Ca(2+)/calmodulin-dependent protein kinases II (CaMKII) have important functions in regulating cardiac excitability and contractility. In the present study, we examined whether CaMKII regulated the transient outward K(+) current (I(to)) in whole-cell patch-clamped human atrial myocytes. We found that a specific CaMKII inhibitor, KN-93 (20 micromol/L), but not its inactive analog, KN-92, accelerated the inactivation of I(to) (tau(fast): 66.9+/-4.4 versus 43.0+/-4.4 ms, n=35; P<0.0001) and inhibited its maintained component (at +60 mV, 4.9+/-0.4 versus 2.8+/-0.4 pA/pF, n = 35; P<0. 0001), leading to an increase in the extent of its inactivation. Similar effects were observed by dialyzing cells with a peptide corresponding to CaMKII residues 281 to 309 or with autocamtide-2-related inhibitory peptide and by external application of the calmodulin inhibitor calmidazolium, which also suppressed the effects of KN-93. Furthermore, the phosphatase inhibitor okadaic acid (500 nmol/L) slowed I(to) inactivation, increased I(sus), and inhibited the effects of KN-93. Changes in [Ca(2+)](i) by dialyzing cells with approximately 30 nmol/L Ca(2+) or by using the fast Ca(2+) buffer BAPTA had opposite effects on I(to). In BAPTA-loaded myocytes, I(to) was less sensitive to KN-93. In myocytes from patients in chronic atrial fibrillation, characterized by a prominent I(sus), KN-93 still increased the extent of inactivation of I(to). Western blot analysis of atrial samples showed that delta-CaMKII expression was enhanced during chronic atrial fibrillation. In conclusion, CaMKII control the extent of inactivation of I(to) in human atrial myocytes, a process that could contribute to I(to) alterations observed during chronic atrial fibrillation.

The effects of tyrosine protein kinases (TK) on the L-type Ca(2+) current (I(Ca)) were examined in whole cell patch-clamped human atrial myocytes. The TK inhibitors genistein (50 microM), lavendustin A (50 microM), and tyrphostin 23 (50 microM) stimulated I(Ca) by 132 +/- 18% (P < 0.001), 116 +/- 18% (P < 0.05), and 60 +/- 6% (P < 0.001), respectively. After I(Ca) stimulation by genistein, external application of isoproterenol (1 microM) caused an additional increase in I(Ca). Dialyzing the cells with a protein kinase A inhibitor suppressed the effect of isoproterenol on I(Ca) but not that of genistein. Inhibition of protein kinase C (PKC) by pretreatment of cells with 100 nM staurosporine or 100 nM calphostin C prevented the effects of genistein on I(Ca). The PKC activator phorbol 12-myristate 13-acetate (PMA), after an initial stimulation (75 +/- 17%, P < 0.05), decreased I(Ca) (-36 +/- 5%, P < 0.001). Once the inhibitory effect of PMA on I(Ca) had stabilized, genistein strongly stimulated the current (323 +/- 25%, P < 0.05). Pretreating myocytes with genistein reduced the inhibitory effect of PMA on I(Ca). We conclude that, in human atrial myocytes, TK inhibit I(Ca) via a mechanism that involves PKC.

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