Abstract
Parkinson’s disease (PD) presents with characteristic movement deficits that are thought to evolve as a consequence of degeneration of dopamine neurons in the brain. The degree of PD motor symptoms strongly correlates to altered activity in the basal ganglia (BG). However, BG dysfunction does not fully explain PD pathogenesis and a system level approach that considers the involvement of other motor structures, such as the cerebellum, is required. Imaging studies show significant grey matter atrophy and hyperactivity in the cerebellum of PD patients but there is limited understanding of the significance of these changes at the neuronal level. Therefore, the aim of the experiments presented in this thesis was to establish the anatomical and physiological changes in the major output nucleus of the cerebellum, the deep cerebellar nuclei (DCN), in the 6-hydroxydopamine (6-OHDA) rat model of PD. Furthermore, the downstream consequences of cerebellar pathology on the motor thalamus were investigated to begin to understand how cerebellar dysfunction may be contributing to dysfunctional movement in PD.
To investigate anatomical changes in the DCN, immunohistochemical labelling of GABAergic and putative glutamatergic neurons using antibodies against glutamic acid decarboxylase 67 (GAD67) and calmodulin-dependent protein kinase type II alpha chain (CaMKIIα), respectively, was performed in the DCN of 6-OHDA and sham-lesioned rats. Neuronal numbers were stereologically quantified using the Cavalieri’s and optical disector methods. Results demonstrated a significant reduction in GAD67+ and CaMKIIα+ neurons in the DCN of 6-OHDA rats. CaMKIIα+ DCN neuronal numbers in 6-OHDA rats showed a significant relationship with behavioural motor performance. CaMKIIα+ neuronal changes following dopamine depletion could have an important role in PD movement deficits.
To investigate if the altered cellular composition of the DCN was accompanied by physiological activity changes, single unit and local field potential (LFP) recordings were performed in urethane anesthetised 6-OHDA and sham-lesioned rats. DCN neuronal firing rate was significantly reduced in 6-OHDA lesioned rats. The incidence of bursting was significantly increased. Furthermore, the timing of spiking and burst onset was significantly earlier with respect to the LFP. These results could have significant consequences on movement as the timing and pattern of DCN firing correlates with the timing of behavioural responses and muscle contraction.
To begin to understand the potential consequences of altered anatomy and physiology of the DCN on downstream motor programming, light stimulation of the DCN was performed while simultaneously recording single-unit and LFP activity in the ventrolateral (VL) thalamus in urethane anesthetised 6-OHDA and sham-lesioned rats. The lentiviral vector CaMKIIα-ChR2(h124R)-mCherry was injected into the DCN to induce selective expression of channelrhodopsin in glutamatergic neurons, which project to the VL thalamus. Activity of the VL thalamus was significantly altered in 6-OHDA rats. Baseline recordings demonstrated reduced general and low threshold calcium spike (LTS) bursts in the VL thalamus of 6-OHDA rats. Light stimulation of the DCN did not elicit significant short latency responses in VL thalamus neurons, which contradicts previous anatomical and physiological evidence that predicts the cerebellum is a driver of motor thalamus activity. During the light stimulation period, the firing rate was reduced and general burst firing increased in VL thalamus neurons, compared to baseline recordings. Further research is required in awake behaving animals to understand the consequences of DCN stimulation effects on motor behaviour.
For the first time, significant changes in neuronal number and physiological activity were shown in the DCN of 6-OHDA rats. Results would suggest these changes have downstream effects on VL thalamus activity. These findings support cerebellar pathology contributing to PD motor symptoms through altered activity in the VL thalamus. Further research is required to fully understand the role that the cerebellum has in altered VL thalamus activity and how this is associated with PD movement deficits.