We have investigated the interaction of rat primary calvarial bone cells and a mouse osteoblast-like cell line MC3T3-E1 with basement membrane components. On a reconstituted gel of basement membrane, both cell types attached and formed isolated clusters that developed long interconnecting cell processes similar to the canalicular network observed in bone. The differentiation of the osteoblastic phenotype was stimulated as determined by increased alkaline phosphatase production and the deposition of mineral. Antibodies to laminin and to a 32/67 kd laminin receptor blocked this differentiation. Cell morphology was altered by the addition of active laminin-derived synthetic peptides, YIGSR-NH2 and CSRARKQAASIKVAVSADR-NH2, but not by an active RGD-containing peptide. When coated directly on plastic, all three peptides promoted cell adhesion, demonstrating that bone cells interact with specific molecular domains of laminin. These data demonstrate that basement membrane plays a key role in formation of a network of cytoplasmic processes resembling the osteocyte canalicular network in bone.