The oDGal Mouse: A Novel, Physiologically Relevant Rodent Model of Sporadic Alzheimer's Disease.

Sporadic Alzheimer's disease (sAD) represents a serious and growing worldwide economic and healthcare burden. Almost 95% of current AD patients are associated with sAD as opposed to patients presenting with well-characterized genetic mutations that lead to AD predisposition, i.e., familial AD (fAD). Presently, the use of transgenic (Tg) animals overexpressing human versions of these causative fAD genes represents the dominant research model for AD therapeutic development. As significant differences in etiology exist between sAD and fAD, it is perhaps more appropriate to develop novel, more sAD-reminiscent experimental models that would expedite the discovery of effective therapies for the majority of AD patients. Here we present the oDGal mouse model, a novel model of sAD that displays a range of AD-like pathologies as well as multiple cognitive deficits reminiscent of AD symptomology. Hippocampal cognitive impairment and pathology were delayed with N-acetyl-cysteine (NaC) treatment, which strongly suggests that reactive oxygen species (ROS) are the drivers of downstream pathologies such as elevated amyloid beta and hyperphosphorylated tau. These features demonstrate a desired pathophenotype that distinguishes our model from current transgenic rodent AD models. A preclinical model that presents a phenotype of non-genetic AD-like pathologies and cognitive deficits would benefit the sAD field, particularly when translating therapeutics from the preclinical to the clinical phase.

A dual druggable genome-wide siRNA and compound library screening approach identifies modulators of parkin recruitment to mitochondria.

Genetic and biochemical evidence points to an association between mitochondrial dysfunction and Parkinson's disease (PD). PD-associated mutations in several genes have been identified and include those encoding PTEN-induced putative kinase 1 (PINK1) and parkin. To identify genes, pathways, and pharmacological targets that modulate the clearance of damaged or old mitochondria (mitophagy), here we developed a high-content imaging-based assay of parkin recruitment to mitochondria and screened both a druggable genome-wide siRNA library and a small neuroactive compound library. We used a multiparameter principal component analysis and an unbiased parameter-agnostic machine-learning approach to analyze the siRNA-based screening data. The hits identified in this analysis included specific genes of the ubiquitin proteasome system, and inhibition of ubiquitin-conjugating enzyme 2 N (UBE2N) with a specific antagonist, Bay 11-7082, indicated that UBE2N modulates parkin recruitment and downstream events in the mitophagy pathway. Screening of the compound library identified kenpaullone, an inhibitor of cyclin-dependent kinases and glycogen synthase kinase 3, as a modulator of parkin recruitment. Validation studies revealed that kenpaullone augments the mitochondrial network and protects against the complex I inhibitor MPP+. Finally, we used a microfluidics platform to assess the timing of parkin recruitment to depolarized mitochondria and its modulation by kenpaullone in real time and with single-cell resolution. We demonstrate that the high-content imaging-based assay presented here is suitable for both genetic and pharmacological screening approaches, and we also provide evidence that pharmacological compounds modulate PINK1-dependent parkin recruitment.

Transcription and pathway analysis of the superior temporal cortex and anterior prefrontal cortex in schizophrenia.

The molecular basis of schizophrenia is poorly understood; however, different brain regions are believed to play distinct roles in disease symptomology. We have studied gene expression in the superior temporal cortex (Brodmann area 22; BA22), which may play a role in positive pathophysiology, and compared our results with data from the anterior prefrontal cortex (BA10), which shows evidence for a role in negative symptoms. Genome-wide mRNA expression was determined in the BA22 region in 23 schizophrenics and 19 controls and compared with a BA10 data set from the same subjects. After adjustments for confounding sources of variation, we carried out GeneGO pathway enrichment analysis in each region. Significant differences were seen in age-related transcriptional changes between the BA22 and the BA10 regions, 21.8% and 41.4% of disease-associated transcripts showing age association, respectively. After removing age associated changes from our data, we saw the highest enrichment in processes mediating cell adhesion, synaptic contact, cytoskeletal remodelling, and apoptosis in the BA22 region. For the BA10 region, we observed the strongest changes in reproductive signalling, tissue remodelling, and cell differentiation. Further exploratory analysis also identified potentially disease-relevant processes that were undetected in our more stringent primary analysis, including autophagy in the BA22 region and the amyloid process in the BA10 region. Collectively, our analysis suggests disruption of many common pathways and processes underpinning synaptic plasticity in both regions in schizophrenia, whereas individual regions emphasize changes in certain pathways that may help to highlight pathway-specific therapeutic opportunities to treat negative or positive symptoms of the disease.

ADAMTSL3 as a candidate gene for schizophrenia: gene sequencing and ultra-high density association analysis by imputation.

We previously reported an association with a putative functional variant in the ADAMTSL3 gene, just below genome-wide significance in a genome-wide association study of schizophrenia. As variants impacting the function of ADAMTSL3 (a disintegrin-like and metalloprotease domain with thrombospondin type I motifs-like-3) could illuminate a novel disease mechanism and a potentially specific target, we have used complementary approaches to further evaluate the association. We imputed genotypes and performed high density association analysis using data from the HapMap and 1000 genomes projects. To review all variants that could potentially cause the association, and to identify additional possible pathogenic rare variants, we sequenced ADAMTSL3 in 92 schizophrenics. A total of 71 ADAMTSL3 variants were identified by sequencing, many were also seen in the 1000 genomes data, but 26 were novel. None of the variants identified by re-sequencing was in strong linkage disequilibrium (LD) with the associated markers. Imputation analysis refined association between ADAMTSL3 and schizophrenia, and highlighted additional common variants with similar levels of association. We evaluated the functional consequences of all variants identified by sequencing, or showing direct or imputed association. The strongest evidence for function remained with the originally associated variant, rs950169, suggesting that this variant may be causal of the association. Rare variants were also identified with possible functional impact. Our study confirms ADAMTSL3 as a candidate for further investigation in schizophrenia, using the variants identified here. The utility of imputation analysis is demonstrated, and we recommend wider use of this method to re-evaluate the existing canon of suggestive schizophrenia associations.

Abnormalities in α/ß-CaMKII and related mechanisms suggest synaptic dysfunction in hippocampus of LPA1 receptor knockout mice.

Lysophosphatidic acid (LPA) is a natural lysophospholipid that regulates neuronal maturation. In mice, the deletion of the LPA1 receptor causes some phenotypic defects partly overlapping with those found in schizophrenia. In this study, we identified molecular abnormalities in hippocampal synaptic mechanisms involved in glutamatergic neurotransmission, which allow further characterization of synaptic aberrations in LPA1 knockout (KO) mice. At the synaptic level, we found dysregulation of Ca2+/calmodulin (CaM)-dependent kinase II (CaMKII) activity and phosphorylation, with markedly higher Ca2+-dependent kinase activity, probably related to increased expression levels of the ß isoform of CaMKII. Conversely, although the synaptic Ca2+-independent activity of the enzyme was unchanged, autophosphorylation levels of both α and ß isoforms were significantly increased in LPA1 KO mice. Moreover, in LPA1 KO mice the α/ß isoform ratio of CaMKII, which plays a key role in neuronal maturation during development, was markedly decreased, as found previously in schizophrenia patients. At post-synaptic level, LPA1 KO mice showed changes in expression, phosphorylation and interactions of NMDA and AMPA receptor subunits that are consistent with basal strengthening of glutamatergic synapses. However, we measured a reduction of nuclear cAMP responsive element-binding protein phosphorylation, suggesting that activation of the NMDA receptor does not occur at the intracellular signalling level. At the presynaptic level, in line with previous evidence from schizophrenia patients and animal models of pathology, LPA1 KO mice showed accumulation of SNARE protein complexes. This study shows that CaMKII and related synaptic mechanisms at glutamatergic synapses are strongly dysregulated in LPA1 KO mice.

Investigation of the H(+)-myo-inositol transporter (HMIT) as a neuronal regulator of phosphoinositide signalling.

Phosphoinositide signalling regulates a series of important neuronal processes that are thought to be altered in mood disorders. Furthermore, mood-stabilizing drugs inhibit key enzymes that regulate phosphoinositide production and alter neuronal growth cone morphology in an inositol-reversible manner. Inositol is taken up by neurons from the extracellular fluid, presumably via membrane transporters; it can also be synthesized by the enzyme MIP-synthase (myo-inositol-1-phosphate synthase) and, in addition, it is generated by inositol phospholipid hydrolysis. The neuronal-specific HMIT (H(+)-myo-inositol transporter) represents a potential regulator of inositol signalling in neurons that warrants further investigation.

Analyzing the effects of psychotropic drugs on metabolite profiles in rat brain using 1H NMR spectroscopy.

The mechanism of action of standard drug treatments for psychiatric disorders remains fundamentally unknown, despite intensive investigation in academia and the pharmaceutical industry. So far, little is known about the effects of psychotropic medications on brain metabolism in either humans or animals. In this study, we investigated the effects of a range of psychotropic drugs on rat brain metabolites. The drugs investigated were haloperidol, clozapine, olanzapine, risperidone, aripiprazole (antipsychotics); valproate, carbamazapine (mood stabilizers) and phenytoin (antiepileptic drug). The relative concentrations of endogenous metabolites were determined using high-resolution proton nuclear magnetic resonance (1H NMR) spectroscopy. The results revealed that different classes of psychotropic drugs modulated a range of metabolites, where each drug induced a distinct neurometabolic profile. Some common responses across several drugs or within a class of drug were also observed. Antipsychotic drugs and mood stabilizers, with the exception of olanzapine, consistently increased N-acetylaspartate (NAA) levels in at least one brain area, suggesting a common therapeutic response on increased neuronal viability. Most drugs also altered the levels of several metabolites associated with glucose metabolism, neurotransmission (including glutamate and aspartate) and inositols. The heterogenic pharmacological response reflects the functional and physiological diversity of the therapeutic interventions, including side effects. Further study of these metabolites in preclinical models should facilitate the development of novel drug treatments for psychiatric disorders with improved efficacy and side effect profiles.

Evaluation of expression and function of the H+/myo-inositol transporter HMIT.

BACKGROUND: The phosphoinositide (PIns) signalling pathway regulates a series of neuronal processes, such as neurotransmitter release, that are thought to be altered in mood disorders. Furthermore, mood-stabilising drugs have been shown to inhibit key enzymes that regulate PIns production and alter neuronal growth cone morphology in an inositol-reversible manner. Here, we describe analyses of expression and function of the recently identified H+/myo-inositol transporter (HMIT) investigated as a potential regulator of PIns signalling. RESULTS: We show that HMIT is primarily a neuronal transporter widely expressed in the rat and human brain, with particularly high levels in the hippocampus and cortex, as shown by immunohistochemistry. The transporter is localised at the Golgi apparatus in primary cultured neurones. No HMIT-mediated electrophysiological responses were detected in rat brain neurones or slices; in addition, inositol transport and homeostasis were unaffected in HMIT targeted null-mutant mice. CONCLUSION: Together, these data do not support a role for HMIT as a neuronal plasma membrane inositol transporter, as previously proposed. However, we observed that HMIT can transport inositol triphosphate, indicating unanticipated intracellular functions for this transporter that may be relevant to mood control.

Prolyl oligopeptidase binds to GAP-43 and functions without its peptidase activity.

Inhibitors of the enzyme prolyl oligopeptidase (PO) improve performance in rodent learning and memory tasks. PO inhibitors are also implicated in the action of drugs used to treat bipolar disorder: they reverse the effects of three mood stabilizers on the dynamic behaviour of neuronal growth cones. PO cleaves prolyl bonds in short peptides, suggesting that neuropeptides might be its brain substrates. PO is located in the cytosol, however, where it would not contact neuropeptides. Here, we show that mice with a targeted PO null-mutation have altered growth cone dynamics. The wild-type phenotype is restored by PO cDNAs encoding either native or a catalytically-dead enzyme. In addition, we show that PO binds to the growth-associated protein GAP-43, which is a key regulator of synaptic plasticity. Taken together, our results show that peptidase activity is not required for PO function in neurons and suggest that PO instead acts by binding to cytosolic proteins that control growth cone and synaptic function.

Antipsychotic treatment alters protein expression associated with presynaptic function and nervous system development in rat frontal cortex.

Haloperidol and olanzapine are widely used antipsychotic drugs in the treatment of schizophrenia and other psychotic disorders. Despite extensive research efforts within the biopharmaceutical industry and academia, the exact molecular mechanisms of their action remain largely unknown. Since the response of patients to existing medications can be variable and often includes severe side effects, it is critical to increase our knowledge on their mechanism of action to guide clinical usage and new drug development. In this study, we have employed the label-free liquid chromatography tandem mass spectrometry (LC-MSE) to identify differentially expressed proteins in rat frontal cortex following subchronic treatment with haloperidol or olanzapine. Subcellular fractionation was performed to increased proteomic coverage and provided insight into the subcellular location involved in the mechanism of drug action. LC-MSE profiling identified 531 and 741 annotated proteins in fractions I (cytoplasmic-) and II (membrane enriched-) in two drug treatments. Fifty-nine of these proteins were altered significantly by haloperidol treatment, 74 by olanzapine and 21 were common to both treatments. Pathway analysis revealed that both drugs altered similar classes of proteins associated with cellular assembly/organization, nervous system development/function (particularly presynaptic function) and neurological disorders, which indicate a common mechanism of action. The top affected canonical signaling pathways differed between the two treatments. The haloperidol data set showed a stronger association with Huntington's disease signaling, while olanzapine treatment showed stronger effects on glycolysis/gluconeogenesis. This could either relate to a difference in clinical efficacy or side effect profile of the two compounds. The results were consistent with the findings reported previously by targeted studies, demonstrating the validity of this approach. However, we have also identified many novel proteins which have not been found previously to be associated with these drugs. Further study of these proteins could provide new insights into the etiology of the disease or the mechanism of antipsychotic medications.

Investigation of G72 (DAOA) expression in the human brain.

BACKGROUND: Polymorphisms at the G72/G30 locus on chromosome 13q have been associated with schizophrenia or bipolar disorder in more than ten independent studies. Even though the genetic findings are very robust, the physiological role of the predicted G72 protein has thus far not been resolved. Initial reports suggested G72 as an activator of D-amino acid oxidase (DAO), supporting the glutamate dysfunction hypothesis of schizophrenia. However, these findings have subsequently not been reproduced and reports of endogenous human G72 mRNA and protein expression are extremely limited. In order to better understand the function of this putative schizophrenia susceptibility gene, we attempted to demonstrate G72 mRNA and protein expression in relevant human brain regions. METHODS: The expression of G72 mRNA was studied by northern blotting and semi-quantitative SYBR-Green and Taqman RT-PCR. Protein expression in human tissue lysates was investigated by western blotting using two custom-made specific anti-G72 peptide antibodies. An in-depth in silico analysis of the G72/G30 locus was performed in order to try and identify motifs or regulatory elements that provide insight to G72 mRNA expression and transcript stability. RESULTS: Despite using highly sensitive techniques, we failed to identify significant levels of G72 mRNA in a variety of human tissues (e.g. adult brain, amygdala, caudate nucleus, fetal brain, spinal cord and testis) human cell lines or schizophrenia/control post mortem BA10 samples. Furthermore, using western blotting in combination with sensitive detection methods, we were also unable to detect G72 protein in a number of human brain regions (including cerebellum and amygdala), spinal cord or testis. A detailed in silico analysis provides several lines of evidence that support the apparent low or absent expression of G72. CONCLUSION: Our results suggest that native G72 protein is not normally present in the tissues that we analysed in this study. We also conclude that the lack of demonstrable G72 expression in relevant brain regions does not support a role for G72 in modulation of DAO activity and the pathology of schizophrenia via a DAO-mediated mechanism. In silico analysis suggests that G72 is not robustly expressed and that the transcript is potentially labile. Further studies are required to understand the significance of the G72/30 locus to schizophrenia.

Comparison between intraperitoneal and subcutaneous phencyclidine administration in Sprague-Dawley rats: a locomotor activity and gene induction study.

In a putative model of acute phencyclidine (PCP)-induced psychosis we evaluated effects of the drug on locomotor activity (LMA) and immediate early gene (IEG) induction in the rat using two routes of drug administration, intraperitoneal (i.p.) and subcutaneous (s.c.). Adult male rats received saline or PCP (1.0-5.0 mg/kg) either i.p or s.c. and were assessed for LMA for 60 min. At the end of the LMA testing animals were culled and blood and brain samples were collected for PCP concentration analysis. Separate cohorts of animals received 5.0 mg/kg PCP (i.p. or s.c.) and were used to investigate (1) the pharmacokinetics of PCP or (2) induction of IEG (Arc, c-fos, BDNF, junB, Krox-20, sgk-1, NURR1, fra-2, Krox-24, and egr-3) mRNA expression in the prefrontal cortex (PFC). Administration of PCP resulted in locomotor hyperactivity which was more robust and longer-lasting in animals dosed s.c. compared to i.p.-treated-animals. Differences in hyperlocomotion were paralleled by higher concentrations of PCP in the blood and in the brain of s.c.-treated animals compared to i.p.-treated animals. The differences in the concentration of PCP between the two routes of administration were detected 30 min after dosing and persisted for up to 4 h. Administration of PCP via the s.c. route resulted in induction of more IEGs and consistently larger magnitudes of induction than that via the i.p. route. Therefore, we have outlined the dosing conditions to induce rapid and robust effect of acute PCP on behaviour, gene induction, and pharmacokinetic profile, to allow investigation of this as a potential animal model of acute psychosis.

Interactions among genes in the ErbB-Neuregulin signalling network are associated with increased susceptibility to schizophrenia.

BACKGROUND: Evidence of genetic association between the NRG1 (Neuregulin-1) gene and schizophrenia is now well-documented. Furthermore, several recent reports suggest association between schizophrenia and single-nucleotide polymorphisms (SNPs) in ERBB4, one of the receptors for Neuregulin-1. In this study, we have extended the previously published associations by investigating the involvement of all eight genes from the ERBB and NRG families for association with schizophrenia. METHODS: Eight genes from the ERBB and NRG families were tested for association to schizophrenia using a collection of 396 cases and 1,342 blood bank controls ascertained from Aberdeen, UK. A total of 365 SNPs were tested. Association testing of both alleles and genotypes was carried out using the fast Fisher's Exact Test (FET). To understand better the nature of the associations, all pairs of SNPs separated by >or= 0.5 cM with at least nominal evidence of association (P < 0.10) were tested for evidence of pairwise interaction by logistic regression analysis. RESULTS: 42 out of 365 tested SNPs in the eight genes from the ERBB and NRG gene families were significantly associated with schizophrenia (P < 0.05). Associated SNPs were located in ERBB4 and NRG1, confirming earlier reports. However, novel associations were also seen in NRG2, NRG3 and EGFR. In pairwise interaction tests, clear evidence of gene-gene interaction was detected for NRG1-NRG2, NRG1-NRG3 and EGFR-NRG2, and suggestive evidence was also seen for ERBB4-NRG1, ERBB4-NRG2, ERBB4-NRG3 and ERBB4-ERBB2. Evidence of intragenic interaction was seen for SNPs in ERBB4. CONCLUSION: These new findings suggest that observed associations between NRG1 and schizophrenia may be mediated through functional interaction not just with ERBB4, but with other members of the NRG and ERBB families. There is evidence that genetic interaction among these loci may increase susceptibility to schizophrenia.

Evaluation of the mGlu8 receptor as a putative therapeutic target in schizophrenia.

Aberrant glutamatergic neurotransmission may underlie the pathogenesis of schizophrenia and metabotropic glutamate receptors (mGluRs) have been implicated in the disease. We have established the localization of the group III mGluR subtype, mGluR8, in the human body and investigated the biological effects of the selective mGluR8 agonist (S)-3,4-dicarboxyphenylglycine ((S)-3,4-DCPG) in schizophrenia-related animal models. The mGlu8 receptor has a widespread CNS distribution with expression observed in key brain regions associated with schizophrenia pathogenesis including the hippocampus. (S)-3,4-DCPG inhibited synaptic transmission and increased paired-pulse facilitation in rat hippocampal slices supporting the role of mGluR8 as a presynaptic autoreceptor. Using the rat Maximal Electroshock Seizure Threshold (MEST) test, (S)-3,4-DCPG (30 mg/kg, i.p.) reduced seizure activity confirming the compound to be centrally active following systemic administration. (S)-3,4-DCPG did not reverse (locomotor) hyperactivity induced by acute administration of phenylcyclidine (PCP, 1-32 mg/kg, i.p.) or amphetamine (3-30 mg/kg, i.p.) in Sprague-Dawley rats. However, 10 nmol (i.c.v.) (S)-3.4-DCPG did reverse amphetamine-induced hyperactivity in mice although it also inhibited spontaneous locomotor activity at this dose. In addition, mGluR8 null mutant mouse behavioral phenotyping revealed an anxiety-related phenotype but no deficit in sensorimotor gating. These data provide a potential role for mGluR8 in anxiety and suggest that mGluR8 may not be a therapeutic target for schizophrenia.

The common inositol-reversible effect of mood stabilizers on neurons does not involve GSK3 inhibition, myo-inositol-1-phosphate synthase or the sodium-dependent myo-inositol transporters.

We previously showed that the mood stabilizers lithium, valproate (VPA), and carbamazepine (CBZ) have a common, inositol-reversible effect on the dynamic behavior of sensory neurons, suggesting that they all inhibit phosphoinositide (PIns) synthesis. We now report similar effects of the drugs in cortical neurons and show by mRNA analysis that these neurons do not express myo-inositol-1-phosphate synthase (MIP-synthase) or the sodium-dependent myo-inositol transporters (SMIT1 and SMIT2), but they do express the H+/myo-inositol transporter (HMIT) mRNA and protein. We used glycogen synthase kinase-3 (GSK3) inhibitors and Western blotting of GSK3 targets to confirm that the common effects of the drugs on both sensory and cortical neuron growth cones are inositol-dependent and GSK3-independent. Moreover, the anti-convulsant drugs gabapentin and phenytoin do not mimic the mood stabilizers. These results confirm that the common inositol-reversible effect of mood stabilizers on neurons does not involve GSK3 and further show that the effects are independent of MIP-synthase and SMIT transporters.

Region-specific reduction in entorhinal gamma oscillations and parvalbumin-immunoreactive neurons in animal models of psychiatric illness.

Psychiatric illnesses, particularly schizophrenia, are associated with disrupted markers for interneuronal function and interneuron-mediated brain rhythms such as gamma frequency oscillations. Here we investigate a possible link between these two observations in the entorhinal cortex and hippocampus by using a genetic and an acute model of psychiatric illness. Lysophosphatidic acid 1 receptor-deficient (LPA1-deficient) mice show psychomotor-gating deficits and neurochemical changes resembling those seen in postmortem schizophrenia studies. Similar deficits are seen acutely with antagonism of the NMDA subtype of glutamate receptor. Neither model induced any change in power or frequency of gamma rhythms generated by kainate in hippocampal slices. In contrast, a dramatic decrease in the power of gamma oscillations was seen in superficial, but not deep, medial entorhinal cortex layers in both models. Immunolabeling for GABA, parvalbumin, and calretinin in medial entorhinal cortex from LPA1-deficient mice showed an approximately 40% reduction in total GABA- and parvalbumin-containing neurons, but no change in the number of calretinin-positive neurons. This deficit was specific for layer II (LII). No change in the number of neurons positive for these markers was seen in the hippocampus. Acute NMDA receptor blockade, which selectively reduces synaptic drive to LII entorhinal interneurons, also disrupted gamma rhythms in a similar manner in superficial entorhinal cortex, but not in hippocampus. These data demonstrate an area-specific deficit in gamma rhythmogenesis in animal models of psychiatric illness and suggest that loss, or reduction in function, of interneurons having a large NMDA receptor expression may underlie the network dysfunction that is seen.

Neurochemical changes in LPA1 receptor deficient mice--a putative model of schizophrenia.

LPA1 is a Gi-coupled seven transmembrane receptor with high affinity for the ligand lysophosphatidic acid. We have investigated the effect of targeted deletion at the lpa1 locus on evoked release of amino acids from hippocampal slices, using in vitro superfusion techniques, and evoked 5-HT efflux from the dorsal raphe nucleus, using in vitro fast cyclic voltammetry. Superfusion of hippocampal slices revealed that basal levels of tyrosine, aspartate and glutamate release were significantly increased while K+ -evoked release of glutamate and GABA were significantly decreased in lpa1(-/-) mice. Fast cyclic voltammetry measurements in the dorsal raphe nucleus demonstrated significant decreases in electrically evoked 5-HT efflux in lpa1(-/-) mice. In summary, these data demonstrate that the lpa1 mutation produces a number of changes in neurotransmitters that have been associated with a schizophrenic-like pathology.

Comparative analysis of the effects of four mood stabilizers in SH-SY5Y cells and in primary neurons.

OBJECTIVES: The mood-stabilizing drug valproic acid (VPA) exerts a neurotrophic effect on the human neuroblastoma cell line, SH-SY5Y. We aimed to establish whether other mood-stabilizing drugs have a similar action and which signalling pathways mediate this process. METHODS: We analysed the effects of the mood stabilizers VPA, lithium, carbamazepine and lamotrigine on proliferation, survival, neurite outgrowth and extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) activation using the SH-SY5Y cell line. We also compared their effects in primary neurons. RESULTS: We found that VPA promotes neurite outgrowth and prevents cell death in SH-SY5Y cells, but has no effect on cell proliferation. This neurotrophic effect does not involve inhibition of glycogen synthase kinase-3, histone deacetylase or prolyl oligopeptidase; the effect also does not seem to involve protein kinase C. In contrast, VPA activates ERK/MAPK and the survival effect of VPA is blocked by inhibition of the ERK/MAPK signalling pathway. Moreover, other activators of ERK/MAPK, such as epidermal growth factor and phorbol 12-myristate 13-acetate, mimic the neurotrophic effects of VPA. Other mood stabilizers do not activate ERK/MAPK and do not promote neurite outgrowth or survival of SH-SY5Y cells. In contrast, both lithium and VPA activate ERK/MAPK in rat primary cortical neurons. CONCLUSIONS: We investigated four mood stabilizers that are effective in the treatment of bipolar disorder. Our results suggest that, while some mood stabilizers may have additional neuroprotective effects, activation of ERK/MAPK does not appear to be a mechanism common to all mood-stabilizing drugs.

Lipid rafts mediate the interaction between myelin-associated glycoprotein (MAG) on myelin and MAG-receptors on neurons.

The interaction between myelin-associated glycoprotein (MAG), expressed at the periaxonal membrane of myelin, and receptors on neurons initiates a bidirectional signalling system that results in inhibition of neurite outgrowth and maintenance of myelin integrity. We show that this involves a lipid-raft to lipid-raft interaction on opposing cell membranes. MAG is exclusively located in low buoyancy Lubrol WX-insoluble membrane fractions isolated from whole brain, primary oligodendrocytes, or MAG-expressing CHO cells. Localisation within these domains is dependent on cellular cholesterol and occurs following terminal glycosylation in the trans-Golgi network, characteristics of association with lipid rafts. Furthermore, a recombinant form of MAG interacts specifically with lipid-raft fractions from whole brain and cultured cerebellar granule cells, containing functional MAG receptors GT1b and Nogo-66 receptor and molecules required for transduction of signal from MAG into neurons. The localisation of both MAG and MAG receptors within lipid rafts on the surface of opposing cells may create discrete areas of high avidity multivalent interaction, known to be critical for signalling into both cell types. Localisation within lipid rafts may provide a molecular environment that facilitates the interaction between MAG and multiple receptors and also between MAG ligands and molecules involved in signal transduction.