RESUMO
The timing, rate, and quantity of gestational alcohol consumption, collectively referred to here as Maternal Drinking Patterns (MDPs), are of known importance to fetal developmental outcomes. However, few studies have directly evaluated the impact of MDPs on offspring behavior. To do so, we used specialized equipment to record the precise amount and timing of alcohol consumption in pregnant dams, and then characterized MDPs using Principle Component Analysis (PCA). We next tested offspring on behaviors we have previously identified as impacted by prenatal alcohol exposure, and evaluated them where possible in the context of MDPs. Male alcohol exposed mice exhibited longer latencies to fall on the rotarod compared to their controls, which we attribute to a delayed decrease in body weight-gain. This effect was mediated by MDPs within the first 15 min of alcohol access (i.e. alcohol frontloading), where the highest performing male offspring came from dams exhibiting the highest rate of alcohol frontloading. Female alcohol exposed mice displayed reduced locomotor activity in the open field compared to controls, which was mediated by MDPs encompassing the entire drinking session. Surprisingly, total gestational alcohol exposure alone was not associated with any behavioral outcomes. Finally, we observed allodynia in alcohol exposed mice that developed more quickly in males compared to females, and which was not observed in controls. To our knowledge, this report represents the highest resolution assessment of alcohol drinking throughout gestation in mice, and one of few to have identified relationships between specific alcohol MDPs and neurobehavioral outcomes in offspring.
RESUMO
BACKGROUND: Neuroimmune dysregulation from prenatal alcohol exposure (PAE) may contribute to neurological deficits associated with fetal alcohol spectrum disorders (FASD). PAE is a risk factor for developing peripheral immune and spinal glial sensitization and release of the proinflammatory cytokine IL-1ß, which lead to neuropathic pain (allodynia) from minor nerve injury. Although morphine acts on µ-opioid receptors, it also activates immune receptors, TLR4, and the NLRP3 inflammasome that induces IL-1ß. We hypothesized that PAE induces NLRP3 sensitization by morphine following nerve injury in adult mice. METHODS: We used an established moderate PAE paradigm, in which adult PAE and non-PAE control female mice were exposed to a minor sciatic nerve injury, and subsequent allodynia was measured using the von Frey fiber test. In control mice with standard sciatic damage or PAE mice with minor sciatic damage, the effects of the NLRP3 inhibitor, MCC950, were examined during chronic allodynia. Additionally, minor nerve-injured mice were treated with morphine, with or without MCC950. In vitro studies examined the TLR4-NLRP3-dependent proinflammatory response of peripheral macrophages to morphine and/or lipopolysaccharide, with or without MCC950. RESULTS: Mice with standard sciatic damage or PAE mice with minor sciatic damage developed robust allodynia. Blocking NLRP3 activation fully reversed allodynia in both control and PAE mice. Morphine paradoxically prolonged allodynia in PAE mice, while control mice with minor nerve injury remained stably non-allodynic. Allodynia resolved sooner in nerve-injured PAE mice without morphine treatment than in morphine-treated mice. MCC950 treatment significantly shortened allodynia in morphine-treated PAE mice. Morphine potentiated IL-1ß release from TLR4-activated PAE immune cells, while MCC950 treatment greatly reduced it. CONCLUSIONS: In female mice, PAE prolongs allodynia following morphine treatment through NLRP3 activation. TLR4-activated PAE immune cells showed enhanced IL-1ß release with morphine via NLRP3 actions. Similar studies are needed to examine the adverse impact of morphine in males with PAE. These results are predictive of adverse responses to opioid pain therapeutics in individuals with FASD.
