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The effects of high saline concentrations on the plasma osmolytes of the four populations of the Pacific Treefrog, Pseudacris regilla, were investigated. Two of the populations, Chris Wicht and Trabuco Canyon, were in fresh water environments and two populations, Newport Back Bay and Alkaline Marsh, were in brackish water environments. The Newport Backbay treefrogs were most tolerant of increased sodium chloride solutions with 11 of 15 individuals surviving at 240 mM sodium chloride at 72 hrs. The Trabuco Canyon population was the least tolerant with 10 of 15 specimens surviving 120 mM sodium chloride at 72 hrs but none at the 180 mM concentration at 72 hrs. The Chris Wicht and Alkaline Marsh treefrogs were intermediate in their survival times with frogs surviving in 180 mM saline solutions at 72 hrs. The Newport treefrogs appeared to be the most efficient in osmoregulating by mainstreaming the plasma sodium and increasing plasma urea to compensate for the osmotic gradient produced by the increased external saline concentrations. The Trabuco Canyon and the Chris Wicht treefrogs appeared to be intermediate in their osmoregulatory abilities with the Alkaline Marsh treefrogs being least efficient and more likely an osmoconformer. The difference in osmoregulatory responses appears to be a population difference and a function of habitat. Sodium chloride was the major plasma osmolyte in the four populations of treefrogs. Urea increased in importance as the external concentration of the saline media increased, thus the role of urea was significant in maintaining the internal osmoconcentration. These simple physiological mechanisms allow this widespread treefrog to occur in so many different habitats., The Pacific Treefrog, or Pacific Chorus Frog, Pseudacris regilla, formerly Hyla regilla, is the most wide spread Western United States' amphibian. It occurs in a variety of habitats (grasslands, woodlands, [...]

This study investigates the link between adaptation to biocides and antibiotics in Pseudomonas aeruginosa. An enrichment continuous culture of P. aeruginosa NCIMB 10421 (MIC 25 mg BKC [l.sup.-1]) was operated (D=0.04 [h.sup.-1], 792 h) with added benzalkonium chloride (BKC). A derivative, PA-29 (696 h), demonstrated a >12-fold decrease in sensitivity to the biocide (MIC >350 mg BKC [l.sup.-1]). The variant demonstrated a 256-fold increase in resistance to ciprofloxacin, with a mutation in the gyrA gene (Thr-83[right arrow]lle). Similarly, culturing of the original strain in a continuous-culture system with ciprofloxacin selection pressure led to the evolution of BKC-adapted populations (MIC 100 mg BKC [l.sup.-1]). Efflux pump activity predominantly contributed to the developed phenotype of PA-29. An amino acid substitution (Val-51[right arrow]Ala) in nfxB, the Mex efflux system regulator gene, was observed for PA-29. Overexpression of both MexAB-OprM and MexCD-OprJ was recorded for PA-29. Similarly, mexR, a repressor of the Mex system, was downregulated. Competition studies were carried out in continuous culture between PA-29 and the original strain (in the presence of subinhibitory concentrations of RKC). The outcome of competition was influenced by the concentration of biocide used and the nature of limiting nutrient. The inclusion of 1 mg BKC [l.sup.-1] in the medium feed was sufficient to select (S=0.011) for the BKC-adapted strain in magnesium-limited culture. Conversely, the presence of 10 mg BKC [l.sup.-1] in the medium supply was insufficient to select for the same organism (S=-0.017) in the glucose-limited culture. These results indicate the importance of environmental conditions on selection and maintenance of biocide adaptation. DOI 10.1099/mic.0.029751-0

Neurotransmitter:sodium symporters (NSS) have a critical role in regulating neurotransmission and are targets for psychostimulants, anti-depressants and other drugs (1,2). Whereas the non-homologous glutamate transporters mediate chloride conductance (3), in [...]

In airway epithelia, purinergic receptor (P2[Y.sub.2]-R) stimulation of intracellular calcium ([Ca.sup.2+.sub.i])-regulated ion transport is restricted to the membrane domain ipsilateral to receptor activation, implying compartmentalization of [Ca.sup.2+.sub.i] signaling. Because mitochondria can spatially restrict cellular [Ca.sup.2+.sub.i] signals, immunocytochemical, electron microscopic, and fluorescent studies of mitochondria localization were performed in human airway epithelia. Although concentrated at the apical domain, mitochondria were found distributed at both the apical and the basolateral poles and in close association with the endoplasmic reticulum. The role of mitochondria in locally restricting P2[Y.sub.2]-R-induced [Ca.sup.2+.sub.i] signals was investigated by measuring changes in mitochondrial [Ca.sup.2+] ([Ca.sup.2+.sub.m]) in human airway epithelial monolayers. P2[Y.sub.2]-R activation induced [Ca.sup.2+.sub.m] accumulation in mitochondria confined to the domain ipsilateral to P2[Y.sub.2]-R stimulation, which was blocked by mitochondrial uncoupling with 1 [micro]M CCCP and 2.5 [micro] g/ml oligomycin. The role of mitochondria in restricting the cellular cross-talk between basolateral P2[Y.sub.2]-R-dependent [Ca.sup.2+.sub.i] mobilization and apical membrane [Ca.sup.2+]-activated [Cl.sup.-] secretion was investigated in studies simultaneously measuring [Ca.sup.2+.sub.i] and [Cl.sup.-] secretion in cystic fibrosis human airway epithelial monolayers. Activation of basolateral P2[Y.sub.2]-Rs produced similar increases in [Ca.sup.2+.sub.i] in monolayers without and with pretreatment with uncouplers, whereas [Ca.sup.2+.sub.i]-activated [Cl.sup.-] secretion was only efficiently triggered in mitochondria-uncoupled conditions. We conclude that (a) mitochondria function as a [Ca.sup.2+.sub.i]-buffering system in airway epithelia, compartmentalizing [Ca.sup.2+.sub.i]-dependent functions to the membrane ipsilateral to receptor stimulation; and (b) the mitochondria provide structural barriers that protect the airway epithelia against nonspecific activation of [Ca.sup.2+.sub.i]-modulated functions associated with [Ca.sup.2+.sub.i] signals emanating from the apical or the basolateral membrane domains. KEY WORDS: calcium signaling * mitochondria * endoplasmic reticulum * chloride secretion * purinergic receptors

Chloride concentration ([[Cl.sup.-]]) was measured in defined organellar compartments using fluorescently labeled transferrin, [[alpha].sub.2]-macroglobulin, and cholera toxin B-subunit conjugated with [Cl.sup.-]-sensitive and -insensitive dyes. In pulse-chase experiments, [[Cl.sup.-]] in Tf-labeled early/ recycling endosomes in J774 cells was 20 mM just after internalization, increasing to 41 mM over ~10 min in parallel to a drop in pH from 6.91 to 6.05. The low [[Cl.sup.-]] just after internalization (compared with 137 mM solution [[Cl.sup.-]]) was prevented by reducing the interior-negative Donnan potential. [[Cl.sup.-]] in [[alpha].sub.2]-macroglobulin-labeled endosomes, which enter a late compartment, increased from 28 to 58 mM at 1-45 min after internalization, whereas pH decreased from 6.85 to 5.20. [Cl.sup.-] accumulation was prevented by bafilomycin but restored by valinomycin. A [Cl.sup.-] channel inhibitor slowed endosomal acidification and [Cl.sup.-] accumulation by ~2.5-fold. [[Cl.sup.-]] was 49 mM and pH was 6.42 in cholera toxin B subunit-labeled Golgi complex in Vero cells; Golgi compartment [Cl.sup.-] accumulation and acidification were reversed by bafilomycin. Our experiments provide evidence that [[Cl.sup.-] is the principal counter ion accompanying endosomal and Golgi compartment acidification, and that an interior-negative Donnan potential is responsible for low endosomal [[Cl.sup.-]] early after internalization. We propose that reduced [[Cl.sup.-]] and volume in early endosomes permits endosomal acidification and [[Cl.sup.-]] accumulation without lysis.

Indicator molecules for caspase-3 activation have been reported that use fluorescence resonance energy transfer (FRET) between an enhanced cyan fluorescent protein (the donor) and enhanced yellow fluorescent protein (EYFP; the acceptor). Because EYFP is highly sensitive to proton ([H.sup.+]) and chloride ion ([Cl.sup.-]) levels, which can change during apoptosis, this indicator's ability to trace the precise dynamics of caspase activation is limited, especially in vivo. Here, we generated an [H.sup.+]- and [Cl.sup.-]--insensitive indicator for caspase activation, SCAT, in which EYFP was replaced with Venus, and monitored the spatio-temporal activation of caspases in living cells. Caspase-3 activation was initiated first in the cytosol and then in the nucleus, and rapidly reached maximum activation in 10 min or less. Furthermore, the nuclear activation of caspase-3 preceded the nuclear apoptotic morphological changes. In contrast, the completion of caspase-9 activation took much longer and its activation was attenuated in the nucleus. However, the time between the initiation of caspase-9 activation and the morphological changes was quite similar to that seen for caspase-3, indicating the activation of both caspases occurred essentially simultaneously during the initiation of apoptosis.

Research has been conducted on ability of bactria to adapt to disinfectants. The morphological and chemical exposure effects to sublethal levels of benzalkonium chloride on Listeria monocytogenes have been investigated, and the details are reported.

Research has been conducted on gram-positive moderately halophilic bacterium Halobacillus halophilus. The biological functions of chloride in this bacterium have been investigated, and the results suggest that chloride can function as a signal molecule in salt adaptation in moderately halophilic bacterium H. halophilus which may sense salinity via chloride concentration of its environment.

Research has been conducted on the aryl halides. Results demonstrate that the amination of these compounds carried out in the presence of air-stable alkali metal hydroxide bases and a palladium catalyst has produced arylamines.

Research has been conducted on the s-triazine herbicides. Results indicate that these herbicides hydrolytically displace chloride and amino substituents of 2-chloro-4,6-diamino-s-triazine.

Research has been conducted on Staphylococcus aureus cells in biofilms. The disinfectant test developed for these cells and the membrane-active compound benzalkonium chloride and oxidizing agent hypochlorite used in this test are discussed.

Research has been conducted on the alpha-sulfinylamides. The preparation of these compounds has been carried out via condensation of the primary amine with ketone and via the reaction of the resulting imine with ethylsulfenylacetyl chloride followed by the oxidation with sodium periodate.

Research has been conducted on the oxime-derived chloro-bridged palladacycles. The effect of bases, additives and solvents on the containing phenylboronic acids aryl bromide and chloride Suzuki couplings with oxime-derived palladacycles as catalysts has been investigated and the details are reported.

Research has been conducted on the allylstannanes and allylsilanes. The results of the intramolecular reaction of these compounds with alkynes carried out in the presence of Pt(II), Ru(II), Pd(II) and Au(II) chlorides are discussed.

Research has been conducted on the alcohol conversion into formate esters. Results indicate that this conversion can be carried out in methyl chloride at room temperature via the use of 2,4,6-trichloro-1,3,5-triazine and N,N-dimethylformamide in the presence of lithium fluoride.

Research has been conducted on the chiral nonracemic and racemic alpha-unsubstituted and alpha-substituted beta-fluoroalkyl beta amino acid derivatives. The synthesis of these acid derivatives has been carried our via fluorinated imidoyl chlorides and ester enolates and the results indicate that the process takes place with good diastereoselectivity.

Research has been conducted on the functional vinylstannane. The palladium-catalyzed stereoselective annulation of this compound carried out by acyl chlorides has been explored and the results indicate that this annulation proceeds via the Stille reaction/cyclization sequence.

Research has been conducted on the range of pyrazole 1-oxides. The preparation of these oxides has been carried out via the regioselective deprotonation or bromine-magnesium exchange and transmetalation with ZnCl (sub)2 and palladium(0)-catalyzed cross-coupling.

Research has been conducted on the acyl chlorides and nitriles. The insertion reaction of these compounds carried out in producing 3-oxo-1-alkenylphosphonates in high yields is described.

Research has been conducted on the air-stable palladium complexes. Results indicate that these complexes serve as catalysts for the vinyl and aryl chloride cross-coupling reactions with thiols, arylzinc reagents and arylboronic acids.

Research has been conducted on the substituted epoxy ethers. The results of the intramolecular radical cyclization of these ethers carried out via the use of bis(cyclopentadienyl)titanium(III) chloride as the radical source are presented.

Research has been conducted on the aryl chlolides and amines. The study of the nickel-catalyzed aryl amination processes carried out without NaH has demonstrated that the reaction scope could be extended to the primary and aromatic alkylamines.

Research has been conducted on the second indole sulfenylation mechanism. Results indicate that the second sulfenylation occurs via the indolenium 3,3-bis-sulfide intermediate formation followed by the sulfide group migration to the 2-position.

Research has been conducted on S-tert-butyl-beta-(trifluoromethyl)isocysteine diastereoisomers. The synthesis of these diastereoisomers has been performed via the reactions of trifluoroacetimidoyl chloride with the tert-butyl alpha-tert-butylthioacetate's lithium enolate and the details are reported.

Research has been conducted on cyclic thioacetals. The dethioacetalization of thioacetals into carbonyl compounds, ring-expansion annelation and ring-expansion chlorination of cyclic thioacetals carried out via the use of silica chloride in the presence of DMSO are discussed.

Research has been conducted on the bridgehead compounds. Results indicate that no redox catalysis is observed when these compounds bear carbonyl group as a pi acceptor.

Research has been conducted on the 3,4-cyanomethyl substituted thiophenes. The reaction of these compounds with thionyl chloride has been carried out in the presence of base in producing dicyano substituted thieno(3,4-c)thiophenes and the details are reported.

Research has been conducted on the reaction of bromine chloride with alkynes. The use of radical inhibitors in examining the rearrangements during bromination is described.

Research has been conducted on the antropisomeric biphenyl chemistry. The method for introducing an N-thiophthalimide group on the electron-rich phenols has been developed and the results are discussed.

Research has been conducted on the protio and hexadeuterated chlorides. The solvolytic behavior of these chlorides has been examined in investigating the pi-participation of the unsaturated substrate in which the double bond is at C4 from the reaction center and the details are presented.

Research has been conducted on the crown ether-containing macrobicycles. The molecular structure of these macrobicycles has been characterized via NMR spectroscopy and X-ray crystallography and the results demonstrate that the macrobicycle's chloride-binding ability is weak.

Research has been conducted on regioselective alkylation of allylic compounds. The results of the palladium-catalyzed alpha-alkylation of allylic bromides and chloride with unsubstituted and substituted benzylic Grignard reagents are reported.