Evidence for Na+/Ca2+ exchange in isolated smooth muscle cells: a fura-2 study

Abstract

Isolated smooth muscle cells (SMC) from guinea pig taenia coli were employed. Suspension of cells were externally loaded in saline with the fluorescent calcium indicators quin-2/AM or fura-2/AM at 20-40 microM or 4 microM respectively, resulting in an estimated intracellular concentration of 100-200 microM for quin-2 or 10-20 microM fura-2 (free acid). On addition of 100 microM carbachol or high K+o (80 mM) depolarization, fura-2 loaded cells contracted (104 +/- 47 micron, n = 121 rest: 39 +/- 13 micron, n = 59 contracted) identically to control (103 +/- 35 micron, n = 232 rest: 39 +/- 16 micron, n = 89 contracted) cells, whereas quin-2 loaded cells were unresponsive to these protocols and there was no significant length change. The Ca2+i of fura-2 loaded cells was 100 +/- 18 nM (mean +/- SD, n = 15) and was not significantly different from quin-2 loaded cells 107 +/- 26 nM (n = 13). Treatment of fura-2 loaded cells with 100 microM ouabain saline for 10-60 min progressively elevated the Ca2+i to a mean of 266 +/- 83 nM (n = 15). Reduction of Na+o (96% Li+ replaced) significantly increased Ca2+i to 317 +/- 77 nM (n = 8). After pretreatment with ouabain (100 microM), Na+o replacement (Li+) increased Ca2+i at a significantly faster rate [3.6 nM min-1 (control) cf. 19.8 nM min-1 (ouabain)].