We investigated if nonnutritive sucking (NNS) during heelstick procedures alleviates behavioral distress in neonates. In our NICU, 26 neonates without severe complications (mean Minde score 0.8, range 0–3), undergoing heelstick procedures at least twice a day, in the first 2 weeks of life, were enrolled in the trial (mean gestational age 33.9 weeks, range 26–39 weeks, mean birth weight 1,988.5 g, range 1,200–4,010 g, mean Apgar score at the first minute 6.7, range 4–10, at the fifth minute 8.5, range 6–10). Two heelpricks were performed in each neonate with NNS randomly assigned. Behavioral states, transcutaneous oxygen tension (TcPO2), heart rate, and respiratory rate were monitored before, during and after the heelstick procedures. Heelstick procedures lasted for a mean of 109 s (range 50–230 s) with NNS, and a mean of 128.8 s (range 20–420 s) without NNS. Compared with baseline, heart rate and behavioral distress increased and respiratory rate decreased during heelstick and after heelstick. Oxygen tension did not change. Nonnutritive sucking had no effect on respiratory rate or transcutaneous oxygen tension, but reduced the time of crying and the heart rate increase during the procedure. In conclusion, NNS can be recommended to reduce distress in newborns undergoing invasive routine procedures. Further studies are needed to evaluate the effects of NNS on respiratory rate and blood gas levels.