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Development and Characterization of In Vitro Microvessel Network and Quantitative Measurements of Endothelial [Ca2+]i and Nitric Oxide Production
Published on: May 19, 2016
Simultaneous analysis of vascular norepinephrine and ATP release using an integrated microfluidic system
Alexandra D Townsend1, Gerald H Wilken2, Kyle K Mitchell3
1Department of Chemistry, Saint Louis University, St. Louis, MO 63103, United States.
Background:
Sympathetic nerves are known to release three neurotransmitters: norepinephrine, ATP, and neuropeptide Y that play a role in controlling vascular tone. This paper focuses on the co-release of norepinephrine and ATP from the mesenteric arterial sympathetic nerves of the rat.
New Method:
In this paper, a quantification technique is described that allows simultaneous detection of norepinephrine and ATP in a near-real-time fashion from the isolated perfused mesenteric arterial bed of the rat. Simultaneous detection is enabled with 3-D printing technology, which is shown to help integrate the perfusate with different detection methods (norepinephrine by microchip-based amperometery and ATP by on-line chemiluminescence).
Results:
Stimulated levels relative to basal levels of norepinephrine and ATP were found to be 363nM and 125nM, respectively (n=6). The limit of detection for norepinephrine is 80nM using microchip-based amperometric detection. The LOD for on-line ATP detection using chemiluminescence is 35nM.
Comparison With Existing Method:
In previous studies, the co-transmitters have been separated and detected with HPLC techniques. With HPLC, the samples from biological preparations have to be derivatized for ATP detection and require collection time before analysis. Thus real-time measurements are not made and the delay in analysis by HPLC can cause degradation.
Conclusions:
In conclusion, the method described in the paper can be used to successfully detect norepinephrine and ATP simultaneously and in a near-real-time fashion.
