Abstract
Adenosine is a complex signaling molecule with a wide array of functions including the regulation of sleep, breathing, and neuromodulation. Recently adenosine was recognized as a fast acting modulator that can signal on a rapid time scale. Monitoring adenosine on a sub-second time scale with fast-scan cyclic voltammetry (FSCV) allowed for many advancements in understanding adenosine function on a rapid time scale. Our laboratory pioneered the field of adenosine measurements in vivo and in brain slices with FSCV. Using FSCV, I discovered and characterized spontaneous, transient adenosine release in vivo in the caudate-putamen and prefrontal cortex of anesthetized rats. The concentration, duration, and frequency of spontaneous adenosine release is characterized in both brain regions. The prefrontal cortex has a higher frequency of release than the striatum, while the concentration and duration remain constant in both regions.
Next, I describe the clearance of spontaneous, transient adenosine in the caudate-putamen of the in vivo rat model as well as in brain slices. Naturally occurring adenosine release is cleared from the extracellular space in 3-4 seconds, suggesting a rapid form of signaling. I also examine uptake through transporters and metabolism. The equilibrative nucleoside transporter, ENT1, and the enzymes responsible for metabolizing adenosine, adenosine deaminase and adenosine kinase, all play a partial role in clearing adenosine. The rapid regulation in the extracellular space suggests there is a mode of adenosine signaling that is fast and takes place on the second time scale. Next I examine how transient adenosine is regulated. I found that the concentration of spontaneous, transient adenosine is independent of all of the examined receptors: glutamate (ionotropic and metabotropic), GABA, and adenosine. However, the frequency of adenosine release is modulated by the glutamate receptor NMDA and the adenosine receptors A1 and A2a. The results demonstrate there is a feedback loop between released adenosine and adenosine receptors, as well as through the NMDA receptor.
Finally, I describe a new phenomenon that I accidentally discovered in the laboratory. While inserting our carbon-fiber microelectrodes, significant release of adenosine was observed during implantation. Mechanically stimulated adenosine release is characterized in the prefrontal cortex. The release is examined in anesthetized rats and brain slices and the average release lasts around 42 seconds. Using cell staining and multiple movements of our electrodes the results demonstrate that the release of adenosine is not from cell death or damage. This new mode of signaling further suggests that adenosine is neuroprotective and might be involved in protection during mechanical perturbation in the brain. The discovery of two new modes of rapid signaling demonstrate that adenosine modulates neurotransmission in the brain and has the ability to be a therapeutic agent during brain injury.
