We have used brain tissue from clinically well-documented and neuropathologically confirmed cases of sporadic Parkinson's disease to establish the transcriptomic expression profile of the medial and lateral substantia nigra. In addition, the superior frontal cortex was analyzed in a subset of the same cases. DNA oligonucleotide microarrays were employed, which provide whole human genome coverage. A total of 570 genes were found to be differentially regulated at a high level of significance. A large number of differentially regulated expressed sequence tags were also identified. Levels of mRNA sequences encoded by genes of key interest were validated by means of quantitative realtime polymerase chain reaction (PCR). Comparing three different normalization procedures, results based on the recently published GeneChip Robust Multi Array algorithm were found to be the most accurate predictor of realtime PCR results. Several new candidate genes which map to PARK loci are reported. In addition, the DNAJ family of chaperones is discussed in the context of Parkinson's disease pathogenesis.
Sporadic Parkinson's disease (PD) is characterized by progressive death of dopaminergic neurons within the substantia nigra. However, pathological cell death within this nucleus is not uniform. In PD, the lateral tier of the substantia nigra (SNl) degenerates earlier and more severely than the more medial nigral component (SNm). The cause of this brain regional vulnerability remains unknown. We have used DNA oligonucleotide microarrays to compare gene expression profiles from the SNl to those of the SNm in both PD and control cases. Genes expressed more highly in the PD SNl included the cell death gene, p53 effector related to PMP22, the tumour necrosis factor (TNF) receptor gene, TNF receptor superfamily, member 21, and the mitochondrial complex I gene, NADH dehydrogenase (ubiquinone) 1beta subcomplex, 3, 12 kDa (NDUFbeta3). Genes that were more highly expressed in PD SNm included the dopamine cell signalling gene, cyclic adenosine monophosphate-regulated phosphoprotein, 21 kDa, the activated macrophage gene, stabilin 1, and two glutathione peroxidase (GPX) genes, GPX1 and GPX3. Thus, there is increased expression of genes encoding pro-inflammatory cytokines and subunits of the mitochondrial electron transport chain, and there is a decreased expression of several glutathione-related genes in the SNl suggesting a molecular basis for pathoclisis. Importantly, some of the genes that are differentially regulated in the SNl are known to be expressed highly or predominantely in glial cells. These findings support the view that glial cells can be primarily affected in PD emphasizing the importance of using a whole tissue approach when investigating degenerative CNS disease.
There is growing evidence that dysfunction of the mitochondrial respiratory chain and failure of the cellular protein degradation machinery, specifically the ubiquitin-proteasome system, play an important role in the pathogenesis of Parkinson's disease. We now show that the corresponding pathways of these two systems are linked at the transcriptomic level in Parkinsonian substantia nigra. We examined gene expression in medial and lateral substantia nigra (SN) as well as in frontal cortex using whole genome DNA oligonucleotide microarrays. In this study, we use a hypothesis-driven approach in analysing microarray data to describe the expression of mitochondrial and ubiquitin-proteasomal system (UPS) genes in Parkinson's disease (PD). Although a number of genes showed up-regulation, we found an overall decrease in expression affecting the majority of mitochondrial and UPS sequences. The down-regulated genes include genes that encode subunits of complex I and the Parkinson's-disease-linked UCHL1. The observed changes in expression were very similar for both medial and lateral SN and also affected the PD cerebral cortex. As revealed by "gene shaving" clustering analysis, there was a very significant correlation between the transcriptomic profiles of both systems including in control brains. Therefore, the mitochondria and the proteasome form a higher-order gene regulatory network that is severely perturbed in Parkinson's disease. Our quantitative results also suggest that Parkinson's disease is a disease of more than one cell class, i.e. that it goes beyond the catecholaminergic neuron and involves glia as well.
This study provides an expression signature of interferon-gamma (IFN-gamma)-activated microglia. Microglia are macrophage precursor cells residing in the brain and spinal cord. The microglial phenotype is highly plastic and changes in response to numerous pathological stimuli. IFN-gamma has been established as a strong immunological activator of microglial cells both in vitro and in vivo. Affymetrix RG_U34A microarrays were used to determine the effect of IFN-gamma stimulation on migroglia cells isolated from newborn Lewis rat brains. More than 8,000 gene sequences were examined, i.e., 7,000 known genes and 1,000 expressed sequence tag (EST) clusters. Under baseline conditions, microglia expressed 326 of 8,000 genes examined (approximately 4% of all genes, 182 known and 144 ESTs). Transcription of only 34 of 7,000 known genes and 8 of 1,000 ESTs was induced by IFN-gamma stimulation. The majority of the newly expressed genes encode pro-inflammatory cytokines and components of the MHC-mediated antigen presentation pathway. The expression of 60 of 182 identified genes and of 9 of 144 ESTs was increased by IFN-gamma, whereas 29 of 182 known genes and 7 of 144 ESTs were down-regulated or undetectable in IFN-gamma-stimulated cultures. Overall, the activating effect of IFN-gamma on the microglial transcriptome showed restriction to pathways involved in antigen presentation, protein degradation, actin binding, cell adhesion, apoptosis, and cell signaling. In comparison, down-regulatory effects of IFN-gamma stimulation appeared to be confined to pathways of growth regulation, remodeling of the extracellular matrix, lipid metabolism, and lysosomal processing. In addition, transcriptomic profiling revealed previously unknown microglial genes that were de novo expressed, such as calponin 3, or indicated differential regulatory responses, such as down-regulation of cathepsins that are up-regulated in response to other microglia stimulators.
Neurons that fire in relation to licking, in the ventral part of the dorsolateral striatum (DLS), were studied during acquisition and performance of a licking task in rats for 14 sessions (2 hours/day). Task learning was indicated by fewer errors of omission of licking and improved movement efficiency (i.e., shorter lick duration) over sessions. Number of licks did not change over sessions. Overtraining did not result in habit formation, as indicated by similar reduction of licking responses following devaluation by satiety in both early and late sessions. Twenty-nine lick neurons recorded and tracked over sessions exhibited a significant linear decrease in average firing rate across all neurons over sessions, correlating with concurrent declines in lick duration. Individually, most neurons (86%) exhibited decreased firing rates, while a small proportion (14%) exhibited increased firing rates, during lick movements that were matched over sessions. Reward manipulations did not alter firing patterns over sessions. Aside from the absence of habit formation, striatal processing during unconditioned movements (i.e., licking) was characterized by high activity of movement-related neurons during early performance and decreased activity of the same neurons during overtraining, similar to our previous report of head movement neurons during acquired, skilled, instrumental head movements that ultimately became habitual (Tang et al., 2007). Decreased activity in DLS neurons may reflect a common neural mechanism underlying improvement in movement efficiency with overtraining. Nonetheless, the decreased striatal firing in relation to a movement that did not become habitual demonstrates that not all DLS changes reflect habit formation.
Over the past two decades, there has been a flurry of government papers and policy reports worldwide calling for increased number and diversity of doctoral researchers and a broadening of the curriculum to meet the developing needs of respective national ‘knowledge-driven’ economies. This has been followed by position papers and best practice examples of employability skills development in boundary-crossing doctoral programmes, especially in response to these initiatives. However, there is a disassociation between this ample literature expounding the new doctorate with its broader remit, inclusivity and production of ‘industry-ready’ graduates and the comparatively sparse literature on the doctoral candidates’ experiences of their programmes and career readiness. Within this review, we briefly outline international government initiatives and examples of the responses by Life Science and Biomedical doctoral programmes to address these various challenges. Furthermore, we explore the recent literature on the lived experience of doctoral researchers by examining their perception of the recent changes to the research context to make recommendations for universities and supervisors on how to better support an ever more diverse doctoral population for a wide range of career opportunities. Examples of how doctoral researchers themselves can make the best of currently available opportunities are also provided.
The identification of mutations that cause familial Parkinson's disease (PD) provides a framework for studies into pathways that may be perturbed also in the far more common, non-familial form of the disorder. Following this hypothesis, we have examined the gene regulatory network that links alpha-synuclein and parkin pathways with dopamine metabolism in neuropathologically verified cases of sporadic PD. By means of an in silico approach using a database of eukaryotic molecular interactions and a whole genome transcriptome dataset validated by qRT-PCR and histological methods, we found parkin and functionally associated genes to be upregulated in the lateral substantia nigra (SN). In contrast, alpha-synuclein and ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) gene expression levels were significantly reduced in both the lateral and medial SN in PD. Gene expression for Septin 4, a member of the GTP-binding protein family involved in alpha-synuclein metabolism was elevated in the lateral parkinsonian SN. Additionally, catalase and mitogen-activated protein kinase 8 and poly (ADP-ribose) polymerase family member 1 (PARP1) known to function in DNA repair and cell death induction, all members of the dopamine synthesis pathway, were up-regulated in the lateral SN. In contrast, two additional PD-linked genes, glucocerebrosidase and nuclear receptor subfamily 4, group A, member 2 (NR4A2) showed reduced expression. We show that in sporadic PD, parkin, alpha-synuclein and dopamine pathways are co-deregulated. Alpha-synuclein is a member of all three gene regulatory networks. Our analysis results support the view that alpha-synuclein has a central role in the familial as well as the non-familial form of the disease and provide steps towards a pathway definition of PD.
To examine the role of striatal mechanisms in cocaine-induced stereotyped licking, we investigated the acute effects of cocaine on striatal neurons in awake, freely moving rats before and after cocaine administration (0, 5, 10, or 20 mg/kg). Stereotyped licking was induced only by the high dose. Relative to control (saline), cocaine reduced lick duration and concurrently increased interlick interval, particularly at the high dose, but it did not affect licking rhythm. Firing rates of striatal neurons phasically related to licking movements were compared between matched licks before and after injection, minimizing any influence of sensorimotor variables on changes in firing. Both increases and decreases in average firing rate of striatal neurons were observed after cocaine injection, and these changes exhibited a dose-dependent pattern that strongly depended on predrug firing rate. At the middle and high doses relative to the saline group, the average firing rates of slow firing neurons were increased by cocaine, resulting from a general elevation of movement-related firing rates. In contrast, fast firing neurons showed decreased average firing rates only in the high-dose group, with reduced firing rates across the entire range for these neurons. Our findings suggest that at the high dose, increased phasic activity of slow firing striatal neurons and simultaneously reduced phasic activity of fast firing striatal neurons may contribute, respectively, to the continual initiation of stereotypic movements and the absence of longer movements.
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