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

Oxidative potential associated with urban aerosol deposited into the respiratory system and relevant elemental and ionic fraction contributions

Size-segregated aerosol measurements were carried out at an urban and at an industrial site. Soluble and insoluble fractions of elements and inorganic ions were determined. Oxidative potential (OP) was assessed on the soluble fraction of Particulate Matter (PM) by ascorbic acid (AA), dichlorofluorescein (DCFH) and dithiothreitol (DTT) assays. Size resolved elemental, ion and OP doses in the head (H), tracheobronchial (TB) and alveolar (Al) regions were estimated using the Multiple-Path Particle Dosimetry (MPPD) model. The total aerosol respiratory doses due to brake and soil resuspension emissions were higher at the urban than at the industrial site. On the contrary, the doses of anthropic combustion tracers were generally higher at the industrial site. In general, the insoluble fraction was more abundantly distributed in the coarse than in the fine mode and vice versa for the soluble fraction. Consequently, for the latter, the percent of the total respiratory dose deposited in TB and Al regions increased. Oxidative potential assay (OPAA) doses were distributed in the coarse region; therefore, their major contribution was in the H region. The contribution in the TB and Al regions increased for OPDTT and OPDCFH

Element levels and predictors of exposure in the hair of Ethiopian children

Children’s development and health may be affected by toxic heavy metal exposure or suboptimal essential element intake. This study aimed to provide updated information regarding the concentrations of 41 elements in children’s hair (aged under 18) living in a rural area of the Benishangul-Gumuz region, Ethiopia. The highest average levels (as a geometric mean) for toxic heavy metals were obtained for Al (1 mg kg−1), Pb (3.1 mg kg−1), and Ni (1.2 mg kg−1), while the lowest concentrations among the essential elements were found for Co (0.32 mg kg−1), Mo (0.07 mg kg−1), Se (0.19 mg kg−1), and V (0.8 mg kg−1). Hair analysis was combined with a survey to evaluate relationships and variations among subgroups and potential metal exposure predictors. Females showed significantly higher concentrations for most hair elements, excluding Zn, than males, and the 6–11 years age group reported the highest levels for Be, Ce, Co, Fe, La, Li, Mo, and Na. The main predictors of exposure to toxic elements were fish consumption for Hg and drinking water for Ba, Be, Cs, Li, Ni, Tl, and U. The data from this study can be used to develop prevention strategies for children’s health and protection in developing countries

Dendritic Spike Saturation of Endogenous Calcium Buffer and Induction of Postsynaptic Cerebellar LTP

The architecture of parallel fiber axons contacting cerebellar Purkinje neurons retains spatial information over long distances. Parallel fiber synapses can trigger local dendritic calcium spikes, but whether and how this calcium signal leads to plastic changes that decode the parallel fiber input organization is unknown. By combining voltage and calcium imaging, we show that calcium signals, elicited by parallel fiber stimulation and mediated by voltage-gated calcium channels, increase non-linearly during high-frequency bursts of electrically constant calcium spikes, because they locally and transiently saturate the endogenous buffer. We demonstrate that these non-linear calcium signals, independently of NMDA or metabotropic glutamate receptor activation, can induce parallel fiber long-term potentiation. Two-photon imaging in coronal slices revealed that calcium signals inducing long-term potentiation can be observed by stimulating either the parallel fiber or the ascending fiber pathway. We propose that local dendritic calcium spikes, evoked by synaptic potentials, provide a unique mechanism to spatially decode parallel fiber signals into cerebellar circuitry changes

In situ physical and chemical characterisation of the Eyjafjallajökull aerosol plume in the free troposphere over Italy

Abstract. Continuous measurements of physical and chemical properties at the Mt. Cimone (Italy) GAW-WMO (Global Atmosphere Watch, World Meteorological Organization) Global Station (2165 m a.s.l.) have allowed the detection of the volcanic aerosol plume resulting from the Eyjafjallajökull (Iceland) eruption of spring 2010. The event affected the Mt. Cimone site after a transport over a distance of more than 3000 km. Two main transport episodes were detected during the eruption period, showing a volcanic fingerprint discernible against the free tropospheric background conditions typical of the site, the first from April 19 to 21 and the second from 18 to 20 May 2010. This paper reports the modification of aerosol characteristics observed during the two episodes, both characterised by an abrupt increase in fine and, especially, coarse mode particle number. Analysis of major, minor and trace elements by different analytical techniques (ionic chromatography, particle induced X-ray emission–particle induced gamma-ray emission (PIXE–PIGE) and inductively coupled plasma mass spectrometry (ICP-MS)) were performed on aerosols collected by ground-level discrete sampling. The resulting database allows the characterisation of aerosol chemical composition during the volcanic plume transport and in background conditions. During the passage of the volcanic plume, the fine fraction was dominated by sulphates, denoting the secondary origin of this mode, mainly resulting from in-plume oxidation of volcanic SO2. By contrast, the coarse fraction was characterised by increased concentration of numerous elements of crustal origin, such as Fe, Ti, Mn, Ca, Na, and Mg, which enter the composition of silicate minerals. Data analysis of selected elements (Ti, Al, Fe, Mn) allowed the estimation of the volcanic plume's contribution to total PM10, resulting in a local enhancement of up to 9.5 μg m−3, i.e. 40% of total PM10 on 18 May, which was the most intense of the two episodes. These results appear significant, especially in light of the huge distance of Mt. Cimone from the source, confirming the widespread diffusion of the Eyjafjallajökull ashes over Europe

In situ physical and chemical characterisation of the Eyjafjallajökull aerosol plume in the free troposphere over Italy

Continuous measurements of physical and chemical properties at the Mt. Cimone (Italy) GAW-WMO (Global Atmosphere Watch, World Meteorological Organization) Global Station (2165 m a.s.l.) have allowed the detection of the volcanic aerosol plume resulting from the Eyjafjallajökull (Iceland) eruption of spring 2010. The event affected the Mt. Cimone site after a transport over a distance of more than 3000 km. Two main transport episodes were detected during the eruption period, showing a volcanic fingerprint discernible against the free tropospheric background conditions typical of the site, the first from April 19 to 21 and the second from 18 to 20 May 2010. This paper reports the modification of aerosol characteristics observed during the two episodes, both characterised by an abrupt increase in fine and, especially, coarse mode particle number. Analysis of major, minor and trace elements by different analytical techniques (ionic chromatography, particle induced X-ray emission–particle induced gamma-ray emission (PIXE–PIGE) and inductively coupled plasma mass spectrometry (ICP-MS)) were performed on aerosols collected by ground-level discrete sampling. The resulting database allows the characterisation of aerosol chemical composition during the volcanic plume transport and in background conditions. During the passage of the volcanic plume, the fine fraction was dominated by sulphates, denoting the secondary origin of this mode, mainly resulting from in-plume oxidation of volcanic SO2. By contrast, the coarse fraction was characterised by increased concentration of numerous elements of crustal origin, such as Fe, Ti, Mn, Ca, Na, and Mg, which enter the composition of silicate minerals. Data analysis of selected elements (Ti, Al, Fe, Mn) allowed the estimation of the volcanic plume's contribution to total PM10, resulting in a local enhancement of up to 9.5 μg m−3, i.e. 40% of total PM10 on 18 May, which was the most intense of the two episodes. These results appear significant, especially in light of the huge distance of Mt. Cimone from the source, confirming the widespread diffusion of the Eyjafjallajökull ashes over Europe

Mapping the spatiotemporal dynamics of calcium signaling in cellular neural networks using optical flow

An optical flow gradient algorithm was applied to spontaneously forming net- works of neurons and glia in culture imaged by fluorescence optical microscopy in order to map functional calcium signaling with single pixel resolution. Optical flow estimates the direction and speed of motion of objects in an image between subsequent frames in a recorded digital sequence of images (i.e. a movie). Computed vector field outputs by the algorithm were able to track the spatiotemporal dynamics of calcium signaling pat- terns. We begin by briefly reviewing the mathematics of the optical flow algorithm, and then describe how to solve for the displacement vectors and how to measure their reliability. We then compare computed flow vectors with manually estimated vectors for the progression of a calcium signal recorded from representative astrocyte cultures. Finally, we applied the algorithm to preparations of primary astrocytes and hippocampal neurons and to the rMC-1 Muller glial cell line in order to illustrate the capability of the algorithm for capturing different types of spatiotemporal calcium activity. We discuss the imaging requirements, parameter selection and threshold selection for reliable measurements, and offer perspectives on uses of the vector data.Comment: 23 pages, 5 figures. Peer reviewed accepted version in press in Annals of Biomedical Engineerin

Integrated evaluation of indoor particulate exposure. The viepi project

Despite the progress made in recent years, reliable modeling of indoor air quality is still far from being obtained. This requires better chemical characterization of the pollutants and airflow physics included in forecasting tools, for which field observations conducted simultaneously indoors and outdoors are essential. The project “Integrated Evaluation of Indoor Particulate Exposure” (VIEPI) aimed at evaluating indoor air quality and exposure to particulate matter (PM) of humans in workplaces. VIEPI ran from February 2016 to December 2019 and included both numerical simulations and field campaigns carried out in universities and research environments located in urban and non-urban sites in the metropolitan area of Rome (Italy). VIEPI focused on the role played by micrometeorology and indoor airflow characteristics in determining indoor PM concentration. Short-and long-term study periods captured diurnal, weekly, and seasonal variability of airflow and PM concentration. Chemical characterization of PM10, including the determination of elements, ions, elemental carbon, organic carbon, and bioaerosol, was also carried out. Large differences in the composition of PM10 were detected between inside and outside as well as between different periods of the day and year. Indoor PM composition was related to the presence of people, to the season, and to the ventilation regime

Ultrafast force-clamp spectroscopy of single molecules reveals load dependence of myosin working stroke

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