Common components of patch-clamp internal recording solutions can significantly affect protein kinase A activity.
Common components of whole-cell internal recording solutions were tested both in vitro and in patch-clamp experiments for their effects on the activity of cAMP-dependent protein kinase. Potassium fluoride (KF), 440 mM trimethylamine chloride and exclusion of bovine serum albumin (BSA) decreased the activity of the enzyme, while ethylene glycol-bis (beta-aminoethyl ether) N,N,N',N'-tetraacetic acid (EGTA) and the potassium salts of aspartate, gluconate, methylsulfate and monobasic phosphate increased its activity. Addition of KF to the internal solution produced a hyperpolarizing shift in the V1/2 of Ih channel activation, consistent with the KF-induced reduction of protein kinase A activity. Therefore, consideration of the composition of internal solutions is warranted when studying channel physiology by patch-clamp techniques.
D000818:Animals; D001224:Aspartic Acid / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D002478:Cells, Cultured; D002586:Cesium / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D002712:Chlorides / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005459:Fluorides / Q000494:pharmacology; D005942:Gluconates / Q000494:pharmacology; D007473:Ion Channels / Q000502:physiology; D007546:Isoquinolines / Q000494:pharmacology; D008744:Methylamines / Q000494:pharmacology; D018034:Olfactory Receptor Neurons / Q000737:chemistry / Q000166:cytology / Q000201:enzymology*; D018408:Patch-Clamp Techniques; D010710:Phosphates / Q000494:pharmacology; D010766:Phosphorylation; D017680:Potassium Compounds / Q000494:pharmacology; D051381:Rats; D015290:Second Messenger Systems / Q000187:drug effects / Q000502:physiology*; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D013449:Sulfonamides; D013463:Sulfuric Acid Esters / Q000494:pharmacology; D019789:Tetraethylammonium / Q000494:pharmacology