Trazodone-induced parkinsonism: A case report.

BACKGROUND: Trazodone is prescribed off-label to treat insomnia, especially in older, not depressed adults. At low doses, it blocks histamine-1 (H1) receptors having a hypnotic effect. Unusual but potentially severe side effects of trazodone include suicidal behavior, excess sedation, QT prolongation, and priapism. Three case studies have been published in the last four decades describing trazodone-induced parkinsonism. CASE PRESENTATION: A 78-year-old Caucasian male with a past medical history of paroxysmal atrial fibrillation (on amiodarone), major depressive disorder, chronic obstructive pulmonary disease, hypothyroidism, and obstructive sleep apnea, was prescribed trazodone for his chronic insomnia. After 1 month, he was seen in the emergency department (ED) with complaints of coarse tremors of his upper extremities and could not write with a pen anymore due to shaking. He noticed dragging of his feet while walking for over a month, which caused him to have multiple falls and significantly impacted his activities of daily living. On clinical exam, the patient had bilateral cogwheel rigidity in ankles and shuffling gait. Trazodone was discontinued, and his symptoms resolved within a week. CONCLUSION: Trazodone was likely causing parkinsonism in our patient. Amiodarone may have hindered trazodone metabolism causing higher levels in blood. Multiple mechanisms of trazodone's effect on dopamine have been suggested, but the serotonin-dopamine system interaction remains significant. Physicians need to contemplate the benefits and detriments before adding more medications to the list for older adults. Polypharmacy can amplify the adverse effects of a drug that might not be seen in everyday practice.

Choice of cover crop influences soil fungal and bacterial communities in Prince Edward Island, Canada.

Soil fungal and bacterial communities play various roles in agroecosystems and are significantly influenced by agricultural management practices. Currently, little is known about the effects of selected cover crops on soil fungal and bacterial communities in no-till systems. In this study, eight cover crops, three mixed crops, and an unmanaged fallow control were evaluated over 2 years for their effects on the soil microbiome. Internal transcribed spacer (ITS) and 16S rRNA amplicon sequencing was performed to characterize fungal and bacterial communities in the soil during the cover crop growing season, and in the subsequent year. Fungal and bacterial alpha diversity significantly increased over time and were influenced in the subsequent growing season by choice of cover crops. Some fungal and bacterial trophic and functional groups were also affected by crop choice. Fungal pathotroph abundance was positively associated with oilseed radish, alfalfa, and phacelia, but negatively associated with sorghum-sudangrass. Beneficial symbiotrophic fungi and functional nitrification-related bacterial groups were also associated with sorghum-sudangrass and buckwheat. These findings suggest that choice of cover crops influences the soil microbial community composition and may impact plant health in the subsequent crops.

Prolonged Brackish Water Exposure: A Case Report.

Exposure to and consumption of brackish water are associated with an elevated risk of infection, hypernatremia, and hypothermia. Minimal data exist to support the diagnosis and treatment of patients with long-term brackish water exposure. We present a case of a patient who spent 5 to 10 d semisubmerged in the Elizabeth River in coastal Virginia. A 55-y-old male presented via ambulance after 5 to 10 d of being "stuck in the mud." He was hypernatremic, with a sodium of 176 mEq·L-1, hypothermic to 34.5°C (94.1°F), and hypotensive at 88/50 mm Hg, with a sodium concentration of 176 mEq·L-1 and an osmolality of 412 mosm·kg-1. He developed pneumonia, with respiratory cultures growing Vibrio parahemolyticus, Klebsiella oxytoca, and Shewanella algae. He had pustules, which grew Aeromonas hydrophilia and Aeromonas caviae. A nasogastric tube was placed. Using suction, 500 mL of coarse sand and gravel was removed from his stomach. Antibiotics and intravenous fluids were given. The patient fully recovered after 3 wk and was discharged to rehabilitation. Exposure to brackish water can present a unique set of infectious and metabolic complications. Initial care should include treatment of metabolic derangements, such as hypovolemia, hypernatremia, and hypothermia, and treatment of infections with antibiotics based on knowledge of the most likely causative organisms.

Riverine discharges to Chesapeake Bay: Analysis of long-term (1927-2014) records and implications for future flows in the Chesapeake Bay basin.

The Chesapeake Bay (CB) basin is under a total maximum daily load (TMDL) mandate to reduce nitrogen, phosphorus, and sediment loads to the bay. Identifying shifts in the hydro-climatic regime may help explain observed trends in water quality. To identify potential shifts, hydrologic data (1927-2014) for 27 watersheds in the CB basin were analyzed to determine the relationships among long-term precipitation and stream discharge trends. The amount, frequency, and intensity of precipitation increased from 1910 to 1996 in the eastern U.S., with the observed increases greater in the northeastern U.S. than the southeastern U.S. The CB watershed spans the north-to-south gradient in precipitation increases, and hydrologic differences have been observed in watersheds north relative to watersheds south of the Pennsylvania-Maryland (PA-MD) border. Time series of monthly mean precipitation data specific to each of 27 watersheds were derived from the Precipitation-elevation Regression on Independent Slopes Model (PRISM) dataset, and monthly mean stream-discharge data were obtained from U.S. Geological Survey streamgage records. All annual precipitation trend slopes in the 18 watersheds north of the PA-MD border were greater than or equal to those of the nine south of that border. The magnitude of the trend slopes for 1927-2014 in both precipitation and discharge decreased in a north-to-south pattern. Distributions of the monthly precipitation and discharge datasets were assembled into percentiles for each year for each watershed. Multivariate correlation of precipitation and discharge within percentiles among the groups of northern and southern watersheds indicated only weak associations. Regional-scale average behaviors of trends in the distribution of precipitation and discharge annual percentiles differed between the northern and southern watersheds. In general, the linkage between precipitation and discharge was weak, with the linkage weaker in the northern watersheds compared to those in the south. On the basis of simple linear regression, 26 of the 27 watersheds are projected to have higher annual mean discharge in 2025, the target date for implementation of the TMDL for the CB basin.

Bringing Barley Back: Analysis of Heritage Varieties for Use as Germplasm Sources to Improve Resistance against the Most Devastating, Contemporary Disease in Canada, Fusarium Head Blight (Fusarium graminearum).

Fusarium head blight (FHB), caused by Fusarium graminearum, is currently the most devastating disease for barley (Hordeum vulgare) in Canada. Associated mycotoxins can compromise grain quality, where deoxynivalenol (DON) is considered particularly damaging due to its frequency of detection. Breeding barley with a lower DON content is difficult, due to the poor adaptation and malt quality of resistance sources. A set of European-derived heritage varieties were screened in an FHB nursery in Charlottetown, PE, with selections tested at Brandon, MB, between 2018-2022. Genetic evaluation demonstrated a distinct clustering of Canadian varieties from the heritage set. At Brandon, 72% of the heritage varieties ranked lower for DON content than did the moderately resistant Canadian check 'AAC Goldman', but resistance was associated with later heading and taller stature. In contrast with Canadian modern malting variety 'AAC Synergy', general deficiencies were observed in yield, enzyme activity, and extract, along with higher protein content. Nonetheless, several resistant varieties were identified with reasonable a heading date and yield, including 'Chevallier Chile', 'Domen', 'Djugay', 'Hannchen', 'Heils Franken', 'Moravian Barley', 'Loosdorfer' with 'Golden Melon', 'Nutans Moskva', and 'Vellavia', these being some of the most promising varieties when malting quality characteristics were also considered. These heritage resources could be used as parents in breeding to develop FHB-resistant malting barley varieties.

Select Cover Crop Residue and Soil Microbiomes Contribute to Suppression of Fusarium Root and Crown Rot in Barley and Soybean.

Fusarium root and crown rot (FRCR) negatively impact several economically important plant species. Cover crops host different soil and residue microbiomes, thereby potentially influencing pathogen load and disease severity. The carryover effect of cover crops on FRCR in barley and soybean was investigated. Field trials were conducted in Prince Edward Island, Canada. Two cover crops from each plant group, including forbs, brassicas, legumes, and grasses, were grown in a randomized complete block design with barley and soybean planted in split plots the following year. Barley and soybean roots were assessed for FRCR through visual disease rating and Fusarium spp. were isolated from diseased tissue. Fungal and bacterial communities in cover crop residues were quantified using amplicon sequencing. The disease-suppressive effects of soil were tested in greenhouse studies. The results indicated that sorghum-sudangrass-associated microbiomes suppress Fusarium spp., leading to reduced FRCR in both barley and soybean. The oilseed radish microbiome had the opposite effect, consequently increasing FRCR incidence in barley and soybean. The results from this study indicate that cover crop residue and the associated soil microbiome influence the incidence and severity of FRCR in subsequent crops. This information can be used to determine cover cropping strategies in barley and soybean production systems.

Proteome Analysis of Rice (Oryza sativa L.) Mutants Reveals Differentially Induced Proteins during Brown Planthopper (Nilaparvata lugens) Infestation.

Although rice resistance plays an important role in controlling the brown planthopper (BPH), Nilaparvata lugens, not all varieties have the same level of protection against BPH infestation. Understanding the molecular interactions in rice defense response is an important tool to help to reveal unexplained processes that underlie rice resistance to BPH. A proteomics approach was used to explore how wild type IR64 and near-isogenic rice mutants with gain and loss of resistance to BPH respond during infestation. A total of 65 proteins were found markedly altered in wild type IR64 during BPH infestation. Fifty-two proteins associated with 11 functional categories were identified using mass spectrometry. Protein abundance was less altered at 2 and 14 days after infestation (DAI) (T1, T2, respectively), whereas higher protein levels were observed at 28 DAI (T3). This trend diminished at 34 DAI (T4). Comparative analysis of IR64 with mutants showed 22 proteins that may be potentially associated with rice resistance to the brown planthopper (BPH). Ten proteins were altered in susceptible mutant (D1131) whereas abundance of 12 proteins including S-like RNase, Glyoxalase I, EFTu1 and Salt stress root protein "RS1" was differentially changed in resistant mutant (D518). S-like RNase was found in greater quantities in D518 after BPH infestation but remained unchanged in IR64 and decreased in D1131. Taken together, this study shows a noticeable level of protein abundance in the resistant mutant D518 compared to the susceptible mutant D1131 that may be involved in rendering enhanced level of resistance against BPH.

Approach to Fatigue: Best Practice.

Owing to the broad differential diagnoses that can present as fatigue, a rational approach to diagnosis is paramount. Performance of a battery of diagnostic tests is unlikely to assist with diagnosis, highlighting the importance of a thorough history and physical examination. Fatigue can be a sequela of an underlying medical disease or exists as a primary condition. Management of secondary fatigue largely depends on treatment of the underlying condition. There are no FDA-approved medications for primary fatigue, now known as system exertion intolerance disease. Treatment is focused on individualized exercise therapy and cognitive behavioral therapy.

Design and synthesis of a hybrid potentiator-corrector agonist of the cystic fibrosis mutant protein DeltaF508-CFTR.

A developing therapy of cystic fibrosis caused by the DeltaF508 mutation in CFTR employs correction of defective CFTR chloride channel gating by a 'potentiator' and of defective CFTR protein folding by a 'corrector'. Based on SAR data for phenylglycine-type potentiators and bithiazole correctors, we designed a hybrid molecule incorporating an enzymatic hydrolysable linker to deliver the potentiator (PG01) fragment 2 and the corrector (Corr-4a) fragment 13. The hybrid molecule 14 contained PG01-OH and Corr-4a-linker-CO(2)H moieties, linked with an ethylene glycol spacer through an ester bond. The potentiator 2 and corrector 13 fragments (after cleavage) had low micromolar potency for restoration of DeltaF508-CFTR channel gating and cellular processing, respectively. Cleavage of hybrid molecule 14 by intestinal enzymes under physiological conditions produced the active potentiator 2 and corrector fragments 13, providing proof-of-concept for small-molecule potentiator-corrector hybrids as a single drug therapy for CF caused by the DeltaF508 mutation.

Phytochemical Characterization of Wild Hops (Humulus lupulus ssp. lupuloides) Germplasm Resources From the Maritimes Region of Canada.

A survey was conducted in the Maritimes region of eastern Canada to measure the phytochemical diversity of prenylchalcone, soft resins (alpha & beta acids), and flavonol constituents from 30 unique wild-growing populations of hops (Humulus lupulus L.). Based on cone chemometrics, the majority of accessions (63.3%) are native Humulus lupulus ssp. lupoloides, with cones containing both xanthogalenol and 4'-O-methyl xanthohumol as chemotaxonomic indicator molecules. Interestingly, the leaves of all verified Humulus lupulus ssp. lupulus accessions accumulated high proportions (>0.20 total flavonols) of two acylated flavonol derivatives (kaempferol-3-O-(6''-O-malonyl)-ß-D-glucopyranoside; quercetin-3-O-(6''-O-malonyl)-ß-D-glucopyranoside), both previously unreported from hops leaves. The native lupuloides accessions examined possess only trace amounts of this compound in their leaves (<0.10 total flavonols), suggesting its potential utility as a novel, leaf-derived chemotaxonomic marker for subspecies identification purposes. A leaf-derived taxonomic marker is useful for identifying wild-growing accessions, as leaves are present throughout the entire growing season, whereas cones are only produced late in summer. Additionally, the collection of cones from 10-meter tall wild plants in overgrown riparian habitats is often difficult. The total levels of alpha acids, beta acids, and prenylchalcones in wild-collected Maritimes lupuloides cones are markedly higher than those previously reported for lupuloides individuals in the westernmost extent of its native range and show potentially valuable traits for future cultivar development, while some may be worthy of immediate commercial release. The accessions will be maintained as a core germplasm resource for future cultivar development.

Small-molecule screen identifies inhibitors of a human intestinal calcium-activated chloride channel.

Calcium-activated chloride channels (CaCCs) are widely expressed in mammalian tissues, including intestinal epithelia, where they facilitate fluid secretion. Potent, selective CaCC inhibitors have not been available. We established a high-throughput screen for identification of inhibitors of a human intestinal CaCC based on inhibition of ATP/carbachol-stimulated iodide influx in HT-29 cells after lentiviral infection with the yellow fluorescent halide-sensing protein YFP-H148Q/I152L. Screening of 50,000 diverse, drug-like compounds yielded six classes of putative CaCC inhibitors, two of which, 3-acyl-2-aminothiophenes and 5-aryl-2-aminothiazoles, inhibited by >95% iodide influx in HT-29 cells in response to multiple calcium-elevating agonists, including thapsigargin, without inhibition of calcium elevation, calcium-calmodulin kinase II activation, or cystic fibrosis transmembrane conductance regulator chloride channels. These compounds also inhibited calcium-dependent chloride secretion in T84 human intestinal epithelial cells. Patch-clamp analysis indicated inhibition of CaCC gating, which, together with the calcium-calmodulin data, suggests that the inhibitors target the CaCC directly. Structure-activity relationships were established from analysis of more than 1800 analogs, with IC(50) values of the best analogs down to approximately 1 muM. Small-molecule CaCC inhibitors may be useful in pharmacological dissection of CaCC functions and in reducing intestinal fluid losses in CaCC-mediated secretory diarrheas.

Cell-based fluorescence screen for K+ channels and transporters using an extracellular triazacryptand-based K+ sensor.

K+ channels and K+-coupled membrane transporters are important targets for drug discovery. We previously developed a triazacryptand (TAC)-based K+ sensor, TAC-Red, and demonstrated its utility to image K+ waves in mouse brain in vivo (Padmawar et al. Nat. Methods. 2005, 2, 825-827). Here, we synthesized a green-fluorescing dextran conjugate of TAC-bodipy ("TAC-Limedex") for use as an extracellular K+ sensor and demonstrated its utility in measuring K+ transport across cell membranes. TAC-Limedex fluorescence increased by 50% with increasing [K+] from 0 to 2 mM and was insensitive to [Na+], [Cl-], or pH. K+ efflux from cells was quantified from increasing extracellular TAC-Limedex fluorescence following cell immersion in K+-free buffer. In HT-29 cells, K+ efflux was 2.0 +/- 0.1 micromol/cm2/s, increasing 8-fold following K+ channel activation by ATP; the increase in K+ efflux was inhibited by a K+ channel blocker or by preventing cytoplasmic calcium elevation. Electroneutral K+/Cl- cotransport was demonstrated in SiHa cells, in which K+ efflux was increased 3-fold by hypotonic challenge; the increase in K+ efflux was fully inhibited by a K+/Cl- transport blocker. K+ efflux measurements were adapted to a commercial fluorescence platereader for automated screening. The fluorescence-based K+ transport assay largely replaces assays requiring radioactive rubidium and is suitable for high-throughput identification of K+ transport modulators.

Oxidative stress caused by pyocyanin impairs CFTR Cl(-) transport in human bronchial epithelial cells.

Pyocyanin (N-methyl-1-hydroxyphenazine), a redox-active virulence factor produced by the human pathogen Pseudomonas aeruginosa, is known to compromise mucociliary clearance. Exposure of human bronchial epithelial cells to pyocyanin increased the rate of cellular release of H(2)O(2) threefold above the endogenous H(2)O(2) production. Real-time measurements of the redox potential of the cytosolic compartment using the redox sensor roGFP1 showed that pyocyanin (100 microM) oxidized the cytosol from a resting value of -318+/-5 mV by 48.0+/-4.6 mV within 2 h; a comparable oxidation was induced by 100 microM H(2)O(2). Whereas resting Cl(-) secretion was slightly activated by pyocyanin (to 10% of maximal currents), forskolin-stimulated Cl(-) secretion was inhibited by 86%. The decline was linearly related to the cytosolic redox potential (1.8% inhibition/mV oxidation). Cystic fibrosis bronchial epithelial cells homozygous for DeltaF508 CFTR failed to secrete Cl(-) in response to pyocyanin or H(2)O(2), indicating that these oxidants specifically target the CFTR and not other Cl(-) conductances. Treatment with pyocyanin also decreased total cellular glutathione levels to 62% and cellular ATP levels to 46% after 24 h. We conclude that pyocyanin is a key factor that redox cycles in the cytosol, generates H(2)O(2), depletes glutathione and ATP, and impairs CFTR function in Pseudomonas-infected lungs.

The effect of sample size in studies of soil microbial community structure.

Replicate soil samples of 0.01, 0.1, 0.25, 1.0 and 10.0 g were taken from a single, large, homogenized sample from a field maintained as continuous meadow. The samples were processed for direct enumeration of bacterial cells and community structure assays by DGGE analysis of PCR-amplified 16S-rDNA fragments from whole community extracts. The goal was to determine the sample size or size range that produced the most consistent results (i.e., mean values) and the lowest variance. Enumeration data were analyzed by ANOVA, and the community composition fingerprints were analyzed by discriminant analysis (DA). Acceptable results were obtained for sample sizes from 0.1 to 1.0 g for both enumeration and community fingerprinting, but the size that yielded the best results for both measures was 0.25 g. The results suggest that for well homogenized silt loam soils with moderate organic matter concentrations, this sample size should produce high quality consistent results. For soils that differ in organic concentrations or clay content, a reconnaissance survey similar to the present examination is recommended.

Bacterial Community Structure and Function along a Heavy Metal Gradient.

The response of the planktonic, sediment, and epilithic bacterial communities to increasing concentrations of heavy metals was determined in a polluted river. None of the communities demonstrated a pollution-related effect on bacterial numbers (viable and total), heterotrophic activity, resistance to Pb or Cu, or species diversity as determined by either the Shannon-Wiener diversity index or rarefaction. The lack of correlation between concentrations of heavy metals and resistance in the sediment bacterial community was investigated and found to be due at least in part to the high pH of the river water and the resultant reduction in heavy metal toxicity. The three different communities demonstrated characteristic profiles based on the relative abundances of bacterial strains grouped according to functional similarities.

Multi-scale variation in spatial heterogeneity for microbial community structure in an eastern Virginia agricultural field.

To better understand the distribution of soil microbial communities at multiple spatial scales, a survey was conducted to examine the spatial organization of community structure in a wheat field in eastern Virginia (USA). Nearly 200 soil samples were collected at a variety of separation distances ranging from 2.5 cm to 11 m. Whole-community DNA was extracted from each sample, and community structure was compared using amplified fragment length polymorphism (AFLP) DNA fingerprinting. Relative similarity was calculated between each pair of samples and compared using geostatistical variogram analysis to study autocorrelation as a function of separation distance. Spatial autocorrelation was found at scales ranging from 30 cm to more than 6 m, depending on the sampling extent considered. In some locations, up to four different correlation length scales were detected. The presence of nested scales of variability suggests that the environmental factors regulating the development of the communities in this soil may operate at different scales. Kriging was used to generate maps of the spatial organization of communities across the plot, and the results demonstrated that bacterial distributions can be highly structured, even within a habitat that appears relatively homogeneous at the plot and field scale. Different subsets of the microbial community were distributed differently across the plot, and this is thought to be due to the variable response of individual populations to spatial heterogeneity associated with soil properties.

A geostatistical analysis of small-scale spatial variability in bacterial abundance and community structure in salt marsh creek bank sediments.

Small-scale variations in bacterial abundance and community structure were examined in salt marsh sediments from Virginia's eastern shore. Samples were collected at 5 cm intervals (horizontally) along a 50 cm elevation gradient, over a 215 cm horizontal transect. For each sample, bacterial abundance was determined using acridine orange direct counts and community structure was analyzed using randomly amplified polymorphic DNA fingerprinting of whole-community DNA extracts. A geostatistical analysis was used to determine the degree of spatial autocorrelation among the samples, for each variable and each direction (horizontal and vertical). The proportion of variance in bacterial abundance that could be accounted for by the spatial model was quite high (vertical: 60%, horizontal: 73%); significant autocorrelation was found among samples separated by 25 cm in the vertical direction and up to 115 cm horizontally. In contrast, most of the variability in community structure was not accounted for by simply considering the spatial separation of samples (vertical: 11%, horizontal: 22%), and must reflect variability from other parameters (e.g., variation at other spatial scales, experimental error, or environmental heterogeneity). Microbial community patch size based upon overall similarity in community structure varied between 17 cm (vertical) and 35 cm (horizontal). Overall, variability due to horizontal position (distance from the creek bank) was much smaller than that due to vertical position (elevation) for both community properties assayed. This suggests that processes more correlated with elevation (e.g., drainage and redox potential) vary at a smaller scale (therefore producing smaller patch sizes) than processes controlled by distance from the creek bank.

The invalidity of disclaimers about the effects of social feedback on self-esteem.

Despite the fact that several theories suggest that people's self-esteem is affected by social approval and disapproval, many individuals steadfastly maintain that how other people regard them has no effect on how they feel about themselves. To examine the validity of these beliefs, two experiments compared the effects of social approval and disapproval on participants who had indicated either that their self-esteem is affected by how other people evaluate them or that their self-esteem is unaffected by interpersonal evaluation. Results of both studies converged to show that approval and disapproval clearly affected the self-esteem of even those individuals who denied that social evaluations affected their feelings about themselves.

Carrageenans, sulphated polysaccharides of red seaweeds, differentially affect Arabidopsis thaliana resistance to Trichoplusia ni (cabbage looper).

Carrageenans are a collective family of linear, sulphated galactans found in a number of commercially important species of marine red alga. These polysaccharides are known to elicit defense responses in plant and animals and possess anti-viral properties. We investigated the effect of foliar application of ι-, κ- and λ-carrageenans (representing various levels of sulphation) on Arabidopsis thaliana in resistance to the generalist insect Trichoplusia ni (cabbage looper) which is known to cause serious economic losses in crop plants. Plants treated with ι- and κ-carrageenan showed reduced leaf damage, whereas those treated with λ- carrageenan were similar to that of the control. In a no-choice test, larval weight was reduced by more than 20% in ι- and κ- carrageenan treatments, but unaffected by λ-carrageenan. In multiple choice tests, carrageenan treated plants attracted fewer T. ni larvae by the fourth day following infestation as compared to the control. The application of carrageenans did not affect oviposition behaviour of T. ni. Growth of T. ni feeding on an artificial diet amended with carrageenans was not different from that fed with untreated control diet. ι-carrageenan induced the expression of defense genes; PR1, PDF1.2, and TI1, but κ- and λ-carrageenans did not. Besides PR1, PDF1.2, and TI1, the indole glucosinolate biosynthesis genes CYP79B2, CYP83B1 and glucosinolate hydrolysing QTL, ESM1 were up-regulated by ι-carrageenan treatment at 48 h post infestation. Gas chromatography-mass spectrometry analysis of carrageenan treated leaves showed increased concentrations of both isothiocyanates and nitriles. Taken together, these results show that carrageenans have differential effects on Arabidopsis resistance to T. ni and that the degree of sulphation of the polysaccharide chain may well mediate this effect.

In situ measurement of airway surface liquid [K+] using a ratioable K+-sensitive fluorescent dye.

The airway surface liquid (ASL) is the thin fluid layer lining airway surface epithelial cells, whose volume and composition are tightly regulated and may be abnormal in cystic fibrosis (CF). We synthesized a two-color fluorescent dextran to measure ASL [K(+)], TAC-Lime-dextran-TMR, consisting of a green-fluorescing triazacryptand K(+) ionophore-Bodipy conjugate, coupled to dextran, together with a red fluorescing tetramethylrhodamine reference chromophore. TAC-Lime-dextran-TMR fluorescence was K(+)-selective, increasing >4-fold with increasing [K(+)] from 0 to 40 mm. In well differentiated human airway epithelial cells, ASL [K(+)] was 20.8 +/- 0.3 mm and decreased by inhibition of the Na(+)/K(+) pump (ouabain), ENaC (amiloride), CF transmembrane conductance regulator (CFTR(inh)-172), or K(+) channels (TEA or XE991). ASL [K(+)] was increased by forskolin but not affected by Na(+)/K(+)/2Cl(-) cotransporter inhibition (bumetanide). Functional and expression studies indicated the involvement of [K(+)] channels KCNQ1, KCNQ3, and KCNQ5 as determinants of ASL [K(+)]. [K(+)] in CF cultures was similar to that in non-CF cultures, suggesting that abnormal ASL [K(+)] is not a factor in CF lung disease. In intact airways, ASL [K(+)] was also well above extracellular [K(+)]: 22 +/- 1 mm in pig trachea ex vivo and 16 +/- 1 mm in mouse trachea in vivo. Our results provide the first noninvasive measurements of [K(+)] in the ASL and indicate the involvement of apical and basolateral membrane ion transporters in maintaining a high ASL [K(+)].