title: Agarose-Based Substrate Modification Technique for Chemical and Physical Guiding of Neurons In Vitro.
abstract: A new low cost and highly reproducible technique is presented that provides patterned cell culture substrates. These allow for selective positioning of cells and a chemically and mechanically directed guiding of their extensions. The patterned substrates consist of structured agarose hydrogels molded from reusable silicon micro templates. These templates consist of pins arranged equidistantly in squares, connected by bars, which mold corresponding wells and channels in the nonadhesive agarose hydrogel. Subsequent slice production with a standard vibratome, comprising the described template pattern, completes substrate production. Invertebrate neurons of locusts and pond snails are used for this application as they offer the advantage over vertebrate cells as being very large and suitable for cultivation in low cell density. Their neurons adhere to and grow only on the adhesive areas not covered by the agarose. Agarose slices of 50 μm thickness placed on glass, polystyrene, or MEA surfaces position and immobilize the neurons in the wells, and the channels guide their neurite outgrowth toward neighboring wells. In addition to the application with invertebrate neurons, the technique may also provide the potential for the application of a wide range of cell types. Long-term objective is the achievement of isolated low-density neuronal networks on MEAs or different culture substrates for various network analysis applications.
authors.affiliation: Institute for Biology II, RWTH Aachen University , Worringerweg 3, 52074 Aachen, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Krumpholz
authors.affiliation: Institute for Biology II, RWTH Aachen University , Worringerweg 3, 52074 Aachen, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Rogal
authors.affiliation: Institute of Materials in Electrical Engineering 1, RWTH Aachen University , Sommerfeldstraße 24, 52074, Aachen, Germany.
authors.forename: Akram
authors.initials: A
authors.lastname: El Hasni
authors.affiliation: Institute of Materials in Electrical Engineering 1, RWTH Aachen University , Sommerfeldstraße 24, 52074, Aachen, Germany.
authors.forename: Uwe
authors.initials: U
authors.lastname: Schnakenberg
authors.affiliation: Institute for Biology II, RWTH Aachen University , Worringerweg 3, 52074 Aachen, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Bräunig
authors.affiliation: Institute for Biology II, RWTH Aachen University , Worringerweg 3, 52074 Aachen, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Bui-Göbbels
mesh_terms: D000818:Animals; D002470:Cell Survival; D002478:Cells, Cultured; D005260:Female; D020100:Hydrogels / Q000737:chemistry*; D007150:Immunohistochemistry; D048188:Locusta migratoria; D008297:Male; D009474:Neurons / Q000166:cytology* / Q000378:metabolism; D012685:Sepharose / Q000737:chemistry*; D012825:Silicon / Q000737:chemistry
chemical_list: D020100:Hydrogels; D012685:Sepharose; D012825:Silicon
keywords: defined network; hydrogel; invertebrates; nonadhesive; patterning; vibratome