From 2018 to 2021, 30 participants with mild to moderate idiopathic PD were recruited from a Movement Disorders Clinic at the Medical University of South Carolina (MUSC) to undergo either taVNS (n = 15) or sham (n = 15) stimulation (Figure 1). The length of participant recruitment was longer than anticipated due to the impact of COVID-19 on in-person visits. Study procedures were reviewed and approved by the MUSC Institutional Review Board and the study was registered on ClinicalTrials.gov (NCT04157621). All participants enrolled in the study were informed of study procedures and provided written consent. Study procedures included an initial screening visit, followed by 10 visits for stimulation over a period of 2 weeks (Figure 1). One week following the final stimulation visit, a follow-up safety assessment was performed. Assessments of motor efficacy were performed in the OFF-medication state in order to avoid any influence of medication response fluctuations on the MDS-UPDRS Part III. Cognitive measures on the contrary were elicited in the ON medication state in order to avoid lack of effort, bradyphrenia, anxiety, and depression, which can be associated with OFF periods, interfering with cognitive assessments.

Randomization of the 30 participants was performed using the REDCap randomization module in which there was a 1:1 ratio in group assignment (taVNS vs. sham stimulation). All participants and study staff were blinded to which treatment assignment with exception of the Principal Investigator and laboratory personnel performing active or sham stimulation. To maintain objectivity of motor assessments a blinded movement disorders neurologist rated videotaped MDS-UPDRS III examinations.

There are no prior studies from which to estimate treatment or placebo effects associated with taVNS in PD. This study was powered to detect a clinically meaningful reduction of 3.25 points on the MDS-UPDRS Part III (29), assuming a small and statistically non-significant change of 1 on the MDS-UPDRS Part III with sham taVNS, and a pooled standard deviation of 2.0. At 80% power and alpha = 0.05, using a two-tailed t-test for difference in means, the total sample size required was 26 (13 per group). Additional participants were recruited to account for attrition.

Individuals meeting UK Brain Bank diagnostic criteria for Parkinson's disease (30) who were between 40 and 79 years of age, taking levodopa three or more times daily, with Hoehn and Yahr staging between 2 and 3 were eligible to participate. Exclusion criteria included prior diagnosis of dementia (31) or mild cognitive impairment on screening [MOCA < 24, (32, 33)], visual hallucinations or other psychotic symptoms, history of ear trauma or facial pain disorder, history of comorbid neurologic disorders or major cardiovascular conditions, history of deep brain stimulation or other brain surgery, neurogenic orthostatic hypotension, chronic respiratory illness, pregnancy, and use of cholinesterase inhibitors or Level 2 and 3 anticholinergic medications (34). All participants were expected to be stable on medication for PD motor and non-motor symptoms for a minimum of 30 days before and for the duration of the trial.

Active taVNS was applied using custom fabricated ear clip electrodes (1 cm round electrode surface) designed to deliver electrical stimulation to the anterior wall of the left outer ear canal landmarked by the tragus (Figure 2). The reasoning for left ear stimulation is based on conventional, surgically implanted VNS trials which primarily target the left cervical bundle of the vagus nerve. In addition, the afferent effects of taVNS have been primarily established in a left-only fashion (35–37). Sham stimulation utilized the same stimulation parameters; however current was delivered to the left earlobe, a region with limited to no innervation of the ABVN. Electrodes were applied to the ear using a conductive Ten20 paste and connected to an FDA 510 k-cleared constant current electrical nerve stimulation device.

After participants were connected to the stimulation system, they remained supine and were instructed to stay awake and maintain still in a comfortable position. Stimulation was administered for 1-h/day for 10 total days spread over 2 weeks. The stimulation parameters were consistent with prior work suggesting activation of vagal afferent network (36): Pulse Width 500 μs; frequency 25 Hz; duty cycle 60 s On, 30 s OFF; current intensity 200% perceptual threshold (Figure 2). Perceptual threshold was defined by the minimum amount of current required to be perceived by the participant.

Safety and tolerability of taVNS stimulation included monitoring of participant reported adverse events, MDS UPDRS Part III examinations, cognitive testing, and the Columbia-Suicide Severity Rating Scale (C-SSRS). Monitoring of vital signs (heart rate and blood pressure) occurred pre-stimulation, 10 min into stimulation and 50 min into stimulation. Neurogenic orthostatic hypotension (NOH) was reported as an adverse event if it was clinically significant. Clinically significant NOH was defined as lightheadedness or syncope in the setting of a 20 mm Hg or more drop in systolic BP or 10 mm Hg or more drop in diastolic BP with standing at 50% or more of assessments. An adverse event was considered related to the study intervention if the event had a reasonable possibility of being causally related to the intervention being administered. Qualitative interviews following study participation were performed to assess participant experiences during the study and feasibility of developing taVNS as a clinical intervention.

As an early indicator for treatment efficacy, we assessed motor function with the Movement Disorder Society Unified Parkinson's Disease Rating Scale (MDS-UPDRS) Part III, a comprehensive scale that rates the severity of motor symptoms in Parkinson's Disease, modified for videotaped assessments (rigidity testing omitted) (38). Both acute (pre to post stimulation change on a given visit) and subacute (baseline to visit 5 and visit 10 change) assessments were evaluated. All MDS-UPDRS Part III evaluations were performed in the OFF-medication state (defined as at least 12 h without PD medication) to capture the standalone benefits of stimulation independent of dopaminergic medication. MDS-UPDRS Parts I, II, and IV were used to evaluate changes in motor and non-motor aspects of daily living and motor complications. Other secondary outcome measures focused on changes in cognitive function as measured by the Delis-Kaplan Executive Function System (DKEFS) to evaluate letter fluency, category fluency and category switching (39) and the Digit Span (Backward and Forward) assessment to evaluate short-term and working memory (40). Cognitive secondary outcome measures were obtained in the medication ON-state at baseline and visit 9. Patient reported outcomes included the Movement Disorders Society Non-Motor Symptoms Scale for Parkinson's Disease (NMSS), the freezing of gait questionnaire (FOG-Q) (41, 42), and the Conners Adult ADHD Rating Scale short form self-report (CAARS-S:S). The CAARS-S:S served as a patient reported outcome of symptoms associated with inattention and executive dysfunction (43, 44). Additionally, the Patient-Reported Outcomes Measurement Information System (PROMIS) was used to evaluate sleep related impairment (PROMIS-Sleep Related Impairment), applied cognitive abilities (PROMIS-Applied Cognition) and fatigue (PROMIS-Fatigue) (45). Self-reported questionnaires and outcome measures were collected at screening and visit 9.

Demographic data, disease severity measures, motor scores at baseline and non-motor scores at baseline were compared between the two treatment groups (active vs. sham taVNS) using chi-squared and two-sided two-sample t-tests. Treatment group differences were considered significant if p-value < 0.05. To assess the effects of taVNS on systolic blood pressure, diastolic blood pressure and heart rate, the change in score before and after 50 min of treatment was compared between the taVNS and sham group using a two-sample t-tests. Data was aggregated from 112 sham sessions and 122 taVNS sessions where complete vital datasets were available. To assess the effects of taVNS on motor (MDS-UPDRS Part III scores) and cognitive outcome measures, the change in score was compared between the active and sham groups using a two sample-sample t-test. Acute effects of taVNS on MDS-UPDRS Part-III scores were evaluated as post-stimulation scores minus pre-stimulation scores at a given visit. Meanwhile subacute effects of taVNS on MDS-UPDRS Part-III scores were evaluated as post-stimulation scores at visit 10 minus pre-stimulation scores at visit 1. The motor outcome measure was considered significant if p-value < 0.05. Cognitive outcome measures were considered significant if p-value < 0.01 (Bonferroni corrected for the five cognitive scores evaluated). To address missing data statistical analyses were performed by excluding participants with missing data and were then repeated using imputation of missing data points.

Table 2 displays the demographics, disease severity and baseline motor and cognitive scores of participants in the taVNS (n = 15) and sham (n = 15) treatment groups. Fifty percent of participants were female and the mean age was 67. Mean time since symptom onset was 7.7 years (median: 6.5) and mean time since PD diagnosis was 5.1 years (median: 4). Treatment groups did not significantly differ in any demographic characteristics, baseline motor symptom severity (MDS-UPDRS Part-III score active group 25.9 ± 10.3 and sham 24.5 ± 7.3), or baseline cognitive function. Stimulation intensity determined by each participant's sensory threshold was greater in the taVNS group (t = 3.32, df = 28, p-value = 0.0025) than the sham group. Sensory thresholds ranged from 1.4 to 3 in the taVNS group and 0.8–2 in the sham group (Supplementary Figure 1).

All 30 participants in the taVNS and sham treatment groups completed the on-site stimulation visits. The frequency of visits to the study center for stimulation was reported as burdensome by participants via qualitative interviews following trial participation. taVNS administration was well-tolerated and feasible without technical issues at the study site. As far as safety, during the 10-day period of stimulation, five (33.3%) participants in the active taVNS group and 3 (20%) participants in the sham group self-reported one or more minor adverse event (AE). The most frequently reported AE in the active taVNS group was difficulty sleeping (n = 2) followed by lightheadedness (n = 1), fatigue (n = 1), nausea (n = 1), ringing in the ear (n = 1), grinding teeth (n = 1), fluid in the ear (n = 1), jitteriness/anxiousness (n = 1), and vertigo (n = 1). The most frequently reported AE in the sham group was lightheadedness (n = 2) followed by difficulty sleeping (n = 1), headache (n = 1), fatigue (n = 1), difficulty concentrating (n = 1), and neck pain (n = 1). No pain at the site of stimulation was reported for either treatment group. No serious adverse events (SAEs) were reported for the duration of the study and follow-up. At the 1-week follow-up, 2 participants from the active taVNS group reported AEs including stomach and hip pain, difficulty sleeping, fatigue, and constipation. At follow-up, 1 participant from the sham group described decreased hearing within the left ear. There was no evidence for the development of suicidal thoughts as monitored by the Columbia-Suicide Severity Rating Scale (C-SSRS) (46). Vital signs were monitored and remained stable throughout the duration of the study. Systolic blood pressure (t=0.495, df=232, p=0.621), diastolic blood pressure (t=1.374, df=232, p=0.171) and heart rate (t=0.183, df=232, p=0.138) changes from pre-treatment to 50 minutes into treatment did not differ between the sham and taVNS groups. In the taVNS group there was a 3.6 mm Hg drop in systolic blood pressure and a 0.9 mm Hg drop in diastolic blood pressure and a 3.4 bpm drop in heart rate. Meanwhile in the sham group there was a 2.7 mm Hg drop in systolic blood pressure, a 0.7 mm Hg increase in diastolic blood pressure and a 1.8 bpm drop in heart rate. Orthostatic hypotension was assessed throughout the trial was identified to be clinically significant in 1 (6.6%) participant receiving active taVNS and 1 (6.6%) participant receiving sham stimulation.

Figure 3 displays individual participant responses with regard to motor symptoms. A detailed summary of results is displayed in Table 3. Participants with missing MDS-UPDRS Part III data at visits 1, 5, and 10 were excluded from the analysis (see Table 3 for sample sizes used in the analysis). Analyses were repeated using imputation of missing data points; however, this is not shown since it did not influence significance of the reported results. Acute effects of stimulation did not significantly differ between groups at any of the timepoints (visits 1, 5, and 10) assessed (p-value > 0.05). Both treatment groups showed small reductions in their MDS-UPDRS Part-III scores (2.0 ± 5.6 points reduction in the taVNS group and 2.2 ± 4.3 points reduction in the sham group); however, there were no significant treatment group differences in subacute score changes (p-value = 0.906).

Cognitive, and patient reported outcomes are presented in Table 3. Cognition: As described for the motor outcome, participants with missing DKEFS (letter fluency, category fluency and category switching) data, and missing Digit Span (Backward and Forward) data were excluded from the analysis (taVNS group n = 1, sham group n = 1). Effects of taVNS on cognitive performance were evaluated by determining treatment group differences in change scores (post-stimulation at visit 9 minus pre-stimulation at baseline). DKEFS letter fluency performance decreased in the taVNS group by 3.8 points but increased in the sham group by 5.4 points. Similarly, DKEFS category fluency performance decreased in the taVNS group by 3.1 points but increased in the sham group by 3.4 points. Change scores in letter fluency (p-value = 0.008, uncorrected) and category fluency (p-value = 0.008, uncorrected) was significantly reduced in the taVNS group relative to the sham group. After Bonferroni correction accounting for number of cognitive tests performed (p-value threshold of 0.01), however, only group differences in category fluency remained significant. DKEFS category switching performance improved in the taVNS group by 1.3 points and was reduced in the sham group by 0.3 points. The Digit Span forward and backward scores increased slightly in the taVNS group (0.2 and 0.1 points, respectively) and increased slightly in the sham group (0.7 and 0.5 point, respectively). There were no significant treatment group differences in change scores for DKEFS category switching, or the digit span forward and backward. Patient reported outcomes: No significant changes from screening to visit 9 were observed for the NMSS, the freezing of gait questionnaire, CAARS or the PROMIS questionnaires.