It has recently been suggested that sodium-calcium exchange current (INa-Ca) may be involved in cardiac pacemaking, for example by contributing to the end of the pacemaker depolarisation in cat sino-atrial (SA) node cells (Huser et al. 2000). The aim of this study was to investigate the possible role of INa-Ca in pacemaking in guinea-pig SA node cells.
Male guinea-pigs were killed by cervical dislocation following stunning. Beating rate was measured either in atrial preparations with an intact SA node, or in single SA node cells isolated enzymatically from the heart. Calcium transients in single SA node cells were measured using either confocal microscopy (cells loaded with 3 mM fluo-4 AM for 10 min), or conventional fluorescence microscopy (1.5 mM indo-1 AM, 10 min). INa-Ca was inhibited by reducing the Na+ concentration of the superfusion solution (2.5 mM Ca2+, 36 °C) from 133 to 73.75 mM (Li2+ as the substitute). Data are quoted as means ± S.E.M. and statistical significance assessed using Student’s paired t test.
In the absence of other drugs, switching to a low Na+ solution caused a decrease in beating rate of 16 ± 2 % in atrial preparations (P < 0.05, n = 5), and 35 ± 5 % in single SA node cells (P < 0.05, n = 5). In SA node cells imaged using confocal microscopy, the rate of generation of spontaneous calcium transients was reduced by 14 ± 4 %, calcium transient amplitude (F/F0) was increased by 91 ± 22 %, and calcium transient duration was increased as a result of a 50 ± 14 % lengthening in the time to peak (all after 1 min in low Na+ solution, P < 0.05, n = 4); there was no significant change in the time for decay of the calcium transient to 50 % of its peak. In three of these four cells, there was also an increase in the minimum diastolic calcium level.
In the presence of 100 nM isoprenaline, switching to a low Na+ solution caused a 41 ± 1 % decrease in beating rate in atrial preparations (P < 0.05, n = 3). In single SA node cells, switching to low Na+ in the presence of 100 nM isoprenaline reduced the rate of generation of spontaneous calcium transients (measured using indo-1) by 17 ± 3 %, increased calcium transient amplitude by 24 ± 7 %, and lengthened time to peak by 34 ± 11 % (P < 0.05, n = 6) with no significant change in the time taken for 50 % decay. The minimum diastolic calcium level was increased by 11 ± 3 % (P < 0.05, n = 6).
These data are consistent with a role for INa-Ca in the pacemaker depolarisation of guinea-pig SA node cells. It remains to be determined how changes in calcium transient characteristics and in minimum diastolic calcium levels contribute to the regulation of pacemaker rate.
This work was supported by BHF and The Wellcome Trust.
All procedures accord with current UK legislation.