Fig. 5
Increasing ambient synaptic glutamate level in the excitatory neurons enables GNE-8324 potentiation of NMDAR EPSCs. a Bath application of the GLT-1 inhibitor DHK revealed significant potentiation of NMDAR EPSCs in the excitatory neurons. Sample traces of NMDAR EPSCs before and after GNE-8324 in the presence of DHK. Scale bars, 10 pA/100 ms. GNE-8324 potentiation was only observed in excitatory neurons following DHK treatment. *P (peak) < 0.05; **P (area) < 0.01, one-way ANOVA with Dunnett's correction, both compared to vehicle. Significant GNE-8324 potentiation was observed at 32 °C in the presence of DHK. b TBS in the excitatory neurons revealed significant GNE-8324 potentiation. (Left) Sample traces showing prominent GNE-8324 potentiation. (Right) Expanding the portion of EPSC traces around the TBS portion revealed increasingly larger baseline responses with successive TBS pulses (indicated by the arrow). Scale bars, 10 pA/100 ms (left); 20 pA/10 ms (right). TBS-induced NMDAR EPSC areas were significantly enhanced by GNE-8324, compared to vehicle. *P < 0.05, unpaired t test. c Bath-applied NMDA enabled GNE-8324 potentiation in the excitatory neurons. Sample traces of holding current after bath application of 0, 5, or 7 μM NMDA. Scale bars, 5 pA/100 ms. Sample traces showing significant GNE-8324 potentiation in the excitatory neurons in the presence of 7 μM NMDA. Scale bars, 10 pA/100 ms. Quantification of GNE-8324 potentiation in the presence of NMDA. For 5 μM NMDA, GEN-8324, 122.2 ± 9.05% (area); Veh, 92.33 ± 5.39 (area); **P (area) < 0.01; N = 10 (GNE-8324) and N = 7 (Veh). For 7 μM NMDA, GNE-8324, 163.3 ± 18.53% (area), 101.6 ± 17.94% (area); *P (area) < 0.05; N = 9 (GNE-8324) and N = 5 (Veh), unpaired t test. d GNE-8324 potentiated NMDAR responses in excitatory neurons in the absence of synaptic stimulation. Sample traces. Puffing of GNE-8324 (3 mM) is indicated by the arrow. Scale bars, 10 pA/1 s. Quantification of responses to puffing of GNE-8324. N = 4 (GNE-8324), 4 (DMSO), 4 (GNE-8324 in APV), and 4 (GNE-8324 after APV washout). **P < 0.01, paired t test. The same neurons are connected by lines. Error bars represent SEM
