title: Isolation and Kv channel recordings in murine atrial and ventricular cardiomyocytes.
abstract: KCNE genes encode for a small family of Kv channel ancillary subunits that form heteromeric complexes with Kv channel alpha subunits to modify their functional properties. Mutations in KCNE genes have been found in patients with cardiac arrhythmias such as the long QT syndrome and/or atrial fibrillation. However, the precise molecular pathophysiology that leads to these diseases remains elusive. In previous studies the electrophysiological properties of the disease causing mutations in these genes have mostly been studied in heterologous expression systems and we cannot be sure if the reported effects can directly be translated into native cardiomyocytes. In our laboratory we therefore use a different approach. We directly study the effects of KCNE gene deletion in isolated cardiomyocytes from knockout mice by cellular electrophysiology - a unique technique that we describe in this issue of the Journal of Visualized Experiments. The hearts from genetically engineered KCNE mice are rapidly excised and mounted onto a Langendorff apparatus by aortic cannulation. Free Ca(2+) in the myocardium is bound by EGTA, and dissociation of cardiac myocytes is then achieved by retrograde perfusion of the coronary arteries with a specialized low Ca(2+) buffer containing collagenase. Atria, free right ventricular wall and the left ventricle can then be separated by microsurgical techniques. Calcium is then slowly added back to isolated cardiomyocytes in a multiple step comprising washing procedure. Atrial and ventricular cardiomyocytes of healthy appearance with no spontaneous contractions are then immediately subjected to electrophysiological analyses by patch clamp technique or other biochemical analyses within the first 6 hours following isolation.
authors.affiliation: Experimental and Clinical Research Center, Charité Medical Faculty and Max-Delbrück Center for Molecular Medicine.
authors.forename: Clemens
authors.initials: C
authors.lastname: Köhncke
authors.forename: Ulrike
authors.initials: U
authors.lastname: Lisewski
authors.forename: Leonhard
authors.initials: L
authors.lastname: Schleußner
authors.forename: Carolin
authors.initials: C
authors.lastname: Gaertner
authors.forename: Saskia
authors.initials: S
authors.lastname: Reichert
authors.forename: Torsten K
authors.initials: TK
authors.lastname: Roepke
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D006325:Heart Atria / Q000166:cytology / Q000378:metabolism; D006352:Heart Ventricles / Q000166:cytology / Q000378:metabolism; D051379:Mice; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D024642:Potassium Channels, Voltage-Gated / Q000032:analysis / Q000378:metabolism*
chemical_list: D024642:Potassium Channels, Voltage-Gated; D002118:Calcium