Heifers express G‐protein coupled receptor 153 in anterior pituitary gonadotrophs in stage‐dependent manner

Abstract

We recently found that orphan G-protein-coupled receptor (GPR)153 is expressed in the anterior pituitary (AP) of heifers, leading us to speculate that GPR153 colocalizes with gonadotropin-releasing hormone receptor (GnRHR) in the plasma membrane of gonadotrophs and is expressed at specific times of the reproductive cycle. To test this hypothesis, we examined the coexpression of GnRHR, GPR153, and either luteinizing hormone or follicle-stimulating hormone in AP tissue and cultured AP cells by immunofluorescence microscopy. GPR153 was detected in the gonadotrophs, and was colocalized with GnRHR in the plasma membrane. GPR153 was also detected in the cytoplasm of cultured gonadotrophs. Real-time PCR and western blot analyses found that expression was lower (P < 0.05) in AP tissues during early luteal phase as compared to pre-ovulation or late luteal phases. The 5 0 -flanking region of the GPR153 gene contained a consensus response element sequence for estrogen, but not for progesterone. These data suggest that some, but not all GPR153 colocalizes with GnRHR in the plasma membrane of gonadotrophs, and its expression changes stage-dependently in the bovine AP.

Discovery of new receptors regulating luteinizing hormone and follicle-stimulating hormone secretion by bovine gonadotrophs to explore a new paradigm for mechanisms regulating reproduction

“…6c) colocalize with GnRHRs in the limited area of the plasma membrane in bovine gonadotrophs. The positive overlap coefficient calculated by confocal data analysis software was very strong (more than 0.70) between GnRHR and GPR61, between GnRHR and GPR153, and between GnRHR and AMHR2 on the cell-surface of gonadotrophs [33][34][35].…”

“…Real-time PCR and western blot analyses found that GPR153 expression was significantly lower in AP tissues during the early luteal phase as compared to pre-ovulation or the late luteal phases [34]. The 5ꞌ-flanking region of the GPR153 gene contained a consensus response element sequence for estrogen, but not for progesterone [34]. None of the few reports published describe the function of GPR153 or identify its ligand(s).…”

“…Therefore, I speculated that GPR61, GPR153, and AMHR2 might colocalize with GnRHR in the limited area of the plasma membrane of gonadotrophs. To test this hypothesis, I examined the colocalization of GnRHR with each one of these receptors (and with either the LHβ subunit or FSHβ subunit) in both AP tissue and cultured AP cells using immunofluorescence microscopy [33][34][35]. I found that GPR61 (Fig.…”

Anti-Müllerian hormone receptor type 2 is expressed in gonadotrophs of postpubertal heifers to control gonadotrophin secretion

“…The lipid rafts are distinct, relatively insoluble regions that have lower density and are less fluid than surrounding membrane (Simons et al 2000;Head et al 2014), and they facilitate signaling by allowing colocalization of membrane receptors and their downstream signaling components (Simons et al 2000;Head et al 2014). We recently discovered that two orphan receptors, GPR61 and GPR153, are colocalized with GnRHR in gonadotroph plasma membrane lipid rafts (Pandey et al 2017a(Pandey et al , 2017b. Therefore, gonadotroph lipid rafts containing GnRHR may contain AMHR2.…”

Bovine gonadotrophs express anti-Müllerian hormone (AMH): comparison of AMH mRNA and protein expression levels between old Holsteins and young and old Japanese Black females

“…We used also a specific guinea pig polyclonal anti-bovine GnRHR (Kadokawa et al 2014), a specific mouse monoclonal anti-LH β (LHβ) subunit antibody (clone 518-B7; Matteri et al 1987;Iqbal et al 2009), and a specific mouse monoclonal anti-FSH β (FSHβ) subunit antibody (clone A3C12; Borromeo et al 2004) for immunohistochemical analysis of AP tissue. These antibodies are utilized for immunohistochemical analysis of AP tissue in our previous papers (Kadokawa et al 2014;Pandey et al 2017aPandey et al , 2017bKereilwe et al 2018) Western Blotting for AMH detection Proteins were extracted from the AP (n = 4) or ovary samples (n = 4, used as positive controls) and western blotting was performed as previously described (Kereilwe et al 2018). The extracted protein sample (33.4µg of total protein in 37.5µl) was mixed in…”