Bombesin is a 14 amino acid peptide originally isolated from amphibian skin; its mammalian homolog is gastrin-releasing peptide (GRP). GRP is found in a high proportion of human small cell lung cancer (SCLC) cell lines. [Tyr4]bombesin caused an increase in intracellular Ca2+ ([Ca2+]i) in 5/11 SCLC cell lines tested. Bombesin action was not inhibited by agents known to alter the plasma membrane potential, nor did replacement of external Na+ with choline affect the bombesin-induced signal. [Tyr4]bombesin did not itself affect the membrane potential. Chelation of external Ca2+ reduced but did not prevent the bombesin-evoked increase in [Ca2+]i. This suggested that in SCLC, bombesin congeners not only promote an influx of extracellular Ca2+ but also release Ca2+ from intracellular stores. [Tyr4]bombesin increased levels of inositol 1,4,5-trisphosphate within seconds of addition to SCLC cell cultures and enhanced the accumulation of inositol 1-phosphate and inositol 4-phosphate in the presence of Li+. The SCLC cell lines responsive to bombesin constitutively expressed L-myc and did not express c-myc or N-myc. In contrast, SCLC cells non-responsive to bombesin had prominent constitutive expression of c-myc or N-myc with or without L-myc expression. Responding cell lines also had constitutive expression of the preproGRP gene, while non-responding cell lines showed no evidence of GRP gene expression. These data support the concept that SCLC which constitutively express the GRP gene and L-myc but not c-myc or N-myc can be stimulated in an autocrine fashion by GRP or its congeners to increase [Ca2+]i by a pathway involving phosphatidylinositol turnover.