Neurochemical monitoring by silicon nanodialysis probe coupled with mass spectrometry detection

The brain is an extraordinarily complex and heterogeneous organ, with its functions varying dynamically across time, space, and chemistry. Understanding how the brain responds to stimuli and undergoes chemical changes during behavior is crucial for unraveling the underlying mechanisms of neurological processes and disorders. However, obtaining this information with high spatial and temporal resolutions remains a significant challenge. This dissertation presents a silicon microfluidic platform coupled with mass spectrometry (MS) for in vivo neurochemical monitoring. The silicon platforms contain several independently developed modules, including droplet generator, nanoelectrospray ionization (nESI) emitter, and nanodialysis sampling probe. These components are integrated on demand, characterized individually, and optimized for their coupling with MS detection workflow. Chapter 2 focuses on the integration of a picoliter-scale droplet generator with an on-chip nESI emitter for quantitative analysis of neurochemicals. A miniaturized T-junction microfluidic channel stably segments analytes into picoliter compartments and delivers them to the integrated nESI emitter tip for MS detection. Several parameters were optimized as crucial for the stable generation of droplet flow and droplet peak integrity. The developed system enabled effective, multiplexed detection of neurochemicals encapsulated within droplets down to low picoliter volumes. Quantitative measurements for each neurochemical demonstrate limits of detection at the 1 – 10 amol range. Next, a silicon nanodialysis probe was presented for localized sampling of extracellular fluid in vivo. The probe’s membrane-free design and in-plane, ultra-slow flow configuration minimize tissue disruption and maintain high analyte recovery. Histological analysis confirmed minimal tissue damage and a confined depletion depth of approximately 200 μm. Coupling this sampling strategy with capillary electrophoresis-mass spectrometry (CE-MS) allows for the quantification of multiple neurotransmitters and metabolites in microliter-scale dialysate samples with in vivo recovery rates between 60% and 93%. Application of the probe in the primary somatosensory cortex of the mouse revealed spatial gradients in neurochemical composition between neighboring subregions, highlighting the platform’s ability to resolve fine-scale heterogeneity in the extracellular chemical landscape. The membrane-free design of the nanodialysis probe also overcomes many of the limitations associated with conventional microdialysis for extracellular neuropeptide sampling, including low recovery, membrane adsorption, and contamination from intracellular components. Therefore, the nanodialysis probe was applied to study the extracellular neuropeptidome in the mouse somatosensory cortex (S1) with spatial resolution as small as 100 μm. Coupling with nanoLC-MS/MS, a number of extracellular peptides from various precursors, including secretogranin-1, ProSAAS, and pro-opiomelanocortin (POMC), were identified. To enhance detection sensitivity for prohormone-derived peptides, a discovery-to-targeted peptidomic approach was developed, further identifying 46 peptides from 24 proteins in dialysate samples. Among all the detected proteins, 11 exhibit low local expression, suggesting these peptides may be transported to S1 from distant sites. These findings unveil a complex extracellular peptide landscape in S1, including local and long-distance signaling. Altogether, this dissertation presents a modular and versatile platform for neurochemical monitoring that combines localized sampling with ultrasensitive MS detection. It enables spatially resolved, multiplexed, and minimally invasive analysis of the brain’s extracellular chemical environment and holds promise for further integration of additional modules for broader applications in neuroscience research, from basic neurophysiology to the study of neurological disease.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Neuroscience

Grantor

University of Illinois Urbana-Champaign

Year dc:date

2025

Author dc:creator

• Li, Keyin

Contributors dc:contributor

• Sweedler, Jonathan V.
• Sweedler, Jonathan V
• Vlasov, Yurii A
• Gillette, Martha L
• Christian-Hinman, Catherine

• Neurochemicals
• In Vivo Sampling
• Mass Spectrometry

Statement dc:rights

• © 2025 Keyin Li

Language dc:language

en, eng

Handle dc:identifier

https://hdl.handle.net/2142/130177

OAI record oai:identifier

https://www.ideals.illinois.edu/oai-pmh?verb=GetRecord&metadataPrefix=etdms&identifier=oai%3Awww.ideals.illinois.edu%3A2142%2F130177