Cleavage state of γENaC in mouse and rat kidneys.

Extracellular proteases can activate the epithelial Na channel (ENaC) by cleavage of the γ subunit. Here, we investigated the cleavage state of the channel in the kidneys of mice and rats on a low-salt diet. We identified the cleaved species of channels expressed in Fisher rat thyroid cells by coexpressing the apical membrane-bound protease channel-activating protease 1 (CAP1; prostasin). To compare the peptides produced in the heterologous system with those in the mouse kidney, we treated both lysates with PNGaseF to remove N-linked glycosylation. The apparent molecular mass of the smallest COOH-terminal fragment of γENaC (52 kDa) was indistinguishable from that of the CAP1-induced species in Fisher rat thyroid cells. Similar cleaved peptides were observed in total and cell surface fractions of the rat kidney. This outcome suggests that most of the subunits at the surface have been processed by extracellular proteases. This was confirmed using nonreducing gels, in which the NH2- and COOH-terminal fragments of γENaC are linked by a disulfide bond. Under these conditions, the major cleaved form in the rat kidney had an apparent molecular mass of 56 kDa, ∼4 kDa lower than that of the full-length form, consistent with excision of a short peptide by two proteolytic events. We conclude that the most abundant γENaC species in the apical membrane of rat and mouse kidneys on a low-Na diet is the twice-cleaved, presumably activated form.NEW & NOTEWORTHY We have identified the major aldosterone-dependent cleaved form of the epithelial Na channel (ENaC) γ subunit in the kidney as a twice-cleaved peptide. This form appears to be identical in size with a subunit cleaved in vitro by the extracellular protease channel-activating protease 1 (prostasin). In the absence of reducing agents, it has an overall molecular mass less than that of the intact subunit, consistent with the excision of an inhibitory domain.

The Ca2+-activated chloride channel ANO1/TMEM16A: An emerging therapeutic target for epithelium-originated diseases?

Anoctamin 1 (ANO1) or TMEM16A gene encodes a member of Ca2+ activated Cl- channels (CaCCs) that are critical for physiological functions, such as epithelial secretion, smooth muscle contraction and sensory signal transduction. The attraction and interest in ANO1/TMEM16A arise from a decade long investigations that abnormal expression or dysfunction of ANO1 is involved in many pathological phenotypes and diseases, including asthma, neuropathic pain, hypertension and cancer. However, the lack of specific modulators of ANO1 has impeded the efforts to validate ANO1 as a therapeutic target. This review focuses on the recent progress made in understanding of the pathophysiological functions of CaCC ANO1 and the current modulators used as pharmacological tools, hopefully illustrating a broad spectrum of ANO1 channelopathy and a path forward for this target validation.

Elevated Tau Level in Aged Rat Cerebrospinal Fluid Reduced by Treatment with a Neurotrophic Compound.

Alzheimer's disease (AD) is the single major cause of dementia in middle- to old-age individuals, and, as of yet, no disease-modifying therapeutic drug is available for its treatment. A major obstacle in the successful development of disease-modifying therapeutic drugs has been the lack of suitable animal models of the sporadic form of AD as well as a biomarker that can be used both for therapeutic preclinical studies and for human clinical trials. Previously we showed neurogenesis and neuronal plasticity deficits and cognitive impairment and their rescue with a neurotrophic peptidergic compound, DGGLAG named P021, in aged Fisher rats. Here we show that P021 is blood-brain-barrier-permeable, and chronic oral treatment with this compound can reduce the brain level of total tau in the aged rats. Furthermore, cerebrospinal fluid (CSF) levels of both tau and Aß/AßPP are elevated in the aged animals, and chronic treatment with P021 can reduce tau but not Aß/AßPP to that of the levels found in young adult rats. Importantly, P021 does not induce any detectable immune reaction in rats. Collectively, these studies show the therapeutic potential of P021 as a disease-modifying compound and the suitability of the aged Fisher rats as a model of cerebral aging in which the therapeutic efficacy of a tau-reducing compound can be monitored in the CSF.

Differential impact of stress on hypothalamic-pituitary-adrenal axis: gene expression changes in Lewis and Fisher rats.

The aim of the present work was to study the influence of variable stress on the expression of 11ß-hydroxysteroid dehydrogenase type 1 (11HSD1) and the neuropeptides corticotropin-releasing hormone (CRH), urocortins 2 and 3(UCN2, UCN3), arginine vasopressin (AVP), oxytocin (OXT) and adenylate cyclase-activating polypeptide (PACAP) in two inbred rat strains: stress hypo-responsive Lewis (LEW) and hyper-responsive Fisher 344 (F344) rats. We found site-specific and strain-dependent differences in the basal and stress-stimulated expression of 11HSD1, CRH, UCN2, UCN3 and PACAP. In LEW rats, stress upregulated 11HSD1 in the prefrontal cortex and lateral amygdala, whereas in F344 rats 11HSD1 was upregulated in the central amygdala and hippocampal CA2 and ventral but not dorsal CA1 region; no effect was observed in the paraventricular nucleus, pituitary gland and adrenal cortex of both strains. The expression of glucocorticoid receptors did not parallel the upregulation of 11HSD1. Stress also stimulated the expression of paraventricular OXT, CRH, UCN3 and PACAP in both strains but amygdalar CRH only in LEW and UCN2/UCN3 in F344 rats, respectively. The upregulation of PACAP and CRH was paralleled only by increased expression of PACAP receptor PAC1 but not CRH receptor type 1. These observations provide evidence that inbred F344 and LEW rats exhibit not only the well-known phenotypic differences in the activity of the HPA axis but also strain- and stress-dependent differences in the expression of genes encoding 11HSD1 and neuropeptides associated with the HPA axis activity. Moreover, the differences in 11HSD1 expression suggest different local concentration of corticosterone and access to GR in canonical and noncanonical structures of the HPA axis.

Obtención y Análisis del Espectro de Absorción Óptico de Sangre de Rata Fisher con Daño Hepático Mediante Espectroscopia Foto acústica / Obtaining and Analysis of Optical Absorption Spectrum of Fisher Rat Blood with Hepatic Damage by Photoacoustic Spectroscopy

Resumen: La Espectroscopia Fotoacústica (EFA), ha destacado de entre las técnicas fototérmicas por su versatilidad para la caracterización de diversos tipos de muestras, incluyendo las de tipo biológicas. En este estudio, se compararon y analizaron mediante EFA, las diferencias entre los espectros de absorción ópticos de sangre de ratas macho Fisher en tres casos, control (ratas sanas), con daño hepático, y con daño hepático tratadas con Curcuma longa. Los resultados experimentales demostraron diferencias en los espectros de absorción ópticos de cada caso analizado, por lo que EFA se propondría como una técnica complementaria, no convencional para el estudio del efecto de la Curcuma longa en daño hepático inducido en modelo animal.

Abstract: Photoacoustic Spectroscopy (PAS) stand outs among Photothermal techniques due to its versatility for the characterization of different types of materials, including biological samples. In this study, were compared and analyzed by PAS, the differences between the optical absorption spectra of blood of male Fisher rats, in three cases, control (healthy rats), liver damage and liver damage treated with Curcuma longa. The experimental results show differences in the optical absorption spectra of each analysed case, therefore PAS would be proposed as a non conventional complementary technique, to study the effect of Curcuma longa in induced liver damage for an animal model.