Culturing thick brain slices: an interstitial 3D microperfusion system for enhanced viability

Area of Science:

• Neuroscience
• Biomedical Engineering
• Tissue Engineering

Background:

• Organotypic brain slice cultures are valuable neuroscience research models.
• Existing methods struggle to support thick brain slices (>150 microm) due to nutrient and oxygen diffusion limitations, leading to necrosis.
• Previous perfusion techniques offer limited viability for acute, thinner slices.

Purpose of the Study:

• To develop and validate a novel interstitial microfluidic perfusion technique for culturing thick (700 microm) organotypic brain slices.
• To assess the viability and cyto-architecture preservation of these thick slices over time in vitro.
• To determine optimal flow rates for enhanced slice survival and function.

Main Methods:

• Development of a custom-made microperfusion chamber enabling interstitial perfusion.
• Culturing of 700 microm thick organotypic brain slices using the developed perfusion system.
• Assessment of slice viability and cyto-architecture at 2 and 5 days in vitro (DIV) compared to unperfused controls.

Main Results:

• The interstitial microfluidic perfusion technique successfully cultured thick (700 microm) organotypic brain slices.
• Viability reached up to 84.6% (p<0.01) after 5 DIV, significantly higher than controls.
• Cultures maintained in vivo cyto-architecture and functional activity.

Conclusions:

• This novel perfusion method overcomes limitations of existing techniques for thick brain slice cultures.
• It enables prolonged viability and functional integrity of thick organotypic brain slices.
• This advancement supports reliable investigation of intact neuronal network properties in vitro.