A role for estrogen receptor (ER) in branching morphogenesis in the ventral prostate (VP) has previously been demonstrated: in the VP of ER^-/- mice, there are fewer side branches than in wild-type littermates. In the present study we show that in the postnatal VP, fibroblast growth factor (FGF) 10 is expressed in wild-type mice but not in ER^-/- mice, and, since branching involves proliferation pathways also used in malignant growth, we investigated whether branching during re-growth of the VP after castration involves ER and FGF10. ER was not detectable in the prostates of sham-operated or castrated mice but was expressed in the prostatic epithelium between days 3–5 after testosterone replacement. Blocking either ER or ER with ICI 182,780 had no detectable effects on epithelial cell proliferation during regrowth by testosterone. The ER agonist, PPT, did not induce re-growth by itself, but exposure to PPT on days 3 to 5 of testosterone replacement resulted in cyclin D1-positive cells in the ductal epithelium, invasion of FGF10-positive immune cells in the re-growing prostate, and in budding 14 days later. Testosterone replacement alone induced neither cyclin D1, FGF10 nor bud formation. These results indicate that stimulation of ER is essential for ductal branching during postnatal prostate growth. During re-growth after castration there is a window in time when selective stimulation of ER can also induce ductal branching. The FGF10 for this growth comes from the immune system, not from the prostatic mesenchyme.