Chronic activation of the glucocorticoid stress pathway decreased NR4A1 phosphorylation and cytosolic distribution in mouse PFC. A, Endogenous NR4A1 protein is both nuclear and cytosolic in pyramidal cortical neurons of PFC (thy1-YFP). Proportion of a total of 10,600 cells with NR4A1 localized equally in nucleus and cytosol (N = C), more in nucleus than cytosol (N > C), and less in nucleus than cytosol (N < C). Mean ± SEM of N = 10 controls, N = 10 acute stress mice, N = 7 CORT mice, and N = 9 chronic stress mice; unpaired t test: *p = < 0.0001 comparing indicated group to controls; and #p = 0.0006 comparing acute to chronic stress mice. B, Western blots of PFC lysates from P31 mice subjected to acute stress, chronic stress, or CORT injections compared with controls. C, Phosphorylation of NR4A1 (S350-P) expressed as a percentage of controls. Mean ± SEM of N = 12 control mice, N = 12 acute stress mice, N = 6 CORT mice, and N = 6 chronic stress mice; unpaired t test: *p = 0.013, op = 0.0094, §p = 0.014 comparing indicated treatment to controls; #p = 0.0153 comparing acute and chronic stress mice. D, Cytosolic NR4A1 expressed as a percentage of controls. Mean ± SEM of N = 7 control mice, N = 7 acute stress mice, N = 6 CORT mice, and N = 6 chronic stress mice; unpaired t test: *p = 0.025, op < 0.0001, §p = 0.0009 comparing indicated treatment to controls; #p = 0.0025 comparing acute and chronic stress mice. E, Model of NR4A1 functional transport in cortical neurons. Induction of NR4A1 by acute stress increased nuclear export and phosphorylation. Chronic stress prolonged NR4A1 nuclear residency, which failed to contain its activity.