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Fig. 3 | Molecular Brain

Fig. 3

From: SCAMP5 mediates activity-dependent enhancement of NHE6 recruitment to synaptic vesicles during synaptic plasticity

Fig. 3

Forskolin-induced cLTP does not change glutamatergic quantal release at individual presynaptic boutons in cultured hippocampal neurons. a Representative images of cultured hippocampal neurons co-transfected with DsRed and iGluSnFR. Images of iGluSnFR represent resting condition and in the response of glutamate released spontaneously. Scale bar: 5 μm. b Representative traces of the iGluSnFR signal during spontaneous glutamate release at one presynaptic bouton. c Amplitude histogram graph of spontaneous iGluSnFR response, n = 135 (boutons). The graph was fitted with a single Gaussian distribution. d, h Representative images of cultured hippocampal neurons co-transfected with mCherry-synaptophysin (SYP) or DsRed-shRNA SCAMP5, and iGluSnFR. Response signal of iGluSnFR due to spontaneous glutamate release was obtained before and after treatment of DMSO or Forskolin, respectively. Scale bar: 5 μm. e, i Representative traces of individual response (thin colored lines) and average trace (thick colored lines) of the iGluSnFR signal during spontaneous glutamate release at one presynaptic bouton before and after treatment of DMSO or Forskolin, respectively. f, g Changes of normalized peak intensity of iGluSnFR due to spontaneous glutamate release before and after treatment of DMSO or Forskolin in control neurons. Before (DMSO), 0.316 ± 0.226 ΔF/F0, n = 20 (boutons, 3 coverslips); After (DMSO), 0.315 ± 0.225 ΔF/F0, n = 20 (boutons, 3 coverslips). p = 0.954, Before (Forskolin), 0.322 ± 0.177 ΔF/F0, n = 20 (boutons, 3 coverslips); After (Forskolin), 0.333 ± 0.182 ΔF/F0, n = 20 (boutons, 3 coverslips). p = 0.075, analyzed paired samples t-test. j, k Changes of normalized peak intensity of iGluSnFR due to spontaneous glutamate release before and after treatment of DMSO or Forskolin in SCAMP5 KD neurons. Before (DMSO), 0.237 ± 0.113 ΔF/F0, n = 24 (boutons, 3 coverslips); After (DMSO), 0.236 ± 0.104 ΔF/F0, n = 24 (boutons, 3 coverslips). p = 0.772, Before (Forskolin), 0.246 ± 0.129 ΔF/F0, n = 28 (boutons, 3 coverslips); After (Forskolin), 0.249 ± 0.130 ΔF/F0, n = 28 (boutons, 3 coverslips). p = 0.670, analyzed paired samples t-test. Values are indicated as mean ± SD. n.s. non-significance. *p < 0.05, **p < 0.01, ***p < 0.001

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