A tool for sustainability impact assessment (ToSIA)of forest-wood chains linked with a database of sustainability indicators collected within the EFORWOOD project

Within the EFORWOOD project new approaches to assess the sustainability impacts of forest-wood chains (FWC) using indicators of environmental, social and economic sustainability were developed

Religious Existentialism as a Countermeasure to Moralistic Therapeutic Deism

Moralistic Therapeutic Deism (MTD) is a new spiritual trend that is increasingly changing the religious landscape of in the Euro-American cultural provenience. Though appealing to the generation of 'emerging adults' (age 18-25), MTD results in undesirable life-outcomes that prompt religious scholars, theologians, and sociologists to carefully study its roots, expressions, and possible alternatives. Using S. Kierkegaard's religious existentialism as a case study, the authors argue that religious existentialism has the potential to face the detrimental effects of MTD and to reinvigorate stagnating religious communities

Le « Pays de Samson ». Eshwa : la grotte dite de Samson ('Araq Isma'in) / Père Raphaël Tonneau

Numérisé par le partenaireAppartient à l’ensemble documentaire : BbLevt0Numérisé par le partenair

Determination of μ-, δ- and κ-opioid receptors in forebrain cortex of rats exposed to morphine for 10 days: Comparison with animals after 20 days of morphine withdrawal

<div><p>Background</p><p>Chronic exposure of mammalian organism to morphine results in adaption to persistent high opioid tone through homeostatic adjustments. Our previous results indicated that in the frontal brain cortex (FBC) of rats exposed to morphine for 10 days, such a compensatory adjustment was detected as large up-regulation of adenylylcyclases I (8-fold) and II (2.5–fold). The other isoforms of AC (III-IX) were unchanged. Importantly, the increase of ACI and ACII was reversible as it disappeared after 20 days of morphine withdrawal. Changes of down-stream signaling molecules such as G proteins and adenylylcyclases should respond to and be preceded by primary changes proceeding at receptor level. Therefore in our present work, we addressed the problem of reversibility of the long-term morphine effects on μ-, δ- and κ-OR protein levels in FBC.</p><p>Methods</p><p>Rats were exposed to increasing doses of morphine (10–40 mg/kg) for 10 days and sacrificed either 24 h (group +M10) or 20 days (group +M10/−M20) after the last dose of morphine in parallel with control animals (groups −M10 and −M10/−M20). Post-nuclear supernatant (PNS) fraction was prepared from forebrain cortex, resolved by 1D-SDS-PAGE under non-dissociated (−DTT) and dissociated (+DTT) conditions, and analyzed for the content of μ-, δ- and κ-OR by immunoblotting with C- and N-terminus oriented antibodies.</p><p>Results</p><p>Significant down-regulation of δ-OR form exhibiting M_w ≈ 60 kDa was detected in PNS prepared from both (+M10) and (+M10/−M20) rats. However, the total immunoblot signals of μ-, δ- and κ-OR, respectively, were unchanged. Plasma membrane marker Na, K-ATPase, actin and GAPDH were unaffected by morphine in both types of PNS. Membrane-domain marker caveolin-1 and cholesterol level increased in (+M10) rats and this increase was reversed back to control level in (+M10/−M20) rats.</p><p>Conclusions</p><p>In FBC, prolonged exposure of rats to morphine results in minor (δ-OR) or no change (μ- and κ-OR) of opioid receptor content. The reversible increases of caveolin-1 and cholesterol levels suggest participation of membrane domains in compensatory responses during opioid withdrawal.</p><p>General significance</p><p>Analysis of reversibility of morphine effect on mammalian brain.</p></div

Na, K-ATPase, caveolin-1 and actin in PNS fractions preparad from experimental groups (±M10) and (±M10/─M20).

<p>Na, K-ATPase <b>(A),</b> [³H]ouabain binding <b>(B),</b> caveolin-1 <b>(C)</b> and actin <b>(D)</b> levels were determined in (+M10), (─M10), (+M10/─M20) and (─M10/─M20) samples of PNS. PNS proteins (20 μg per lane) were resolved under dissociated conditions (+DTT) by standard SDS-PAGE in 10% w/v acrylamide/0.26% w/v bis-acrylamide gels, and immunoblotted. Antibodies H-300 (sc-28800) and I-19 (sc-1616) from Santa Cruz were used for recognition of α subunit of Na, K-ATPase and actin, respectively. [³H]ouabain binding assay was performed at saturating, 18 nM concentration as described in Methods. Caveolin-1 (↓) was identified by Ab C13630-050 from Transduction Laboratories. The human endothelial cell lysate was used as a positive control for caveolin-1 (Transduction Laboratories, in last lane of immunoblots). Results represent the average of three immunoblots of Na, K-ATPase or [³H]ouabain binding assays ± SEM, each performed in quadruplicates (upper columns). Analysis of caveolin-1 and actin was based on signals collected from five and three immunoblots, each performed with four control + four morphine-treated samples of PNS, respectively. 100% on y-axis (upper panels) represents the average intensity of a given immunoblot signal determined in PNS prepared from control, (─M10) rats. The significance of difference between the control and morphine-treated samples was analyzed by Student´s <i>t</i>-test using GraphPad<i>Prizm4</i>. In the lower panels, typical immunoblots are shown (NS, p > 0.05; ***, p < 0.001).</p

Determination of μ-OR in PNS prepared from FBC of experimental groups (+M10) and (─M10).

<p>PNS fractions (20 μg protein per lane) prepared from morphine-treated (+M10) and control (— M10) rats were resolved under non-dissociated (— DTT) or dissociated (+DTT) conditions by 1D-SDS-PAGE in 10% w/v acrylamide/0.26% w/v bis-acrylamide (<b>A, C)</b> or in 4–12% NuPAGE gels (<b>B, D</b>); μ-OR was recognized by C-terminus-oriented Ab C-20 (sc-7488-R). When resolved under non-dissociated conditions (<b>A</b> and <b>B</b>), Ab C-20 distinguished the single protein with M_w ≈ 60 kDa. The average intensity of this immunoblot signal was not significantly different when compared in control (— M10) and morphine-treated (+M10) PNS samples (NS, p>0.05). Resolution under dissociated conditions (<b>C</b> and <b>D</b>) revealed the presence of multiple protein bands exhibiting a wide range of M_w ≈ 20–90 kDa. These immunoblot signals were also unchanged by morphine (NS, p>0.05). The results (upper columns) represent the average signal of five immunoblots, each performed with four control + four morphine-treated samples of PNS ± SEM. 100% on y-axis represents the average intensity of a given immunoblot signal determined in PNS prepared from control, (─M10) rats. The significance of difference between the control and morphine-treated PNS samples was analyzed by Student´s <i>t</i>-test using GraphPad<i>Prizm4</i>. In the lower panels, typical immunoblots are presented.</p

Detection of μ-OR, δ-OR and κ-OR in PNS fractions (±M10) treated with <i>N</i>-glycosidase F.

<p>PNS fractions prepared from FBC of rats in groups (+M10) and (─M10) were treated with SDS, Triton X-100 and <i>N</i>-glycosidase F (NGF) as described in Methods. 20 μg of protein was applied per each lane of 10% w/v acrylamide/0.26% w/v bis-acrylamide gels (four + four) and resolved under the dissociated (+DTT) conditions. The OR were recognized by antibodies from Santa Cruz: μ-OR (C-20, sc-7488-R, C-terminus), δ-OR (H-60, sc-9111, N-terminus), and κ-OR (H-70, sc-9112, N-terminus). Results represent the typical immunoblots of NGF-untreated <b>(A)</b> and NGF-treated <b>(C)</b> samples. The significance of difference between NGF-untreated (+M10) versus (─M10) PNS samples <b>(B)</b> and NGF-treated (+M10) versus (─M10) PNS samples <b>(D)</b> was analyzed by Student´s <i>t</i>-test using GraphPad<i>Prizm4</i> (NS, p>0.05). The results represent the average signal of three immunoblots, each performed with four control + four morphine-treated samples of PNS ± SEM. 100% on y-axis (lower panels) represents the average intensity of a given immunoblot signal determined in PNS prepared from control, (─M10) rats.</p

Determination of μ-, δ- and κ-OR in PNS prepared from experimental groups (+M10/─M20) and (─M10/─M20).

<p>PNS fractions prepared from (+M10/─M20) and (─M10/─M20) groups of rats were resolved under dissociated (+DTT) conditions by 1D-SDS-PAGE in 10% w/v acrylamide/0.26% w/v bis-acrylamide gels and the opioid receptors were recognized by antibodies from Santa Cruz: μ-OR (C-20, sc-7488-R, C-terminus), δ-OR (H-60, sc-9111, N-terminus), and κ-OR (H-70, sc-9112, N-terminus). As before, 20 μg of PNS protein was applied per each lane. The results (upper columns) represent the average signal of four immnoblots ± SEM. 100% on y-axis represents the average intensity of a given immunoblot signal determined in PNS prepared from control, (─M10) rats. The significance of difference between the (+M10/─M20) and (─M10/─M20) PNS samples was analyzed by Student´s <i>t</i>-test using GraphPad<i>Prizm4</i>. In the lower panels, typical immunoblots are shown (NS, p>0.05; *, p<0.05).</p

Detection of μ-OR, δ-OR and κ-OR in PNS fractions (±M10/─M20) treated with <i>N</i>-glycosidase F.

<p>PNS fractions prepared from FBC of rats in groups (+M10/─M20) and (— M10/─M20) were treated with SDS, Triton X-100 and <i>N</i>-glycosidase F (NGF) as described in Methods. 20 μg of protein was applied per each lane of 10% w/v acrylamide/0.26% w/v bis-acrylamide gels (four + four) and resolved under dissociated (+DTT) conditions. The OR were recognized by antibodies from Santa Cruz: μ-OR (C-20, sc-7488-R, C-terminus), δ-OR (H-60, sc-9111, N-terminus), and κ-OR (H-70, sc-9112, N-terminus). Results represent the typical immunoblots of NFG-untreated <b>(A)</b> and NGF-treated <b>(C)</b> samples. The significance of difference between NGF-untreated (+M10/─M20) versus (─M10/─M20) PNS samples <b>(B)</b> and NGF-treated (+M10/─M20) versus (─M10/─M20) PNS samples <b>(D)</b> was analyzed by Student´s <i>t</i>-test using GraphPad<i>Prizm4</i> (NS, p>0.05). The results represent the average signal of three immunoblots, each performed with four control + four morphine-treated samples of PNS ± SEM. 100% on y-axis (lower panels) represents the average intensity of a given immunoblot signal determined in PNS prepared from control, (─M10) rats.</p

Cholesterol amount in PNS prepared from FBC of control (─M10) and morphine-treated rats (+M10); comparison with animals sacrificed 20 days since the morphine withdrawal (±M10/─M20).

<p>Cholesterol amount in PNS prepared from FBC of control (─M10) and morphine-treated rats (+M10); comparison with animals sacrificed 20 days since the morphine withdrawal (±M10/─M20).</p