Presynaptic Fer-mediated eEPSCs. Paired neurons with soma–soma distance around 100 μm were selected for dual recording. Neurons expressing control (Vec) or Fer shRNA (shRNA) were identified by the expression of GFP, which is contained in the same constructs. Non-GFP–expressing neurons were considered as WT. Recordings were performed at 14–17 DIV. (A) Persistent eEPSCs at vector to WT synapses. Five constant stimuli (200 mV for 1 ms) delivered into the presynaptic cell (pre) reliably evoke robust EPSCs at the postsynaptic cell (post). The time interval between stimuli is 10 s. (B) Examples of eEPSCs at Fer shRNA to WT synapse. Unlike the control, stimuli with the same strength delivered at Fer shRNA neurons initiate only weak or no eEPSCs by neighboring WT neurons. (C) Substantial reduction of EPSC size by presynaptic Fer shRNA. Lines and circles indicate the average values from five stimulation trials. It is worth noting that postsynaptic knockdown of Fer (i.e., WT to Fer synapses) does not have a statistically significant effect on the eEPSC amplitude. (D) Increased variation on the EPSC amplitude at Fer shRNA to WT synapses. C.V., coefficient of variation. (E) Low success rate of the synaptic transmission rate at Fer shRNA to WT synapses. Synaptic transmission is defined as successful when an evoked response is five times larger than the root mean square of the noise (∼10–13 pA). n = 20 (vector→WT), 15 (shRNA→WT), and 8 (WT→shRNA). *, P < 0.025; **, P < 0.01; one-way analysis of variance. Error bars indicate SEM.