OBJECTIVE-To determine whether maternal intrauterine endotoxin administration leads to neurobehavioral deficits in newborn rabbits. STUDY DESIGN-PregnantNew Zealand white rabbits were injected with 1ml saline (n=8) or 20μg/kg of lipolysaccharide in saline (LPS) (n=8) into the uterine wall on day 28/31 of gestation. On postnatal day 1, kits [saline (n=30) and LPS (n=18) from 4 consecutive litters] underwent neurobehavioral testing. Neonatal brains were stained for microglial cells and myelin.RESULTS-LPS-group kits were hypertonic and demonstrated significant impairment in posture, righting reflex, locomotion and feeding, along with neuroinflammation indicated by activated microglia, and hypomyelination in the periventricular regions. A greater mortality was noted in the LPS group (16 stillbirths from 3 litters vs. 3 from 1 litter).CONCLUSION-Maternal intrauterine endotoxin administration leads to white matter injury and motor deficits in the newborn rabbit resulting in a phenotype that resembles those found in periventricular leukomalacia and cerebral palsy.

Intrauterine infection can lead to a fetal inflammatory response syndrome that has been implicated as one of the causes of perinatal brain injury leading to periventricular leukomalacia (PVL) and cerebral palsy. The presence of activated microglial cells has been noted in autopsy specimens of patients with PVL and in models of neonatal hypoxia and ischemia. Activated microglial cells can cause oligodendrocyte damage and white matter injury by release of inflammatory cytokines and production of excitotoxic metabolites. We hypothesized that exposure to endotoxin in utero leads to microglial activation in the fetal brain that can be monitored in vivo by 11 positron-emitting ligand that binds peripheral benzodiazepine receptor sites in activated microglia-using small-animal PET. Methods: Pregnant New Zealand White rabbits underwent laparotomy and were injected with 20 and 30 mg/kg of Escherichia coli lipopolysaccharide along the length of the uterus on day 28 of gestation. The pups were born spontaneously at term (31 d) and were scanned using small-animal PET after intravenous administration of 11 C-(R)-PK11195 and by MRI on postnatal day 1. The standard uptake values (SUVs) of the tracer were calculated for the whole brain at 10-min intervals for 60 min after tracer injection. The pups were euthanized after the scan, and brains were fixed, sectioned, and stained for microglial cells using biotinylated tomato lectin. Results: There was increased brain retention of 11 C-(R)-PK11195-as determined by a significant difference in the slope of the SUV over time-in the endotoxintreated pups when compared with that of age-matched controls. Immunohistochemical staining showed dose-dependent changes in activated microglia (increased number and morphologic changes) in the periventricular region and hippocampus of the brain of newborn rabbit pups exposed to endotoxin in utero. Conclusion: Intrauterine inflammation leads to activation of microglial cells that may be responsible for the development of brain injury and white matter damage in the perinatal period.PET with the tracer 11 C-(R)-PK11195 can be used as a noninvasive, sensitive tool for determining the presence and progress of neuroinflammation due to perinatal insults in newborns.

Intrauterine inflammation is known to be a risk factor for the development of periventricular leukomalacia (PVL) and cerebral palsy. In recent years, activated microglial cells have been implicated in the pathogenesis of PVL and in the development of white matter injury. Clinical studies have shown the increased presence of activated microglial cells diffusely throughout the white matter in brains of patients with PVL. In vitro studies have reported that activated microglial cells induce oligodendrocyte damage and white matter injury by release of inflammatory cytokines, reactive nitrogen and oxygen species and the production of excitotoxic metabolites. PK11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinoline carboxamide] is a ligand that is selective for the 18-kDa translocator protein expressed on the outer mitochondrial membrane of activated microglia and macrophages. When labeled with carbon-11, [¹¹C]PK11195 can effectively be used as a ligand in positron emission tomography (PET) studies for the detection of activated microglial cells in various neuroinflammatory and neurodegenerative conditions. In this study, we hypothesized that the magnitude of [¹¹C]-(R)-PK11195 uptake in the newborn rabbit brain, as measured using a small-animal PET scanner, would match the severity of motor deficits resulting from intrauterine inflammation-induced perinatal brain injury. Pregnant New Zealand white rabbits were intrauterinely injected with endotoxin or saline at 28 days of gestation. Kits were born spontaneously at 31 days and underwent neurobehavioral testing and PET imaging following intravenous injection of the tracer [¹¹C]-(R)-PK11195 on the day of birth. The neurobehavioral scores were compared with the change in [¹¹C]PK11195 uptake over the time of scanning, for each of the kits. Upon analysis using receiver operating characteristic curves, an optimal combined sensitivity and specificity for detecting abnormal neurobehavioral scores suggestive of cerebral palsy in the neonatal rabbit was noted for a positive change in [¹¹C]PK11195 uptake in the brain over time on PET imaging (sensitivity of 100% and area under the curve of >0.82 for all parameters tested). The strongest agreements were noted between a positive uptake slope – indicating increased [¹¹C]PK11195 uptake over time – and worsening scores for measures of locomotion (indicated by hindlimb movement, forelimb movement, circular motion and straight- line motion; Cohen’s ĸ >0.75 for each) and feeding (indicated by ability to suck and swallow and turn the head during feeding; Cohen’s ĸ >0.85 for each). This was also associated with increased numbers of activated microglia (mean ratio ± SD of activated to total microglia: 0.96 ± 0.16 in the endotoxin group vs. 0.13 ± 0.08 in controls; p < 0.001) in the internal capsule and corona radiata. Our findings indicate that the magnitude of [¹¹C]PK11195 binding measured in vivo by PET imaging matches the severity of motor deficits ...