Changes in the cholinergic innervation pattern of porcine ovaries with cysts induced by dexamethasone administration

Abstract

We revealed earlier that induction of ovarian cysts in gilts by dexamethasone phosphate disodium salt (DXM) administration from the follicular phase of the estrous cycle (EC) changed the cholinergic innervation of the gonad. In the present study, the innervation of porcine ovaries by vesicular acetylcholine transporter (VAChT)-, neuronal nitric oxide synthase (nNOS)-, vasoactive intestinal peptide (VIP)- and somatostatin (SOM)-immunoreactive (IR) fibres, after induction of cystic changes from the middle luteal phase of the EC, was determined. The cystic changes were induced by DXM injections from days 7 to 21 of the EC, and 11 days later, the ovaries were collected. In the cystic ovaries, VAChT-, nNOS- and SOM-IR fibres were found around cysts and small tertiary follicles; nNOS-IR and also VAChT-IR fibres were observed near secondary follicles and veins; and VAChT- and nNOS-IR fibres were not found around cortical arteries. The number of VIP-IR fibres increased near the cysts and within the ground plexus, while the number of VAChT-IR fibres decreased within the medullar part of this structure. Thus, our study showed changes in the cholinergic innervation pattern of the porcine cystic ovaries induced from the middle phase of the cycle and confirmed that cystic ovary innervation depends partly on the phase of the EC in which the induction of cysts was started.

Figs. 1–7

Immunohistochemical localization of VAChT- and nNOS-IR nerve fibres in the cortical part of the ovary of the control (CON) and DXM-treated (DXM) gilts. Lack of VAChT and/or nNOS-IR nerve fibres around the secondary (1a and b, respectively) and tertiary follicles (up to 3 mm; 3a and b, respectively; 4–6 mm in diameter; 5a and b, respectively) in the CON group. Nerve fibres containing VAChT and/or nNOS near the secondary (2a and b, respectively) and tertiary up to 3 mm (4a and b, respectively) follicles and cyst (6a and b, respectively) in the DXM group. The co-localization of VAChT and nNOS in the same nerve fibres supplying secondary (2a and b) and tertiary up to 3 mm (4a and b) follicles and cyst (6a and b) in the cystic-changed ovaries. Negative control for VAChT (7a) and nNOS (7b) in the ovary of the CON gilt. Arrows, nerve fibres; SF secondary follicle, TF <3 tertiary follicle up to 3 mm in diameter, TF 4–6 tertiary follicle 4–6 mm in diameter, C cyst. Scale bar 25 μm

Figs. 8–11

Immunohistochemical localization of VAChT- and nNOS-IR nerve fibres in the medullar part of the ovary of control (CON) and DXM-treated (DXM) gilts. A decrease in the number of VAChT-IR nerve fibres in the area of the ground plexus in the DXM group (9a) compared to the CON group (8a). Nerve fibres containing nNOS observed in the area of this structure in the CON group (8b) and the same number of these nerve fibres forming bundles (9b) found after DXM injections. The presence nNOS-IR nerve fibres in the vicinity of an artery in the control gilt (10) and their lack in the ovary of DXM-treated animal (11). Co-localization of VAChT and nNOS in the same nerve fibres in the area of the ground plexus (8a and b) in the CON group. Arrows, nerve terminal; A artery. Scale bar 25 μm

Figs. 12–22

Immunohistochemical localization of VIP-IR nerve fibres in the cortical part of the ovary of the control (CON) and DXM-treated (DXM) gilts. An increase in the number of VIP-positive nerve fibres, formed bunches, in the area of the ground plexus (13) as well as near the cyst (19) in the ovary of the DXM-treated animal compared to the CON group (12 and 18, respectively). A lack of VIP-IR nerve fibres around the primordial (15) and secondary (17) follicles as well as the vein (21) after DXM administration and their presence in the CON group (14, 16 and 20, respectively). Negative control for VIP in the ovary of the CON gilt (22). Arrows, nerve fibres; PF primordial follicle, SF secondary follicle, TF 4–6 tertiary follicle 4–6 mm in diameter, C cyst, V vein. Scale bar 25 μm

Figs. 23–24

Immunohistochemical localization of VIP-IR nerve fibres in the medullar part of the ovary of the control (CON) and DXM-treated (DXM) gilts. An increase in the number of VIP-IR nerve fibres, often forming bundles, in the area of the ground plexus in the cystic-changed ovary (24) compared to the control gonad (23). Arrows, nerve fibres. Scale bar 25 μm

Figs. 25–29

Immunohistochemical localization of SOM-IR nerve fibres in the cortical part of the control (CON) and DXM-treated (DXM) gilts. SOM-IR nerve fibres were not observed within the ground plexus (25) as well as around the tertiary follicles (4–6 mm in diameter; 27) in the control gilts, while they were present in the ovaries of DXM-treated gilts (26 and 28, respectively). Negative control for SOM (29) in the ovary of the CON gilt. Arrows, nerve fibres; TF 4–6 tertiary follicle 4–6 mm in diameter, C cyst. Scale bar 25 μm

Figs. 30–31

Immunohistochemical localization of SOM-IR nerve fibres in the medullar part of the control (CON) and DXM-treated (DXM) gilts. An increase in the number of the SOM-IR nerve fibres in the area of the ground plexus in the DXM group (31) compared to the CON group (30). Arrows, nerve fibres. Scale bar 25 μm