Electron microscopy of cultured P388 leukemia cells revealed morphological differences between the Adriamycin-resistant (P388/ADR) and -sensitive (P388/0) cell lines. P388/ADR cells showed longer villus-like protrusions and large foldings of the plasma cell membrane, whereas P388/0 cells had only short membrane protrusions. Western blot analysis of cells revealed the increased expression of a glycoprotein of 170-180 kDa (P-glycoprotein) in P388/ADR cells as compared with P388/0 cells, which is consistent with the findings of other studies. A modulation of the membrane morphology of in vitro P388/ADR cells was evident from one i.p. in vivo passage for 10 days, by a reversion to the non-ruffled sensitive-cell morphology. Long-term P388/ADR culture cells showed a reduction in membrane folding with as little as a 4-h exposure to the peritoneal ascitic fluid obtained when the tumor was harvested. However, there was no alteration in the expression of P-glycoprotein or in the sensitivity to Adriamycin in either of the P388 cell lines when grown in vivo or when exposed to ascites fluid from an i.p. tumor. Thus, the modulation of membrane morphology was related to the tumor cell environment rather than the abundance of P-glycoprotein in the plasma membrane.