Combining Membrane Potential Imaging with l-Glutamate or GABA Photorelease

Combining membrane potential imaging using voltage sensitive dyes with photolysis of l-glutamate or GABA allows the monitoring of electrical activity elicited by the neurotransmitter at different sub-cellular sites. Here we describe a simple system and some basic experimental protocols to achieve these measurements. We show how to apply the neurotransmitter and how to vary the dimension of the area of photolysis. We assess the localisation of photolysis and of the recorded membrane potential changes by depolarising the dendrites of cerebellar Purkinje neurons with l-glutamate photorelease using different experimental protocols. We further show in the apical dendrites of CA1 hippocampal pyramidal neurons how l-glutamate photorelease can be used to calibrate fluorescence changes from voltage sensitive dyes in terms of membrane potential changes (in mV) and how GABA photorelease can be used to investigate the phenomenon of shunting inhibition. We also show how GABA photorelease can be used to measure chloride-mediated changes of membrane potential under physiological conditions originating from different regions of a neuron, providing important information on the local intracellular chloride concentrations. The method and the proof of principle reported here open the gateway to a variety of important applications where the advantages of this approach are necessary

Cryo Electron Tomography of Native HIV-1 Budding Sites

The structure of immature and mature HIV-1 particles has been analyzed in detail by cryo electron microscopy, while no such studies have been reported for cellular HIV-1 budding sites. Here, we established a system for studying HIV-1 virus-like particle assembly and release by cryo electron tomography of intact human cells. The lattice of the structural Gag protein in budding sites was indistinguishable from that of the released immature virion, suggesting that its organization is determined at the assembly site without major subsequent rearrangements. Besides the immature lattice, a previously not described Gag lattice was detected in some budding sites and released particles; this lattice was found at high frequencies in a subset of infected T-cells. It displays the same hexagonal symmetry and spacing in the MA-CA layer as the immature lattice, but lacks density corresponding to NC-RNA-p6. Buds and released particles carrying this lattice consistently lacked the viral ribonucleoprotein complex, suggesting that they correspond to aberrant products due to premature proteolytic activation. We hypothesize that cellular and/or viral factors normally control the onset of proteolytic maturation during assembly and release, and that this control has been lost in a subset of infected T-cells leading to formation of aberrant particles

Marine Cyanobacteria Compounds with Anticancer Properties: Implication of Apoptosis

Marine cyanobacteria have been proved to be an important source of potential anticancer drugs. Although several compounds were found to be cytotoxic to cancer cells in culture, the pathways by which cells are affected are still poorly elucidated. For some compounds, cancer cell death was attributed to an implication of apoptosis through morphological apoptotic features, implication of caspases and proteins of the Bcl-2 family, and other mechanisms such as interference with microtubules dynamics, cell cycle arrest and inhibition of proteases other than caspases

Benign external hydrocephalus: a review, with emphasis on management

Benign external hydrocephalus in infants, characterized by macrocephaly and typical neuroimaging findings, is considered as a self-limiting condition and is therefore rarely treated. This review concerns all aspects of this condition: etiology, neuroimaging, symptoms and clinical findings, treatment, and outcome, with emphasis on management. The review is based on a systematic search in the Pubmed and Web of Science databases. The search covered various forms of hydrocephalus, extracerebral fluid, and macrocephaly. Studies reporting small children with idiopathic external hydrocephalus were included, mostly focusing on the studies reporting a long-term outcome. A total of 147 studies are included, the majority however with a limited methodological quality. Several theories regarding pathophysiology and various symptoms, signs, and clinical findings underscore the heterogeneity of the condition. Neuroimaging is important in the differentiation between external hydrocephalus and similar conditions. A transient delay of psychomotor development is commonly seen during childhood. A long-term outcome is scarcely reported, and the results are varying. Although most children with external hydrocephalus seem to do well both initially and in the long term, a substantial number of patients show temporary or permanent psychomotor delay. To verify that this truly is a benign condition, we suggest that future research on external hydrocephalus should focus on the long-term effects of surgical treatment as opposed to conservative management