Investigating the mechanisms underlying cholinergic regulation of dopamine release

Dopamine (DA) is a critical regulator of movement and motivation and disruptions to its signalling underlie various psychomotor disorders, including Parkinson’s disease (PD) and addiction. DA neurons projecting from the midbrain to the striatum form extensive axonal arborisation which are subject to local neuromodulation, gating how action potential firing at the level of DA neuron soma is translated into axonal DA release. Unravelling these mechanisms is therefore fundamental to better understand DA regulation of striatal output in health and disease.
Acetylcholine (ACh) is one of the major local modulators of striatal DA release. ACh mainly originates from striatal cholinergic interneurons (ChIs) and dysregulation to striatal cholinergic signalling is also implicated in a range of neuropsychiatric and neurodegenerative disorders. ACh acts at nicotinic ACh receptors (nAChRs) localised on DA axons and modulates DA release in a multiplicity of ways, from directly driving DA release bypassing action potential firing by DA somata, to promoting short-term depression of DA released during burst stimulation. Despite this fundamental role, striatal ACh release dynamics are currently incompletely characterised, mainly due to a lack of tools for direct, real-time measurement of ACh release, only recently filled by the development of genetically encoded fluorescent sensors (GRABACh). In this thesis, I used GRABACh in mouse striatal slices to characterise the major properties of striatal ACh release, a fundamental step to help unravel the mechanisms underlying cholinergic modulation of DA release. I found that ACh in dorsal and ventral striatum is dynamically released according to activity in ChIs and that acetylcholinesterase (AChE) modulates the spatiotemporal dynamics of evoked ACh release and tonic ambient levels of ACh. Furthermore, I show that ACh release is modulated by GABA acting at GABAA and GABAB receptors and inhibited by endogenous DA acting at D2- receptors (D2Rs). I then investigated the complex relationship between changes in ACh release and how these can dynamically gate DA release via nAChRs, combining GRABACh sensors and FCV to measure ACh and DA release in mouse striatal slices. I found that the magnitude of ACh-driven DA release is sensitive to changes in ACh levels but insensitive to activity in ChIs. Furthermore, I found that nAChRs regulate short-term plasticity of DA but not ACh release and are not functionally associated with the calcium binding protein calbindin D-28K in DA axons. Finally, I’ve explored maladaptive alterations in striatal cholinergic signalling in a human a-synuclein overexpressing mouse model of early parkinsonism. In this model, we found lower ACh release and stronger regulation of tonic ACh by AChE. In the early parkinsonian model, we also reported stronger D2R-mediated regulation of ACh release and no changes in tonic GABAergic inhibition.
Overall, this thesis aimed at investigating the dynamic interplay between ACh and DA release in the striatum. The data here presented provide a novel characterisation of ACh release dynamics in health and early Parkinson’s disease, together with previously unappreciated aspects of the dynamic gating of DA release by ACh.