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. Key Words: Smell-G protein-Protein kinesis-Protein foldingMembrane trafficking. J. Neurochem. 72, 2301Neurochem. 72, -2311Neurochem. 72, (1999.The cloning of mammalian olfactory receptors (Buck and Axel, 1991) provided molecular tools that helped answer several significant questions about the olfactory system. We have learned that expression of most receptors is constrained to one of four zones within the epithelium, that the olfactory receptor neurons expressing a given receptor project to only two ipsilateral glomeruli, that the receptor participates in the targeting of the olfactory receptor neuron axons to glomeruli, that receptors can be detected at the plasma membrane in olfactory cilia, and that a single mammalian olfactory receptor neuron probably expresses only one olfactory receptor (Strotmann et al

The continuous replacement of cells in the spiny lobster olfactory organ depends on proliferation of new cells at a specific site, the proximal proliferation zone (PPZ). Using representational difference analysis of cDNA, we identified transcripts enriched in the PPZ compared to the mature zone (MZ) of the organ. The 12 clones identified included four novel sequences, three exoskeletal proteins, a serine protease, two protease inhibitors, a putative growth factor, and a sequence named PET-15 that has similarity to antimicrobial proteins of the crustin type. PET-15 mRNA was only detected in epithelial cells. It was abundant in all epithelial cells of the PPZ, but was only detected in the MZ at sites of damage to the olfactory organ. PET-15 mRNA was increased by types of damage that are known to induce proliferation of new olfactory sensory neurons in the olfactory organ. It increased in the PPZ after partial ablation of the olfactory organ and in the MZ after shaving of aesthetasc sensilla. These ipsilateral effects were mirrored by smaller increases in the undamaged contralateral olfactory organ. These contralateral effects are most parsimoniously explained by the action of a diffusible signal. Because epithelial cells are the source of proliferating progenitors in the olfactory organ, the same diffusible signal may stimulate increases in both cellular proliferation and PET-15 mRNA. The uniformity of expression of PET-15 in the PPZ epithelium suggests that the epithelial cells that give rise to new olfactory sensory neurons are a subset of cells that express PET-15.

The role of the sensory neuropeptide calcitonin gene-related peptide (CGRP) was studied in preterm and term neonates with sepsis and shock. CGRP levels in blood were measured by RIA. The identity of immunoreactive CGRP (irCGRP) in adult and infant human blood was confirmed by reverse phase-HPLC. CGRP levels were analyzed in a total of 189 samples (95 from cord blood and 94 from neonates). The gestational ages ranged from 24 to 43 wk, and the birth weights ranged from 520 to 4445 g. Cord samples were collected immediately after delivery and infant blood samples were collected within 12 h of birth. Samples were coded, and the data were assigned to groups after determination of CGRP levels. There was a weight- and gestation-dependent increase in irCGRP in the newborn population. The direct correlation of circulating CGRP with ascending birth weight and gestation may have significance in the development of the fetus. Infants with and without certain complications were grouped in 500-g intervals. CGRP levels in cord blood were significantly elevated when certain stressful situations existed in the mother. These included culture-positive chorioamnionitis, placental abruption, and severe preeclampsia. There was a similar elevation in CGRP in patient blood in infants with culture-positive sepsis and/or shock with blood pressure <2 SD from the mean for corresponding gestation. CGRP levels did not differ between male and female infants and did not appear to be influenced by type of delivery (vaginal versus cesarean section). There was no significant difference in CGRP level between cord and patient blood in preterm neonates, but at term gestation cord blood levels were slightly higher than those in the patient blood. These results suggest that inflammation and hemodynamic imbalance (e.g. shock) are associated with increased in CGRP levels in the circulation in neonates. Future studies will focus on the biologic effects of elevated CGRP during neonatal complications and will examine the utility of CGRP measurement for diagnosis and treatment of disease in preterm infants.

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