The distribution of glucose-6-phosphatase (G6Pase) activity in the epithelium of the small intestine in mouse embryos (the last 4 days of gestation) was studied by EM cytochemistry and by enzymatic assays. At 16 days, the lead phosphate deposited by the cytochemical reaction is localized on the rough endoplasmic reticulum (RER) and nuclear envelope of very few cells in the duodenum and jejunum. Positive cells are more frequently seen in the upper part of the developing villi. At 17 days of gestation, a tremendous burst in RER differentiation is noticed in all parts of the small intestine and concomitantly glycogen disappears. At 18 days of gestation all the principal cells of the intestinal mucosa show a well differentiated positive RER and the enzyme is also present in the smooth endoplasmic reticulum. G6Pase activity is detected in the proximal 2/3 of the small intestine at 17 days of gestation and appears at 18 days in the last 1/3. Afterwards the activity increases up until birth. These results suggest the following: the endoplasmic reticulum may differentiate very late in the intestinal mucosa of mouse embryo; the differentiation with respect to G6Pase is asynchronous between the enterocytes; for a given cell all the cisternae of RER are involved in G6Pase synthesis at the same moment; and the enterocytes of the duodenum differentiate sooner and faster than those of the jejunum and ileum.