p63 Regulates adult neural precursor and newly born neuron survival to control hippocampal-dependent Behavior.

The molecular mechanisms that regulate adult neural precursor cell (NPC) survival, and thus maintain adult neurogenesis, are not well defined. Here, we investigate the role of p63, a p53 family member, in adult NPC function in mice. Conditional ablation of p63 in adult NPCs or p63 haploinsufficiency led to reduced numbers of NPCs and newborn neurons in the neurogenic zones of the hippocampus and lateral ventricles and in the olfactory bulb. These reductions were attributable to enhanced apoptosis of NPCs and newborn neurons and were rescued by inhibition of caspase activity, p53, or the p53 apoptotic effector PUMA (p53-upregulated modulator of apoptosis). Moreover, these cellular deficits were functionally important because they led to perturbations in hippocampus-dependent memory formation. These results indicate that p63 regulates the numbers of adult NPCs and adult-born neurons as well as neural stem cell-dependent cognitive functions, and that it does so, at least in part, by inhibiting p53-dependent cell death.

Coffin-Lowry syndrome: a role for RSK2 in mammalian neurogenesis.

Coffin-Lowry Syndrome (CLS) is an X-linked genetic disorder associated with cognitive and behavioural impairments. CLS patients present with loss-of-function mutations in the RPS6KA3 gene encoding the mitogen-activated protein kinase (MAPK)-activated kinase p90 ribosomal S6 kinase 2 (Rsk2). Although Rsk2 is expressed in the embryonic brain, its function remains largely uncharacterized. To this end, we isolated murine cortical precursors at embryonic day 12 (E12), a timepoint when neuronal differentiation is initiated, and knocked-down Rsk2 expression levels using shRNA. We performed similar experiments in vivo using in utero electroporations to express shRNA against Rsk2. Rsk2 knockdown resulted in a significant decrease in neurogenesis and an increase in the proportion of proliferating Pax6-positive radial precursor cells, indicating that Rsk2 is essential for cortical radial precursors to differentiate into neurons. In contrast, reducing Rsk2 levels in vitro or in vivo had no effect on the generation of astrocytes. Thus, Rsk2 loss-of-function, as seen in CLS, perturbs the differentiation of neural precursors into neurons, and maintains them instead as proliferating radial precursor cells, a defect that may underlie the cognitive dysfunction seen in CLS.

p63 antagonizes p53 to promote the survival of embryonic neural precursor cells.

The molecular mechanisms that regulate survival of embryonic neural precursors are still relatively ill-defined. Here, we have asked whether the p53 family member p63 plays any role during this developmental window, focusing on the embryonic cerebral cortex. We show that genetic knockdown of p63 either in culture or in the embryonic telencephalon causes apoptosis of cortical precursors and newly born cortical neurons, and that this can be rescued by expression of DeltaNp63, but not TAp63 isoforms. This cortical precursor apoptosis is the consequence of deregulated p53 activity, since both basal precursor apoptosis and that induced by loss of p63 are rescued by coincident genetic silencing of p53. Finally, we demonstrate that the third p53 family member, DeltaNp73, does not regulate survival of cortical precursor cells, but that it collaborates with DeltaNp63 to ensure the survival of newly born cortical neurons. Thus, the balance of DeltaNp63 versus p53 determines the life versus death of embryonic cortical precursors, a role that these p53 family members may well play in other populations of developing and/or adult neural precursors.

Selective regulation of the perinuclear distribution of glucose transporter 4 (GLUT4) by insulin signals in muscle cells.

Insulin regulates glucose transporter 4 (GLUT4) availability at the surface of muscle and adipose cells. In L6 myoblasts, stably expressed GLUT4myc is detected mostly in a perinuclear region. In unstimulated cells, about half of perinuclear GLUT4myc colocalizes with the transferrin receptor (TfR). Insulin stimulation selectively decreased the perinuclear colocalization of GLUT4myc with TfR determined by 3D-reconstruction of fluorescence images. Perinuclear GLUT4myc adopted two main distributions defined morphometrically as 'conical' and 'concentric'. Insulin rapidly reduced the proportion of cells with conical in favor of concentric perinuclear GLUT4myc distributions in association with the gain in surface GLUT4myc. Upon removal of insulin, the GLUT4myc perinuclear distribution and surface levels reversed in parallel. In contrast, hypertonicity (which like insulin elevates surface GLUT4myc) did not elicit perinuclear GLUT4myc redistribution. Insulin also caused redistribution of perinuclear vesicle-associated membrane protein-2 (VAMP2), without alteration of perinuclear TfR and VAMP3. Inhibitory mutants of phosphatidylinositol-3 kinase (Deltap85) or Akt substrate AS160 (AS160-4P) prevented insulin-mediated perinuclear GLUT4myc redistribution. Tetanus toxin expression did not prevent the perinuclear GLUT4myc redistribution, suggesting that redistribution is independent of GLUT4myc fusion with the plasma membrane. We propose that insulin causes selective, dynamic relocalization of perinuclear GLUT4myc and VAMP2 and perinuclear GLUT4myc redistribution is a direct target of insulin-derived signals.

Might astrocytes play a role in maintaining the seizure-prone state?

The amygdala-kindling model is used to study complex partial epilepsy with secondary generalization. The present study was designed to (A) quantify astrocytic changes in the piriform cortex of amygdala-kindled subjects over time and (B) investigate the role that astrocytes might play in maintaining the seizure-prone state. In Study A, once the experimental subjects reached five stage 5 seizures, stimulation was stopped, and both kindled and control rats were allowed to survive for the interval appropriate to their group (7, 18, 30, or 90 days). Following each interval, the kindled and control animals were given 10 intraperitoneal injections of bromodeoxyuridine (BrdU) and sacrificed 24 h following the last injection. Significantly higher numbers of dividing astrocytes (identified by co-labeling for BrdU and to one of the astrocytic intermediate filament proteins glial fibrillary acidic protein or vimentin) were found in the kindled brains. All kindled groups had significantly higher numbers of double-labeled cells on the side contralateral to the stimulation site, except for those in the 90 day survival group. In Study B, rats were implanted with chemotrodes, were kindled as in Study A, and were subsequently infused with either saline or with L alpha-AA (to lesion astrocytes) during a further 25 stimulations (1/day). L alpha-AA infused rats had significantly diminished levels of behavioral seizures, higher after discharge thresholds, lower after discharge durations, and decreased numbers of double-labeled astrocytes in piriform cortex than did saline infused rats. Together, the data indicate that astrocytes may play a role in maintaining the seizure-prone state.

Astrocytic proliferation in the piriform cortex of amygdala-kindled subjects: a quantitative study in partial versus fully kindled brains.

Complex partial epilepsy is a seizure disorder in which attacks frequently arise from foci located in the temporal lobes. The amygdala-kindling model is a widely used model of complex partial epilepsy with secondary generalization. The present study was designed to quantitatively assess astrocytic changes in the rat piriform cortex in the amygdala-kindling model of epilepsy. Bromodeoxyuridine-injected subjects were sacrificed 24 h after the first stage 1 or fifth stage 5 seizure. Brain sections were prepared and examined quantitatively. A significantly higher number of dividing astrocytes (identified by co-labeling with antibodies to bromodeoxyuridine and to one of the astrocytic intermediate filament proteins glial fibrillary acidic protein or vimentin) was found in both partially kindled (stage 1) and fully kindled (stage 5) brains. The partially kindled brains had a significantly higher number of double-labeled cells on the side ipsilateral to stimulation. The opposite trend was observed in the fully kindled brains. Differences between the ipsilateral and contralateral sides of the kindled brain may suggest different role(s) for astrocytes in the development and progression of the seizure-prone state.

Assessing the long-range transport potential of polybrominated diphenyl ethers: a comparison of four multimedia models.

Data from a comprehensive literature search of environmentally relevant physical-chemical properties for nine polybrominated diphenyl ethers (PBDE), ranging from a monobrominated congener to the fully brominated decabromodiphenyl ether, were evaluated and adjusted to achieve both internal and interhomologue consistency. These data were then used in four model-based long-range transport potential (LRTP) assessment methods. The models TaPL3-2.10, ELPOS-1.1.1, Chemrange-2, and Globo-POP-1.1 were found to yield comparable predictions. A comparison of the LRTP estimates for the PBDEs with those of benchmark chemicals (polychlorinated biphenyls [PCBs]) suggest that the lower-brominated congeners have a LRTP comparable to that of PCBs known to be subject to significant LRT, whereas the highly brominated congeners have a very low potential to reach remote areas. This is in agreement with field measurements in remote regions that indicate that the lighter components of commercially produced PBDE mixtures predominate. Deviations between Chemrange and the models based on the concept of a characteristic travel distance were due to differences in the assumed height of the air compartment, which influences the relative importance of atmospheric degradation and deposition processes. The three models assuming a uniform temperature of 25 degrees C may underestimate the LRTP of the smaller congeners. Only atmospheric parameters had a notable influence on the LRTP estimates by TaPL3, ELPOS, and Chemrange. whereas the relative enrichment of chemicals in the Arctic calculated by Globo-POP is additionally sensitive to the parameters related to the interaction of temperature with air-surface exchange and degradation in surface compartments.

TAp73 acts via the bHLH Hey2 to promote long-term maintenance of neural precursors.

Increasing evidence suggests that deficits in adult stem cell maintenance cause aberrant tissue repair and premature aging [1]. While the mechanisms regulating stem cell longevity are largely unknown, recent studies have implicated p53 and its family member p63. Both proteins regulate organismal aging [2-4] as well as survival and self-renewal of tissue stem cells [5-9]. Intriguingly, haploinsufficiency for a third family member, p73, causes age-related neurodegeneration [10]. While this phenotype is at least partially due to loss of the ΔNp73 isoform, a potent neuronal prosurvival protein [11-16], a recent study showed that mice lacking the other p73 isoform, TAp73, have perturbations in the hippocampal dentate gyrus [17], a major neurogenic site in the adult brain. These findings, and the link between the p53 family, stem cells, and aging, suggest that TAp73 might play a previously unanticipated role in maintenance of neural stem cells. Here, we have tested this hypothesis and show that TAp73 ensures normal adult neurogenesis by promoting the long-term maintenance of neural stem cells. Moreover, we show that TAp73 does this by transcriptionally regulating the bHLH Hey2, which itself promotes neural precursor maintenance by preventing premature differentiation.

Glucose transporter 4: cycling, compartments and controversies.

Insulin promotes glucose uptake into muscle and adipose tissues through glucose transporter 4 (GLUT4). In unstimulated cells, rapid endocytosis, slow exocytosis and dynamic or static retention cause GLUT4 to concentrate in early recycling endosomes, the trans-Golgi network and vesicle-associated protein 2-containing vesicles. The coordinated action of phosphatidylinositol 3-kinase effectors, protein kinase Akt, atypical protein kinase C (aPKC) and Akt substrate of 160-kDa (AS160), regulates the GLUT4 cycle by affecting its translocation, fusion with the plasma membrane, internalization and sorting. We review the evidence that supports such cycling, evaluate current models proposing static or dynamic retention, and highlight how distinct steps of GLUT4 transport are regulated by insulin signals. In particular, fusion seems to be regulated by aPKC (via munc18) and Akt (via syntaxin4-interacting protein (synip)). AS160 participates in GLUT4 intracellular retention, and possibly fusion, through candidate ras-related GTP-binding protein (Rab)2, Rab8, Rab10 and/or Rab14. The localization of the insulin-sensitive GLUT4 compartment and the precise target of insulin-derived signals remain open for future investigation.

Turning signals on and off: GLUT4 traffic in the insulin-signaling highway.

Insulin stimulation of glucose uptake into skeletal muscle and adipose tissues is achieved by accelerating glucose transporter GLUT4 exocytosis from intracellular compartments to the plasma membrane and minimally reducing its endocytosis. The round trip of GLUT4 is intricately regulated by diverse signaling molecules impinging on specific compartments. Here we highlight the key molecular signals that are turned on and off by insulin to accomplish this task.