Sampling Probes Microfabricated from Parylene‑C for In Vivo Neurochemical Monitoring

Monitoring chemistry
in the central nervous system in
vivo is an
important route to better understand chemical signals and metabolism
underlying brain function. Sampling from the brain extracellular space
allows for in depth and multiplexed analysis of this compartment.
Miniaturized sampling devices prepared by using microfabrication are
emerging as tools that allow high spatial resolution measurements
with low invasiveness. In this work, we describe a microfabrication
method for Parylene-C-based push–pull sampling probes. Parylene-C
is an attractive material because its biocompatibility and flexibility
may allow for chronic measurements. Probes with 3 mm long shanks and
45 × 166 μm cross sections were prepared and tested for
suitability for in vivo experiments. Two designs, one with push–pull
orifices on the probe tip and the other with the orifices recessed
∼40 mm within a sheath of Parylene-C, were investigated. The
sheathed probes successfully flowed in tofu (n =
5) and ex vivo brain tissue (n = 5) for at least
3 h at 100 nL/min. In contrast, only 60% (n = 5)
of the end-on probes flowed successfully in tofu. In vitro recoveries
for the sheathed and design were 66 ± 2 and 91 ± 7%, respectively,
in stirred solution. The sheathed probes were tested by sampling from
the cortex of anesthetized rats. In 13 of 19 experiments, push–pull
fractions were successfully collected. Of the six failures, all had
a sheath shallower than the 30 μm design depth, suggesting that
this depth is necessary for reliable sampling flow. Basal concentrations
of 20 neurotransmitters and metabolites determined by liquid chromatography
with tandem mass spectrometry (LC-MS/MS) were comparable with concentrations
recorded using microdialysis sampling in prior work. Infusion of 2.0
mM nipecotic acid through the probe resulted in a significant increase
in GABA with little effect on other compounds. Untargeted metabolomics
analysis by LC-MS/MS identified 141 metabolites within the push–pull
perfusates. The results demonstrate the feasibility of using Parylene-C
push–pull probes with a sheathed tip design for multiplexed
and temporally resolved monitoring of brain chemistry in vivo.