Simultaneous oxytocin and arg-vasopressin measurements in microdialysates using capillary liquid chromatography–mass spectrometry

Oxytocin (OXT) and arg-vasopressin (AVP) are nonapeptides with many important functions both peripherally and centrally. Intracerebral microdialysis has helped characterize their importance in regulating complex social and emotional processes. Radioiummunoassay is the most commonly used analytical method used for OXT and AVP measurements in microdialysates. These measurements have several well-known issues including single peptide per assay limit, possible cross-reactivity between structurally related peptides, and laborious sample preparation with radioactive materials. Here we demonstrate the use of capillary LC–MS3 for measuring OXT and AVP simultaneously in dialysates at a 10 min sampling frequency. Microdialysate samples required no preparation and instrumentation was commercially available. Microdialysis probes made with polyacrylonitrile membranes were suitable for high level recovery of the peptides in vitro and in vivo. Responses were linear from 1 to 100 pM. Matrix effect was assessed by standard addition experiments and by comparing signal intensities of OXT and AVP standards made in aCSF or dialysate. It was determined that the online washing step used on this setup was adequate for removing contaminants which interfere with electrospray ionization efficiency. In vivo, both peptides were stimulated by high K+ (75 mM) aCSF perfusion in the paraventricular nucleus (PVN). Also, a systemic injection of high Na+ (2 M) caused a rapid and transient increase in PVN OXT while AVP increased only after 1.5 h. Our findings suggest that capillary LC–MS3 is a straightforward method for monitoring OXT and AVP simultaneously from complex samples such as dialysates.