Fig. 5: Enhanced AMPAR trafficking and function in the vmPFC were associated with chronic pain-induced anxiety. | Nature Communications

Fig. 5: Enhanced AMPAR trafficking and function in the vmPFC were associated with chronic pain-induced anxiety.

From: nNOS-expressing neurons in the vmPFC transform pPVT-derived chronic pain signals into anxiety behaviors

Fig. 5

a Representative traces of sEPSCs in layer 2/3 pyramidal neurons of the vmPFC 3 days after CFA or NS injection. b, c Cumulative fraction plots of sEPSC amplitude (b, mean amplitude in inset) and interevent intervals (c, mean frequency in inset). n = 14 (from 5 mice) and 15 (from 6 mice) neurons for the NS and CFA groups, respectively. d Representative traces of evoked EPSCs (eEPSCs) in the presence of BMI in layer 2/3 pyramidal neurons of the vmPFC 3 days after CFA or NS injection. Arrows indicate that AMPAR-EPSCs were measured at the peak at a holding potential of −70 mV, and NMDAR-EPSCs were measured 50 ms after stimulation at a holding potential of +40 mV. e Dot plot showing the AMPA/NMDA ratio from (d). n = 10 neurons (from 4 mice). (f–h) Representative western blots (f) and dot plots showing total (g) and surface (h) protein levels of GluA1 and GluA2 at day 3 after CFA or NS injection. i Scheme indicating the vmPFC area where AAV-hSyn-DIO-hM3Dq-eGFP (0.2 μl) and AAV-CaMKIIα-mCherry (0.2 μl) were microinjected into nNOS-Cre mice. n = 5. j Immunofluorescent image of vmPFC showing mCherry+/eGFP− neurons (arrows indicated), form which sEPSCs and eEPSCs were recorded. This experiment was conducted in triplicate with 5 mice and similar results were observed. k Representative traces of sEPSCs in layer 2/3 pyramidal neurons of the vmPFC 3 weeks after AAV injection into nNOS-Cre mice. l, m Cumulative fraction plots of sEPSC amplitude (l, mean amplitude in inset) and interevent intervals (m, mean frequency in inset). n = 15 neurons (from 6 mice). n Representative traces of eEPSCs in the presence of BMI in layer 2/3 pyramidal neurons of the vmPFC 3 weeks after AAV injection into nNOS-Cre mice. Arrows indicate that AMPAR-EPSCs were measured at their peak at a holding potential of −70 mV, and NMDAR-EPSCs were measured 50 ms after stimulation at a holding potential of +40 mV. o Bar graph showing the AMPA/NMDA ratio. n = 10 neurons (from 4 mice). p–r Representative western blots (p) and dot plots showing total (q) and surface (r) protein levels of GluA1 and GluA2 30 min after CNO (2 mg per kg, i.p.) or NS treatment at day 21 after AAV microinjection. n = 4. Data are the mean ± SEM; *p < 0.05, **p < 0.01, and ***p < 0.001 (unpaired two-tailed Student’s t-test for (b), (c), (e), (g), (h), (q), and (r); one-way ANOVA followed by Tukey’s post hoc test for (i), (m), and (o)). Scale bar, 50 μM in (j). Source data are provided as a Source data file. Exact p values and additional statistical information can be found in Source data.

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