This is an RSS newsfeed from BioMed Central It is intended to be used with an RSS reader. For more information about RSS newsfeeds from BioMed Central, visit http://www.biomedcentral.com/info/about/rss/ http://www.molecularbrain.com The latest articles from Molecular Brain (ISSN 1756-6606) published by BioMed Central alpha-synuclein (alpha-syn) is a main component of Lewy bodies (LB) that occur in many neurodegenerative diseases, including Parkinson's disease (PD), dementia with LB (DLB) and multi-system atrophy. alpha-syn mutations or amplifications are responsible for a subset of autosomal dominant familial PD cases, and overexpression causes neurodegeneration and motor disturbances in animals. To investigate mechanisms for alpha-syn accumulation and toxicity, we studied a mouse model of lysosomal enzyme cathepsin D (CD) deficiency, and found extensive accumulation of endogenous alpha-syn in neurons without overabundance of alpha-syn mRNA. In addition to impaired macroautophagy, CD deficiency reduced proteasome activity, suggesting an essential role for lysosomal CD function in regulating multiple proteolytic pathways that are important for alpha-syn metabolism. Conversely, CD overexpression reduces alpha-syn aggregation and is neuroprotective against alpha-syn overexpression-induced cell death in vitro. In a C. elegans model, CD deficiency exacerbates alpha-syn accumulation while its overexpression is protective against alpha-syn-induced dopaminergic neurodegeneration. Mutated CD with diminished enzymatic activity or overexpression of cathepsins B (CB) or L (CL) is not protective in the worm model, indicating a unique requirement for enzymatically active CD. Our data identify a conserved CD function in alpha-syn degradation and identify CD as a novel target for LB disease therapeutics. http://www.molecularbrain.com/content/1/1/17 Liyan Qiao, Shusei Hamamichi, Kim A Caldwell, Guy A Caldwell, Talene A Yacoubian, Scott Wilson, Zuo-Lei Xie, Lisa D Speake, Rachael Parks, Donna Crabtree, Qiuli Liang, Stephen Crimmins, Lonnie Schneider, Yasuo Uchiyama, Takeshi Iwatsubo, Yi Zhou, Lisheng Peng, YouMing Lu, David G Standaert, Ken C Walls, John J Shacka, Kevin A Roth and Jianhua Zhang Molecular Brain 2008, 1:17 2008-11-21 doi:10.1186/1756-6606-1-17 Molecular Brain 1756-6606 1 17 2008-11-21 Background: Phenylethanolamines selectively bind to NR2B subunit-containing N-methyl-D-aspartate-subtype of ionotropic glutamate receptors and negatively modulate receptor activity. To investigate the structural and functional properties of the ifenprodil binding domain on the NR2B protein, we have purified a soluble and functional recombinant rat NR2B protein fragment comprising the first ~400 amino acid amino-terminal domain (ATD2B) expressed in E. coli. Spectral measurements on refolded ATD2B protein demonstrated specific binding to ifenprodil. We have used site-directed mutagenesis, circular dichroism spectroscopy and molecular modeling to obtain structural information on the interactions between critical amino acid residues and ifenprodil of our soluble refolded ATD2B proteins. Ligand-induced changes in protein structure were inferred from changes in the circular dichroism spectrum, and the concentration dependence of these changes was used to determine binding constants for ifenprodil and its analogues. Results: Ligand binding of ifenprodil, RO25,6981 and haloperidol on soluble recombinant ATD2B determined from circular dichroism spectroscopy yielded low-to-high micromolar constants which concurred with functional IC50 measurement determined in heterologously expressed NR1/NR2B receptors in Xenopus oocytes. Amino acid residue substitutions of Asp101, Ile150 and Phe176 with alanine residue within the ATD2B protein altered the recombinant protein dissociation constants for ifenprodil, mirroring the pattern of their functional phenotypes. Molecular modeling of ATD2B as a clam-shell-like structure places these critical residues near a putative ligand binding site. Conclusions: We report for the first time biochemical measurements show that the functional measurements actually reflect binding to the ATD of NR2B subunit. Insights gained from this study help advance the theory that ifenprodil is a ligand for the ATD of NR2B subunit. http://www.molecularbrain.com/content/1/1/16 Fui-Mee Ng, Matthew T Geballe, James P Snyder, Stephen F Traynelis and Chian-Ming Low Molecular Brain 2008, 1:16 2008-11-18 doi:10.1186/1756-6606-1-16 Molecular Brain 1756-6606 1 16 2008-11-18 Background: Deposition of amyloid beta protein (Abeta) is a major pathological hallmark of Alzheimer's disease (AD). Abeta is generated from gamma-secretase cleavage of amyloid precursor protein (APP). In addition to APP, gamma-secretase also cleaves other type I integral membrane proteins, including the Notch receptor, a key molecule involved in embryonic development. Results: To explore selective gamma-secretase inhibitors, a combination of five methods was used to systematically determine these inhibitors' profiles on the gamma-secretase cleavage of APP and Notch. When two potent gamma-secretase inhibitors, compound E (cpd E) and DAPT, were used in a conventional in vitro gamma-secretase activity assay, cpd E completely blocked Abeta generation from the cleavage of substrate APP C100, but only had a minor effect on Notch cleavage and NICD generation. Next, cpd E and DAPT were applied to HEK293 cells expressing a truncated Notch substrate Notch deltaE. Both cpd E and DAPT were more potent in blocking Abeta generation than NICD generation. Third, a reporter construct was created that carried the NICD targeting promoter with three Su(H) binding sequences followed by the luciferase gene. We found that the inhibition of NICD generation by cpd E and DAPT was consistent with the reduced expression of luciferase gene driven by this Notch targeting promoter. Fourth, levels of "Notch-Abeta-like" (Nbeta*) peptide derived from two previously reported chimeric APP with its transmembrane domain or the juxtamembrane portion replaced by the Notch sequence were quantified. Measurement of Nbeta* peptides by ELISA confirmed that EC50's of cpd E were much higher for Nbeta* than Abeta. Finally, the expression levels of Notch target gene her6 in cpd E or DAPT-treated zebrafish were correlated with the degree of tail curvature due to defective somitogenesis, a well characterized Notch phenotype in zebrafish. Conclusion: our ELISA-based quantification of Abeta and Nbeta* in combination with the test in zebrafish provides a novel approach for efficient cell-based screening and in vivo validation of APP selective gamma-secretase inhibitors. http://www.molecularbrain.com/content/1/1/15 Ting Yang, Dilyara Arslanova, Yongli Gu, Corinne Augelli-Szafran and Weiming Xia Molecular Brain 2008, 1:15 2008-11-04 doi:10.1186/1756-6606-1-15 Molecular Brain 1756-6606 1 15 2008-11-04 Background: Ca2+-activated Cl- channels (CaCCs) participate in many important physiological processes. However, the lack of effective and selective blockers has hindered the study of these channels, mostly due to the lack of good assay system. Here, we have developed a reliable drug screening method for better blockers of CaCCs, using the endogeneous CaCCs in Xenopus laevis oocytes and two-electrode voltage-clamp (TEVC) technique. Results: Oocytes were prepared with a treatment of Ca2+ ionophore, which was followed by a treatment of thapsigargin which depletes Ca2+ stores to eliminate any contribution of Ca2+ release. TEVC was performed with micropipette containing chelerythrine to prevent PKC dependent run-up or run-down. Under these conditions, Ca2+-activated Cl- currents induced by bath application of Ca2+ to oocytes showed stable peak amplitude when repetitively activated, allowing us to test several concentrations of a test compound from one oocyte. Inhibitory activities of commercially available blockers and synthesized anthranilic acid derivatives were tested using this method. As a result, newly synthesized N-(4-trifluoromethylphenyl)anthranilic acid with trifluoromethyl group (-CF3) at para position on the benzene ring showed the lowest IC50. Conclusion: Our results provide an optimal drug screening strategy suitable for high throughput screening, and propose N-(4-trifluoromethylphenyl)anthranilic acid as an improved CaCC blocker. http://www.molecularbrain.com/content/1/1/14 Soo-Jin Oh, Jung Hwan Park, Sungyu Han, Jae Kyun Lee, Eun Joo Roh and C Justin Lee Molecular Brain 2008, 1:14 2008-10-29 doi:10.1186/1756-6606-1-14 Molecular Brain 1756-6606 1 14 2008-10-29 Background: Although long-term potentiation (LTP) of synaptic strength is very persistent, current studies have provided evidence that various manipulations or pharmacological treatment when applied shortly after LTP induction can reverse it. This kind of reversal of synaptic strength is termed as depotentiation and may have a function to increase the flexibility and storage capacity of neuronal networks. Our previous studies have demonstrated that an increase in extracellular levels of adenosine and subsequent activation of adenosine A1 receptors are important for the induction of depotentiation; however, the signaling downstream of adenosine A1 receptors to mediate depotentiation induction remains elusive. Results: We confirm that depotentiation induced by low-frequency stimulation (LFS) (2 Hz, 10 min, 1200 pulses) was dependent on adenosine A1 receptor activation, because it was mimicked by bath-applied adenosine A1 receptor agonist N6-cyclopentyladenosine (CPA) and was inhibited by the selective adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). Pretreatment of the hippocampal slices with the selective p38 mitogen-activated protein kinase (MAPK) inhibitors, 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl]-5-(4-pyrudyl)-1H-imidazole (SB203580) or trans-1-(4-hydroxycyclohexyl)-4-(fluorophenyl)-5-(2-methoxypyrimidin-4-yl)imidazole (SB239063), prevented the induction of depotentiation by LFS and CPA. In agreement with electrophysiological observation, both LFS- and CPA-induced depotentiation are associated with an increase in p38 MAPK activation, which are blocked by DPCPX or SB203580 application. Conclusion: These results suggest that activation of adenosine A1 receptor and in turn triggering p38 MAPK signaling may contribute to the LFS-induced depotentiation at hippocampal CA1 synapses. http://www.molecularbrain.com/content/1/1/13 Ying-Ching Liang, Chiung-Chun Huang and Kuei-Sen Hsu Molecular Brain 2008, 1:13 2008-10-23 doi:10.1186/1756-6606-1-13 Molecular Brain 1756-6606 1 13 2008-10-23 Background: In previous work, we investigated dieldrin cytotoxicity and signaling cell death mechanisms in dopaminergic PC12 cells. Dieldrin has been reported to be one of the environmental factors correlated with Parkinson's disease and may selectively destroy dopaminergic neurons. Methods: Here we further investigated dieldrin toxicity in a dopaminergic neuronal cell model of Parkinson's disease, namely N27 cells, using biochemical, immunochemical, and flow cytometric analyses. Results: In this study, dieldrin-treated N27 cells underwent a rapid and significant increase in reactive oxygen species followed by cytochrome c release into cytosol. The cytosolic cytochrome c activated caspase-dependent apoptotic pathway and the increased caspase-3 activity was observed following a 3 hr dieldrin exposure in a dose-dependent manner. Furthermore, dieldrin caused the caspase-dependent proteolytic cleavage of protein kinase C delta (PKCδ) into 41 kDa catalytic and 38 kDa regulatory subunits in N27 cells as well as in brain slices. PKCδ plays a critical role in executing the apoptotic process in dieldrin-treated dopaminergic neuronal cells because pretreatment with the PKCδ inhibitor rottlerin, or transfection and over-expression of catalytically inactive PKCδK376R, significantly attenuates dieldrin-induced DNA fragmentation and chromatin condensation. Conclusion: Together, we conclude that caspase-3-dependent proteolytic activation of PKCδ is a critical event in dieldrin-induced apoptotic cell death in dopaminergic neuronal cells. http://www.molecularbrain.com/content/1/1/12 Anumantha G Kanthasamy, Masashi Kitazawa, Yongjie Yang, Vellareddy Anantharam and Arthi Kanthasamy Molecular Brain 2008, 1:12 2008-10-22 doi:10.1186/1756-6606-1-12 Molecular Brain 1756-6606 1 12 2008-10-22 Background: Schizophrenia is a complex genetic disorder caused by multiple genetic and environmental factors. The dystrobrevin-binding protein 1 (DTNBP1: dysbindin-1) gene is a major susceptibility gene for schizophrenia. Genetic variations in DTNBP1 are associated with cognitive functions, general cognitive ability and memory function, and clinical features of patients with schizophrenia including negative symptoms and cognitive decline. Since reduced expression of dysbindin-1 has been observed in postmortem brains of patients with schizophrenia, the sandy (sdy) mouse, which has a deletion in the Dtnbp1 gene and expresses no dysbindin-1 protein, could be an animal model of schizophrenia. To address this issue, we have carried out a comprehensive behavioral analysis of the sdy mouse in this study. Results: In a rotarod test, sdy mice did not exhibit motor learning whilst the wild type mice did. In a Barnes circular maze test both sdy mice and wild type mice learned to selectively locate the escape hole during the course of the training period and in the probe trial conducted 24 hours after last training. However, sdy mice did not locate the correct hole in the retention probe tests 7 days after the last training trial, whereas wild type mice did, indicating impaired long-term memory retention. A T-maze forced alternation task, a task of working memory, revealed no effect of training in sdy mice despite the obvious effect of training in wild type mice, suggesting a working memory deficit. Conclusion: Sdy mouse showed impaired long-term memory retention and working memory. Since genetic variation in DTNBP1 is associated with both schizophrenia and memory function, and memory function is compromised in patients with schizophrenia, the sdy mouse may represent a useful animal model to investigate the mechanisms of memory dysfunction in the disorder. http://www.molecularbrain.com/content/1/1/11 Keizo Takao, Keiko Toyama, Kazuo Nakanishi, Satoko Hattori, Hironori Takamura, Masatoshi Takeda, Tsuyoshi Miyakawa and Ryota Hashimoto Molecular Brain 2008, 1:11 2008-10-22 doi:10.1186/1756-6606-1-11 Molecular Brain 1756-6606 1 11 2008-10-22 Several myelin-associated factors that inhibit axon growth of mature neurons, including Nogo66, myelin-associated glycoprotein (MAG) and oligodendrocyte myelin glycoprotein (OMgp), can associate with a common GPI-linked protein Nogo-66 receptor (NgR). Accumulating evidence suggests that myelin inhibitors also signal through unknown NgR-independent mechanisms. Here we show that MAG, a RGD tri-peptide containing protein, forms a complex with β1-integrin to mediate axonal growth cone turning responses of several neuronal types. Mutations that alter the RGD motif in MAG or inhibition of β1-integrin function, but not removal of NgRs, abolish these MAG-dependent events. In contrast, OMgp-induced repulsion is not affected by inhibition of b1-integrin function. We further show that MAG stimulates tyrosine phosphorylation of focal adhesion kinase (FAK), which in turn is required for MAG-induced growth cone turning. These studies identify β1-integrin as a specific mediator for MAG in growth cone turning responses, acting through FAK activation. http://www.molecularbrain.com/content/1/1/10 Eyleen LK Goh, Ju Kim Young, Kenichiro Kuwako, Marc Tessier-Lavigne, Zhigang He, John W Griffin and Guo-li Ming Molecular Brain 2008, 1:10 2008-10-15 doi:10.1186/1756-6606-1-10 Molecular Brain 1756-6606 1 10 2008-10-15 Long-term potentiation (LTP) in the hippocampal CA1 region requires the activation of N-methyl-D-aspartate receptors (NMDARs). Studies using genetic and pharmacological approaches have reported inconsistent results of the requirement of NR2B-containing NMDARs in LTP in the CA1 region. Pharmacological studies showed that NR2B-containing NMDARs are not required for LTP, while genetic studies reported that over-expression of NR2B-NMDARs enhances LTP and hippocampus-dependent memory. Here, we provide evidence showing that the functional role of NR2B-NMDARs in hippocampal LTP and memory depends on LTP-inducing and behavior-conditioning protocols. Inhibition of NR2B-NMDARs with the NR2B selective antagonist ifenprodil or Ro25-6981 suppressed LTP induced by spike-timing protocol, with no impact on LTP induced by pairing protocol or two-train high-frequency stimulation (HFS) protocol. Inhibition of NR2B-NMDARs did not affect the late phase LTP induced by four-train HFS. Ca2+ imaging showed that there was difference in kinetics of intracellular Ca2+ signals induced by spiking-timing and pairing protocols. Pre-training intra-CA1 infusion of ifenprodil or Ro25-6981 impaired the contextual fear memory induced by five CS-US pairings, with no effect on the memory induced by one CS-US pairing. http://www.molecularbrain.com/content/1/1/9 Xue-Han Zhang, Long-Jun Wu, Bo Gong, Ming Ren, Bao-Ming Li and Min Zhuo Molecular Brain 2008, 1:9 2008-09-30 doi:10.1186/1756-6606-1-9 Molecular Brain 1756-6606 1 9 2008-09-30 The midbrain dopaminergic (mDA) neurons of the substantia nigra and the ventral tegmental area play a fundamental role in the control of voluntary movement and the regulation of emotion, and are severely affected in Parkinson's disease. Recent advances in mouse genetics and vertebrate development have provided us with insight into the genetic cascades involved in the development of mDA neurons, including the induction of mDA neuron progenitors in the ventral mesencephalon, the specification of the mDA neuronal fate and the maintenance of postmitotic mDA neurons. In parallel, rapid progress has been made in the generation of DA neurons from pluripotent stem cells and the development of stem cell-based therapies for Parkinson's disease. Here, we summarize the new findings via the developmental progression of mDA neurons and outline how this knowledge has been exploited to develop novel paradigms for the in vitro generation of these neurons from embryonic stem cells. http://www.molecularbrain.com/content/1/1/8 Emily Gale and Meng Li Molecular Brain 2008, 1:8 2008-09-30 doi:10.1186/1756-6606-1-8 Molecular Brain 1756-6606 1 8 2008-09-30 Background: Calpastatin is an endogenous inhibitor of calpain, intracellular calcium-activated protease. It has been suggested to be involved in molecular mechanisms of long-term plasticity and excitotoxic pathways. However, functions of calpastatin in vivo are still largely unknown. To examine the physiological roles of calpastatin, we subjected calpastatin-knockout mice to a comprehensive behavioral test battery. Results: Calpastatin-knockout mice showed decreased locomotor activity under stressful environments, and decreased acoustic startle response, but we observed no significant change in hippocampus-dependent memory function. Conclusion: These results suggest that calpastatin is likely to be more closely associated with affective rather than cognitive aspects of brain function. http://www.molecularbrain.com/content/1/1/7 Ryuichi Nakajima, Keizo Takao, Shu-Ming Huang, Jiro Takano, Nobuhisa Iwata, Tsuyoshi Miyakawa and Takaomi C Saido Molecular Brain 2008, 1:7 2008-09-15 doi:10.1186/1756-6606-1-7 Molecular Brain 1756-6606 1 7 2008-09-15