Prenatal lipopolysaccharide does not accelerate progressive dopamine neuron loss in the rat as a result of normal aging.

We previously demonstrated that in utero exposure to the bacteriotoxin lipopolysaccharide (LPS) led to the birth of rat pups with fewer than normal dopamine (DA) neurons. These animals exhibited significant neuroinflammation in the nigrostriatal pathway creating the possibility that they could exhibit further, progressive DA neuron loss over their lives. To study this possibility, we injected gravid female rats i.p. at 10,000 endotoxin units (EUs) of LPS per kg or saline at embryonic (E) day 10.5 and assigned pups to sacrifice groups at 4, 14 and 17 months such that littermates were sacrificed at each end point. The effects of prenatal LPS on DA cell counts and striatal DA were significantly reduced relative to controls whereas DA activity and numbers of activated microglia (OX-6ir cell) were statistically increased. However, the progressive DA neuron loss was parallel to that of the controls suggesting that prenatal LPS does not produce an accelerated rate of DA neuron loss. Interestingly, locomotor activity was increased after 3 months in animals exposed to LPS prenatally, but by 16 months, was significantly reduced relative to controls. Additionally, animals exposed to LPS prenatally exhibited Lewy body-like inclusions that were first seen in 14 month old animals. These data broadly support previous studies demonstrating that prenatal exposure to LPS, as frequently occurs in humans as part of Bacterial Vaginosis, leads to the birth of animals with fewer than normal DA neurons. The progressive DA neuron loss seen in these animals is, however, primarily a result of normal aging.

Progressive dopamine neuron loss following supra-nigral lipopolysaccharide (LPS) infusion into rats exposed to LPS prenatally.

Toxin-induced animal models of Parkinson's disease (PD) exhibit many of the same neuroinflammatory changes seen in patients suggesting a role for inflammation in DA neuron loss. Yet, despite this inflammation, the progressive loss of DA neurons that characterizes PD is rarely seen in animals. We infused lipopolysaccharide (LPS) or saline into 7-month-old rats that had been exposed to LPS or saline prenatally and assessed them for DA neuron loss and inflammatory measures (interleukin 1 beta, tumor necrosis factor-alpha, glutathione, and activated microglia) over a period of 84 days to examine the role of pre-existing inflammation in progressive DA neuron loss. LPS infusion into both prenatal treatment groups produced neuroinflammation during the 14 days of LPS infusion that subsequently reverted toward normal over the next 70 days. In animals with pre-existing inflammation (i.e., prenatal LPS), however, the acute changes seen were attenuated, but took much longer to return to normal suggesting a prolonged inflammatory response. These inflammatory changes were consistent with the greater acute DA neuron loss seen in the prenatal saline controls and the progressive DA neuron loss seen only in the animals exposed to LPS prenatally. Interestingly, both prenatal treatment groups exhibited increases in microglia over the entire 84-day course of the study. These data suggest that pre-existing neuroinflammation prolongs the inflammatory response that occurs with a second toxic exposure, which may be responsible for progressive DA neuron loss. This provides further support for the "multiple hit" hypothesis of PD.

Rotenone potentiates dopamine neuron loss in animals exposed to lipopolysaccharide prenatally.

We previously demonstrated that treating gravid female rats with the bacteriotoxin lipopolysaccharide (LPS) led to the birth of offspring with fewer than normal dopamine (DA) neurons. This DA neuron loss was long-lived and associated with permanent increases in the pro-inflammatory cytokine tumor necrosis factor alpha (TNFalpha). Because of this pro-inflammatory state, we hypothesized that these animals would be more susceptible to subsequent exposure of DA neurotoxins. We tested this hypothesis by treating female Sprague-Dawley rats exposed to LPS or saline prenatally with a subtoxic dose of the DA neurotoxin rotenone (1.25 mg/kg per day) or vehicle for 14 days when they were 16 months old. After another 14 days, the animals were sacrificed. Tyrosine hydroxylase-immunoreactive (THir) cell counts were used as an index of DA neuron survival. Animals exposed to LPS prenatally or rotenone postnatally exhibited a 22% and 3%, respectively, decrease in THir cell counts relative to controls. The combined effects of prenatal LPS and postnatal rotenone exposure produced a synergistic 39% THir cell loss relative to controls. This loss was associated with decreased striatal DA and increased striatal DA activity ([HVA]/[DA]) and TNFalpha. Animals exposed to LPS prenatally exhibited a marked increase in the number of reactive microglia that was further increased by rotenone exposure. Prenatal LPS exposure also led to increased levels of oxidized proteins and the formation of alpha-Synuclein and eosin positive inclusions resembling Lewy bodies. These results suggest that exposure to low doses of an environmental neurotoxin like rotenone can produce synergistic DA neuron losses in animals with a preexisting pro-inflammatory state. This supports the notion that Parkinson's disease (PD) may be caused by multiple factors and the result of "multiple hits" from environmental toxins.

In utero bacterial endotoxin exposure causes loss of tyrosine hydroxylase neurons in the postnatal rat midbrain.

We investigated whether in utero exposure to the Gram(-) bacteriotoxin lipopolysaccharide (LPS) induces dopamine (DA) neuron loss in rats. The proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) kills DA neurons and is elevated in the brains of patients with Parkinson's disease (PD). LPS is a potent inducer of TNF-alpha, and both are increased in the chorioamniotic environment of women who have bacterial vaginosis (BV) during pregnancy, suggesting that BV might interfere with the normal development of fetal DA neurons. Gravid female rats were injected intraperitoneally with either LPS or normal saline at embryonic day 10.5 and their pups were killed at postnatal day 21. The brains of the pups were assessed for DA and TNF-alpha levels and DA cell counts in the mesencephalon using tyrosine hydroxylase immunoreactive (THir) cells as a DA neuron marker. Prenatal LPS exposure significantly reduced striatal DA (29%) and increased DA activity (72%) as well as TNF-alpha (101%). Stereological cell counts in the mesencephalon were also significantly reduced (27%) by prenatal LPS exposure. Prenatal exposure to LPS, as might occur in humans with BV, produces a significant loss of THir cells in rats that is still present 33 days following a single injection of LPS. Since this cell loss is well past the normal phase of DA neuron apoptosis that occurs in early postnatal life, rats so exposed may have a permanent loss of DA neurons, suggesting that prenatal infections may represent risk factors for PD.