Fig 2: GLT‐1 overexpression attenuated neuropathological injuries induced by hyperexcitability in hippocampus and improved cognitive function in T2DM mice suffering anesthesia/surgery. (A) Representative immunofluorescent images of c‐Fos and Camkllα staining at 3 h after surgery. Scale bar: 100 μm. (B) Quantification of c‐Fos immunoreactivity in Camkllα+ cells (n = 9). (C) Representative Western blot of SOD2 at day 4. (D) Quantitative analysis of SOD2 expression compared with the control group (n = 6). (E) Analysis of MMP in the hippocampus region among the two groups at day 4 (n = 6). (F) Representative images of Tunel and Camkllα staining at day 4 after anesthesia/surgery. Scale bar: 100 μm. (G) Quantitation of co‐labeling rate of Tunel and Camkllα (n = 6). (H) LTP recording in the hippocampal CA1 region. The arrow indicated the time point of TBS application. fEPSP slope before TBS was recorded as representative waveform 1 and fEPSP slope after TBS was recorded as representative waveform 2. (I) The average fEPSP slope during the last 20 min after TBS (n = 6). (J) The exploration preference in NORT training (n = 10). (K) The exploration preference in NORT test (n = 10). (l) The exploration preference in OLT training (n = 10). (M) The exploration preference in OLT test (n = 10). All data are expressed as the mean ± SEM. *p < 0.05, **p < 0.01.

Fig 3: GLT‐1 expression in hippocampal CA1 astrocytes was downregulated in T2DM mice. (A) Representative Western blot of GLT‐1 at 3 h after anesthesia/surgery. (B) Quantitative analysis of GLT‐1 expression compared with the control group (n = 6). (C) Quantification of GLT‐1 immunoreactivity in GFAP+ cells (n = 9). (D) Representative immunofluorescent images of GLT‐1 and GFAP staining at 3 h after anesthesia/surgery. Scale bar: 100 μm. All data are expressed as the mean ± SEM. *p < 0.05, **p < 0.01.

Fig 4: GLT1 overexpressed in the hippocampal CA1 astrocytes. (A) Timeline of the experiments. (B) The location of viral microinjection with a rAAV‐Ef1a‐DIO‐GLT1‐EGFP‐WPRE‐pA (AAV‐GLT1) or a control rAAV‐Ef1a‐DIO‐EGFP‐WPRE (AAV‐VEH). Fluorescence images showing efficient expression of the AAV‐GLT1 vector in the CA1 region. Scar bar = 200 μm. (C) GLT1 (green) was mainly expressed in astrocytes. No co‐localization with the microglia marker Iba1 or the neuron marker NeuN was detected. Scale bar = 100 μm. (D) Percentage of GFAP cells that were EGFP labeled. (E) Percentage of EGFP+ cells expressing GFAP (n = 6). (F) Representative immunofluorescent images of GLT‐1 and GFAP staining. Scar bar = 100 μm. (G) Representative Western blot of GLT‐1. (H) Quantification of GLT‐1 immunoreactivity in GFAP+ cells (n = 9). (I) Quantitative analysis of GLT‐1 expression compared with the EGFP group (n = 6). All data are expressed as the mean ± SEM. *p < 0.05, **p < 0.01.

Fig 5: The schematic diagram shows that GLT‐1 plays a key role in T2DM‐induced cognitive impairment in mice after anesthesia/surgery. Due to the downregulation of GLT‐1 expression in hippocampal astrocytes caused by diabetes, the increased excitability of glutamatergic neurons induced by anesthesia/surgery was amplified in the local hippocampal, which led to oxidative stress, apoptosis and finally cognitive dysfunction.