Patch clamp analysis of chemically activated and modulated ionic channels in isolated mammalian cardiomyocytes

Techniques routinely utilized in this laboratory for recording currents through single ionic channels of isolated atrial and ventricular rat cardiomyocytes are described. Emphasis is placed in two main areas: first, on methods for obtaining a sufficient yield of Ca++-tolerant myocytes suitable for patch clamp experiments, and secondly, on methods for analyzing the temporal characteristics of 'patched' ionic channels. These methods were used on acetylcholine activated K+ channels in isolated atrial myocytes and on an inwardly-rectifying K+ channel in ventricular myocytes. The latter is an example of a hormonally modulated K+ channel, since its activity could be substantially increased by norepinephrine. Analysis of the closed and open time distributions suggested that one of the closed states of this channel is markedly abbreviated by norepinephrine, whereas the open state is nearly unaffected. Norepinephrine was effective when channel activity was recorded from on-cell patches and the hormone was added to the solution bathing the cell membrane outside of the patched area. This indicates that a second messenger substance is probably mediating the action of norepinephrine.