For many G-protein-coupled receptors (GPCRs), including cannabinoid receptor 1 (CB 1 R), desensitization has been proposed as a principal mechanism driving initial tolerance to agonists. GPCR desensitization typically requires phosphorylation by a G-proteincoupled receptor kinase (GRK) and interaction of the phosphorylated receptor with an arrestin. In simple model systems, CB 1 R is desensitized by GRK phosphorylation at two serine residues (S426 and S430). However, the role of these serine residues in tolerance and dependence for cannabinoids in vivo was unclear. Therefore, we generated mice where S426 and S430 were mutated to nonphosphorylatable alanines (S426A/S430A). S426A/S430A mutant mice were more sensitive to acutely administered delta-9-tetrahydrocannabinol (⌬ 9 -THC), have delayed tolerance to ⌬ 9 -THC, and showed increased dependence for ⌬ 9-THC. S426A/S430A mutants also showed increased responses to elevated levels of endogenous cannabinoids. CB 1 R desensitization in the periaqueductal gray and spinal cord following 7 d of treatment with ⌬ 9 -THC was absent in S426A/S430A mutants. ⌬ 9 -THC-induced downregulation of CB 1 R in the spinal cord was also absent in S426A/S430A mutants. Cultured autaptic hippocampal neurons from S426A/S430A mice showed enhanced endocannabinoid-mediated depolarization-induced suppression of excitation (DSE) and reduced agonist-mediated desensitization of DSE. These results indicate that S426 and S430 play major roles in the acute response to, tolerance to, and dependence on cannabinoids. Additionally, S426A/S430A mice are a novel model for studying pathophysiological processes thought to involve excessive endocannabinoid signaling such as drug addiction and metabolic disease. These mice also validate the approach of mutating GRK phosphorylation sites involved in desensitization as a general means to confer exaggerated signaling to GPCRs in vivo.

Enhanced neuronal synchronization of the subthalamic nucleus (STN) is commonly found in PD patients and corresponds to decreased motor ability. Coordinated reset (CR) was developed to decouple synchronized states causing long lasting desynchronization of neural networks. Vibrotactile CR stimulation (vCR) was developed as non-invasive therapeutic that delivers gentle vibrations to the fingertips. A previous study has shown that vCR can desynchronize abnormal brain rhythms within the sensorimotor cortex of PD patients, corresponding to sustained motor relief after 3 months of daily treatment. To further develop vCR, we created a protocol that has two phases. Study 1, a double blinded randomized sham-controlled study, is designed to address motor and non-motor symptoms, sensorimotor integration, and potential calibration methods. Study 2 examines dosing effects of vCR using a remote study design. In Study 1, we will perform a 7-month double-blind sham-controlled study including 30 PD patients randomly placed into an active vCR or inactive (sham) vCR condition. Patients will receive stimulation for 4 h a day in 2-h blocks for 6 months followed by a 1-month pause in stimulation to assess long lasting effects. Our primary outcome measure is the Movement Disorders Society-Unified Parkinson's Disease Rating Scale (MDS-UPDRS) part III off medication after 6 months of treatment. Secondary measures include a freezing of gait (FOG) questionnaire, objective motor evaluations, sensorimotor electroencephalography (EEG) results, a vibratory temporal discrimination task (VTDT), non-motor symptom evaluations/tests such as sleep, smell, speech, quality of life measurements and Levodopa Equivalent Daily Dose (LEDD). Patients will be evaluated at baseline, 3, 6, and 7 months. In the second, unblinded study phase (Study 2), all patients will be given the option to receive active vCR stimulation at a reduced dose for an additional 6 months remotely. The remote MDS-UPDRS part III off medication will be our primary outcome measure. Secondary measures include sleep, quality of life, objective motor evaluations, FOG and LEDD. Patients will be evaluated in the same time periods as the first study. Results from this study will provide clinical efficacy of vCR and help validate our investigational vibrotactile device for the purpose of obtaining FDA clearance.Clinical Trial Registration:ClinicalTrials.gov, identifier: NCT04877015.

Δ9‐THC and other cannabinoid agonists have been reported to induce rewarding effects and physical dependence in several different animal models. In this study, desensitization–resistant CB1 “knock‐in” mice were used to determine whether this “hyper‐sensitive” form of CB1 would cause more sever symptoms associated with precipitated withdrawal (physical dependence). These mice contain two point mutations in C‐terminus of the CB1 receptor where serines at residues 426 and 430 have been substituted for non‐phosphorylatable alanines (S426/430A). Withdrawal was measured in wild‐type, CB1 knock‐out and S426/430A mutant mice after administering two daily injections of 50 mg/kg Δ9–THC over a period of six consecutive days. Withdrawal symptoms were precipitated using 10 mg/kg of rimonabant, a selective inverse agonist for CB1. Behavioral manifestations of withdrawal (head shakes, paw tremors, jumps) were recorded and scored by a trained observer blind to genotype. Our results demonstrate a significant increase in paw tremors and jumps, two measures of cannabinoid withdrawal, in S426/430A mutant mice relative to wild‐type littermate controls. In addition, all symptoms of precipitated withdrawal are absent in mice lacking the CB1 receptor. Our results suggest that that dependence for Δ9– THC is enhanced in desensitization‐resistant S426/430A mutant mice.