Research conducted in the Pharmacology and Physiology of the Substantia Nigra and Basal Ganglia Project over the past year in the Neurophysiological Pharmacology Section has continued to focus on the sources and consequences of changes in basal ganglia function in Parkinsons disease (PD). We are currently engaged in completing four manuscripts under this project, and are involved in two collaborative studies. These endeavors have taken advantage of a strategy developed in our lab for studying ongoing activity in motor circuits while the rat performs a continuous walking task. We can quantitate unilateral changes in gait in conjunction with LFP and spiking activity through the use of a circular treadmill. A paddle is lowered over the rotating circular track to encourage the rat to keep walking. The rats ability to walk in the opposite contralateral direction to the dopamine cell lesion with his affected paws on the inside of the track is typically very limited and provides a read-out of the motor disability. We can monitor basal ganglia circuit spiking and local field potential (LFP) activity continuously from the intact and lesioned hemispheres as the animal walks and rests. A major accomplishment this year has been the completion and recent successful submission of Heysol Burmudezs PhD thesis (Brown Univ. NIH GPP). The goal of this study was to explore the role of the external segment of the globus pallidus (GPe) in mediating the increases in oscillatory activity in the subthalamic nucleus (STN) in the hemiparkinsonian rat model. It has only been relatively recently appreciated that there are different subtypes of GABAergic neurons in the GPe, and that their interconnections with each other and with the reciprocally connected STN provide a possible basis for generating the abnormal beta range oscillations emerging in this network after loss of dopamine. A previous postdoc (Ana Cruz) explored the logical idea that these oscillations are transmitted from the cortex through the striatum and globus pallidus to the STN and back to the cortex. However, in that study, which Ana Cruz is currently writing up for publication, although the exaggerated beta range LFP rhythms do appear in LFPs recorded from some parts of the striatum, we failed to find correlated spiking in the striatal output neurons in the parkinsonian rats. Thus, the hyperdirect pathways from cortex to STN and GPe-STN subcircuits became critical sites to probe with respect to these questions. Unfortunately, Heysols study showed that the most promising chemogenetic tools viruses delivering Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) failed to effectively infect the desired GABAergic subpopulations (CRE-expressing parvalbumin GABAergic neurons) projecting from the GPe to the STN in the rat. These failures are important to note, as a cautionary tale, as others have also found these tools more difficult it use in rat models than in mice. Heysols study did, however, provide a novel, more precise characterization of the dynamic and transient nature of the exaggerated beta oscillatory activity in the rodent parkinsonian model, and insight into correlations, and lack thereof, between power in the high beta range and improvements in motor function related to the function of GABAergic inputs into the STN. Traditionally, LFP data is averaged across trials and subjects, giving the impression of continuously robust beta power during a given behavioral state. However, on a trial-by-trial basis, beta power recorded from human patients has been shown to emerge as transient bursts (Tinkhauser. G, et al. Brain, 2017) which may be useful for closed loop feedback control of deep brain stimulation and information about underlying pathology. Analysis of the LFP activity recorded from the MCx of the lesioned hemisphere of our hemiparkinsonian rats showed that during circular treadmill walking, the rats display bursts of activity similar to those seem in human patients; the distribution of beta bursts and their features periodically fluctuate as the animals walk. Specifically, bursts are less likely to occur around the time that an animal exhibits a step with the hindlimb contralateral to their lesioned hemisphere. These results suggest that desynchronization of beta activity coincides with initiation of hindlimb stepping. Interestingly, Heysols study showed that infusion of GABA antagonist picrotoxin into the STN during treadmill walking had no significant effects on total beta power; however, this treatment did induce significant modulation of beta burst features coincident with improved motor function. Infusion of the GABA agonist, muscimol, however, resulted in a significant reduction of high beta total power. These results show that amelioration of motor deficits can be induced by both blockade of GABA-A input and stimulation of GABA-A receptors in the STN, but this effect does not require a robust reduction of beta power in the awake, hemi-parkinsonian rat model. These results support the view that beta burst duration is an effective biomarker for parkinsonian dysfunction and are consistent with observations in parkinsonian patients showing that adaptive deep brain stimulation of the STN both ameliorates motor dysfunction and reduces the average duration of beta bursts. This study is currently being written up for publication and will be presented as a poster at the Society for Neuroscience meeting in Nov. 2018. Another study close to submission describes the changes in high beta range oscillatory activity over the first week following 6-hydroxydopamine induced dopamine cell lesion in the rat hemiparkinsonian model. This study shows motor deficits are fully evident within hours after the 6-OHDA-mediated dopamine cell lesion in hemiparkinsonian rats, as the rat fails to walk effectively in the contraversive direction on the treadmill and does poorly on stepping tests, while increases in power in LFP activity in motor cortex and substantia nigra emerge more slowly over the first days post lesion. A second strategy for studying the effect of loss of dopamine receptor stimulation, treatment twice daily with D1/D2 dopamine receptor antagonists, provided similar results. Thus, this study also shows the emergence of the high beta oscillatory power, per se, is not well correlated with the expression of catalepsy. Notably, by the first hours post lesion, when the rats do show clear difficulty walking, the peak LFP frequency in the motor cortex shifts from 40 Hz toward the 30 Hz range, initiating the emergence of coherence in this frequency range with respect to spiking and LFPs in substantia nigra and motor cortex. This results call for better understanding of the spike pattern alterations initiating the shift in cortical peak frequencies that best correlate with the early emergence of motor deficits. Two exciting collaborative studies have been underway during the past year. We are currently testing a novel dopamine D3 receptor agonist in our Parkinsonian rats in collaboration with Dr. Sibleys lab. It has previously been difficult to conduct such a study as drugs with affinity for D3 receptors are typically effective at D2 receptors. This newly developed drug, will show, for the first time, whether an agent highly selective for the D3 dopamine receptor subtype will reduce motor deficits without inducing dyskinesias, a common side effect of drugs used to treat Parkinsons disease. In addition, collaborative studies with Dr. Alexandra Nelson, Johathan Shor and Dr. du Hoffmann are underway, examining the open question of whether the mouse model of hemiparkinsonism shows similar beta range oscillatory activity during walking. To date, preliminary studies suggest there is a distinct difference between LFP patterns in the mouse and rat models.
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