Functions and Pathogenic Mechanisms of DAGLB and the Endocannabinoid System in Parkinson's Disease

National Institute On Aging

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Abstract

Recently, we identified four loss-of-function mutations in the DAGLB gene in patients with familial early-onset recessive Parkinson’s disease (PD). Analysis of RNA sequencing data from dopaminergic neurons in the substantia nigra pars compacta (SNc)—a brain region critical for movement—revealed that DAGLB is the primary enzyme responsible for synthesizing 2-arachidonoyl-glycerol (2-AG), the most abundant endocannabinoid in the brain, in both human and mouse SNc neurons. These findings underscore the importance of DAGLB in maintaining normal motor function. To investigate how DAGLB dysfunction might contribute to PD, we reduced Daglb expression in mouse SNc neurons, resulting in lower 2-AG levels and impaired motor skill learning in the rotarod test. Conversely, pharmacological inhibition of 2-AG breakdown elevated 2-AG levels, enhanced neuronal activity and dopamine release, and improved motor performance. These results identify a novel mechanism by which DAGLB deficiency could contribute to PD pathogenesis and suggest that enhancing endocannabinoid (eCB) signaling, such as boosting 2-AG levels, may offer a potential therapeutic approach. To avoid the developmental compensations seen in traditional knockout models, we employed CRISPR/saCas9-mediated gene editing to selectively reduce Daglb expression in adult mouse SNc neurons. This approach enabled more accurate testing of motor skill learning and real-time tracking of 2-AG release via fiber photometry, allowing correlation with motor performance. To improve knockdown efficiency, we recently generated conditional Daglb knockout mice. By crossing these mice with strains expressing Cre in dopaminergic neurons, we can delete Daglb in all midbrain dopamine neurons or specifically in ALDH1A1+ neurons. This strategy is expected to produce more pronounced behavioral phenotypes. Because this also affects ventral tegmental area (VTA) neurons, we will perform non-motor behavioral assays to assess learning, motivation, and stress responses. In addition, we will generate mice lacking both Dagla and Daglb in dopamine neurons to determine the combined role of 2-AG synthesis in motor and non-motor functions. Given the established role of 2-AG in modulating neuroinflammation, we will also delete Daglb specifically in microglia and astrocytes to investigate its function in these immune-responsive brain cell types. Furthermore, we aim to examine the regulation of DAGLB activity. Notably, direct pathway spiny projection neurons (dSPNs) form specialized connections with ALDH1A1+ SNc neurons, potentially influencing their activity. We will test whether dendritic calcium elevations in these neurons trigger 2-AG production, reducing inhibitory inputs and enhancing firing. In summary, our studies provide novel insights into how DAGLB mutations may contribute to PD pathogenesis. Through innovative genetic, physiological, and behavioral approaches, we aim to define the role of 2-AG and endocannabinoid signaling in both motor and non-motor functions, paving the way for new therapeutic strategies targeting the ECS in Parkinson’s disease.