The oxytocin-arginine vasopressin (OT-AVP) ligand-receptor family influences a variety of physiological, behavioral, and social behavioral processes in the brain and periphery. The OT-AVP family is highly conserved in mammals, but recent discoveries have revealed remarkable diversity in OT ligands and receptors in New World Monkeys (NWMs) providing a unique opportunity to assess the effects of genetic variation on pharmacological signatures of peptide ligands. The consensus mammalian OT sequence has leucine in the 8^th position (Leu⁸-OT), whereas a number of NWMs, including the marmoset, have proline in the 8^th position (Pro⁸-OT) resulting in a more rigid tail structure. OT and AVP bind to OT’s cognate G-protein coupled receptor (OTR), which couples to various G-proteins (G_i/o, G_q, G_s) to stimulate diverse signaling pathways. CHO cells expressing marmoset (mOTR), titi monkey (tOTR), macaque (qOTR), or human (hOTR) OT receptors were used to compare AVP and OT analog-induced signaling. Assessment of G_q-mediated increase in intracellular calcium (Ca²⁺) demonstrated that AVP was less potent than OT analogs at OTRs from species whose endogenous ligand is Leu⁸-OT (tOTR, qOTR, hOTR), relative to Pro⁸-OT. Likewise, AVP-induced membrane hyperpolarization was less potent at these same OTRs. Evaluation of (Ca²⁺)-activated potassium (K⁺) channels using the inhibitors apamin, paxilline, and TRAM-34 demonstrated that both intermediate and large conductance Ca²⁺-activated K⁺ channels contributed to membrane hyperpolarization, with different pharmacological profiles identified for distinct ligand-receptor combinations. Understanding more fully the contributions of structure activity relationships for these peptide ligands at vasopressin and OT receptors will help guide the development of OT-mediated therapeutics.