Neuronal Ndrg4 Is Essential for Nodes of Ranvier Organization in Zebrafish.

Axon ensheathment by specialized glial cells is an important process for fast propagation of action potentials. The rapid electrical conduction along myelinated axons is mainly due to its saltatory nature characterized by the accumulation of ion channels at the nodes of Ranvier. However, how these ion channels are transported and anchored along axons is not fully understood. We have identified N-myc downstream-regulated gene 4, ndrg4, as a novel factor that regulates sodium channel clustering in zebrafish. Analysis of chimeric larvae indicates that ndrg4 functions autonomously within neurons for sodium channel clustering at the nodes. Molecular analysis of ndrg4 mutants shows that expression of snap25 and nsf are sharply decreased, revealing a role of ndrg4 in controlling vesicle exocytosis. This uncovers a previously unknown function of ndrg4 in regulating vesicle docking and nodes of Ranvier organization, at least through its ability to finely tune the expression of the t-SNARE/NSF machinery.

Identification of differentially expressed proteins and validation of the changes of N-ethylmaleimide-sensitive factor in rats with focal cerebral ischemia after transection of the cervical sympathetic trunk.

Stellate ganglion blockade (SGB) protects patients from focal cerebral ischemic injury, and transection of the cervical sympathetic trunk (TCST) in a rat model can mimic SGB in humans. The purpose of this study was to investigate the mechanisms underlying the neuroprotective effects of TCST on neuronal damage in the hippocampus in a rat model of middle cerebral artery occlusion (MCAO) in an attempt to elucidate the neuroprotective effects of SGB. The modified method of Zea Longa was used to establish the permanent MCAO model. Male Wistar rats were randomly divided into three groups: sham-operated group, MCAO group, and TCST group. The animals in TCST group were sacrificed 48 h after TCST which was performed after the establishment of the MCAO model. Proteins were extracted from the ipsilateral hippocampus and analyzed by two-dimensional difference gel electrophoresis (2D-DIGE) and peptide mass fingerprinting (PMF). The levels of N-ethylmaleimide-sensitive factor (NSF) were measured as well. The results showed that 11 types of proteins were identified by 2D-DIGE. The expressions of eight proteins were changed both in the sham-operated and TCST groups, and the expressions of the other three proteins were changed in all three groups. Moreover, the expression of NSF was higher in the TCST group than in the MCAO group but lower in the MCAO group than in sham-operated group. The ratio of NSF expression between the MCAO group and shamoperated group was -1.37 (P<0.05), whereas that between the TCST group and MCAO group was 1.35 (P<0.05). Our results imply that TCST increases the expression of NSF in the hippocampus of adult rats with focal cerebral ischemia, which may contribute to the protection of the injured brain. Our study provides a theoretical basis for the therapeutic application of SGB to patients with permanent cerebral ischemia.

Interactions between N-ethylmaleimide-sensitive factor and GluR2 in the nucleus accumbens contribute to the expression of locomotor sensitization to cocaine.

Many studies have reported a withdrawal-dependent increase in synaptic AMPA receptor (AMPAR) levels in the nucleus accumbens (NAc) of cocaine-sensitized rats; however, the exact relationship between the expression of sensitization and altered AMPAR surface expression in the NAc has not yet been investigated. We demonstrated that the expression of behavioral sensitization was negatively controlled by N-ethylmaleimide-sensitive factor (NSF)-GluR2 interactions in the NAc. The upregulation of NSF-GluR2 interactions, which may be resulted by the increase in NSF S-nitrosylation after withdrawal from cocaine, was associated with the changes in the expression of behavioral sensitization. Disruption of NSF-GluR2 interactions in the NAc with a specific peptide, TAT-pep-R845A, increased the locomotor response of rats to cocaine by decreasing GluR2 surface insertion. In contrast, prevention of GluR2-containing AMPARs removal from synapses with Pep2-EVKI attenuated the expression of behavioral sensitization. Similarly, treatment with the nitric oxide donor, S-Nitroso-N-acetyl-DL-penicillamine (SNAP), attenuated the expression of locomotor sensitization by promoting GluR2 surface expression. This effect was mediated by the binding of S-nitrosylated NSF to GluR2, which promoted the surface expression of AMPARs. Noticeably, exogenous injection of SNAP into NAc also attenuated the expression of cocaine-induced conditioned place preference. Thus, these results indicate that increased NSF-GluR2 interactions in the NAc after withdrawal from cocaine attenuated the expression of behavioral sensitization and serves as a negative regulatory mechanism in drug-exposed individuals.

Proteomic profiling of the hypothalamus in a mouse model of cancer-induced anorexia-cachexia.

BACKGROUND: Anorexia-cachexia is a common and severe cancer-related complication but the underlying mechanisms are largely unknown. Here, using a mouse model for tumour-induced anorexia-cachexia, we screened for proteins that are differentially expressed in the hypothalamus, the brain's metabolic control centre. METHODS: The hypothalamus of tumour-bearing mice with implanted methylcholanthrene-induced sarcoma (MCG 101) displaying anorexia and their sham-implanted pair-fed or free-fed littermates was examined using two-dimensional electrophoresis (2-DE)-based comparative proteomics. Differentially expressed proteins were identified by liquid chromatography-tandem mass spectrometry. RESULTS: The 2-DE data showed an increased expression of dynamin 1, hexokinase, pyruvate carboxylase, oxoglutarate dehydrogenase, and N-ethylmaleimide-sensitive factor in tumour-bearing mice, whereas heat-shock 70 kDa cognate protein, selenium-binding protein 1, and guanine nucleotide-binding protein Gα0 were downregulated. The expression of several of the identified proteins was similarly altered also in the caloric-restricted pair-fed mice, suggesting an involvement of these proteins in brain metabolic adaptation to restricted nutrient availability. However, the expression of dynamin 1, which is required for receptor internalisation, and of hexokinase, and pyruvate carboxylase were specifically changed in tumour-bearing mice with anorexia. CONCLUSION: The identified differentially expressed proteins may be new candidate molecules involved in the pathophysiology of tumour-induced anorexia-cachexia.

Molecular characterization and functional distribution of N-ethylmaleimide-sensitive factor in Helicoverpa armigera.

N-ethylmaleimide-sensitive fusion protein (NSF) is an essential component for the neurotransmitter or neurohormone release apparatus present in all eukaryotic cells. Here, a new NSF orthologue was characterized from the cotton bollworm, Helicoverpa armigera (Har). Northern blot exhibited a high expression in larval brain. Southern analysis indicated that a single copy of the gene is present in a haploid genome. Using antibodies labeled with fluoresceins, we directly proved that NSF is co-localized with two crucial neurohormones, prothoracicotropic hormone and diapause hormone, both of which regulate insect development. These findings suggest that Har-NSF may be involved in regulating insect neurohormone release.