cAMP-dependent chloride secretion mediates tubule enlargement and cyst formation by cultured mammalian collecting duct cells.

Polycystic kidney diseases result from disruption of the genetically defined program that controls the size and geometry of renal tubules. Cysts which frequently arise from the collecting duct (CD) result from cell proliferation and fluid secretion. From mCCD(cl1) cells, a differentiated mouse CD cell line, we isolated a clonal subpopulation (mCCD-N21) that retains morphogenetic capacity. When grown in three-dimensional gels, mCCD-N21 cells formed highly organized tubular structures consisting of a palisade of polarized epithelial cells surrounding a cylindrical lumen. Subsequent addition of cAMP-elevating agents (forskolin or cholera toxin) or of membrane-permeable cAMP analogs (CPT-cAMP) resulted in rapid and progressive dilatation of existing tubules, leading to the formation of cystlike structures. When grown on filters, mCCD-N21 cells exhibited a high transepithelial resistance as well as aldosterone- and/or vasopressin-induced amiloride-sensitive and -insensitive current. The latter was in part inhibited by Na(+)-K(+)-2Cl(-) cotransporter (bumetanide) and chloride channel (NPPB) inhibitors. Real-time PCR analysis confirmed the expression of NKCC1, the ubiquitous Na(+)-K(+)-2Cl(-) cotransporter and cystic fibrosis transmembrane regulator (CFTR) in mCCD-N21 cells. Tubule enlargement and cyst formation were prevented by inhibitors of Na(+)-K(+)-2Cl(-) cotransporters (bumetanide or ethacrynic acid) or CFTR (NPPB or CFTR inhibitor-172). These results further support the notion that cAMP signaling plays a key role in renal cyst formation, at least in part by promoting chloride-driven fluid secretion. This new in vitro model of tubule-to-cyst conversion affords a unique opportunity for investigating the molecular mechanisms that govern the architecture of epithelial tubes, as well as for dissecting the pathophysiological processes underlying cystic kidney diseases.

In vitro functionality of isolated embryonic hypothalamic vasopressinergic and oxytocinergic neurons: modulatory effects of brain-derived neurotrophic factor and angiotensin II.

There are only a few studies on the ontogeny and differentiation process of the hypothalamic supraoptic-paraventriculo-neurohypophysial neurosecretory system. In vitro neuron survival improves if cells are of embryonic origin; however, surviving hypothalamic neurons in culture were found to express small and minimal amounts of arginine-vasopressin (AVP) and oxytocin (OT), respectively. The aim of this study was to develop a primary neuronal culture design applicable to the study of magnocellular hypothalamic system functionality. For this purpose, a primary neuronal culture was set up after mechanical dissociation of sterile hypothalamic blocks from 17-day-old Sprague-Dawley rat embryos (E17) of both sexes. Isolated hypothalamic cells were cultured with supplemented (B27)-NeuroBasal medium containing an agent inhibiting non-neuron cell proliferation. The neurosecretory process was characterized by detecting AVP and OT secreted into the medium on different days of culture. Data indicate that spontaneous AVP and OT release occurred in a culture day-dependent fashion, being maximal on day 13 for AVP, and on day 10 for OT. Interestingly, brain-derived neurotrophic factor (BDNF) and Angiotensin II (A II) were able to positively modulate neuropeptide output. Furthermore, on day 17 of culture, non-specific (high-KCl) and specific (Angiotensin II) stimuli were able to significantly (P < 0.05) enhance the secretion of both neuropeptides over respective baselines. This study suggests that our experimental design is useful for the study of AVP- and OT-ergic neuron functionality and that BDNF and A II are positive modulators of embryonic hypothalamic cell development.

Simultaneous Optical Recording in Multiple Cells by Digital Holographic Microscopy of Chloride Current Associated to Activation of the Ligand-Gated Chloride Channel GABA(A) Receptor.

Chloride channels represent a group of targets for major clinical indications. However, molecular screening for chloride channel modulators has proven to be difficult and time-consuming as approaches essentially rely on the use of fluorescent dyes or invasive patch-clamp techniques which do not lend themselves to the screening of large sets of compounds. To address this problem, we have developed a non-invasive optical method, based on digital holographic microcopy (DHM), allowing monitoring of ion channel activity without using any electrode or fluorescent dye. To illustrate this approach, GABA(A) mediated chloride currents have been monitored with DHM. Practically, we show that DHM can non-invasively provide the quantitative determination of transmembrane chloride fluxes mediated by the activation of chloride channels associated with GABA(A) receptors. Indeed through an original algorithm, chloride currents elicited by application of appropriate agonists of the GABA(A) receptor can be derived from the quantitative phase signal recorded with DHM. Finally, chloride currents can be determined and pharmacologically characterized non-invasively simultaneously on a large cellular sampling by DHM.

Ammonium chloride as nitrogen source in sugarcane harvested without burning

The great difficulty of incorporation of N fertilizers into the "green sugarcane" system causes concern and since urea is the most commonly used source, there is the risk of loosing NH3 through volatilization. For this reason, a field experiment was undertaken (in a Hapludox Typic) with the objective of evaluating the agronomic efficiency of ammonium chloride on stubble of the second ratoon (SP89 1115), as well as its residual effect on the subsequent cycle (third ratoon). The experimental design was randomized blocks with four replications. Treatments consisted of three N rates (60, 120 and 180 kg ha-1 N) in the form of NH4Cl, in addition to a control treatment without the addition of N fertilizer. The ratoon cane of the second cutting was harvested in November 2006 and the treatments were applied in December 2006. The second ratoon was harvested mechanically in November 2007 and in December 2007, 450 kg ha-1 of the NPK mixture 20-05-19 was applied, providing 90, 22 and 86 kg ha-1 N, P2O5 and K2O, respectively, for the purpose of evaluating the effect of residual-N from the treatments implanted in December 2006. An increase in the rates of N-NH4Cl had a positive effect on the leaf concentrations of P, Mg and S. Stalk yield (MSS - Mg ha-1 of sugarcane stalks) and sugar (MSH - Mg ha-1 of sucrose) in the November 2006 harvest responded linearly to the increase of N doses in the form of NH4Cl. In relation to the effect of residual-N in the 2007/2008 harvest, it was observed, in general, that the concentrations of macronutrients in the sugarcane leaf +1 were within the range considered adequate in the state of São Paulo, Brazil. The residual-N of the NH4Cl doses resulted in a significant reduction in stalk (MSS) and sugar (MSH) production. It may be concluded that the NH4Cl source at a dose of 120 kg ha-1 N in ratoon fertilization of the second cutting was agronomically efficient, presenting, however, less efficiency of residual-N in the subsequent cycle.

Relação entre o alumínio extraível com KCl e oxalato de amônio e a mineralogia da fração argila, em solos ácidos brasileiros

Em solos ácidos de ambiente mais tropical com esmectitas, assim como em solos altamente tamponados de ambiente subtropical, contendo esmectita com hidroxi-Al entrecamadas (EHE) e, ou, vermiculita com hidroxi-Al entrecamadas (VHE), os teores de Al-KCl podem ser excepcionalmente altos; entretanto, em alguns casos não se manifestam efeitos fitotóxicos do elemento nas culturas. O Al "trocável" é tradicionalmente quantificado no extrato da solução de KCl 1 mol L-1 (Al-KCl), mas nem sempre esse elemento provém unicamente de formas trocáveis. Este trabalho objetivou investigar relações entre o Al extraído com solução de KCl e de oxalato de amônio com a mineralogia da fração argila. A quantificação do Al nos extratos de KCl e oxalato de amônio foi feita, respectivamente, por titulação com NaOH 0,02 mol L-1 padronizado e por espectrofotometria de absorção atômica. Foram utilizadas amostras de dois horizontes (A e B) de 12 perfis de solo de cinco estados brasileiros (AC, PE, BA, RS e SC), com diferentes características mineralógicas, todos com teores de Al-KCl superior a 4 cmol c kg-1 no horizonte B. Dois perfis de SC (Rancho Queimado e Curitibanos), com níveis mais baixos de Al-KCl, foram incluídos para comparação. Os altos teores de Al-KCl evidenciaram-se relacionados com a mineralogia dos solos estudados. Nos solos ácidos com mais esmectitas, drenagem moderada ou imperfeita e oscilação do lençol freático houve evidências morfológicas, confirmadas pelas análises mineralógicas, de que no clima atual ocorre um processo de destruição de argilas, liberando Al que precipita como compostos amorfos. A alta concentração salina da solução de KCl dissolve parcialmente tais compostos, superestimando as formas trocáveis desse elemento, principalmente nos horizontes subsuperficiais. Nos solos ácidos do ambiente subtropical, os teores de Al-KCl também foram altos, embora com valores mais baixos do que nos com predomínio de esmectitas. Parte do Al-KCl nesses solos parece provir da dissolução de compostos orgânicos e de Al presente em entrecamadas de argilominerais 2:1. Os resultados demonstraram que o KCl 1 mol L-1 não foi adequado para estimar o Al "trocável" na maioria dos solos estudados.

Effect of manganese chloride on the neurochemical profile of the rat hypothalamus.

Manganese (Mn(2+))-enhanced magnetic resonance imaging studies of the neuronal pathways of the hypothalamus showed that information about the regulation of food intake and energy balance circulate through specific hypothalamic nuclei. The dehydration-induced anorexia (DIA) model demonstrated to be appropriate for studying the hypothalamus with Mn(2+)-enhanced magnetic resonance imaging. Manganese is involved in the normal functioning of a variety of physiological processes and is associated with enzymes contributing to neurotransmitter synthesis and metabolism. It also induces psychiatric and motor disturbances. The molecular mechanisms by which Mn(2+) produces alterations of the hypothalamic physiological processes are not well understood. (1)H-magnetic resonance spectroscopy measurements of the rodent hypothalamus are challenging due to the distant location of the hypothalamus resulting in limited measurement sensitivity. The present study proposed to investigate the effects of Mn(2+) on the neurochemical profile of the hypothalamus in normal, DIA, and overnight fasted female rats at 14.1 T. Results provide evidence that γ-aminobutyric acid has an essential role in the maintenance of energy homeostasis in the hypothalamus but is not condition specific. On the contrary, glutamine, glutamate, and taurine appear to respond more accurately to Mn(2+) exposure. An increase in glutamine levels could also be a characteristic response of the hypothalamus to DIA.

Neutral atoms in ionic lattices: Excited states of KCl:Ag(0)

The optical-absorption spectrum of a cationic Ag0 atom in a KCl crystal has been studied theoretically by means of a series of cluster models of increasing size. Excitation energies have been determined by means of a multiconfigurational self-consistent field procedure followed by a second-order perturbation correlation treatment. Moreover results obtained within the density-functional framework are also reported. The calculations confirm the assignment of bands I and IV to transitions of the Ag-5s electron into delocalized states with mainly K-4s,4p character. Bands II and III have been assigned to internal transitions on the Ag atom, which correspond to the atomic Ag-4d to Ag-5s transition. We also determine the lowest charge transfer (CT) excitation energy and confirm the assignment of band VI to such a transition. The study of the variation of the CT excitation energy with the Ag-Cl distance R gives additional support to a large displacement of the Cl ions due to the presence of the Ag0 impurity. Moreover, from the present results, it is predicted that on passing to NaCl:Ag0 the CT onset would be out of the optical range while the 5s-5p transition would undergo a redshift of 0.3 eV. These conclusions, which underline the different character of involved orbitals, are consistent with experimental findings. The existence of a CT transition in the optical range for an atom inside an ionic host is explained by a simple model, which also accounts for the differences with the more common 3d systems. The present study sheds also some light on the R dependence of the s2-sp transitions due to s2 ions like Tl+.

A novel in vitro metabolomics approach for neurotoxicity testing, proof of principle for methyl mercury chloride and caffeine

There is a need for more efficient methods giving insight into the complex mechanisms of neurotoxicity. Testing strategies including in vitro methods have been proposed to comply with this requirement. With the present study we aimed to develop a novel in vitro approach which mimics in vivo complexity, detects neurotoxicity comprehensively, and provides mechanistic insight. For this purpose we combined rat primary re-aggregating brain cell cultures with a mass spectrometry (MS)-based metabolomics approach. For the proof of principle we treated developing re-aggregating brain cell cultures for 48h with the neurotoxicant methyl mercury chloride (0.1-100muM) and the brain stimulant caffeine (1-100muM) and acquired cellular metabolic profiles. To detect toxicant-induced metabolic alterations the profiles were analysed using commercial software which revealed patterns in the multi-parametric dataset by principal component analyses (PCA), and recognised the most significantly altered metabolites. PCA revealed concentration-dependent cluster formations for methyl mercury chloride (0.1-1muM), and treatment-dependent cluster formations for caffeine (1-100muM) at sub-cytotoxic concentrations. Four relevant metabolites responsible for the concentration-dependent alterations following methyl mercury chloride treatment could be identified using MS-MS fragmentation analysis. These were gamma-aminobutyric acid, choline, glutamine, creatine and spermine. Their respective mass ion intensities demonstrated metabolic alterations in line with the literature and suggest that the metabolites could be biomarkers for mechanisms of neurotoxicity or neuroprotection. In addition, we evaluated whether the approach could identify neurotoxic potential by testing eight compounds which have target organ toxicity in the liver, kidney or brain at sub-cytotoxic concentrations. PCA revealed cluster formations largely dependent on target organ toxicity indicating possible potential for the development of a neurotoxicity prediction model. With such results it could be useful to perform a validation study to determine the reliability, relevance and applicability of this approach to neurotoxicity screening. Thus, for the first time we show the benefits and utility of in vitro metabolomics to comprehensively detect neurotoxicity and to discover new biomarkers.

Effects of sodium sulphate and potassium chloride fertilizers on the nutritive value of timothy grown on different soils

Selostus: Natrium- ja kaliumlannoituksen vaikutus timotein ravintoarvoon

Studies on Soil-Aggregate-Sodium Chloride Stabilized Roads in Franklin County, Iowa, HR-33

This investigation was conducted to study the performance characteristics of low cost roadway surfaces of soil-aggregate-sodium chloride mixtures. Many roads have been successfully stabilized with sodium chloride. However, little information is available on either the properties of the road materials or the effects of sodium chloride on the materials. The performance of some of the sodium chloride stabilized roads in Franklin County, Iowa, and the performance of some near-by nonchemically treated roads has been studied. The study of sodium chloride stabilized roads was restricted to the roads in which the binder soil used in construction came from the same source. The effects of sodium chloride on some of the engineering properties of the soil and soil-aggregate mixtures used were studied in the laboratory.

Lime or Chloride Treatment of Granular Base Course Materials, HR-99

Results are presented of triaxial testing of three crushed limestones to which either hydrated high-calcium lime, sodium chloride or calcium chloride had been added. Lime was added at rates of 1, 3, 10 and 16 percent, chlorides were added at 0.5 percent rate only. Speciments were compacted using vibratory compaction apparatus and were tested in triaxial compression using lateral pressures from 10 to 100 psi. Triaxial test results indicate that: (1) sodium chloride slightly decreased the angle of internal friction and increased cohesion, (2) calcium chloride slightly increased the angle of internal friction and decreased cohesion, and (3) lime had no appreciable effect on angle of internal friction but increased cohesion, decreased density and increased pore water pressure.

Evaluation of Rapid Determination of the Chloride Permeability of Portland Cement Concrete by AASHTO T277-83, MLR-86-11, 1987

An Iowa D.O.T. Laboratory built machine was constructed for the chloride permeability testing of concrete by measuring electric current through a specimen between a salt solution and a base solution. This study had two purposes. The first was to evaluate the machine's performance. To do this, three concrete mixes were made consisting of different cement factors and water/cement ratios. Each mix was tested for chloride ion content by the 90- day salt ponding method and for chloride permeability at a 28-day cure by the permeability machine. The results from each test were evaluated to see if there was correlation between chloride ion content and the chloride permeability. It was determined that there was a correlation and that the permeability machine was satisfactory for determining chloride permeability in concrete. The second purpose of this study was to examine the effects that pozzolans have on the chloride permeability of concrete. Four mixes were made: one without any pozzolans as a control, one with class C fly ash, one with class F fly ash, and one with silica fume. Specimens from each mix were evaluated for chloride ion content by the 90-day salt ponding test and by the laboratory built machine for chloride permeability after curing 28 days. Specimens from these mixes were also taken from the salt ponding slabs after completion of the ponding test to examine the effect chloride ion content has on the operation of the chloride permeability machine. Specimens containing pozzolans were also examined for chloride permeability after a cure of 180 days. It was determined that the addition of pozzolans to concrete lowers the chloride permeability as measured by the permeability machine. Class F fly ash and silica fume in the concrete had a major effect in lowering the chloride permeability in concrete as measured by the permeability machine.

Chlormequat chloride and ethephon affect growth and yield formation of conventional, naked and dwarf oat

Selostus: Kasvunsääteiden vaikutukset tavanomaisen, paljasjyväisen ja kääpiökauran kasvuun ja sadonmuodostukseen

Chloride Penetration Into LSDC (Iowa System) Resurfacing Mixes, MLR-77-05 and R-267, 1977

The purpose of this investigation was to determine the comparative effectiveness of standard D-57 concrete and Iowa system Low Slump Dense Concrete in preventing threshold levels of chloride from penetrating the concrete slabs to the reinforcing steel.