Heart failure is a debilitating and fatal condition characterised by sympathetic activation and parasympathetic withdrawal. Current therapies target sympathetic activation (e.g. beta-blockers) and have improved the clinical outcome of heart failure. However, the prevalence of heart failure is increasing, demanding the development of new therapeutic approaches. Tackling the underlying autonomic imbalance through parasympathetic stimulation can improve cardiac function in heart failure patients, however, the current method of cervical vagus nerve stimulation (VNS) is invasive and associated with side effects1. This study investigated the autonomic effects of a non-invasive method of VNS – transcutaneous stimulation of the auricular branch of the vagus nerve (tVNS) – in healthy humans (n=89; 49 female, 40 male; aged 20-66 years). The study was approved by the University of Leeds Ethics Committee. Heart rate (ECG), blood pressure and respiration were recorded continuously. Heart rate variability (HRV) was calculated using spectral analysis of beat-to-beat (R-R) intervals derived from ECG data. Low frequency (LF; 0.04-0.15Hz) and high frequency (HF; 0.15-0.40Hz) power were calculated. HF power represents parasympathetic/vagal modulation of heart rate and the ratio of low to high frequency power (LF/HF) can be used as an indicator of sympathovagal balance, such that a decrease in LF/HF indicates a shift in autonomic balance towards parasympathetic dominance. tVNS was applied at either low pulse width and pulse frequency (L-tVNS) or high pulse width and pulse frequency (H-tVNS). Data was analysed at baseline, during tVNS and during recovery. Repeated measures ANOVAs were used to analyse results (data presented as mean ± S.E.M). There was a significant difference in HRV response between L-tVNS and H-tVNS groups (p=0.045). L-tVNS had no significant effect on HRV (n=63) whereas HRV improved significantly in the H-tVNS group with a decrease in LF/HF ratio from 1.45 ± 0.22 at baseline to 1.21 ± 0.15 during H-tVNS (n=26; p=0.048). HRV improved in 49% of the L-tVNS group compared to 63% of the H-tVNS group. Baseline LF/HF was significantly higher in subjects whose HRV improved during L-tVNS or H-tVNS compared to those who did not (p=0.038 and p=0.012 respectively). Based on the results of this study, tVNS can improve HRV in healthy subjects although the effects may be underestimated due to the limitation of using subjects with healthy autonomic balance. This is supported by the fact that baseline LF/HF was significantly higher in responders to tVNS indicating that these subjects had higher sympathetic and/or lower parasympathetic activity at baseline. tVNS could be a practical, non-invasive and economical therapy for heart failure patients and warrants further investigation.