Prophylactic ketamine alters nucleotide and neurotransmitter metabolism in brain and plasma following stress

Abstract

Recently, we have shown that ketamine given prior to stress exposure protects against the development of depressive-like behavior in mice. These data suggest that it may be possible to prevent the induction of affective disorders before they develop by administering prophylactic pharmaceuticals, a relatively nascent and unexplored strategy for psychiatry. Here, we performed metabolomics analysis of brain and plasma following prophylactic ketamine treatment in order to identify markers of stress resilience enhancement. We administered prophylactic ketamine in mice to buffer against fear expression. Following behavioral analyses, untargeted metabolomic profiling was performed on both hemispheres of the prefrontal cortex (PFC) and the hippocampus (HPC), and plasma. We found that prophylactic ketamine attenuated learned fear. Eight metabolites were changed in the PFC and HPC upon ketamine treatment. Purine and pyrimidine metabolism were most significantly changed in the HPC, PFC, and, interestingly, plasma of mice two weeks after prophylactic administration. Moreover, most precursors to inhibitory neurotransmitters were increased whereas precursors to excitatory neurotransmitters were decreased. Strikingly, these long-term metabolomic changes were not observed when no stressor was administered. Our results suggest that prophylactic treatment differentially affects purine and pyrimidine metabolism and neurotransmission in brain and plasma following stress, which may underlie the long-lasting resilience to stress induced by a single injection of ketamine. These data may provide novel targets for prophylactic development, and indicate an interaction effect of prophylactic ketamine and stress. To our knowledge, this is the first study that identifies metabolomic alterations and biomarker candidates for prophylactic ketamine efficacy in mice.

Fig. 1

Prophylactic ketamine buffers the fear response and alters a significant number of metabolites in the brain. a Experimental design. b Prophylactic ketamine does not alter CFC training behavior as measured by freezing. c Prophylactic ketamine administration decreases freezing behavior upon context re-exposure when compared with prophylactic saline administration. (n = 9–10 male mice per group). Error bars represent ± SEM. *p < 0.05. Sal saline, K ketamine, CFC contextual fear conditioning, RE re-exposure, HPC hippocampus, PFC prefrontal cortex, Sac sacrifice, QToF quadrupole-time-of-flight, LCMS liquid chromatography mass spectrometry

Fig. 2

Prophylactic ketamine significantly alters metabolites in the PFC and HPC following stress. a A significant number of metabolites were changed in both brain regions and in both hemispheres. A total number of eight metabolites were changed in both brain regions and in both hemispheres. b A pathway analysis of changed metabolites in the PFC. Purine metabolism; phenylalanine, tyrosine, and tryptophan metabolism; and phenylalanine metabolism are most significantly changed in the PFC following prophylactic ketamine administration. c A pathway analysis of changed metabolites in the HPC. Purine metabolism; alanine, aspartate, and glutamate metabolism; and glutamine and glutamate metabolism are most significantly changed in the HPC following prophylactic ketamine administration. d A heat map of metabolites changed in the PFC. e A heat map of metabolites changed in the HPC. (n = 9–10 male mice per group). HPC hippocampus, PFC prefrontal cortex, ATP adenosine triphosphate, NADH nicotinamide adenine dinucleotide, UTP uridine triphosphate, GDP guanosine diphosphate, GTP guanosine triphosphate, AMP adenosine monophosphate, IMP inosine monophosphate, dUDP deoxyuridine-diphosphate, 13-HOTE 13-OH-9Z,11E,15Z-octadecatrienoic acid, NAD nicotinamide adenine dinucleotide, CDP cytidine diphosphate, ADP adenosine diphosphate

Fig. 3

Prophylactic ketamine significantly alters purine metabolism in the PFC and HPC following stress. Purine precursors are significantly decreased and nucleotides are significantly increased in both hemispheres of the a–g PFC and h–l HPC following prophylactic ketamine administration. (n = 9–10 male mice per group). Error bars represent ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001. Sal saline, K ketamine

Fig. 4

Prophylactic ketamine significantly alters pyrimidine metabolism in the PFC and HPC following stress. a–e Pyrimidine metabolites are significantly altered in both hemispheres of the PFC following prophylactic ketamine administration. f–h Pyrimidine metabolites are significantly altered in both hemispheres of the HPC following prophylactic ketamine administration. i, j The amount of dUDP in both hemispheres of the PFC, but not the HPC, is positively correlated with freezing levels upon context re-exposure in CFC in mice administered prophylactic ketamine and stress. (n = 9–10 male mice per group). Error bars represent ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001. Sal saline, K ketamine, PFC prefrontal cortex, HPC hippocampus, dUDP deoxyuridine-diphosphate

Fig. 5

Prophylactic ketamine significantly alters purine and pyrimidine metabolism in plasma following stress. a A pathway analysis of changed metabolites in the plasma. Purine metabolism; pyrimidine metabolism; and the TCA cycle were most significantly changed following prophylactic ketamine administration. b A heat map of changed metabolites in the plasma. c–l Purine metabolites significantly increased in the plasma following prophylactic ketamine administration. m–q Pyrimidine metabolites significantly increased in the plasma following prophylactic ketamine administration. (n = 9–10 male mice per group). Error bars represent ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001. TCA tricarboxylic acid, Glu glutamate, Asp aspartate, GABA gamma-aminobutyric acid, Ser serine, 5-HTP 5-hydroxytryptophan, PEP phosphoenolpyruvic acid, TPP thiamine pyrophosphate, AMP adenosine monophosphate, Ade adenosine, MCA monochloroacetic acid, CMP cytidine monophosphate, GSSG glutathione disulfide, PE phosphatidylethanolamine, 5′-UMP 5′-uridine monophosphate; RLA, R-(+)-enantiomer lipoic acid, Ino inosine, GMP guanosine monophosphate, 3-PGA 3-phosphoglyceric acid, ATP adenosine triphosphate, NANA N-acetylneuraminic acid, SAMe S-adenosyl methionine, GDP guanosine diphosphate, PRPP 5-phospho-alpha-D-ribosyl 1-pyrophosphate, TRA tiaramide, 5′-UDP 5′-uridine diphosphate, CYSSG cysteine-glutatione disulfide