Introduction: Optimism has been associated with reduced cardiovascular (CV) risk, while cynical hostility has been linked with worse CV risk. The pathophysiological mechanism(s) underlying these associations are largely unexplored. We sought to examine associations between optimism, hostility, and heart rate variability (HRV). Hypothesis: Greater optimism will be associated with higher HRV, while greater hostility will be associated with lower HRV. Methods: Participants (n=3,372) enrolled in the Myocardial Ischemia and Migraine Study, a Women’s Health Initiative (WHI) ancillary study, underwent a 24-hour ambulatory ECG monitoring (AECG) 3 years after WHI enrollment. Optimism (Life Orientation Test-Revised), cynical hostility (Cook-Medley questionnaire), demographics, and CV risk factors were assessed at baseline. HRV measures included standard deviation of all analyzed N-N intervals (SDNN); standard deviation of average N-N interval over 5 min (SDANN); and average heart rate (HR). Multivariable linear regression models examined the relationship between optimism, hostility and HRV indices adjusted for demographics and coronary risk factors. Results: After excluding women with previous CV disease, missing data, and AECG <12 hours, 2655 were included in the analysis. At baseline, more optimistic women were more likely to be younger, white, educated, non-smokers, physically active, and to have lower systolic blood pressure (SBP). More hostile women were more likely to be older, Hispanic or African American, less educated, diabetic, obese, smokers, and to have high cholesterol and high SBP. We found a significant inverse association between hostility and HRV (SDANN: adjusted β= -0.54; 95% CI -0.97, -0.11; SDNN: -0.49; 95% CI -0.93, -0.05). Optimism was not associated with HRV indices (SDANN: adjusted β= 0.10; 95% CI -0.25, 0.44; SDNN: 0.03; 95% CI -0.32, 0.39). Average HR was not associated with optimism or hostility. Conclusions: A worse autonomic balance may play a role in the physiopathology of unfavorable CV outcomes observed among women with higher cynical hostility. The lack of association between HRV and optimism suggests that other (behavioral?) factors may play a role in the protective association between optimism and CV outcomes.