Brain Slice Stimulation Using a Microfluidic Network and Standard Perfusion Chamber

Summary

We demonstrate fabrication of a simple microfluidic device that can be integrated with standard electrophysiology setups to expose microscale surfaces of a brain slice in a well controlled manner to different neurotransmitters.

Abstract

We have demonstrated the fabrication of a two-level microfluidic device that can be easily integrated with existing electrophysiology setups. The two-level microfluidic device is fabricated using a two-step standard negative resist lithography process ¹. The first level contains microchannels with inlet and outlet ports at each end. The second level contains microscale circular holes located midway of the channel length and centered along with channel width. Passive pumping method is used to pump fluids from the inlet port to the outlet port ². The microfluidic device is integrated with off-the-shelf perfusion chambers and allows seamless integration with the electrophysiology setup. The fluids introduced at the inlet ports flow through the microchannels towards the outlet ports and also escape through the circular openings located on top of the microchannels into the bath of the perfusion. Thus the bottom surface of the brain slice placed in the perfusion chamber bath and above the microfluidic device can be exposed with different neurotransmitters. The microscale thickness of the microfluidic device and the transparent nature of the materials [glass coverslip and PDMS (polydimethylsiloxane)] used to make the microfluidic device allow microscopy of the brain slice. The microfluidic device allows modulation (both spatial and temporal) of the chemical stimuli introduced to the brain slice microenvironments.

Protocol

SU-8 mold fabrication

Master preparation

1. The SU-8 master on silicon wafer substrate is prepared using a two-step standard negative resist lithography process.
2. The alignment marks on the silicon wafer are removed using a razor blade as the height of these structures (located along the outer periphery of the wafer) is more than the actual device structures.
3. The silicon wafer is then cleaned using isopropyl alcohol and dried in a stream of N₂. Support pillars made out of tape with thickness less than the tallest device structure replace the alignment marks on four sides of the wafer.
   
   Not....

Discussion

Existing macroscale or microscale brain slice perfusion chambers are limited in terms of the spatial resolution they provide to expose brain slices with neurotransmitters. The microfluidic device technology demonstrated here overcomes this limitation using simple bioMEMS techniques. It is anticipated that the simplicity in the fabrication of the microfluidic device and the ease in integrating it with existing electrophysiology setups will allow widespread application of the demonstrated device technology. Interesting exp.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
RC-26GPL	Tool	Warner Instruments	W2-64-0236	Low Profile Large Bath RC-26GLP Recording Chamber
SHD-26GH/10	Tool	Warner Instruments	W2-64-0253	Stainless steel slice hold-down for RC-26G, 1.0 mm thread spacing
PDMS (polydimethylsiloxane)	Reagent	Dow Corning	Sylgard 184	Silicone Elastomer Kit
Plasma Preen-II 862	Tool	Plasmatic Systems, Inc.		Microwave plasma system
Model P-1	Tool	Warner Instruments	W2-64-0277	Series 20 Plain Platform, Model P-1
SA-NIK	Tool	Warner Instruments	W2-64-0291	Adapter for Nikon Diaphot/TE200/TE2000, SA-NIK
Oxygenated, heated ACSF (Artificial cerebro-spinal fluid)	Reagent			Exact composition will vary with application