Epidermal growth factor (EGF), a mitogen in vitro for hepatocytes, produces in various cell lines changes similar to those observed very rapidly in hepatocytes after partial hepatectomy (PH). These changes include ion movements, membrane hyperpolarization and proto-oncogene expression. A stimulatory effect of EGF on liver regeneration can therefore tentatively be associated with the events occurring within the first 3 hours after a PH, sometimes referred to as the “priming phase.” To assess this hypothesis, we examined in Wistar rats the effect of EGF deprivation produced by sialoadenectomy (SX) performed before or after a PH of 70%. SX at the time of PH significantly decreased the 3H-thymidine uptake in the DNA 24 hours later (147 ± 14 DPM per microgram of DNA, mean ± SE) compared with a simple PH (322 ± 16; P < .01), but also compared with results obtained when PH is combined with a sham sialoadenectomy (SSX) or in rats pair-fed with the sialoadenectomized rats. This incomplete inhibition was confirmed by a decreased rise in thymidine kinase (TK) activity and by reduced proliferating cell nuclear antigen (PCNA) labeling and mitotic indices 30 hours after PH. By contrast, SX did not inhibit the early expression of c-jun and c-fos, or of c-myc, 30 or 120 minutes after PH, respectively. A reduction of DNA synthesis was also obtained when SX was performed 3 hours after PH (127 ± 15 DPM per microgram of DNA vs. 350 ± 21 in SSX; P < .001) but not when SX was delayed until the 6th or the 17th hour after PH. It was sufficient to administer EGF (40 microg) from the third to the ninth hour to correct the reduction of [3H]thymidine uptake in rats sialoadenectomized before PH. These results indicate that the diminished EGF availability following SX decreases or at least delays liver regeneration, and that the effect of EGF on liver regeneration does not seem related to the early changes of proto-oncogene expression, but rather to events occuring later, at the time of reported internalization and binding of EGF to its nuclear receptors.