Objective To unravel the mechanism of aggravation of neuroinflammation induced by microglial pyroptosis following chronic infection with the Toxoplasma gondii Chinese 1 genotype Wh6 strain (TgCtwh6). Methods Twenty C57BL/6 mice were divided into a control group and an infection group randomly, of 10 mice each group. Mice in the infection group were each administered by oral gavage with 10 TgCtwh6 strain cysts suspended in 200 μl PBS to establish a chronic T. gondii infection model, while mice in the control group received an equal volume of PBS via oral gavage. Mouse cerebral cortex was collected, and the differential expression profiles of pyroptosis-related genes were screened in the mouse cerebral cortex using transcriptome sequencing 6 weeks post-infection. Sixty C57BL/6 mice were divided into a control group, an infection group, a VX-765 control group, and a VX-765 treatment group randomly, of 15 mice each group. Mice in the infection group and VX-765 treatment group were each administered by oral gavage with 10 TgCtwh6 strain cysts. Mice in the VX-765 control group and VX-765 treatment groups were intraperitoneally injected with the caspase-1 inhibitor VX-765 (at a dose of 50 mg/kg) once every 2 days since 4 weeks post-infection for 7 injections, and mouse cerebral cortex was collected from each group 8 weeks post-infection. BV2 mouse microglial cells were assigned to a control group, an infection group, a VX-765 control group, and a VX-765 treatment group, of 5 × 10⁵ cells each group. Cells in the infection group and VX-765 treatment group were infected with an equal amount of TgCtwh6 strain tachyzoites, while cells in the VX-765 control and VX-765 treatment groups were treated with 20 μmol/L VX-765. Cells were harvested 24 h following culture. RNA was extracted from mouse cerebral cortex and BV2 cells, and the relative RNA expression of pyroptosis-related genes caspase-1 and gasdermin D (GSDMD) and pro-inflammatory cytokines interleukin-18 (IL-18), IL-1β, IL-6, tumor necrosis factor-α (TNF-α) were quantified using real-time quantitative reverse transcription PCR (RT-qPCR) assay. Protein was extracted from mouse cerebral cortex and BV2 cells, and the expression of caspase-1, caspase-1 p20 subunit (caspase-1 p20), GSDMD, and GSDMD N-terminal fragment (GSDMD-N) were determined using Western blotting assay. Comparisons of means between groups were done using independent sample t-test, and multi-group comparisons were conducted with one-way analysis of variance (ANOVA) and Tukey’s post hoc test. Results Transcriptome sequencing revealed upregulation of pyroptosis-related genes NOD-like receptor thermal protein domain associated protein 3 (NLRP3), caspase-1, caspase-4, GSDMD, IL-1β and IL-18 expression in the mouse cerebral cortex in the infection group, with relatively higher caspase-1 (16.48 ± 6.40 vs. 1.33 ± 0.42; t = 4.09, P < 0.05) and GSDMD (12.80 ± 5.62 vs. 0.59 ± 0.20; t = 3.76, P < 0.05) expression in the infection group than in the control group. RT-qPCR assay quantified relatively higher mRNA levels of caspase-1 (4.04 ± 0.38 vs. 0.88 ± 0.18), GSDMD (9.67 ± 0.27 vs. 1.00 ± 0.26), IL-18 (1.49 ± 0.16 vs. 0.97 ± 0.16), IL-1β (7.50 ± 0.27 vs. 0.94 ± 0.21), IL-6 (4.96 ± 0.79 vs. 0.92 ± 0.22) and TNF-α (9.97 ± 1.77 vs. 0.82 ± 0.42) in the mouse cerebral cortex in the infection group than in the control group [Tukey’s Honest Significant Difference test (Tukey’s HSD test), all P < 0.01], and lower relative mRNA expression of caspase-1 (1.13 ± 0.13), GSDMD (0.87 ± 0.25), IL-18 (0.77 ± 0.05), IL-1β (0.89 ± 0.11), IL-6 (1.03 ± 0.05), and TNF-α (0.93 ± 0.43) in the VX-765 treatment group than in the infection group (Tukey’s HSD test, all P < 0.05). Western blotting assay determined higher relative expression of GSDMD (1.49 ± 0.14 vs. 0.41 ± 0.29), caspase-1 (1.38 ± 0.24 vs. 0.50 ± 0.29), GSDMD-N (1.60 ± 0.17 vs. 0.70 ± 0.30), and caspase-1 p20 (0.89 ± 0.11 vs. 0.17 ± 0.06) in the mouse cerebral cortex in the infection group than in the control group (Tukey’s HSD test, all P < 0.05), and lower relative expression of GSDMD (0.76 ± 0.11), caspase-1 (0.43 ± 0.15), GSDMD-N (0.72 ± 0.29), and caspase-1 p20 (0.43 ± 0.14) in the VX-765 treatment group than in the infection group (Tukey’s HSD test, all P < 0.05). RT-qPCR assay detected relatively higher mRNA expression of caspase-1 (1.64 ± 0.03 vs. 0.94 ± 0.05), GSDMD (2.17 ± 0.40 vs. 0.81 ± 0.21), IL-18 (3.01 ± 0.31 vs. 1.02 ± 0.01), IL-1β (3.47 ± 0.05 vs. 0.99 ± 0.08), IL-6 (3.64 ± 0.15 vs. 0.94 ± 0.09), and TNF-α (2.23 ± 0.20 vs. 0.99 ± 0.03) in BV2 cells in the infection group than in the control group (Tukey’s HSD test, all P < 0.01), and significantly lower mRNA expression of caspase-1 (0.70 ± 0.06), GSDMD (1.29 ± 0.27), IL-18 (1.07 ± 0.01), IL-1β (0.98 ± 0.03), IL-6 (0.52 ± 0.03), and TNF-α (1.26 ± 0.03) in the VX-765 treatment group than in the infection group (Tukey’s HSD test, all P < 0.05). Western blotting assay determined higher relative expression of GSDMD (1.43 ± 0.34 vs. 0.67 ± 0.23), caspase-1 (1.45 ± 0.14 vs. 0.48 ± 0.02), GSDMD-N (1.29 ± 0.40 vs. 0.56 ± 0.22), and caspase-1 p20 (1.25 ± 0.11 vs. 0.41 ± 0.18) in BV2 cells in the infection group than in the control group (Tukey’s HSD test, all P < 0.05), and lower relative expression of GSDMD (0.78 ± 0.20), caspase-1 (0.61 ± 0.07), GSDMD-N (0.56 ± 0.22), and caspase-1 p20 (0.47 ± 0.17) in the VX-765 treatment group than in the infection group (Tukey’s HSD test, all P < 0.05). Conclusion Chronic infection with the TgCtwh6 strain induces microglial pyroptosis and promotes the release of pro-inflammatory cytokines, thereby exacerbating neuroinflammatory responses, while VX-765 inhibitor is effective to suppress pyroptosis and inflammatory responses.