Unveiling Dopamine and Met-Enkephalin Dynamics: Simultaneous Co-Detection in Rat Striatum

Endogenous opioid peptides have been linked to numerous
physiological
functions, including pain perception and the motivational drive associated
with substance use disorders, but many fundamental aspects of transmission
remain ambiguous. The kinetics of endogenous opioid peptides are thought
to be slower and to last longer than those of more classical small-molecule
neurotransmitters, like dopamine; however, a direct comparison of
the release and diffusive spread of these molecules in the brain is
lacking. Here, fast-scan cyclic voltammetry was coupled with carbon
microelectrodes for co-detection of dopamine and met-enkephalin at
single recording sites in rat striatal slices. The measurements used
a voltammetric waveform that was specifically designed to minimize
sensitivity to dopamine, maximize sensitivity to enkephalin, and minimize
biofouling. Both neurotransmitter (dopamine) and neuropeptide (met-enkephalin,
M-ENK) release scaled with stimulation duration. Interestingly, ENK
dynamics in striatum displayed a unique biphasic profile with a significant
latency to peak that occurred ∼30 s after stimulation, suggesting
a sphere of influence that was ∼3x larger than that of dopamine.
Mathematical modeling of the evoked M-ENK concentration profile suggests
that multiple forms of ENK were released at once, such that some of
the five-amino-acid form of M-ENK was released in exocytosis, and
some was generated in the extracellular space by enzymatic cleavage
of a larger form of ENK. Finally, a series of experiments combined
solid-phase extraction with liquid-chromatography mass spectrometry
to independently verify ENK release. The findings provide direct evidence
to support widely held assumptions regarding neuropeptide release,
and they demonstrate how different classes of signaling molecules
can potentially affect distinct cellular populations in striatumeven
when released at the same site.