Ketamine ameliorates depressive-like behaviors in mice through increasing glucose uptake regulated by the ERK/GLUT3 signaling pathway

Abstract

To investigate the effects of ketamine on glucose uptake and glucose transporter (GLUT) expression in depressive-like mice. After HA1800 cells were treated with ketamine, 2-[N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino]-2-Deoxyglucose (2-NBDG) was added to the cells to test the effects of ketamine on glucose uptake, production of lactate, and expression levels of GLUT, ERK1/2, AKT, and AMPK. Adult female C57BL/6 mice were subjected to chronic unpredictable mild stress (CUMS), 27 CUMS mice were randomly divided into the depression, ketamine (i.p.10 mg/kg), and FR180204 (ERK1/2 inhibitor, i.p.100 mg/kg) + ketamine group. Three mice randomly selected from each group were injected with ¹⁸F-FDG at 6 h after treatment. The brain tissue was collected at 6 h after treatment for p-ERK1/2 and GLUTs. Treatment with ketamine significantly increased glucose uptake, extracellular lactic-acid content, expression levels of GLUT3 and p-ERK in astrocytes and glucose uptake in the prefrontal cortex (P < 0.05), and the immobility time was significantly shortened in depressive-like mice (P < 0.01). An ERK1/2 inhibitor significantly inhibited ketamine-induced increases in the glucose uptake in depressive-like mice (P < 0.05), as well as prolonged the immobility time (P < 0.01). The expression levels of p-ERK1/2 and GLUT3 in depressive-like mice were significantly lower than those in normal control mice (P < 0.01). Ketamine treatment in depressive-like mice significantly increased the expression levels of p-ERK1/2 and GLUT3 in the prefrontal cortex (P < 0.01), whereas an ERK1/2 inhibitor significantly inhibited ketamine-induced increases (P < 0.01).Our present findings demonstrate that ketamine mitigated depressive-like behaviors in female mice by activating the ERK/GLUT3 signal pathway, which further increased glucose uptake in the prefrontal cortex.

Figure 1

Ketamine increases the 2-NBDG uptake, protein levels of GLUT3 and P-ERK1/2, and production of lactate in HA1800 cells. (A) The intracellular fluorescence of 2-NBDG was examined by confocal microscopy. (B) The expression of GLUT1, GLUT3 and GLUT4 were examined by Western blots at 6 h post ketamine treatment with different concentration (0, 10, 25, 50 and 100 μM); (C) Western blots analysis was used to test the levels of P-ERK1/2, P-AKT and P-AMPK at 6 h post ketamine treatment with different concentration (0, 10, 25, 50 and 100 μM); (D) The subcellular location of P-ERK1/2 response to ketamine treatment was detected by immunofluorescent staining; (E) The production of lactate was tested by a lactate detection kit. Vs control, *P < 0.05 and **P < 0.01. Immunoblotting signals were quantified via Image J software.

Figure 2

Effects of ketamine on depressive-like behaviors in CUMS mice. (A) The percentage of the consumption of sucrose-sweetened water in the SPT. (B) The immobility time in the TST. (C) The immobility time in the FST. (D) The latency to feed in the NSF test. NC normal control group, D depression group, K ketamine group, FR ERK1/2 inhibitor (FR180204) group; n = 6. *P < 0.05, **P < 0.01.

Figure 3

Ketamine increases glucose uptake in the prefrontal cortices of depressive-like mice. (A) ¹⁸F-FDG PET CT brain imaging; the more red, the more glucose uptake. (B) The SUV was prefrontal-cortex SUV (normalized to that of the whole-brain SUV). NC normal control group, D depression group, K ketamine group, FR ERK1/2 inhibitor (FR180204) group; n = 3; *P < 0.05, **P < 0.01. Images were analyzed using ASI Pro VM software (Siemens Medical Solutions, USA).

Figure 4

Ketamine upregulates the protein levels of P-ERK1/2 and GLUT3 in the prefrontal cortices. (A) Western-blot detection of related protein expression. (B) Relative expression levels of P-ERK1/2, T-ERK1/2, and GLUT3; NC normal control group, D depression group, K ketamine group, FR: ERK1/2 inhibitor (FR180204) group; n = 6; *P < 0.05, **P < 0.01. Protein levels of P-ERK1/2, T-ERK1/2, and GLUT3 in the prefrontal cortices were quantified by Image J software.

Figure 5

Immunofluorescence micrographs in the prefrontal cortices of depressive-like mice (A,B): Stained with the P-ERK1/2 antibodies (green) and GFAP antibodies (red). (C): Stained with the P-ERK1/2 antibodies (green) and neuron antibodies (red). (D): Stained with the P-ERK1/2 antibodies (green) and GLUT3 antibodies (red). (E): Stained with the P-ERK1/2 antibodies (green) and GLUT3 antibodies (red). NC normal control group, K ketamine group, GFAP astrocyte marker, NeuN neuron marker; scale bar = 25 μM.

Figure 6

The ANLS (Astrocyte-neuron lactate shuttle) model.

Figure 7

Flowchart of experimental setup.