Figure 1. Acute antipsychotic treatment increases Akt-mTORC1 pathway signaling in D₂R-positive neurons.

Primary DIV7 mouse striatal neurons were treated with haloperidol (Hal) or DMSO (Veh), in the presence of an Akt inhibitor (Akti) or rapamycin (Rap) for 20 minutes. (A) Western blot analysis of phosphorylated and total Akt in lysates exposed to the indicated conditions (left, representative blot; right quantification of n =4, One Way ANOVA p < 0.0001, post hoc analysis Veh vs Hal p <0.01, Hal vs Akti, p < 0.001). (B) Western blot analysis of phosphorylated and total S6 and 4E-BP in lysates exposed to the indicated conditions [left, representative blot; right quantification of S6 (n=3, One Way ANOVA p < 0.0008, Veh vs Hal p < 0.05, Hal vs Akti and Hal vs Rap p < 0.01) and 4E-BP (n = 3, One Way ANOVA p < 0.0003, Veh vs Hal p <0.05, Hal vs Akti p < 0.001, Hal vs Rap p <0.01). All graphs shown are average ± SEM. (C) Changes in the abundance of the indicated phosphorylated proteins in striatal neurons exposed to the indicated treatments as detected by immunofluorescence. Arrows indicate responding neurons, stars indicate cells that are not responding. Quantification of pS6 (n= 4, One Way ANOVA p < 0.05, all indicated single comparisons p < 0.05) and p4E-BP (n=4, One Way ANOVA p < 0.0001, Veh vs Hal p < 0.01, Hal vs Akti and Hal vs Rap p < 0.001). (D) D₂Rs are detected in nearly all cells with increased pAkt (arrow). A nonresponding cell is indicated by asterisk. Quantification of responding cells and D₂R-positive (D₂R+) and D₂R-negative (D₂R−) cells (n=3, 109 cells counted). D₂R image underwent Iterative deconvolution in ImageJ to increase clarity of signal. (E) The abundance of phosphorylated Akt and S6 in striatal neurons exposed to amisulpride (1 μM) or vehicle for 20 minutes (pAkt, n = 3, p < 0.06; pS6, n = 3, p < 0.03). Scale bars indicate 50 μm.