Development of Patch-Clamp Chips for Mammalian Cell Applications

Dolores      Martinez; Marzia      Martina; Laura      Kremer; Robert      Monette; Tanya      Comas; Danish      Salim; Christophe      Py; Mike      W. Denhoff; Geoff      Mealing

doi:10.2174/1876402911002040274

Abstract

We have previously described the designs of two planar patch-clamp neurochips and their application to the electrophysiological study of molluscan neurons cultured on-chip. Neuron attachment and growth over apertures on the neurochip surface permitted the acquisition of whole-cell patch-clamp recordings. To broaden the application of these neurochips from molluscan to mammalian neurons, we conducted a study of cell-to-aperture interaction to optimize conditions for these smaller, more fragile cells. For this purpose, we designed a “sieve” chip having multiple apertures on its surface. Random growth of rat cortical neurons resulted in a 32% (n = 324) probability of cell growth over 2 μm diameter apertures; larger diameters resulted in growth through the aperture. Based on these findings, single-aperture neurochips were fabricated having 2 μm diameter aperture and preliminary electrophysiological recordings from cortical cultures at 14 DIV are presented. The implications of this study for the next-generation neurochips are discussed.

Keywords: Mammalian cells, planar patch-clamp, neurochip, whole-cell recordings, apertures, poly(dimethylsiloxane), PDMS, Lymnaea neurons, dissecting microscope, Sieve Chip, polyimide, Plexiglas, Minimal Essential Medium, L-glutamine, Normal Bath Media, Calcein-AM, RH-237, LSM-410 Zeiss, confocal microscope, krypton/argon, North-ern Eclipse Software, Electrophysiology, DIV, scanning electron microscopy, SEM, focused, FIB