Nicotinamide adenine dinucleotide (NAD⁺) has emerged as a critical co-substrate for enzymes involved in the beneficial effects of regular calorie restriction on healthspan. As such, the use of NAD⁺ precursors to augment NAD⁺ bioavailability has been proposed as a strategy for improving cardiovascular and other physiological functions with aging in humans. Here we provide the evidence in a 2 × 6-week randomized, double-blind, placebo-controlled, crossover clinical trial that chronic supplementation with the NAD⁺ precursor vitamin, nicotinamide riboside (NR), is well tolerated and effectively stimulates NAD⁺ metabolism in healthy middle-aged and older adults. Our results also provide initial insight into the effects of chronic NR supplementation on physiological function in humans, and suggest that, in particular, future clinical trials should further assess the potential benefits of NR for reducing blood pressure and arterial stiffness in this group.