ImportanceInternalizing (anxiety and/or depressive) behaviors are prevalent in children born very preterm (24-32 weeks’ gestation). Procedural pain-related stress in the neonatal intensive care unit (NICU) is associated with long-term internalizing problems in this population; however, whether positive parenting during toddlerhood attenuates development of internalizing behaviors across childhood is unknown.ObjectiveTo investigate whether neonatal pain-related stress is associated with trajectories of internalizing behaviors across 1.5, 3.0, 4.5, and 8.0 years, and whether supportive parenting behaviors and lower parenting stress at 1.5 and 3.0 years attenuate this association.Design, Setting, and ParticipantsIn this prospective longitudinal cohort study, preterm neonates (born at 24-32 weeks’ gestation) were recruited from August 16, 2006, to September 9, 2013, with follow-up visits at ages 1.5, 3.0, 4.5, and 8.0 years. The study was conducted at BC Women’s Hospital, Vancouver, Canada, with recruitment from a level III neonatal intensive care unit and sequential developmental assessments performed in a Neonatal Follow-up Program. Data analysis was performed from August to December 2021.Main Outcomes and MeasuresParental report of child internalizing behaviors on the Child Behavior Checklist at 1.5, 3.0, 4.5, and 8.0 years.ResultsA total of 234 neonates were recruited, and 186 children (101 boys [54%]) were included in the current study across ages 1.5 (159 children), 3.0 (169 children), 4.5 (162 children), and 8.0 (153 children) years. After accounting for clinical factors associated with prematurity, greater neonatal pain-related stress was associated with more internalizing behaviors across ages (B = 4.95; 95% CI, 0.76 to 9.14). Higher parenting stress at age 1.5 years (B = 0.17; 95% CI, 0.11 to 0.23) and a less supportive parent environment (less sensitivity, structure, nonintrusiveness, nonhostility, and higher parenting stress; B = −5.47; 95% CI, −9.44 to −1.51) at 3.0 years were associated with greater internalizing problems across development to age 8.0 years.Conclusions and RelevanceIn this cohort study of children born very preterm, exposure to repetitive neonatal pain-related stress was associated with persistent internalizing behavior problems across toddlerhood to age 8.0 years. Supportive parenting behaviors during early childhood were associated with better long-term behavioral outcomes, whereas elevated parenting stress was associated with more child anxiety and/or depressive behaviors in this population. These findings reinforce the need to prevent pain in preterm neonates and inform future development of targeted parent-led behavioral interventions.