Glucocorticoids promote spine formation and pruning through distinct signaling pathways. (a) Corticosterone (cort) increased spine formation rapidly (F1,40 = 124.0, P < 0.001). (b) Cort caused a delayed increase in spine elimination (F1,40 = 102.3, P < 0.001). (c) Glucocorticoids promote spine formation rapidly through a GR-dependent, non-transcriptional mechanism. Direct cortical application of cort caused rapid increases in spine formation (t = 14.6, P < 0.001), with comparable effects evident after cortical application of cort plus actinomycin D (50 μg ml−1; t = 18.8, P < 0.001) or a membrane-impermeant cort-BSA conjugate (t = 4.83, P < 0.001). Co-administration of the GR antagonist (antag) mifepristone (100 μM) blocked this effect. (d) Mifepristone (20 mg kg−1 i.p.) administered immediately after training reduced learning-dependent spine formation (t = 8.46, P < 0.001). (e) Glucocorticoids cause a delayed increase in spine elimination through a MR-dependent, transcriptional mechanism. A selective MR antagonist (spironolactone, 10 μM) reduced 24-h spine elimination (t = 6.19, P < 0.001), whereas a selective MR agonist (aldosterone, 10 μM) had the opposite effect (t = 18.3, P < 0.001). This effect was blocked by cotreatment with either actinomycin D or anisomycin, consistent with a transcriptional mechanism of action. (f) Spironolactone administered during the circadian trough on days 4, 5 and 6 after training interfered with learning-induced spine elimination (t = 4.84, P = 0.002). *P < 0.05 versus corresponding control. Error bars, s.e.m. See for statistics and details.