Marilyn L. Getchell

Perireceptor and receptor events in vertebrate olfaction

In this article we have summarized the basic information which identifies several key issues in the study of perireceptor and receptor events in vertebrate olfaction. We have emphasized the biophysical and biochemical data which have established a pivotal role for the olfactory mucus in the access of odorants to receptor sites as well as their clearance from the micro-environment. In addition, based on initial reports in the literature, we have postulated that the uptake of odorants by cells in the olfactory epithelium and their subsequent enzymatic degradation is an important mechanism in odorant removal. Hence, the pre- and post-interactive events in vertebrate olfaction play a key role in molecular recognition, sensory transduction and receptor desensitization. Study of the primary events in vertebrate olfaction is an increasingly active area of research in neurobiology. Application of contemporary techniques in cell and molecular biology as well as biochemistry and cellular biophysics is yielding new insights into the process and into establishing new hypotheses to be tested.

Molecular basis of age-associated cytokine dysregulation in LPS-stimulated macrophages

Aged humans and rodents are susceptible to infection with Streptococcus pneumoniae bacteria as a result of an inability to make antibodies to capsular polysaccharides. This is partly a result of decreased production of proinflammatory cytokines and increased production of interleukin (IL)‐10 by macrophages (MΦ) from aged mice. To understand the molecular basis of cytokine dysregulation in aged mouse MΦ, a microarray analysis was performed on RNA from resting and lipopolysaccharide (LPS)‐stimulated MΦ from aged and control mice using the Affymetrix Mouse Genome 430 2.0 gene chip. Two‐way ANOVA analysis demonstrated that at an overall P < 0.01 level, 853 genes were regulated by LPS (169 in only the young, 184 in only the aged, and 500 in both). Expression analysis of systematic explorer revealed that immune response (proinflammatory chemokines, cytokines, and their receptors) and signal transduction genes were specifically reduced in aged mouse MΦ. Accordingly, expression of Il1 and Il6 was reduced, and Il10 was increased, confirming our previous results. There was also decreased expression of interferon‐γ. Genes in the Toll‐like receptor‐signaling pathway leading to nuclear factor‐κB activation were also down‐regulated but IL‐1 receptor‐associated kinase 3, a negative regulator of this pathway, was increased in aged mice. An increase in expression of the gene for p38 mitogen‐activated protein kinase (MAPK) was observed with a corresponding increase in protein expression and enzyme activity confirmed by Western blotting. Low doses of a p38 MAPK inhibitor (SB203580) enhanced proinflammatory cytokine production by MΦ and reduced IL‐10 levels, indicating that increased p38 MAPK activity has a role in cytokine dysregulation in the aged mouse MΦ.

Immunolocalization of two cytochrome P450 isozymes in rat nasal chemosensory tissue

The NMa and NMb isoforms of cytochrome P450 enzymes are expressed in three nasal chemosensory organs: the olfactory, septal and vomeronasal mucosae. The NMa isoform is widely distributed throughout the nasal mucosa whereas the NMb isoform is present primarily in the chemosensory mucosae. The localization of cytochromes P450 demonstrates that sustentacular cells in the olfactory and septal epithelia, the mucus of the vomeronasal organ and the acinar cells of glands in the lamina propria of all three chemosensory systems engage in xenobiotic metabolism and participate in odorant/pheromone clearance, a perireceptor process associated with chemosensory transduction.

Immunohistochemical Localization of a Cytochrome P-450 Isozyme in Human Nasal Mucosa: Age-Related Trends

Immunoperoxidase staining with an antibody to cytochrome P-450 (NMa) was used to investigate the localization of this isozyme in the human nasal mucosa. Olfactory mucosa was identified by staining of olfactory receptor cells with an antibody to olfactory marker protein. Immunoreactivity to NMa was localized in sustentacular cells in the olfactory epithelium, and in Bowmans gland acinar cells and vascular endothelial cells in the lamina propria. In the respiratory mucosa, ciliated epithelial cells, as well as serous gland acinar cells and vascular endothelial cells in the lamina propria, were immunoreactive for this isozyme. An age-related decrement in the intensity and extent of immunoreactivity within these cells was noted in nasal tissue from patients 60 years of age and over when compared with that of patients under 60 years of age. These results identify sites of xenobiotic metabolism or activation in human nasal mucosa.

Vomeronasal epithelial cells of the adult human express neuron-specific molecules.

IMMUNOHISTOCHEMICAL localization of three molecular markers, neuron-specific enolase (NSE) and protein gene product (PGP) 9.5 for neurons and neuroendocrine cells, and olfactory marker protein (OMP) for olfactory receptor neurons (ORNs) was investigated in the vomeronasal epithelium (VNE) of adult humans. NSE- and PGP 9.5-immunoreactive cells were identified in the VNE. ORNs in the olfactory epithelium of approximately age-matched controls were immunoreactive for the three markers. Most NSE-immunoreactive cells in the VNE were bipolar and similar in shape to the NSE- and PGP 9.5-immuno-reactive ORNs. The results indicate that the adult human VNE contains cells expressing two molecular markers characteristic of neurons and that these cells bear a striking morphological similarity to ORNs.

Localization of substance P and neurofilament immunoreactive fibers in the lumbar facet joint capsule and supraspinous ligament of the rabbit

An indirect immunofluorescence method was utilized to identify substance P-like immunoreactive (SPLI) and neurofilament protein immunoreactive (NFIR) fibers in the lumbar facet joint capsule and supraspinous ligament of the rabbit. The results demonstrated a large population of NFIR fibers, indicating that these tissues are richly innervated, and a smaller population of SPLI fibers. In some fibers, neurofilament protein and substance P were colocalized. The data suggest that the facet joint capsule and the supraspinous ligament contain SPLI nociceptive fibers that could be a source of low back pain.

Spectrophotometric determination of cation concentrations in olfactory mucus

Spectrophotometric techniques were used to determine the concentrations of Na+, K+ and Ca2+ in the olfactory mucus of frogs. The mean concentrations in mEq/l were: [Na+], 52.7 +/- 4.1; [K+], 10.6 +/- 1.9 and [Ca2+], 10.7 +/- 1.7. Topical application of the odorant cineole was associated with statistically significant increases in [Na+] and [Ca2+]; the secretagogues methacholine and isoproterenol induced transient increases in [Na+]. Cineole and methacholine caused sustained increases in [Na+]/[K+] from the control value of 5:1, while isoproterenol caused a transient increase followed by a decline. The results indicate that the cation concentrations in olfactory mucus samples are more similar to those derived from secretory tissue than to those found in the extracellular fluids surrounding typical neural tissue.

Human olfactory receptor neurons express heat shock protein 70: age-related trends

Immunocytochemical methods were used to investigate the cellular distribution and age-related trends in the expression of constitutive and/or inducible forms of heat shock protein (hsp) 70 in the human nasal mucosa of 22 subjects who ranged in age from 16 weeks prenatal to 90 years, including 3 subjects with Alzheimers disease. The olfactory mucosa was characterized by the presence of olfactory marker protein—immunoreactive olfactory receptor neurons. The hsp 70 immunoreactivity was localized in olfactory receptor neurons and the supranuclear region of sustentacular cells in the olfactory epithelium, and in the acinar cells of the Bowmans glands in the lamina propria. A systematic age-related decrement in the expression of hsp 70 immunoreactivity was observed in the olfactory receptor neurons. This trend was not apparent in sustentacular cells and Bowmans glands. A marked decrement in hsp 70 immunoreactivity was also noted in the olfactory receptor neurons of subjects with Alzheimers disease when compared to age-matched controls. These results suggest that the age-dependent decrease in hsp 70 in olfactory receptor neurons of older subjects and those with Alzheimers disease may be attributable to their greater susceptibility to stress.

Odorant and Autonomic Regulation of Secretion in the Olfactory Mucosa

The mechanisms associated with sensory transduction and subsequent membrane events in sensory reception are influenced by activity in ancillary cells within the sense organs. It is well documented that sheath cells in the pacinian corpuscle, Muller cells in the retina, and sustentacular cells in the organ of Corti influence the corresponding sensory receptor cells in somatosensory, visual, and auditory systems. The influence of perireceptor events in chemoreception (i.e., the olfactory, vomeronasal, and gustatory systems) is an emerging area of intense research activity.

Age-dependent phenotypic switching of mast cells in NGF-transgenic mice

Effects of overexpression of nerve growth factor (NGF) on mast cell phenotype and numbers were investigated in nasal and oral mucosae and skin of 3- and 6-week-old transgenic mice in which NGF expression in epithelial basal cells was driven by the keratin-14 promoter. Mast cell phenotypes were identified by Alcian blue/safranin and berberine sulfate histochemistry. In the 3-week-old transgenic mice, NGF overexpression had no effect on phenotype except in tongue, where mast cells exhibited mixed or connective tissue phenotypes compared with the mucosal phenotype in the non-transgenic. In 6-week-old transgenic animals, NGF overexpression resulted in the mucosal phenotype in tissues which contained connective tissue or mixed mast cells in non-transgenics. Mast cell hyperplasia occurred at both ages. NGF effects on mast cell phenotype were age-dependent and involve complex microenvironmental interactions.