This study examined whether neural processing of infant cry sounds changes across the transition to fatherhood (i.e., from the prenatal to postnatal period), and examined whether an interaction-based prenatal intervention modulated these changes. Furthermore, we explored whether postnatal activation in brain regions showing transition or intervention effects was associated with sensitive care and involvement. In a randomized controlled trial, 73 first-time expectant fathers were enrolled, of whom 59 had at least 1 available fMRI scan. Intervention and transition effects on cry processing were analyzed in the amygdala and superior frontal gyrus (SFG) using linear mixed effect models with all available data and with intent-to-treat analyses. Further, exploratory whole-brain analyses were performed. ROI analyses suggest that the transition to fatherhood is characterized by decreasing activation in response to cry vs control sounds in the amygdala but not SFG. Exploratory whole-brain analyses also show a decrease in activation over the transition to fatherhood in the sensorimotor cortex, superior lateral occipital cortex, hippocampus, and regions of the default mode network. In the putamen and insula, larger decreases were found in fathers with more adverse childhood caregiving experiences. In regions showing transitional changes, higher postnatal activation was associated with more concurrent parenting sensitivity. No effects of the intervention were found. The decrease in activation from the pre- to postnatal period may reflect fathers’ habituation to cry sounds over repeated exposures. The positive association between postnatal neural activation and paternal sensitive care suggest that continued sensitivity to cry sounds may be conducive to parenting quality.