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The presence of substance P (SP) in the immature rat ovary was determined by radioimmunoassay (RIA) of acidic extracts. The extracts produced an inhibition-displacement curve of 125I-SP binding parallel to that generated by authentic SP in the SP RIA. Initial chromatographic characterization of ovarian SP in Sephadex G-25 revealed the presence of a molecular form that coeluted with authentic SP and a more abundant component that eluted earlier, suggesting the presence of a heavier peptide, immunologically similar to SP. Nevertheless, further characterization of these two seemingly different components by reversed-phase high-performance liquid chromatography (HPLC) demonstrated that both of them had a retention time similar to that of authentic SP. The ovarian concentration of SP-like immunoreactivity (SP-LI) varied in relation to the onset of puberty, with values increasing significantly between the late juvenile phase and the day of first proestrus. Substance P seems to be devoid of steroidogenic capacity since SP itself and its stable analog [pGlu5,MePhe8,Sar9]-SP5-11 (SP-A) failed to stimulate steroid secretion from either granulosa cells in culture or ovarian fragments in short-term incubation. Substance P also failed to stimulate prostaglandin E2 release from whole ovaries and to modify the steroidal response of cultured granulosa cells to follicle-stimulating hormone and to the beta 2-adrenergic agonist Zinterol. Production of SP-LI from granulosa cells in culture could not be detected under either basal or gonadotropin-stimulated conditions. These observations and the distribution of the peptide within the ovary presented in the companion paper (Dees et al., this issue) strongly suggest that SP is not directly involved in regulating steroidogenesis. Instead, SP may be a component of the so-called sensory innervation of the ovary, and among other undisclosed functions it may contribute to the regulation of ovarian blood flow.

Evidence for the Existence of Substance P in the Prepubertal Rat Ovary. I. Biochemical and Physiologic Studies1