Table 8.

Developmental EDC Effects on GnRH-Gonadotropin and Kisspeptin Systems in Vivo

EDC	Animal	Sex	Exposure Period	Dosage, Route	Age at Analysis	Endpoint(s)	GnRH/LH Effect	Kisspeptin Effect	Ref.
BPA, MXC	Sheep	F	E30–E90 (full term is 147 d)	5 mg/kg/d via daily sc injections to ewe	Adulthood, ∼21 mo	GnRH mRNA in anterior hypothalamus and POA	GnRH mRNA was decreased by both MXC and BPA ↓		1117
BPA, MXC	Sheep	F	E30–E90 (full term is 147 d)	5 mg/kg/d via daily sc injections to ewe	OVX at 21 mo of age	LH response to steroid feedback and to GnRH challenge	LH and FSH response to steroid negative feedback and LH response to positive feedback were unaffected by the EDCs. GnRH-induced LH release was similarly unaffected		1156
EDC mix from sewage sludge	Sheep	Both sexes combined	E0–E110	Sewage sludge was applied to pastures; control pastures received conventional inorganic fertilizer	Ewes and fetuses were euthanized at E110.	Kiss1, Kiss1r, Esr1 mRNA in rostral, medial and caudal hypothalamus; colocalization of kisspeptin and LHβ and ERα in the pituitary gland by dual IHC	Numbers of LHβ cells, and those co-expressing ERα in pituitary, were decreased in the sewage sludge group ↓.	Kiss1 was significantly decreased ↓ through the hypothalamus and in the pituitary of sewage-sludge exposed fetuses. Kiss1r (GPR54), and Esr1 mRNA were unaffected in hypothalamus and pituitary . Numbers of kisspeptin cells, double-labeled kisspeptin/LHβ cells, and double-labeled kisspeptin/ERα cells, were decreased in the sewage sludge fetuses ↓	1158
PCBs	Sheep	F	E0 to birth	Ewes were gavaged 3 × weekly with PCB153 (98 μg/kg/d or PCB118 (49 μg/kg/d	Lambs	GnRH-induced LH and FSH release	Basal LH and FSH concentrations did not differ among groups . The PCB153 group had significantly greater GnRH-induced LH release, and slightly but not significantly higher FSH release ↑. No effect of PCB 118 was found for this endpoint		419
BPA	Mouse (C57BL/6J)	M	E15–P21	Gavage of dams 50 μg or 5 mg/kg/d	8 wk	Kisspeptin immunolabeling in the MPOA and AVPV		Kisspeptin immunoreactive neuron numbers were unaffected by BPA	1110
BPA	Mouse (C57bl/6J)	F	E15–P21	0.05 or 5 mg/kg/d by gavage to dam	8 wk (young adult)	GnRH, kisspeptin, and ERα immunolabeling	Numbers of GnRH immunoreactive neurons and the percentage of those with kisspeptin apposition were unchanged by BPA	Numbers of kisspeptin neurons were unchanged in AVPV and caudal periventricular nucleus , and increased in rostral periventricular nucleus ↑	1115
BPA	Rat (Wistar)	F	P1–P7	0.05 or 20 mg/kg, sc	P100, OVX + estradiol to induce the GnRH/LH surge	Gnrh1 mRNA and primary transcript; serum LH; ERα protein by IHC in AVPV and ARC	Gnrh1 mRNA was increased by the low-dose BPA↑. GnRH primary transcript (presence of intron A) was decreased by both BPA dosages ↓. Serum LH was decreased by high-dose BPA ↓		572
BPA	Rat (Long-Evans)	F/M	P0–P3	50 μg or 50 mg/kg/d	Adults (P148 or older), females OVX and hormone primed with estradiol + progesterone. Males were intact	Kisspeptin fiber density in the AVPV and ARC		Hormone priming in females reduced kisspeptin fiber density in AVPV, but not in BPA females ↓. In the ARC, hormones reduced kisspeptin immunoreactivity, but not in the low-dose BPA group ↓. No effect in males	1154
BPA	Rat (Long-Evans)	F/M	P0–P3	50 μg or 50 mg/kg/d	Adults (P148 or older). Females were OVX and hormone primed with estradiol + progesterone	GnRH-Fos double labeling by IHC, and number of GnRH neurons	Number of GnRH neurons was unaffected; % of GnRH neurons that co-expressed Fos was unaffected		545
BPA	Rat (GnRH-GFP transgenic Wistar)	F/M	P0–P3	50 μg or 50 mg/kg/d	Female (P17, P21, P24, P28, P33); male (P21, P33)	GnRH was detectable by GFP labeling. Kisspeptin and RFRP3 were detected by IHC	Kisspeptin appositions on GnRH neurons were unaffected by BPA . Appositions of RFRP3 on GnRH neurons were decreased in the low-BPA females ↓	In AVPV, kisspeptin fiber density was unaffected by BPA . In ARC, in P33 females, the high-BPA exposure group had decreased kisspeptin ↓, similar to that in males. RFRP3 fiber density was decreased by low-dose BPA in the females at P28 and P33 ↓	1155
BPA	Mouse (CD-1)	M/F	E1 to P20 to dams. Some pups continued gavage from P21 to 49	12, 25, or 50 mg/kg/d to dames via gavage	P50 (females on proestrus)	GnRH, Kisspeptin, GPR54 (hypothalamus); and pituitary mRNAs (including LHβ, FSHβ, and others)	Gnrh1: increased in both sexes ↑. Pituitary LH was unaffected ; FSH was increased in both sexes ↑. TSH, GH, PRL were unaffected	Kiss1 was increased in both sexes ↑. Kiss1r was unaffected	434
BPA	Rat (S-D)	M/F	E10 to P7	2 μg/kg/d sc to dam	Male at P30, P50, P90. Males from BPA dams, and males and females from vehicle dams, were also gonadectomized and given estradiol for up to 7 d. Intact rats were also used	Number of kisspeptin immunoreactive cells in the AVPV. Number of GnRH cells in the POA	BPA slightly but significantly increased GnRH+ cells in the male POA ↑. When the LH surge was measured, females and BPA males, but not control males, exhibited an LH surge ↑	In gonadectomized, estradiol-treated rats, BPA increased AVPV kisspeptin neuron numbers at all 3 ages ↑, making them closer to the female (higher) levels. In intact rats, BPA kisspeptin cell numbers were also higher in both estradiol-treated and untreated males ↑. The LH surge in females and in BPA males was attenuated by the GPR54 inhibitor P234 ↓	1153
BPA	Rat (S-D)	F	P1–P10	BPA 50 (2.5–6.2) or BPA 500 (25–62 mg/kg/d) given sc to pups	P13 or P120 (in estrus)	GnRH pulsatility, GnRH-induced LH release	At P13, in vivo GnRH-induced LH release was decreased by BPA 500 ↓, the number of GnRH peaks/hour in an in vitro perifusion increased ↑, and the interpulse interval decreased ↓. At P120, GnRH-induced LH release was decreased at 15 and 50 min post-BPA ↓		525
DBP	Rat (S-D)	F	P1–P5 (neonatal) or P26–P30 (peripubertal)	0.5, 5, or 50 mg/kg/d given sc to pups	Adulthood (on proestrus)	mRNA of Gnrh1 and Kiss1r in the AVPV and ARC	GnRH1 mRNA in the AVPV and ARC was increased by DBP (5 mg/kg group only) in the neonatal group ↑. For the pubertal group, Gnrh1 was increased (0.5 mg/kg) ↑ or decreased (5 mg/kg) ↓ in AVPV, and increased (0.5 mg/kg) ↑ or decreased (5, 50 mg/kg) ↓ in the ARC	Kiss1r in AVPV was increased by DBP (5 mg/kg) in the neonatal group ↑, and increased (0.5 mg/kg) ↑ or decreased (50 mg/kg) ↓ in the peripubertal group. In the ARC, Kiss1r was decreased (all doses) ↓ in the neonatal group, and increased (0.5, 50 mg/kg) ↑ or decreased (5 mg/kg) ↓ in the peripubertal group. Kisspeptin immunoreactivity in AVPV was increased (0.5 mg/kg, neonatal group) ↑. In the ARC, kisspeptin immunoreactivity was increased (5 mg/kg, neonatal) ↑ or decreased (all dosages, peripubertal) ↓	555
PCBs	Rat (S-D)	M/F	E16, E18	1 mg/kg of A1221 or a mix of PCBs 138, 153, 180, given ip to dam on E16 and E18	P1	Gene expression in the POA	GnRH1 mRNA was unaffected by PCB treatment	Kiss1 mRNA was unaffected . Kiss1r mRNA was significantly decreased by PCBs in males ↓ and increased in females ↑	1157
PCBs	Rat (S-D)	M/F	E16, E18	1 mg/kg of A1221 or a mix of PCBs 138, 153, 180, ip to dam on E16 and E18	P60 (females on proestrus)	Gene expression in the POA, and IHC for kisspeptin, GnRH-Fos, and ERα in the AVPV	GnRH-Fos double labeling was significantly decreased by PCBs in females on proestrus ↓. mRNA for Gnrh1 was unaffected in both sexes	Kisspeptin immunofluorescence density was significantly decreased by PCBs in the female ↓, but not the male , AVPV. mRNA for Kiss1 and Kiss1r were unaffected, both sexes	567
PCBs	Rat (S-D)	M/F	E16, E18	1 mg/kg of A1221, ip to dam on E16 and E18	P15, P30, P45, P90	mRNA of 48 genes in the AVPV and ARC	In females, A1221 masculinized Gnrh1 in the AVPV ↓	In females, A1221 masculinized expression of Kiss1r in the AVPV ↓, and Kiss1 was increased by A1221 in the ARC ↑	1145

Abbreviations: F, female; IHC, immunohistochemistry; M, male; S-D, Sprague-Dawley. Directionality of responses: ↓, down-regulated; ↑, up-regulated; , no change.