The int-2 proto-oncogene encodes several products related to the fibroblast growth factor (FGF) family. FGFs have been associated with mesoderm induction in the amphibian embryo and int-2 has a distinct pattern of expression throughout development in vertebrates. But evidence for a function of int-2 in embryo-genesis has been lacking. In the mouse embryo, int-2 transcripts have been detected in the rhombencephalon at a developmental stage where classical experiments showed that the induction of the inner ear occurs. This raises the possibility that int-2 may constitute a signal for the induction of the otic vesicle, the primordium of the inner ear. We provide direct evidence for this view by showing that (1) the formation of the otic vesicle is inhibited by antisense oligonucleotides targeted to the secreted form of int-2, and by antibodies against int-2 oncoproteins, and (2) basic FGF (bFGF) can mimic the inductive signal in the absence of the rhombencephalon.

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The role of insulin-like growth factors (IGF) was investigated during the early development of the inner ear. IGF-I stimulated growth of otic vesicles that were isolated and cultured in vitro. IGF-I induced DNA synthesis, increased cell number, and mitotic rate in a dose-dependent manner at concentrations between 0.1-10 nM. IGF-II also induced growth but with a lower potency, whereas insulin had no effect. In the presence of IGF-I, otic vesicles developed from stage 18 to stage 21 in 24-h cultures, mimicking the normal mitotic pattern and morphogenesis in vivo. IGF-I also stimulated growth in the cochleovestibular ganglion. Binding of 125I-IGF-I to specific receptors occurred with high affinity. An autoradiographic study of sections from otic vesicles showed radiolabeled IGF-I in the epithelium. Immunoreactivity to IGF-I was detected in the otic vesicle and in the cochleovestibular ganglion. Intracellular signaling mechanisms of IGF were explored by studying the turnover of glycosylated phosphatidylinositols and the expression of Fos oncoprotein. IGF-I rapidly increased Fos levels in cultured otic vesicles. Furthermore, antisense oligonucleotides complementary to c-fos were able to inhibit IGF-I-induced growth. Both IGF-I-induced cell proliferation and Fos expression were blocked by an antiinositol phosphoglycan (alpha-IPG) antibody. This work suggests that IGF-I may be a candidate to regulate proliferative growth of the otic primordium during normal development and that this action requires the sequential modulation of glycosyl-phosphatidylinositol turnover and Fos expression.