NOS inhibitors increase cell number, brain volume, and cell density in the tadpole brain
| Control, 3 d | l-NAME, 3 d | Change (% of control) | Control, 7 d | l-NAME, 7 d | Change (% of control) | ETU, 7 d | Change (% of control) | |
|---|---|---|---|---|---|---|---|---|
| Cell number | 3644 ± 158 | 5136 ± 339* | 140.9 | 8373 ± 928 | 13742 ± 510* | 164.1 | 12870 ± 690* | 153.7 |
| Volume (mm3 × 10−4) | 52.7 ± 3.8 | 67.8 ± 9.1 | 128.7 | 99.4 ± 4.9 | 107.0 ± 3.7 | 107.6 | 123.0 ± 4.3* | 123.7 |
| Cell density (cells/mm3 × 10−4) | 69 ± 7 | 76 ± 11 | 110.1 | 84 ± 6 | 128 ± 10* | 152.4 | 105 ± 3* | 125.0 |
| BrdU-positive cells (total) | 133 ± 13 | 499 ± 402-160 | 375.2 | 141 ± 10 | 424 ± 40* | 300.7 | 537 ± 25* | 380.9 |
| BrdU-positive cells/103 cells (BrdU index) | 37 ± 3 | 97 ± 52-160 | 262.1 | 17 ± 2 | 31 ± 3* | 182.4 | 42 ± 3* | 247.1 |
| TUNEL-positive cells (total) | 115 ± 7 | 168 ± 162-160 | 146.1 | 83 ± 8 | 123 ± 12* | 148.2 | 179 ± 25* | 215.7 |
| TUNEL-positive cells/103 cells (TUNEL index) | 32 ± 3 | 33 ± 4 | 103.1 | 10 ± 1 | 9 ± 1 | 90.0 | 14 ± 4 | 140.0 |
Tadpole brains were implanted with pieces of Elvax impregnated with NOS inhibitors l-NAME and ETU or saline solution as a control. Data were collected as described in Table 1. The number of sections encompassing the optic tectum did not differ significantly between control and treated animals after 3 d (4 ± 1 in all groups; p = 0.2) or 7 d (5 ± 1 in all groups; p = 0.3) of treatment. Changes after 3 d of treatment with l-NAME or saline were analyzed as described in Table 1 (*p < 0.05;
↵F2-160 p < 0.01). Data collected after 7 d of treatment with l-NAME, ETU, or saline were subjected to one-way ANOVA with treatment (F(2,6) = 15.58; p = 0.004) as the main effect. Two pair-wise subsequent planned comparisons were adjusted for experiment-wise error rate (α = 0.05) by using a critical value of α′ = 0.025. For this experiment,
↵* p < 0.025. Inhibition of NOS during Xenopus brain development resulted in multiple changes. Proliferation as measured by BrdU index was significantly increased. After 3 d of treatment, the BrdU index of the brain treated with l-NAME was 262.1% larger (p< 0.01; F = 107) than in the control animals. It remained higher after 7 d of treatment with both inhibitors: 182.4% of control after treatment with l-NAME (p < 0.025; F = 5.0) and 247.1% after treatment with ETU (p < 0.01; F = 15). Total cell number in the treated brain was also significantly increased: to 140.9% of control (p < 0.05;F = 15.9) after 3 d of treatment withl-NAME, to 164.1% of control (p < 0.01;F = 27) after 7 d of treatment withl-NAME, and to 153.7% of control (p < 0.01; F = 18.9) after 7 d of treatment with ETU. Volume was significantly increased after 7 d of treatment with ETU to 123.7% of control (p < 0.025; F = 7). Cell density significantly increased after 7 d of treatment with l-NAME to 152.4% of control (p < 0.01; F = 19.5) or with ETU to 125% of control (p < 0.025; F = 4). Changes in cell number were not accompanied by significant changes in the rate of programmed cell death as measured by TUNEL index.