scholarly journals Monocarboxylate Transport in Drosophila Larval Brain during Low and High Neuronal Activity

2019 ◽  
Author(s):  
Andres Gonzalez-Gutierrez ◽  
Andrés Ibacache ◽  
Andrés Esparza ◽  
L. Felipe Barros ◽  
Jimena Sierralta
Keyword(s):  
Glial Cells ◽  
Motor Neurons ◽  
Ex Vivo ◽  
Larval Brain ◽  
Metabolic Coupling ◽  
Monocarboxylate Transport ◽  
Lactate And Pyruvate ◽  
Increased Activity

ABSTRACTThe transport of lactate and pyruvate between glial cells and neurons plays an important role in the nervous system metabolic coupling. However, the mechanisms and characteristics that underlie the transport of monocarboxylates (MC-T) in vivo are poorly described. Here we use Drosophila expressing genetically-encoded FRET sensors to provide an ex vivo characterization of the MC-T in motor neurons and glial cells from the ventral nerve cord. We show that lactate/pyruvate transport on glial cells is coupled to protons and is more efficient than in neurons. Glial cells maintain higher levels of intracellular lactate generating a positive gradient towards neurons. Moreover, our results show that under increased activity lactate and pyruvate rise on motor neurons and suggest that this depends on the transfer of lactate from glial cells mediated in part by the previously described MC transporter Chaski, giving support to the in vivo glia to neurons lactate shuttling during activity.

scholarly journals Consequences of SUR2[A478V] Mutation in Skeletal Muscle of Murine Model of Cantu Syndrome

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1791
Author(s):  
Rosa Scala ◽  
Fatima Maqoud ◽  
Nicola Zizzo ◽  
Giuseppe Passantino ◽  
Antonietta Mele ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Katp Channel ◽  
Ex Vivo ◽  
Channel Modulation ◽  
Flexor Digitorum Brevis ◽  
Inflammatory Edema ◽  
Flexor Digitorum ◽  
Channel Currents

(1) Background: Cantu syndrome (CS) arises from gain-of-function (GOF) mutations in the ABCC9 and KCNJ8 genes, which encode ATP-sensitive K+ (KATP) channel subunits SUR2 and Kir6.1, respectively. Most CS patients have mutations in SUR2, the major component of skeletal muscle KATP, but the consequences of SUR2 GOF in skeletal muscle are unknown. (2) Methods: We performed in vivo and ex vivo characterization of skeletal muscle in heterozygous SUR2[A478V] (SUR2wt/AV) and homozygous SUR2[A478V] (SUR2AV/AV) CS mice. (3) Results: In SUR2wt/AV and SUR2AV/AV mice, forelimb strength and diaphragm amplitude movement were reduced; muscle echodensity was enhanced. KATP channel currents recorded in Flexor digitorum brevis fibers showed reduced MgATP-sensitivity in SUR2wt/AV, dramatically so in SUR2AV/AV mice; IC50 for MgATP inhibition of KATP currents were 1.9 ± 0.5 × 10−5 M in SUR2wt/AV and 8.6 ± 0.4 × 10−6 M in WT mice and was not measurable in SUR2AV/AV. A slight rightward shift of sensitivity to inhibition by glibenclamide was detected in SUR2AV/AV mice. Histopathological and qPCR analysis revealed atrophy of soleus and tibialis anterior muscles and up-regulation of atrogin-1 and MuRF1 mRNA in CS mice. (4) Conclusions: SUR2[A478V] “knock-in” mutation in mice impairs KATP channel modulation by MgATP, markedly so in SUR2AV/AV, with atrophy and non-inflammatory edema in different skeletal muscle phenotypes.

scholarly journals Characterization of Intrinsically Radiolabeled Poly(lactic-co-glycolic acid) Nanoparticles for ex Vivo Autologous Cell Labeling and in Vivo Tracking

2021 ◽  
Author(s):  
Massis Krekorian ◽  
Gerwin G. W. Sandker ◽  
Kimberley R. G. Cortenbach ◽  
Oya Tagit ◽  
N. Koen van Riessen ◽  
...  
Keyword(s):  
Glycolic Acid ◽  
Ex Vivo ◽  
Cell Labeling ◽  
Autologous Cell

About Puncture Testing Applied for Mechanical Characterization of Fetal Membranes

2014 ◽  
Vol 136 (11) ◽  
Author(s):  
Wilfried Bürzle ◽  
Edoardo Mazza ◽  
John J. Moore
Keyword(s):  
Deformation Behavior ◽  
Mechanical Characterization ◽  
Ex Vivo ◽  
Fetal Membrane ◽  
Membrane Rupture ◽  
Resistance To Deformation ◽  
Ex Vivo Testing ◽  
Insight Into

Puncture testing has been applied in several studies for the mechanical characterization of human fetal membrane (FM) tissue, and significant knowledge has been gained from these investigations. When comparing results of mechanical testing (puncture, inflation, and uniaxial tension), we have observed discrepancies in the rupture sequence of FM tissue and significant differences in the deformation behavior. This study was undertaken to clarify these discrepancies. Puncture experiments on FM samples were performed to reproduce previous findings, and numerical simulations were carried out to rationalize particular aspects of membrane failure. The results demonstrate that both rupture sequence and resistance to deformation depend on the samples' fixation. Soft fixation leads to slippage in the clamping, which reduces mechanical loading of the amnion layer and results in chorion rupturing first. Conversely, the stiffer, stronger, and less extensible amnion layer fails first if tight fixation is used. The results provide a novel insight into the interpretation of ex vivo testing as well as in vivo membrane rupture.

In vivo characterization of the transitional bronchioles by aerosol-derived airway morphometry

1999 ◽  
Vol 87 (3) ◽  
pp. 920-927 ◽  
Author(s):  
Kirby L. Zeman ◽  
Gerhard Scheuch ◽  
Knut Sommerer ◽  
James S. Brown ◽  
William D. Bennett
Keyword(s):  
Ex Vivo ◽  
Normal Subjects ◽  
Characteristic Line ◽  
Chronic Obstructive ◽  
Obstructive Pulmonary Disease ◽  
Lung Capacity ◽  
Single Breath ◽  
In Vivo Characterization

Effective airway dimensions (EADs) were determined in vivo by aerosol-derived airway morphometry as a function of volumetric lung depth (VLD) to identify and characterize, noninvasively, the caliber of the transitional bronchiole region of the human lung and to compare the EADs by age, gender, and disease. By logarithmically plotting EAD vs. VLD, two distinct regions of the lung emerged that were identified by characteristic line slopes. The intersection of proximal and distal segments was defined as VLDtransand associated EADtrans. In our normal subjects ( n = 20), VLDtrans [345 ± 83 (SD) ml] correlated significantly with anatomic dead space (224 ± 34 ml) and end of phase II of single-breath nitrogen washout (360 ± 53 ml). The corresponding EADtranswas 0.42 ± 0.07 mm, in agreement with other ex vivo measurements of the transitional bronchioles. VLDtrans was smaller (216 ± 64 ml) and EADtrans was larger (0.83 ± 0.04 mm) in our patients with chronic obstructive pulmonary disease ( n = 13). VLDtrans increased with age for children (age 8–18 yr; P = 0.006, n = 26) and with total lung capacity for age 8–81 yr ( P < 0.001, n = 61). This study extends the usefulness of aerosol-derived airway morphometry to in vivo measurements of the transitional bronchioles.

scholarly journals Ex Vivo Isolation and Characterization of Cd4+Cd25+ T Cells with Regulatory Properties from Human Blood

2001 ◽  
Vol 193 (11) ◽  
pp. 1303-1310 ◽  
Author(s):  
Detlef Dieckmann ◽  
Heidi Plottner ◽  
Susanne Berchtold ◽  
Thomas Berger ◽  
Gerold Schuler
Keyword(s):  
T Cells ◽  
Cytotoxic T Lymphocyte ◽  
Ex Vivo ◽  
Cell Receptor ◽  
Regulatory Cells ◽  
Suppressor T Cells ◽  
Isolation And Characterization ◽  
Regulatory Properties

It has been known for years that rodents harbor a unique population of CD4+CD25+ “professional” regulatory/suppressor T cells that is crucial for the prevention of spontaneous autoimmune diseases. Here we demonstrate that CD4+CD25+CD45RO+ T cells (mean 6% of CD4+ T cells) are present in the blood of adult healthy volunteers. In contrast to previous reports, these CD4+CD25+ T cells do not constitute conventional memory cells but rather regulatory cells exhibiting properties identical to their rodent counterparts. Cytotoxic T lymphocyte–associated antigen (CTLA)-4 (CD152), for example, which is essential for the in vivo suppressive activity of CD4+CD25+ T cells, was constitutively expressed, and remained strongly upregulated after stimulation. The cells were nonproliferative to stimulation via their T cell receptor for antigen, but the anergic state was partially reversed by interleukin (IL)-2 and IL-15. Upon stimulation with allogeneic (but not syngeneic) mature dendritic cells or platebound anti-CD3 plus anti-CD28 the CD4+CD25+ T cells released IL-10, and in coculture experiments suppressed the activation and proliferation of CD4+ and CD8+ T cells. Suppression proved IL-10 independent, yet contact dependent as in the mouse. The identification of regulatory CD4+CD25+ T cells has important implications for the study of tolerance in man, notably in the context of autoimmunity, transplantation, and cancer.

scholarly journals AAV9-mediated delivery of miR-23a reduces disease severity in Smn2B/−SMA model mice

2019 ◽  
Vol 28 (19) ◽  
pp. 3199-3210 ◽  
Author(s):  
Kevin A Kaifer ◽  
Eric Villalón ◽  
Benjamin S O'Brien ◽  
Samantha L Sison ◽  
Caley E Smith ◽  
...  
Keyword(s):  
Motor Neuron ◽  
Motor Neurons ◽  
Molecular Mechanisms ◽  
Viral Vector ◽  
Muscular Atrophy ◽  
Induced Pluripotent Stem Cell ◽  
Survival Motor Neuron ◽  
Virus Serotype

Abstract Spinal muscular atrophy (SMA) is a neuromuscular disease caused by deletions or mutations in survival motor neuron 1 (SMN1). The molecular mechanisms underlying motor neuron degeneration in SMA remain elusive, as global cellular dysfunction obscures the identification and characterization of disease-relevant pathways and potential therapeutic targets. Recent reports have implicated microRNA (miRNA) dysregulation as a potential contributor to the pathological mechanism in SMA. To characterize miRNAs that are differentially regulated in SMA, we profiled miRNA levels in SMA induced pluripotent stem cell (iPSC)-derived motor neurons. From this array, miR-23a downregulation was identified selectively in SMA motor neurons, consistent with previous reports where miR-23a functioned in neuroprotective and muscle atrophy-antagonizing roles. Reintroduction of miR-23a expression in SMA patient iPSC-derived motor neurons protected against degeneration, suggesting a potential miR-23a-specific disease-modifying effect. To assess this activity in vivo, miR-23a was expressed using a self-complementary adeno-associated virus serotype 9 (scAAV9) viral vector in the Smn2B/− SMA mouse model. scAAV9-miR-23a significantly reduced the pathology in SMA mice, including increased motor neuron size, reduced neuromuscular junction pathology, increased muscle fiber area, and extended survival. These experiments demonstrate that miR-23a is a novel protective modifier of SMA, warranting further characterization of miRNA dysfunction in SMA.

Ex vivo and in vivo characterization of cold preserved cartilage for cell transplantation

2016 ◽  
Vol 17 (4) ◽  
pp. 721-734 ◽  
Author(s):  
Wichaya Sriuttha ◽  
Nantawat Uttamo ◽  
Apisek Kongkaew ◽  
Jongkolnee Settakorn ◽  
Suchanan Rattanasalee ◽  
...  
Keyword(s):  
Cell Transplantation ◽  
Ex Vivo ◽  
In Vivo Characterization

Profiling substrate fluxes in the isolated working mouse heart using 13C-labeled substrates: focusing on the origin and fate of pyruvate and citrate carbons

2004 ◽  
Vol 286 (4) ◽  
pp. H1461-H1470 ◽  
Author(s):  
Maya Khairallah ◽  
François Labarthe ◽  
Bertrand Bouchard ◽  
Gawiyou Danialou ◽  
Basil J. Petrof ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Ex Vivo ◽  
Heart Function ◽  
Acid Oxidation ◽  
Mouse Heart ◽  
Relative Contribution ◽  
Cardiac Functions ◽  
Lactate And Pyruvate

The availability of genetically modified mice requires the development of methods to assess heart function and metabolism in the intact beating organ. With the use of radioactive substrates and ex vivo perfusion of the mouse heart in the working mode, previous studies have documented glucose and fatty acid oxidation pathways. This study was aimed at characterizing the metabolism of other potentially important exogenous carbohydrate sources, namely, lactate and pyruvate. This was achieved by using 13C-labeling methods. The mouse heart perfusion setup and buffer composition were optimized to reproduce conditions close to the in vivo milieu in terms of workload, cardiac functions, and substrate-hormone supply to the heart (11 mM glucose, 0.8 nM insulin, 50 μM carnitine, 1.5 mM lactate, 0.2 mM pyruvate, 5 nM epinephrine, 0.7 mM oleate, and 3% albumin). The use of three differentially 13C-labeled carbohydrates and a 13C-labeled long-chain fatty acid allowed the quantitative assessment of the metabolic origin and fate of tissue pyruvate as well as the relative contribution of substrates feeding acetyl-CoA (pyruvate and fatty acids) and oxaloacetate (pyruvate) for mitochondrial citrate synthesis. Beyond concurring with the notion that the mouse heart preferentially uses fatty acids for energy production (63.5 ± 3.9%) and regulates its fuel selection according to the Randle cycle, our study reports for the first time in the mouse heart the following findings. First, exogenous lactate is the major carbohydrate contributing to pyruvate formation (42.0 ± 2.3%). Second, lactate and pyruvate are constantly being taken up and released by the heart, supporting the concept of compartmentation of lactate and glucose metabolism. Finally, mitochondrial anaplerotic pyruvate carboxylation and citrate efflux represent 4.9 ± 1.8 and 0.8 ± 0.1%, respectively, of the citric acid cycle flux and are modulated by substrate supply. The described 13C-labeling strategy combined with an experimental setup that enables continuous monitoring of physiological parameters offers a unique model to clarify the link between metabolic alterations, cardiac dysfunction, and disease development.

scholarly journals Characterization of LAMP1-labeled nondegradative lysosomal and endocytic compartments in neurons

2018 ◽  
Vol 217 (9) ◽  
pp. 3127-3139 ◽  
Author(s):  
Xiu-Tang Cheng ◽  
Yu-Xiang Xie ◽  
Bing Zhou ◽  
Ning Huang ◽  
Tamar Farfel-Becker ◽  
...  
Keyword(s):  
Motor Neurons ◽  
Distribution Patterns ◽  
Confocal Imaging ◽  
Lysosomal Hydrolases ◽  
Differential Distribution ◽  
Gradient Fractionation ◽  
Endocytic Compartments ◽  
Lateral Sclerosis

Despite widespread distribution of LAMP1 and the heterogeneous nature of LAMP1-labeled compartments, LAMP1 is routinely used as a lysosomal marker, and LAMP1-positive organelles are often referred to as lysosomes. In this study, we use immunoelectron microscopy and confocal imaging to provide quantitative analysis of LAMP1 distribution in various autophagic and endolysosomal organelles in neurons. Our study demonstrates that a significant portion of LAMP1-labeled organelles do not contain detectable lysosomal hydrolases including cathepsins D and B and glucocerebrosidase. A bovine serum albumin–gold pulse–chase assay followed by ultrastructural analysis suggests a heterogeneity of degradative capacity in LAMP1-labeled endolysosomal organelles. Gradient fractionation displays differential distribution patterns of LAMP1/2 and cathepsins D/B in neurons. We further reveal that LAMP1 intensity in familial amyotrophic lateral sclerosis–linked motor neurons does not necessarily reflect lysosomal deficits in vivo. Our study suggests that labeling a set of lysosomal hydrolases combined with various endolysosomal markers would be more accurate than simply relying on LAMP1/2 staining to assess neuronal lysosome distribution, trafficking, and functionality under physiological and pathological conditions.

In Vivo and Ex Vivo characterization of a novel Er fiber laser system for fractional treatment of soft oral tissues

2018 ◽  
Author(s):  
Ksenia V. Shatilova ◽  
Georgii A. Aloian ◽  
Ilya V. Yaroslavsky ◽  
Gregory B. Altshuler ◽  
Maria Karabut ◽  
...  
Keyword(s):  
Fiber Laser ◽  
Ex Vivo ◽  
Laser System
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