Mechanical obstruction of the olfactory cleft reveals differences between orthonasal and retronasal olfactory functions

Following up on recent observations in patients with nasal polyposis (NP), the present study aimed to investigate whether a mechanical obstruction of the anterior olfactory cleft (OC) would produce differential effects on orthonasal and retronasal olfactory functions. To this end, we studied 33 healthy subjects in a randomized trial. Sponges with high content of saline were either placed in the OC or on the respiratory epithelium, such that this was blinded to both subject and observer. The results indicated that orthonasal (P = 0.04) but not retronasal (P = 0.15) olfactory identification ability was lower when the OC was blocked. This confirms the idea that differences between orthonasal and retronasal olfactory functions, as observed in NP patients, are, at least to some degree, due to mechanical obstruction of the anterior portion of the OC. The present data also suggest that mechanical obstruction is a means to induce reversible hyposmia void of side effects which can be performed in a blinded fashion. This might become a valuable model of hyposmia for future investigations.     

Naris occlusion alters transductory protein immunoreactivity in olfactory epithelium

We have recently shown that unilateral naris occlusion (UNO) causes an increase in olfactory marker protein (OMP) immunoreactivity (IR) in mouse olfactory sensory neurons (OSN) from the occluded side of the nasal cavity and a decrease in OMP-IR on the non-occluded side, relative to controls. Given OMP's demonstrated role in olfactory modulation, these OMP-IR changes have been interpreted as a compensatory response by OSNs to odor deprivation on the occluded side and to supernormal exposure to odor on the non-occluded side of the nasal cavity. In the current study, we examined the developmental timing and the regional distribution of this process throughout the nasal cavity using immunocytochemistry. Results demonstrate that OMP-IR diverges in OSNs from the occluded side relative to the non-occluded side of the nasal cavity within eleven days after UNO, with statistically significant differences measurable after 17 days (n=16). We also measured relative levels of the Type 4 phosphodiesterase (PDE4A), another potential olfactory modulator, in nasal cavity tissue from UNO (n=8) and untreated mice (n=9) using western blots and immunocytochemistry. Like OMP, PDE4A-IR increased on the occluded side of the nasal cavity after UNO. Finally, we used immunocytochemistry to assess relative levels of olfactory-specific adenylyl cyclase (ACIII, n=4) and G-protein (Golf, n=2) in OSNs from the occluded and non-occluded sides of the nasal cavity of UNO mice. Following UNO, ACIII but not Golf -IR levels diverged comparing the occluded to the non-occluded sides of the nasal cavity. Taken together, our findings provide support for the previously unknown phenomenon of compensatory responses by OSNs to odor environment.     

Bilateral projection of the olfactory mucosa in the frog

Bulbar potentials wer bilaterally recorded in the frog following electrical stimulation of one olfactory nerve bundle. The general features of the contralateral evoked response were very similar to those of ipsilateral ones. The contralateral response was shown to be produced in situ, not being electronically transmitted from the bulb on the stimulated side. Its response disappeared after section of the olfactory interbulbar adhesion but was not affected by sectioning through either the anterior or the habenular commissure. It was concluded that messages from the neuroreceptors belonging to either the ventral or the dorsal olfactory mucosa on one side, reach both olfactory bulbs.     

Laterality of the olfactory event-related potential response

In experimental practice, odors are commonly applied to only one nostril for recordings of olfactory event-related potentials (OERPs), but the lateralization aspect of the OERP response is unclear regarding both stimulated nostril and cortical topography. The purpose of the present study was to investigate whether stimulated-nostril side affects OERP amplitudes and latencies and whether these potentials indicate lateralization of brain response in healthy, right-handed, young adults. OERPs were recorded from nine electrode sites in response to monorhinal stimulation with amyl acetate in 28 participants. The results showed a general increase in amplitude from frontal to parietal electrode sites (in particular for N1/P3) and generally larger amplitudes on the left hemisphere and midline than on the right hemisphere. There was no main effect of stimulated-nostril side on amplitude. Interactions indicated that N1/P2 amplitude was larger for left- than right-nostril stimulation and larger on the left hemisphere and midline than on the right hemisphere in left-nostril stimulation. No main effect or interactions of stimulated-nostril side over latencies were found and no effects on latencies of sagittal or coronal sites. These findings suggest a general parietal, left-hemisphere predominance in response amplitude to odorous stimulation and imply that either the left or the right nostril may be sufficient for accurate assessment of OERP latency in right-handed, young adults.     

The quantitative relationship between olfactory axons and mitral/tufted cells in developing Xenopus with partially deafferented olfactory bulbs

Partial deafferentation of the olfactory bulb in Xenopus embryos was performed to analyze the effects of afferent innervation on the development of the central olfactory structure. In an attempt to analyze a possible early inductive role of the olfactory axons, one olfactory placode was removed before differentiation of the neural tube began (stages 26–31). A morphological and quantitative analysis was performed on larvae at the onset of metamorphic climax (stage 58). When the single olfactory nerve innervated one side of the rostral telencephalon, a single olfactory bulb developed on that side and no olfactory bulb formed on the contralateral side. When the nerve innervated the midline of the rostral telencephalon, a smaller-than-normal, fused olfactory bulb developed. Partial deafferentation at these early stages resulted in a significant reduction in the number of olfactory axons (to approximately one-half of control values) and a corresponding decrease in the number of mitral/tufted cells (output neurons of the olfactory bulb). To control for possible damage to the neural tube during olfactory-placode removal, a portion of the neural tube directly beneath one of the olfactory placodes was removed in embryos. In these animals, the neural tube regenerated within 24 h and formed a normal olfactory bulb; olfactory axon and mitral/tufted-cell numbers were not significantly different from controls. In conclusion, olfactory-afferent innervation was critical for differentiation of the olfactory bulb, and decreasing the number of olfactory axons resulted in a reduction in the number of output neurons of the olfactory bulb. © 1993 John Wiley & Sons, Inc.     

Cell death of olfactory receptor neurons in a rat with nasosinusitis infected artificially with Staphylococcus

Nasosinusitis is a common cause of acquired hyposmia or anosmia. To study the apoptotic death of olfactory receptor neurons in nasosinusitis, we made an inflammation model in rat infected with STAPHYLOCOCCUS: The histochemical changes in olfactory epithelium were examined using antibodies against protein gene product 9.5 (PGP 9.5), single-strand DNA (ssDNA), Bcl-2 and Bax that might be involved in the apoptosis of olfactory receptor neurons. The thickness of olfactory epithelium and the number of ssDNA-labeled cells were evaluated in each post-treatment group and the results were analyzed by two-way analysis of variance (ANOVA) and post hoc tests. Hematoxylin-eosin staining showed that a severe inflammatory reaction had occurred on the infected side of the nasal cavity and sinus, but not on the non-infected side. However, apoptosis of olfactory receptor neurons occurred on both sides; the apoptosis on the non-infected side started later and behaved like a shadow curve similarly to the infected side. Repeated measures ANOVA showed significant differences of both the thickness of olfactory epithelium (P < 0.0001) and the number of ssDNA-labeled cells (P = 0.0339) in the epithelium between the infected side and non-infected side comparing treatment, time and their interactions. Bcl-2 and Bax were detected only on the infected side in the early stages. Thus, nasosinusitis induced the apoptosis of olfactory receptor neurons. However, the apoptosis occurred not only on the infected side, but also on the non-infected side with no significant inflammation. The Bcl-2/Bax family seems to play an important role in the apoptosis induced by infection, but not in the apoptosis on the non-infected side. The results suggest that mechanisms of apoptosis of olfactory receptor neurons on the infected side may differ from those on the non-infected side.     

Breeding history and olfactory discrimination in the Syrian hamster

The ability of outbred female Syrian hamsters to differentially respond to urine samples obtained from either inbred or outbred Syrian males was examined. The odors obtained from the males were introduced into a Y-maze olfactometer. On the basis of time spent in proximity to the stimuli, estrous females evidenced a preference for the urine of the outbred males. The same females when diestrus exhibited no apparent preferences. It is possible that the ability of females to make such subtle discriminations may impact on mate selection leading to positive assortment.     

Heteromorphic antennules protect the olfactory midbrain from atrophy following chronic antennular ablation in freshwater crayfish

Posthatch larval crayfish (Procambarus clarkii) were unilaterally antennulectomized and maintained in the laboratory for a 6-month period, during which time all regenerating antennular stumps were periodically excised. In another group of animals at a similar developmental stage a heteromorphic antennule was induced on the side of the head ipsilateral to the chronically sectioned normal antennule. After 6 months, all experimental animals were sacrificed and their brains were fixed and sectioned. Computer-aided quantitative measurements were obtained for the volumes of the olfactory lobes on both the experimental and control sides of the brains of both groups of crayfish. In the brains of the crayfish group in which only chronic antennualectomy had been performed, the olfactory lobe ipsilateral to the lesion was reduced in volume by about 80% compared to the olfactory lobe on the control side. In animals in which the normal antennule had been chronically ablated, but which possessed a heteromorphic antennule on the same side, the olfactory lobe on the lesioned side differed in volume from the control side by a mean value of only 28%. We conclude that afferent fibers from a heteromorphic antennule ipsilateral to a chronically lesioned normal antennule can assume some of the central trophic functions of the afferents from the normal antennule; thus, the presence of a heteromorphic antennule offers some measure of protection from the dystrophic effects of chronic ablation of the normal antennule during development and growth in crayfish.     

Decline and recovery of olfactory receptor expression following unilateral bulbectomy

The effects of unilateral olfactory bulb ablation upon the odorant receptor expression were studied during the degeneration/regeneration process in the olfactory epithelium of adult rats. Using the in situ hybridization approach, we compared the time course of decay and recovery of expression for three different receptor subtypes (OR14, OR5, OR124). The number of neurons expressing receptor subtypes dramatically decreased in the olfactory epithelium on the lesioned side and reached a minimum at day 5 postsurgery. A progressive recovery was then observed from day 5 to day 15 postlesion, when a plateau was reached. Noticeable differences in the recovery level of receptor expression were observed according to the zonal patterning: the recovery level for neurons located in the lateral zone reached 70% of the control side value while the recovery levels in the dorsal and medial zones represented 35% and 53% of this value, respectively. Axotomy experiments suggest that zone-specific differences in receptor reexpression reported after bulbectomy might be related to the trophic influence of the olfactory bulb.     

Rhinotopy is disrupted during the re-innervation of the olfactory bulb that follows transection of the olfactory nerve

Re-innervation of the olfactory bulb was investigated after transection of the olfactory nerve using monoclonal antibody RB-8 to assess whether rhinotopy of the primary olfactory projection is restored. In normal animals RB-8 heavily stains the axons, and their terminals, that project from the ventrolateral olfactory epithelium onto glomeruli of the ventrolateral bulb (termed RB-8(+)). In contrast, axons from dorsomedial epithelium are unlabeled (RB-8(-)) and normally terminate in the dorsomedial bulb. Sprague-Dawley rats underwent unilateral olfactory nerve transection and survived for 6 weeks prior to perfusion, sectioning and immunostaining with RB-8. Nerve lesion does not shift the position of the boundary between RB-8(+) and RB-8(-) regions of the epithelium. However, following transection and bulb re-innervation, the distribution of RB-8(+) and RB-8(-) axons is markedly abnormal. First, in all 10 experimental animals RB-8(-) axons displace RB-8(+) axons from anterior glomeruli. Furthermore, the usual target of the RB-8(-) fibers, i.e. the dorsomedial bulb at more posterior levels of the bulb, remains denervated, judging by the lack of staining with antibodies that label axons derived from all epithelial zones. Finally, RB-8(+) fibers invade foreign territory in the dorsolateral bulb on the lesioned side in some cases. The shifts in terminal territory in the bulb after transection contrast with the restoration of the normal zonal patterning of the projection after recovery from methyl bromide lesion, but is consistent with reports of mistargeting by a receptor-defined subset of neurons after transection.     

The number of functional olfactory receptor genes and the relative size of olfactory brain structures are poor predictors of olfactory discrimination performance with enantiomers

The ability of four squirrel monkeys and three pigtail macaques to distinguish between nine enantiomeric odor pairs sharing an isopropenyl group at the chiral center was investigated in terms of a conditioning paradigm. All animals from both species were able to discriminate between the optical isomers of limonene, carvone, dihydrocarvone, dihydrocarveole and dihydrocarvyl acetate, whereas they failed to distinguish between the (+)- and (-)-forms of perillaaldehyde and limonene oxide. The pigtail macaques, but not the squirrel monkeys, also discriminated between the antipodes of perillaalcohol and isopulegol. A comparison of the across-task patterns of discrimination performance shows a high degree of similarity among the two primate species and also between these nonhuman primates and human subjects tested in an earlier study on the same tasks. These findings suggest that between-species comparisons of the relative size of olfactory brain structures or of the number of functional olfactory receptor genes are poor predictors of olfactory discrimination performance with enantiomers.     

Metamorphic remodeling of the primary olfactory projection in Xenopus: Developmental independence of projections from olfactory neuron subclasses

In adult Xenopus, the nasal cavity is divided into separate middle (MC) and principal (PC) cavities; the former is used to smell water-borne odorants, the latter air-borne odorants. Recent work has shown that olfactory neurons of each cavity express a distinct subclass of odorant receptors. Moreover, MC and PC axons project to distinct regions of the olfactory bulb. To examine the developmental basis for this specificity in the olfactory projection, we extirpated the developing MC from early metamorphic (stage 54–57) tadpoles and raised the animals through metamorphosis. In most lesioned animals, the MC partly regenerated. Compared with the unlesioned side, reduction of the region of the glomerular layer of the olfactory bulb receiving MC afferents ranged from 70% to 95%. PC afferents did not occupy regions of the olfactory bulb deprived of MC afferents. These results support a model in which intrinsic cues in the olfactory bulb control the projection pattern attained by ingrowing olfactory axons. © 1997 John Wiley & Sons, Inc. J Neurobiol 32: 213–222, 1997.     

Amitriptyline reduces olfactory neuron adenylyl cyclase activity

Adenylyl cyclase plays an important role in olfactory signaltransduction. Recently, a novel type III adenylyl cyclase hasbeen localized in olfactory neurons (Pfeuffer et al., 1989;Bakalyar and Reed, 1990). Because amitriptyline (AMI), a tricyclicantidepressant, appears to have an inhibitory effect on adenylylcyclase activity in other in other neuronal tissue (Yamaokaet al., 1988; Wong et al., 1991), we measured the effect ofAMI on forskolin-stimulated adenylyl cyclase activity in membranepreparations of olfactory mucosa from adult rats. In the presenceof 5'-guanylyl-imidodiphosphate, AMI (0.5–8.0 µM)inhibited forskolin-stimulated adenylyl cyclase activity ina dose-dependent manner. To determine whether this effect wasspecific for olfactory neurons, as opposed to other cells inthe olfactory epithelium, rats were unilaterally bulbectomizedin order to reduce selectively the number of olfactory neuronson the side ipsilateral to the bulbectomy. In membrane preparationsfrom unilaterally bulbectomized animals we saw significantlylower adenylyl cyclase activity in ipsilateral olfactory mucosa,compared with adenylyl cyclase activity from non-bulbectomizedmucosa. These results indicate that AMI inhibition of adenylylcyclase activity is primariy localized in olfactory neurons.     

Effect of Ninjin-yoei-to (Rensheng-Yangrong-Tang) on olfactory behavior after olfactory nerve transection

Ninjin-yoei-to (NYT), a Japanese traditional medicine, is used to treat athrepsia due to surgery, anorexia, cold constitution, and anemia. There are reports of the effects of NYT on the nervous system; however, there have been no behavioral studies of the effect of NYT on olfactory function. The olfactory system undergoes continuous replacement of sensory neurons. Morphologic and behavioral studies have shown that the olfactory system recovers after bilateral olfactory nerve transection (BNX). However, in the humans, olfactory function does not always recover. In this study, we examined the effect of oral NYT on behavioral recovery after BNX. Fourteen mice were subjected to BNX. The regular diet was mixed with 2% NYT (NYT diet). Mice were separated into two groups; seven mice were fed the regular diet (control group), and seven mice were fed the NYT diet (NYT group). NYT was administered beginning 7 days prior to BNX and continuing for 35 days after BNX. Mice in both groups had free access to food and water. Olfactory function was evaluated by testing each mouse's ability to avoid cotton balls treated with acetic acid. After BNX, mice lost their ability to avoid cotton balls treated with acetic acid. In the control group, the time for behavioral recovery after BNX was 28 days. In the NYT group, the time for behavioral recovery after BNX was 21 days. NYT hastened behavioral recovery after BNX. NYT may have therapeutic benefits for patients with olfactory disorders.     

Experimental evolution of olfactory memory in Drosophila melanogaster

In order to address the nature of genetic variation in learning performance, we investigated the response to classical olfactory conditioning in "high-learning" Drosophila melanogaster lines previously subject to selection for the ability to learn an association between the flavor of an oviposition medium and bitter taste. In a T-maze choice test, the seven high-learning lines were better at avoiding an odor previously associated with aversive mechanical shock than were five unselected "low-learning" lines originating from the same natural population. Thus, the evolved improvement in learning ability of high-learning lines generalized to another aversion learning task involving a different aversive stimulus (shock instead of bitter taste) and a different behavioral context than that used to impose selection. In this olfactory shock task, the high-learning lines showed improvements in the learning rate as well as in two forms of consolidated memory: anesthesia-resistant memory and long-term memory. Thus, genetic variation underlying the experimental evolution of learning performance in the high-learning lines affected several phases of memory formation in the course of olfactory aversive learning. However, the two forms of consolidated memory were negatively correlated among replicate high-learning lines, which is consistent with a recent hypothesis that these two forms of consolidated memory are antagonistic.     

Neural mechanisms in the anterior olfactory nucleus of the rabbit and their possible relevance to localization of an olfactory source

Extracellular recordings were obtained from 40 Anterior OlfactoryNucleus (A.O.N.) units in response to birhinal olfactory stimulations.The units located in the A.O.N. pars dorsalis responded differentlyaccording to the time interval between the onsets of right andleft puffs during birhinal olfactory stimulation. On the contrary,the greater part of the units located in the A.O.N. pars ventraliswas not sensitive to time intervals variations. These data indicatethe ability of some A.O.N. cells to encode temporal characteristicsof an olfactory stimulation.     

An enhanced olfactory marker protein immunoreactivity in individual olfactory receptor neurons following olfactory bulbectomy may be related to increased neurogenesis

Olfactory marker protein (OMP) is a 19-kD acidic protein found throughout the cytoplasm of mature olfactory receptor neurons (ORNs). Its function remains unknown. Following olfactory bulbectomy, the proportion of ORNs mature enough to express OMP declines greatly. However, in the few remaining mature ORNs, it has been observed that the intensity of OMP immunoreactivity (IR) appears to increase over that of ORNs on the unoperated side. We have now investigated this phenomenon quantitatively in rats subjected to unilateral olfactory bulbectomy. Results show that at all postbulbectomy survival periods examined quantitatively (3 days to 6 months), a significant decrease (19–37%) occurs in the transmission of incident light through OMP(+)-ORNs in bulbectomized versus unoperated olfactory epithelium (OE). Further, we also observed a consistent side-to-side difference in OMP IR in control unoperated animals. Possible explanations for these observations and their relation to the still unknown function of OMP are discussed. To test the possibility that OMP might serve a mitogenic role in the OE, recombinant OMP was added to organotypic explant cultures of fetal olfactory mucosa. Addition of OMP resulted in a dose-dependent increase in the density of bromodeoxyuridine-positive cells in the cultures, with a 50% increase occurring at the plateau OMP concentration of 25 nM. © 1998 John Wiley & Sons, Inc. J Neurobiol 34: 377–390, 1998     

Ligand binding specificity of a neutral L-amino acid olfactory receptor

1. The ligand binding specificity of the L-[3H]alanine binding site was investigated in isolated cilia preparations from the olfactory epithelium of channel catfish (Ictalurus punctatus) by competitive binding experiments. 2. Approximately 45 amino acids, derivatives and enantiomers were tested for the ability to compete with radiolabeled L-alanine for common binding sites. 3. Acidic and basic L-amino acids and imino acids did not compete as effectively as L-alanine for the receptor, while long-chain neutral ligands were only partially effective inhibitors of L-alanine binding. 4. D-Alanine and L-alanine derivatives with substituted alpha-amino or carboxyl groups exhibited decreased ability to compete for the receptor, paralleling their lower neurophysiological potency. 5. In combination, the ligand binding results were consistent with previous electrophysiological data in catfish, and suggest the presence of an olfactory receptor site that selectively recognizes short-chain neutral amino acids.     

SNP genotypes of olfactory receptor genes associated with olfactory ability in German Shepherd dogs

To find out the relationship between SNP genotypes of canine olfactory receptor genes and olfactory ability, 28 males and 20 females from German Shepherd dogs in police service were scored by odor detection tests and analyzed using the Beckman GenomeLab SNPstream. The representative 22 SNP loci from the exonic regions of 12 olfactory receptor genes were investigated, and three kinds of odor (human, ice drug and trinitrotoluene) were detected. The results showed that the SNP genotypes at the OR10H1‐like:c.632C>T, OR10H1‐like:c.770A>T, OR2K2‐like:c.518G>A, OR4C11‐like:c.511T>G and OR4C11‐like:c.692G>A loci had a statistically significant effect on the scenting abilities (P < 0.001). The kind of odor influenced the performances of the dogs (P < 0.001). In addition, there were interactions between genotype and the kind of odor at the following loci: OR10H1‐like:c.632C>T, OR10H1‐like:c.770A>T, OR4C11‐like:c.511T>G and OR4C11‐like:c.692G>A (P < 0.001). The dogs with genotype CC at the OR10H1‐like:c.632C>T, genotype AA at the OR10H1‐like:c.770A>T, genotype TT at the OR4C11‐like:c.511T>G and genotype GG at the OR4C11‐like:c.692G>A loci did better at detecting the ice drug. We concluded that there was linkage between certain SNP genotypes and the olfactory ability of dogs and that SNP genotypes might be useful in determining dogs' scenting potential.     

Computation of frequency-to-spatial transform by olfactory bulb glomeruli

A physiological simulation of 2.5% of the input and inhibitory neurons and 25% of the primary mitral/tufted cells in a single mammalian olfactory bulb glomerulus was constructed. This physiological simulation used the integrate-and-fire paradigm with realistic activation curves and synaptic delays. The dendritic integration incorporated non-linear interactive effects of individual cell excitatory and inhibitory post-synaptic potentials (PSPs) from both axodendritic and dendrodendritic synaptic contacts. Refractory periods for granule-cell inhibition of mitral/tufted cell activity lead to relatively fixed-frequency rhythmic activity in the glomerulus, independent of the input frequency from the olfactory nerve. Though the frequency of mitral/ tufted cell firing in bulb was approximately independent of input frequency, the number of cells active in the glomerulus was a roughly-linear function of input frequency to the glomerulus, indicating the mechanism's ability to function as a frequency-to-spatial encoder.