LH receptor mRNA and cytochrome P450 side-chain cleavage expression in bovine theca and granulosa cells luteinized by LH or forskolin

Abstract

This study was undertaken to characterize the cDNA sequence encoding bovine LH receptor (LHR), and study the effect of different luteinization protocols on the steroidogenic capacity and LHR mRNA in bovine luteal cells. The bovine LHR cDNA sequence reported here showed high sequence identity with that of other species. Several mRNA splice variants were expressed in bovine corpus luteum (CL). Reverse transcription-polymerase chain reaction conducted with primers spanning exons 2–11 revealed, in addition to the full-length sequence, a shorter fragment homologous to splice variant B reported in porcine and ovine LHR cDNA sequences. Granulosa and theca cells derived from bovine preovulatory follicles were cultured with either forskolin (10 μM, for 8 d culture—Protocol 1) or LH (100 ng/ml, for 12 hr followed by a 7.5-d culture with 2 ng/ml—Protocol 2). LHR mRNA was not detected in luteal granulosa cells (LG); in contrast, luteal theca cells (LT) contained LHR mRNA at similar levels when cultured under Protocol 1 or 2. cAMP responses to a short challenge with LH were in good agreement with LHR mRNA levels. Cytochrome P450 side-chain-cleavage (P450scc) contents were lower in luteal cells cultured with LH as compared with cells cultured with forskolin; however, the difference between the two luteinization protocols was much larger in LT (P < 0.05) than in LG. This study suggests that a) LHR mRNA is mainly present in the theca-derived luteal cell and b) LHR mRNA and cytochrome P450scc expression in each of the luteal cell types seems to be under different control.