Erythrina mulungu Alkaloids Are Potent Inhibitors of Neuronal Nicotinic Receptor Currents in Mammalian Cells

Crude extracts and three isolated alkaloids from Erythrina mulungu plants have shown anxiolytic effects in different animal models. We investigated whether these alkaloids could affect nicotinic acetylcholine receptors and if they are selective for different central nervous system (CNS) subtypes. Screening experiments were performed using a single concentration of the alkaloid co-applied with acetylcholine in whole cell patch-clamp recordings in three different cell models: (i) PC12 cells natively expressing α3* nicotinic acetylcholine receptors; (ii) cultured hippocampal neurons natively expressing α7* nicotinic acetylcholine receptors; and (iii) HEK 293 cells heterologoulsy expressing α4β2 nicotinic acetylcholine receptors. For all three receptors, the percent inhibition of acetylcholine-activated currents by (+)-11á-hydroxyerysotrine was the lowest, whereas (+)-erythravine and (+)-11á-hydroxyerythravine inhibited the currents to a greater extent. For the latter two substances, we obtained concentration-response curves with a pre-application protocol for the α7* and α4β2 nicotinic acetylcholine receptors. The IC₅₀ obtained with (+)-erythravine and (+)-11á-hydroxyerythravine were 6 µM and 5 µM for the α7* receptors, and 13 nM and 4 nM for the α4β2 receptors, respectively. Our data suggest that these Erythrina alkaloids may exert their behavioral effects through inhibition of CNS nicotinic acetylcholine receptors, particularly the α4β2 subtype.     

Endocytic SNAREs are involved in optimal Coxiella burnetii vacuole development

Coxiella burnetii is a Gram‐negative intracellular bacterium. As previously described, both the endocytic and the autophagic pathways contribute to the maturation of Coxiella replicative vacuoles (CRVs). The large CRVs share the properties of both phagolysosomal and autophagolysosomal compartments. Vamp3, Vamp7 and Vamp8 are v‐SNAREs involved in the endocytic pathway which participate mainly in the fusion between endosomes and lysosomes. In the present study we observed that Vamp7 interacts with C. burnetii at different infection times (1 h–48 h p.i.). We have determined that a truncated mutant of Vamp7 (Vamp7 NT) and a siRNA against this SNARE protein affects the optimal development of CRVs, suggesting that Vamp7 mediates fusion events that are required for the biogenesis of CRVs. Indeed, we have observed that overexpression of Vamp7 NT inhibited the heterotypic fusion with lysosomes and the homotypic fusion between individual Coxiella phagosomes and CRVs. Moreover, we have detected in the vacuole membrane, at different infection times, the Vamp7 partners (Vti1a and Vti1b). Interestingly, treatment with chloramphenicol reduced the colocalization between C. burnetii and Vamp7, Vti1a or Vti1b, indicating that the recruitment of these SNAREs proteins is a bacteria‐driven process that favours the CRV biogenesis, likely by facilitating the interaction with the endolysosomal compartment.     

Energy Nonequipartition in a Sheared Granular Mixture

The kinetic granular temperatures of a binary granular mixture in simple shear flow are calculated by means of the Direct Simulation Monte Carlo method. The results show that the temperature ratio is clearly different from unity (as may be expected since the system is out of equilibrium) and strongly depends on the restitution coefficients as well as on the parameters of the mixture. The influence of the temperature differences on the rheological properties is also discussed. The results are compared with the theoretical predictions obtained from the Boltzmann kinetic theory by using a Sonine polynomial expansion. The comparison shows an excellent agreement over the range of parameters investigated.     

Isolation and identification of a pepsitensin

1. From ox plasma incubated with pepsin a highly purified pepsitensin has been isolated by fractional precipitation, solvent extraction, column chromatography, countercurrent distribution and paper chromatography. 2. Comparison of the properties of this substance with those of synthetic valyl-5 angiotensin I showed: (a) the same specific pressor activity; (b) the same amino acid composition; (c) identical paper-electrophoretic mobilities at various pH values. 3. N- and C-Terminal studies carried out on the intact polypeptide and on the products of chymotrypsin digestion established the following sequence for the amino acids present in the molecule: Asp-Arg-Val-Tyr-Val-His-Pro-Phe-His-Leu. This structure is identical with that of valyl-5 angiotensin I.     

Effects of thyroidectomy and submaxillary sialoadenectomy on physiological parameters in the female rat

The effects of interaction of the thyroid function with the submaxillary glands on the female rat, on the body growth, weight evolution of adrenal glands, ovary and parotid glands, puberty onset, estrous cycle, fertility, gestation and offspring size, plasmatic levels of glucose, glucagon, Na+, K+ and total protein have been studied. The effects of sialadenectomy and thyroid function are both independent in all parameters, except in ovary growth, which is depressed by thyroid function when submaxillary glands are removed, and in circulating glucose, since sialadenectomy stabilizes to hypoglycemia which results from thyroidectomy.     

Seasonal physiological responses of Argania spinosa tree from Mediterranean to semi-arid climate

Argania spinosa (the argan tree) is a slow-growing tree endemic of Morocco, growing on semi-arid areas where no other tree species can live. With the aim of predicting temporal changes in A. spinosa woodlands under a probable increase in aridity, we set off to investigate these questions: how do A. spinosa physiological attributes respond to variations in climatic conditions and seasonality, and which is the set of attributes that most affects tree response to environmental conditions? In three study sites, Beni Snassen (North), High-Atlas (Mountain) and Admine Forest in Agadir (Coastal), gas exchange measurements, photochemical efficiency, leaf water potential and different leaf attributes were monitored in February, July and November of 2006. The Mountain site presents the most continental climate. Trees in this site were the most stressed in summer, having the lowest midday leaf water potential values, photochemical efficiency and assimilation rates. We found a Ψ_md threshold around -4 MPa, below which stomatal conductance responds linearly to Ψ_md. Plants from the North area never reached this threshold during the study period. Although leaf pigments presented a clear seasonal pattern, leaves from Coastal trees exhibit the highest content for each season. The three study sites were separated by two discriminate functions obtained by canonical discriminant analysis. In summer, the Mountain population is separated from the other sites mainly by assimilation rate and F_v/F_m, while in winter transpiration rates and chlorophyll content are the main discriminant variables. Our study shows that A. spinosa trees adjust their physiological status and leaf attributes to environmental conditions allowing plants to thrive under a dry climate. Under a scenario of global change, the distribution of the argan tree likely shifts to milder areas.     

AFLPs Reveal Different Population Genetic Structure under Contrasting Environments in the Marine Snail Nucella lapillus L.

Dispersal has received growing attention in marine ecology, particularly since evidence obtained with up-to-date techniques challenged the traditional view. The dogwhelk Nucella lapillus L., a sedentary gastropod with direct development, is a good example: dispersal was traditionally assumed to be limited until studies with microsatellites disputed this idea. To shed some light on this controversy, the genetic structure of dogwhelk populations in northwest Spain was investigated with highly polymorphic AFLP markers giving special attention to the influence of hydrodynamic stress. In agreement with the expectations for a poor disperser, our results show a significant genetic structure at regional (<200 km) and areal scales (<15 km). However, the spatial genetic structure varied with wave-exposure in the present case study: IBD was evident under sheltered conditions but absent from the exposed area where genetic differentiation was stronger. Our results provide evidence that differences in wave-exposure can exert a detectable influence on the genetic structure of coastal organisms, even in species without a planktonic larva.     

Clonal Human Fetal Ventral Mesencephalic Dopaminergic Neuron Precursors for Cell Therapy Research

A major challenge for further development of drug screening procedures, cell replacement therapies and developmental studies is the identification of expandable human stem cells able to generate the cell types needed. We have previously reported the generation of an immortalized polyclonal neural stem cell (NSC) line derived from the human fetal ventral mesencephalon (hVM1). This line has been biochemically, genetically, immunocytochemically and electrophysiologically characterized to document its usefulness as a model system for the generation of A9 dopaminergic neurons (DAn). Long-term in vivo transplantation studies in parkinsonian rats showed that the grafts do not mature evenly. We reasoned that diverse clones in the hVM1 line might have different abilities to differentiate. In the present study, we have analyzed 9 hVM1 clones selected on the basis of their TH generation potential and, based on the number of v-myc copies, v-myc down-regulation after in vitro differentiation, in vivo cell cycle exit, TH⁺ neuron generation and expression of a neuronal mature marker (hNSE), we selected two clones for further in vivo PD cell replacement studies. The conclusion is that homogeneity and clonality of characterized NSCs allow transplantation of cells with controlled properties, which should help in the design of long-term in vivo experiments.     

α-Synuclein Levels in Blood Plasma from LRRK2 Mutation Carriers

The diagnosis of Parkinson’s disease (PD) remains primarily a clinical issue, based mainly on phenotypic patterns. The identification of biomarkers capable of permitting the preclinical detection of PD is critically needed. α-Synuclein is a key protein in PD, with missense and multiplication mutations in the gene encoding α-synuclein (SNCA) having been reported in familial cases of PD, and accumulation of the protein identified in Lewy bodies (LBs) and Lewy neurites (LNs) in affected brain regions. With the objective of validating the use of α-synuclein as a clinical or progressive biomarker in an accessible tissue, we used an enzyme-linked immunosorbent assay (ELISA) to measure α-synuclein levels in the peripheral blood plasma of idiopathic PD and LRRK2 mutation carrier patients and compared our findings with healthy control subjects. Compared to healthy controls, we found a significant decrease in plasma total α-synuclein levels in idiopathic PD (iPD) patients (n = 134, p = 0.010). However, the reduction was less significant in patients who were LRRK2 mutation carriers (n = 32, p = 0.133). This lack of significance could be due to the small number of individuals employed in this group. No predictive value of total α-synuclein in the diagnosis of PD was found in a receiver operating characteristic (ROC) curve analysis. Although this is a pilot study requiring corroboration on a larger cohort of patients, our results highlight the possible use of plasma α-synuclein as a biomarker for PD.