Once microdissection is completed, the tissue chunks were warmed to 34°C for 10 min before trituration. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. What company does Daniel Moreira work for? Un but de Daniel Moreira donne la Coupe de la Ligue à Lens.Lens offensif offense Metz", http://www.eurosport.fr/football/matches-amicaux/2015/avant-portugal-france-ces-bleus-qui-ont-la-selecao-dans-le-coeur_sto4894055/story.shtml, https://en.wikipedia.org/w/index.php?title=Daniel_Moreira&oldid=977230185, Sportspeople from Nord (French department), Short description is different from Wikidata, Creative Commons Attribution-ShareAlike License, * Senior club appearances and goals counted for the domestic league only, This page was last edited on 7 September 2020, at 17:41. However, further work is needed to understand whether and how differential CO2/H+ vascular reactivity in these chemoreceptor regions contributes to breathing problems in disease states. Moreira was born in France, and is of Portuguese descent. The investigators significantly advance their previous work by applying a number of new experimental approaches to provide evidence that P2Y2 receptors in RTN vascular smooth muscle cells are responsible for the purinergic mechanism mediating the vascular reactivity and specifically contribute to RTN chemosensitivity. One symbol = p < 0.05, two symbols = p < 0.01, three symbols = p < 0.001, four symbols = p < 0.0001. United States, The University of Illinois at Chicago is a major research university located in the heart of one of the world's greatest cities. P2ry2 transcript was not detected (n.d.) in smooth muscle cells from P2Y2 cKO mice (N = 3 runs/9 animals). Slices were then incubated in a 0.5% Triton-X/PBS solution for 45 min to permeabilize the tissue. communities in the world, Get the week's mostpopular data scienceresearch in your inbox -every Saturday, Towards a Mathematical Understanding of Neural Network-Based Machine The cellular and transmitter basis for CO2/H+-induced constriction of RTN arterioles appears to involve CO2/H+-evoked ATP release from local astrocytes. ∙ We have added an expanded Discussion section that includes experimental limitations section. Data are reported as mean ± SE. *, Different from 0% CO2 in condition as assessed by Tukey’s post-hoc multiple comparison test. The P2 transcript profile of vascular cells in the RTN was fairly similar to the ROb (Figure 1A–B) but differed from the cNTS in several ways including lower expression of P2ry6 in endothelial cells (Figure 1A) and higher expression of P2rx1, P2rx4, and P2ry14 in smooth muscle cells (Figure 1B). After the anterior neck muscles were removed, a basio-occipital craniotomy exposed the ventral medullary surface, and the dura was resected. A single-cell suspension was achieved by trituration using a 25G and 30G needle sequentially, attached to a 3 mL syringe. Assessment of scientific research integrity through the detection of image tampering in scientific papers. This is a direction we plan to pursue in the future. This is interesting because in other brain regions activation of endothelial P2Y2 receptors favors nitric-oxide-induced vasodilation (Marrelli, 2001), whereas activation of this same receptor in vascular smooth muscle cells mediates vasoconstriction (Brayden et al., 2013; Lewis et al., 2000). ∙ If the mechanism underlying vascular control of RTN chemoreception involves maintenance of tissue CO2/H+ levels, then by this same logic, vasodilation in the cNTS and ROb should buffer tissue CO2/H+ and limit contributions of these regions to respiratory output. The left carotid artery was exposed and quickly cannulated to allow for arterial blood to be collected and analyzed by the blood gas analyzer; no more than 5 s was spent between blood collection and analysis on the blood gas analyzer. Consistent with the possibly that smooth muscle P2Y2 receptor-mediated vasoconstriction in the RTN augments the ventilatory response to CO2, we found that P2Y2 cKO mice exhibit normal respiratory activity under room air conditions (Table 1) but pronounced hypoventilation during exposure to graded increases in CO2 (balance O2 to minimize input from peripheral chemoreceptors). Animal use was in accordance with guidelines approved by the University of São Paulo Animal Care and Use Committee. The authors thank Drs. For example, myogenic tone increases with increased perfusion pressure and relaxes with decreased pressure. Gapdh showed fairly consistent and repeatable Ct values across all regions of interest, and so was used an internal control for results shown in Figures 1A-B and 4A. The second injection was made 1–2 min later at the same level on the contralateral side. Our discovery that CO2/H+-dependent regulation of vascular tone in the RTN is the opposite to the rest of the cerebral vascular tree is novel and fundamentally important for understanding how regulation of vascular tone is tailored to support neural function and behavior, in this case the drive to breathe. Which industry does Daniel Moreira work in? Gustavo Daniel tiene 7 empleos en su perfil. Indeed, this is thought to be the cause of unstable breathing in patients with global deficits in CO2/H+ vascular reactivity due to congestive heart failure or cerebrovascular disease (Cherniack et al., 2005; Cherniack and Longobardo, 2006). The right cheek of the animal was shaved, and an incision was made to expose the right marginal mandibular branch of the facial nerve. We found that injections of U46619 (1 μM) into the cNTS alone caused an increase in breathing frequency but together with the ROb resulted in unstable breathing as evidenced by frequent bouts of hyperventilation followed by apnea (Figure 3A) (1.2 ± 0.6, vs. saline: 0.4 ± 0.2 apneas/min; p<0.001) and increased breath to breath variability (Figure 3C–E). However, the relationship between protein synthesis and presynaptic structural plasticity remains unclear. After enzyme incubation, slices were washed three times in cold dissociation solution and then transferred to an enzyme inhibitor mix containing trypsin inhibitor (10 mg/mL, Sigma), bovine serum albumin (BSA, 10 mg/mL, Sigma), and sodium nitroprusside (SNP, 2 mg/mL, ThermoFisher) in cold sucrose dissociation solution. 3) Additional information on measurements of vascular diameters would be useful. Additional funds were also provided by the American Heart Association (17SDG33670237 and 19IPLOI34660108 to TAL), the São Paulo Research Foundation (FAPESP; grants: 2019/01236-4 to ACT; 2015/23376-1 to TSM) and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq; grant: 408647/2018–3 to ACT). It is important to note that this work was performed using isoflurane-anesthetized mice which require 2-3% CO2 in order to maintain respiratory activity (this level of CO2 is defined as the apneic threshold). We are also offering, if you choose, to post the manuscript to bioRxiv (if it is not already there) along with this decision letter and a formal designation that the manuscript is "in revision at eLife". These results identify P2Y2 receptors in RTN smooth muscle cells as a novel vascular element of respiratory chemoreception. We also provide evidence that, similar to brain neural stem cells, the activation and neuronal differentiation of enteric glia are regulated by Notch signalling. Here, we show that CO2/H+ vascular reactivity in the RTN is unique compared to other chemoreceptor regions. Moreover, while CB1-iLTD increased ubiquitin/proteasome activity, ubiquitination but not proteasomal degradation was critical for structural and functional presynaptic CB1-iLTD. *,difference from 0% CO2; #, differences between genotypes under an experimental condition (two-way ANOVA with Tukey's multiple comparison test). As previously described (Hawkins et al., 2017), we assessed arteriole viability at the end of each experiment by inducing a large vasoconstriction with a 60 mM K+ solution and then a large vasodilation with a Ca+2-free solution containing EGTA (5 mM), a phosphodiesterase inhibitor (papaverine, 200 mM), and an L-type Ca+2 channel blocker (diltiazem, 50 mM). Baseline parameters were allowed to stabilize for 30 min prior to recording. (Di-Dii) The CO2/H+ response of RTN vessels was blocked by a selective P2Y2 receptor antagonist (AR-C118925; 10 μM) #, difference from baseline (one sample t-test). communities, © 2019 Deep AI, Inc. | San Francisco Bay Area | All rights reserved. This approach is commonly used to study vascular regulation in the brain slice preparation (PMID: 18456724). share, Prior art has shown it is possible to estimate, through image processing... Daniel Moreira works in the industry of Education, Colleges & Universities. Also, mechanisms underlying this specialized CO2/H+ regulation of vascular tone in the RTN are largely unknown. ∙ All three technical replicates were averaged to create one raw Ct values per Taqman assay. ... Aug 2020 – Present 1 month. ∙ ∙ Évoluant au poste de milieu offensif puis d'attaquant, il est professionnel de 1996 à décembre 2010. The baseline activity was normalized to 100%, and the percent change was used to compare the magnitude of increases or decreases across muscle from those physiological baselines. This suggests that CO2/H+-dependent regulation of vascular tone and roles of astrocytes in this process are fundamentally different in the RTN compared to other chemoreceptor regions. 01/17/2020 ∙ by William Theisen, et al. To control for RNA contamination in FACS droplets, allophycocyanin (APC) beads were sorted based on green fluorescence and were treated alongside experimental samples (data not shown). Transverse brainstem slices (400 μm thick) were prepared and then immediately enzymatically treated at 34°C with an initial incubation in a papain (6 mg/mL, Sigma) and dithioerythritol (10 mg/mL, ThermoFisher) mixture for 30 min in sucrose dissociation solution containing (in mM): 185 sucrose, 10 glucose, 30 Na2SO4, 2 K2SO4, 10 HEPES, 0.5 CaCl2, 6 MgCl2, 5 L-ascorbic acid, pH 7.4, 320 mOsm, followed by a collagenase IV (10 mg/mL, ThermoFisher) enzyme treatment for 6 min. ∙ We have added a new supplementary figure (Figure 4—figure supplement 4) that provides several examples of viral transduced smooth muscle cells (eGFP+ and Acta2+). Rats were placed supine onto a stereotaxic apparatus (Type 1760; Harvard Apparatus) on a heating pad and core body temperature was maintained at a minimum of 36.5°C via a thermocouple. Individual brain slices containing either the caudal NTS (cNTS), raphe obscurus (ROb), RTN, or somatosensory cortex were transferred to a Plexiglas recording chamber mounted on a fixed-stage upright fluorescent microscope (Zeiss Axioskop FS) and perfused with 37°C aCSF bubbled with a 5% CO2-21% O2 (balance N2). TaglnCre and Myh11Cre/eGFP mice were crossed with P2ry2 floxed mice (gifted by Dr. Cheike Seye at Indiana University) to generate double floxed smooth muscle-specific P2Y2 cKO mice for in vitro and in vivo experimentation. This level of isoflurane minimally affects blood gases (Constantinides et al., 2011; Loeven et al., 2018). ∙ Mice were placed on a heated pad until consciousness was regained. Join Facebook to connect with Daniel Moreira and others you may know. For example, previous work showed that (i) CO2/H+ caused discrete release of ATP near the RTN region (Gourine et al., 2005); (ii) astrocytes in the RTN show Ca2+ responses to mild acidification (Gourine et al., 2010); and (iii) activation of astrocytes with tACPD, an mGluR agonist widely used to elicit Ca2+ elevations in astrocytes (Howarth et al., 2017), constricted RTN arterioles by a purinergic-dependent mechanism (Hawkins et al., 2017; Figure 2B), whereas application of an ATP receptor blocker to the ventral surface blunted CO2/H+-induced vasoconstriction and the ventilatory response to CO2 (18). Three days before the metabolic experiment, each animal was placed in the CLAMS housing cage with metered water and waste collection. Crowthorne. Meloxicam was administered 24 and 48 hr postoperatively. Our evidence also suggests that CO2/H+ constriction in the RTN enhances chemoreceptor activity and respiratory output, while simultaneous dilation in other chemoreceptor regions may serve to limit chemoreceptor activity and stabilize breathing. En septembre 2002, il marque également en Ligue des Champions face au Milan AC8. (A) Trace of external intercostal muscle EMG (IntEMG) activity shows respiratory activity of an anesthetized wild type mouse breathing 2.5% CO2 following injections of saline or U46619 (1 μM; 30 nL/region) into the cNTS and ROb. We also used a large stimulus (15% CO2) to characterize CO2/H+ vascular reactivity in vitro. These new results have been added as a new Figure 3F. View complete Tapology profile, bio, rankings, photos, news and record. The addition of CO2 was monitored to reach a maximum end-expiratory CO2 between 9% and 10%, which corresponds with an estimated arterial pH of 7.2 based on the following equation: pHa = 7.955–0.7215 × log10 (EtCO2).
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