Demonstration of direct olfactory projections in the hypothalamus, the mesencephalon and the metencephalon of Microcebus murinus (Primate, Lemurian)

The efferent projections of the olfactory bulb in Microcebus murinus were identified after transection of the olfactory peduncle and after the revelation of the degenerating fibers by different silver staining methods. Total and partial sections have allowed demonstrating the importance of the two olfactory tracts in the olfactory projection areas. Degenerated fibers and endings were evidence not only in the different telencephalic regions, as classically known, but also in various hypothalamic nuclei (lateral hypothalamus, the suprachiasmatic, posterior supraoptic and mammillary nuclei and in the median eminence) and in several mesencephalic and metencephalic nuclei (ventral tegmental area, interpeduncular and raphe nuclei, and locus coeruleus). In all these structures the degenerate fibers were seen on both sides. The olfactory projections appeared not to be limited to the telencephalic areas. Moreover, the olfactory bulbs seem to be directly connected especially with the vegetative and integrative areas localized in the hypothalamus and the brainstem and particularly with the major aminergic nuclei that play an essential role in the neurovegetative, neuroendocrine and behavioral regulations.     

The rodent accessory olfactory system

The accessory olfactory system contributes to the perception of chemical stimuli in the environment. This review summarizes the structure of the accessory olfactory system, the stimuli that activate it, and the responses elicited in the receptor cells and in the brain. The accessory olfactory system consists of a sensory organ, the vomeronasal organ, and its central projection areas: the accessory olfactory bulb, which is connected to the amygdala and hypothalamus, and also to the cortex. In the vomeronasal organ, several receptors—in contrast to the main olfactory receptors—are sensitive to volatile or nonvolatile molecules. In a similar manner to the main olfactory epithelium, the vomeronasal organ is sensitive to common odorants and pheromones. Each accessory olfactory bulb receives input from the ipsilateral vomeronasal organ, but its activity is modulated by centrifugal projections arising from other brain areas. The processing of vomeronasal stimuli in the amygdala involves contributions from the main olfactory system, and results in long-lasting responses that may be related to the activation of the hypothalamic–hypophyseal axis over a prolonged timeframe. Different brain areas receive inputs from both the main and the accessory olfactory systems, possibly merging the stimulation of the two sensory organs to originate a more complex and integrated chemosensory perception.     

LHRH-systems in the brain of the golden hamster

Summary Vibra tome sections of male hamster brains were treated immunohistochemically with LHRH antiserum, and the anatomical distribution of LHRH immunoreactive cells and nerve fibers was assessed. LHRH-cell bodies are found in the ventral hypothalamus that includes its preoptic, anterior and central parts, in the septum, the olfactory tubercle, the main and accessory olfactory bulb, and the prepiriform cortex. In addition, extracerebral LHRH-neurons and ganglia exist in LHRH-positive nerves at the ventromedial surface of the olfactory tubercle and bulb as well as in olfactory nerves. Dense networks of LHRH-immunoreactive fibers are found in all regions where LHRH-cell bodies exist. Intraseptal connections reach the organum vasculosum of the lamina terminalis, the subfornical organ, and the lateral ventricle. Dorsolateral projections from the septum can be traced via the fimbria hippocampi and alveus to the ventral hippocampus, via the stria terminalis to the amygdala and piriform cortex. Ventrolateral projections extend from the level of the olfactory tubercle and preoptic-anterior hypothalamic area via the ventral amygdalofugal pathway to the prepiriform and piriform cortex as well as the amygdala. Dorsal supracallosal projections via the stria longitudinalis are seen in the induseum griseum and the cingulate cortex. Caudal efferents reach the habenula, interpeduncular nucleus, midbrain raphe, and central gray of the rostral fourth ventricle via the stria medullaris and fasciculus retroflexus and by a ventral projection via the periventricular and subventricular hypothalamus. A major portion of this ventrocaudal projection gives rise to a dense network in the median eminence. Anatomical relationships of LHRH-fibers to certain regions of the inner ventricular and outer brain surface are noted.Postdoctoral fellow of the Deutsche ForschungsgemeinschaftSupported by US PHS grant NS09914 and NRCHD grant HD03110     

Extra-bulbar primary olfactory projection in teleost fishes

Immunohistochemical investigations with anti-substance P antiserum demonstrate the existence of an extensive extrabulbar primary olfactory projection in several gymnotid teleost fish. This projection, never described before, originates in particular primary olfactory bundles which enter with the olfactory nerve into the olfactory bulb. While the bulk of the olfactory fibers end with glomeruli in the glomerular layer of the olfactory bulb, two particular bundles penetrate into the telencephalon and end, without forming glomeruli, in several telencephalic and diencephalic regions. A few fibers run as far as to the hypothalamus. In the light of these findings, the general notion that the primary olfactory projection is limited to the olfactory bulb and forms only glomeruli-like terminals, should be reconsidered.     

Central connections of the olfactory bulb in the bichir,Polypterus palmas,reexamined

Summary Central connections of the olfactory bulb of Polypterus palmas were studied with the use of horseradish peroxidase and cobalt-tracing techniques. The olfactory bulb projects to subpallial and palliai areas in the ipsilateral telencephalon; a projection to the contralateral subpallium is noted via the habenular commissure. A further target of secondary olfactory fibers is a caudal olfactory projection area in the ipsilateral hypothalamus. No labeling was seen in the anterior commissure and in the contralateral olfactory bulb. The medial and the lateral pallium receive secondary olfactory fibers in distinct areas. Neurons projecting to the bulb are found in the ipsilateral subpallium, mainly in one dorsal longitudinal nucleus. The main connection with the tel- and diencephalon is mediated via the medial olfactory tract. This tract also contains fibers to the contralateral telencephalon, and to the hypothalamus. The smaller lateral olfactory tract mediates fibers to the lateral pallium. The organization of pathways of secondary olfactory fibers in the telencephalon is described. The present findings are compared to those obtained in species possessing an inverted forebrain.This investigation was supported by grants from the Deutsche Forschungsgemeinschaft to DLM     

Oxidative metabolism of main and accessory olfactory bulbs, limpic system and hypothalamus during the estral cycle of the rat (author's transl)

The in vitro oxidative metabolism of hypothalamus, olfactory and limbic systems from female rats in the estrous cycle have been measured. The accessory olfactory bulb becomes most active during diestrous when the hypothalamus reaches its lowest values.     

Expression of orexin receptors in the brain and peripheral tissues of the male sheep

Orexins exert their effects through two specific receptors (OX1R and OX2R) that have been found mainly in the brain and also in peripheral tissues of rats and humans. Here, we demonstrate expression of mRNA encoding for ovine OX1R and OX2R in central and peripheral tissues of sheep. Gene expression for orexin receptors in the hypothalamus and the preoptic area was localised by in situ hybridisation. OX1R was detected in arcuate nuclei (ARC), median eminence (ME), the lateral hypothalamic nuclei and preoptic area (POA) and it was scattered along the third ventricle from the paraventricular (PVN) to the ventromedial hypothalamic nuclei (VMH). OX2R was localised in the PVN, ARC, ME, ventral VMH and a small region of the ventral POA. Gene expression for OX1R and OX2R in central and peripheral tissues was analysed using quantitative real time RT-PCR. Both orexin receptor genes were expressed in the hypothalamus, POA, hippocampus, amygdala, olfactory bulb, pineal gland and recess and pituitary gland, whereas only OX1R mRNA was detected in the testis, kidney and adrenal gland. The expression of the genes for orexin receptors in this range of ovine tissues suggests roles for orexins in multiple physiological functions, with actions at both central and peripheral levels.     

Electrophysiological evidence for a topographical projection of the nasal mucosa onto the olfactory bulb of the frog

Three olfactory nerve branches respectively subserving either a medial, an intermediate, or a lateral region of the dorsal olfactory receptor sheet of the bullfrog Rana catesbeiana were electrically stimulated with bipolar platinum hook electrodes. Extracellular single unit responses from 93 second-order cells in different regions of the olfactory bulb were recorded with metal-filled glass micropipets. The excitatory responsiveness of each unit to the stimulation of each of the three nerve branches (response profile) was determined. Some units were sensitive to stimulation of each of the three nerve branches, thus suggesting a wide projection from the entire receptor sheet. On the other hand, other units were more selective. Of this latter group, units in the lateral bulb were excited by nerve branches subserving the more lateral regions of the receptor sheet; units in the medial bulb were excited by the nerve branches subserving the more medial regions of the receptor sheet. These data provide electrophysiological evidence for a topographical projection of the olfactory receptor sheet onto the olfactory bulb, and further suggest that the projections onto different bulbar cells vary in degree of localization.     

Accessory chemosignaling mechanisms in primates

Accessory olfaction is defined as the chemoreceptive system that employs the vomeronasal complex (VNC) and its distinct central projections to the accessory olfactory bulb (AOB) and limbic/cortical systems. Comparisons of the structural and functional features of primate accessory olfaction can now be made at many levels. Advances in the understanding of molecular mechanisms of odorant transfer and detection, physiological analyses of signal processing, and appreciation of ontogenetic timetables have clarified the contribution of accessory chemoreception to the sensory map. Two principal functions dominate: the decoding of social information through the uptake of signals (often fluid-borne), and the provision of an essential pathway for the "migration" of presumptive neurocrine (GnRH) cells from the olfactory placode to the hypothalamus. VN "smelling" (vomerolfaction) is now seen to overlap with primary olfaction. Both systems detect signal compounds along the spectrum of volatility/molecular weight, and neither is an exclusive sensor. Both main and accessory chemoreception seem to require collaborative molecular devices to assist in odorant transfer (binding proteins) and (for the VNO) signal recognition (MHC1 proteins). Most adaptive-selective features of primate chemocommunication variously resemble those of other terrestrial mammals. VN function, along with its genome, has been maintained within the Strepsirrhines and tarsiers, reduced in Platyrrhines, and nearly extinguished at the Catarrhine up to hominin levels. It persists as an intriguing ancient sense that retains key features of past evolutionary events.     

Involvement of accessory neurosecretory nuclei of hypothalamus in the formation of hypothalamohypophyseal system during prenatal and postnatal development in rats

We studied the development of direct axonal connections of the accessory neurosecretory hypothalamic nuclei with the posterior pituitary lobe on the fixed rat brain from day 15 of embryogenesis until day 10 of postnatal development using the retrograde diffusion method of the lipophilic fluorescent carbocyanine dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate along the neuronal membranes. The marker was applied onto the posterior pituitary lobe and, after incubation in a fixative, fluorescing bodies of nerve cells were visualized in the hypothalamus. Neuronal axons of the retrochiasmatic nucleus were the first of the accessory nuclei to ingrow in the posterior pituitary lobe (on days 16-17 of embryogenesis). Neurons of the circular and dorsolateral nuclei and the nuclei of the median bundle of the forebrain sent their axons to the posterior pituitary lobe starting from the first days of postnatal development. No direct connections of the anterior commissure and perifornical accessory nuclei with the posterior pituitary lobe were found in perinatal development. These facts are discussed in the light of concepts about the different functional role of accessory peptidergic hypothalamic nuclei in rats.     

The central pathway of primary olfactory axons is abnormal in mice lacking the N-CAM-180 isoform

Although N-CAM has previously been implicated in the growth and fasciculation of axons, the development of axon tracts in transgenic mice with a targeted deletion of the 180-kD isoform of the neural cell adhesion molecule (N-CAM-180) appears grossly normal in comparison to wild-type mice. We examined the organization of the olfactory nerve projection from the olfactory neuroepithelium to glomeruli in the olfactory bulb of postnatal N-CAM-180 null mutant mice. Immunostaining for olfactory marker protein revealed the normal presence of fully mature primary olfactory neurons within the olfactory neuroepithelium of mutant mice. The axons of these neurons form an olfactory nerve, enter the nerve fiber layer of the olfactory bulb, and terminate in olfactory glomeruli as in wild-type control animals. The olfactory bulb is smaller and the nerve fiber layer is relatively thicker in mutants than in wild-type mice. Previous studies have revealed that the plant lectin Dolichos biflorus agglutinin (DBA) clearly stains the perikarya and axons of a subpopulation of primary olfactory neurons. Thus, DBA staining enabled the morphology of the olfactory nerve pathway to be examined at higher resolution in both control and mutant animals. Despite a normal spatial pattern of DBA-stained neurons within the nasal cavity, there was a distorted axonal projection of these neurons onto the surface of the olfactory bulb in N-CAM-180 null mutants. In particular, DBA-stained axons formed fewer and smaller glomeruli in the olfactory bulbs of mutants in comparison to wild-type mice. Many primary olfactory axons failed to exit the nerve fiber layer and contribute to glomerular formation. These results indicate that N-CAM-180 plays an important role in the growth and fasciculation of primary olfactory axons and is essential for normal development of olfactory glomeruli. © 1997 John Wiley & Sons, Inc. J Neurobiol 32 : 643–658, 1997     

Involvement of Accessory Neurosecretory Nuclei of Hypothalamus in the Formation of Hypothalamohypohysial System during Prenatal and Postnatal Development in Rats

We studied the development of direct axonal connections of the accessory neurosecretory hypothalamic nuclei with the posterior pituitary lobe on the fixed rat brain from day 15 of embryogenesis until day 10 of postnatal development using the retrograde diffusion method of the lipophilic fluorescent carbocyanine dye 1,1"-dioctadecyl-3,3,3",3"-tetramethylindocarbocyanine perchlorate along the neuronal membranes. The marker was applied onto the posterior pituitary lobe and, after incubation in a fixative, fluorescing bodies of nerve cells were visualized in the hypothalamus. Neuronal axons of the retrochiasmatic nucleus were the first of the accessory nuclei to ingrow in the posterior pituitary lobe (on days 16–17 of embryogenesis). Neurons of the circular and dorsolateral nuclei and the nuclei of the median bundle of the forebrain sent their axons to the posterior pituitary lobe starting from the first days of postnatal development. No direct connections of the anterior commissure and perifornical accessory nuclei with the posterior pituitary lobe were found in perinatal development. These facts are discussed in the light of concepts about the different functional role of accessory peptidergic hypothalamic nuclei in rats.     

A neuropeptide FF-related gene is expressed selectively in neurons of the terminal nerve in Danio rerio

RFamides constitute a large family of neuromodulatory peptides. We have cloned a zebrafish gene, which is presumably a homologue to the mammalian PQRF subfamily of RFamides, and named it zfPQRF for its species and subfamily allocation. We report that in contrast to its mammalian counterparts zfPQRF is expressed in the olfactory bulb and the nucleus olfactoretinalis in the telencephalon, but absent in more caudal regions, including hypothalamus, brain stem and spinal cord. zfPQRF-expressing neurons originate in the vicinity of the olfactory placode and populate the nuclei of the terminal nerve during later development, as demonstrated by co-expression of zebrafish salmon-type gonadotropin releasing hormone, which was found to exclusively label terminal nerve neurons.     

In vitro TRH release from hypothalamus slices varies during the diurnal cycle

We have previously described a daily rhythm in thyrotropin releasing hormone (TRH) and TRH mRNA in the rat hypothalamus. To determine whether TRH release fluctuates in a diurnal manner, we have measured basal and potassium stimulated release from hypothalamic slices, and compared it to release from olfactory bulb slices, during the diurnal cycle. Basal TRH release was higher at 7:00 h than at any other time (1:00, 13:00 or 19:00 h) in either hypothalamus or olfactory bulb. The ratio of stimulated over basal release was higher in the hypothalamus at 19:00 h, when TRH content was highest. Potassium stimulated TRH release from olfactory bulb was not different from basal release at any time. TRH release fluctuations were not due to a rhythm of extracellular inactivation: the activity of pyroglutamyl aminopeptidase II, an ectoenzyme responsible for TRH inactivation, was constant throughout the cycle. Our data demonstrate that diurnal variations of TRH release occur in vitro and that the enhanced responsiveness to potassium stimulation in hypothalamus is correlated with increased levels of peptide.     

Localization of neurokinin B in the central nervous system of the rat.

The distribution of neurokinin B (NKB) was determined by immunocytochemistry with antisera directed toward its amino terminus. Immunoreactive perikarya were detected in the main and accessory olfactory bulbs, cortical regions, the olfactory tubercle, the bed nucleus of the stria terminalis, the diagonal band of Broca, the nucleus accumbens, the septum, the neostriatum, several hypothalamic nuclei, the superior colliculus, the central gray, the substantia nigra, the medullary reticular formation, and the external cuneate nucleus. The distribution of NKB-containing perikarya revealed by immunocytochemistry was similar to the distribution of protachykinin B-containing cells previously visualized by in situ hybridization. Immunoreactive nerve fibers and terminals were detected in all major subdivisions of the brain. The levels of NKB measured by radioimmunoassay were highest in the hypothalamus. The distribution of NKB in the rat brain was similar to the distribution of substance P; however, there were several regions where the two distributions were clearly different.     

Goα expression in the vomeronasal organ and olfactory bulb of the tammar wallaby

The vomeronasal organ (VNO) detects pheromones via 2 large families of receptors: vomeronasal receptor 1, associated with the protein Giα2, and vomeronasal receptor 2, associated with Goα. We investigated the distribution of Goα in the developing and adult VNO and adult olfactory bulb of a marsupial, the tammar wallaby. Some cells expressed Goα as early as day 5 postpartum, but by day 30, Goα expressing cells were distributed throughout the receptor epithelium of the VNO. In the adult tammar, Goα appeared to be expressed in sensory neurons whose nuclei were mostly basally located in the vomeronasal receptor epithelium. Goα expressing vomeronasal receptor cells led to all areas of the accessory olfactory bulb (AOB). The lack of regionally restricted projection of the vomeronasal receptor cell type 2 in the tammar was similar to the uniform type, with the crucial difference that the uniform type only shows expression of Giα2 and no expression of Goα. The observed Goα staining pattern suggests that the tammar may have a third accessory olfactory type that could be intermediate to the segregated and uniform types already described.     

Loss of smelling is an early marker of aging and is associated with inflammation and DNA damage in C57BL/6J mice

Olfactory dysfunction is a prevalent symptom and an early marker of age-related neurodegenerative diseases in humans, including Alzheimer's and Parkinson's Diseases. However, as olfactory dysfunction is also a common symptom of normal aging, it is important to identify associated behavioral and mechanistic changes that underlie olfactory dysfunction in nonpathological aging. In the present study, we systematically investigated age-related behavioral changes in four specific domains of olfaction and the molecular basis in C57BL/6J mice. Our results showed that selective loss of odor discrimination was the earliest smelling behavioral change with aging, followed by a decline in odor sensitivity and detection while odor habituation remained in old mice. Compared to behavioral changes related with cognitive and motor functions, smelling loss was among the earliest biomarkers of aging. During aging, metabolites related with oxidative stress, osmolytes, and infection became dysregulated in the olfactory bulb, and G protein coupled receptor-related signaling was significantly down regulated in olfactory bulbs of aged mice. Poly ADP-ribosylation levels, protein expression of DNA damage markers, and inflammation increased significantly in the olfactory bulb of older mice. Lower NAD⁺ levels were also detected. Supplementation of NAD⁺ through NR in water improved longevity and partially enhanced olfaction in aged mice. Our studies provide mechanistic and biological insights into the olfaction decline during aging and highlight the role of NAD⁺ for preserving smelling function and general health.     

Mating Increases Neuronal Tyrosine Hydroxylase Expression and Selectively Gates Transmission of Male Chemosensory Information in Female Mice

Exposure to chemosensory signals from unfamiliar males can terminate pregnancy in recently mated female mice. The number of tyrosine hydroxylase-positive neurons in the main olfactory bulb has been found to increase following mating and has been implicated in preventing male-induced pregnancy block during the post-implantation period. In contrast, pre-implantation pregnancy block is mediated by the vomeronasal system, and is thought to be prevented by selective inhibition of the mate’s pregnancy blocking chemosignals, at the level of the accessory olfactory bulb. The objectives of this study were firstly to identify the level of the vomeronasal pathway at which selective inhibition of the mate’s pregnancy blocking chemosignals occurs. Secondly, to determine whether a post-mating increase in tyrosine hydroxylase-positive neurons is observed in the vomeronasal system, which could play a role in preventing pre-implantation pregnancy block. Immunohistochemical staining revealed that mating induced an increase in tyrosine-hydroxylase positive neurons in the arcuate hypothalamus of BALB/c females, and suppressed c-Fos expression in these neurons in response to mating male chemosignals. This selective suppression of c-Fos response to mating male chemosignals was not apparent at earlier levels of the pregnancy-blocking neural pathway in the accessory olfactory bulb or corticomedial amygdala. Immunohistochemical staining revealed an increase in the number of tyrosine hydroxylase-positive neurons in the accessory olfactory bulb of BALB/c female mice following mating. However, increased dopamine-mediated inhibition in the accessory olfactory bulb is unlikely to account for the prevention of pregnancy block to the mating male, as tyrosine hydroxylase expression did not increase in females of the C57BL/6 strain, which show normal mate recognition. These findings reveal an association of mating with increased dopaminergic modulation in the pregnancy block pathway and support the hypothesis that mate recognition prevents pregnancy block by suppressing the activation of arcuate dopamine release.     

Functional anatomy of the peripheral olfactory system of the African lungfish Protopterus annectens owen: development of the primary olfactory pathway during postembryonic growth

Seven specimens of the African lungfish ranging in size from 4 to 350 gm were used for the study of changes in the olfactory nerve and bulb during postembryonic growth as expressed by the increasing number of lamellae of the olfactory organ. Serial semithin sections were used, and the data were studied mathematically. The number of axons of the olfactory nerve, the surface area of the bulb, and the number of mitral cells increase exponentially; the mitral cell density decreases exponentially; and the calculated average convergence (counted axons/counted mitral cells) of axons onto the cells of the bulbar relay increases exponentially. Convergence figures obtained are smaller than those cited by other authors; two factors might explain this: first the mode of calculation, and second the fact that every year this fish undergoes a 6-month starvation stage marked by degeneration of the primary olfactory structure. A preliminary view of the primary olfactory pathway using serial-section reconstructions is proposed: An orderly projection of the mucosa onto the bulb takes place after a 90 degrees rotation of the system of two perpendicular axes which apply to both the olfactory organ and the olfactory bulb. This study suggests an original attempt at relating the postembryonic change in convergence and projection to the primary processing of the olfactory message.