Microcutting of living brain slices by a pulsed ultrafine water jet which allows simultaneous electrophysiological recordings (micromingotome).

Abstract

Up to now microsurgical dissections in living nervous tissue (e.g. in slices or cell cultures) are performed either by micro-scalpels or by laser beams. As an alternative technique, a device for cutting with an ultrafine pulsed water jet was developed to allow precise, visually controled dissections in neuronal circuits even during electrophysiological recordings. Water is ejected by pressure (20-30 bar) from patch pipettes with tip diameters of 10-12 microm. By means of a piezo-element the pipette and the water jet are forced to oscillate vertically with a frequency of 200-400 Hz with an adjustable amplitude. These oscillations facilitate the transsection of neuronal connections even in thick slice preparations. Best results were obtained when the tip of the pipette was about 500 microm above the surface of the submerged slice tissue. This micromingotome offers the following advantages: (i) histological studies show that the water jet cleans the cutting surface, thus avoiding debris and its uncontrolable effects on cells underneath; (ii) the arrangement enables ongoing electrophysiological recordings from hippocampal slices during the cutting procedure and thus facilitates studies of the functions of neuronal connections; (iii) the device allows even disconnection in cultured nervous tissue overgrowing polyamid grids with 50 microm wide meshes.