Walking pattern analysis after unilateral 6-OHDA lesion and transplantation of foetal dopaminergic progenitor cells in rats

Abstract

Functional sensorimotor recovery after transplantation of mesencephalic dopaminergic (DAergic) neurons has been well documented in the rat 6-hydroxydopamine (6-OHDA) model of Parkinson's disease. However, the functional restoration of more specific gait-related patterns such as skilled walking, balance, and individual limb movements have been insufficiently studied. The purpose of this study was to investigate the behavioural effects of intrastriatal DA grafts on different aspects of normal and skilled walking in rats following unilateral 6-OHDA lesions of the medial forebrain bundle. Rats were subjected to drug-induced rotation, detailed footprint analysis, and assessment of skilled walking in the ladder rung walking test prior and after the transplantation of E14 ventral mesencephalon-derived progenitor cells. Good DAergic graft survival, as revealed by immunohistochemistry, was accompanied by a compensation of drug-induced rotational asymmetries. Interestingly, the analysis of walking patterns displayed a heterogeneous graft-induced response in skilled and non-skilled limb use. Grafted animals made fewer errors with their contralateral limbs in skilled walking than the sham-transplanted rats, and they improved their ipsi- and contralateral limb rotation. However, the parameter distance between feet showed a delayed recovery, and the stride length was not affected by the DA grafts at all. These findings indicate that ectopic intrastriatal transplantation of E14 ventral mesencephalon-derived cells promotes recovery of gait balance and stability, but does not ameliorate the shuffling gait pattern associated with 6-OHDA lesions. A full restoration of locomotor gait pattern might require a more complete and organotypic reconstruction of the mesotelencephalic DAergic pathway.

Introduction

Shuffling gait is one of the most prominent motor impairments in Parkinson's disease (PD; [26], [35], [36]). The death of predominantly dopaminergic (DAergic) neurons, projecting from the substantia nigra pars compacta (SNc) to the striatum (Str), leads to an impoverishment of motor movements in general [5], [19], [22].

To mimic Parkinsonian symptoms, the unilateral injection of the neurotoxin 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle (MFB) is commonly used. This results in the depletion of striatal DA on the ipsilateral side of the brain [30], [58]. The lesioned animals show a rotational bias when injected with direct or indirect DA agonists [59], and display a number of motor impairments that resemble those of human PD [21], [60]. For instance, a unilateral 6-OHDA lesion causes loss of skilled reaching accuracy and abnormal movement patterns [33]. The lesion also triggers other more subtle – and less studied – impairments in locomotion, such as short shuffling steps and muscle co-contractions [30].

Current drug-based therapies (l-DOPA and DA agonists) have only a limited success in improving motor impairments and cannot stop the progression of neurodegenerative events during the course of the disease [25]. As an alternative treatment the cell-replacement therapy has been used to compensate for DA cell loss by ectopically implanting foetal progenitor cells into the DA-depleted striatum [25], [50], [54].

Intracranial cell transplantation has shown to reverse some of the functional deficits observed in rodent models of neurodegeneration. The literature provides a large amount of research into the intrastriatal transplantation of embryonic 14 ventral mesencephalon-derived progenitor cells (E14 VM-derived cells), particularly into cell survival, morphological graft integration and development, and into general aspects of motor behaviour [12], [14], [38], [56], [57], [61]. In a parallel report Klein et al. [24] observed that the DA grafts indeed led to an improvement of skilled limb movements after quantitative evaluation of reaching behaviour. However, the qualitative measurements of the individual reaching movements revealed no graft-related effects. It has been suggested that compensatory movement strategies within the subjects might be the reason for motor improvement after transplantation. Considering that gait impairments are amongst the cardinal symptoms of PD, it is surprising that only little data can be found about the effects of intrastriatal DAergic grafts on walking patterns in hemiparkinsonian rats in the literature.

The objective of the present study was to further investigate the functional effects of E14 VM-derived grafts on pre- and post-transplantational walking patterns in hemiparkinsonian rats. The rats were tested in the drug-induced rotation paradigm to measure the degree of lesion and graft outcome in vivo. The footprint analysis was carried out in order to assess general locomotion and limb control; skilled walking was assessed with the help of the ladder rung walking test. The results provide further understanding of the functional restorative capacity of the neurotransplantation approach and will help to further optimize clinically relevant cell-based therapies.