Background: Increased arterial stiffness is a risk factor for cardiovascular disease. Slow, deep breathing decreases blood pressure related to arterial stiffness. The objective of the present study was to determine the acute effects of a single session of slow breathing on arterial stiffness, blood pressure, and cardiac autonomic function. Methods: Fifteen healthy men (20 ± 0 years) were administered (a) a slow breathing condition (12 consecutive breaths of 4 s of inhalation, 4 s of pause, and 8 s of exhalation through the nose, approximately 5 min per breath) and (b) a control, two-condition crossover design. Carotid-femoral artery pulse wave velocity (cfPWV), brachial-ankle PWV (baPWV), brachial blood pressure, high frequency (HF) and low frequency (LF) were measured at baseline, 30 min, 60 min and 24 h after respiratory control. Results: Brachial-ankle PWV and brachial systolic pressure on the 4-4-8 breathing trial decreased after 30 min of respiratory control compared to baseline (p < 0.05), but did not change on the CON trial. Carotid-femoral PWV on both trials was unchanged; HF on the 4-4-8 breathing trial increased (p < 0.05) and LF decreased (p < 0.05) after 30 min of respiratory control compared to baseline, but was unchanged on the CON trial. Conclusions: These results suggest that slow breathing techniques may be effective in modulating autonomic function and improving arterial stiffness in healthy young adults.