Opioid peptides are critically involved in a variety of physiological functions necessary for adaptation and survival, and as such they show tremendous promise as therapeutic targets. However there is a critical gap in understanding when and where these molecules are released, because there is a paucity of detection methods for monitoring opioid peptides in the extracellular space. We have designed a novel waveform that employs two distinct scan rates in each voltammetric sweep to detect enkephalin dynamics in live tissue with sub-second temporal resolution using carbon fiber microelectrodes. Combining two scan rates in a single voltammetric scan is unprecedented in molecular monitoring. It exploits fundamental principles to offer a combination of temporal and spatial resolution, sensitivity, and chemical selectivity that has the potential to advance the voltammetric detection of many classes of molecules. Electrochemical detection is further enhanced by combining this approach with a microelectrode sensing substrate engineered entirely of multi-walled carbon nanotubes spun into a yarn, replacing the standard carbon fiber altogether. The high aspect ratio and distinct electronic properties of the nanotubes result in markedly improved selectivity, sensitivity, and spatial resolution, as well as faster electron transfer kinetics that are manifested as sharper voltammetric peaks when compared to conventional carbon-fiber microelectrodes. By combining a groundbreaking voltammetric approach with a novel tool engineered from nanoscale materials, we are enabling a shift from interferential measures of endogenous opioid activity to direct molecular measurements.