In Vitro Assessment of Developmental Neurotoxicity: Use of Microelectrode Arrays to Measure Functional Changes in Neuronal Network Ontogeny1

Because the Developmental Neurotoxicity Testing Guidelines require large numbers of animals and is expensive, development of in vitro approaches to screen chemicals for potential developmental neurotoxicity is a high priority. Many proposed approaches for screening are biochemical or morphological, and do not assess function of neuronal networks. In this study, microelectrode arrays (MEAs) were used to determine if chemical-induced changes in function could be detected by assessing the development of spontaneous network activity. MEAs record individual action potential spikes as well as groups of spikes (bursts) in neuronal networks, and activity can be assessed repeatedly over days in vitro (DIV). Primary cultures of rat cortical neurons were prepared on MEAs and spontaneous activity was assessed on DIV 2, 6, 9, 13, and 20 to determine the in vitro developmental profile of spontaneous spiking and bursting in cortical networks. In addition, 5 μM of the protein kinase C inhibitor bisindolylmaleamide-1 (Bis-1) was added to MEAs (n = 9–18) on DIV 5 to determine if changes in spontaneous activity could be detected in response to inhibition of neurite outgrowth. A clear profile of in vitro activity development occurred in control MEAs, with the number of active channels increasing from 0/MEA on DIV 2 to 37 ± 5/MEA by DIV 13; the rate of increase was most rapid between DIV 6 and 9, and activity declined by DIV 20. A similar pattern was observed for the number of bursting channels, as well as the total number of bursts. Bis-1 decreased the number of active channels/MEA and the number of bursting channels/MEA. Burst characteristics, such as burst duration and the number of spikes in a burst, were unchanged by Bis-1. These results demonstrate that MEAs can be used to assess the development of functional neuronal networks in vitro, as well as chemical-induced dysfunction

Characterization of Early Cortical Neural Network Development in Multiwell Microelectrode Array Plates.

We examined neural network ontogeny using microelectrode array (MEA) recordings made in multiwell MEA (mwMEA) plates over the first 12 days in vitro (DIV). In primary cortical cultures, action potential spiking activity developed rapidly between DIV 5 and 12. Spiking was sporadic and unorganized at early DIV, and became progressively more organized with time, with bursting parameters, synchrony, and network bursting increasing between DIV 5 and 12. We selected 12 features to describe network activity; principal components analysis using these features demonstrated segregation of data by age at both the well and plate levels. Using random forest classifiers and support vector machines, we demonstrated that four features (coefficient of variation [CV] of within-burst interspike interval, CV of interburst interval, network spike rate, and burst rate) could predict the age of each well recording with >65% accuracy. When restricting the classification to a binary decision, accuracy improved to as high as 95%. Further, we present a novel resampling approach to determine the number of wells needed for comparing different treatments. Overall, these results demonstrate that network development on mwMEA plates is similar to development in single-well MEAs. The increased throughput of mwMEAs will facilitate screening drugs, chemicals, or disease states for effects on neurodevelopment.EC was supported by a Wellcome Trust PhD studentship and NIHR Cambridge Biomedical Research Centre studentship. DH was supported by student services contract #EP-13-D-000108 and by a travelling fellowship from the Company of Biologists.This is the final version of the article. It first appeared from SAGE Publications via https://doi.org/10.1177/108705711664052

Minor versus major mergers: the stellar mass growth of massive galaxies from z=3 using number density selection techniques

We present a study on the stellar mass growth of the progenitors of local massive galaxies with a variety of number density selections with n≤1×10−4 Mpc−3 (corresponding to M*=1011.24 M⊙ at z=0.3) in the redshift range 0.3<z<3.0. We select the progenitors of massive galaxies using a constant number density selection, and one which is adjusted to account for major mergers. We find that the progenitors of massive galaxies grow by a factor of 4 in total stellar mass over this redshift range. On average the stellar mass added via the processes of star formation, major and minor mergers account for 24±8, 17±15 and 34±14per cent, respectively, of the total galaxy stellar mass at z=0.3. Therefore 51±20per cent of the total stellar mass in massive galaxies at z=0.3 is created externally to their z=3 progenitors. We explore the implication of these results on the cold gas accretion rate and size evolution of the progenitors of most massive galaxies over the same redshift range. We find an average gas accretion rate of∼66±32 M⊙ yr−1 over the redshift range of 1.5<z<3.0. We find that the size evolution of a galaxy sample selected this way is on average lower than the findings of other investigation

Transcriptional response of rat frontal cortex following acute In Vivo exposure to the pyrethroid insecticides permethrin and deltamethrin

<p>Abstract</p> <p>Background</p> <p>Pyrethroids are neurotoxic pesticides that interact with membrane bound ion channels in neurons and disrupt nerve function. The purpose of this study was to characterize and explore changes in gene expression that occur in the rat frontal cortex, an area of CNS affected by pyrethroids, following an acute low-dose exposure.</p> <p>Results</p> <p>Rats were acutely exposed to either deltamethrin (0.3 – 3 mg/kg) or permethrin (1 – 100 mg/kg) followed by collection of cortical tissue at 6 hours. The doses used range from those that cause minimal signs of intoxication at the behavioral level to doses well below apparent no effect levels in the whole animal. A statistical framework based on parallel linear (SAM) and isotonic regression (PIR) methods identified 95 and 53 probe sets as dose-responsive. The PIR analysis was most sensitive for detecting transcripts with changes in expression at the NOAEL dose. A sub-set of genes (<it>Camk1g</it>, <it>Ddc</it>, <it>Gpd3</it>, <it>c-fos </it>and <it>Egr1</it>) was then confirmed by qRT-PCR and examined in a time course study. Changes in mRNA levels were typically less than 3-fold in magnitude across all components of the study. The responses observed are consistent with pyrethroids producing increased neuronal excitation in the cortex following a low-dose <it>in vivo </it>exposure. In addition, Significance Analysis of Function and Expression (SAFE) identified significantly enriched gene categories common for both pyrethroids, including some relating to branching morphogenesis. Exposure of primary cortical cell cultures to both compounds resulted in an increase (~25%) in the number of neurite branch points, supporting the results of the SAFE analysis.</p> <p>Conclusion</p> <p>In the present study, pyrethroids induced changes in gene expression in the frontal cortex near the threshold for decreases in ambulatory motor activity <it>in vivo</it>. The penalized regression methods performed similarly in detecting dose-dependent changes in gene transcription. Finally, SAFE analysis of gene expression data identified branching morphogenesis as a biological process sensitive to pyrethroids and subsequent <it>in vitro </it>experiments confirmed this predicted effect. The novel findings regarding pyrethroid effects on branching morphogenesis indicate these compounds may act as developmental neurotoxicants that affect normal neuronal morphology.</p

The evolution of galaxies at constant number density: a less biased view of star formation, quenching, and structural formation

Due to significant galaxy contamination and impurity in stellar mass selected samples (up to 95 per cent from z = 0–3), we examine the star formation history, quenching time-scales, and structural evolution of galaxies using a constant number density selection with data from the United Kingdom Infra-Red Deep Sky Survey Ultra-Deep Survey field. Using this methodology, we investigate the evolution of galaxies at a variety of number densities from z= 0–3. We find that samples chosen at number densities ranging from 3 × 10−4 to 10−5 galaxies Mpc−3 (corresponding to z ∼ 0.5 stellar masses of M∗ = 1010.95−11.6 M0) have a star-forming blue fraction of ∼50 per cent at z ∼ 2.5, which evolves to a nearly 100 per cent quenched red and dead population by z ∼ 1. We also see evidence for number density downsizing, such that the galaxies selected at the lowest densities (highest masses) become a homogeneous red population before those at higher number densities. Examining the evolution of the colours for these systems furthermore shows that the formation redshift of galaxies selected at these number densities is zform > 3. The structural evolution through size and S´ersic index fits reveal that while there remains evolution in terms of galaxies becoming larger and more concentrated in stellar mass at lower redshifts, the magnitude of the change is significantly smaller than for a mass-selected sample. We also find that changes in size and structure continues at z < 1, and is coupled strongly to passivity evolution.We conclude that galaxy structure is driving the quenching of galaxies, such that galaxies become concentrated before they become passive

The Spitzer c2d Survey of Large, Nearby, Interstellar Clouds. VIII. Serpens Observed with MIPS

We present maps of 1.5 deg^2 of the Serpens dark cloud at 24, 70, and 160 μm observed with the Spitzer Space Telescope MIPS camera. We describe the observations and briefly discuss the data processing carried out by the c2d team on these data. More than 2400 compact sources have been extracted at 24 μm, nearly 100 at 70 μm, and four at 160 μm. We estimate completeness limits for our 24 μm survey from Monte Carlo tests with artificial sources inserted into the Spitzer maps. We compare source counts, colors, and magnitudes in the Serpens cloud to two reference data sets: a 0.50 deg^2 set on a low-extinction region near the dark cloud, and a 5.3 deg^2 subset of the SWIRE ELAIS N1 data that was processed through our pipeline. These results show that there is an easily identifiable population of young stellar object candidates in the Serpens cloud that is not present in either of the reference data sets. We also show a comparison of visual extinction and cool dust emission illustrating a close correlation between the two and find that the most embedded YSO candidates are located in the areas of highest visual extinction

The Spitzer c2d Survey of Weak-line T Tauri Stars II: New Constraints on the Timescale for Planet Building

One of the central goals of the Spitzer Legacy Project ``From Molecular Cores to Planet-forming Disks'' (c2d) is to determine the frequency of remnant circumstellar disks around weak-line T Tauri stars (wTTs) and to study the properties and evolutionary status of these disks. Here we present a census of disks for a sample of over 230 spectroscopically identified wTTs located in the c2d IRAC (3.6, 4.5, 4.8, and 8.0 um) and MIPS (24 um) maps of the Ophiuchus, Lupus, and Perseus Molecular Clouds. We find that ~20% of the wTTs in a magnitude limited subsample have noticeable IR-excesses at IRAC wavelengths indicating the presence of a circumstellar disk. The disk frequencies we find in these 3 regions are ~3-6 times larger than that recently found for a sample of 83 relatively isolated wTTs located, for the most part, outside the highest extinction regions covered by the c2d IRAC and MIPS maps. The disk fractions we find are more consistent with those obtained in recent Spitzer studies of wTTs in young clusters such as IC 348 and Tr 37. From their location in the H-R diagram, we find that, in our sample, the wTTs with excesses are among the younger part of the age distribution. Still, up to ~50% of the apparently youngest stars in the sample show no evidence of IR excess, suggesting that the circumstellar disks of a sizable fraction of pre-main-sequence stars dissipate in a timescale of ~1 Myr. We also find that none of the stars in our sample apparently older than ~10 Myrs have detectable circumstellar disks at wavelengths < 24 um. Also, we find that the wTTs disks in our sample exhibit a wide range of properties (SED morphology, inner radius, L_DISK/L*, etc) which bridge the gaps observed between the cTTs and the debris disk regimes.Comment: 54 pages, 13 figures, Accepted by Ap

The Spitzer c2d Survey of Large, Nearby, Interstellar Clouds. VII. Ophiuchus Observed with MIPS

We present maps of 14.4 deg^2 of the Ophiuchus dark clouds observed by the Spitzer Space Telescope Multiband Imaging Photometer for Spitzer (MIPS). These high-quality maps depict both numerous point sources and extended dust emission within the star-forming and non–star-forming portions of these clouds. Using PSF-fitting photometry, we detect 5779 sources at 24 μm and 81 sources at 70 μm at the 10 σ level of significance. Three hundred twenty-three candidate young stellar objects (YSOs) were identified according to their positions on the MIPS/2MASS K versus color-magnitude diagrams, as compared to 24 μm detections in the SWIRE extragalactic survey. We find that more than half of the YSO candidates, and almost all those with protostellar Class I spectral energy distributions, are confined to the known cluster and aggregates

The Spitzer c2d Survey of Large, Nearby, Interstellar Clouds. IV. Lupus Observed with MIPS

We present maps of 7.78 square degrees of the Lupus molecular cloud complex at 24, 70, and $160\:\mu$m. They were made with the Spitzer Space Telescope's Multiband Imaging Photometer for Spitzer (MIPS) instrument as part of the Spitzer Legacy Program, ``From Molecular Cores to Planet-Forming Disks'' (c2d). The maps cover three separate regions in Lupus, denoted I, III, and IV. We discuss the c2d pipeline and how our data processing differs from it. We compare source counts in the three regions with two other data sets and predicted star counts from the Wainscoat model. This comparison shows the contribution from background galaxies in Lupus I. We also create two color magnitude diagrams using the 2MASS and MIPS data. From these results, we can identify background galaxies and distinguish them from probable young stellar objects. The sources in our catalogs are classified based on their spectral energy distribution (SED) from 2MASS and Spitzer wavelengths to create a sample of young stellar object candidates. From 2MASS data, we create extinction maps for each region and note a strong corresponence between the extinction and the $160\:\mu$m emission. The masses we derived in each Lupus cloud from our extinction maps are compared to masses estimated from $^{13}$CO and C$^{18}$O and found to be similar to our extinction masses in some regions, but significantly different in others. Finally, based on our color-magnitude diagrams, we selected 12 of our reddest candidate young stellar objects for individual discussion. Five of the 12 appear to be newly-discovered YSOs.Comment: 15 pages, 17 figures, uses emulateapj.cls. Accepted for publication in ApJ. A version with high-quality figures can be found at http://peggysue.as.utexas.edu/SIRTF