Radioactive Chloride Research Articles

Evaluation of anti-inflammatory diphenyldihaloketone EF24 in transient ischemic stroke model

ABSTRACT Objectives Revascularization is necessary in patients with ischemic stroke, however it does not address inflammation that contribute to reperfusion injury and the early growth of ischemic core. We investigated EF24, an anti-inflammatory agent, in a stroke model. Methods Ischemic stroke was induced in mice by occluding middle cerebral artery for 1 h followed by reperfusion. EF24 was given either 10 min post-reperfusion (EF24Post) or 10 min before occlusion (prophylactic, EF24Pro). Survival, ipsilateral uptake of radioactive infarct marker 18F-fluoroglucaric acid (FGA), inflammatory cytokines, and tetrazolium chloride (TTC) staining were assessed. Results Survival was increased in both EF24-treated groups compared to the stroke+vehicle group. Ipsilateral 18F-FGA uptake increased 2.6-fold in stroke+vehicle group compared to sham group (p < 0.05); the uptake in EF24-treated groups and sham group was not significantly different. TTC-staining also showed reduction in infarct size by EF24 treatment. Plasma IL-6, TNF-α, and corticosterone did not show significant changes among groups. However, ipsilateral tissue in stroke+vehicle mice showed increased IL-6 (>90-fold) and TNF-α (3-fold); the tissue IL-6 and TNF-α were significantly reduced in stroke+EF24Pro and stroke+EF24Post groups. 18F-FGA uptake significantly correlated with tissue IL-6 levels. Conclusions EF24 controls infarct growth and suppresses tissue inflammation in ischemic stroke, which can be monitored by 18F-FGA uptake.

Dechlorination of molten chloride waste salt from electrorefining via ion-exchange using pelletized ultra-stable H-Y zeolite in a fluidized particle reactor

Dechlorination of eutectic LiCl–KCl based electrorefiner (ER) salt is reported via ion-exchange reaction with protonated ultrastable Y-type (USHY) zeolite bound into mechanically fluidized 45–250 μm diameter particles. Evidence of exchange of cations from the salt (Li+, K+, and fission product cations) into the zeolite lattice replacing H+ ions was found based on a change in unit cell size, ICP-MS, XRD and TEM–EDS in addition to detection of HCl off gas. Ion exchange reaction was carried out at 625 and 650 °C, temperatures above the melting point of eutectic LiCl–KCl. Experiments were carried out to optimize zeolite drying temperature, estimate maximum ion-exchange capacity, and determine the thermal stability of USHY zeolite. The results indicate over 90% dechlorination can be achieved without zeolite structure collapse at 625 °C. This provides a promising route to stabilizing waste from radioactive chloride salts into dechlorinated waste forms for permanent geologic disposal.

Thermal Stability of Glass with Simulators of Chloride Highly Radioactive Wastes

High temperature behavior of sodium–aluminum fluorophosphate glass—a potential matrix for immobilisation of waste salt electrolyte from pyrochemical reprocessing of irradiated nuclear fuel (INF) has been studied. The glass crystallizes between 430 and 640°C with formation of phosphate phases, which dissolve above 640°C leading to homogenization of the glass. Similar transformations of the glass matrix due to heating from the decay of short-lived fission products may have a negative effect on properties of vitrified radioactive chloride wastes during storage in a repository.

Radiolabeling of aromatic compounds using K[*Cl]Cl and OXONE®

We report a novel labeling method for the rapid radiochlorination of aromatic compounds using no-carrier-added short-half-life radioactive chloride ions ([38,39Cl]Cl− and [34mCl]Cl−) and OXONE® (Sigma-Aldrich, Japan) as an oxidation reagent. The reaction of radioactive chloride ions with anisole in the presence of OXONE gave a mixture of p-[*Cl]chloroanisole and o-[*Cl]-chloroanisole, with a radiochemical conversion yield of more than 85% (not decay corrected) and selectivity of 7:3, within 10 min. Regioselective radioactive chlorination was achieved by chlorodestannylation of tributylstannyl compounds to afford the corresponding radiochlorinated compounds (p-chloroanisole and m-chloroanisole) in satisfactory radiochemical yields (80–95% and 46–65%, respectively). Copyright © 2012 John Wiley & Sons, Ltd.

Sensitivity of Chloride Efflux vs. Transepithelial Measurements in Mixed CF and Normal Airway Epithelial Cell Populations

Background/Aims: While the Cl^- efflux assays are relatively straightforward, their ability to assess the efficacy of phenotypic correction in cystic fibrosis (CF) tissue or cells may be limited. Accurate assessment of therapeutic efficacy, i.e., correlating wild type CF transmembrane conductance regulator (CFTR) levels with phenotypic correction in tissue or individual cells, requires a sensitive assay. Methods: Radioactive chloride (³⁶Cl) efflux was compared to Ussing chamber analysis for measuring cAMP-dependent Cl^- transport in mixtures of human normal (16HBE14o-) and cystic fibrosis (CF) (CFTE29o- or CFBE41o-, respectively) airway epithelial cells. Cell mixtures with decreasing amounts of 16HBE14o- cells were evaluated. Results: Efflux and Ussing chamber studies on mixed populations of normal and CF airway epithelial cells showed that, as the number of CF cells within the population was progressively increased, the cAMP-dependent Cl^- decreased. The ³⁶Cl efflux assay was effective for measuring Cl^- transport when ≥ 25% of the cells were normal. If < 25% of the cells were phenotypically wild-type (wt), the ³⁶Cl efflux assay was no longer reliable. Polarized CFBE41o- cells, also homozygous for the ΔF508 mutation, were used in the Ussing chamber studies. Ussing analysis detected cAMP-dependent Cl^- currents in mixtures with ≥1% wild-type cells indicating that Ussing analysis is more sensitive than ³⁶Cl efflux analysis for detection of functional CFTR. Conclusions: Assessment of CFTR function by Ussing analysis is more sensitive than ³⁶Cl efflux analysis. Ussing analysis indicates that cell mixtures containing 10% 16HBE14o- cells showed 40-50% of normal cAMP-dependent Cl^- transport that drops off exponentially between 10-1% wild-type cells.

Male transgenic glycine receptor α 1 (S267Q) mutant mice display a hyperekplexia-like increase in acoustic startle responses

Glycine receptors (GlyR) are ligand-gated ion channels that inhibit neurotransmission in the spinal cord and brainstem, and mutations in GlyR can cause the human disease hyperekplexia, which is characterized by elevated startle responses. Recently, the GlyR α 1S267Q mutation was shown to disrupt normal GlyR function, and knock-in mice harboring this mutation displayed profoundly increased acoustic startle responses and reduced glycine-stimulated the chloride flux [Findlay, G.S., Phelan, R., Roberts, M.T., Homanics, G.E., Bergeson, S.E., Lopreato, G.F., Mihic, S.J., Blednov, Y.A., Harris, R.A. 2003. Glycine receptor knock-in mice and hyperekplexia: comparisons with the null mutant. J Neurosci 23, 8051–8059.]. In this study, a transgenic mouse model expressing this S267Q mutation was evaluated using similar techniques to determine if these mice are similarly affected. Male transgenic mice displayed increased acoustic startle responses. However, decreases in glycine-stimulated strychnine-sensitive radioactive chloride ( 36Cl −) uptake were not observed in spinal cord and brainstem synaptoneurosomes from transgenic mice. No changes in habituation or prepulse inhibition of startle responses or spontaneous locomotion in response to taurine were observed as a result of presence of the transgene. Consistent with previous studies using immunoblotting and strychnine binding [Findlay, G.S., Wick, M.J., Mascia, M.P., Wallace, D., Miller, G.W., Harris, R.A., Blednov, Y.A. 2002. Transgenic expression of a mutant glycine receptor decreases alcohol sensitivity of mice. J Pharmacol Exp Ther 300, 526–534.], the glycine-stimulated strychnine-sensitive chloride flux of cortical microsacs in transgenic mice confirmed the ectopic expression of transgenic GlyR. These results support both the idea that transgenic expression of the S267Q mutation produces a less dramatic phenotype as compared to the knock-in mouse model as well as the idea that the in vivo acoustic startle test (as compared to the in vitro chloride flux assay) is particularly sensitive to disruptions in GlyR function.

Glycine-gated chloride channels in neutrophils attenuate calcium influx and superoxide production.

Recently, it was demonstrated that liver injury and TNF-alpha production as a result of endotoxin (lipopolysaccharide, LPS) were attenuated by feeding animals a diet enriched with glycine. This phenomenon was shown to be a result of, at least in part, activation of a chloride channel in Kupffer cells by glycine, which hyperpolarizes the cell membrane and blunts increases in intracellular calcium concentrations ([Ca(2+)](i)) similar to its action in the neuron. It is well known that hepatotoxicity due to LPS has a neutrophil-mediated component and that activation of neutrophils is dependent on increases in [Ca(2+)](i). Therefore, the purpose of this study was to determine if glycine affected agonist-induced increases in [Ca(2+)](i) in rat neutrophils. The effect of glycine on increases in [Ca(2+)](i) elicited either by the bacterial-derived peptide formyl-methionine-leucine-phenylalanine (FMLP) or LPS was studied in individual neutrophils using Fura-2 and fluorescence microscopy. Both FMLP and LPS caused dose-dependent increases in [Ca(2+)](i), which were maximal at 1 microM FMLP and 100 microgram/ml LPS, respectively. LPS increased intracellular calcium in the presence and absence of extracellular calcium. Glycine blunted increases in [Ca(2+)](i) in a dose-dependent manner with an IC(50) of approximately 0.3 mM, values only slightly higher than plasma levels. Glycine was unable to prevent agonist-induced increases in [Ca(2+)](i) in chloride-free buffer. Moreover, strychnine (1 microM), an antagonist of the glycine-gated chloride channel in the central nervous system, reversed the effects of glycine (1 mM) on FMLP- or LPS-stimulated increases in [Ca(2+)](i). To provide hard evidence for a glycine-gated chloride channel in the neutrophil, the effect of glycine on radioactive chloride uptake was determined. Glycine caused a dose-dependent increase in chloride uptake into neutrophils with an ED(50) of approximately 0.4 mM, an effect also prevented by 1 microM strychnine. Glycine also significantly reduced the production of superoxide anion from FMLP-stimulated neutrophils. Taken together, these data provide clear evidence that neutrophils contain a glycine-gated chloride channel that can attenuate increases in [Ca(2+)](i) and diminish oxidant production by this important leukocyte.

Measurement of glycine receptor function by radioactive chloride uptake

The glycine receptor is an important inhibitory receptor for both spinal and supraspinal functions, and mutations in receptor subunits are responsible for neurological disorders in several species, including humans. However, functional measurements of glycine receptors have generally been restricted to electrophysiological analysis of immature, cultured neurons. We developed a 36Cl − flux assay to measure glycine receptor function using membrane vesicles from spinal cord and brainstem of adult mice. The uptake of 36Cl − stimulated by glycine was characterized by a glycine EC 50 of 22 μM for the major component and an EC 50 of 0.5 μM for a minor component. Strychnine inhibited the glycine-stimulated uptake with an IC 50 of 0.4 μM. The uptake was not affected by picrotoxin, bicuculline, or pentobarbital. Glycine-stimulated uptake reached a maximum by 10 s. This technique should prove useful for genetic and pharmacological analysis of the function of glycine receptors.

Calcium distribution in soybean during seed‐filling in relation to moisture stress

Abstract The distribution of calcium (Ca) within stem, leaves, and seeds of soybean was studied with an infused radioactive calcium chloride (45CaCl2), solution during seed‐filling under well‐watered and water stressed regimes in a greenhouse. The kinetics of infusion volume showed a quadratic reduction in absorption which was reduced to zero on the sixth day for non‐irrigated plants. The concentration of 45Ca increased quadratically from the point of injection towards the apex for both water status and plant parts. The difference in concentration of 45Ca between irrigated and non‐irrigated plants was significant (p<0.05) and concentrations attained maximum values at the sixth node from the plant base. Seeds contained considerably less 45Ca than either stem or leaves.

Chloride absorption and distribution in chlorine-deficient Pharbitis nil

Determination and distribution of radioactive chloride in Pharbitis nil was investigated by a bio-imaging analyzer. When leaves that contained various amounts of 36Cl were analyzed with the imaging analyzer and then each sample was homogenized and its radioactivity measured in a liquid scintillation counter, radioactive levels recorded by the analyzer were directly proportional to the radioactivity determined by the counter. When plants that had been grown in full nutrient solution were incubated in [36Cl]-containing solution, more activity was found in young leaves than in mature leaves, while little radioactivity was detected in shrivelled leaves and the nonsymptomatic cusp of young leaves of plants that had been grown in chlorine-deficient solution.

The GABAA receptor channel mediated chloride ion translocation through the plasma membrane: new insights from 36Cl- ion flux measurements.

GABAA receptors in plasma membranes of neurons are integral oligomers which form chloride channels. The binding of GABA molecules at recognition sites for channel opening triggers a transient increase in transmembrane chloride ion flux. The multiplicity and drug specificity of GABAA receptor, kinetics of channel opening, and desensitization of GABAA receptor and its short- and long-term regulation have been investigated by the use of tracer amounts of the radioactive chloride isotope, 36Cl- ion. Results and new insights from 36Cl- ion flux measurements have been reviewed.

Slow release of chloride from 36Cl-labeled photosystem II membranes

Photosystem II (PS II) membrane fragments isolated from spinach cultured on a medium containing 36Cl − retain a significant amount of radioactive chloride corresponding to at least one Cl − per PS II unit. The release of chloride from these sites occurs with halftimes of several hours. Treatments which cause the release of the manganese and/or the extrinsic proteins also result in the loss of chloride from these sites, indicating a possible role for this chloride in photosynthetic oxygen evolution.

Kinetics of isotopic exchange between chloramine-B and radioactive chloride

The kinetics of chlorine isotope exchange between chloramine-B /CAB/ and chloride has been studied using ion-exchange separation and tracer technique. McKay's plot are linear. The exchange reaction is fast in acidic medium, very slow in neutral medium and does not take place in alkaline medium. In the acidic range the exchange is maximum at pH 3.3. The rate of exchange decreases at pH >3.3 and 5. Addition of neutral salt or parent compound has no effect on the rate of exchange. Activation energy and activation entropy for this exchange reaction have been calculated.

Ion-adsorption by specialized epithelial regions in Musca domestica larvae

In experiments using labelled chloride solutions in combination with a histochemical chloride precipitation technique, and a combination of the latter technique with autoradiography it has been shown that the semiaquatic larvae of the terrestrial dipteran, Musca domestica, adsorb chloride ions from the external medium specifically by well-defined, specialized epithelial regions. By the use of radioactive chloride in extremely hypotonic concentrations applied to normal larvae and to larvae which were ligated just behind their mouth, it was demonstrated that this adsorption occurs even in highly diluted, hypotonic solution. These experiments simultaneously verify the silver chloride nature of the precipitates caused by histochemical treatment with silver nitrate exclusively along the species-specific, cutaneous differentiation. With regard to these facts, the designation of those specialized epithelial structures of Musca domestica larvae as chloride cells and anal chloride epithelia in accordance with comparable differentiations in other insect species proved to be absolutely correct. The results in conjunction with supporting preliminary data of ultrastructural features suggest that these epithelial structures are involved in osmoregulation by active ion transport.

Anion movements across lamprey ( Lampetra fluviatilis) red cell membrane

Chloride and bicarbonate movements across lamprey red cell membrane were investigated. The halftime for equilibration of radioactive chloride across the red cell membrane was 2.46 h, and apparent permeability for chloride-36 was approximately 10 −9 cm · s −1, a value similar to that observed for lipid bilayers. Chloride movements were not affected by the anion exchange inhibitor, 4,4′-diisothiocyano-stilbene-2,2′-disulfonic acid (DIDS). Furthermore, intracellular buffering is effectively isolated from the extracellular compartment, as shown by the fact that practically no pH recovery occurred in the unbuffered extracellular medium after either acidification or alkalinization. These observations show that lamprey red cell membrane is quite impermeable to bicarbonate and other acid/base equivalents.

Membrane Transport in Isolated Vesicles from Sugarbeet Taproot

The effects of fluoride on the tonoplast type ATPase and transport activities associated with sealed membrane vesicles isolated from sugarbeet (Beta vulgaris L.) storage tissue were examined. This anion had two distinct effects upon the proton-pumping vesicles. When ATP hydrolysis was measured in the presence of gramicidin D, significant inhibition (approximately 50%) only occurred when the fluoride concentration approached 50 millimolar. In contrast, the same degree of inhibition of proton transport occurred when the fluoride concentration was about 24 millimolar. Effects on proton pumping at this concentration of fluoride could be attributed to an inhibition of chloride movement which serves to dissipate the vesicle membrane potential. Valinomycin could partially restore ATPase activity in sealed vesicles which were inhibited by fluoride and this restoration occurred with a reduction in the membrane potential. Fluoride demonstrated a competitive interaction with chloride-stimulation of proton transport and inhibited the uptake of radioactive chloride into sealed vesicles. When the vesicles were allowed to develop a pH gradient in the absence of KCl, and KCl was subsequently added, fluoride reduced enhancement of the existing pH gradient by KCl. The results are consistent with a chloride carrier that is inhibited by fluoride.

Kinetics of cellular uptake of tracers used in myocardial scintigraphy.

The purpose of this study was to obtain more information about the role of the plasma membrane in cellular uptake of tracers used in myocardial scintigraphy (radioactive thallium chloride and palmitic acid). This knowledge is of importance for an adequate interpretation of the results obtained under clinical conditions.

Effects of arginine vasopressin on the thin ascending limb of Henle's loop of hamsters.

Arginine vasopressin (AVP) has been shown to stimulate active Cl transport across the medullary thick ascending limb of Henle's loop (MAL) in association with an increase in adenylate cyclase activity. To determine whether the failure to demonstrate active Cl transport across the thin ascending limb of Henle's loop (TAL) in previous in vitro perfusion studies was due to the absence of AVP in the preparation, we examined the effect of AVP on adenylate cyclase activity and Cl transport in the hamsters TAL. AVP (1 mU/ml) increased adenylate cyclase activity in the hamster TAL (20.7 +/- 5.2 control vs. 46.2 +/- 10.1 fmol . mm-1 . 30 min-1, n = 6, P less than 0.05) but not in the descending limb (27.8 +/- 7.0 control vs. 20.4 +/- 2.7, n = 4, P less than 0.05). When both MAL and TAL were perfused, a lumen-positive transepithelial voltage (Vt) was observed. The Vt was increased by adding 1 or 10 mU/ml AVP to the bath. When only the TAL was perfused, the Vt was not different from zero. Similar results were obtained in mouse renal tubules. In other experiments, AVP did not affect the diffusion potential generated when a transepithelial NaCl gradient was present. AVP or dibutyryl cAMP caused little or no change in efflux of radioactive chloride across the hamster TAL. These findings suggest that electrogenic chloride transport is not demonstrable in the TAL even in the presence of AVP. The physiologic role of AVP-sensitive adenylate cyclase in the TAL remains to be established.

Radiotracer studies on ion-selective electrode membranes containing metal complexes of a nonylphenoxypoly(ethyleneoxy)ethanol in poly(vinyl chloride) matrices

Radiotracer studies with 133Ba, 45Ca and 36Cl are reported for PVC matrix membranes containing 2-nitrophenyl phenyl ether and the tetraphenylborate of a barium (or calcium) complex with a nonylphenoxypoly(ethyleneoxy)ethanol (NP), Antarox CO880. The results show that there is very limited permeation of radioactive barium and calcium ions through the membrane systems. However, the continued uptake (with time) of radioactive barium ions by the membranes (the uptake of calcium ion is less) suggests that in relation to selective electrode response the stabilization of ions by the NP ligand within the membrane is important in addition to the simple availability of membrane pathways for primary-ion transport through the membrane. Permselectivity to counter-ions is indicated by the non-permeation of radioactive chloride ions.

Methylmercury sexual dimorphism in the mouse.

5–8 days after a single nontoxic dose of radioactive methylmercury chloride adult male mouse kidneys contain twice as much radioactive mercury per unit wet wt as do kidneys of similarly dosed adult females.