Measuring the release of ATP from the mouse carotid body

Adenosine Triphosphate (ATP) is considered to be an essential excitatory neurotransmitter in the cat and rat carotid body (CB). However, the role of ATP in the mouse CB has yet to be determined. Therefore, we have developed a system to measure in vitro release of ATP from the mouse CB using bioluminescence techniques. The initial focus was to determine an optimal in vitro environment. We tried various chamber systems and methods of integrating the CB within these systems. These attempts have elevated our understanding of the sensitivity of the measurement, the sample collection procedures, and the quantification of sample concentration. We have found that the methods that have been previously used to measure the ATP release from the cat and rat CB cannot be directly applied to the mouse CB due to the small tissue volume. Further, due to the small amount of the ATP release, possible bacterial contamination has impeded upon accurate measurements. Thus, experiments were performed aseptically. The incubation chamber was made of Spin‐X® Centrifuge Tube Filter. The filter was coated with Martigel to hold four CBs. Krebs (120 μL) was added and CBs were incubated for 7 min at 37 °C. By using trichloroacetic acid, we estimated the maximum release of ATP from the CB of DBA/2J mice was 289 pmoles/min. In hyperoxic/normocapnic conditions, the release was 7.88 pmoles/min. Supported by AHA 09GRNT2080158, NHLBI HL81345