channels. Other inward currents such as sodium currents or inward rectifier currents were absent. We conclude that undifferentiated hMSC express a distinct pattern of ion channel mRNA and functional ion channels that might contribute to physiological cell function.

Objective: In chronic atrial fibrillation (cAF) the potassium current I K,ACh develops agonist-independent constitutive activity. We hypothesized that abnormal phosphorylation-dependent regulation underlies the constitutive I K,ACh activity. Methods: We used voltage-clamp technique and biochemical assays to study I K,ACh regulation in atrial appendages from 61 sinus rhythm (SR), 11 paroxysmal AF (pAF), and 33 cAF patients. Results: Compared to SR basal current was higher in cAF only, whereas the muscarinic receptor (2 μmol/L carbachol)-activated I K,ACh was smaller in pAF and cAF. In pAF the selective I K,ACh blocker tertiapin abolished the muscarinic receptor-activated I K,ACh but excluded agonist-independent constitutive I K,ACh activity. Blockade of type-2A phosphatase and the subsequent shift to increased muscarinic receptor phosphorylation (and inactivation) reduced muscarinic receptor-activated I K,ACh in SR but not in cAF, pointing to an impaired function of Gprotein-coupled receptor kinase. Using subtype-selective kinase inhibitors we found that in SR the muscarinic receptor-activated I K,ACh requires phosphorylation by protein kinase G (PKG), protein kinase C (PKC), and calmodulin-dependent protein kinase II (CaMKII), but not by protein kinase A (PKA). In cAF, constitutive I K,ACh activity results from abnormal channel phosphorylation by PKC but not by PKG or CaMKII, whereas the additional muscarinic receptor-mediated I K,ACh activation occurs apparently without involvement of these kinases. In cAF, the higher protein level of PKCε but not PKCα, PKCβ 1 or PKCδ is likely to contribute to the constitutive I K,ACh activity. Conclusions: The occurrence of constitutive I K,ACh activity in cAF results from abnormal PKC function, whereas the muscarinic receptormediated I K,ACh activation does not require the contribution of PKG, PKC or CaMKII. Selective drug targeting of constitutively active I K,ACh channels may be suitable to reduce the ability of AF to become sustained.

This work gives evidence for a differential recruitment of human PKC isoforms in various forms of myocardial hypertrophy and heart failure.

Freshly isolated and primary cultured adult rat cardiomyocytes were used to elucidate the mechanism of action of the new oral antidiabetic agent (+/-)-5-[4-(6-hydroxy-2, 5, 7, 8-tetramethyl-chroman-2-yl-methoxy)benzyl]-2,4-thiazolidinedione (troglitazone) on the heart. Interaction with protein kinase C (PKC) and regulation of glucose transport were evaluated as possible sites of drug action. Acute treatment (30 min) of cardiomyocytes with troglitazone did not affect the phorbolester-induced membrane association of PKC-delta and PKC-epsilon, which represent the major isoforms present in these cells. However, under these conditions the phorbolester-mediated increase in membrane associated PKC activity was inhibited by 43 +/- 4% (n = 4) without affecting the basal distribution of PKC activity. In contrast to these findings, troglitazone had no acute effect on basal or insulin-stimulated glucose transport in freshly isolated cardiomyocytes; even after 120 min treatment an unaltered release of lactate was determine in the presence of the drug. After 20 h in serum-free culture troglitazone induced a dose-dependent increase in 2-deoxyglucose uptake reaching a 40-fold stimulation at 5 mumol/l. This was paralleled by a dose-dependent increase of glucose transporter-1 (GLUT1) and GLUT4 protein expression to 320 +/- 80 and 156 +/- 15% of control, respectively. In addition, chronic exposure to troglitazone increased the GLUT4 abundance in a plasma membrane fraction about twofold. These data show that troglitazone exerts multiple effects on cardiomyocytes involving inhibition of PKC and regulation of glucose transporter expression and distribution. We suggest that an increased glucose supply may be beneficial for the diabetic heart and that modulation of PKC-activity could be relevant for improving insulin action in muscle tissue.

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