Using Microfluidic Devices to Investigate Peptide Release from Neuronal Cells with Mass Spectrometry

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Using Microfluidic Devices to Investigate Peptide Release from Neuronal Cells with Mass Spectrometry
Ming Zhong1, Kyubong Jo2, and Jonathan V. Sweedler1
1 Department of Chemistry and Beckman Institute, University of Illinois at Urbana-Champaign, Urbana, IL 2 Sogang University, Seoul, South Korea

Different regions of neurons within the brain experience distinct chemical environments and the overall cellular response depends on these chemical microenvironments. Providing precise chemical control of small-volume regions around neuronal cells remains a challenge. By placing neurons in small chambers in a microfluidic device, the chemical environment can be controlled, and materials released by the neuron in response to chemical stimulations can be efficiently collected for off-line characterization. In this device, the cellular releasates are collected as they flow along a serpentine channel that contains a surface derivatized with octadecyltrichlorosilane designed to collect the peptides released from chemically-simulated neurons. Next the released peptides are characterized by matrix-assisted laser desorption/ionization (MALDI)-based mass spectrometry (MS). We validated this approach using Aplysia californica bag cell neurons stimulated by elevated potassium chloride. After the stimulation, several neuropeptides, such as egg laying hormone (ELH) and acidic peptide (AP) were detected. Distinct peptides were deposited at different locations of the channel due to differential partitioning of the peptides onto the surface. This approach demonstrates the capability of microfluidic devices to manipulate mass-limited samples and interface these samples with the high sensitivity and information content of MALDI MS. Besides identifying the peptides, characterizing the amount of neuropeptides released is also important, especially when characterizing neuron dynamics. We describe an enhancement to the microfluidic device described above that allows us to quantify peptide release. This is done by measuring the length of channel covered by each peptide. A larger amount of a peptide covers a longer section of the channel as the ability of the surface to collect / bind a peptide is constant. This method was validated using samples containing known amounts of a standard neuropeptide--substance P; the results show that the length of channel occupied by substance P increases in proportion to the amount of peptide present. Our microfluidic device can provide quantitative information on peptide release from neurons in various states and environments.