David B. Edelman

HDAC6 regulates mitochondrial transport in hippocampal neurons.

Background 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-GSK3β signaling pathway. Here, we demonstrate a role for HDAC6 in this signaling pathway. Methodology/Principal Findings We found that the presence of tubacin, a specific HDAC6 inhibitor, dramatically enhanced mitochondrial movement in hippocampal neurons, whereas niltubacin, an inactive tubacin analog, had no effect. Compared to control cultures, higher levels of acetylated tubulin were found in neurons treated with tubacin, and more kinesin-1 was associated with mitochondria isolated from these neurons. Inhibition of GSK3β decreased cytoplasmic deacetylase activity and increased tubulin acetylation, whereas blockade of Akt, which phosphorylates and down-regulates GSK3β, increased cytoplasmic deacetylase activity and decreased tubulin acetylation. Concordantly, the administration of 5-HT, 8-OH-DPAT (a specific 5-HT1A receptor agonist), or fluoxetine (a 5-HT reuptake inhibitor) increased tubulin acetylation. GSK3β was found to co-localize with HDAC6 in hippocampal neurons, and inhibition of GSK3β resulted in decreased binding of antibody to phosphoserine-22, a potential GSK3β phosphorylation site in HDAC6. GSK3β may therefore regulate HDAC6 activity by phosphorylation. Conclusions/Significance This study demonstrates that HDAC6 plays an important role in the modulation of mitochondrial transport. The link between HDAC6 and GSK3β, established here, has important implications for our understanding of neurodegenerative disorders. In particular, abnormal mitochondrial transport, which has been observed in such disorders as Alzheimers disease and Parkinsons disease, could result from the misregulation of HDAC6 by GSK3β. HDAC6 may therefore constitute an attractive target in the treatment of these disorders.

Serotonin stimulates mitochondrial transport in hippocampal neurons

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.

Animal consciousness: a synthetic approach

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.

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

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.

Dopamine Inhibits Mitochondrial Motility in Hippocampal Neurons

Background 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 (GSK3β) 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. Methodology/Principal Findings Here, we show that dopamine, like 5-HT, regulates mitochondrial motility in cultured hippocampal neurons through the Akt-GSK3β signaling cascade. But, in contrast to the stimulatory effect of 5-HT, administration of exogenous dopamine or bromocriptine, a dopamine 2 receptor (D2R) agonist, caused an inhibition of mitochondrial movement. Moreover, pretreatment with bromocriptine blocked the stimulatory effect of 5-HT on mitochondrial movement. Conversely, in cells pretreated with 5-HT, no further increases in movement were observed after administration of haloperidol, a D2R antagonist. In contrast to the effect of the D2R agonist, addition of SKF38393, a dopamine 1 receptor (D1R) agonist, promoted mitochondrial transport, indicating that the inhibitory effect of dopamine was actually the net summation of opposing influences of the two receptor subtypes. The most pronounced effect of dopamine signals was on mitochondria that were already moving directionally. Western blot analysis revealed that treatment with either a D2R agonist or a D1R antagonist decreased Akt activity, and conversely, treatment with either a D2R antagonist or a D1R agonist increased Akt activity. Conclusions/Significance Our observations strongly suggest a role for both dopamine and 5-HT in regulating mitochondrial movement, and indicate that the integrated effects of these two neuromodulators may be important in determining the distribution of energy sources in neurons.

The homeobox transcription factor Barx2 regulates chondrogenesis during limb development

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.

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

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.

Charting out the octopus connectome at submicron resolution using the knife-edge scanning microscope

The common octopus, or Octopus vulgaris, has the largest nervous system of any invertebrate, and has been shown to possess learning and memory capabilities that in many ways rival those of some vertebrates [1]. Nevertheless, the neural architecture of this cephalopod mollusk differs markedly from that of any vertebrate. Investigating the differences and similarities between the neural architecture—or connectome—of the octopus and mammals, such as the mouse, may lead to deep insights into the computational principles underlying animal cognition. The octopus brain provides some unique advantages for anatomical research, since its axons are generally thick and unmyelinated, allowing traditional staining methods, such as Golgi, to be used effectively. With this in mind, we first imaged the brain using the Knife-Edge Scanning Microscope [2], a custom serial sectioning microscope that can image large blocks of tissue (1 cm3) at sub-micrometer resolution. We imaged large portions of the octopus subesophageal mass (SUB) and the optic lobe (OL) which were stained using Golgi. In order to extract the geometry of the neuronal morphology, we used our Maximum Intensity Projection (MIP)-based tracing algorithm [3]. The imaging results are shown in Figure 1(a-d), and tracing results are shown in 1(e). Although quite preliminary, to our knowledge this is the first time large volumes of the octopus brain have been imaged at sub-micrometer resolution, allowing us to resolve many of the processes that make up the neural network. We expect that this pilot study and the more detailed investigations to follow will allow fruitful comparisons of the neural circuitries of individual octopuses with different ecological life histories, as well as of animals that have been exposed to a variety of neurodegenerative insults. Moreover, such explorations will engender a greater understanding of how functional neural architecture is altered by learning in invertebrates such as the octopus and vertebrates such as the mouse. In sum, this approach should contribute greatly to our understanding of the computational architecture of invertebrates and ultimately provide insights into the differences between invertebrate and vertebrate cognitive capabilities. Figure 1 Octopus subesophageal mass (SUB) and optic lobe (OL) imaged with the KESM (a–d), and tracing results (e). Scale (block width): (a) 1.44 mm, (b) 0.72 mm, (c) 1.44 mm, (d-f) 76.8 μm. Voxel resolution: 0.6 μm x 0.7 μm x 1.0 ...

Neuromorphic and Brain-Based Robots: The case for using brain-based devices to study consciousness

Introduction Within the past few decades, the nature of consciousness has become a central issue in neuroscience, and it is increasingly the focus of both theoretical and empirical work. Studying consciousness is vital to developing an understanding of human perception and behavior, of our relationships with one another, and of our relationships with other potentially conscious animals. Although the study of consciousness through the construction of artificial models is a recent innovation, the advantages of such an approach are clear. First, models allow us to investigate consciousness in ways that are currently not feasible using human subjects or other animals. Second, an artifact that exhibits the necessary and sufficient properties of consciousness may conceivably be the forerunner of a new and very useful class of neuromorphic robots. A model of consciousness must take into account current theories of its biological bases. Although the field of artificial consciousness is a new one, it is striking how little attention has been given to modeling mechanisms. Instead, great – and perhaps undue – emphasis has been placed on purely phenomenological models. Many of these models are strongly reductionist in aim and fail to specify neural mechanisms.

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

In this important and thought-provoking review, the author takes up what is certainly the central challenge to thescientific study of consciousness: namely, the apparent irreducibility of core aspects of subjective experience. Theauthor rightly identifies four core features of consciousness that defy attempts at reduction: the referral of mentalstates, mental unity (taken here to be the same as ‘perceptual unity’), qualia, and mental causation [1, this issue].For the sake of this commentary, I will consider an overarching claim made in the target review about the nature ofreduction in biological systems generally.In framing the problem of irreducibility in the study of consciousness, the author begins with the observation thatall biological systems are hierarchically organized. In such hierarchical systems, certain novel properties are emergentat the highest levels of organization (i.e., properties that differ markedly from the features that collectively generatethem); at the same time, high-level components