Modafinil Reduces Neuronal Pyroptosis and Cognitive Decline After Sleep Deprivation

Abstract

Sleep deprivation (SD) induces systemic inflammation that promotes neuronal pyroptosis. The purpose of this study was to investigate the effect of an antioxidant modafinil on neuronal pyroptosis and cognitive decline following SD. Using a mouse model of SD, we found that modafinil improved learning and memory, reduced proinflammatory factor (IL-1β, TNF-α, and IL-6) production, and increased the expression of anti-inflammatory factors (IL-10). Modafinil treatment attenuated inflammasome activity and reduced neuronal pyroptosis involving the NLRP3/NLRP1/NLRC4-caspase-1-IL-1β pathway. In addition, modafinil induced an upregulation of brain-derived neurotrophic factor (BDNF) and synaptic activity. These results suggest that modafinil reduces neuronal pyroptosis and cognitive decline following SD. These effects should be further investigated in future studies to benefit patients with sleep disorders.

Keywords: inflammasome; modafinil; pyroptosis; sleep deprivation; synaptic plasticity.

FIGURE 1

The schematic representation of the experimental design. In the Morris Water Maze (MWM) test, mice were trained for adaptability on the first day, followed by formal training for 2–6 days, positioning navigation testing for 7–10 days (also at the SD stage), and additional space exploration testing after positioning navigation testing on the 10th day. The aim of testing all mice on the morning of day 7 was to exclude those with large individual differences. At the end of the day, they were randomly divided into four groups: the normal group, the Control + MD group, the SD group, and the SD + MD group; subsequently, the sleep deprivation stage was initiated, and the Control + MD group and SD + MD group were treated with modafinil. After the last spatial exploration test, all mice were sacrificed for tissue collection.

FIGURE 2

The effect of modafinil on learning and memory of SD. (A) The escape latency to reach the platform during the training sessions of the MWM test was detected. *P < 0.05, ^****P < 0.0001, significantly different from Control; ^#P < 0.05, ^##P < 0.01, ^###P < 0.001, significantly different from Control + MD group; ^HP < 0.05, significantly different from SD group; ^uP < 0.05, significantly different from SD + MD group. (B) The number of entries in the spatial acquisition trial was decreased. (C) The time spent in the target quadrant in the probe trail was increased by modafinil treatment. (D) The swimming path length was recorded after SD. (E) There was no significant difference in swimming speed among the four groups. (F) Representative swimming tracks of mice after SD for 72 h. All data presented are means ± SD; N = 9–15 mice per group. *P < 0.05, ^**P < 0.01, ^***P < 0.001.

FIGURE 3

Pyroptosis induced by SD was inhibited by modafinil. (A,B) Western blot data showing that modafinil suppressed the increase in the expression of NLRP3, NLRC4, NLRP1, ASC, caspase-1, GSDMD, and the downstream pro-inflammatory cytokines IL-1β and IL-18 in mice subjected to SD. All data presented are means ± SD; N = 4–5 mice per group. *P < 0.05, ^**P < 0.01, ^***P < 0.001, ^****P < 0.0001.

FIGURE 4

Modafinil decreased the expression of caspase-1 in the hippocampi of mice subjected to SD mice. (A) Representative pictures of double staining of caspase-1 and NeuN in the hippocampi of mice obtained by a fluorescence microscope. (B,C) Western blot analysis of caspase-1 expression in the hippocampi of mice subjected to SD and treated with vehicle or modafinil. All data presented are means ± SD; N = 4–5 mice per group. (D) Immunofluorescence analysis and quantification of the expression level of caspase-1. All data presented are means ± SD; N = 5–10 mice per group. *P < 0.05, ^**P < 0.01, ^***P < 0.001.

FIGURE 5

Modafinil altered the expression of inflammatory cytokines in the hippocampi of mice subjected to SD. Modafinil upregulated the expression of IL-1β IL-6 and TNF-α and downregulated the expression of IL-10. All data presented are means ± SEM; N = 5–8 mice per group. *P < 0.05, ^**P < 0.01, ^****P < 0.0001.

FIGURE 6

Modafinil decreased BDNF expression and alleviated dendritic spine loss in hippocampal CA3 pyramidal neurons in mice subjected to SD. (A,B) Western bot analysis of BDNF expression in the hippocampus. All data presented are means ± SD; N = 3–5 mice per group. (C,D) The density of CA3 pyramidal neurons, as measured by Golgi staining, was decreased in mice subjected to SD, and modafinil reversed this decrease. All data presented are means ± SD; N = 7–8 mice per group. *P < 0.05, ^**P < 0.01, ^****P < 0.0001.

FIGURE 7

The canonical inflammasome pathway. SD can activate inflammasomes in cells and further recruit ASC and pro-caspase-1 to form inflammasome complex. Active caspase-1 cleaves GSDMD to produce GSDMD pores on the cell membrane; active caspase-1 can activate pro-IL-1β and pro-IL-18, and then IL-1β and IL-18 are released from the GSDMD pores.