Our previous finding of appreciable quantities of a gastrin-releasing peptide (GRP)-like immunoreactive (GRPLI) entity in ovine fetal and maternal plasma led us to examine the ovine pregnant uterus as a possible source of this material. At term, intense immunohistochemical staining for GRPLI occurred in the endometrial epithelial cells, and the term ovine uterus also contained abundant GRP messenger RNA (mRNA). In contrast, GRP mRNA was not detected in fetal membranes. GRP mRNA was present in the uterus on gestational day 63; a significant increase in GRP mRNA had occurred by day 100. Thereafter, levels remained elevated until term, but 3 months postpartum, GRP mRNA levels were greatly reduced. As previous studies suggested the GRPLI entity to be of greater molecular size than GRP-(1-27), we deduced the primary structure of ovine uterus GRP by sequencing a complementary DNA clone isolated from a complementary DNA library constructed from term ovine uterus polyadenylated RNA. Ovine uterine GRP is composed of 27 amino acid residues and has a conserved C-terminal region, similar to GRP structures in other species. We conclude that during pregnancy, the ovine uterus produces considerable quantities of GRP, which may play an important but hitherto unrecognized role in utero-placental development and possibly in fetal development after transfer to the fetus.

Although the excitatory amino acid (EAA) receptor agonist N-methyl-D-aspartate (NMDA) can exert profound stimulatory effects on the neuroendocrine reproductive axis of Syrian hamsters, the exact relationship between NMDA receptors and LHRH neurones is unclear. In the present study, in situ hybridization histochemistry was performed on sections of hamster brain using an 35S-labelled riboprobe to the EAA receptor gene, NMDAR1. A high content of NMDA receptor mRNA was detected not only in brain areas classically associated with specific NMDA binding (for example, hippocampus and cerebral cortex) but also in the hypothalamus, in particular the ventromedial-arcuate area; diffuse hybridization of the riboprobe also occurred in the medial-septal area and diagonal band of Broca, regions of the hamster brain in which the LHRH neuronal perikarya are primarily located. In a separate experiment, RNA was extracted from immortalized LHRH neurones (GT1-1 and GT1-7 cells) and used for northern analysis with a 32P-labelled NMDAR1 riboprobe. Clear-cut hybridization occurred with RNA bands of approximately 4.2 and 4.4 kb from the two LHRH neuronal subtypes. These findings suggest that at least some of the stimulatory action of EAAs on LHRH secretion is likely to be exerted directly at the level of the LHRH neurones rather than being mediated through interneurones. Furthermore, the demonstration of abundant NMDA receptor gene expression within hypothalamic areas that lie outside the blood-brain barrier adds plausibility to the concern that EAAs of dietary origin, such as monosodium glutamate, have the capacity to perturb the normal secretory activity of neuroendocrine circuits of the hypothalamus.

Preimplantation rabbit embryos collected at the early morula stage were cultured to blastocysts in the presence of [3H]inositol. The blastocysts were lysed, and both the aqueous and lipid portions were analysed for incorporated radioactivity. Thin-layer chromatographic separation of the lipid portion indicated that [3H]inositol was incorporated into phosphatidylinositol, phosphatidylinositol 4-phosphate, and phosphatidylinositol 4,5-bisphosphate. HPLC anion-exchange chromatography indicated that [3H]inositol was incorporated into inositol phosphates, including the two second messengers, inositol 1,4,5-triphosphate and inositol 1,3,4,5-tetrakisphosphate, and also inositol monophosphate and inositol 1,4-bisphosphate. These results provide evidence that rabbit blastocysts may have an active phosphatidylinositol second messenger system, which may be responsive to intrauterine factors or intraembryonic paracrine factors.

The effect of different concentrations (0, 0.6, 3, 15, 75 and 375 microM) of myo-inositol on the development of rabbit morulae to expanded blastocysts was investigated in terms of blastocyst expansion and synthesis of DNA and protein, as measured by incorporation of [3H]thymidine and [14C]amino acids into acid-precipitable material. A concentration of 15 microM inositol caused a 2.8-fold increase in blastocyst expansion (P less than 0.01), a 9.9-fold increase in thymidine incorporation into DNA (P less than 0.01) and a 3.6-fold increase in amino acid incorporation into protein (P less than 0.01). There were no significant differences in the range from 15 to 375 microM inositol.