Leptin counteracts the hypoxia-induced inhibition of spontaneously firing hippocampal neurons: a microelectrode array study.

Besides regulating energy balance and reducing body-weight, the adipokine leptin has been recently shown to be neuroprotective and antiapoptotic by promoting neuronal survival after excitotoxic and oxidative insults. Here, we investigated the firing properties of mouse hippocampal neurons and the effects of leptin pretreatment on hypoxic damage (2 hours, 3% O(2)). Experiments were carried out by means of the microelectrode array (MEA) technology, monitoring hippocampal neurons activity from 11 to 18 days in vitro (DIV). Under normoxic conditions, hippocampal neurons were spontaneously firing, either with prevailing isolated and randomly distributed spikes (11 DIV), or with patterns characterized by synchronized bursts (18 DIV). Exposure to hypoxia severely impaired the spontaneous activity of hippocampal neurons, reducing their firing frequency by 54% and 69%, at 11 and 18 DIV respectively, and synchronized their firing activity. Pretreatment with 50 nM leptin reduced the firing frequency of normoxic neurons and contrasted the hypoxia-induced depressive action, either by limiting the firing frequency reduction (at both ages) or by increasing it to 126% (in younger neurons). In order to find out whether leptin exerts its effect by activating large conductance Ca(2+)-activated K(+) channels (BK), as shown on rat hippocampal neurons, we applied the BK channel blocker paxilline (1 µM). Our data show that paxilline reversed the effects of leptin, both on normoxic and hypoxic neurons, suggesting that the adipokine counteracts hypoxia through BK channels activation in mouse hippocampal neurons

Heterogeneous distribution of exocytotic microdomains in adrenal chromaffin cells resolved by high-density diamond ultra-microelectrode arrays

Here we describe the ability of a high-density diamond microelectrode array targeted to resolve multi-site detection of fast exocytotic events from single cells. The array consists of nine boron-doped nanocrystalline diamond ultra-microelectrodes (9-Ch NCD-UMEA) radially distributed within a circular area of the dimensions of a single cell. The device can be operated in voltammetric or chronoamperometric configuration. Sensitivity to catecholamines, tested by dose–response calibrations, set the lowest detectable concentration of adrenaline to ∼5 μm. Catecholamine release from bovine or mouse chromaffin cells could be triggered by electrical stimulation or external KCl-enriched solutions. Spikes detected from the cell apex using carbon fibre microelectrodes showed an excellent correspondence with events measured at the bottom of the cell by the 9-Ch NCD-UMEA, confirming the ability of the array to resolve single quantal secretory events. Subcellular localization of exocytosis was provided by assigning each quantal event to one of the nine channels based on its location. The resulting mapping highlights the heterogeneous distribution of secretory activity in cell microdomains of 12–27 μm(2). In bovine chromaffin cells, secretion was highly heterogeneous with zones of high and medium activity in 54% of the cell surface and zones of low or no activity in the remainder. The ‘non-active’ (‘silent’) zones covered 24% of the total and persisted for 6–8 min, indicating stable location. The 9-Ch NCD-UMEA therefore appears suitable for investigating the microdomain organization of neurosecretion with high spatial resolution

Two firing modes and well-resolved Na+, K+, and Ca2+ currents at the cell-microelectrode junction of spontaneously active rat chromaffin cell on MEAs

We recorded spontaneous extracellular action potentials (eAPs) from rat chromaffin cells (CCs) at 37 °C using microelectrode arrays (MEAs) and compared them with intracellularly recorded APs (iAPs) through conventional patch clamp recordings at 22 °C. We show the existence of two distinct firing modes on MEAs: a ~ 4 Hz irregular continuous firing and a frequent intermittent firing mode where periods of high-intraburst frequency (~ 8 Hz) of ~ 7 s duration are interrupted by silent periods of ~ 12 s. eAPs occurred either as negative- or positive-going signals depending on the contact between cell and microelectrode: either predominantly controlled by junction-membrane ion channels (negative-going) or capacitive/ohmic coupling (positive-going). Negative-going eAPs were found to represent the trajectory of the Na(+), Ca(2+), and K(+) currents passing through the cell area in tight contact with the microelectrode during an AP (point-contact junction). The inward Nav component of eAPs was blocked by TTX in a dose-dependent manner (IC(50) ~ 10 nM) while the outward component was strongly attenuated by the BK channel blocker paxilline (200 nM) or TEA (5 mM). The SK channel blocker apamin (200 nM) had no effect on eAPs. Inward Nav and Cav currents were well-resolved after block of Kv and BK channels or in cells showing no evident outward K(+) currents. Unexpectedly, on the same type of cells, we could also resolve inward L-type currents after adding nifedipine (3 μM). In conclusion, MEAs provide a direct way to record different firing modes of rat CCs and to estimate the Na(+), Ca(2+), and K(+) currents that sustain cell firing and spontaneous catecholamines secretion

Logistic regression of attitudes and coverage for influenza vaccination among Italian Public Health medical residents

Introduction. A few number of literature specifically addresses vaccination uptake among Public Health Residents (PHRs). Influ- enza vaccine attitudes and risk perceptions of PHRs across Italy were studied, contributing to literature on influenza vaccination uptake predictors, in particular among young physicians. Methods. An online survey was conducted in 25 Schools of Public Health in Italy in 2011-2012. Results were analysed using preva- lence and logistic regression methods. Results. A total of 365 Italian public health residents were included in the study. Vaccination uptake was confirmed by 22.2 and 33.2% of PHRs in 2010-2011 and 2011-2012, respectively. For the 2010-2011 influenza season, vaccination was associated with male sex (adj-OR 3.43; 95%CI = 1.5-7.84) and vaccination history (adj-OR 29.44; 95%CI = 6.4-135.04). For the 2011-2012 season, vaccination was significantly associated with having had between one and three influenza vaccinations in the previ- ous five years (adj-OR 11.56; 95%CI = 6.44-20.75) or more than three (adj-OR 136.43; 95%CI = 30.8-604.7) and with individual participation in general population vaccination campaigns (adj- OR 1.85; 95%CI = 1.01-3.41). Discussion. Italian residents in public health have no confidence and a low personal risk perception about vaccinations therefore taking no measures to protect patients, general population and themselves. Annual influenza vaccination acceptance is associ- ated with influenza vaccine uptake in the previous years and per- sonal involvement in general population vaccination campaigns. These factors should be considered for the design of future cam- paigns targeting public health residents