Determining the gene expression pattern underlying L-DOPA-induced dyskinesisas in the 6-OHDA hemiparkinsonian rat model of Parkinson's disease

Abstract

L-DOPA is the gold standard treatment for Parkinson’s disease. However, with prolonged use the majority of patients will develop abnormal involuntary movements (dyskinesias) as a side effect of L-DOPA use. These movements are known as L-DOPA-induced dyskinesias (LIDs). Currently there is no widely effective treatment for alleviating LIDs. The purpose of this study was to examine the genes expressed in dorsal striatum in the dyskinetic and non-dyskinetic state.

In order to minimise the variables between the dyskinetic and non-dyskinetic state, an animal model of LIDs with no differences in surgical, behavioural, or pharmacological treatment was developed using the 6-OHDA rat model of Parkinson’s disease. Rats were subject to a unilateral 6-OHDA lesion of the medial forebrain bundle to induce hemiparkinsonism. After a two-week recovery period, each rat received twice daily subcutaneous injections of 4 mg/kg of L-DOPA (with 7.5 mg/kg benserazide). Using this protocol 50% of the treated rats developed LIDs, as observed using the rodent Abnormal Involuntary Movements scoring method. Because these rats received identical treatment throughout the experiment, these changes may be associated solely with the expression or absence of LIDs. To investigate the gene expression profile associated with LIDs, dyskinetic and non-dyskinetic rats – along with untreated parkinsonian controls – were subject to RNAseq gene expression profiling in two experiments. The first experiment investigated the gene expression profile observed at the peak of L-DOPA in the brain, and the peak of LID expression (peak-dose study). 255 genes were differentially expressed between two or more of the groups (dyskinetic, non-dyskinetic, and untreated parkinsonian). Of those, 110 changes were observed between the dyskinetic and non-dyskinetic rats. The majority of the genes showed decreases in gene expression in dyskinetic rats compared with either the non-dyskinetic or untreated parkinsonian rats. The second RNAseq experiment investigated the gene expression profile associated with dyskinesia expression when L-DOPA was not present, i.e. the chronic gene expression changes not associated with the performance of dyskinetic movements (off-dose study). This experiment found 412 genes differentially expressed between two or more groups. In contrast to the peak-dose study, in the off-dose study only a fifth of the observed changes in gene expression were between the dyskinetic and non-dyskinetic rats (80). The majority (266) of gene expression changes observed were between the L-DOPA-treated rats and the untreated parkinsonian controls. Furthermore, the majority of changes observed were increases in gene expression in the dyskinetic and non-dyskinetic rats compared with the untreated parkinsonian controls (239). A number of potential therapeutic targets were identified. Intervention in the NO/cGMP/PKG pathway is suggested, either through direct modulation of Pde9a and Prkg1 expression or ion channels associated with the pathway. The dynein complex associated with transport and the cAMP/DARPP32/MAPK pathway are suggested. Interestingly, comparison of the differentially expressed genes in the two RNAseq experiments revealed nine genes that showed changes in expression in both conditions. Furthermore, only two of those genes showed the same pattern of gene expression in the peak-dose and off-dose experiments, arc and cyp7b1. In addition, cAMP/DARRP32/MAPK signalling is down-regulated between the dyskinetic and non-dyskinetic states at the peak of LID expression but up-regulated when L-DOPA has worn off. Overall, this study suggest a number of therapeutic targets, but also that when treating L-DOPA-induced dyskinesias the timing of the treatment may be just as important as the treatment itself, when interpreting the patterns of changes in gene expression, and for designing and administering novel treatments for dyskinesias.