Background: Exposure to maternal stress in utero negatively impacts cognitive and behavioral outcomes of children born at term. The neonatal intensive care unit (NICU) can be stressful for preterm infants during a developmental period corresponding to the third trimester of gestation. It is unknown whether stress in the NICU contributes to adverse neurodevelopment among NICU graduates. The aim was to examine the association between salivary cortisol and early neurodevelopment in preterm infants. Methods: We examined the association between cortisol levels during the NICU hospitalization and subsequent performance on the NICU Network Neurobehavioral Scales (NNNS), estimating time-specific associations and considering sex differences. Results: Eight hundred and forty salivary cortisol levels were measured from 139 infants. Average cortisol levels were inversely associated with NNNS Regulation scores for both male and female infants (β = −0.19; 95% CI: −0.44, −0.02). Critical developmental windows based on postmenstrual age were identified, with cortisol measured <30 weeks PMA positively associated with Habituation and Lethargy scores (β = 0.63–1.04). Critical developmental windows based on chronological age were identified, with cortisol measured in the first week of life inversely associated with Attention score (β = −1.01 for females; −0.93 for males). Conclusions: Stress in the NICU at specific developmental time points may impact early preterm infant neurodevelopment. Impact: Stress in the neonatal intensive care unit can impact the neurodevelopmental trajectory of premature infants.The impact of stress is different at different points in development.The impact of stress is sexually dimorphic.