sequence strongly resembles a prenylation motif, human ECE-1 did not appear to be prenylated when labeled in vivo using various isoprenoid precursors in cell lines expressing ECE-1. We used site-directed mutagenesis to investigate the role of the CXAW motif and determined that the conserved cysteine residue of the CXAW motif in ECE-1, Cys(755), is critical for proper folding of the enzyme, its export from the endoplasmic reticulum, and its maturation in the secretory pathway. In addition, site-directed mutagenesis revealed that the conserved tryptophan residue of the sequence CEVW appears to be important for endoplasmic reticulum export and is essential for enzyme activity. Deletion of Trp(758) or substitution with alanine greatly slowed maturation of the enzyme, and resulted in more than a 90% loss of enzyme activity relative to the wild type. Conservative substitution of the tryptophan with phenylalanine did not reduce activity, whereas replacement with tyrosine, methionine, or leucine reduced enzyme activity by 50%, 75%, and 85%, respectively. Together, these data indicate that the conserved CEVW sequence does not serve as a prenylation signal and that both the conserved cysteine and tryptophan residues are necessary for proper folding and maturation of the enzyme. Furthermore, the conserved tryptophan appears to be critical for enzyme activity.

Non-invasive markers of airway inflammation as predictors of oral steroid responsiveness in asthma.

BACKGROUND: Sputum eosinophil counts and exhaled nitric oxide (NO) levels are increased in asthma and both measurements fall in response to corticosteroids. METHODS: Exhaled NO levels and sputum eosinophil counts were assessed as non-invasive markers of the response to an oral steroid in 37 patients (19 women) with stable chronic asthma (mean (SD) age 48.6 (12.2) years, asthma duration 25. 9 (17.3) years, and baseline forced expiratory volume in one second (FEV(1)) 76.3 (21.9)% predicted). Spirometric tests, with reversibility to a beta agonist (2.5 mg nebulised salbutamol), and induced sputum (using nebulised 3% saline) were performed at recruitment and following treatment with 30 mg prednisolone/day for 14 days. RESULTS: Baseline NO levels correlated with the percentage improvement in FEV(1) from baseline to the post-steroid, post-bronchodilator value (r(s) = 0.47, p = 0.003), with an NO level of >10 ppb at baseline having a positive predictive value of 83% for an improvement in FEV(1) of > or =15% (sensitivity 59%, specificity 90%). Sputum eosinophilia (> or =4%) had a positive predictive value of 68% (sensitivity 54%, specificity 76%) for an increase in FEV(1) of > or =15%. A combination of sputum eosinophilia and increased NO levels resulted in a positive predictive value of 72% and a negative predictive value of 79% (sensitivity 76%, specificity 75%). CONCLUSION: Exhaled NO levels and sputum eosinophilia may be useful in predicting the response to a trial of oral steroid in asthma.

Cysteine scanning mutagenesis of the noncatalytic nucleotide binding site of the yeast V-ATPase.

To investigate residues involved in the formation of the noncatalytic nucleotide binding sites of the vacuolar proton-translocating adenosine triphosphatase (V-ATPase), cysteine scanning mutagenesis of the VMA2 gene that encodes the B subunit in yeast was performed. Replacement of the single endogenous cysteine residue at position 188 gave rise to a Cys-less form of the B subunit (Vma2p) which had near wild-type levels of activity and which was used in the construction of 16 single cysteine-containing mutants. The ability of adenine nucleotides to prevent reaction of the introduced cysteine residues with the sulfhydryl reagent 3-(N-maleimidopropionyl)biocytin (biotin-maleimide) was evaluated by Western blot. Biotin-maleimide labeling of the purified V-ATPase from the wild-type and the mutants S152C, L178C, N181C, A184C, and T279C was reduced after reaction with the nucleotide analog 3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate (BzATP). These results suggest the proximity of these residues to the nucleotide binding site on the B subunit. In addition, we have examined the level of endogenous nucleotide bound to the wild-type V-ATPase and to a mutant (the A subunit mutant R483Q) which is postulated to be altered at the noncatalytic site and which displays a marked nonlinearity in ATP hydrolysis (MacLeod, K. J., Vasilyeva, E., Baleja, J. D., and Forgac, M. (1998) J. Biol. Chem. 273, 150-156). The R483Q mutant contained 2.6 mol of ATP/mol of V-ATPase compared with the wild-type enzyme, which contained 0.8 mol of ATP/mol of V-ATPase. These results suggest that binding of additional ATP to the noncatalytic sites may modulate the catalytic activity of the enzyme.

Photoaffinity labeling of wild-type and mutant forms of the yeast V-ATPase A subunit by 2-azido-[(32)P]ADP.

Molecular modeling studies have previously suggested the possible presence of four aromatic residues (Phe(452), Tyr(532), Tyr(535), and Phe(538)) near the adenine binding pocket of the catalytic site on the yeast V-ATPase A subunit (MacLeod, K. J., Vasilyeva, E., Baleja, J. D., and Forgac, M. (1998) J. Biol. Chem. 273, 150-156). To test the proximity of these aromatic residues to the adenine ring, the yeast V-ATPase containing wild-type and mutant forms of the A subunit was reacted with 2-azido-[(32)P]ADP, a photoaffinity analog that stably modifies tyrosine but not phenylalanine residues. Mutant forms of the A subunit were constructed in which the two endogenous tyrosine residues were replaced with phenylalanine and in which a single tyrosine was introduced at each of the four positions. Strong ATP-protectable labeling of the A subunit was observed for the wild-type and the mutant containing tyrosine at 532, significant ATP-protectable labeling was observed for the mutants containing tyrosine at positions 452 and 538, and only very weak labeling was observed for the mutants containing tyrosine at 535 or in which all four residues were phenylalanine. These results suggest that Tyr(532) and possibly Phe(452) and Tyr(538) are in close proximity to the adenine ring of ATP bound to the A subunit. In addition, the effects of mutations at Phe(452), Tyr(532), Tyr(535), and Glu(286) on dissociation of the peripheral V(1) and integral V(0) domains both in vivo and in vitro were examined. The results suggest that in vivo dissociation requires catalytic activity while in vitro dissociation requires nucleotide binding to the catalytic site.

Sputum cellular and cytokine responses to inhaled endothelin-1 in asthma.

BACKGROUND: Endothelin (ET)-1 is a 21-amino acid peptide which has potent bronchoconstrictor activity. Animal studies show elevation of ET-1 during experimental airway inflammation, and inhibition of inflammation by endothelin-antagonists, suggesting pro-inflammatory activity for ET-1. OBJECTIVE: We wanted to assess any acute influence that bronchoconstrictor doses of inhaled ET-1 might have on cells, tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, nitrite (NO2) and albumin in induced sputum in asthma. METHODS: Bronchial challenge was performed using nebulized ET-1 (nebulized dose range 0.96-15.36 nmol) and placebo in 10 adult asthmatic subjects in a randomized double-blind placebo-controlled cross-over study. Sputum induction was performed 30 min and 4 h after placebo or ET-1 bronchial challenge. RESULTS: All subjects experienced dose-dependent bronchoconstriction to inhaled ET-1 with a mean (range) PC15 forced expiratory volume in 1 s (FEV1) to ET-1 of 9.45 (1.2-21.7) nmol. Comparing ET-1 with placebo inhalation, there was no change in sputum differential cell counts, TNFalpha, IL-1beta, NO2 or albumin at 30 min or 4 h after inhalation, nor was there a difference in these parameters at 4 h compared with 30 min after ET-1 inhalation. There was no fall in FEV1 at 4 h after ET-1 inhalation, suggesting that ET-1 inhalation is not associated with a late bronchoconstrictor response. CONCLUSIONS: We conclude that inhaled ET-1 does not appear to stimulate an acute inflammatory response in asthma as assessed by differential cell count, TNFalpha, IL-1beta, NO2 and albumin concentrations in induced sputum.

Sputum endothelin-1 is increased in cystic fibrosis and chronic obstructive pulmonary disease.

Many patients with cystic fibrosis (CF) have airflow obstruction, with peribronchial and peribronchiolar fibrosis. Endothelin (ET)-1 is a potent bronchoconstrictor with mitogenic activity for airway smooth muscle. Do the levels of ET-1 in sputum support the putative role of ET-1 in contributing to airway remodelling with airflow obstruction in CF? The levels of ET-1 in plasma, saliva and sputum from 12 adult patients with CF not in exacerbation (spontaneous sputum), 17 normal control subjects (induced sputum) and as an additional control population, nine patients with stable chronic obstructive pulmonary disease (COPD) (seven spontaneous sputum) were measured. Total and differential sputum cell counts were performed. Median (interquartile range) sputum ET-1 level was elevated in CF (77.6 (29.0-122.8) pg x mL(-1)) compared to normal subjects (6.00 (2.8-14.8) pg x mL(-1)) and COPD (16.4 (6.8-38.2) pg x mL(-1)), and in COPD compared to normal subjects. There was a slight elevation of plasma ET-1 level in CF (5.3 (3.2-6.0) pg x mL(-1)) compared to normal subjects (3.1 (1.7-4.4) pg x mL(-1)) and COPD (3.3 (2.7-4.2) pg x mL(-1)). Sputum and saliva ET-1 levels were significantly higher than plasma levels in all groups, suggesting local production or release in the respiratory tract. Sputum differential cell counts revealed pronounced neutrophilia in CF and COPD compared to normal subjects. Sputum endothelin-1 concentrations are elevated in cystic fibrosis sputum compared to chronic obstructive pulmonary disease, and in cystic fibrosis and chronic obstructive pulmonary disease compared to normal subjects. The role of endothelin-1 in contributing to airflow obstruction through bronchoconstriction and mitogenesis in cystic fibrosis needs now to be explored.

Mutational analysis of the nucleotide binding sites of the yeast vacuolar proton-translocating ATPase.

To further define the structure of the nucleotide binding sites on the vacuolar proton-translocating ATPase (V-ATPase), the role of aromatic residues at the catalytic sites was probed using site-directed mutagenesis of the VMA1 gene that encodes the A subunit in yeast. Substitutions were made at three positions (Phe452, Tyr532, and Phe538) that correspond to residues observed in the crystal structure of the homologous beta subunit of the bovine mitochondrial F-ATPase to be in proximity to the adenine ring of bound ATP. Although conservative substitutions at these positions had relatively little effect on V-ATPase activity, replacement with nonaromatic residues (such as alanine or serine) caused either a complete loss of activity (F452A) or a decrease in the affinity for ATP (Y532S and F538A). The F452A mutation also appeared to reduce stability of the V-ATPase complex. These results suggest that aromatic or hydrophobic residues at these positions are essential to maintain activity and/or high affinity binding to the catalytic sites of the V-ATPase. Site-directed mutations were also made at residues (Phe479 and Arg483) that are postulated to be contributed by the A subunit to the noncatalytic nucleotide binding sites. Generally, substitutions at these positions led to decreases in activity ranging from 30 to 70% relative to wild type as well as modest decreases in Km for ATP. Interestingly, the R483E and R483Q mutants showed a time-dependent increase in ATPase activity following addition of ATP, suggesting that events at the noncatalytic sites may modulate the catalytic activity of the enzyme.

Endothelin-1 levels in induced sputum samples from asthmatic and normal subjects.

BACKGROUND: Endothelin-1 (ET-1) is a potent bronchoconstrictor which may have a role in the pathogenesis of asthma. The levels of ET-1 in saliva, induced sputum, and plasma from asthmatic and non-asthmatic subjects were compared. METHODS: Sputum induction was performed on 28 asthmatic subjects and nine normal volunteers. ET-1 levels were measured in plasma, saliva, and sputum samples and reversed phase high performance liquid chromatography (RP-HPLC) was performed on saliva and sputum samples. RESULTS: ET-1 was present in the following order of concentration in both normal and asthmatic subjects: saliva > sputum > plasma (saliva, median 30.1 and 23.9 pg/ ml, respectively; sputum, median 15.5 and 11.2 pg/ml; plasma, median 3.1 and 3.6 pg/ ml). There were no differences between asthmatic and normal subjects in the levels of ET-1 in each fluid. The levels of ET-1 in asthmatic subjects were not influenced by whether or not they were taking inhaled steroids. RP-HPLC of sputum and saliva confirmed the presence of ET-1 in these fluids. CONCLUSIONS: Levels of ET-1 can be measured in saliva and sputum obtained by sputum induction in asthmatic and healthy subjects and, although no difference was found in basal levels of ET-1 in sputum, saliva and plasma between normal subjects and asthmatics without bronchoconstriction, it is apparent that ET-1 is produced or released locally within the respiratory tract in concentrations higher than those in plasma.

Visual information processing during controlled hypoglycaemia in humans.

A general impairment of cognitive performance occurs during acute insulin-induced hypoglycaemia, but little objective evidence is available for disruption of more specific cognitive processes. The effect of controlled hypoglycaemia on the early stages of visual information processing and contrast sensitivity was examined in a homogeneous group of 20 nondiabetic human subjects. Hypoglycaemia caused a significant disruption in general cognitive performance as assessed by a digit symbol task (P < 0.001) and the trail making B task (P < 0.05). Hypoglycaemia also produced a highly significant deterioration in performance on all of the visual information processing tasks, namely inspection time (IT) (P = 0.01), visual change detection (VCD) (P < 0.005) and visual movement detection (VMD) (P < 0.005). A significant deterioration in contrast sensitivity was observed during hypoglycaemia (P < 0.005). In contrast, no significant effect of hypoglycaemia was demonstrated on standard clinical measures of visual acuity or stereoscopic vision. Thus, although hypoglycaemia caused no detectable deterioration in visual acuity as measured by Snellen-type tests, a marked deterioration occurred in the speed of visual information processing and in contrast sensitivity. As many decisions are made under conditions of limited perceptual time and low visual contrast (e.g. when driving), the disruptive effect of moderate insulin-induced hypoglycaemia on visual perception will have important practical implications in diabetic humans exposed to this metabolic stress. The present results are congruent with other evidence which shows that the early stages of visual information processing are susceptible to deterioration by general cerebral insults.