The pancreas of a child with intractable neonatal hypoglycemia was explanted in tissue culture on plasma clot and in monolayer culture after enzymatic dissociation. 1. Cytological and immunoenzymatic studies of pancreas before explantation showed a very altered structure, suggesting a polyendocrine tumor composed mainly of B cells. Endocrine cells were present in the epithelium of duct-like structures. Large islets were often in continuity with these structures, suggesting islet budding from ducts, and supporting the hypothesis of the persistence of embryonic characteristics. 2. In vitro, the pancreatic endocrine cells survived and proliferated; they were maintained for 362 days. During this time, they maintained their secretory capacity, as shown by radioimmunoassays of the culture media: the cells released insulin in rapidly decreasing amounts, then continued excreting low levels (5 to 60 muU/flask/day), in alternative periods of secretion and absence of secretion. 3. Cytological study by light and electron microscopy of the cells in tissue and in monolayer cultures shows that they can undergo morphological changes, and may become epithelioid, fibroblastoid or round, while retaining their secreting activity. 4. In long-term culture, the cells did not contain typical mature secretion granules. The hormone might be released into the medium by a clasmatosis mechanism. On the other hand, hormone excretion by vesicles originating from the rough endoplasmic reticulum is possible.