Maternal Intrauterine Inflammation Induces Kynurenine Pathway Activation and Leads to Decreased Cortical Serotonin In the Newborn Rabbit Brain: Implications for Maternal Infection and Autism

Background:  

Maternal intrauterine infection and inflammation have been implicated in neurodevelopmental disorders, including autism spectrum disorders. Activation of the kynurenine pathway of tryptophan metabolism may play a crucial role in the development of these disorders, by causing serotonin depletion and increased production of neurotoxic kynurenine metabolites in the fetal and neonatal brain.

Objectives:  

The objective of this study was to determine if maternal endotoxin administration is associated with increased expression of kynurenine pathway enzymes and formation of neurotoxic kynurenine metabolites, decreased cortical serotonin and increased pro-inflammatory microglia in the periventricular regions of the newborn rabbit brain.

Methods:  

All animal procedures were approved by the institutional animal care and use committee. Pregnant New Zealand White rabbits were injected with saline (control-saline group) or 20µg/kg of E.Coli endotoxin (endotoxin group) along the length of the uterus at 28 days gestation. A third group comprised of rabbits that had no intervention (control) (N=4 per group). Newborn rabbits (denoted entotoxin-kits, control saline kits and control kits) were delivered spontaneously at term (31 days gestation). The newborn kits underwent PET imaging with alpha[¹¹C]methyl-L-tryptophan (AMT) to evaluate tryptophan metabolism in vivo on day 1 of life. Following the imaging, the kits were euthanized and rabbit brains processed for (1) serotonin concentration in the cortex and hippocampus, (2) concentration of kynurenine metabolites quinolinic acid (QA), kynurenic acid (KA) and 3-hydroxykynurenine (3HK) by HPLC (3) mRNA expression of the kynurenine enzymes indoleamine 2,3 dioxygenase (IDO) and kynurenine 3 monooxygenase (KMO) (3) immunohistochemistry for presence of activated microglia indicated by CD11b staining and for serotonin immunoreactivity in the cortex.

Results:  

Decrease in standard uptake value for AMT and decrease in serotonin concentration was noted in the frontal and parietal cortices of endotoxin kits when compared to controls (x² = 43.24, df = 1, p<0.001 for endotoxin vs. both control groups). In addition, a significant decrease in cortical serotonin concentration  (Mean±SD= 0.188±0.042 ng/ml for endotoxin, 0.546±0.049 ng/ml for control and 0.586±0.002 ng/ml for control-saline; p<0.001 for endotoxin vs controls) and decrease in serotonin immunoreactive fibers was measured in the somatosensory cortex. A significant increase in formation of the neurotoxic metabolite QA was noted in the brain of endotoxin kits (2646 ± 21 pg/g of tissue in endotoxin vs. 6 ± 3 pg/g in controls; p<0.001) along with a 62% increase in expression of KMO, the upstream enzyme for QA formation. This was associated with an increase in CD11b expression in the periventricular region of the endotoxin kit brain indicating an increase in pro-inflammatory microglial phenotype. Increased 3HK staining was also noted in this region in the endotoxin kits.

Conclusions:  

These results indicate that maternal intrauterine endotoxin exposure is associated with increased tryptophan metabolism by the kynurenine pathway, decreased cortical serotonin and increased formation of neurotoxic metabolites. This model shows potential mechanisms by which intrauterine infection/inflammation may produce brain injury associated with autism and other neurodevelopmental disorders in the neonate.