Adult weight management across the community: population-level impact of the LOSE IT to WIN IT challenge.

Objective: Excess body weight negatively impacts health, but there are few evaluations of low-intensity weight management challenge programs in defined populations. This study examined weight change in adults who participated in the LOSE IT to WIN IT (LIWI) health challenge in a US community. The community-level impact on body mass index was also explored. Methods: Body weight was analysed over 1 year in the cohort of LIWI enrolees, stratified by participants who were healthy weight or overweight/obese at baseline. Secondarily, a multiple cross-sectional analysis compared the 2.5-year trends in body mass index between community adults who did vs. did not participate in LIWI. Results: LOSE IT to WIN IT participants who were overweight/obese lost a mean (95% confidence interval) 1.6 (1.2, 2.0) kg (~2%) over 1 year (p < 0.001), whereas healthy weight participants lost 0.7 (0.3, 1.1) kg. Across the community, LIWI participants and non-participants both gained 0.4 kg m-2 over the 2.5-year study period (p = 0.884). Conclusions: LOSE IT to WIN IT was modestly effective among enrolees, resulting in a small weight loss of 2% over 1 year among those who were overweight/obese. However, LIWI did not impact weight gain in the community. To slow such community-level weight gain trends, weight management challenges must reach larger fractions of the populations that they target.

Syntaxin 4 is concentrated on plasma membrane of astrocytes.

Syntaxins are a family of transmembrane proteins that participate in SNARE complexes to mediate membrane fusion events including exocytosis. Different syntaxins are thought to participate in exocytosis in different compartments of the nervous system such as the axon, the soma/dendrites or astrocytes. It is well known that exocytosis of synaptic vesicles at axonal presynaptic terminals involves syntaxin 1 but distributions of syntaxins on neuronal somal and dendritic, postsynaptic or astroglial plasma membranes are less well characterized. Here, we use pre-embedding immunogold labeling to compare the distribution of two plasma membrane-enriched syntaxins (1 and 4) in dissociated rat hippocampal cultures as well as in perfusion-fixed mouse brains. Comparison of Western blots of neuronal cultures, consisting of a mixture of hippocampal neurons and glia, with glial cultures, consisting of mostly astrocytes, shows that syntaxin 1 is enriched in neuronal cultures, whereas syntaxin 4 is enriched in glial cultures. Electron microscopy (EM)-immunogold labeling shows that syntaxin 1 is most abundant at the plasma membranes of axons and terminals, while syntaxin 4 is most abundant at astroglial plasma membranes. This differential distribution was evident even at close appositions of membranes at synapses, where syntaxin 1 was localized to the plasma membrane of the presynaptic terminal, including that at the active zone, while syntaxin 4 was localized to nearby peri-synaptic astroglial processes. These results show that syntaxin 4 is available to support exocytosis in astroglia.

In situ metal imaging and Zn ligand-speciation in a soil-dwelling sentinel: complementary electron microprobe and synchrotron microbeam X-ray analyses.

Understanding the relationships between accumulated metal speciation in cells and tissues of ecologically significant taxa such as earthworms will improve risk assessments. Synchrotron-based µ-focus X-ray spectroscopy was used to detect, localize, and determine ligand-speciation of Zn and Pb in thin sections of two epigeic earthworm species collected from a Pb/Zn-mine soil. The findings indicated that Zn and Pb partition predominantly as typical hard acids (i.e., strong affinities for O-donors) within liverlike chloragocytes. Moreover, Zn speciation was very similar in the chloragog and intestinal epithelia but differed subtly in the kidneylike nephridial tubules; neither Zn nor Pb was detectable in the ventral nerve cord. High resolution X-ray mapping of high pressure-frozen, ultrathin, freeze-substituted sections in a transmission electron microscope (TEM), combined with conventional TEM structural analysis, identified a new cell type packed with highly organized rough endoplasmic reticulum and containing deposits of Cd (codistributed with S); there was no evidence that these cells are major depositories of Zn or Pb. These data may be used in a systems biology approach to assist in the interpretation of metal-evoked perturbations in whole-worm transcriptome and metabolome profiles.

Expressions of depression in rural women with chronic illness.

INTRODUCTION: Globally, chronic conditions have become the most prevalent and costly of health problems, imposing a growing drain on healthcare delivery systems and healthcare financing. Depressive symptoms and disorders are one of the most common complications of chronic illness and negatively impact one's perceived quality of life. In recent years, depression has been recognized as a major health problem for rural women. The purpose of this article is to describe the experience of depression in a sample of chronically ill rural women who participated in an online social-support and health education research project. METHODS: Middle-aged rural women with at least one chronic condition were recruited from the western USA to participate in the Women to Women (WTW) project, a 22 week computer-based intervention of virtual support and health education. The presence of depression was measured quantitatively using the Center for Epidemiologic Studies Depression Scale (CES-D). Messages posted by the women (n=82) to the online support forum were carefully examined for evidence of depressive symptomatology, perceptions of the relationship of depression to their chronic illnesses, and their strategies for coping with their depression. RESULTS: Of the 82 women who participated in the support intervention, 47 (57%) demonstrated clinically significant psychological distress at the time of enrollment into the WTW project by scoring 16 or above on the CES-D (range=0-48; x=19.27; sd=11.25). At the end of the computer intervention, complete data were available on 57 women. Of these, 24 (42%) scored 16 or above on the CES-D (range=0-49; x=15.74; sd=11.55) indicating continuing psychological distress. In all, 59 messages were coded 'depression'. The women's messages included descriptions of symptoms consistent with the literature (feelings of worthlessness and guilt; helplessness and hopelessness; alterations in sleep patterns; loss of energy). The interrelationship of depression and illness, pain, and seasonal weather variations was acknowledged; traditional and complementary healthcare treatments were discussed; relationships with healthcare providers and family and friends were described; and a variety of strategies used in coping with their depression were shared. CONCLUSIONS: Rural women with chronic illness struggle with depression. The description of their depressive symptomatology provides insight into the experience and may facilitate healthcare providers' ability to recognize depression and identify strategies to ameliorate the negative impact of depression in chronically ill rural women.

Rapid turnover of spinules at synaptic terminals.

Spinules found in brain consist of small invaginations of plasma membranes which enclose membrane evaginations from adjacent cells. Here, we focus on the dynamic properties of the most common type, synaptic spinules, which reside in synaptic terminals. In order to test whether depolarization triggers synaptic spinule formation, hippocampal slice cultures (7-day-old rats, 10-14 days in culture) were exposed to high K+ for 0.5-5 min, and examined by electron microscopy. Virtually no synaptic spinules were found in control slices representing a basal state, but numerous spinules appeared at both excitatory and inhibitory synapses after treatment with high K+. Spinule formation peaked with approximately 1 min treatment at 37 degrees C, decreased with prolonged treatment, and disappeared after 1-2 min of washout in normal medium. The rate of disappearance of spinules was substantially slower at 4 degrees C. N-methyl-D-aspartic acid (NMDA) treatment also induced synaptic spinule formation, but to a lesser extent than high K+ depolarization. In acute brain slices prepared from adult mice, synaptic spinules were abundant immediately after dissection at 4 degrees C, extremely rare in slices allowed to recover at 28 degrees C, but frequent after high K(+) depolarization. High pressure freezing of acute brain slices followed by freeze-substitution demonstrated that synaptic spinules are not induced by chemical fixation. These results indicate that spinules are absent in synapses at low levels of activity, but form and disappear quickly during sustained synaptic activity. The rapid turnover of synaptic spinules may represent an aspect of membrane retrieval during synaptic activity.

Identifying individual scaffolding molecules in the postsynaptic density.

Extended abstract of a paper presented at Microscopy and Microanalysis 2008 in Albuquerque, New Mexico, USA, August 3 - August 7, 2008.

Access to and use of research by rural nurses.

INTRODUCTION: The use of relevant research findings to inform clinical practice is important for nurses, regardless of setting. Although there have been studies addressing the use of research among various practitioners, little is known about how nurses in rural areas access health information (specifically research findings), nor how such findings are incorporated into daily practice. The purpose of this study was to explore rural nurses' access, use and perceived usefulness of research for rural practice. METHODS: The study was conducted in a sparsely populated state located in the western part of the USA. An ethnographic method was chosen to answer the research questions for this descriptive study. Semi-structured interviews were conducted with 29 rural nurses from nine communities by graduate nursing students enrolled in a rural nursing course following in-class instruction and practice. Field notes taken by the students supplemented the interview data. The students' notes included a windshield survey or description of the context and location within which the participants lived and/or practiced as well as the interviewers' observations, thoughts and impressions about the research project. Interviews were audiotaped and transcribed verbatim. Once transcribed, the interview narratives, windshield data and field notes were analyzed by the students for common themes; the students then wrote and submitted papers to the faculty addressing the themes that emerged from their interviews. The analysis conducted by the faculty members included four sources of data: transcriptions of interviews; field notes; windshield data; and students' papers. The process of identifying themes was facilitated by using the software program NUD*IST (QSR International; Melbourne, VIC, Australia). Demographic information was entered into the Statistical Package for Social Scientists (SPSS Inc; Chicago, IL, USA) to compile descriptive information about the sample. FINDINGS: Twenty-seven female and two male nurses participated in the study. The nurses' ages ranged from 31-72 years and their experience in nursing spanned 3-50 years with a range of 1 to 35 years in rural nursing. The interviews revealed that most of the nurses used the term 'research' to mean 'gathering information'. When asked how often they used 'research' the responses ranged from 2-3 times per day to 2-3 times per month. The preferred means of obtaining information was asking a colleague. Additional resources included work-place journals, books, in-services, conferences and the internet. Twenty-three of the nurses reported having internet access at work; 25 had internet access at home. Supportive supervisors and articles in general nursing journals were identified as helpful. Barriers to using research included: lack of knowledge of research methods; lack of time at work or at home to look up information; and the lack of computers and internet access on the nursing units. When computers were available, the nurses reported that poor computer literacy decreased their ability to quickly find and evaluate information. Additional barriers included diminishing financial support from employers and the long travel distances required to attend conferences. The nurses reported finding little clinical research specifically related to rural practice. CONCLUSIONS: Education and mentorship is needed about how to evaluate the types and strength of evidence, access research using the internet, interpret findings, and incorporate evidence in clinical practice. Interventions that foster the appreciation and use of research by staff nurses and managers are needed in order to build an evidence based culture. Research is needed, specifically as related to rural clinical practice.

Ligand arsenic complexation and immunoperoxidase detection of metallothionein in the earthworm Lumbricus rubellus inhabiting arsenic-rich soil.

Although earthworms have been found to inhabit arsenic-rich soils in the U.K., the mode of arsenic detoxification is currently unknown. Biochemical analyses and subcellular localization studies have indicated that As3+-thiol complexes may be involved; however, it is not known whether arsenic is capable of inducing the expression of metallothionein (MT) in earthworms. The specific aims of this paper were (a) to detect and gain an atomic characterization of ligand complexing by X-ray absorption spectrometry (XAS), and (b) to employ a polyclonal antibody raised against an earthworm MT isoform (w-MT2) to detect and localize the metalloprotein by immunoperoxidase histochemistry in the tissues of earthworms sampled from arsenic-rich soil. Data suggested that the proportion of arsenate to sulfur-bound species varies within specific earthworm tissues. Although some arsenic appeared to be in the form of arsenobetaine, the arsenic within the chlorogogenous tissue was predominantly coordinated with S in the form of -SH groups. This suggests the presence of an As::MT complex. Indeed, MT was detectable with a distinctly localized tissue and cellular distribution. While MT was not detectable in the surface epithelium or in the body wall musculature, immunoperoxidase histochemistry identified the presence of MT in chloragocytes around blood vessels, within the typhlosolar fold, and in the peri-intestinal region. Focal immunostaining was also detectable in a cohort of cells in the intestinal wall. The results of this study support the hypothesis that arsenic induces MT expression and is sequestered by the metalloprotein in certain target cells and tissues.

Inhibition of phosphatase activity facilitates the formation and maintenance of NMDA-induced calcium/calmodulin-dependent protein kinase II clusters in hippocampal neurons.

The majority of hippocampal neurons in dissociated cultures and in intact brain exhibit clustering of calcium/calmodulin-dependent protein kinase II (CaMKII) into spherical structures with an average diameter of 110 nm when subjected to conditions that mimic ischemia and excitotoxicity [Neuroscience 106 (2001) 69]. Because clustering of CaMKII would reduce its effective concentration within the neuron, it may represent a cellular strategy to prevent excessive CaMKII-mediated phosphorylation during episodes of Ca2+ overload. Here we employ a relatively mild excitatory stimulus to promote sub-maximal clustering for the purpose of studying the conditions for the formation and disappearance of CaMKII clusters. Treatment with 30 microM N-methyl-D-aspartic acid (NMDA) for 2 min produced CaMKII clustering in approximately 15% of dissociated hippocampal neurons in culture, as observed by pre-embedding immunogold electron microscopy. These CaMKII clusters could be labeled with antibodies specific to the phospho form (Thr286) of CaMKII, suggesting that at least some of the CaMKII molecules in clusters are autophosphorylated. To test whether phosphorylation is involved in the formation and maintenance of CaMKII clusters, the phosphatase inhibitors calyculin A (5 nM) or okadaic acid (1 microM) were included in the incubation medium. With inhibitors more neurons exhibited CaMKII clusters in response to 2 min NMDA treatment. Furthermore, 5 min after the removal of NMDA and Ca2+, CaMKII clusters remained and could still be labeled with the phospho-specific antibody. In contrast, in the absence of phosphatase inhibitors, no clusters were detected 5 min after the removal of NMDA and Ca2+ from the medium. These results suggest that phosphatases type 1 and/or 2A regulate the formation and disappearance of CaMKII clusters.

Differential metallothionein expression in earthworm (Lumbricus rubellus) tissues.

This article describes the immunoperoxidase localization of metallothionein (MT) in the major organs and tissues of the earthworm Lumbricus rubellus sampled from a mine soil heavily polluted with Pb, Zn, and Cd. The polyclonal antiserum used was raised against the MT isoform (wMT2), the molecular characteristics and focal subcellular distribution of which indicate a primary role for it in the sequestration of certain nonessential metals, such as Cd. Intense MT immunostaining was detectable in chloragogenous tissue throughout the body: around the intestine, in the typhlosolar infolding, around blood vessels anterior and posterior to the crop/gizzard, and around the calciferous gland. Electron probe X-ray microanalysis of neutral red-labeled vesicular structures in the chloragogenous tissue indicated that this acidic compartment, probably lysosomal, yielded the strong Cd and S signals associated with Cd-MT. MT expression was also detected in the apical cytoplasm of intestinal epithelial cells; in coelomocytes contiguous with chloragocytes attached to the gut; within the narrow tubular region of nephridia, in the secretory epithelia of the calciferous gland, but not anywhere in the body wall. We concluded that (a) the main route of Cd uptake is probably via absorptive alimentary surfaces, and not across the external epidermal layer; (b) nephridia are involved with Cd excretion and/or are a major target of Cd-induced pathological damage; (c) tentatively, a combination of immunohistochemistry and proton-induced X-ray emission analysis indicated that the calciferous gland is probably not a major "heavy metal" excretory route.

Chloride channels in basolateral TAL membranes. XVIII. Phenylglyoxal induces functional mcCIC-Ka activity in basolateral MTAL membranes.

Cultured mouse MTAL cells contain more mRNA encoding the Cl- channel mcCIC-Ka, which mediates CTAL Cl- absorption, than mRNA encoding the Cl- channel mmCIC-Ka, which mediates MTAL Cl- absorption. mmCIC-Ka and mcCIC-Ka have three functional differences: 1) mmCIC-Ka open time probability, Po, increases with increasing cytosolic Cl-, but variations in cytosolic Cl- do not affect Po in mcCIC-Ka; 2) mmCIC-Ka is gated by (ATP + PKA), while (ATP + PKA) have no effect on Po in mcCIC-Ka; and 3) mmCIC-Ka channels have single-ion occupancy, while mcCIC-Ka channels have multi-ion occupancy. Using basolateral vesicles from MTAL cells fused into bilayers, we evaluated the effects of 1 mM cytosolic phenylglyoxal (PGO), which binds covalently to lysine or arginine, on Cl- channels. With PGO pretreatment, Cl- channels were uniformly not gated either with increases in cytosolic-face Cl- or with (ATP + PKA) at 2 mM cytosolic-face Cl-; and they exhibited multi-ion occupancy kinetics typical for mcCIC-Ka channels. Thus, in basolateral MTAL membranes, blockade of Cl- access to arginine or lysine residues on mmCIC-Ka by PGO results in Cl- channels having the functional characteristics of mcCIC-Ka channels.

Cl- channels in basolateral TAL membranes. XIX. Cytosolic Cl- regulates mmCIC-Ka and mcCIC-Ka channels.

We evaluated the effects of culturing mouse MTAL cells under conditions that suppressed steady-state cytosolic Cl- on chloride channels fused into bilayers from basolateral vesicles of cultured MTAL cells. We used two agents to suppress Cl- entry: 10(-6) M PGE2 and 10(-4) M bumetanide. Basolateral Cl- channels from control cultured MTAL cells exhibited the signature characteristics of mmCIC-Ka channels: increased open-time probability (Po) either by raising cytosolic-face [Cl-] or, at 2 mM cytosolic Cl-, by adding (ATP + PKA), and first-order conductance kinetics. Either 10(-6) M PGE2 or 10(-4) M bumetanide in culture media reduced steady-state MTAL cytosolic Cl-. Chloride channels from these cells exhibited characteristics unique to CTAL mcCIC-Ka channels, namely: no augmentation of Po either by raising cytosolic Cl- or with cytosolic (ATP + PKA), and multi-ion occupancy. Semi-quantitative RT-PCR and real-time quantitative PCR showed that culturing MTAL cells with 10(-6) M PGE2 or 10(-4) M bumetanide reduced mRNA levels encoding mmCIC-Ka but not mRNA levels encoding mcCIC-Ka. However, when MTAL cells were cultured under control conditions, and then pre-incubated for 60 minutes with 10(-4) M bumetanide, cytosolic Cl- fell acutely but Cl- channels exhibited characteristics of mmCIC-Ka channels. Thus PGE2 and bumetanide, both of which lower steady-state MTAL cytosolic Cl- concentrations, inhibit either the transcriptional and/or the translational processes for mmCIC-Ka synthesis.

Cl channels in basolateral TAL membranes. XVII. Kinetic properties of mcClC-Ka, a basolateral CTAL Cl- channel.

This paper describes the kinetics of Cl- flux through mcClC-Ka Cl- channels from basolateral membranes of mouse CTAL cells. We have cloned two separate but highly homologous Cl- channels, mmClC-Ka from cultured mouse MTAL cells and mcClC-Ka from cultured mouse CTAL cells. The mmClC-Ka and mcClC-Ka channels appear to mediate net Cl- absorption in the MTAL and CTAL, respectively. The kinetics of Cl- permeation through mmClC-Ka channels exhibit traditional criteria for a first-order process, including saturation kinetics. Thus mmClC-Ka channels operate functionally as if the channels were occupied by a single Cl- ion at any given time. In the present studies, we examined conductance-concentration relations in mcClC-Ka channels, and compared both mole-fraction effects and ion selectivity characteristics in mmClC-Ka and mcClC-Ka channels. In mcClC-Ka channels, we observed both self-block at high external Cl- concentrations and, at constant ionic strength, an anomalous mole-fraction effect using external solutions containing varying F-/Cl- concentrations. Neither effect was obtained in mmClC-Ka channels. These data are consistent with the possibility that Cl- permeation through mcClC-Ka channels involved multi-ion occupancy channels that expressed single-file behavior.

Inhibition of phosphatase activity prolongs NMDA-induced modification of the postsynaptic density.

NMDA-induced modification of postsynaptic densities (PSDs) was studied by immunoelectron microscopy. Treatment of cultured hippocampal neurons with NMDA for 2 min promotes a 2.3 fold thickening of the PSD and a 4 fold increase in PSD-associated CaMKII immunolabel. These changes are reversed 5 min after the removal of NMDA and Ca2+ from the medium. In addition, following NMDA treatment, PSDs exhibit a 7.5 fold increase in labeling with an antibody specific to the (Thr286) phospho-form of CaMKII, indicating that CaMKII translocated to the PSD is phosphorylated. When the phosphatase inhibitors, calyculin A or okadaic acid, are included in the medium, the NMDA-induced thickening of the PSD as well as the increase in PSD-associated CaMKII immunolabeling are largely maintained (75% and 88% of the peak values respectively) at 5 min after removal of NMDA and Ca2+ from the medium. These results imply that NMDA receptors can mediate activity-induced changes in the PSD and that phosphatases of type 1 and/or 2A are involved in the reversal of these changes.

Sustained elevation of calcium induces Ca(2+)/calmodulin-dependent protein kinase II clusters in hippocampal neurons.

Treatment of cultured hippocampal neurons with the mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) in the absence of glucose mimics ischemic energy depletion and induces formation of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) clusters, spherical structures with diameters of 75-175 nm [Dosemeci et al., J. Neurosci. 20 (2000) 3076-3084]. The demonstration that CaMKII clustering occurs in the intact, adult rat brain upon interruption of blood flow indicates that clustering is not confined to cell cultures. Application of N-methyl-D-aspartate (250 microM, 15 min) to hippocampal cultures also induces cluster formation, suggesting a role for Ca(2+). Indeed, intracellular Ca(2+) monitored with Fluo3-AM by confocal microscopy reaches a sustained high level within 5 min of CCCP treatment. The appearance of immunolabeled CaMKII clusters, detected by electron microscopy, follows the onset of the sustained increase in intracellular Ca(2+). Moreover, CaMKII does not cluster when the rise in intracellular Ca(2+) is prevented by the omission of extracellular Ca(2+) during CCCP treatment, confirming that clustering is Ca(2+)-dependent. A lag period of 1-2 min between the onset of high intracellular Ca(2+) levels and the formation of CaMKII clusters suggests that a sustained increase in Ca(2+) level is necessary for the clustering. CaMKII clusters disappear within 2 h of returning the cultures to normal incubation conditions, at which time no significant cell death is detected. These results indicate that pathological conditions that promote sustained episodes of Ca(2+) overload result in a transitory clustering of CaMKII into spherical structures. CaMKII clustering may represent a cellular defense mechanism to sequester a portion of the CaMKII pool, thereby preventing excessive protein phosphorylation.

Glutamate-induced transient modification of the postsynaptic density.

Depolarization of rat hippocampal neurons with a high concentration of external potassium induces a thickening of postsynaptic densities (PSDs) within 1.5-3 min. After high-potassium treatment, PSDs thicken 2.1-fold in cultured neurons and 1.4-fold in hippocampal slices compared with their respective controls. Thin-section immunoelectron microscopy of hippocampal cultures indicates that at least part of the observed thickening of PSDs can be accounted for by an accumulation of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) on their cytoplasmic faces. Indeed, PSD-associated gold label for CaMKII increases 5-fold after depolarization with potassium. The effects of high-potassium treatment on the composition and structure of the PSDs are mimicked by direct application of glutamate. In cultures, glutamate-induced thickening of PSDs and the accumulation of CaMKII on PSDs are reversed within 5 min of removal of glutamate and Ca(2+) from the extracellular medium. These results suggest that PSDs are dynamic structures whose thickness and composition are subject to rapid and transient changes during synaptic activity.

Metal ion trafficking in earthworms. Identification of a cadmium-specific metallothionein.

Exposure to cadmium poses a considerable risk to human health and environmental safety. Earthworms reside in the most contaminated sites on earth, displaying a phenomenal tolerance to toxic heavy metals. They exhibit a distinct metabolic pathway that allows the bio-accumulation of cadmium to yield body burdens in excess of 1/1000th of total dry body weight, a most impressive figure by any standard. However, the precise molecular mechanism underlying this phenomenon remains to be unraveled. This study meets this challenge by fully characterizing the major metal-binding protein in earthworms, namely the two isoforms of metallothionein. Chemical analysis of recombinant protein showed that although both isoforms bind equimolar amounts of cadmium (6 mol), wMT-2 is more stable during proton competition. Furthermore, isoform-specific transcript analysis demonstrated that only wMT-2 is responsive to cadmium in a dose and temporal manner. The specific sequestration of cadmium to wMT-2 protein was confirmed in situ using polyclonal antisera. The latter also provided the means for mapping the cellular and intracellular distribution of metallothionein, thus yielding a holistic insight into its involvement in cadmium transit during absorption, storage, and excretion. The structure-function relationship of wMT-2 and its role in cadmium detoxification through sequestration and compartmentalization is discussed.

Chloride channels in the loop of Henle.

Cl- transport in the loop of Henle is responsible for reclamation of 25-40% of the filtered NaCl load and for the formation of dilute urine. Our understanding of the physiologic and molecular mechanisms responsible for Cl- reabsorption in both the thin ascending limb and thick ascending limb of Henle's loop has increased greatly over the last decade. Plasma membrane Cl- channels are known to play an integral role in transcellular Cl- transport in both the thin and thick ascending limbs. This review focuses on the functional characteristics and molecular identities of these Cl- channels, as well as the role of these channels in the pathophysiology of disease.