Perinatal exposure to the fungicide ketoconazole alters hypothalamic control of puberty in female rats

Abstract

IntroductionEstrogenic endocrine disrupting chemicals (EDCs) such as diethylstilbestrol (DES) are known to alter the timing of puberty onset and reproductive function in females. Accumulating evidence suggests that steroid synthesis inhibitors such as ketoconazole (KTZ) or phthalates may also affect female reproductive health, however their mode of action is poorly understood. Because hypothalamic activity is very sensitive to sex steroids, we aimed at determining whether and how EDCs with different mode of action can alter the hypothalamic transcriptome and GnRH release in female rats.DesignFemale rats were exposed to KTZ or DES during perinatal (DES 3-6-12μg/kg.d; KTZ 3-6-12mg/kg.d), pubertal or adult periods (DES 3-12-48μg/kg.d; KTZ 3-12-48mg/kg.d).ResultsEx vivo study of GnRH pulsatility revealed that perinatal exposure to the highest doses of KTZ and DES delayed maturation of GnRH secretion before puberty, whereas pubertal or adult exposure had no effect on GnRH pulsatility. Hypothalamic transcriptome, studied by RNAsequencing in the preoptic area and in the mediobasal hypothalamus, was found to be very sensitive to perinatal exposure to all doses of KTZ before puberty with effects persisting until adulthood. Bioinformatic analysis with Ingenuity Pathway Analysis predicted “Creb signaling in Neurons” and “IGF-1 signaling” among the most downregulated pathways by all doses of KTZ and DES before puberty, and “PPARg” as a common upstream regulator driving gene expression changes. Deeper screening ofRNAseq datasets indicated that a high number of genes regulating the activity of the extrinsic GnRH pulse generator were consistently affected by all the doses of DES and KTZ before puberty. Several, including MKRN3, DNMT3 or Cbx7, showed similar alterations in expression at adulthood.ConclusionnRH secretion and the hypothalamic transcriptome are highly sensitive to perinatal exposure to both DES and KTZ. The identified pathways should be exploredfurther to identify biomarkers for future testing strategies for EDC identification and when enhancing the current standard information requirements in regulation.

Comparison of female rat reproductive effects of pubertal versus adult exposure to known endocrine disruptors

“…The same pattern was seen in animals exposed to DES-0.048 during adulthood. Hypothalamic GnRH secretion was not affected in these females ( 16 ). Unfortunately, we do not have measurements of luteinizing hormone/follicle stimulating hormone and it is therefore difficult to deduce if observed ovarian changes are associated with local and/or central effects of DES.…”

“…Using two compounds that we know have endocrine disrupting effects in humans enables comparison of the two exposure windows, as well as endpoint sensitivity using an established OECD test guideline ( 3 ). Perinatal exposure to these endocrine disruptors has been shown to affect reproductive development ( 1 , 2 , 16 , 17 ) and they are expected to display different effect patterns depending on age at exposure start.…”

Endocrine disrupting chemicals and male fertility: from physiological to molecular effects

“…Several experimental studies on rats have reported that perinatal, juvenile, or adult exposure to EDCs, disrupt the hypothalamic control of pituitary gonadotropin synthesis leading to a disruption of gonadal steroid production and androgens cyclicity ( 58–62 ). These studies suggest that EDCs exposure affects the expression of GnRH and kisspeptin, influences the pulsatile launch of GnRH, and interferes with the regulation of gonadotropin production from the pituitary axis ( 63 ).…”