Caffeine Modulates Vesicle Release and Recovery at Cerebellar Parallel Fibre Terminals,Independently of Calcium and Cyclic AMP Signalling

BackgroundCerebellar parallel fibres release glutamate at both the synaptic active zone and at extrasynaptic sites—a process known as ectopic release. These sites exhibit different short-term and long-term plasticity, the basis of which is incompletely understood but depends on the efficiency of vesicle release and recycling. To investigate whether release of calcium from internal stores contributes to these differences in plasticity, we tested the effects of the ryanodine receptor agonist caffeine on both synaptic and ectopic transmission.MethodsWhole cell patch clamp recordings from Purkinje neurons and Bergmann glia were carried out in transverse cerebellar slices from juvenile (P16-20) Wistar rats.ConclusionsWe conclude that caffeine increases release probability and inhibits vesicle recovery at parallel fibre synapses, independently of known pharmacological targets. This complex effect would lead to potentiation of transmission at fibres firing at low frequencies, but depression of transmission at high frequency connections.     

The conserved GTPase LepA contributes mainly to translation initiation in Escherichia coli

LepA is a paralog of EF-G found in all bacteria. Deletion of lepA confers no obvious growth defect in Escherichia coli, and the physiological role of LepA remains unknown. Here, we identify nine strains (ΔdksA, ΔmolR1, ΔrsgA, ΔtatB, ΔtonB, ΔtolR, ΔubiF, ΔubiG or ΔubiH) in which ΔlepA confers a synthetic growth phenotype. These strains are compromised for gene regulation, ribosome assembly, transport and/or respiration, indicating that LepA contributes to these functions in some way. We also use ribosome profiling to deduce the effects of LepA on translation. We find that loss of LepA alters the average ribosome density (ARD) for hundreds of mRNA coding regions in the cell, substantially reducing ARD in many cases. By contrast, only subtle and codon-specific changes in ribosome distribution along mRNA are seen. These data suggest that LepA contributes mainly to the initiation phase of translation. Consistent with this interpretation, the effect of LepA on ARD is related to the sequence of the Shine–Dalgarno region. Global perturbation of gene expression in the ΔlepA mutant likely explains most of its phenotypes.     

The role of phosphoenolpyruvate carboxykinase in neuronal steroidogenesis under acute inflammation

Phosphoenolpyruvate carboxykinase (PEPCK) is a key gluconeogenic enzyme found in many tissues throughout the body including brain. In the present study, we have investigated the effect of bacterial lipopolysaccharide (LPS) on PEPCK and its role in neuronal steroidogenesis. Adult female albino rats were administered LPS (5 mg/kg body weight) to induce acute inflammation. LPS administration resulted in a significant increase of PEPCK mRNA expression with concomitant increase in mRNA levels of steroidogenic acute regulatory (StAR) protein and other steroidogenic enzymes including 3β-hydroxysteroid dehydrogenase (3β-HSD), 17β-hydroxysteroid dehydrogenase (17β-HSD) and aromatase in brain tissue. Further, the inhibition of PEPCK expression by glipizide significantly decreased the mRNA expression of steroidogenic proteins and concurrently increased the mRNA levels of proinflammatory cytokines under LPS administration. The results of this study suggest a novel finding that PEPCK may have an important role in neuronal steroidogenesis; which serves as an adaptive response under inflammation.     

Evaluation of the nutritional profile and antioxidant and anti-cholinesterase activities of Padina gymnospora (Phaeophyceae)

The use of cholinesterase inhibitors and antioxidants is currently considered as an effective therapeutic strategy for the treatment of Alzheimer’s disease (AD). There is also a growing trend to use nutraceuticals for cognitive impairment. Since natural product-based drugs offer better hope for the treatment of AD, the present study was aimed at evaluating the nutritional profile of the brown seaweed Padina gymnospora and investigating the antioxidant and inhibitory effect of different solvent extracts of P. gymnospora on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) enzymes. The nutritional profile of P. gymnospora includes large amounts of carbohydrate, protein, lipid, proline, chlorophyll, fibre, minerals, fatty acids and amino acids, along with antioxidants such as vitamins C and E. Antioxidant activities of various solvent extracts of P. gymnospora were assessed by the DPPH scavenging assay, nitric oxide scavenging assay, reducing power assay and FRAP assay. Acetone extracts showed significant DPPH radical and nitric oxide scavenging activity with IC₅₀ values of 402 ± 9.12 and 441 ± 48.16 µg ml^–1 respectively and aqueous extracts had better reducing potential. Among the different solvent extracts, the acetone extract showed the highest inhibitory activity with IC₅₀ values <150 µg ml^–1 for both AChE and BuChE. Qualitative phytochemical screening of P. gymnospora revealed the presence of flavonoids and cardiac glycosides. Overall the results suggest that the seaweed Padina gymnospora may be a good nutraceutical candidate for discovering drugs against AD.     

Development of an antibody against a 170-kDa fragment of fibroin isolated from cocoon fibres of Bombyx mori.

A simple method for the isolation of denatured fragments of the fibroin protein from cocoon fibres by alkali solubilization is discussed. This 170-kDa antigen has been purified and used to raise the polyclonal antibody in rabbit. The specificity of this antibody to the purified cocoon protein has suggested strong immunoreactivity up to a titre of 1:5000 dilution of the antibody. Further, dot-blot analysis with the tissue extracts from silk glands of different larval stages (3rd to 5th) reveals that this antibody reacts showing a stage-specific increase in the intensity of the colour, correlating well with the in-vivo expression of the silk protein. This study suggests the availability of a specific polyclonal antibody that detects the native fibroin with no crossreactivity with other tissue proteins.     

Algorithm to find distant repeats in a single protein sequence

Distant repeats in protein sequence play an important role in various aspects of protein analysis. A keen analysis of the distant repeats would enable to establish a firm relation of the repeats with respect to their function and three-dimensional structure during the evolutionary process. Further, it enlightens the diversity of duplication during the evolution. To this end, an algorithm has been developed to find all distant repeats in a protein sequence. The scores from Point Accepted Mutation (PAM) matrix has been deployed for the identification of amino acid substitutions while detecting the distant repeats. Due to the biological importance of distant repeats, the proposed algorithm will be of importance to structural biologists, molecular biologists, biochemists and researchers involved in phylogenetic and evolutionary studies.     

A new mutation, 3905insT, accounts for 4.8% of 1173 CF chromosomes in Switzerland and causes a severe phenotype

We have analysed 1173 cystic fibrosis (CF) chromosomes from Switzerland for eight mutations in the CF transmembrane conductance regulator (CFTR) gene. This permitted the identification of 88.5% of all mutations present. A novel insertion mutation in exon 20 of the CFTR gene, 3905insT, was discovered. This mutation accounted for 4.8% of CFTR gene mutations in Switzerland and has since been identified in other populations of probable Swiss descent. It is associated with a highly variable clinical phenotype but always with pancreatic insufficiency. Haplotype analysis with three intragenic microsatellites in the CFTR gene showed that the mutation is associated with a haplotype rarely identified on other CFTR alleles and, therefore, that the frequency of the mutation in Switzerland is explained by a founder effect of a relatively recent mutation event. Received: 17 February 1997 / Accepted: 26 March 1977