Stress-Induced Genetic Expression of a Selective Corticotropin-Releasing Factor-Receptor Subtype Within the Rat Ovaries: An Effect Dependent on the Ovulatory Cycle1

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Abstract
The identification of several sources of corticotropin-releasing factor (CRF) outside the brain, including the gonads, has suggested the intriguing possibility that CRF of systemic origin can influence gonadal function under both normal and stressful conditions. However, the exact sites of action and the type of cells targeted by this stress-related neuropeptide in the ovaries remain unknown. The present study therefore investigated the effect of acute immobilization stress on the distribution of the recently cloned CRF receptor (CRF1, CRF2 alpha, and CRF2 beta) genes in the ovaries of adult cycling female rats (200-250 g; 14 h light, lights-on at 0600 h). Reproductive stages were verified by daily vaginal smears taken each morning for a minimum of 3-4 cycles prior to the experiment. Three hours after the start of a 90-min immobilization session, rats were deeply anesthetized and transcardially perfused with a solution of 4% paraformaldehyde on the morning (1100 h) and on the afternoon (1700 h) of proestrus and diestrus-2. Frozen ovaries were mounted on a microtome, cut into 30-microns slices, and then processed for the detection of mRNAs encoding CRF1, CRF2 alpha, or CRF2 beta receptors by in situ hybridization histochemistry using 35S-labeled riboprobes. Whereas the ovaries displayed barely detectable levels of CRF1 receptor mRNA in control and in stressed animals on the morning of proestrus and the day of diestrus-2, a positive signal for this transcript was detected in stroma cells and in the theca surrounding the ovulatory follicles during the afternoon of proestrus. Excluding the cumulus oophorus, which showed a light expression of the mRNA encoding the type 1 CRF receptor, granulosa cells were completely devoid of transcript in Graafian follicles as well as in CL, regardless of the stage of maturation. Interestingly, immobilization stress induced a marked expression of CRF1 receptor mRNA in the stroma cells in the afternoon of proestrus, suggesting that the ovaries may be sensitive to acute neurogenic challenge during the preovulatory stage. On the other hand, CRF2 alpha and CRF2 beta receptor mRNAs were undetectable both in control and stressed animals throughout the estrous cycle. These results provide clear evidence that the gene encoding the CRF1 receptor but not the type 2 receptors can be finely induced in selective ovarian compartments in both control and stressful conditions during the gonadal life cycle. The temporal and anatomic selectivity of the ovarian periovulatory CRF1 receptor gene expression may suggest a critical biological action of CRF during the ovulatory process and suggests that the intraovarian environment may influence the stress-induced transcription of a selective CRF receptor subtype within the ovary.

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