ABSTRACT
Engrailed-2 (En2), a homeodomain transcription factor involved in regionalization and patterning of the midbrain and hindbrain regions has been associated to autism spectrum disorders (ASDs). En2 knockout (En2(-/-)) mice show ASD-like features accompanied by a significant loss of GABAergic subpopulations in the hippocampus and neocortex. Brain-derived neurotrophic factor (BDNF) is a crucial factor for the postnatal development of forebrain GABAergic neurons, and altered GABA signaling has been hypothesized to underlie the symptoms of ASD. Here we sought to determine whether interneuron loss in the En2(-/-) forebrain might be related to altered expression of BDNF and its signaling receptors. We first evaluated the expression of different BDNF mRNA isoforms in the neocortex and hippocampus of wild-type (WT) and En2(-/-) mice. Quantitative RT-PCR showed a marked down-regulation of several splicing variants of BDNF mRNA in the neocortex but not hippocampus of adult En2(-/-) mice, as compared to WT controls. Accordingly, levels of mature BDNF protein were lower in the neocortex but not hippocampus of En2(-/-) mice, as compared to WT. Increased levels of phosphorylated TrkB and decreased levels of p75 receptor were also detected in the neocortex of mutant mice. Accordingly, the expression of low density lipoprotein receptor (LDLR) and RhoA, two genes regulated via p75 was significantly altered in forebrain areas of mutant mice. These data indicate that BDNF signaling alterations might be involved in the anatomical changes observed in the En2(-/-) forebrain and suggest a pathogenic role of altered BDNF signaling in this mouse model of ASD.
Subject(s)
Hippocampus/metabolism , Homeodomain Proteins/metabolism , Neocortex/metabolism , Nerve Tissue Proteins/metabolism , Receptors, Nerve Growth Factor/metabolism , Alternative Splicing , Animals , Autism Spectrum Disorder/metabolism , Brain-Derived Neurotrophic Factor/metabolism , Corpus Striatum/metabolism , Corpus Striatum/pathology , Disease Models, Animal , Female , Hippocampus/pathology , Homeodomain Proteins/genetics , Male , Mice, 129 Strain , Mice, Inbred C57BL , Mice, Knockout , Neocortex/pathology , Nerve Tissue Proteins/genetics , Phosphorylation , RNA, Messenger/metabolism , Receptor, trkB/metabolism , Receptors, LDL/metabolism , rhoA GTP-Binding Protein/metabolismABSTRACT
Mice lacking the homeodomain transcription factor Engrailed-2 (En2(-/-) mice) are a well-characterized model for autism spectrum disorders (ASD). En2(-/-) mice present molecular, neuropathological and behavioral deficits related to ASD, including down-regulation of ASD-associated genes, cerebellar hypoplasia, interneuron loss, enhanced seizure susceptibility, decreased sociability and impaired cognition. Specifically, impaired spatial learning in the Morris water maze (MWM) is associated with reduced expression of neurofibromin and increased phosphorylation of extracellular-regulated kinase (ERK) in the hippocampus of En2(-/-) adult mice. In the attempt to better understand the molecular cascades underlying neurofibromin-dependent cognitive deficits in En2 mutant mice, we investigated the expression and phosphorylation of synapsin I (SynI; a major target of neurofibromin-dependent signaling) in the hippocampus of wild-type (WT) and En2(-/-) mice before and after MWM. Here we show that SynI mRNA and protein levels are down-regulated in the hippocampus of naïve and MWM-treated En2(-/-) mice, as compared to WT controls. This down-regulation is paralleled by reduced levels of SynI phosphorylation at Ser549 and Ser553 residues in the hilus of mutant mice, before and after MWM. These data indicate that in En2(-/-) hippocampus, neurofibromin-dependent pathways converging on SynI phosphorylation might underlie hippocampal-dependent learning deficits observed in En2(-/-) mice.
Subject(s)
Child Development Disorders, Pervasive/genetics , Child Development Disorders, Pervasive/metabolism , Hippocampus/metabolism , Homeodomain Proteins/genetics , Nerve Tissue Proteins/genetics , Spatial Learning/physiology , Synapsins/metabolism , Animals , Child Development Disorders, Pervasive/psychology , Disease Models, Animal , Down-Regulation , Female , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Phosphorylation , Signal TransductionABSTRACT
The En2 gene, coding for the homeobox-containing transcription factor Engrailed-2 (EN2), has been associated to autism spectrum disorder (ASD). Due to neuroanatomical and behavioral abnormalities, which partly resemble those observed in ASD patients, En2 knockout (En2(-/-)) mice have been proposed as a model for ASD. In the mouse embryo, En2 is involved in the specification of midbrain/hindbrain regions, being predominantly expressed in the developing cerebellum and ventral midbrain, and its expression is maintained in these structures until adulthood. Here we show that in the adult mouse brain, En2 mRNA is expressed also in the hippocampus and cerebral cortex. Hippocampal En2 mRNA content decreased after seizures induced by kainic acid (KA). This suggests that En2 might also influence the functioning of forebrain areas during adulthood and in response to seizures. Indeed, a reduced expression of parvalbumin and somatostatin was detected in the hippocampus of En2(-/-) mice as compared to wild-type (WT) mice, indicating an altered GABAergic innervation of limbic circuits in En2(-/-) mice. In keeping with these results, En2(-/-) mice displayed an increased susceptibility to KA-induced seizures. KA (20 mg/kg) determined more severe and prolonged generalized seizures in En2(-/-) mice, when compared to WT animals. Seizures were accompanied by a widespread c-fos and c-jun mRNA induction in the brain of En2(-/-) but not WT mice. Long-term histopathological changes (CA1 cell loss, upregulation of neuropeptide Y) also occurred in the hippocampus of KA-treated En2(-/-) but not WT mice. These findings suggest that En2(-/-) mice might be used as a novel tool to study the link between epilepsy and ASD.
Subject(s)
Disease Susceptibility , Excitatory Amino Acid Agonists/toxicity , Kainic Acid/toxicity , Nerve Tissue Proteins/deficiency , Seizures/chemically induced , Seizures/genetics , Animals , Brain/anatomy & histology , Brain/drug effects , Brain/metabolism , Gene Expression Regulation/drug effects , Gene Expression Regulation/genetics , Homeodomain Proteins/genetics , Mice , Mice, Inbred C57BL , Mice, Knockout , Nerve Tissue Proteins/genetics , Parvalbumins/metabolism , Proto-Oncogene Proteins c-fos/genetics , Proto-Oncogene Proteins c-fos/metabolism , Proto-Oncogene Proteins c-jun/genetics , Proto-Oncogene Proteins c-jun/metabolism , RNA, Messenger/metabolism , Reaction Time/drug effects , Somatostatin/metabolism , Time FactorsABSTRACT
In order to reach a deeper insight into the mechanism of diethyldithiocarbamate (DDC)-induced enhancement of MPTP toxicity in mice, we showed that CYP450 (2E1) inhibitors, such as diallyl sulfide (DAS) or phenylethylisothiocyanate (PIC), also potentiate the selective DA neuron degeneration in C57/bl mice. Furthermore we showed that CYP 2E1 is present in the brain and in the basal ganglia of mice (Vaglini et al., 2004). However, because DAS and PIC are not selective CYP 2E1 inhibitors and in order to provide direct evidence for CYP 2E1 involvement in the enhancement of MPTP toxicity, CYP 2E1 knockout mice (GONZ) and wild type animals (SVI) of the same genetic background were treated with MPTP or the combined DDC + MPTP treatment. In CYP 2E1 knockout mice, DDC pretreatment completely fails to enhance MPTP toxicity, although enhancement of MPTP toxicity was regularly present in the SVI control animals. The immunohistochemical study confirms our results and suggests that CYP 2E1 may have a detoxifying role.
Subject(s)
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine/pharmacology , Cytochrome P-450 CYP2E1/genetics , Ditiocarb/pharmacology , Neurotoxins/pharmacology , Parkinsonian Disorders/chemically induced , Animals , Cricetinae , Cytochrome P-450 CYP2E1 Inhibitors , Dopamine/metabolism , Drug Synergism , Inactivation, Metabolic/genetics , Mice , Mice, Knockout , Parkinsonian Disorders/genetics , Polymerase Chain Reaction , Premedication , Substantia Nigra/drug effects , Substantia Nigra/pathology , Tyrosine 3-Monooxygenase/metabolism , Ventral Tegmental Area/drug effects , Ventral Tegmental Area/pathologyABSTRACT
An important goal in Parkinson's Disease research is to identify neuroprotective therapy, and the interaction between basic science and clinical research is needed to discover drugs that can slow or halt the disorder progression. At present there is not a perfect animal model of PD to test neuroprotective strategies, however the models that portray the basic characteristics needed are toxin-induced and gene-based models. The first group comprehends 6-OHDA e MPTP and recently rotenone, paraquat and epoxomicin treated animals that shows some of human disease characteristics. Gene-based models are various and, even if with limits, they seem suitable models to test neuroprotection in PD since they present replicable lesions, a predictable pattern of neurodegeneration and a well-characterized behavior, biochemistry and morphology to assist in the understanding of induced changes. In clinical trials researchers have first used as marker of disease progression clinical scores and motor tasks which are limited by the potential symptomatic effect of tested drugs and are not useful in the pre-clinical phases of PD. Recently has emerged the important role of neuroimaging (Dopamine Transporter SPECT, 18FDopa-PET) as surrogate biomarker of PD progression. Even if there are still concerns about the influence of regulatory effects of tested drugs, neuroimaging features could represent a good outcome measure to evaluate PD progression and putative neuroprotective effect of pharmacological and non-pharmacological manipulations.
Subject(s)
Diagnostic Imaging/methods , Neurodegenerative Diseases , Neuroprotective Agents/therapeutic use , Parkinson Disease , Animals , Disease Models, Animal , Humans , Neurodegenerative Diseases/diagnosis , Neurodegenerative Diseases/drug therapy , Neurodegenerative Diseases/etiology , Parkinson Disease/complications , Parkinson Disease/etiology , Parkinson Disease/pathologyABSTRACT
The PC3 gene is a marker of dividing neuroepithelial (NE) cells. We transduced single cortical precursors of the ventricular zone (VZ) with a PC3-carrying retroviral vector at E16 stage, and analysed the effects of transgene expression on their progeny in 3-week-old animals. Unlike control-transduced cells, all viable PC3-transduced cells remained close to the ventricle and displayed a round-shaped, undifferentiated morphology.
