Simultaneous analysis of vascular norepinephrine and ATP release using an integrated microfluidic system

- A method for the simultaneous detection of norepinephrine and ATP was devised.
- Microfluidics and amperometry were employed to detect norepinephrine, and online chemiluminescence to detect ATP.
- 3-D printing technology was utilized to interface key components.
- The method was confirmed using the model of the isolated perfused mesenteric arterial bed of the rat as a model of the sympathetic neuroeffector junction.

BackgroundSympathetic 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 methodIn 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).ResultsStimulated levels relative to basal levels of norepinephrine and ATP were found to be 363 nM and 125 nM, respectively (n = 6). The limit of detection for norepinephrine is 80 nM using microchip-based amperometric detection. The LOD for on-line ATP detection using chemiluminescence is 35 nM.Comparison with existing methodIn 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.ConclusionsIn conclusion, the method described in the paper can be used to successfully detect norepinephrine and ATP simultaneously and in a near-real-time fashion.