Binding and Signaling Properties of the Leu8 and Pro8 Isoforms of Oxytocin for Oxytocin Receptors from Four Primate Species

Abstract

Among New World monkeys (NWM), there is a remarkable correlation between those species exhibiting social monogamy and biparental behavior and their expression of an oxytocin (OT) variant with proline at the eighth position (Pro8OT) instead of the common mammalian OT ligand that has leucine at position 8 (Leu8OT). In light of evidence for OT involvement in normative and dysfunctional social behavior, a better understanding of the impact of ligand variation on OT binding and signaling could provide insights into the pharmacological modulation of sociality. We tested the hypothesis that Leu8OT and Pro8OT would bind differently to the OT receptors (OTRs) from Leu8OT‐versus Pro8OT‐expressing species. OTRs from four primate species were expressed in Chinese hamster ovary cells, each species representing a unique combination of ancestral lineage, breeding system, and native OT ligand: marmoset (Pro8OT, monogamous, NWM), macaque (Leu8OT, nonmonogamous, Old World monkey), Titi monkey (Leu8OT, monogamous, NWM; an exception to the pattern), and human (Leu8OT). Saturation assays of binding of the OTR antagonist radioligand 125I‐OVTA to intact cells on ice yielded Bmax values (fmol per well of a 96‐well plate) of approximately 100 for marmoset, 10 for macaque, 10 for human, and 30 for Titi. KD values in pM were 350 for marmoset, 150 for macaque, 290 for Titi, and 140 for human. Competition binding assays were conducted with Pro8OT, Leu8OT, and the closely related nonapeptide hormone arginine vasopressin (AVP) for OTRs from all of four species. Contrary to our hypothesis, all of the OTRs exhibited higher affinity for Pro8OT than for Leu8OT. IC50 values in nM for Pro8OT, Leu8OT, and AVP, respectively, were 150, 400, and 2900 for marmoset, 40, 70, and 460 for macaque, 190, 960, and 3900 for Titi, and 20, 90, and 520 for human. Preference for Pro8OT versus Leu8OT was about 2‐fold for marmoset and macaque and about 4‐fold for Titi and human. All of the receptors exhibited a similar 4‐ to 7‐fold lower affinity for AVP than for Leu8OT, as expected. Calcium signaling downstream of OTRs was assessed with FlexStation assays. EC50 values in pM for Pro8OT, Leu8OT, and AVP, respectively, were 50, 160, 430 for marmoset, 1340, 2100, and 7800 for macaque, 600, 1000, and 4900 for Titi, and 70,130, and 230 for human. The higher potencies for marmoset and Titi are likely due to the higher OTR expression in those cells. Preference for Pro8OT versus Leu8OT for calcium signaling was less than 2‐fold for all species except marmoset, with a 3.2‐fold preference. The human receptor showed the smallest preference for Leu8OT versus AVP. Together these data show that differences in OTR ligand binding affinities and/or calcium signaling potencies do not provide an obvious explanation for the different behaviors of Pro8OT‐versus Leu8OT‐expressing species. Differential signaling to other downstream responses remains to be investigated.Support or Funding InformationSupported by NIH grant 1‐R01‐HD089147 to JAF, TFM, and MLT.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.