Basolateral plasma membrane localization of ouabain-sensitive sodium transport sites in the secretory epithelium of the avian salt gland

The distribution of Na+ pump sites (Na+-K+-ATPase) in the secretory epithelium of the avian salt gland was demonstrated by freeze-dry autoradiographic analysis of [(3)H] ouabain binding sites. Kinetic studies indicated that near saturation of tissue binding sites occurred when slices of salt glands from salt-stressed ducks were exposed to 2.2 μM ouabain (containing 5 μCi/ml [(3)H]ouabain) for 90 min. Washing with label-free Ringer's solution for 90 min extracted only 10% of the inhibitor, an amount which corresponded to ouabain present in the tissue spaces labeled by [(14)C]insulin. Increasing the KCl concentration of the incubation medium reduced the rate of ouabain binding but not the maximal amount bound. In contrast to the low level of ouabain binding to salt glands of ducks maintained on a freshwater regimen, exposure to a salt water diet led to a more than threefold increase in binding within 9-11 days. This increase paralleled the similar increment in Na+-K+-ATPase activity described previously. [(3)H]ouabain binding sites were localized autoradiographically to the folded basolateral plasma membrane of the principal secretory cells. The luminal surfaces of these cells were unlabeled. Mitotically active peripheral cells were also unlabeled. The cell-specific pattern of [(3)H]ouabain binding to principal secretory cells and the membrane-specific localization of binding sites to the nonluminal surfaces of these cells were identical to the distribution of Na+-K+-ATPase as reflected by the cytochemical localization of ouabain-sensitive and K+-dependent nitrophenyl phosphatase activity. The relationship between the nonluminal localization of Na+-K+-ATPase and the possible role of the enzyme n NaCl secretion is considered in the light of physiological data on electrolyte transport in salt glands and other secretory epithelia.     

Intercellular communication in the rat anterior pituitary gland: an in vivo and in vitro study

The concept of "stimulus-secretion coupling" suggested by Douglas and co-workers to explain the events related to monamine discharge by the adrenal medulla (5, 7) may be applied to other endocrine tissues, such as adrenal cortex (36), pancreatic islets (4), and magnocellular hypothalamic neurons (6), which exhibit a similar ion-dependent process of hormone elaboration. In addition, they share another feature, that of joining neighbor cells via membrane junctions (12, 26, and Fletcher, unpublished observation). Given this, and the reports that hormone secretion by the pars distalis also involves a secretagogue-induced decrease in membrane bioelectric potential accompanied by a rise in cellular [Ca++] (27, 34, 41), it was appropriate to test the possibility that cells of the anterior pituitary gland are united by junctions.     

Information Structure Influences Depth of Syntactic Processing: Event-Related Potential Evidence for the Chomsky Illusion

Information structure facilitates communication between interlocutors by highlighting relevant information. It has previously been shown that information structure modulates the depth of semantic processing. Here we used event-related potentials to investigate whether information structure can modulate the depth of syntactic processing. In question-answer pairs, subtle (number agreement) or salient (phrase structure) syntactic violations were placed either in focus or out of focus through information structure marking. P600 effects to these violations reflect the depth of syntactic processing. For subtle violations, a P600 effect was observed in the focus condition, but not in the non-focus condition. For salient violations, comparable P600 effects were found in both conditions. These results indicate that information structure can modulate the depth of syntactic processing, but that this effect depends on the salience of the information. When subtle violations are not in focus, they are processed less elaborately. We label this phenomenon the Chomsky illusion.     

High-throughput screen identifies disulfiram as a potential therapeutic for triple-negative breast cancer cells: Interaction with IQ motif-containing factors

Triple-negative breast cancer (TNBC) represents an aggressive subtype, for which radiation and chemotherapy are the only options. Here we describe the identification of disulfiram, an FDA-approved drug used to treat alcoholism, as well as the related compound thiram, as the most potent growth inhibitors following high-throughput screens of 3185 compounds against multiple TNBC cell lines. The average IC₅₀ for disulfiram was ~300 nM. Drug affinity responsive target stability (DARTS) analysis identified IQ motif-containing factors IQGAP1 and MYH9 as direct binding targets of disulfiram. Indeed, knockdown of these factors reduced, though did not completely abolish, cell growth. Combination treatment with 4 different drugs commonly used to treat TNBC revealed that disulfiram synergizes most effectively with doxorubicin to inhibit cell growth of TNBC cells. Disulfiram and doxorubicin cooperated to induce cell death as well as cellular senescence, and targeted the ESA⁺/CD24^-/low/CD44⁺ cancer stem cell population. Our results suggest that disulfiram may be repurposed to treat TNBC in combination with doxorubicin.     

Repeated oral administration of capsaicin increases anxiety-like behaviours with prolonged stress-response in rats

This study was conducted to examine the psycho-emotional effects of repeated oral exposure to capsaicin, the principal active component of chili peppers. Each rat received 1 mL of 0.02% capsaicin into its oral cavity daily, and was subjected to behavioural tests following 10 daily administrations of capsaicin. Stereotypy counts and rostral grooming were significantly increased, and caudal grooming decreased, in capsaicin-treated rats during the ambulatory activity test. In elevated plus maze test, not only the time spent in open arms but also the percent arm entry into open arms was reduced in capsaicin-treated rats compared with control rats. In forced swim test, although swimming duration was decreased, struggling increased in the capsaicin group, immobility duration did not differ between the groups. Repeated oral capsaicin did not affect the basal levels of plasma corticosterone; however, the stress-induced elevation of plasma corticosterone was prolonged in capsaicin treated rats. Oral capsaicin exposure significantly increased c-Fos expression not only in the nucleus tractus of solitarius but also in the paraventricular nucleus. Results suggest that repeated oral exposure to capsaicin increases anxiety-like behaviours in rats, and dysfunction of the hypothalamic-pituitary-adrenal axis may play a role in its pathophysiology.     

Differential Sensitivities of Fast- and Slow-Cycling Cancer Cells to Inosine Monophosphate Dehydrogenase 2 Inhibition by Mycophenolic Acid

As uncontrolled cell proliferation requires nucleotide biosynthesis, inhibiting enzymes that mediate nucleotide biosynthesis constitutes a rational approach to the management of oncological diseases. In practice, however, results of this strategy are mixed and thus elucidation of the mechanisms by which cancer cells evade the effect of nucleotide biosynthesis restriction is urgently needed. Here we explored the notion that intrinsic differences in cancer cell cycle velocity are important in the resistance toward inhibition of inosine monophosphate dehydrogenase (IMPDH) by mycophenolic acid (MPA). In short-term experiments, MPA treatment of fast-growing cancer cells effectively elicited G0/G1 arrest and provoked apoptosis, thus inhibiting cell proliferation and colony formation. Forced expression of a mutated IMPDH2, lacking a binding site for MPA but retaining enzymatic activity, resulted in complete resistance of cancer cells to MPA. In nude mice subcutaneously engrafted with HeLa cells, MPA moderately delayed tumor formation by inhibiting cell proliferation and inducing apoptosis. Importantly, we developed a lentiviral vector–based Tet-on label-retaining system that enables to identify, isolate and functionally characterize slow-cycling or so-called label-retaining cells (LRCs) in vitro and in vivo. We surprisingly found the presence of LRCs in fast-growing tumors. LRCs were superior in colony formation, tumor initiation and resistance to MPA as compared with fast-cycling cells. Thus, the slow-cycling compartment of cancer seems predominantly responsible for resistance to MPA.     

Processing of cell‐surface signalling anti‐sigma factors prior to signal recognition is a conserved autoproteolytic mechanism that produces two functional domains

Cell‐surface signalling (CSS) enables Gram‐negative bacteria to transduce an environmental signal into a cytosolic response. This regulatory cascade involves an outer membrane receptor that transmits the signal to an anti‐sigma factor in the cytoplasmic membrane, allowing the activation of an extracytoplasmic function (ECF) sigma factor. Recent studies have demonstrated that RseP‐mediated proteolysis of the anti‐sigma factors is key to σ^ECF activation. Using the Pseudomonas aeruginosa FoxR anti‐sigma factor, we show here that RseP is responsible for the generation of an N‐terminal tail that likely contains pro‐sigma activity. Furthermore, it has been reported previously that this anti‐sigma factor is processed in two separate domains prior to signal recognition. Here, we demonstrate that this process is common in these types of proteins and that the processing event is probably due to autoproteolytic activity. The resulting domains interact and function together to transduce the CSS signal. However, our results also indicate that this processing event is not essential for activity. In fact, we have identified functional CSS anti‐sigma factors that are not cleaved prior to signal perception. Together, our results indicate that CSS regulation can occur through both complete and initially processed anti‐sigma factors.     

Neuronal dynamics underlying high- and low-frequency EEG oscillations contribute independently to the human BOLD signal

Work on animals indicates that BOLD is preferentially sensitive to local field potentials, and that it correlates most strongly with gamma band neuronal synchronization. Here we investigate how the BOLD signal in humans performing a cognitive task is related to neuronal synchronization across different frequency bands. We simultaneously recorded EEG and BOLD while subjects engaged in a visual attention task known to induce sustained changes in neuronal synchronization across a wide range of frequencies. Trial-by-trial BOLD fluctuations correlated positively with trial-by-trial fluctuations in high-EEG gamma power (60-80 Hz) and negatively with alpha and beta power. Gamma power on the one hand, and alpha and beta power on the other hand, independently contributed to explaining BOLD variance. These results indicate that the BOLD-gamma coupling observed in animals can be extrapolated to humans performing a task and that neuronal dynamics underlying high- and low-frequency synchronization contribute independently to the BOLD signal.     

Mapping and association studies of diabetes related genes in the pig

The mitogen-activated protein kinase 8 (MAPK8), resistin (RETN), 11 beta hydroxysteroid dehydrogenase isoform 1 (HSD11B1) and protein kinase B Akt2 (AKT2) genes are all genes known to affect insulin signalling and have been implicated in the progression of obesity and type 2 diabetes in humans. In this study, polymorphisms in the porcine diabetes related MAPK8, RETN, HSD11B1 and AKT2 genes were identified, mapped and their associations with phenotypic measurements in swine were analysed. Polymorphisms detected in the MAPK8, RETN and HSD11B1 loci were used to genotype a Berkshire-Yorkshire pig breed reference family. Using linkage analysis, RETN, HSD11B1 and MAPK8 genes were mapped to pig chromosomes 2, 9 and 14, respectively, while the AKT2 gene was physically mapped to pig chromosome 6q21. Results presented here suggest associations between the polymorphisms in the MAPK8, RETN and HSD11B1 genes with several phenotypic measurements, including fat deposition traits in the pig. Because these genes have been implicated in obesity and diabetes in humans, and this study suggests associations with fat related traits, further research on these genes in swine may provide useful information on genetic factors underlying lean pork production.