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We previously reported that injection of the Gram (−) bacteriotoxin, lipopolysaccharide (LPS), into gravid females at embryonic day 10.5 led to the birth of animals with fewer than normal dopamine (DA) neurons when assessed at postnatal days (P) 10 and 21. To determine if these changes continued into adulthood, we have now assessed animals at P120. As part of the previous studies, we also observed that the pro-inflammatory cytokine tumor necrosis factor α (TNFα) was elevated in the striatum, suggesting that these animals would be more susceptible to subsequent DA neurotoxin exposure. In order to test this hypothesis, we injected (at P99) 6-hydroxydopamine (6OHDA) or saline into animals exposed to LPS or saline prenatally. The results showed that animals exposed to prenatal LPS or postnatal 6OHDA alone had 33% and 46%, respectively, fewer DA neurons than controls, while the two toxins combined produced a less than additive 62% loss. Alterations in striatal DA were similar to, and significantly correlated with (r2=0.833) the DA cell losses. Prenatal LPS produced a 31% increase in striatal TNFα, and combined exposure with 6OHDA led to an 82% increase. We conclude that prenatal exposure to LPS produces a long-lived THir cell loss that is accompanied by an inflammatory state that leads to further DA neuron loss following subsequent neurotoxin exposure. The results suggest that individuals exposed to LPS prenatally, as might occur had their mother had bacterial vaginosis, would be at increased risk for Parkinson''s disease. [Copyright &y& Elsevier]

Olfactory receptors are difficult to express functionally in heterologous cells. We found that olfactory receptors traffic poorly to the plasma membrane even in cells with neuronal phenotypes, including cell lines derived from the olfactory epithelium. Other than mature olfactory receptor neurons, few cells appear able to traffic olfactory receptors to the plasma membrane. In human embryonic kidney 293 cells and Xenopus fibroblasts, olfactory receptor immunoreactivity overlapped with a marker for the endoplasmic reticulum (ER) but not with markers for the Golgi apparatus or endosomes. Except for the ER, olfactory receptors were therefore absent from organelles normally involved in the plasma membrane trafficking of receptors. Olfactory receptors truncated prior to transmembrane domain VI were expressed in the plasma membrane, however. Co-expression of the missing C-terminal fragment with these truncated receptors prevented their expression in the plasma membrane. Intramolecular interactions between N- and C-terminal domains joined by the third cytoplasmic loop appear to be responsible for retention of olfactory receptors in the ER of heterologous cells. Our results are consistent with misfolding of the receptors but could also be explained by altered trafficking of the receptors.

We have isolated from the olfactory organ of the American lobster (Homarus americanus) two cDNA clones with homology to beta subunits of G proteins. LobGbeta1 contained a complete open reading frame that predicted an amino acid sequence with >80% identity to Gbeta sequences from other species. LobGbeta2 was a fragment of an open reading frame whose predicted amino acid sequence had 65-69% identity to other Gbeta sequences. LobGbeta2 mRNA was not detectable in the brain, eye plus eyestalk, leg, dactyl, olfactory organ, or tail muscle. In contrast, lobGbeta1 was expressed in all these tissues as a single mRNA species of 6.4 kb and a protein of 37 kD. In the brain and olfactory organ, Gbeta immunoreactivity was almost exclusively confined to neurites: the neuropil regions of the brain and the outer dendrites of the olfactory receptor neurons. Coimmunoprecipitation revealed that lobster Gbeta interacted with both Galpha s and Galpha q. LobGbeta1 is likely to be involved in a wide range of signaling events including olfactory transduction and synaptic transmission in the brain.

We have isolated from an American lobster (Homarus americanus) olfactory organ cDNA library a clone, lobG alphaS, with >70% identity to mammalian and arthropod G alphaS sequences. In genomic Southern blots, a fragment of lobG alphaS detected only one band, suggesting the lobsters have a single G alphaS gene. In brain and olfactory organ, lobG alphaS mRNA was expressed predominantly in neurons, including many of the neuronal cell body clusters of the brain. G alphaS protein was also expressed broadly, appearing on western blots as a band of 51.8 kDa in brain, eyestalk, pereiopod, dactyl, tail muscle, olfactory organ, and aesthetasc hairs. These results suggest that lobG alphaS plays a role in a wide variety of signal transduction events. Its presence in the olfactory aesthetasc hairs, which are almost pure preparations of the outer dendrites of the olfactory receptor neurons, and the expression of lobG alphaS mRNA in the olfactory receptor neurons of the olfactory organ indicate that lobG alphaS may mediate olfactory transduction. That virtually all ORNs express lobG alphaS mRNA equally predicts that hyperpolarizing odor responses mediated by cyclic AMP are a property of all lobster olfactory receptor neurons.

We have isolated from an American lobster (Homarus americanus) olfactory organ cDNA library a clone, hG alpha(q), with >80% identity to mammalian and arthropod G alpha(q) sequences. In brain and olfactory organ, hG alpha(q) mRNA was expressed predominantly in neurons, including virtually all the neuronal cell body clusters of the brain. G alpha(q) protein was also expressed broadly, appearing on western blots as a single band of 46 kDa in brain, eyestalk, pereiopod, dactyl, tail muscle, olfactory organ, and aesthetasc hairs. These results suggest that hG alpha(q) plays a role in a wide variety of signal transduction events. Its presence in the olfactory aesthetasc hairs, which are almost pure preparations of the outer dendrites of the olfactory receptor neurons, the expression of a single hG alpha(q) mRNA species (6 kb) in the olfactory organ, and the localization of hG alpha(q) mRNA predominantly in the olfactory receptor neurons of the olfactory organ strongly suggest that one function of hG alpha(q) is to mediate olfactory transduction.

Neuronal expression of the ALZ-50 epitope was investigated in hippocampus and medulla from infants dying of sudden infant death syndrome or known causes (controls). Hippocampal studies include data from 31 infants dying of known causes between 32 weeks' gestation and 16 months postpartum and 46 infants who died of sudden infant death syndrome. The medulla at the level of the mid olivary protuberance was investigated in 22 infants with sudden infant death syndrome and 11 controls matched for age and postmortem interval. Medullary sections were also examined using immunohistochemical methods to demonstrate reactivity to glial fibrillary acidic protein antibody. The density of ALZ-50-immunodecorated neurons in control hippocampus rises from the level observable in utero to a maximum between 1 and 4 months of age and declines thereafter. The density of ALZ-50-immunoreactive neurons in hippocampus is significantly increased in infants with sudden infant death syndrome at all ages. Significant regionally specific increases in the number of ALZ-50-immunoreactive neurons, and glial fibrillary acidic protein-reactive cells were found in sudden infant death syndrome medulla; coincidental increases were observed in only the solitary nucleus. Neurons exhibiting the ALZ-50 epitope may reflect apoptotic neuron death of normal development, and increased numbers of immunoreactive neurons may suggest enhanced neurodegeneration in sudden infant death syndrome.