Patch-clamping of primary cardiac cells with micro-openings in polyimide films.
Patch-clamping is a powerful method for investigating the function and regulation of ionic channels. Currently, great efforts are being made to automate this method. As a step towards this goal, the feasibility of patch-clamping primary cells with a microscopic opening in a planar substrate was tested. Using standard microfabrication and ion beam technology, small-diameter openings (2 and 4 microm) were formed in polyimide films (thickness 6.5 microm). Single cells (sheep Purkinje heart cells, Chinese hamster ovary cells) in a suspension were positioned on top of the opening and sucked towards the opening to improve adhesion of the cell to the planar substrate, hence increasing the seal resistance. Voltage/current measurements yielded a median seal resistance of 1.3 Mohms with 4 microm openings (n=24) and 26.0 Mohms with 2 microm openings (n = 75), respectively. With 2 microm openings, successful loose-patch recordings of TTX-sensitive inward currents and action potentials in sheep Purkinje heart cells (n = 18) were made. In rare cases, gigaseals (n = 4) were also measured, and a whole-cell configuration (n = 1) could be established. It was concluded that the simple planar patch approach is suitable for automated loose-patch recordings from cells in suspension but will hardly be suitable for high-throughput whole-cell patch-clamping with high-resistance seals.
Stett
A
A
NMI Natural & Medical Science Institute, Reutlingen, Germany. stett@nmi.de
Bucher
V
V
Burkhardt
C
C
Weber
U
U
Nisch
W
W
D000818:Animals; D016466:CHO Cells / Q000502:physiology; D006224:Cricetinae; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005240:Feasibility Studies; D007473:Ion Channels / Q000502:physiology; D018408:Patch-Clamp Techniques / Q000379:methods*; D011689:Purkinje Cells / Q000502:physiology*; D012117:Resins, Synthetic; D012756:Sheep
D007473:Ion Channels; D012117:Resins, Synthetic; C029945:polyimide resin