In Vivo Chemical Monitoring at High Spatiotemporal Resolution Using Microfabricated Sampling Probes and Droplet-Based Microfluidics Coupled to Mass Spectrometry

Abstract

An essential approach for in vivo chemical monitoring is to use sampling probes coupled with analytical methods; however, this method traditionally has limited spatial and temporal resolution. To address this problem, we developed an analytical system that combines microfabricated push-pull sampling probes with droplet-based microfluidics. The microfabricated probe provides spatial resolution approximately 1000-fold better than that of common microdialysis probes. Microfabrication also facilitated integration of an extra channel into the probe for microinjection. We created microfluidic devices and interfaces that allowed manipulation of nanoliter droplet samples collected from the microfabricated probe at intervals of a few seconds. Use of droplet-based microfluidics prevented broadening of collected zones, yielding 6 s temporal resolution at 100 nL/min perfusion rates. Resulting droplets were analyzed by direct infusion nanoelectrospray ionization (nESI) mass spectrometry for simultaneous determination of glutamine, glutamate, γ-aminobutyric acid, and acetylcholine. Use of low infusion rates that enabled nESI (50 nL/min) was critical to allowing detection in the complex samples. Addition of C-labeled internal standards to the droplet samples was used for improved quantification. Utility of the overall system was demonstrated by monitoring dynamic chemical changes evoked by microinjection of high potassium concentrations into the brain of live rats. The results showed stimulated neurochemical release with rise times of 15 s. This work demonstrates the potential of coupling microfabricated sampling probes to droplet-based mass spectrometric assays for studying chemical dynamics in a complex microenvironment at high spatiotemporal resolution.

Novel microwire-based biosensor probe for simultaneous real-time measurement of glutamate and GABA dynamics in vitro and in vivo

“…One of the most common methods for the acquisition of longitudinal measurements is microdialysis, but analyte measurement is limited to large cell populations. In addition, its low sampling rate, on the order of minutes, prevents its use in measuring faster, real-time transients 2 , 3 , 31 – 35 , introducing a low-pass filtering effect into recorded data 36 . Precision, micromachined dialysis probes have increased the sampling rate to several seconds, but this resolution is still slow relative to the expected dynamics of GLU and GABA in the extracellular space.…”

“…Precision, micromachined dialysis probes have increased the sampling rate to several seconds, but this resolution is still slow relative to the expected dynamics of GLU and GABA in the extracellular space. Furthermore, microdialysis appears to underreport neurotransmitter concentrations, possibly because artificial cerebral fluid is perfused into the dialysis cannula which dilutes the neurotransmitters 35,37 . Alternatively, enzyme-based biosensors based on microelectrode arrays (MEAs) may be able to improve spatial and temporal resolution, as well as biocompatibility 10,32 .…”

Recent Advances in In Vivo Neurochemical Monitoring

“…Microdialysis typically leads to a total delay time of approximately 10–20 min [ 217 ]. Recent advances in system set-ups, capillary-scale HPLC columns, microfluidic systems, nL-level precision sample loop valves, and improvements in optimization have brought temporal resolution down to the sub-minute range [ 218 , 219 , 220 , 221 , 222 , 223 ] Therefore, microdialysis is a good choice for neuronal processes that last from minutes to hours, but not as good as electrochemical or optical sensors in capturing the fast dynamics of neurochemistry. Another issue regarding the use of microdialysis is accuracy of measurement.…”

“…Significant reductions in electrically-evoked DA releases were recorded by a CFE attached to a microdialysis probe as soon as 2 h after probe implantation [ 227 ]. Recent advances in anti-inflammatory drug delivery through the perfusate [ 228 , 229 , 230 ] and ultrasmall microfabricated nano-probe development [ 219 ] have improved the tissue/probe interface, resulting in longer-lasting in vivo measurements. By perfusing the anti-inflammatory corticosteroid dexamethasone through the microdialysis probe, glucose levels in TBI model rats have been continuously monitored for upwards of 10 days without glial barrier formation [ 231 ].…”

Dexamethasone-Enhanced Microdialysis and Penetration Injury

“…Thus, over time, various workers have improved the temporal resolution of microdialysis by developing refined analytical methods with lower detection limits. For example, the Kennedy group has developed numerous rapid dialysate analyses by on-line capillary liquid chromatography or on-line capillary electrophoresis ( Watson et al, 2006 ; Li et al, 2009 ; Perry et al, 2009 ; Wang et al, 2010 ; Lee et al, 2013 ; Kennedy et al, 2018 ; Ngernsutivorakul et al, 2018 ). The Andrews and Weber groups have used capillary liquid chromatography to analyze dopamine and serotonin in brain dialysates with minute and sub-minute temporal resolution ( Liu et al, 2010 ; Yang et al, 2013 ; Zhang et al, 2013 ; Gu et al, 2015 ; Wilson and Michael, 2017 ).…”