Functional Ultrasound imaging (fUS) has recently demonstrated the feasibility of assessing functional connectivity (fC) in neonates at bedside1, facilitating brain monitoring in this vulnerable population. Here, we validate fUS in a larger cohort of preterm neonates to investigate fC in a music exposure paradigm for which MRI studies have shown improved structural and functional maturation2,3. Our study involved n=21 preterm neonates (27.6±2.2 gestational week (GW)) born at Geneva University Hospitals : a control group with standard care (n=10) and a second group repeatedly exposed (2 to 3 times a day) to a 8-minutes long music from 33 GW (n= 11). fUS acquisitions were performed at 37GW using the Aixplorer ultrasound system and a 6.5MHz custom probe maintained over the fontanel with an adapted headset. The 24-minute 1s-resolution fUS sequence enabled to measure cerebral blood volume (CBV) fluctuations in a coronal plane. Connectivity matrices were built from correlation between mean CBV time courses of defined functional area. To refine spatial pattern of fC we also investigated homotopic connectivity. The coefficients of the mean connectivity matrices are higher for the music group (p<0.02) suggesting long-term global connectivity enhancement. Homotopic connectivity shows stronger correlation between 1 and 2 mm depth, which could correspond to gray matter. These results show that fUS is able to detect subtle fC modifications and holds promise for monitoring brain development during neonatal care.1. Baranger J, et al. Nat Commun. 20212. Sa de Almeida J, et al. Neuroimage. 20193. Lordier L, et al. PNAS 2019