Simulated weightlessness affects the expression and activity of neuronal nitric oxide synthase in the rat brain

Abstract

Spaceflight induces pathophysiological alterations in various organs. To study pathophysiological adaptations to weightlessness on the ground, the tail suspension (TS) rat model has been used to simulate the effects of weightlessness. There is currently little information on the effect of TS on the expression and activity of nitric oxide synthase (NOS) in the brain. In this study, we examined time-dependent alterations in the expression and activity of neuronal NOS (nNOS) in the brains of TS rats. Male Sprague-Dawley rats were tail-suspended for 1 (TS1), 7 (TS7), and 14 (TS14) days or rested on the ground for 3 days after 14 days of TS. TS1 and TS7 rats exhibited no significant alterations in the expression of nNOS compared to control rats, whereas nNOS expression in TS14 rats was significantly upregulated compared to control rats. Normalized expression of nNOS mRNA and protein in TS14 rats (1.86 ± 0.48 and 1.84 ± 0.29, respectively) were significantly higher than that of control rats (P < 0.001 and P < 0.001, respectively). Consistent with these results, significant elevations in NOS activity and NO production were observed in TS14 rats. Thus, we demonstrated a significant upregulation of nNOS expression, accompanied by significant increases in NOS activity and NO production, in the brain of rats exposed to simulated weightlessness.

Keywords: Pathology Section; brain; neuronal nitric oxide synthase; rat; simulated weightlessness; tail suspension.

Figure 1. Scheme of the experimental design

The experimental groups were exposed to simulated weightlessness by tail suspension (TS) for 1 (TS1), 7 (TS7), or 14 (TS14) days or rested on the ground for 3 days after 14 days of TS (TS14+3). The number of rats per group was 8.

Figure 2. Effects of simulated weightlessness by tail suspension on body weight

Box and whisker diagram. The band inside each box is the median value of the indicated group. Body weight exhibited an increasing trend during the observation period, but the differences among groups were not statistically significant.

Figure 3. Effects of simulated weightlessness by tail suspension (TS) on neuronal nitric oxide synthase (nNOS) expression

A. nNOS mRNA expression was significantly elevated at 14 days of TS (P < 0.001) and remained significantly high after 3 days post-suspension (P = 0.002). B. Significantly elevated nNOS protein levels were observed in the TS14 (P < 0.001) and TS14+3 (P < 0.001) rats. The expression levels of both mRNA and protein were slightly decreased in the TS14+3 rats compared to those in the TS14 rats, but the differences between the two groups were not statistically significant.

Figure 4. Effects of simulated weightlessness by tail suspension (TS) on nitric oxide synthase (NOS) activity and nitric oxide (NO) production

Data are presented as box and whisker diagrams, and the band inside each box is the median value of the indicated group. An L-[¹⁴C]-citrulline assay and a nitrate plus nitrite assay revealed similar trends with respect to the levels of NOS enzymatic activity and NO production, both of which were significantly increased after 14 days of TS. A. NOS activity was significantly upregulated in TS14 (P < 0.001) and TS14+3 (P < 0.001) rats. B. The maximal nitrate plus nitrite level was observed after 14 days of TS. The time-dependent alteration pattern was consistent with that of NOS expression and activity.