Discipline: Biological Sciences
Subcategory: Physiology and Health
Kierra A. Goins - Auburn University
Co-Author(s): Jeff Reese, Elaine L. Shelton, and Naoko Brown, Dept. of Pediatrics, Vanderbilt University Med Ctr, Nashville, TN Michael T. Yarboro, Dept. of Cell and Developmental Biology, Vanderbilt University Med Ctr, Nashville, TN
Background: PDA is a common cardiac condition in premature neonates and occurs when the ductus arteriosus (DA), a muscular arterial shunt, does not close after birth as it should. Clinical studies show that less incidence of PDA is associated with caffeine treatment for apnea of prematurity, a condition in which premature neonates stop breathing. Caffeine works by binding to adenosine receptors and inhibiting adenosine activity, suggesting a potential mechanism for reduction in PDA. Since current treatments for PDA have adverse side effects, it is crucial that better treatment for PDA is found.
Objective: This study will define caffeine’s role in ductal closure. I will test the hypothesis that caffeine has a direct constrictive effect on the DA.
Design/Methods: CD-1 mice were delivered via C-section on day 19 (full term) of gestation. The DA was isolated and mounted in a chamber filled with Krebs buffer bubbled with 2% oxygen to simulate fetal conditions (pO2 <40). Pressurized myography illustrated the diameter of the DA in response to increasing concentrations of caffeine citrate (10-7M-10-2M) or adenosine (10-9M-10-3M), both dissolved in aqueous solution. I also performed reverse transcription-polymerase chain reaction (RT-PCR) and quantitative PCR (qPCR) to evaluate adenosine receptor expression in the DA. Results: RT-PCR demonstrated expression of all four adenosine receptors (A1, A2a, A2b, and A3) in day 19 DA and control tissue (fetal brain). qPCR demonstrated that adenosine receptor expression is upregulated in the day 19 DA relative to other stages of gestation. Dose response experiments showed that neither adenosine nor caffeine significantly affected DA diameter until the highest concentrations caused dilation. Pretreatment studies showed that pretreating vessels with caffeine decreased adenosine-induced dilation. Conclusions: Caffeine did not directly affect ductal tone. Although caffeine did not directly constrict the DA, caffeine may work to reduce PDA by binding to adenosine receptors and blocking adenosine’s dilating effects. Future studies will investigate whether caffeine mediates Ca2+ ion channels to induce vasoconstriction.
Funder Acknowledgement(s): This research was sponsored by the NIH Short Term Training for Minority Students: R25 HL096223-06 grant
Faculty Advisor: Jeff Reese, firstname.lastname@example.org
Role: Dr. Reese assigned me my project as an individual summer project. We had frequent meetings in which we discussed the best plan of action for me for the following week. On a typical day, I would run the 5-7.5 hour experiments. This included setting up the experiment, warming up the ductus arteriosus (DA) to 37 degrees Celsius for 40 minutes, bringing the DA to 20 mmHg pressure, exposing the DA to KCl, washing the DA with Krebs solution, and exposing the DA to drugs of interest (caffeine citrate or adenosine). I also recorded the results and used GraphPad Prism to convert data into statistically significant graphs. I performed reverse-transcription polymerase chain reaction, and I also dissected fetal mice, isolated their DA, and mounted the DA in the pressurized myography chambers.