(A) Images of newly formed gephyrin puncta (green arrowheads) and dendritic spines (pink arrowheads) in vitro and in vivo. (B and C) Success rate of de novo gephyrin and spine formation by GABA and glutamate HFU in vitro [(B); GABA, n = 29 trials, 16 cells; glutamate, n = 24 trials, 12 cells; GABA in younger, n = 17 trials, 7 cells; GABA in old, n = 36 trials, 16 cells] and in vivo [(C); GABA, n = 61 trials, 55 cells, 5 mice; mock stimulation, n = 52 trials, 45 cells, 5 mice; spontaneous, n = 71 trials, 52 cells, 6 mice]. (D) Summary plots of distance-dependent de novo gephyrin and spine formation. (E) Time course for size changes of new gephyrin puncta (n = 18) and spines (n = 24). (F) Time-lapse images of GABA HFU–induced gephyrin puncta and spines. (G) Stability of newly formed gephyrin puncta (12 of 15) and dendritic spines (15 of 24). **P < 0.01; n.s., not significant. Error bars in (E) denote SEM.