Photoperiod and reproductive condition are associated with changes in RFamide-related peptide (RFRP) expression in Syrian hamsters (Mesocricetus auratus)

Abstract

To conserve scarce energetic resources during winter, seasonal breeders inhibit reproduction and other nonessential behavioral and physiological processes. Reproductive cessation is initiated in response to declining day lengths, a stimulus represented centrally as a long-duration melatonin signal. The melatonin signal is not decoded by the reproductive axis directly, but by an unidentified neurochemical system upstream of gonadotropin-releasing hormone (GnRH). The dorsomedial nucleus of the hypothalamus (DMH) has been implicated in seasonal changes in reproductive function in Syrian hamsters (Mesocricetus auratus), although the specific-cell phenotype decoding photoperiodic information remains unknown. RFamide-related peptide (RFRP; the mammalian homolog of the gonadotropin-inhibitory hormone (GnIH) gene identified in birds) has emerged as a potent inhibitory regulator of the reproductive axis and, significantly, its expression is localized to cell bodies of the DMH in rodents. In the present study, the authors explored the relationship between RFRP expression, photoperiod exposure, and reproductive condition/hormonal status. In male hamsters that respond to short days with reproductive inhibition, RFRP-ir and mRNA expression are markedly reduced relative to long-day animals. Replacement of testosterone in short-day animals did not affect this response, suggesting that alterations in RFRP expression are not a result of changing sex steroid concentrations. A subset of the hamster population that ignores day length cues and remains reproductively competent in short days (nonresponders) exhibits RFRP-ir expression comparable to long-day hamsters. Analysis of cell body and fiber density suggests a potential interplay between peptide production and release rate in differentially regulating the reproductive axis during early and late stages of reproductive regression. Together, the present findings indicate that photoperiod-induced suppression of reproduction is associated with changes in RFRP and mRNA expression, providing opportunity for further exploration on the role that RFRP plays in this process.

Figure 1

Body and reproductive organ masses from animals held in long (LD 16:8) or short (LD 8:16) day lengths. Animals held in short days for 8 weeks were separated into “responders” (R) or “nonresponders” (NR) based on gonadal size. Body size (A) was unaffected by photoperiodic treatment. Testes mass (B) and epididymal mass (C) were significantly diminished in R hamsters held in short days for 8 weeks. NR animals held in short days for 8 weeks had significantly decreased testes and epididymal masses than either long-day or short-day 3-week animals. Seminal vesicle mass (D) was significantly less in short-day 8-week animals than all other groups.

Figure 2

RFRP immunohistochemical labeling. Representative photomicrographs of GnIH-ir staining in the DMH of hamsters held in long-day (LD 16:8) photoperiod (A) for 8 weeks, short-day (LD 8:16) photoperiod (B) for 3 weeks, short-day nonresponders (NR; C) held for 8 weeks, and short-day responders (D) held for 8 weeks. High power (200×) photomicrographs are shown for each condition.

Figure 3

RFRP-ir cell numbers and optical density are associated with reproductive condition, independent of photoperiod. Mean (± SEM) number of RFRP-ir cells (top) and cellular optical density (bottom) of animals held in long days for 8 weeks (LD 16:8) or short days (LD 8:16) for 3 weeks or 8 weeks (responders (R) and nonresponders (NR)). Short-day R hamsters held for 8 weeks in short days display significantly fewer number of RFRP-ir cells. Cellular optical density was unaffected by photo-period. *p < 0.05.

Figure 4

RFRP-ir fiber density is associated with changes in photoperiod and reproductive condition in some brain targets. Fiber optical density in the (A) lateral septum, (B) preoptic area, (D) anterior hypothalamus, and (E) paraventricular nucleus of the thalamus was significantly affected by photoperiod in short-day responsive (R) animals held for 8 weeks in short days. In these same nuclei, fiber density was significantly decreased in animals held in short days for 3 weeks. Fiber density was unaffected by photoperiod in the (C) bed nucleus of the stria terminalis and the (F) arcuate nucleus. ^*Significantly less than all groups, excluding those bearing a single asterisk (p < 0.05). ^**Significantly less than all groups (p < 0.05).

Figure 5

RFRP mRNA labeling. Representative photomicrographs depicting RFRP mRNA expression across long-day (LD) and short-day (SD) photoperiods in gonadectomized animals receiving either testosterone replacement (+T) or an empty capsule (sham).

Figure 6

RFRP mRNA expression is affected by photoperiod, independent of testosterone levels. Mean (± SEM) number of RFRP mRNA-expressing cells of gonadectomized animals held in long-day (LD 16:8) and short-day (LD 8:16) photoperiods and receiving either testosterone replacement (+T) or empty capsules (sham). Hamsters held in short days (LD 8:16) exhibited significantly fewer (p < 0.05) RFRP mRNA-expressing cells than groups held in long days, regardless of testosterone treatment. There was no significant effect of testosterone in either photoperiod. *p < 0.05.