Table 2.
Quantitation of ASC morphology from confocal images
| Apical Vacuole Diameter L, μm | Apical Vacuole Diameter W, μm | Diameter L/Diameter W | |
|---|---|---|---|
| Control | 4.8 ± 0.69 (110;3) | 4.6 ± 0.51 (110;3) | 1.04 |
| Histamine | 20.8 ± 0.73 (102;3)* | 21.1 ± 1 (102;3)* | 0.99 |
| ML-7 | 2.7 ± 0.29 (108;3)* | 2.4 ± 0.25 (108;3)* | 1.10 |
| ML-7 + Histamine | 2.6 ± 0.3 (100;3)† | 2.8 ± 0.33 (100;3)† | 0.93 |
| Histamine + ML-7 | 21.8 ± 1.2 (96;3)* | 20.1 ± 0.95 (96;3)* | 1.08 |
| Histamine + NaSCN− | 6.9 ± 1.2 (74;3)† | 7.2 ± 0.95 (74;3)† | 0.95 |
| Y-27632 | 4.0 ± 0.51 (112;3) | 3.8 ± 0.23 (112;3) | 1.05 |
| Y-27632 + Histamine | 21.2 ± 0.73 (105;3)* | 19.3 ± 0.93 (105;3)* | 1.10 |
| Blebbistatin | 3.9 ± 0.33 (110;3) | 4.3 ± 0.72 (110;3) | 0.91 |
| Blebbistatin + Histamine | 22.0 ± 0.9 (113;3)* | 21.3 ± 0.81 (113;3)* | 1.03 |
| Jasplakinolide | 12.7 ± 1.44 (101;3)* | 3.4 ± 0.45 (101;3)* | 3.70* |
| Jasplakinolide +Histamine | 13.2 ± 1.3 (104;3)† | 3.7 ± 0.29(104;3)† | 3.60† |
| Histamine + Jasplakinolide | 19.5 ± 2.0 (90;3)* | 20.5 ± 2.2 (90;3)* | 0.95 |
| ML-7 + Jasplakinolide | 2.1 ± 0.19 (97;3)*‡ | 2.3 ± 0.31 (97;3)*‡ | 0.91 |
| Blebbistatin + Jasplakinolide | 4.3 ± 0.39 (78;3)‡ | 3.9 ± 0.45 (78;3)‡ | 1.10 |
| Nocodazole + Jasplakinolide | 3.6 ± 0.31 (75;3)*‡ | 1.5 ± 0.21 (75;3)*‡ | 2.40*‡ |
Cells were treated as follows: Control (100 μM cimetidine), Histamine (100 μM histamine for 30 min), ML-7 (10 μM ML-7 for 30 min), ML-7 + Histamine (10 μM ML-7 for 30 min followed by 100 μM histamine for 30 min), Histamine + ML-7 (100 μM histamine for 30 min followed by 10 μM ML-7 for 30 min), Histamine + NaSCN (100 μM histamine +3 mM NaSCN together for 30 min), Y-27632 (10 μM Y-27632 for 30 min), Y-27632 + Histamine (10 μM Y-27632 for 30 min followed by 100 μM histamine for 30 min), Blebbistatin (50 μM blebbistatin for 30 min), Blebbistatin + Histamine (50 μM blebbistatin for 30 min followed by 100 μM histamine for 30 min), Jasplakinolide (3 μM jasplakinolide for 60 min), Jasplakinolide + Histamine (3 μM jasplakinolide for 30 min followed by 100 μM histamine for 30 min), Histamine + Jasplakinolide (100 μM histamine for 30 min followed by 3 μM jasplakinolide for 30 min), ML-7 + Jasplakinolide (10 μM ML-7 for 30 min followed by 3 μM jasplakinolide for 30 min), Blebbistatin + Jasplakinolide (50 μM blebbistatin for 30 min followed by 3 μM jasplakinolide for 30 min), Nocodazole + Jasplakinolide (20 μM nocodazole for 30 min followed by 3 μM jasplakinolide for 30 min). Following these treatments, cells were fixed and stained (see materials and methods). Diameters of apical secretory canaliculus (ASC) were measured (from ezrin or myosin IIB staining) along the long axis and then at 90° to this in confocal micrographs of ASCs, and the ratio of the 2 measurements was calculated. Averages ± SD are reported for 75–113 ASCs measured in parietal cells from 3 different experiments each. Statistical comparisons:
P < 0.05 for comparison to control;
P < 0.05 for comparsion to histamine;
P < 0.05 for comparison to jasplakinolide.