Skip to main content
. 2011 Jul 25;108(32):E440–E449. doi: 10.1073/pnas.1104977108

Fig. 2.

Fig. 2.

Hevin-induced synapses are ultrastructurally normal but postsynaptically silent. (A) EM images of synapses formed between rat RGCs cultured in the presence of astrocyte feeder layer inserts (astrocytes, Left) or purified hevin (Right). Hevin-induced synapses appear ultrastructurally normal and resemble astrocyte-induced synapses. Pre, presynaptic; post, postsynaptic site. (Scale bar: 250 nm.) (B) Quantification of number of synapses formed onto cultured rat RGCs by EM. RGCs treated with astrocyte feeder layers or hevin formed three- to fivefold more synapses compared with RGCs cultured alone (*P < 0.05; n = 8 cells per condition, error bars indicate SEM). (C) Quantification of the number of synaptic vesicles per presynaptic terminal per EM section. Black bars represent the total number of synaptic vesicles per synapse per section. Gray bars represent the number of docked vesicles (within 50 nm distance of the active zone; P > 0.05, not significant; n = 15 synapses for hevin and n = 17 synapses for astrocyte condition; error bars indicate SEM). (D) Hevin-induced synapses are postsynaptically silent. Representative traces from whole-cell patch-clamp recordings of mEPSCs from RGCs cultured alone, with astrocytes, with 30 nM hevin, or with 100 nM SPARC. Only the RGCs cultured with astrocyte feeder layer inserts (i.e., astrocytes) exhibited an increase in postsynaptic events versus RGCs alone. Quantification of the frequency (E) and amplitude (F) of mEPSCs from RGCs cultured alone, with astrocytes, with 30 nM hevin, or with 100 nM SPARC (*P < 0.01; n = 12 cells per condition; error bars indicate SEM). (G) Cumulative probability plot of the amplitude of mEPSCs recorded from RGCs cultured alone (control), with astrocytes, with 30 nM hevin, or with 100 nM SPARC. Only the mEPSCs from RGCs cultured with astrocytes are larger than those under control conditions. Inset: Average waveforms of mEPSCs from RGCs cultured alone (control), with astrocytes, with 30 nM hevin, or with 100 nM SPARC (n = 12 cells per condition).