OBJECTIVES: Evaluate physiological and behavioral parameters to verify if newborns feel pain when submitted to gastric suctioning. METHODS: 50 healthy newborns over 33 weeks gestational age, weighing more than 1999g, were submitted to gastric suctioning, randomly selected, and divided into two groups: Friction/Lancing and Lancing/Friction. The newborns were assessed through the Neonatal Infant Pain Scale (NIPS) (0-7, pain>3) by two independent observers in three different moments: one minute before, during, and one minute after gastric suctioning, heel lancing and foot friction, with simultaneous monitoring of heart rate, respiratory frequency, and hemoglobin oxygen saturation. RESULTS: The respiratory frequency only decreased during gastric suctioning (P=0.004). The heart rate was lower during gastric suctioning (P<0.0001), during the heel lacing procedure in the Friction/Lancing group (P=0.01), and during friction in the Friction/Lancing group (P=0.022). The hemoglobin oxygen saturation suffered no alterations during the three procedures. The results obtained through NIPS revealed that both groups felt pain during gastric suctioning and heel lancing, and did not feel pain during friction (P<0.0001). CONCLUSION: During the gastric suctioning procedure, newborns responded as if it were a painful stimulus. Physiological alterations were neither specific nor sensitive to pain assessment in newborn infants.