title: Temporal profile of arginine vasopressin release from the neurohypophysis in response to hypertonic saline and hypotension measured using a fluorescent fusion protein.
abstract: Methods currently employed to study the release of hormones such as arginine vasopressin (AVP), while sensitive, suffer from a low temporal resolution such that the monitoring of AVP release on a moment-to-moment basis is not possible. Here, we describe a new approach to indirectly monitor the temporal profile of AVP release from the neurohypophysis of transgenic rats expressing an AVP-eGFP fusion gene. Using fibre-optic probes (termed 'optrodes') we were able to indirectly monitor AVP release via a reporter moiety in real-time. This method is a major advance over current methods used to monitor AVP release. Intravenous administration of hypertonic saline (3M NaCl) induced a rapid (latency of 2-3s) increase in fluorescence detected in the neurohypophysis that lasted on average for 60s - a response that was highly reproducible. Infusion of sodium nitroprusside induced a rapid fall in blood pressure accompanied by a rapid, stimulus-locked increase in fluorescent signal that returned to baseline with the recovery of blood pressure to pre-stimulus levels - again this response was highly reproducible. Withdrawal of blood (to simulate haemorrhage) also resulted in a stimulus-locked increase in fluorescence that return to baseline after the withdrawn blood was returned to the animal. In conclusion, we developed a highly sensitive approach that allows the indirect measurement of AVP release via the monitoring of a reporter gene in real-time. This technology can be adapted to permit the study of a whole array of neurohormones/chemicals in transgenic animals expressing a fluorescent reporter construct.
authors.affiliation: Henry Wellcome Laboratories for Integrative Neuroscience and Endocrinology, University of Bristol, Bristol BS1 3NY, UK. song.yao@florey.edu.au
authors.forename: Song T
authors.initials: ST
authors.lastname: Yao
authors.forename: Vagner R
authors.initials: VR
authors.lastname: Antunes
authors.forename: Peter M J
authors.initials: PM
authors.lastname: Bradley
authors.forename: Sergey
authors.initials: S
authors.lastname: Kasparov
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Ueta
authors.forename: Julian F R
authors.initials: JF
authors.lastname: Paton
authors.forename: David
authors.initials: D
authors.lastname: Murphy
mesh_terms: D000818:Animals; D001127:Arginine Vasopressin / Q000032:analysis / Q000378:metabolism*; D005260:Female; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000378:metabolism*; D007022:Hypotension / Q000175:diagnosis / Q000378:metabolism*; D008297:Male; D010904:Pituitary Gland, Posterior / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D055647:Rats, Transgenic; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000378:metabolism*; D012462:Saline Solution, Hypertonic / Q000494:pharmacology*; D013997:Time Factors
chemical_list: D011993:Recombinant Fusion Proteins; D012462:Saline Solution, Hypertonic; D001127:Arginine Vasopressin; D049452:Green Fluorescent Proteins