A Microflow Superfusion System for Use with Excised Membrane Patches

A new method for rapidly changing the solution superfusing excised membrane patches is described. The main advantage of this method over previous rapid exchange methods is that it is possible to use very small quantities of solution (minimal volume about 100 microliters). The method consists of inserting the pipette tip, after obtaining an excised patch, into a small tube which can be independently perfused. Air bubbles are used to separate different solutions and to produce rapid "step" changes of solution. To test this method we examined the rate of change of: the pipette resistance produced in response to changing to a solution of lower conductivity, the amplitude of single K-channel currents in response to changing external K. The results suggest that the solution exchange is essentially a step function and is complete within 10-20 ms. The recording is briefly interrupted during the time taken for the passage of the air bubble past the pipette tip (less than 250 ms). In general, the passage of the air bubble did not affect the seal resistance. This method will be particularly useful for studies of the interactions of single ionic channels with chemicals which can only be obtained in small quantities.