•Neonatal social isolation in mice increased PSD-95-negative immature spines of the layer 2/3 pyramidal neuron in the somatosensory cortex (SSC).•Neonatal social isolation in mice didn’t alter spine dynamics of the layer 2/3 pyramidal neurons in the SSC.•Socially isolated mice showed modest change in dendritic morphology and spine structure of the layer 2/3 pyramidal neurons in the SSC. Social isolation during the juvenile period is postulated to leave specific sequelae, such as attention deficits and emotion recognition. Miswiring of the cortical neuronal circuit during postnatal development may underlie such behavioral impairments, but the details of the circuit-level impairment associated with social isolation have not yet been clarified. In this study, we evaluated the possibility that environmental factors may induce alternation in spine characteristics and dynamics. We isolated mice from the mother and siblings from postnatal day 7 to 11 for 6 h per day. Both dynamics and structural properties of spines in the layer 2/3 pyramidal neurons of the somatosensory cortex were measured at postnatal 3 weeks by in vivo two-photon microscopy. We found decrease in the ratio of PSD-95-positive dendritic spines in the mice after social isolation. These mice did not show alteration in spine dynamics. Those results suggest that the neonatal social isolation results in less mature spines, with normal rate of their turnover, which is distinct from spine phenotype seen in multiple models of autism spectrum disorders.