IN-SITU HYBRIDIZATION OF LUTEINIZING-HORMONE HUMAN CHORIONIC-GONADOTROPIN RECEPTOR MESSENGER-RIBONUCLEIC-ACID DURING HORMONE-INDUCED DOWN-REGULATION AND THE SUBSEQUENT RECOVERY IN RAT CORPUS-LUTEUM

Previous studies have shown that injection of a pharmacological dose of hCG to rats primed with PMSG/hCG results in a loss of hCG binding in the luteinized ovary, which is closely coupled with the loss of LH/hCG receptor (LH/hCG R) messenger RNA (mRNA). The time course of down-regulation of the receptor mRNA reveals that mRNA totally disappears 24 h after the hormone injection, but fully recovers by 72 h. The purpose of this study was to determine by in situ hybridization whether the recovery of the receptor mRNA occurs in preexisting or newly formed corpora lutea. Twenty-one-day-old female rats were treated with 50 IU PMSG, followed 56 h later by a single injection of hCG. On day 5 after the hCG injection, one group of rats was treated with a desensitizing dose of hCG, and a control group received saline. The ovaries were collected 6, 12, 24, 48, 72, and 96 h after the treatments and were processed for in situ hybridization using S-35 antisense RNA synthesized from a 750-mer LH/hCG-R complementary DNA. In control ovaries, heavily labeled LH/hCG-R mRNA-containing cells were observed in the numerous corpora lutea. Ribonuclease pretreatment of the sections eliminated the signal, and no specific hybridization was observed when sense strand probe was used. No hybridization to the granulosa cells was seen. Some hybridization occurred to the theca interna, but the intensity was lower than that in the corpora lutea. Time-course studies showed a marked decline in the hCG-R mRNA signals in corpora lutea as early as 6 h after hCG injection, with a maximum loss of receptor mRNA by 24 h. After 48 h, hCG-R mRNA reappeared in preexisting corpora lutea, with the intensity of the hybridization signal equaling that in corpora lutea of controls. New corpora lutes could not be identified at any time after injection of the down-regulating dose of hCG. As down-regulated receptor mRNA recovered in preexisting, not new, corpora lutes, hormone-induced loss of luteal cell receptors would appear to be reversible.