Aberrant expression of FBXO22 is associated with propofol-induced synaptic plasticity and cognitive dysfunction in adult mice

Abstract

Recent observation demonstrated that prolonged anesthesia modifies brain synaptic architecture in all ages, including adult. Propofol is the most commonly utilized anesthetics at clinic. Whether repeated administration of propofol modulates cognitive impairment in adults and changes synaptic plasticity remains, however, to be explored. In this study, we first discovered that repeated and prolonged exposure to propofol-induced cognitive impairment in adult rodents. Then, we examined the property of hippocampal primary neurons and slices after propofol treatment in mice, including synaptic protein profile, dendritic spine density, as well as synaptic transmission. We found the distinctive change of the F-box only protein 22 (FBXO22), an F-box E3 ligase, during this process and further explored its role. Knockdown experiments showed the downregulation of FBXO22 restored the changes by propofol treatment on hippocampal primary neurons and attenuated propofol-induced hippocampal dependent cognitive dysfunction. Our results showed that FBXO22 is involved in the regulation of repeated propofol treatment induced changes of synaptic plasticity and cognitive dysfunction in adult mice. Repeated propofol treatment leads to cognitive dysfunction by regulating FBXO22 in adult rodents.

Keywords: FBXO22; general anesthesia; hippocampus; neurocognition; propofol.

Figure 1

Diagram of timeline of experimental procedures. (A) Effects of different doses of propofol anesthesia on spatial learning and memory in mice. 80 mice were randomly allocated to two behavioral experiments: Morris water maze (MWM): 4 groups (control, propofol 25 mg/kg, propofol 50 mg/kg, propofol 100 mg/kg) × 10 mice/group. The novel object recognition test (NORT) and object location test (OLT): 4 groups (the same group allocation as MWM) × 10 mice/group. (B) The effect of FBXO22 regulation on cognitive dysfunction induced by propofol. 120 mice were randomly allocated to three behavioral experiments. MWM: 4 groups (control+FBXO22-SCM, control+FBXO22-shRNA, propofol+FBXO22-SCM, propofol+ FBXO22-shRNA) × 10 mice/group; NORT and OLT: 4 groups (the same group allocation as morris water maze) × 10 mice/group; Contextual fear conditioning (CFC), the same group allocation as MWM: 4 groups×10 mice/group. (C) The schematic time course and experimental designs of spine density model P: postnatal.

Figure 2

Effects of different doses of propofol (25, 50, and 100 mg/kg) anesthesia on spatial learning and memory in mice. (A) The typical trace of mice on the 5th training day. (B) The latency of finding the hidden platform from day 1 to day 5 during the training period. (C) The escape latency in the memory retrieval tests on the 6th day. (D) The typical paths of mice in the memory retrieval tests on the 6th day. (E) The time spent in the target quadrant on the test day. (F) The number of crossing over the previous platform on the test day. (G,L) The schematic diagram and representative paths traveled in NORT and OLT. (H,M) The typical paths of mice in training and test of NORT and OLT. (I,N) The investigation time for novel location in NORT and OLT. (J,K) The total object exploration time in NORT when training and testing. (O,P) The total object exploration time in OLT when training and testing. Data are presented as medians with interquartile range in panel F and are expressed as means ± SEM in other panels, and n = 10 mice per group; *p < 0 0.05, **p < 0 0.01, compared to the control group.

Figure 3

Western blot analysis of FBXO22, NMDAR1 and PSD95 protein level in hippocampus treated with different dose of propofol (10, 25, 50, 100, and 200 mg/kg). The protein level of FBXO22, NMDAR1 and PSD95 was determined by western blot (A) and the bands intensity corresponding to the target protein was analyzed (B–D). Representative blots are shown. Data are expressed as means ± SEM, and n = 3 mice per group; *p < 0.05, **p < 0.01, compared to the control group.

Figure 4

Downregulation of FBXO22 restored the defects of glutamatergic synaptic transmission induced by propofol. (A) Representative image of the mice’s brain slice injected with AAV-FBXO22-shRNA-GFP. (B) The FBXO22 protein level of mice injected with AAV-FBXO22-Scramble-GFP (control) and AAV-FBXO22-shRNA-GFP (FBXO22). (C) Representative sEPSC recordings for the control or propofol (100 mg/kg)treated, AAV-FBXO22-SCM-injected and AAV-FBXO22-shRNA-injected mice. (D,E) The quantitative analysis of mEPSC frequency and amplitude. Data are expressed as means ± SEM, and n = 13 neurons from three mice per group; *p < 0.05, **p < 0.01.

Figure 5

Downregulation of FBXO22 restored the defects of spine density in hippocampal primary neurons induced by propofol. (A) Representative images of the dendrites of primary hippocampal neurons transfected with either FBXO22-SCM or FBXO22-shRNA. Scale bars: upper panels, 25 μm; lower panels, 10 μm. (B–D) The summary of spine density, neck length, and head width. Data are expressed as means ± SEM, and n = 13 neuron from three mice per group; *p < 0.05, **p < 0.01.

Figure 6

Downregulation of FBXO22 expression partially restored learning and memory ability induced by propofol (100 mg/kg). (A) The typical paths of mice on the 5th training day. (B) The latency of finding the hidden platform from day 1 to day 5 during the training period (Control+ FBXO22-SCM vs. Propofol+ FBXO22-SCM: *p < 0.05, **p < 0.01; Propofol+ FBXO22-SCM vs. Propofol+ FBXO22-shRNA: #p < 0.05). (C) The escape latency in the memory retrieval tests on the 6th day. (D) The typical paths of mice in the memorial retrieval test on the test day. (E) The time spent in the target quadrant in the memory retrieval tests. (F) The number of crossing over the previous platform position on the test day. (G,L) The schematic diagram and representative paths traveled in NORT and OLT. (H,M) The typical paths of mice in training and test of NORT and OLT. (I,N) The investigation time for novel location in NORT and OLT. (J,K) The total object exploration time in NORT when training and testing. (O,P) The total object exploration time in OLT when training and testing. Data are presented as medians with interquartile range in panel F and are expressed as means ± SEM in other panels, and n = 10 mice/ group. *p < 0.05, **p < 0.01.

Figure 7

Downregulation of FBXO22 expression partially restored contextual memory induced by propofol (100 mg/kg). (A) The diagrammatic sketch of fear conditioning and context discrimination. (B) The baseline levels of fear behavior (freezing time)among four groups. (C) The freezing time during testing. Data are expressed as means ± SEM, and n = 10 mice per group; *p < 0.05, **p < 0.01.