Previous work has shown that Na⁺-Ca²⁺ exchange activity appears to occur mainly within the t-tubules of cardiac ventricular myocytes (Yang et al. 2002). We have, therefore, investigated the effect of detubulation on the Na⁺-Ca²⁺ exchange-mediated increase of spontaneous contractile activity that occurs when extracellular Na⁺ concentration ([Na⁺]_o) is decreased, or intracellular Na⁺ concentration ([Na⁺]_i) increased using strophanthidin.

Ventricular myocytes were isolated from the hearts of Wistar rats killed humanely. Cells were detubulated as described previously (Yang et al. 2002). Spontaneous contractile activity was assessed visually in control solution (140 mM Na⁺), and in solutions with reduced [Na⁺] (replaced with Li⁺) and in the presence of strophanthidin (100 µM); observations were started 1 min after decreasing [Na⁺]_o from 140 mM. Cells were considered spontaneously active if they showed spontaneous contractile activity within 5 s of being observed.

Figure 1 shows that reducing [Na⁺]_o increased spontaneous contractile activity in control cells. Calculation of E_Na/Ca ([Na⁺]_i = 10 mM, [Ca²⁺]_i = 100 nM, [Ca²⁺]_o = 1 mM) suggests that the exchanger will reverse, to produce Ca²⁺ influx, when [Na⁺]_o is less than ~70 mM, the concentration below which spontaneous contractile activity increased significantly (P < 0.05). Figure 1 also shows that the increase in spontaneous activity caused by decreasing [Na⁺]_o was significantly reduced in detubulated cells. In both normal and detubulated cells, the increase in spontaneous activity was inhibited by Ni²⁺ (which inhibits Na⁺-Ca²⁺ exchange; Fig. 1) and the sarcoplasmic reticulum inhibitor ryanodine (1 µM), but was unaffected by the Ca²⁺ channel blocker nifedipine (20 µM; not shown). The increase in spontaneous contractile activity caused by strophanthidin was also reduced in detubulated cells.

These data provide functional evidence for the loss of some, but not all, Na⁺-Ca²⁺ exchange activity in detubulated ventricular myocytes.

This work was supported by the Wellcome Trust, British Heart Foundation and School of Biomedical Sciences