Premium
Effects of Oxytocin Analogs on Neurite Outgrowth and Cellular Signaling in Human SH‐SY5Y Neuroblastoma Cell Line
Author(s) -
Pierce Marsha,
Vattem Nishita,
Scoles Madeline,
Gore Ashleigh,
Aziz Areej,
Gehr Marissa
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.03838
Subject(s) - neurite , receptor , biology , oxytocin , neuropeptide , neuroscience , sh sy5y , vasopressin , oxytocin receptor , signal transduction , endocrinology , microbiology and biotechnology , medicine , neuroblastoma , biochemistry , cell culture , in vitro , genetics
In the central nervous system, arginine vasopressin (AVP) and oxytocin (OT) stimulate neural networks that regulate social behaviors, including social attachment, aggression, and complex social cognition. AVP and OT are neuropeptides that are expressed in the social behavioral network and bind to G‐protein coupled receptors, stimulating cellular signaling patterns that ultimately affect social behaviors at the organismal level. Consensus mammalian AVP and OT sequences are highly conserved. In addition to sequence homology between AVP and OT, there is ~85% structural homology between the OT receptor (OTR) and vasopressin 1a receptor (V1aR) resulting in significant cross‐reactivity between the ligands and receptors. Perturbations in OT and/or OT receptor expression results in social behavioral deficits, and is associated with a number of psychopathologies including autism spectrum disorder, schizophrenia, anxiety, and depression. Methods The human neuroblastoma derived SH‐SY5Y cell line demonstrates morphological characteristics of neurons including neurites, endogenously expresses OTR, and is often used as an in vitro model for human neurons. Functional assays were performed using Fluo8‐AM to measure ligand‐induced Ca 2+ mobilization. Sigmoidal dose‐response curves for OT analogs were generated using three‐parameter nonlinear regression analysis in GraphPad Prism 8.3 software. To assess effects on neurite outgrowth, cells were treated with logarithmic doses of the consensus mammalian OT sequence (Cys‐Tyr‐Ile‐Gln‐Asn‐Cys‐Pro‐ Leu ‐Gly; Leu8‐OT) and a new world monkey analog (Cys‐Tyr‐Ile‐Gln‐Asn‐Cys‐Pro‐ Pro ‐Gly; Pro8‐OT) for 24 hours, fixed and stained with phalloidin and DAPI. The Nikon A1R confocal microscope was used to collect 60X images of GFP‐expressing neurons assessed for effects on neurite outgrowth using the Image J package FIJI with the NeuronJ plugin. Results In SH‐SY5Y cells, both OT analogs were more potent than AVP at inducing Ca 2+ mobilization, which is consistent with OT being more potent and efficacious at the human OTR. Specificity of the signaling at the OTR were confirmed using the vasopressin 1a receptor antagonist SR49059 and the oxytocin receptor antagonist L‐371,257. Additionally, both OT analogs show a dose‐dependent increase in neurite outgrowth. Conclusions Together, these data suggest that OTR mediated signaling pathways lead to neurite outgrowth in SH‐SY5Y cells. Integrative studies of behavior, genetics and ligand‐receptor interaction are crucial for translating signaling activation at the cellular level to effects of AVP and OT ligands on social behavior. Knowledge of how OT alters neuronal structure and function has the potential to both identify mechanisms that produce social dysfunction and to inform the development of therapeutic agents.