Long-term estradiol-17β exposure decreases the cholinergic innervation pattern of the pig ovary

Elevated levels of endogenous estrogens in the course of pathological states of ovaries, as well as xenoestrogens, may lead to hyperestrogenism. It has previously been demonstrated that long-term estradiol-17β (E₂) administration in adult gilts affected the population of sympathetic intraovarian nerve fibers. The aim of this study has been to determine the effect of long-term E₂ exposure on the cholinergic innervation pattern of porcine ovaries. Intraovarian distribution and the density of nerve fibers immunoreactive (IR) to vesicular acetylocholine transporter (VAChT) and/or neuronal isoform of nitric oxide synthase (nNOS), vasoactive intestinal polypeptide (VIP), somatostatin (SOM) were determined. From day 4 of the first estrous cycle to day 20 of the second studied cycle, experimental gilts were intramuscularly injected with E₂, while control gilts received corn oil. The ovaries were then collected and processed for double-labelling immunofluorescence. After E₂ administration, the total number of fibers IR to VAChT, nNOS and VIP decreased significantly. The numbers of VAChT-, nNOS- and VIP-IR fibers within the ground plexus were significantly lower, while they were significantly higher around small or medium tertiary follicles. In the E₂-affected ovaries, the numbers of nNOS- and VIP-IR fibers were significantly higher near secondary follicles and VAChT-IR in the vicinity of medullar blood vessels. In turn, around the latter structures there were significantly lowered populations of nNOS- and VIP-IR nerve fibers. These results suggest that the elevated E₂ levels that occur during pathological states may affect the cholinergic innervation pattern of ovaries and their function(s).