A novel two-compartment culture dish allows microscopic evaluation of two different treatments in one cell culture simultaneously

Abstract

A new type of culture dish containing two separate compartments is described, that can be used in high-magnification microscopy. Using the dish, two halves of a single-cell culture, grown on a standard coverslip, can be exposed to different treatments simultaneously, allowing the effect of one treatment to be compared with that of the other treatment in the same culture. This way, the natural variability that might exist between different individual cultures is circumvented. In addition, by simultaneously conducting two experiments per dish, the number of experiments needed can be decreased. This both reduces the time to complete a series of experiments and allows the optimal use of specimens that are difficult to obtain, such as human material. We found there is an excellent barrier between the two compartments for lipophilic and hydrophilic compounds, and for low-molecular-mass cations. To illustrate the use of the dish we describe the influence of external pH on the activity of the Na⁺/Ca²⁺ exchanger in intact cultured neonatal rat ventricular cardiomyocytes. The intracellular free calcium concentration ([Ca²⁺]_i) in the cardiomyocytes, measured using fura-2 and imaging fluorescence microscopy, was studied during sodium-free incubation. The resulting rise in [Ca²⁺]_i at pH 7.4 in one compartment was compared with that in the other compartment in which the pH was either 6.0, 7.0, 7.4 or 8.0. It was found that below pH 7.4, Na⁺/Ca²⁺ exchanger activity was diminished, whereas at pH higher than 7.4 the Na⁺/ Ca²⁺ exchanger activity was increased. We conclude that the two-compartment culture dish offers researchers a valuable new tool in the manipulation and the observation of single cells in culture.