Abstract An autoclavable chamber and associated medium circulation system has been constructed to provide a stable environment for mammalian cultures for long periods of time. The chamber was specifically designed for (a) multichannel electrophysiological recording from monolayer networks with photoetched multielectrode matrices, (b) for microscope observations of networks in the living state and (c) for the manipulation of such networks with laser cell surgery. The chamber components can be assembled under sterile conditions in less than 30 s to minimize pH and osmolarity stress to the monolayer cultures. An open chamber version provides a constant medium bath for pharmacological studies. The closed chamber version, attached to a 10-ml medium reservoir and a peristaltic pump, has so far provided constant conditions for continual recording over an 8-day period. Flow characteristics within the closed chamber, optical properties, pH maintenance, and schemes for drug addition are described.