Efficient modulation of of γ-aminobutyric acid type A (GABAA) receptors by piperine derivatives

[Image: see text] Piperine activates TRPV1 (transient receptor potential vanilloid type 1 receptor) receptors and modulates γ-aminobutyric acid type A receptors (GABA(A)R). We have synthesized a library of 76 piperine analogues and analyzed their effects on GABA(A)R by means of a two-microelectrode voltage-clamp technique. GABA(A)R were expressed in Xenopus laevis oocytes. Structure–activity relationships (SARs) were established to identify structural elements essential for efficiency and potency. Efficiency of piperine derivatives was significantly increased by exchanging the piperidine moiety with either N,N-dipropyl, N,N-diisopropyl, N,N-dibutyl, p-methylpiperidine, or N,N-bis(trifluoroethyl) groups. Potency was enhanced by replacing the piperidine moiety by N,N-dibutyl, N,N-diisobutyl, or N,N-bistrifluoroethyl groups. Linker modifications did not substantially enhance the effect on GABA(A)R. Compound 23 [(2E,4E)-5-(1,3-benzodioxol-5-yl)-N,N-dipropyl-2,4-pentadienamide] induced the strongest modulation of GABA(A) (maximal GABA-induced chloride current modulation (I(GABA-max) = 1673% ± 146%, EC(50) = 51.7 ± 9.5 μM), while 25 [(2E,4E)-5-(1,3-benzodioxol-5-yl)-N,N-dibutyl-2,4-pentadienamide] displayed the highest potency (EC(50) = 13.8 ± 1.8 μM, I(GABA-max) = 760% ± 47%). Compound 23 induced significantly stronger anxiolysis in mice than piperine and thus may serve as a starting point for developing novel GABA(A)R modulators

Physiological and biochemical responses to low non-freezing temperature of two Eucalyptus globulus clones differing in drought resistance

Abstract – We have compared the metabolic responses of leaves and roots of two Eucalyptus globulus L. clones CN5 and ST51 that differ in their sensitivity to water deficits (ST51 is more drought sensitive), with regard to the effect of chilling (10/5 ◦C, day/night). We studied changes in growth, osmotic potential and osmotically active compounds, soluble proteins, leaf pigments, and membrane lipid composition. Our data showed that both clones have the ability to acclimatize to chilling temperatures. As a result of 10 days of acclimation, an increase of soluble sugars in leaves of treated plants of both clones was observed that disappeared later on. Differences between clones were observed in the photosynthetic pigments and soluble protein content which were more stable in CN5 under chilling. It also was apparent that CN5 presented a less negative predawn water potential (ψpd) and a higher leaf turgor than ST51 throughout the chilling treatment. In the case of the CN5, increased total lipids (TFA) and concomitant increase of linolenic acid (C18:3) in leaves after acclimatization may be related to a better clone performance under chilling temperatures. Moreover, a higher constitutive investment in roots in the case of CN5 as compared to ST51 may benefit new root regeneration under low temperatures favoring growth after cold Mediterranean winter

Scorpion sting: a public health problem in El Kelaa des Sraghna (Morocco)

The present study aimed at verifying the impact of a Moroccan strategy against scorpion stings and specifically at identifying the epidemiological features of patients envenomed or just stung by scorpions. It included 11,907 patients from El Kelaa des Sraghna Province, Morocco, who were evaluated over five years (2001-2005). Most stings occurred during the hot period and mainly at night. The average incidence was 3.2 per 1,000 inhabitants; patients <15 years accounted for 34%, and the envenomation rate was 12%. Average lethality rate was 0.7%. Our work evaluated the efficacy of the adopted strategy based on indicators of follow-up, morbidity and lethality due to scorpion sting and envenomation

Dynamics of Responses in Compatible Potato - Potato virus Y Interaction Are Modulated by Salicylic Acid

To investigate the dynamics of the potato – Potato virus Y (PVY) compatible interaction in relation to salicylic acid - controlled pathways we performed experiments using non-transgenic potato cv. Désirée, transgenic NahG-Désirée, cv. Igor and PVYNTN, the most aggressive strain of PVY. The importance of salicylic acid in viral multiplication and symptom development was confirmed by pronounced symptom development in NahG-Désirée, depleted in salicylic acid, and reversion of the effect after spraying with 2,6-dichloroisonicotinic acid (a salicylic acid - analogue). We have employed quantitative PCR for monitoring virus multiplication, as well as plant responses through expression of selected marker genes of photosynthetic activity, carbohydrate metabolism and the defence response. Viral multiplication was the slowest in inoculated potato of cv. Désirée, the only asymptomatic genotype in the study. The intensity of defence-related gene expression was much stronger in both sensitive genotypes (NahG-Désirée and cv. Igor) at the site of inoculation than in asymptomatic plants (cv. Désirée). Photosynthesis and carbohydrate metabolism gene expression differed between the symptomatic and asymptomatic phenotypes. The differential gene expression pattern of the two sensitive genotypes indicates that the outcome of the interaction does not rely simply on one regulatory component, but similar phenotypical features can result from distinct responses at the molecular level

Four-week short chain fructo-oligosaccharides ingestion leads to increasing fecal bifidobacteria and cholesterol excretion in healthy elderly volunteers

<p>Abstract</p> <p>Background</p> <p>Short-chain fructo-oligosaccharides (scFOS) are increasingly used in human diet for their prebiotic properties. We aimed at investigating the effects of scFOS ingestion on the colonic microflora and oro-fecal transit time in elderly healthy humans.</p> <p>Methods</p> <p>Stools composition, oro-fecal transit time, and clinical tolerance were evaluated in 12 healthy volunteers, aged 69 ± 2 yrs, in three consecutive periods: basal period (2 weeks), scFOS (Actilight^®) ingestion period (8 g/d for 4 weeks) and follow-up period (4 weeks). Two-way ANOVA, with time and treatment as factors, was used to compare the main outcome measures between the three periods.</p> <p>Results</p> <p>Fecal bifidobacteria counts were significantly increased during the scFOS period (9.17 ± 0.17 log cfu/g vs 8.52 ± 0.26 log cfu/g during the basal period) and returned to their initial values at the end of follow-up (8.37 ± 0.21 log cfu/g; P < 0.05). Fecal cholesterol concentration increased during the scFOS period (8.18 ± 2.37 mg/g dry matter vs 2.81 ± 0.94 mg/g dry matter during the basal period) and returned to the baseline value at the end of follow-up (2.87 ± 0.44 mg/g dry matter; P < 0.05). Fecal pH tended to decrease during scFOS ingestion and follow-up periods compared to the basal period (P = 0.06). Fecal bile acids, stool weight, water percentage, and oro-fecal transit time did not change throughout the study. Excess flatus and bloating were significantly more frequent during scFOS ingestion when compared to the basal period (P < 0.05), but the intensity of these symptoms was very mild.</p> <p>Conclusion</p> <p>Four-week 8 g/d scFOS ingestion is well tolerated and leads to a significant increase in fecal bifidobacteria in healthy elderly subjects. Whether the change in cholesterol metabolism found in our study could exert a beneficial action warrants further studies.</p

Pharmacokinetics and metabolism of dietary kaempferol and its metabolite 4-hydroxy-phenylacetic acid in rats

Scope Kaempferol is a major flavonoid in the human diet and in medicinal plants. The compound exerts anxiolytic activity when administered orally in mice, while no behavioural changes were observed upon intraperitoneal administration, or upon oral administration in gut sterilized animals. 4-Hydroxyphenylacetic acid (4-HPAA), which possesses anxiolytic effects when administered intraperitoneally, is a major intestinal metabolite of kaempferol. Pharmacokinetic properties of the compounds are currently not clear. Methods and results UHPLC-MS/MS methods were validated to support pharmacokinetic studies of kaempferol and 4-HPAA in rats. Non-compartmental and compartmental analyses were performed. After intravenous administration, kaempferol followed a one-compartment model, with a rapid clearance (4.40–6.44 l/h/kg) and an extremely short half-life of 2.93–3.79 min. After oral gavage it was not possible to obtain full plasma concentration–time profiles of kaempferol. Pharmacokinetics of 4-HPAA was characterized by a two-compartment model, consisting of a quick distribution phase (half-life 3.04–6.20 min) followed by a fast elimination phase (half-life 19.3–21.1 min). Conclusion Plasma exposure of kaempferol is limited by poor oral bioavailability and extensive metabolism. Both compounds are rapidly eliminated, so that effective concentrations at the site of action do not appear to be reached. At present, it is not clear how the anxiolytic-like effects reported for the compounds can be explained

Protein and carbohydrate analyses of abiotic stress underlying cryopreservation in potato

Cryopreservation complements classical conservation methods, which are carried out in the field or in vitro. It involves the storage of biological material in liquid nitrogen (- 196°C). At this temperature all chemical and physical processes are stopped, allowing a safe storage over an unlimited period of time. The aim of our work is to understand the effects of pretreatments, by increasing the osmotic pressure of the medium, on the cryopreservation ability of potato shoot tips and more specifically to try to evaluate the effects of such pretreatments on the metabolism of potato. Indeed, drought acclimation is known to improve recovery after cryopreservation in potato and other species In vitro Désirée potato shoots were precultured for 21 days on regular MS and MS complemented with 0.055M, 0.11M, and 0.22M sorbitol. Directly after pre-treatment, leaf and shoot tip samples were taken and stored at 80°C for proteomic and carbohydrate analyses. In addition, cryopreservation was carried on the precultured shoot-tips. For the cryopreservation, potato shoot tips were cut from pre-treated plants and incubated in a high osmotic Loading Solution. Afterwards, shoot tips were placed on an aluminum foil strip in droplets of Plant Vitrification Solution 2 and plunged into liquid nitrogen (Agrawal, 2004). Thawing was done in a highly osmotic Recovery Solution at room temperature, to prevent osmotic shock. After cryopreservation, shoot tips were transferred into the dark for 1 week (Panis, 2005). During the initial days of post-culture, shoot tips were maintained on MS media containing 0.3M sucrose. Afterwards, regular MS media were used. After 30 days, recovery was calculated as the percentage of shoot-tips forming new shoot. Proteins from shoot tip and leaf samples were extracted, using a TCA/Acetone ex-traction method. After quantification, 40 mg protein was labelled using three different fluorescent dyes (Cy2, Cy3 and Cy5). In this way, 2 samples and an internal standard - containing a mix of all the samples - was loaded on the same IEF strip (pI 4-7). Isoelectric focusing was carried out using the GE Healthcare Ettan IPGphor. The second dimension was run in a GE Healthcare Ettan Dalt Six electrophoresis system. Gels were scanned using the Typhoon 9400 scanner and subsequently analysed with the GE Healthcare DeCyder program and EDA module. As such, differences in protein expression could be quantified and differentially expressed spots picked by the GE Healthcare Ettan Spot Handling Workstation (Renaut, 2006). In a following step, spots were digested, using Trypsin and spotted on a MALDI plate. Protein identification was done, using the Applied Biosystems MALDI 4800 TOF/TOF analyzer. For the analysis of carbohydrates and polyols, roughly 100 mg leaf samples was used for carbohydrate and polyol extractions. The following carbohydrates and polyos were measured: sucrose, glucose, fructose, galactose, stachyose, arabinose, melibiose, maltose, trehalose, inositol, mannitol, galactinol and sorbitol. Carbohydrates were analyzed on a Dionex HPLC ICS2500-Bio LC, using a Carbopac PA-20 column. HPAEC-PAD analyses for polyols was conducted on a Dionex DX-500 chromatograph, using a Dionex Car-bopac MA1 column. Recovery rates without sorbitol were around 50% but increased with increasing concentration of sorbitol pre-treatment up to a recovery rate up to 80%. For 2D-DIGE proteomics, preliminary analysis of the gels showed differences in protein patterns, when plants were precultured on different sorbitol media. Fifteen up- or downregulated proteins were isolated and identified. Interestingly, these preliminary results indicate strong alteration of the primary metabolism and more precisely in carbon fixation. These results are sustained by the carbohydrate and polyol analyses. Indeed, sucrose, glucose, fructose, mannitol, arabinose, galactinol, meli-biose and stachyose increased with increasing concentrations of exogenousely supplied sorbitol up to 0.11M sorbitol. At 0.22M sorbitol their concentrations decreased. Trehalose and sorbitol concentrations increased exponentially with an increasing molarity of sorbitol pretreatment. Maltose was only observed when plantlets were treated with 0.22M sorbitol. When plants were treated with up to 0.11M sorbitol during 21 days, carbohydrate and polyol concentrations increased. When 0.22M sorbitol was applied as pre-treatment, most sugar concentrations decreased. Since high intracellular osmolyte concentrations are needed to allow successful recovery after cryopreservation, the results from carbohydrate and polyol analysis (Bhandal, 1985) may explain the higher recovery rate after cryopreservation observed after sorbitol treatment. Extra cryopreservation experiments are needed to confirm these results. Beside the sugar analysis, these changes affecting the primary metabolism have been observed at the protein level by using differential in gel electrophoresis. Further experiments including also chilling pretreatments will lead to a better understanding of the physiological status of the tissue and will hopefully explain the reason of the better results of cryopreservation.vokMyyynti MTT, Tietopalvelut 31600 Jokioine