Mcnallyrosario1501

Cyclosporine and systemic corticosteroids, but not antihistamines or omalizumab, are used less during the pandemic. CU does not affect the course of COVID-19, but COVID-19 results in CU exacerbation in one of three patients, with higher rates in patients with severe COVID-19.

The COVID-19 pandemic brings major changes and challenges for CU patients and their physicians. The long-term consequences of these changes, especially the increased use of remote consultations, require careful evaluation.

The COVID-19 pandemic brings major changes and challenges for CU patients and their physicians. The long-term consequences of these changes, especially the increased use of remote consultations, require careful evaluation.Extensive efforts have been undertaken in the northern Gulf of Mexico to restore coastal wetlands that have been lost rapidly. The evaluation of these restorations mostly focused on individual-project scales. A modeling framework that can coherently synthesize multi-scale monitoring data and account for various uncertainties would improve quantitative evaluations at broader spatial scales needed for regional decision-making. We aim to develop such a framework to investigate the impact of different restoration methods (hydrological alteration, breakwater infrastructure, vegetative planting, or marsh creation using dredged materials) on wetland loss on the outermost mainland coastlines in Louisiana. We did this by implementing multi-level Bayesian models to predict areal wetland loss (1996-2005 before Hurricane Katrina) as a function of local geophysical variables (relative sea-level rise, wave height, tidal range) and a dummy variable indicating presence/absence of restoration. We assumed the effects of these variables varied by broader watershed scales. The restoration's effect also depended on temporal scales of implementation. The results indicate the sites with hydrological alteration, when implemented for longer than 7 years, had significantly smaller areal wetland loss, compared to the reference sites controlled for the local geophysical variables, in the Chenier Plain watershed, but not in the lower Mississippi River watershed. The effects of the other restoration methods on wetland loss were not significant based on limited numbers of sites. The Bayesian modeling framework we developed can integrate monitoring data/key drivers across projects with uncertainties accounted for, it is adaptable, and presents a useful tool in restoration evaluations spatially and temporally.A fundamental feature of circadian clock neurons across species is that they express circadian rhythms in spontaneous spike frequency. Spike frequency rhythms serve as both output timing signals of clock neurons as well as resonant elements of rhythms generation. Importantly, optogenetics, as applied to clock neurons, can enable investigation of the roles of clock neuron electrical activity in circadian timing. Here we describe protocols for using both in vitro and in vivo optogenetics directed to mammalian clock neurons in the suprachiasmatic nucleus to study circadian physiology and behavior. Optogenetic stimulation via channelrhodopsin, or inhibition via halorhodopsin, allows for the precise manipulation of neuronal firing rates across the SCN, and within specific neuronal subpopulations thereof, and can be combined with actigraphy and gene expression analysis.In mammals, the part of the nervous system responsible for most circadian behavior can be localized to a bilaterally paired structure in the hypothalamus known as the suprachiasmatic nucleus (SCN). Understanding the mammalian circadian system will require a detailed multilevel analysis of neural SCN circuits ex vivo and in vivo. Many of the techniques and approaches that are used for the analysis of the circuitry driving circadian oscillations in the SCN are similar to those employed in other brain regions. There is, however, one fundamental difference that needs to be taken into consideration, that is, the physiological, cell, and molecular properties of SCN neurons vary with the time of day. find more In this chapter, we will consider the preparations and electrophysiological techniques that we have used to analyze the SCN circuit focusing on the acute brain slice and intact, freely moving animal.Advances in imaging technology, combined with the development of bioluminescent reporters for core clock genes, has enabled the observation of spatiotemporal patterns of circadian rhythms in the suprachiasmatic nuclei (SCN). In particular, the PERIOD2luciferase (PER2LUC) knockin mouse has led to novel approaches for studying the heterogeneous circadian network in the SCN. This chapter describes how to automate the processing of PER2LUC imaging data from SCN slices for generating spatiotemporal maps of circadian parameters like phase, period, and amplitude. These maps can be aligned and averaged to produce composite maps displaying common features across multiple slices. In addition, we describe a method for automated detection of cell-like regions of interest, to support the study of the neural network in the SCN.Circadian rhythms in cellular function can be monitored in real time with bioluminescence imaging. In this approach, bioluminescence is produced by an enzymatic reaction, which can be used to report dynamic changes in gene or protein expression in living cells. Bioluminescence imaging in circadian experiments typically uses an ex vivo slice preparation, with the most commonly studied structure being the master clock in the suprachiasmatic nucleus (SCN) of the anterior hypothalamus. Here we describe procedures for dissecting and collecting SCN slices for bioluminescence imaging experiments.The ability to record ensemble action potential (AP) discharge frequencies from large populations of neurons over extended periods of time in vitro offers clear advantages in neuroscience and circadian biology research. Here, we provide an overview of a step-by-step method to perform multisite extracellular AP activity recordings in suprachiasmatic and mediobasal hypothalamic nuclei brain slices, using a state-of-the-art perforated multielectrode array system. Further, we describe in detail a setup architecture which systematically delivers stable, high-quality recordings with excellent anatomical accuracy and consistency. We also provide some procedural, technical, and methodological troubleshooting notes and examples of good quality recordings.The discovery of the suprachiasmatic nucleus (SCN) as the master mammalian pacemaker has since opened up a variety of alternative methods for assessing how external timing cues influence the clock. One powerful approach for understanding how sensory inputs influence the SCN is to monitor acute changes in SCN electrophysiological activity via in vivo extracellular recording. This methodology offers the ability to monitor stimulus-evoked changes in SCN function at very fine timescales and to rapidly test multiple stimuli and/or stimulus repeats within a single animal. In this chapter we describe our methods for acute in vivo multielectrode recording in head-fixed, anesthetized rodents. These allow for monitoring of single-cell/population activity for >12 h; enable the delivery of carefully controlled sensory stimuli; can be used alongside an array of established or novel experimental tools; and are easily adapted to study activity in any other brain region.Circadian rhythms are 24-h cycles in physiology and behavior that occur in virtually all organisms. These processes are not simply driven by changes in the external environment as they persist under constant conditions, providing evidence for an internal biological clock. In mammals, this clock is located in the hypothalamic suprachiasmatic nuclei (SCN) and is based upon an intracellular mechanism composed of a transcriptional-translational feedback loop composed of a number of core clock genes. However, a clock is of no use unless it can be set to the correct time. The primary time cue for the molecular clock in the SCN is light detected by the eye. The photoreceptors involved in this process include the rods and cones that mediate vision, as well as the recently identified melanopsin-expressing photosensitive retinal ganglion cells (pRGCs). Light information is conveyed to the SCN via the retinohypothalamic tract, resulting in an intracellular signaling cascade which converges on cAMP-response elements in the promoters of several key clock genes. Over the last two decades a number of studies have investigated the transcriptional response of the SCN to light stimuli with the aim of further understanding these molecular signaling pathways. Here we provide an overview of these studies and provide protocols for studying the molecular responses to light in the SCN clock.Drosophila melanogaster is a powerful model organism used to study circadian rhythms, historically for elucidating the molecular basis of the clock and, more recently, for allowing for dissection of neural circuits underlying rhythmic behavior. The fly can be used to investigate the neuronal basis of complex behaviors at single-neuron resolution. Patch clamp electrophysiology permits single-neuron recording of resting membrane potential and action potential firing in response to genetic or environmental manipulations or application of drugs and neurotransmitters. Here we describe a protocol for dissecting Drosophila brains for electrophysiology, setting up and using a patch clamp system, and analyzing firing data around the circadian day and in stimulation-response experiments to test for functional neuronal connectivity in circadian circuits.Live imaging of the molecular clockwork within the circadian pacemaker neurons offers the unique possibility to study complex interactions between the molecular clock and neuronal communication within individual neurons and throughout the entire circadian circuitry. Here we describe how to establish brain explants and dissociated neuron culture from Drosophila larvae, guidelines for time-lapse fluorescence microscopy, and the method of image analysis. This approach enables the long-term monitoring of fluorescence signals of circadian reporters at single-cell resolution and can be also applicable to analyze real-time expression of other fluorescent probes in Drosophila neurons.Daily rhythms of behaviors and physiologies are driven by transcriptional-translational negative feedback loops of clock genes and encoded clock proteins (Bass and Takahashi Science 3301349-1354, 2010; Brown et al. Dev Cell 22477-487, 2012). Posttranslational modifications of clock proteins, including protein phosphorylation, play an essential role for normal oscillation of the circadian clock through regulation of their activities, stabilities, interactions, and intracellular localization (Gallego and Virshup Nat Rev Mol Cell Biol 8139-148, 2007; Hirano et al. Nat Struct Mol Biol 231053-1060, 2016). In this chapter, we describe detailed methods for quantitative analysis of phosphorylation levels of clock proteins, particularly focusing on circadian phosphorylation of CLOCK, BMAL1, and their complex (Yoshitane et al. Mol Cell Biol 293675-3686, 2009).Recent advances in mass spectrometry (MS)-based quantitative proteomics now allow the identification and quantification of deep proteomes and post-translational modifications (PTMs) in relatively short times. Therefore, in the last few years, this technology has proven successful in the circadian field to characterize temporal oscillations of the proteome and more recently PTMs in cellular systems and in tissues. In this chapter, we describe a robust and simple protocol, based on the EasyPhos workflow, to enable preparation of large number of proteomes and phosphoproteomes from mouse tissues for MS-based quantitative analysis. We additionally discuss computational methods to analyze proteome and phosphoproteome time series to determine circadian oscillations.