scholarly journals Mitochondrial Ca2+ Dynamics in MCU Knockout C. elegans Worms

2020 ◽  
Vol 21 (22) ◽  
pp. 8622
Author(s):  
Pilar Álvarez-Illera ◽  
Paloma García-Casas ◽  
Rosalba I Fonteriz ◽  
Mayte Montero ◽  
Javier Alvarez
Keyword(s):  
Cell Activation ◽  
Physiological Activity ◽  
Large Cell ◽  
Atp Production ◽  
Physiological Conditions ◽  
C Elegans ◽  
Neuronal Stimulation ◽  
Low Amplitude ◽  
Stimulation Of

Mitochondrial [Ca2+] plays an important role in the regulation of mitochondrial function, controlling ATP production and apoptosis triggered by mitochondrial Ca2+ overload. This regulation depends on Ca2+ entry into the mitochondria during cell activation processes, which is thought to occur through the mitochondrial Ca2+ uniporter (MCU). Here, we have studied the mitochondrial Ca2+ dynamics in control and MCU-defective C. elegans worms in vivo, by using worms expressing mitochondrially-targeted YC3.60 yellow cameleon in pharynx muscle. Our data show that the small mitochondrial Ca2+ oscillations that occur during normal physiological activity of the pharynx were very similar in both control and MCU-defective worms, except for some kinetic differences that could mostly be explained by changes in neuronal stimulation of the pharynx. However, direct pharynx muscle stimulation with carbachol triggered a large and prolonged increase in mitochondrial [Ca2+] that was much larger in control worms than in MCU-defective worms. This suggests that MCU is necessary for the fast mitochondrial Ca2+ uptake induced by large cell stimulations. However, low-amplitude mitochondrial Ca2+ oscillations occurring under more physiological conditions are independent of the MCU and use a different Ca2+ pathway.

scholarly journals A day in the life of mitochondria reveals shifting workloads

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Tobias W. Weinrich ◽  
Jaimie Hoh Kam ◽  
Bill T. Ferrara ◽  
Elinor P. Thompson ◽  
John Mitrofanis ◽  
...  
Keyword(s):  
Complex I ◽  
Energy Capacity ◽  
Complex Activity ◽  
Spare Capacity ◽  
Atp Production ◽  
Cellular Energy ◽  
Changing Patterns ◽  
Differential Transcription ◽  
Stimulation Of

Abstract Mitochondria provide energy for cellular function. We examine daily changing patterns of mitochondrial function and metabolism in Drosophila in vivo in terms of their complex (I-IV) activity, ATP production, glycolysis, and whole fly respiration in the morning, afternoon and night. Complex activity and respiration showed significant and unexpected variation, peaking in the afternoon. However, ATP levels by contrast are >40% greater in the morning and lowest at night when glycolysis peaks. Complex activity modulation was at the protein level with no evidence for differential transcription over the day. Timing differences between increased ATP production and peaks of complex activity may result from more efficient ATP production early in the day leaving complex activity with spare capacity. Optical stimulation of mitochondria is only possible in the mornings when there is such spare capacity. These results provide first evidence of shifts in cellular energy capacity at the organism level. Understanding their translation may be significant to the chosen timing of energy demanding interventions to improve function and health.

scholarly journals In vivo stimulation of I kappa B phosphorylation is not sufficient to activate NF-kappa B.

1995 ◽  
Vol 15 (3) ◽  
pp. 1294-1301 ◽  
Author(s):  
I Alkalay ◽  
A Yaron ◽  
A Hatzubai ◽  
S Jung ◽  
A Avraham ◽  
...  
Keyword(s):  
T Cell Activation ◽  
Cell Activation ◽  
Extended Period ◽  
Activation Process ◽  
Nf Kappa B ◽  
Inflammatory Reactions ◽  
I Kappa B Alpha ◽  
Cytoplasmic Complex ◽  
Stimulation Of

NF-kappa B is a major inducible transcription factor in many immune and inflammatory reactions. Its activation involves the dissociation of the inhibitory subunit I kappa B from cytoplasmic NF-kappa B/Rel complexes, following which the Rel proteins are translocated to the nucleus, where they bind to DNA and activate transcription. Phosphorylation of I kappa B in cell-free experiments results in its inactivation and release from the Rel complex, but in vivo NF-kappa B activation is associated with I kappa B degradation. In vivo phosphorylation of I kappa B alpha was demonstrated in several recent studies, but its role is unknown. Our study shows that the T-cell activation results in rapid phosphorylation of I kappa B alpha and that this event is a physiological one, dependent on appropriate lymphocyte costimulation. Inducible I kappa B alpha phosphorylation was abolished by several distinct NF-kappa B blocking reagents, suggesting that it plays an essential role in the activation process. However, the in vivo induction of I kappa B alpha phosphorylation did not cause the inhibitory subunit to dissociate from the Rel complex. We identified several protease inhibitors which allow phosphorylation of I kappa B alpha but prevent its degradation upon cell stimulation, presumably through inhibition of the cytoplasmic proteasome. In the presence of these inhibitors, phosphorylated I kappa B alpha remained bound to the Rel complex in the cytoplasm for an extended period of time, whereas NF-kappa B activation was abolished. It appears that activation of NF-kappa B requires degradation of I kappa B alpha while it is a part of the Rel cytoplasmic complex, with inducible phosphorylation of the inhibitory subunit influencing the rate of degradation.

Disassembly of rat pancreatic acinar cell cytoskeleton during supramaximal secretagogue stimulation

1995 ◽  
Vol 268 (2) ◽  
pp. G328-G338 ◽  
Author(s):  
J. Jungermann ◽  
M. M. Lerch ◽  
H. Weidenbach ◽  
M. P. Lutz ◽  
B. Kruger ◽  
...  
Keyword(s):  
Acinar Cell ◽  
Digestive Enzyme ◽  
Experimental Models ◽  
Pancreatic Acinar Cell ◽  
Structural Proteins ◽  
Beta Tubulin ◽  
Physiological Conditions ◽  
Supramaximal Stimulation ◽  
Stimulation Of

In vivo stimulation of the exocrine pancreas with concentrations of secretagogue in excess of a maximally stimulating dose causes a marked disturbance of the intracellular segregation, transport, and exocytosis of digestive enzyme zymogens. Under physiological conditions elements of the cytoskeleton, most notably microtubules and microfilaments, are involved in the regulation of these intracellular events. We infused caerulein, a peptide analogue of cholecystokinin, at a supramaximal dose (10 micrograms.kg-1.h-1 for up to 6 h) intravenously in rats. To study the ultrastructural alterations of acinar cell microfilaments and microtubules by immunogold labeling, we used monoclonal antibodies directed against actin and beta-tubulin. As early as 30 min after the start of the secretagogue infusion we found a progressive disassembly of microtubules and microfilaments in exocrine cells. In immunoblot studies this disassembly of the cytoskeleton was paralleled by a degradation of its structural proteins actin and beta-tubulin. Our results indicate that the earliest morphological events during supramaximal secretagogue stimulation of the pancreas involve the disassembly and degradation of microtubules and microfilaments. This cell biological phenomenon offers an explanation for the disturbances of segregation, transport, and exocytosis of digestive enzymes, which are known to be associated with supramaximal stimulation of the pancreas and experimental models of pancreatitis.

scholarly journals Type I Interferon-mediated Stimulation of T Cells by CpG DNA

1998 ◽  
Vol 188 (12) ◽  
pp. 2335-2342 ◽  
Author(s):  
Siquan Sun ◽  
Xiaohong Zhang ◽  
David F. Tough ◽  
Jonathan Sprent
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Type I Interferons ◽  
Type I ◽  
Cpg Dna ◽  
Stimulation Of

Immunostimulatory DNA and oligodeoxynucleotides containing unmethylated CpG motifs (CpG DNA) are strongly stimulatory for B cells and antigen-presenting cells (APCs). We report here that, as manifested by CD69 and B7-2 upregulation, CpG DNA also induces partial activation of T cells, including naive-phenotype T cells, both in vivo and in vitro. Under in vitro conditions, CpG DNA caused activation of T cells in spleen cell suspensions but failed to stimulate highly purified T cells unless these cells were supplemented with APCs. Three lines of evidence suggested that APC-dependent stimulation of T cells by CpG DNA was mediated by type I interferons (IFN-I). First, T cell activation by CpG DNA was undetectable in IFN-IR−/− mice. Second, in contrast to normal T cells, the failure of purified IFN-IR−/− T cells to respond to CpG DNA could not be overcome by adding normal IFN-IR+ APCs. Third, IFN-I (but not IFN-γ) caused the same pattern of partial T cell activation as CpG DNA. Significantly, T cell activation by IFN-I was APC independent. Thus, CpG DNA appeared to stimulate T cells by inducing APCs to synthesize IFN-I, which then acted directly on T cells via IFN-IR. Functional studies suggested that activation of T cells by IFN-I was inhibitory. Thus, exposing normal (but not IFN-IR−/−) T cells to CpG DNA in vivo led to reduced T proliferative responses after TCR ligation in vitro.

scholarly journals Signaling through Toll-Like Receptors Induces Murine Gammaherpesvirus 68 Reactivation In Vivo

2008 ◽  
Vol 83 (3) ◽  
pp. 1474-1482 ◽  
Author(s):  
Lisa M. Gargano ◽  
J. Craig Forrest ◽  
Samuel H. Speck
Keyword(s):  
B Cell ◽  
Cell Activation ◽  
B Cell Activation ◽  
Toll Like Receptors ◽  
Murine Gammaherpesvirus ◽  
Latently Infected ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68 ◽  
Stimulation Of

ABSTRACT Murine gammaherpesvirus 68 (MHV68) establishes a lifelong infection in mice and is used as a model pathogen to study the role of viral and host factors in chronic infection. The maintenance of chronic MHV68 infection, at least in some latency reservoirs, appears to be dependent on the capacity of the virus to reactivate from latency in vivo. However, the signals that lead to MHV68 reactivation in vivo are not well characterized. Toll-like receptors (TLRs), by recognizing the specific patterns of microbial components, play an essential role in the activation of innate immunity. In the present study, we investigated the capacity of TLR ligands to induce MHV68 reactivation, both in vitro and in vivo. The stimulation of latently infected B cell lines with ligands for TLRs 3, 4, 5, and 9 enhanced MHV68 reactivation; the ex vivo stimulation of latently infected primary splenocytes, recovered from infected mice, with poly(I:C), lipopolysaccharide, flagellin, or CpG DNA led to early B-cell activation, B-cell proliferation, and a significant increase in the frequency of latently infected cells reactivating the virus. In vivo TLR stimulation also induced B-cell activation and MHV68 reactivation, resulting in heightened levels of virus replication in the lungs which correlated with an increase in MHV68-specific CD8+ T-cell responses. Importantly, TLR stimulation also led to an increase in MHV68 latency, as evidenced by an increase in viral genome-positive cells 2 weeks post-in vivo stimulation by specific TLR ligands. Thus, these data demonstrate that TLR stimulation can drive MHV68 reactivation from latency and suggests that periodic pathogen exposure may contribute to the homeostatic maintenance of chronic gammaherpesvirus infection through stimulating virus reactivation and reseeding latency reservoirs.

scholarly journals Direct activation of the mitochondrial calcium uniporter by natural plant flavonoids

2004 ◽  
Vol 384 (1) ◽  
pp. 19-24 ◽  
Author(s):  
Mayte MONTERO ◽  
Carmen D. LOBATÓN ◽  
Esther HERNÁNDEZ-SANMIGUEL ◽  
Jaime SANTODOMINGO ◽  
Laura VAY ◽  
...  
Keyword(s):  
Cell Activation ◽  
Fold Increase ◽  
Mitogen Activated Protein Kinase ◽  
Intact Cells ◽  
Natural Plant ◽  
Physiological Conditions ◽  
Calcium Uniporter ◽  
Strong Stimulation ◽  
Mitogen Activated Protein ◽  
Stimulation Of

During cell activation, mitochondria play an important role in Ca2+ homoeostasis due to the presence of a fast and specific Ca2+ channel in its inner membrane, the mitochondrial Ca2+ uniporter. This channel allows mitochondria to buffer local cytosolic [Ca2+] changes and controls the intramitochondrial Ca2+ levels, thus modulating a variety of phenomena from respiratory rate to apoptosis. We have described recently that SB202190, an inhibitor of p38 MAPK (mitogen-activated protein kinase), strongly activated the uniporter. We show in the present study that a series of natural plant flavonoids, widely distributed in foods, produced also a strong stimulation of the mitochondrial Ca2+ uniporter. This effect was of the same magnitude as that induced by SB202190 (an approx. 20-fold increase in the mitochondrial Ca2+ uptake rate), developed without measurable delay and was rapidly reversible. In intact cells, the mitochondrial Ca2+ peak induced by histamine was also largely increased by the flavonoids. Stimulation of the uniporter by either flavonoids or SB202190 did not require ATP, suggesting a direct effect on the uniporter or an associated protein which is not mediated by protein phosphorylation. The most active compound, kaempferol, increased the rate of mitochondrial Ca2+ uptake by 85±15% (mean±S.E.M., n=4) and the histamine-induced mitochondrial Ca2+ peak by 139±19% (mean±S.E.M., n=5) at a concentration of 1 μM. Given that flavonoids can reach this concentration range in plasma after ingestion of flavonoid-rich food, these compounds could be modulating the uniporter under physiological conditions.

scholarly journals Multifunctional Cre-dependent transgenic mice for high-precision all-optical interrogation of neural circuits

2021 ◽  
Author(s):  
Hayley A Bounds ◽  
Masato Sadahiro ◽  
William D Hendricks ◽  
Marta Gajowa ◽  
Ian Antón Oldenburg ◽  
...  
Keyword(s):  
Neural Circuits ◽  
Physiological Activity ◽  
Large Cell ◽  
Optical Systems ◽  
Mammalian Brain ◽  
Neural Ensemble ◽  
Activity Measurements ◽  
All Optical

Causally relating the detailed structure and function of neural circuits to behavior requires the ability to precisely and simultaneously write-in and read-out neural activity. All optical systems that combine two photon (2p) calcium imaging and targeted photostimulation provide such an approach, but require co-expression of an activity indicator, such as GCaMP, and an optogenetic actuator, ideally a potent soma-targeted opsin. In the mammalian brain, such co-expression has so far been achieved by viral transduction, which is invasive and can produce variable, focal, and sometimes toxic overexpression. To overcome this challenge, we developed and validated a Cre-reporter mouse ("Ai203") that conditionally expresses a soma-targeted opsin, ChroME, fused to GCaMP7s. 1p or 2p illumination of expressing neurons in vitro and in vivo produces powerful, precise activation comparable to viral expression of ChroME. The soma-targeted GCaMP7s provides sensitive activity measurements for tracking physiological activity, and the soma-targeted ChroME provides powerful control over neural ensemble activity with holographic optogenetics. We further demonstrate the use of the Ai203 reporter line in 1p optogenetic manipulation of performance on a cortex-dependent visual task and in 2p synaptic connectivity mapping. This new transgenic line could thus greatly facilitate the study of neural circuits by providing a flexible, convenient, and stable tool for all-optical access to large, cell-type specific neural populations throughout the nervous system.

scholarly journals Interplay of target site architecture and miRNA abundance determine miRNA activity and specificity

2017 ◽  
Author(s):  
Giovanna Brancati ◽  
Sarah H. Carl ◽  
Helge Großhans
Keyword(s):  
Mirna Family ◽  
Target Site ◽  
Site Quality ◽  
Seed Match ◽  
Base Pairs ◽  
Physiological Conditions ◽  
C Elegans ◽  
Mirna Seed ◽  
Mirna Abundance

ABSTRACTThe recognition that the miRNA seed sequence is a major determinant of miRNA activity has greatly advanced the ability to predict miRNA targets. However, it has remained unclear to what extent miRNAs act redundantly when they are members of the same family and thus share a common seed. Using in vivo studies in C. elegans, we uncover features that drive specific target repression by individual miRNA family members. We find that seed-distal complementarity to a specific family member promotes specificity. However, the extent and robustness of specificity are greatly increased by seed match ‘imperfections’, such as bulges and G:U wobble base pairs. Depending on the seed match architecture, specificity may be overcome by increasing the levels of a miRNA lacking seed-distal complementarity. Hence, in contrast to a binary distinction between functional and non-functional target sites, our data support a model where functionality depends on a combination of target site quality and miRNA abundance. This emphasizes the importance of studying miRNAs under physiological conditions in their endogenous contexts.

scholarly journals Stimulation of astrocyte fatty acid oxidation by thyroid hormone is protective against ischemic stroke-induced damage

2016 ◽  
Vol 37 (2) ◽  
pp. 514-527 ◽  
Author(s):  
Naomi L Sayre ◽  
Mikaela Sifuentes ◽  
Deborah Holstein ◽  
Sheue-yann Cheng ◽  
Xuguang Zhu ◽  
...  
Keyword(s):  
Ischemic Stroke ◽  
Fatty Acid ◽  
Stroke Volume ◽  
Fatty Acid Oxidation ◽  
Glucose Deprivation ◽  
Acid Oxidation ◽  
Atp Production ◽  
Stimulation Of

We previously demonstrated that stimulation of astrocyte mitochondrial ATP production via P2Y1 receptor agonists was neuroprotective after cerebral ischemic stroke. Another mechanism that increases ATP production is fatty acid oxidation (FAO). We show that in primary human astrocytes, FAO and ATP production are stimulated by 3,3,5 triiodo-l-thyronine (T3). We tested whether T3-stimulated FAO enhances neuroprotection, and show that T3 increased astrocyte survival after either hydrogen peroxide exposure or oxygen glucose deprivation. T3-mediated ATP production and protection were both eliminated with etomoxir, an inhibitor of FAO. T3-mediated protection in vitro was also dependent on astrocytes expressing HADHA (hydroxyacyl-CoA dehydrogenase/3-ketoacyl-CoA thiolase/enoyl-CoA hydratase), which we previously showed was critical for T3-mediated FAO in fibroblasts. Consistent with previous reports, T3-treatment decreased stroke volumes in mice. While T3 decreased stroke volume in etomoxir-treated mice, T3 had no protective effect on stroke volume in HADHA +/− mice or in mice unable to upregulate astrocyte-specific energy production. In vivo, 95% of HADHA co-localize with glial-fibrillary acidic protein, suggesting the effect of HADHA is astrocyte mediated. These results suggest that astrocyte-FAO modulates lesion size and is required for T3-mediated neuroprotection post-stroke. To our knowledge, this is the first report of a neuroprotective role for FAO in the brain.

Influence of thiol substances on in vitro and in vivoL-thyroxine deiodination in rainbow trout, Salmo gairdneri

1984 ◽  
Vol 62 (8) ◽  
pp. 1495-1501 ◽  
Author(s):  
J. G. Eales ◽  
Shirley Shostak ◽  
Catherine G. Flood
Keyword(s):  
Rainbow Trout ◽  
Reduced Glutathione ◽  
Salmo Gairdneri ◽  
Physiological Conditions ◽  
Low Concentrations ◽  
Dtt Dithiothreitol ◽  
High Concentrations ◽  
Stimulation Of

The effects of the thiols DTT (dithiothreitol) and GSH (reduced glutathione) on hepatic in vitro and in vivo T4 (L-thyroxine) deiodination by rainbow trout held at 11 °C were studied. Hepatic deiodination increased progressively over the DTT range of 0.02–20 mM. GSH was less potent than DTT at low concentrations and strongly inhibited deiodination at high concentrations (> 1 mM). Hepatic deiodination was not increased by 1 mM NADPH or anaerobic conditions and was enhanced and not inhibited by the GSH inhibitor, diamide (2.5 mM), indicating that the low T4 deiodination in the absence of DTT is not due to endogenous GSH deficiency. Intraperitoneally injected GSH consistently increased plasma levels of 125I and [125I]-3,5,3′-triiodo-L-thyronine (T3) in fed or starved [125I]T4-injected trout, suggesting a GSH stimulation of extrahepatic T4 deiodination. However, injected GSH did not elevate plasma T3 concentrations. This was probably due to a demonstrated GSH stimulation of plasma T4 and T3 clearance. Force-fed GSH did not increase [125I]T4 deiodination. It is concluded that exogenous thiols can enhance T4 deiodination both in vitro and in vivo. However, availability of neither endogenous nor dietary GSH appears to regulate T4 deiodination under physiological conditions, including altered nutritional state.

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