N. S. Rama Krishna

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.

Age- and Gender-Related Trends in the Expression of Glutathione S-Transferases in Human Nasal Mucosa

The cellular expression of α, μ, and π classes of glutathione S-transferases (GSTs) was investigated in human nasal mucosa by means of immunocytochemical techniques. In the olfactory mucosa, immunoreactivity for GST-α was most intense in the acinar cells of the Bowmans glands, with weak immunoreactivity in the supranuclear region of sustentacular cells. Whereas GST-π was localized only in the sustentacular cells, no GST-μ was detected. In the respiratory mucosa, GST-α and GST-π were detected at the brush borders of ciliated columnar epithelial cells. There were age- and gender-related trends in the expression of GST-α, but not GST-π, in the olfactory mucosa. The intensity of immunoreactivity in the olfactory mucosa was decreased in older subjects. The expression of GST-α in the olfactory mucosa of females consistently exhibited greater intensity than that of males at all the ages studied. These differences were not observed in the respiratory mucosa. These results indicate that acinar cells of the Bowmans glands and sustentacular cells are the major sites of phase II biotransformation in the human nasal mucosa.

Glutathione and γ-glutamyl transpeptidase are differentially distributed in the olfactory mucosa of rats

SummaryComponents of the γ-glutamyl cycle, including thiols, glutathione (GSH) and γ-glutamyl transpeptidase (γ-GT), were localized in the nasal mucosae of rats using histochemical and immunohistochemical methods. In olfactory mucosa, thiols were widely distributed, with intense staining in the mucociliary complex (MC), basal cells, acinar cells of Bowmans glands (BG), and olfactory nerve bundles, and with moderate staining in olfactory receptor neurons (ORNs). GSH was localized in MC, BG acinar cells, nerve bundles and, to a lesser extent, in ORNs. γ-GT immunoreactivity was restricted to the MC and to basolateral and apical membranes of BG acinar and duct cells. The basolateral membrane of BG acinar cells, located in close association with blood vessels and connective tissue, showed granule-like immunoreactivity. Inrespiratory mucosa, all three compounds were localized in the MC and acinar cells of respiratory glands (RG). In the MC, γ-GT immunoreactivity was associated primarily with brush borders of ciliated cells. Granular immunoreactivity was also apparent in the supranuclear region of RG acinar cells. These results demonstrate that components of the γ-glutamyl cycle are localized in olfactory and respiratory glands, and that they are secreted into the mucus, where they may mediate perireceptor events such as detoxification and/or solubilization of air-borne xenobiotics, toxicants and odorants.

Epidermal growth factor receptor mRNA and protein are expressed in progenitor cells of the olfactory epithelium

Olfactory receptor neurons are continuously replaced postnatally through the initiation of the division and terminal differentiation of progenitor cells located in the basal layer of the olfactory epithelium. Although the factors that regulate this process in vivo are not known, recent in vitro studies demonstrated that members of the epidermal growth factor (EGF) family including transforming growth factor‐α (TGFα) and EGF are highly potent in promoting the proliferation of progenitor cells, suggesting a role for the EGF receptor (EGFR), which is the molecular receptor for both mitogens. We have examined the expression of EGFR mRNA and protein in the olfactory epithelium by using reverse transcriptase—polymerase chain reaction (RT‐PCR) and Western blot analysis and have examined their cellular localization with in situ RT‐PCR and immunocytochemistry. RT‐PCR and Southern blot analysis demonstrated that EGFR mRNA is expressed in the olfactory mucosa and also in the positive control tissues, kidney and tongue. The 170‐kDa EGFR protein was identified with Western blot analysis in the olfactory epithelium and control tissues. Our results using in situ RT‐PCR localized EGFR mRNA—expressing cells more extensively in the basal cell layer of the epithelium than did the immunocytochemical methods. These results suggest that EGFR mediates the mitogenic effect of TGFα and/or EGF on the quiescent basal cells to initiate the cell cycle.

Amphibian olfactory receptor neurons express olfactory marker protein

Expression of olfactory marker protein (OMP) in olfactory receptor neurons (ORNs) in two amphibians was investigated by immunohistochemical methods. The OMP immunoreactivity was observed in the cilia, apical dendritic knobs, dendrites and somas of ORNs; the axons of ORNs also showed intense immunoreactivity for OMP throughout their course from the olfactory epithelium to the glomerular layer of the olfactory bulb. Seven days after olfactory nerve transection in salamander, the number of OMP-positive ORNs was markedly reduced in the ipsilateral epithelium. The results demonstrate that amphibian ORNs express OMP and confirm its phylogenetic conservation across diverse species.

Differential distribution of β-glutamyl cycle molecules in the vomeronasal organ of rats

Molecules related to the gamma-glutamyl cycle, including thiols, glutathione (GSH) and gamma-glutamyl transpeptidase (gamma-GT) were identified histochemically and immunohistochemically in the vomeronasal organ of neonatal and adult rats. Thiols and GSH were distributed in the mucomicrovillar complex (MMC), vomeronasal receptor neurons and acinar cells of vomeronasal glands (VNG). gamma-GT was localized in the MMC and in the VNG, where it was associated mainly with the luminal surface of the acinar cells and ducts. The VNO of the neonates exhibited higher staining intensities for all compounds than that of the adults. The data indicate that components of the gamma-glutamyl cycle are present in the VNO and that they are secreted into mucus, where they may be associated with perireceptor events including clearance of pheromones and detoxification of xenobiotics.

Human olfactory receptor neurons contain OMP mRNA in their dendritic and axonal processes

The cellular expression of olfactory marker protein (OMP) mRNA and protein was investigated in the olfactory mucosa of humans ranging in age from 26 weeks of gestation to 85 years using in situ hybridization and immunocytochemistry. OMP mRNA and protein were most abundant in the somas of olfactory receptor neurons (ORNs). The hybridization signal over the ORN somal layer was greater in older subjects than in younger ones, reflecting either a higher neuronal density or more OMP mRNA per cell. In contrast, it was significantly lower in subjects with Alzheimers disease when compared with an age-matched control. Characteristics of older subjects were patchiness in the distribution of OMP-expressing ORNs and the occurrence of subepithelial invaginations containing OMP-positive neurons. In addition, a significant hybridization signal was detected in the apical olfactory epithelium containing the dendrites, dendritic knobs, and cilia of ORNS, and over olfactory nerve bundles in the lamina propria, indicating the occurrence of OMP mRNA in dendritic and axonal domains.

Activation of hypothalamic neurons by intraovarian pressure signals in a teleost fish, Clarias batrachus: role of mechanosensitive channels.

Application of intraovarian pressure is known to trigger profound cytomorphological changes in the neurosecretory cells of nucleus preopticus in the teleost Clarias batrachus. These findings indicate the presence of stretch receptors in the ovaries, perhaps equipped with mechanosensitive channels that transduce the stretch signals to be transmitted to the brain. To test the occurrence of the mechanosensitive channels in the ovaries, we administered a range of pharmacological agents (lignocaine, quinidine, tetraethylammonium chloride, ethylene-diaminetetraacetic acid and gadolinium) known to block the mechanosensitive ion channels, in the ovarian lumen prior to the administration of the intraovarian pressure. Pretreatment with the above agents inhibited the response by the nucleus preopticus neurosecretory cells to intraovarian pressure. The results suggest the occurrence of the mechanosensitive channels in the ovaries of teleostean fishes. In terms of function we speculate that the stretch sensory system and the ensuing pathway connecting the ovaries with the hypothalamus might play a role in apprising the brain of the status of ovarian maturity and in the initiation of the spawning reflex.

Differential Expression of Vomeromodulin mRNA in the Nasal Mucosa of Rats During Ontogeny

Vomeromodulin is a 70-kDa glycoprotein that has been identified and cloned from the nasal mucosa of rats [1]. Its primary protein structure and partial amino acid sequence have been determined. The 2.2-kb mRNA encodes the primary protein of 60 kDa that is posttranslationally modified by N-glycosylation to form the 70-kDa glycoprotein. Immunocytochemical studies with antibodies raised against vomeromodulin showed that this protein is localized primarily in the acinar cells of the lateral nasal glands (LNG) located in the lateral wall of the nasal cavity and, to a lesser extent, in the glandular acini of vomeronasal and posterior glands of the nasal septum. Further, these studies also demonstrated that vomeromodulin is abundantly present in the mucus covering the sensory and nonsensory epithelia of the vomeronasal organ. These observations led to the suggestion that this glycoprotein may function as a molecular transporter for pheromones to vomeronasal receptor neurons; hence, it was called a putative pheromone transporter, vomeromodulin [1]. We have reported the expression of vomeromodulin mRNA in glands of the nasal mucosae of postnatal rats [2]. To obtain further insight into its functional properties, we have used in situ hybridization to investigate the expression of the vomeromodulin gene in the nasal mucosa during ontogeny to correlate its expression with that of other molecular markers signifying functional maturation of the nasal chemosensory systems.