Modern science is characterized by integration and synergy between research fields. Accordingly, as technological advances allow new and more ambitious quests in scientific inquiry, numerous analytical and engineering techniques have become useful tools in biological research. The focus of this review is on cutting edge technologies that aid direct measurement of bioactive compounds in the nervous system to facilitate fundamental research, diagnostics, and drug discovery. We discuss challenges associated with measurement of cell-to-cell signaling molecules in the nervous system, and advocate for a decrease of sample volumes to the nanoliter volume regimen for improved analysis outcomes. We highlight effective approaches for the collection, separation, and detection of such small-volume samples, present strategies for targeted and discovery-oriented research, and describe the required technology advances that will empower future translational science.