Calcium-activated potassium (K_Ca) channel activation is involved in smooth muscle/endothelial hyperpolarization, inducing coronary vasodilatation in vivo largely mediated by bradykinin (BK; Paolocci et al. 2001). However, their presence and role in cardiac myocytes is not yet known. We hypothesized that K_Ca channels are present in the heart playing a major role in modulating contractility. We used Western blot analysis to detect large and intermediate K_Ca channels in isolated myoctes. Performing studies in isolated, perfused rat hearts and myoctes we aimed to test the impact of K_Ca channel blockade on contractility, sarcomere length and myocyte calcium transients. Wistar rats (5-6 months old) were killed by cervical dislocation after pentobarbitone (0.2 g I.P.) anaesthesia. Myocytes were isolated from hearts perfused with gassed, calcium-free bicarbonate buffer, followed by enzymatic digestion. For sarcomere length studies, myocytes were incubated with 10 µM fura-2 AM, continuously perfused with Tyrode solution, and stimulated at 1 Hz. Isolated hearts were retrogradely perfused (10 ml min^-1) with Krebs-Henseleit, and paced at 300 b.p.m. Data are means ± S.E.M.

Using an anti-BK_Ca (large conductance) channel antibody, Western blot analysis of protein purified from isolated myocytes showed a 40 kDa protein, suggesting the presence of K_Ca channel β subunit in myocytes. The antibody pre-incubation with the control antigen led to disappearance of the band validating the specificity of the signal detected. Similarly, a weaker band of the same size was obtained with anti-charybdotoxin (CbTX) antibody, suggesting the intermediate K_Ca channels presence. In perfused hearts, BK (500 ng ml^-1) depressed myocardial contractility: dP/dt_max decreased from 2560 ± 189 to 2317 ± 144 mmHg s^-1 -9 ± 3 % (P = 0.01, n = 6), and ESP -14 ± 2 % (P < 0.001). These changes were prevented by the intermediate and small-K_Ca blockers CbTX and apamin (18.6 and 186 nM, respectively) but not BK-induced vasodilatation. Moreover, toxins alone increased dP/dt_max by 4.3 ± 1.3 % (P = 0.01). Consistently, when isolated myoctes were perfused with K_Ca blockers (100 nM each), sarcomere length shortening was enhanced by 116 ± 47 % (P < 0.05 vs. base, n = 7) as well as peak Ca²⁺ transients (+42 ± 16% P < 0.05). Here we showed for the first time the presence of both large and intermediate K_Ca channels in cardiac myocytes and demonstrate their role in exerting tonic inhibition of cardiac contractility. Thus activity of these channels may participate in the control of cardiac contractility both at baseline and in response to specific autocrine/paracrine agonists (Rastaldo et al. 2001).

Paolocci, N., Pagliao, P., Isoda, T., Saavedra, F.W. & Kass, D.A. (2001). Circulation 103, 119-124.

Rastaldo, R., Paolocci, N., Chiribiri, A., Penna, C., Gattullo, D. & Pagliaro, P. (2001). Am. J. Physiol. 280, H2823-2832.