Detection of silent cells,synchronization and modulatory activity in developing cellular networks

Developing networks in the immature nervous system and in cellular cultures are characterized by waves of synchronous activity in restricted clusters of cells. Synchronized activity in immature networks is proposed to regulate many different developmental processes, from neuron growth and cell migration, to the refinement of synapses, topographic maps, and the mature composition of ion channels. These emergent activity patterns are not present in all cells simultaneously within the network and more immature “silent” cells, potentially correlated with the presence of silent synapses, are prominent in different networks during early developmental periods. Many current network analyses for detection of synchronous cellular activity utilize activity‐based pixel correlations to identify cellular‐based regions of interest (ROIs) and coincident cell activity. However, using activity‐based correlations, these methods first underestimate or ignore the inactive silent cells within the developing network and second, are difficult to apply within cell‐dense regions commonly found in developing brain networks. In addition, previous methods may ignore ROIs within a network that shows transient activity patterns comprising both inactive and active periods. We developed analysis software to semi‐automatically detect cells within developing neuronal networks that were imaged using calcium‐sensitive reporter dyes. Using an iterative threshold, modulation of activity was tracked within individual cells across the network. The distribution pattern of both inactive and active, including synchronous cells, could be determined based on distance measures to neighboring cells and according to different anatomical layers. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 357–374, 2016     

Effectiveness of Partial Sedation to Reduce Stress in Captured Mule Deer

Information garnered from the capture and handling of free-ranging animals helps advance understanding of wildlife ecology and can aid in decisions on wildlife management. Unfortunately, animals may experience increased levels of stress, injuries, and death resulting from captures (e.g., exertional myopathy, trauma). Partial sedation is a technique proposed to alleviate stress in animals during capture, yet efficacy of partial sedation for reducing stress and promoting survival post-capture remains unclear. We evaluated the effects of partial sedation on physiological, biochemical, and behavioral indicators of acute stress and probability of survival post-capture for mule deer (Odocoileus hemionus) that were captured via helicopter net-gunning in the eastern Greater Yellowstone Ecosystem, Wyoming, USA. We administered 10–30 mg of midazolam and 15 mg of azaperone intramuscularly (IM) to 32 mule deer in 2016 and 53 mule deer in 2017, and maintained a control group (captured but not sedated) of 38 mule deer in 2016 and 54 mule deer in 2017. To evaluate indicators of acute stress, we measured heart rate, blood-oxygen saturation, body temperature, respiration rate, and levels of serum cortisol. We recorded number of kicks and vocalizations of deer during handling and evaluated behavior during release. We also measured levels of fecal glucocorticoids as an indicator of baseline stress. Midazolam and azaperone did not reduce physiological, biochemical, or behavioral indicators of acute stress or influence probability of survival post-capture. Mule deer that were administered midazolam and azaperone, however, were more likely to hesitate, stumble or fall, and walk during release compared with individuals in the control group, which were more likely to trot, stot, or run without stumbling or falling. Our findings suggest that midazolam (10–30 mg IM) and azaperone (15 mg IM) may not yield physiological or demographic benefits for captured mule deer as previously assumed and may pose adverse effects that can complicate safety for captured animals, including drug-induced lethargy. Although we failed to find efficacy of midazolam and azaperone as a method for reducing stress in captured mule deer, the efficacy of midazolam and azaperone or other combinations of partial sedatives in reducing stress may depend on the dose of tranquilizer, study animal, capture setting, and how stress is defined. © 2020 The Wildlife Society.     

Shifts in fine root traits within and among species along a fine-scale hydrological gradient

Background and AimsLessons from above-ground trait ecology and resource economics theory may not be directly translatable to below-ground traits due to differences in function, trade-offs and environmental constraints. Here we examine root functional traits within and across species along a fine-scale hydrological gradient. We ask two related questions: (1) What is the relative magnitude of trait variation across the gradient for within- versus among-species variation? (2) Do correlations among below-ground plant traits conform with predictions from resource-economic spectrum theory?MethodsWe sampled four below-ground fine-root traits (specific root length, branching intensity, root tissue density and root dry matter content) and four above-ground traits (specific leaf area, leaf size, plant height and leaf dry matter content) in vascular plants along a fine-scale hydrological gradient within a wet heathland community in south-eastern Australia. Below-ground and above-ground traits were sampled both within and among species.Key ResultsRoot traits shifted both within and among species across the hydrological gradient. Within- and among-species patterns for root tissue density showed similar declines towards the wetter end of the gradient. Other root traits showed a variety of patterns with respect to within- and among-species variation. Filtering of species has a stronger effect compared with the average within-species shift: the slopes of the relationships between soil moisture and traits were steeper across species than slopes of within species. Between species, below-ground traits were only weakly linked to each other and to above-ground traits, but these weak links did in some cases correspond with predictions from economic theory.ConclusionsOne of the challenges of research on root traits has been considerable intraspecific variation. Here we show that part of intraspecific root trait variation is structured by a fine-scale hydrological gradient, and that the variation aligns with among-species trends in some cases. Patterns in root tissue density are especially intriguing and may play an important role in species and individual response to moisture conditions. Given the importance of roots in the uptake of resources, and in carbon and nutrient turnover, it is vital that we establish patterns of root trait variation across environmental gradients.     

CGGBP1 phosphorylation constitutes a telomere-protection signal

The shelterin proteins are required for telomere integrity. Shelterin dysfunction can lead to initiation of unwarranted DNA damage and repair pathways at chromosomal termini. Interestingly, many shelterin accessory proteins are involved in DNA damage signaling and repair. We demonstrate here that in normal human fibroblasts, telomeric ends are protected by phosphorylation of CGG triplet repeat-binding protein 1 (CGGBP1) at serine 164 (S164). We show that serine 164 is a major phosphorylation site on CGGBP1 with important functions. We provide evidence that one of the kinases that can phosphorylate S164 CGGBP1 is ATR. Overexpression of S164A phospho-deficient CGGBP1 exerted a dominant-negative effect, causing telomeric dysfunction, accelerated telomere shortening, enhanced fusion of telomeres, and crisis. However, overexpression of wild-type or phospho-mimicking S164E CGGBP1 did not cause these effects. This telomere damage was associated with reduced binding of the shelterin protein POT1 to telomeric DNA. Our results suggest that CGGBP1 phosphorylation at S164 is a novel telomere protection signal, which can affect telomere-protective function of the shelterin complex.     

Altered GABAA Receptor Expression and Seizure Threshold Following Acute Ethanol Challenge in Mice Lacking the RIIβ Subunit of PKA

Ethanol causes pathological changes in GABA_A receptor trafficking and function. These changes are mediated in part by ethanol activation of protein kinase A (PKA). The current study investigated the expression of the GABA_A α1 and α4 subunits and the kinase anchoring protein AKAP150, as well as bicuculline-induced seizure threshold, at baseline and following acute injection of ethanol (3.5 g/kg IP) in a mouse line lacking the regulatory RIIβ subunit of PKA. Whole cerebral cortices were harvested at baseline, 1 h, or 46 h following injection of ethanol or saline and subjected to fractionation and western blot analysis. Knockout (RIIβ?/?) mice had similar baseline levels of PKA RIIα and GABA_A α1 and α4 subunits compared to wild type (RIIβ+/+) littermates, but had deficits in AKAP150. GABA_A α1 subunit levels were decreased in the P2 fraction of RIIβ?/?, but not RIIβ+/+, mice following 1 h ethanol, an effect that was driven by decreased α1 expression in the synaptic fraction. GABA_A α4 subunits in the P2 fraction were not affected by 1 h ethanol; however, synaptic α4 subunit expression was increased in RIIβ+/+, but not RIIβ?/? mice, while extrasynaptic α4 and δ subunit expression were decreased in RIIβ?/?, but not RIIβ+/+ mice. Finally, RIIβ knockout was protective against bicuculline-induced seizure susceptibility. Overall, the results suggest that PKA has differential roles in regulating GABA_A receptor subunits. PKA may protect against ethanol-induced deficits in synaptic α1 and extrasynaptic α4 receptors, but may facilitate the increase of synaptic α4 receptors.     

Confidently identifying the correct K value using the ΔK method: When does K = 2?

Populations delineated based on genetic data are commonly used for wildlife conservation and management. Many studies use the program structure combined with the ΔK method to identify the most probable number of populations (K). We recently found K = 2 was identified more often when studies used ΔK compared to studies that did not. We suggested two reasons for this: hierarchical population structure leads to underestimation, or the ΔK method does not evaluate K = 1 causing an overestimation. The present contribution aims to develop a better understanding of the limits of the method using one, two and three population simulations across migration scenarios. From these simulations we identified the “best K” using model likelihood and ΔK. Our findings show that mean probability plots and ΔK are unable to resolve the correct number of populations once migration rate exceeds 0.005. We also found a strong bias towards selecting K = 2 using the ΔK method. We used these data to identify the range of values where the ΔK statistic identifies a value of K that is not well supported. Finally, using the simulations and a review of empirical data, we found that the magnitude of ΔK corresponds to the level of divergence between populations. Based on our findings, we suggest researchers should use the ΔK method cautiously; they need to report all relevant data, including the magnitude of ΔK, and an estimate of connectivity for the research community to assess whether meaningful genetic structure exists within the context of management and conservation.