Fig. 3: Alcohol activates mTORC1, increases the translation of Trax, GW182 and CNOT, and decreases the translation of Aldolase A, PPM1E, Rbfox2 in NAc D1+ neurons. | Nature Communications

Fig. 3: Alcohol activates mTORC1, increases the translation of Trax, GW182 and CNOT, and decreases the translation of Aldolase A, PPM1E, Rbfox2 in NAc D1+ neurons.

From: Paradoxical mTORC1-Dependent microRNA-mediated Translation Repression in the Nucleus Accumbens of Male Mice Consuming Alcohol Attenuates Glycolysis

Fig. 3

A D1-Cre mice were crossed with RiboTag mice allowing the expression of RPL10-EGFP in D1-expressing neurons. B Mice underwent 7 weeks of IA20%2BC (Supplementary Table 1). Brains were dissected at the end of the last 24-hour withdrawal session as in Fig. 1A and processed for IHC or biochemical analysis. C IHC analysis of phospho-S6 levels in the NAc shell D1 neurons of drinking mice compared to water controls. Representative images of NAc at 20x magnification labeled with phospho-S6 in red, RPL10-GFP in green and NeuN in magenta. Scale bar 100 µm. D Phospho-S6 and D1 labeled neurons are expressed as % of water controls±SEM. ***p < 0.001. n = 3 mice per group. Significance was determined using two-tailed unpaired t-tests. t(4) = 27.29, p = <0.0001. E Percentage of D1 positive vs. D1 negative NAc shell neurons labeled as phosphoS6 positive neurons. F Affinity purification of ribosomes from D1+ neurons by using anti-GFP magnetic beads followed by RNA isolation and RT-qPCR. Image provided by Servier Medical Art (https://smart.servier.com/), licensed under CC BY 4.0 (https://creativecommons.org/licenses/by/4.0/). G–L Polysomal mRNA levels in D1+ neurons of Trax (G), GW182 (H), CNOT4 (I), Aldolase A (J), PPM1E (K), Rbfox2 (L) after alcohol withdrawal were determined by RT-qPCR. Each data point represents an average of 3 technical replicates. Data are presented as the average ratio of a transcript to GAPDH ± SEM and expressed as % of water control. *p < 0.05, **p < 0.01, ***p < 0.001. G, H, J–L Water: n = 9, Alcohol: n = 10, I n = 5 mice per group. Significance was determined using two-tailed unpaired t-tests. G t(17) = 4.042, p = 0.0008; H t(17) = 2.575, p = 0.0197; I t(8) = 2.322, p = 0.0488; J t(17) = 2.299, p = 0.0345; K t(17) = 3.425, p = 0.0032; L t(17) = 4.26, p = 0.0005. M Alcohol activates mTORC1 signaling in D1+ NAc neurons which in turn increases the translation of GW182, Trax and CNOT4 and represses the translation of Aldolase A, Rbfox2 and PPM1E. Source data are provided as a Source Data file.

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