neuromodulation-mitochondrial-transport-live-imaging-hippocampal

Neuromodulation and Mitochondrial Transport: Live Imaging in Hippocampal Neurons over Long Durations

To understand the relationship between mitochondrial transport and neuronal function, it is critical to observe mitochondrial behavior in live cultured ...

Regulation of vascular smooth muscle cell growth and adhesion by paired-related homeobox genes.

Regulierung der vaskul&#xE4;ren glatten Muskulatur Zellwachstum und Haftung durch gepaarte-Suchbegriffe Hom&#xF6;oboxgene.

Das Protein Hom&#xF6;odom&#xE4;ne Barx2 &#x200B;&#x200B;f&#xF6;rdert myogenen Differenzierung und wird von myogenen regulatorischen Faktoren reguliert.

The homeobox protein Barx2 is expressed in both smooth and skeletal muscle and is up-regulated during differentiation of skeletal myotubes. Here we use antisense-oligonucleotide inhibition of Barx2 expression in limb bud cell culture to show that Barx2 is required for myotube formation. Moreover, overexpression of Barx2 accelerates the fusion of MyoD-positive limb bud cells and C2C12 myoblasts. However, overexpression of Barx2 does not induce ectopic MyoD expression in either limb bud cultures or in multipotent C3H10T1/2 mesenchymal cells, and does not induce fusion of C3H10T1/2 cells. These results suggest that Barx2 acts downstream of MyoD. To test this hypothesis, we isolated the Barx2 gene promoter and identified DNA regulatory elements that might control Barx2 expression during myogenesis. The proximal promoter of the Barx2 gene contained binding sites for several factors involved in myoblast differentiation including MyoD, myogenin, serum response factor, and myocyte enhancer factor 2. Co-transfection experiments showed that binding sites for both MyoD and serum response factor are necessary for activation of the promoter by MyoD and myogenin. Taken together, these studies indicate that Barx2 is a key regulator of myogenic differentiation that acts downstream of muscle regulatory factors.

The homeodomain protein Barx2 promotes myogenic differentiation and is regulated by myogenic regulatory factors.

Extracellular hypertonicity can induce the phosphorylation of mitogen-activated protein kinases (MAPKs). Of these, both extracellular signal-regulated kinases (ERKs) and the stress-activated kinase p38 have been implicated in neuronal cell survival. Resuscitation with hypertonic saline decreases secondary brain injury after trauma, as well as neuronal damage, after ischemia. Since hypertonicity has been shown to support somatic cell survival, we investigated if hypertonicity can also prevent neuronal cell death via MAPK signaling. Death of postnatal rat corticospinal motoneurons (CSMNs) was induced by serum deprivation, and survival in both isotonic and hypertonic media was assessed after 20 h. Addition of NaCl (4-250 mM) to isotonic medium significantly and dose dependently protected CSMN in enriched cultures, increasing cell survival by up to 70% over that in isotonic medium. This response was not restricted to NaCl; addition of KCl, choline chloride, and sucrose had similar effects on cell survival. In addition, hypertonicity supported the survival of pure CSMN populations, albeit with lower potency. In cortical cell suspensions, hypertonic NaCl (20-100 mM) increased basal phosphorylation of p38 and ERK. The activation of both MAPKs, which was induced by 40 mM NaCl, was transient. Cultivation of CSMNs in media containing the specific p38 inhibitor SB203580 abolished the protective effect of hypertonic NaCl, indicating a central role for p38. We therefore conclude that hypertonicity can prevent neuronal cell death via MAPK signaling.

Hypertonicity promotes survival of corticospinal motoneurons via mitogen-activated protein kinase p38 signaling.

Hypertonus f&#xF6;rdert das &#xDC;berleben von Motoneuronen kortikospinalen &#xFC;ber Mitogen-aktivierten Protein-Kinase p38-Signalisierung.

BARX2 is a homeobox transcription factor that influences cellular differentiation in various developmental contexts. To begin to identify the gene targets that mediate its effects, chromatin immunoprecipitation (ChIP) was used to isolate BARX2 binding sites from the human MCF7 breast cancer cell line. Cloning and sequencing of BARX2-ChIP-derived DNA fragments identified 60 potential BARX2 target loci that were proximal to or within introns of genes involved in cytoskeletal organization, cell adhesion, growth factor signaling, transcriptional regulation, and RNA metabolism. The sequences of over half of the fragments showed homology with the mouse genome, and several sequences could be mapped to orthologous human and mouse genes. Binding of BARX2 to 21 genomic loci examined was confirmed quantitatively by replicate ChIP assays. A combination of sequence analysis and electrophoretic mobility shift assays revealed homeodomain binding sites within several fragments that bind to BARX2 in vitro. The majority of BARX2 binding fragments tested (14/19), also affected transcription in luciferase reporter gene assays. Mutation analyses of three fragments showed that their transcriptional activities required the HBS, and suggested that BARX2 regulates gene expression by binding to DNA elements containing paired TAAT motifs that are separated by a poly(T) sequence. Inhibition of BARX2 expression in MCF7 cells led to reduced expression of eight genes associated with BARX2 binding sites, indicating that BARX2 directly regulates their expression. The data suggest that BARX2 can coordinate the expression of a network of genes that influence the growth of MCF7 cells.

Identification of novel binding elements and gene targets for the homeodomain protein BARX2.

Identifizierung neuer Bindungselemente und Gen-Targets f&#xFC;r die Hom&#xF6;odom&#xE4;ne Protein BARX2.

Increasingly, researchers are recognizing the limitations of two-dimensional (2-D), monolayer cell culture and embracing more realistic three-dimensional (3-D) cell culture systems. Currently, 3-D culture techniques are being employed by neuroscientists to grow cells from the central nervous system. From this work, it has become clear that 3-D cell culture offers a more realistic milieu in which the functional properties of neurons can be observed and manipulated in a manner that is not possible in vivo. The implications of this technical renaissance in cell culture for both clinical and basic neuroscience are significant and far-reaching.

A cultural renaissance: in vitro cell biology embraces three-dimensional context.

Eine kulturelle Renaissance: In-vitro-Zellbiologie umarmt dreidimensionalen Kontext.

The standard behavioral index for human consciousness is the ability to report events with accuracy. While this method is routinely used for scientific and medical applications in humans, it is not easy to generalize to other species. Brain evidence may lend itself more easily to comparative testing. Human consciousness involves widespread, relatively fast low-amplitude interactions in the thalamocortical core of the brain, driven by current tasks and conditions. These features have also been found in other mammals, which suggests that consciousness is a major biological adaptation in mammals. We suggest more than a dozen additional properties of human consciousness that may be used to test comparative predictions. Such homologies are necessarily more remote in non-mammals, which do not share the thalamocortical complex. However, as we learn more we may be able to make "deeper" predictions that apply to some birds, reptiles, large-brained invertebrates, and perhaps other species.

Criteria for consciousness in humans and other mammals.

Kriterien f&#xFC;r Bewusstsein bei Menschen und anderen S&#xE4;ugetieren.

Most early studies of consciousness have focused on human subjects. This is understandable, given that humans are capable of reporting accurately the events they experience through language or by way of other kinds of voluntary response. As researchers turn their attention to other animals, "accurate report" methodologies become increasingly difficult to apply. Alternative strategies for amassing evidence for consciousness in non-human species include searching for evolutionary homologies in anatomical substrates and measurement of physiological correlates of conscious states. In addition, creative means must be developed for eliciting behaviors consistent with consciousness. In this paper, we explore whether necessary conditions for consciousness can be established for species as disparate as birds and cephalopods. We conclude that a strong case can be made for avian species and that the case for cephalopods remains open. Nonetheless, a consistent effort should yield new means for interpreting animal behavior.

Identifying hallmarks of consciousness in non-mammalian species.

Identifizieren Markenzeichen des Bewusstseins in Nicht-S&#xE4;uger-Spezies.

Among the many factors involved in regulation of chondrogenesis, bone morphogenetic proteins (BMPs) and members of the Sox and homeobox transcription factor families have been shown to have crucial roles. Of these regulators, the homeobox transcription factors that function during chondrogenesis have been the least well defined. We show here that the homeobox transcription factor Barx2 is expressed in primary mesenchymal condensations, digital rays, developing joints and articular cartilage of the developing limb, suggesting that it plays a role in chondrogenesis. Using retroviruses and antisense oligonucleotides to manipulate Barx2 expression in limb bud micromass cultures, we determined that Barx2 is necessary for mesenchymal aggregation and chondrogenic differentiation. In accordance with these findings, Barx2 regulates the expression of several genes encoding cell-adhesion molecules and extracellular matrix proteins, including NCAM and collagen II (Col2a1) in the limb bud. Barx2 bound to elements within the cartilage-specific Col2a1 enhancer, and this binding was reduced by addition of Barx2 or Sox9 antibodies, or by mutation of a HMG box adjacent to the Barx2-binding element, suggesting cooperation between Barx2 and Sox proteins. Moreover, both Barx2 and Sox9 occupy Col2a1 enhancer during chondrogenesis in vivo. We also found that two members of the BMP family that are crucial for chondrogenesis, GDF5 and BMP4, regulate the pattern of Barx2 expression in developing limbs. Based on these data, we suggest that Barx2 acts downstream of BMP signaling and in concert with Sox proteins to regulate chondrogenesis.

The homeobox transcription factor Barx2 regulates chondrogenesis during limb development.

Die homeobox Transkriptionsfaktor reguliert Barx2 &#x200B;&#x200B;Chondrogenese w&#xE4;hrend der Entwicklung der Extremit&#xE4;ten.

Axonal transport of mitochondria is critical for proper neuronal function. However, little is known about the extracellular signals that regulate this process. In the present study, we show that the neuromodulator serotonin (5-HT) greatly enhances mitochondrial movement in the axons of rat hippocampal neurons in vitro. Administration of a 5-HT1A receptor antagonist inhibited mitochondrial movement, whereas addition of fluoxetine, a selective serotonin reuptake inhibitor, promoted mitochondrial movement. 5-HT receptors are known to activate the Akt/Protein kinase B pathway. Consistent with this, directional mitochondrial movement was almost completely blocked by a specific Akt inhibitor. Moreover, an inhibitor of glycogen synthase kinase-3beta (GSK3beta), a kinase whose activity is blocked by Akt-mediated phosphorylation, promoted mitochondrial movement. These findings show that 5-HT1A receptor activation stimulates mitochondrial movement in hippocampal neurons by inhibiting GSK3beta activity via Akt. Our findings suggest that 5-HT may mediate the redistribution of energy sources within responsive neurons, a possibility that has significant implications for understanding the global biological effects of this important neuromodulator.

Serotonin stimulates mitochondrial transport in hippocampal neurons.

Axonale Transport der Mitochondrien ist kritisch f&uuml;r neuronale Funktion. Jedoch ist wenig bekannt &uuml;ber die extrazellul&auml;re Signale, die diesen Prozess regulieren. In der vorliegenden Studie zeigen wir, dass das oder Neuromodulatoren Serotonin (5-HT) steigert erheblich die mitochondriale Bewegung in die Axone der Ratte Neuronen in-vitro. Verwaltung von einer 5-HT1A-Rezeptor-Antagonist gehemmt mitochondriale Bewegung, w&auml;hrend die Zugabe von Fluoxetin, ein selektiver Serotonin-Wiederaufnahme-Hemmer, mitochondriale Bewegung gef&ouml;rdert. 5-HT-Rezeptoren sind bekannterma&szlig;en die Akt/Protein-Kinase B-Signalweg aktivieren. Einklang mit dieser, direktionale wurde mitochondriale Bewegung fast vollst&auml;ndig von einem bestimmten Akt-Inhibitor blockiert. Zudem schuf ein Inhibitor der Glykogen-Synthase Kinase-3beta (GSK3beta), eine Kinase, dessen Aktivit&auml;t durch Akt-vermittelte Phosphorylierung, blockiert ist, mitochondriale Bewegung. Diese Ergebnisse zeigen, dass 5-HT1A-Rezeptor-Aktivierung mitochondriale Bewegung in Neuronen stimuliert durch die Hemmung der GSK3beta Aktivit&auml;t &uuml;ber Akt. Unsere Forschungsergebnisse legen nahe, dass die 5-HT die Umverteilung von Energiequellen innerhalb reagieren Neuronen eine M&ouml;glichkeit vermitteln kann, die erhebliche Auswirkungen auf das Verst&auml;ndnis der globalen biologischen Auswirkungen von diesem wichtigen oder Neuromodulatoren hat.

Serotonin stimuliert mitochondriale Verkehr in Neuronen.

The trafficking of mitochondria within neurons is a highly regulated process. In an earlier study, we found that serotonin (5-HT), acting through the 5-HT1A receptor subtype, promotes axonal transport of mitochondria in cultured hippocampal neurons by increasing Akt activity, and consequently decreasing glycogen synthase kinase (GSK3beta) activity. This finding suggests a critical role for neuromodulators in the regulation of mitochondrial trafficking in neurons. In the present study, we investigate the effects of a second important neuromodulator, dopamine, on mitochondrial transport in hippocampal neurons.

Dopamine inhibits mitochondrial motility in hippocampal neurons.

Des Handels mit Mitochondrien innerhalb der Neuronen ist ein stark regulierten Prozess. In einer fr&uuml;heren Studie fanden wir, dass Serotonin (5-HT), handelnd durch den 5-HT1A-Rezeptor-Subtyp, axonale Transport von Mitochondrien in kultivierten Neuronen f&ouml;rdert durch Erh&ouml;hung Akt-Aktivit&auml;t, und folglich abnehmender Glykogen-Synthase-Kinaseaktivit&auml;t (GSK3beta). Diese Erkenntnis schl&auml;gt eine entscheidende Rolle f&uuml;r die Neuromodulators bei der Regulierung der mitochondrialen Menschenhandel Neuronen. In der vorliegenden Studie untersuchen wir die Auswirkungen der eine zweite wichtige oder Neuromodulatoren, Dopamin, mitochondriale Verkehr in Neuronen.

Dopamin hemmt mitochondriale Motilit&auml;t in Neuronen.

Obgleich anekdotischer bewusste Staaten in einer Vielzahl von nicht-menschlichen Tiere wurde noch keine systematische neurowissenschaftliche Untersuchung des tierischen Bewusstseins durchgef&uuml;hrt. Weiter setzen wir einen Rahmen f&uuml;r eine solche Untersuchung, die Integration von Daten aus Neuroanatomie, Neurophysiologie und behavioral Studies, Verwendungen Beweise von den Menschen als Ma&szlig;stab umfasst ein und erkennt die wichtige Rolle von expliziten m&uuml;ndlichen Bericht des bewussten Erfahrungen in Studien am Menschen. Wir zeigen unser Framework unter Bezugnahme auf zwei Subphyla: eine relativ nahe S&auml;ugetiere - V&ouml;gel- und eine ziemlich weit - Kopff&uuml;&szlig;ern Weichtiere. In &Uuml;bereinstimmung mit der M&ouml;glichkeit der bewussten Staaten, beide Subphyla zeigen komplexes Verhalten und anspruchsvolle Nervensystem besitzen. Ihre weitere Untersuchungen kann gemeinsame Phyletisches Bedingungen und neuralen Substrate zugrunde liegt die Entstehung des tierischen Bewusstseins offenbaren.

Tierische Bewusstsein: einen synthetischen Ansatz.

Despite anecdotal evidence suggesting conscious states in a variety of non-human animals, no systematic neuroscientific investigation of animal consciousness has yet been undertaken. We set forth a framework for such an investigation that incorporates integration of data from neuroanatomy, neurophysiology, and behavioral studies, uses evidence from humans as a benchmark, and recognizes the critical role of explicit verbal report of conscious experiences in human studies. We illustrate our framework with reference to two subphyla: one relatively near to mammals - birds - and one quite far -cephalopod molluscs. Consistent with the possibility of conscious states, both subphyla exhibit complex behavior and possess sophisticated nervous systems. Their further investigation may reveal common phyletic conditions and neural substrates underlying the emergence of animal consciousness.

Animal consciousness: a synthetic approach.

Tubulin is a major substrate of the cytoplasmic class II histone deacetylase HDAC6. Inhibition of HDAC6 results in higher levels of acetylated tubulin and enhanced binding of the motor protein kinesin-1 to tubulin, which promotes transport of cargoes along microtubules. Microtubule-dependent intracellular trafficking may therefore be regulated by modulating the activity of HDAC6. We have shown previously that the neuromodulator serotonin increases mitochondrial movement in hippocampal neurons via the Akt-GSK3beta signaling pathway. Here, we demonstrate a role for HDAC6 in this signaling pathway.

HDAC6 regulates mitochondrial transport in hippocampal neurons.

Tubulin ist ein wichtiger Substrat der zytoplasmatischen Klasse II Histon Deacetylase HDAC6. Hemmung der HDAC6 resultiert in h&ouml;heren Ebenen der acetylierten Tubulin und verst&auml;rkte Bindung der motor Protein Kinesin-1 Tubulin, die Bef&ouml;rderung von Frachten entlang der Mikrotubuli f&ouml;rdert. Microtubule-abh&auml;ngige intrazellul&auml;re Menschenhandel kann daher durch Modulation der Aktivit&auml;t des HDAC6 geregelt werden. Wir haben bereits gezeigt, dass das Serotonin oder Neuromodulatoren mitochondriale Bewegung in Neuronen &uuml;ber den Akt-GSK3beta-Signalweg erh&ouml;ht. Hier zeigen wir eine Rolle f&uuml;r die HDAC6 in dieser Signalweg.

HDAC6 regelt die mitochondriale Verkehr in Neuronen.

How uniquely irreducible is consciousness? Defining the limits of biological reductionism A commentary on Neuroontology, neurobiological naturalism, and consciousness: A challenge to scientific reduction and a solution, by Todd E. Feinberg, MD.

Wie eindeutig irreduzibel ist Bewusstsein? Definieren die Grenzen des biologischen Reduktionismus A Kommentar auf Neuroontology, neurobiologische Naturalismus und Bewusstsein: eine Herausforderung f&uuml;r wissenschaftliche Reduktion und eine L&ouml;sung von Todd E. Feinberg, MD.

Article	Year
Regulation of vascular smooth muscle cell growth and adhesion by paired-related homeobox genes. Chest\| PubMed ID: 11893718	2002
The homeodomain protein Barx2 promotes myogenic differentiation and is regulated by myogenic regulatory factors. The Journal of biological chemistry\| PubMed ID: 12486129	2003
Hypertonicity promotes survival of corticospinal motoneurons via mitogen-activated protein kinase p38 signaling. Journal of molecular neuroscience : MN\| PubMed ID: 14593211	2003
Identification of novel binding elements and gene targets for the homeodomain protein BARX2. The Journal of biological chemistry\| PubMed ID: 14744868	2004
A cultural renaissance: in vitro cell biology embraces three-dimensional context. Experimental neurology\| PubMed ID: 15698613	2005
Criteria for consciousness in humans and other mammals. Consciousness and cognition\| PubMed ID: 15766894	2005
Identifying hallmarks of consciousness in non-mammalian species. Consciousness and cognition\| PubMed ID: 15766896	2005
The homeobox transcription factor Barx2 regulates chondrogenesis during limb development. Development (Cambridge, England)\| PubMed ID: 15800003	2005
Serotonin stimulates mitochondrial transport in hippocampal neurons. Molecular and cellular neurosciences\| PubMed ID: 17904380	2007
Dopamine inhibits mitochondrial motility in hippocampal neurons. PloS one\| PubMed ID: 18665222	2008
Animal consciousness: a synthetic approach. Trends in neurosciences\| PubMed ID: 19716185	2009
HDAC6 regulates mitochondrial transport in hippocampal neurons. PloS one\| PubMed ID: 20520769	2010
How uniquely irreducible is consciousness? Defining the limits of biological reductionism A commentary on Neuroontology, neurobiological naturalism, and consciousness: A challenge to scienti... Physics of life reviews\| PubMed ID: 22243993	2012

David B. Edelman

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