Abstract
Blood Anti-Müllerian hormone (AMH) level significantly indicates fertility in animals, including women and cows, via unclarified physiological mechanisms. AMH stimulate the LH and FSH secretions from bovine gonadotrophs via its main receptors, AMH receptor type 2 (AMHR2) (Kereilwe et al. 2018). We hypothesized that AMH and AMHR2 are expressed in the most important parts of brain for reproduction: preoptic area (POA), arcuate nucleus (ARC), and median eminence (ME).We collected brain blocks containing either the POA or the ARC and ME (ARC&ME) from Japanese Black heifers (n = 5), following the method of Tanco et al. (2016). We previously developed a new method to quench autofluorescence in bovine brain samples. After our original quenching, we used the free-floating method to stain the POA or ARC&ME coronal sections [50 µm thickness, prepared based on the bovine brain atlas (Okamura 2002))] using the verified anti-human AMH and anti-bovine AMHR2 antibodies with bovine ovaries (Kereilwe et al. 2018; Kereilwe and Kadokawa 2019).We also collected tissues containing either the POA or the ARC&ME from other heifers (n = 5) to perform RT-PCR or western blotting (WB) using previously reported methods.The RT-PCR and WB detected both AMH and AMHR2 expressions in the POA tissues as well as in the ARC&ME tissues.Immunohistochemistry revealed that approximately 65% of GnRH cells express AMHR2, and 50% of GnRH cells express AMH in the POA, ARC, and ME, including the secretory zone to the pituitary portal blood. We also found that GnRH cells were situated around other AMH-positive GnRH and non-GnRH cells.Therefore, both AMH and AMHR2 are expressed in the bovine POA, ARC, and ME.
