A possible mechanism contributing to the synergistic action of vasotocin (VT) and corticotropin-releasing hormone (CRH) receptors on corticosterone release in birds

Arginine vasotocin (AVT) and corticotropin-releasing hormone (CRH) are two neuronal regulators in the hypothalamic-pituitary-adrenal (HPA) axis that modulate biological responses to stress in avian species. When AVT and CRH are administered together in vitro or in vivo, levels of adrenocorticotropic hormone (ACTH) or plasma corticosterone (CORT) are released, respectively, in a synergistic manner. The underlying mechanism of this greater than additive stress response was investigated by expressing the vasotocin receptor type 2 (VT2R) and CRH receptor type 1 (CRH-R1), both G-protein coupled receptors, in HeLa cells. Fluorescence resonance energy transfer (FRET) analysis provided the evidence for heterodimerization of the VT2R/CRH-R1 in the presence of their respective ligands, AVT and CRH. The VT2R and CRH-R1 were tagged at the C-terminal ends with either cyan fluorescent protein (CFP) or yellow fluorescent protein (YFP), and a VT2R chimera was constructed by replacing the fourth transmembrane region (TM4) of the VT2R with TM-IV of the β2-adrenergic receptor (β2AR). When VT2R/β2AR chimera and CRH-R1 were expressed in HeLa cells, heterodimerization was partly disrupted. Taken together, these data indicate that TM-IV of the VT2R may provide an important interface for effective receptor dimerization, suggesting that direct molecular interaction between VT2R and CRH-R1 receptors plays a role in mediating an enhanced interaction between these two receptors. Their interaction at the anterior pituitary level may potentiate the endocrine output of the avian HPA system.