Fig. 4: Labeling the spontaneous pain-related neuronal ensemble in the PL via RAM system. | Nature Communications

Fig. 4: Labeling the spontaneous pain-related neuronal ensemble in the PL via RAM system.

From: A distinct neuronal ensemble of prelimbic cortex mediates spontaneous pain in rats with peripheral inflammation

Fig. 4

a The RAM labeling system. Left: Virus strategy. Right: Timeline of virus injection, Dox switching, activated neuronal labeling and electrophysiological recording. The diagram was created with BioRender.com. b, c Characterization of labeled PL neurons in spontaneous pain induced by CFA using markers of glutamatergic (EAAC1+) and GABAergic (GAD+) neurons. Typical examples (b) and a quantitative summary (c) are shown. Scale bar, 250 µm. Three slices from each rat, n = 3 rats. d The raster plot (upper) and PSTH (bottom) of a representative opto-tagged spike unit activated by 1-Hz light pulse. e The proportion of the neurons responding to spontaneous paw lifting (inner pie charts) and 1-Hz light pulse (outer rings). n = 91 units from 3 rats. f Averaged PSTH of the tagged (green, activated), relevant (blue, inhibited) and irrelevant (orange, no significant change) neuronal subpopulations to a single light pulse. Shaded areas were defined by SEM. n = 133 units from 7 rats. g PSTHs of recorded neurons (left) in the 1-Hz protocol. Neurons were classified by a descending order of firing rate during the 1-s light-on period. Pie graphs (right) showing the proportion of neurons. h PSTHs of recorded neurons in the 3-min, 20-Hz protocol. Neurons were ordered in line with (g). i Average firing rate of the tagged and relevant neurons after optogenetic activation. n = 21 tagged, 56 irrelevant, 46 relevant units, respectively. *p < 0.05, ***p < 0.001, Friedman test. j Evoked nociceptive responses of recorded PL neurons to noxious laser on CFA-injected and uninjected hindpaws with and without optogenetic activation of spontaneous pain ensemble. Bin, 100 ms. k Averaged Z score changes. n = 21 tagged, 56 irrelevant, 46 relevant units respectively. **p < 0.01, two-way repeated measures ANOVA. Evoked spectrogram (l) and gamma power (m) of PL by noxious laser on CFA-injected and uninjected hindpaws with and without optogenetic activation. Bin, 50 ms. n Nociception-evoked gamma power after activating the spontaneous pain neuronal ensemble. n = 7 rats. *p < 0.05, two-way repeated measures ANOVA. Data for I, K and N were presented as mean ± SEM and provided as a Source Data file.

Back to article page