Diss. (sammanfattning) Alnarp : Sveriges lantbruksuniversitet, 2004. / Härtill 5 uppsatser.

Diss. (sammanfattning) Umeå : Umeå universitet, 2008. / Härtill 3 uppsatser.

The human olfactory system can classify new odors in a dynamic environment with varying odor complexity and concentration, while simultaneously reducing the influence of stable background odors. Replication of this capability has remained an active area of research over the past 3 decades and has great potential to advance medical diagnostics, environmental monitoring and industrial monitoring, among others. New methods for rapid dynamic temporal evaluation of chemical sensor arrays for the monitoring of analytes is explored in this work. One such method is high and low bandpass filtering of changing sensor responses; this is applied to reduce the effects of background noise and sensor drift over time. Processed sensor array responses, coupled with principal component analysis (PCA), will be used to develop a novel approach to classify odors in the presence of changing sensor responses associated with evolving odor concentrations. These methods will enable the removal of noise and drift, as well as facilitating the normalization to decouple classification patterns from intensity; lastly, PCA and artificial neural networks (ANNs) will be used to demonstrate the capability of this approach to function under dynamic conditions, where concentration is changing temporally. / February 2016

Olfactory signals are an important means of social communication among felids. However, not much is known about how individual volatiles of body-borne odours influence behavioural responses. 2-acetyl-1pyrroline has recently been identified as a characteristic component of tiger marking fluid, while being absent from lion marking fluid. One pride each of captive Amur tigers (Panthera tigris altaica) and African lions (Panthera leo) were presented with wooden logs impregnated with four different odours and their behaviour was observed. The tigers displayed significantly more interactions towards the marking fluid component (2-acetyl-1-pyrroline), the conspecific urine odour, and the fruity odour (iso-pentyl acetate) than towards the near odourless control (diethyl phthalate). The lions displayed significantly more behaviours towards conspecific urine than towards any of the other odours.  In general all lions interacted more with the logs than tigers. Hence, these results support the notion that 2-acetyl-1-pyrroline is a species-specific odorant for tiger olfactory communication. Furthermore, the results show that a single compound (2-acetyl-1pyrroline) can elicit behavioural responses to the same degree as a complex chemical mixture (tiger urine). The high number of interactions performed by both species towards the wooden logs impregnated with conspecific urine suggests that conspecific odours are suitable to use as olfactory enrichment for captive felids.

Female mice exhibit a hardwired preference to investigate pheromones released by male conspecifics. The neural pathways that convey pheromonal inputs to brain regions controlling motivated behaviors remain largely unknown. One brain region known to process pheromonal information conveyed via main- and accessory olfactory bulb inputs is the Medial Amygdala (Me), a limbic structure comprised of anterior (MeA) and posterior (MeP) subdivisions. Electrolytic lesions of the MeP blocked the normal preference of estrous female mice to investigate urinary odors emitted from breeding as opposed to castrated males whereas lesions of either the MeA or MeP significantly reduced females' display of the receptive lordosis posture in response to male mounts. Quantitative analysis of synaptic puncta in the efferent projection targets of these two amygdaloid subregions, visualized using fluorescent anterograde tract tracing techniques, revealed that the MeA and MeP differentially innervate several forebrain regions. The medial olfactory tubercle (mOT; a component of the ventral striatum) receives dense monosynaptic input from the MeA and responds selectively to breeding male (but not female) soiled bedding volatiles, indexed by augmented FOS expression. Using injections of the retrograde tracer, cholera toxin B (CTb), neurons were identified in the MeA and ventral tegmental area (VTA) that projected to the mOT in female mice and which also co-expressed FOS after exposure to breeding male, but not female, soiled bedding/urinary volatiles. This suggests that the MeA and VTA convey opposite-sex (male) pheromonal information to the mOT. Bilateral dopaminergic lesions of the anteromedial VS (a region which includes the mOT) eliminated females' preference for breeding male vs. female urinary pheromones, suggesting that dopaminergic modulation in the VS is necessary for the display of these behaviors. Lastly, bilateral silencing of mOT neuronal firing by the activation of the inhibitory DREADD receptor, hM4Di, induced by intraperitoneal injection of its ligand (CNO), also disrupted females' preference to investigate urinary odors from breeding males; this deficit was reversed when saline was administered instead of CNO. The Me, VTA, and mOT are essential segments of a neural reward circuit that motivates estrous female mice to seek out male pheromones, thereby facilitating mate recognition and reproductive success.

abstract: Sensory gating is a process by which the nervous system preferentially admits stimuli that are important for the organism while filtering out those that may be meaningless. An optimal sensory gate cannot be static or inflexible, but rather plastic and informed by past experiences. Learning enables sensory gates to recognize stimuli that are emotionally salient and potentially predictive of positive or negative outcomes essential to survival. Olfaction is the only sensory modality in mammals where sensory inputs bypass conventional thalamic gating before entering higher emotional or cognitive brain regions. Thus, olfactory bulb circuits may have a heavier burden of sensory gating compared to other primary sensory circuits. How do the primary synapses in an olfactory system "learn"' in order to optimally gate or filter sensory stimuli? I hypothesize that centrifugal neuromodulator serotonin serves as a signaling mechanism by which primary olfactory circuits can experience learning informed sensory gating. To test my hypothesis, I conditioned genetically-modified mice using reward or fear olfactory-cued learning paradigms and used pharmacological, electrophysiological, immunohistochemical, and optical imaging approaches to assay changes in serotonin signaling or functional changes in primary olfactory circuits. My results indicate serotonin is a key mediator in the acquisition of olfactory fear memories through the activation of its type 2A receptors in the olfactory bulb. Functionally within the first synaptic relay of olfactory glomeruli, serotonin type 2A receptor activation decreases excitatory glutamatergic drive of olfactory sensory neurons through both presynaptic and postsynaptic mechanisms. I propose that serotonergic signaling decreases excitatory drive, thereby disconnecting olfactory sensory neurons from odor responses once information is learned and its behavioral significance is consolidated. I found that learning induced chronic changes in the density of serotonin fibers and receptors, which persisted in glomeruli encoding the conditioning odor. Such persistent changes could represent a sensory gate stabilized by memory. I hypothesize this ensures that the glomerulus encoding meaningful odors are much more sensitive to future serotonin signaling as such arousal cues arrive from centrifugal pathways originating in the dorsal raphe nucleus. The results advocate that a simple associative memory trace can be formed at primary sensory synapses to facilitate optimal sensory gating in mammalian olfaction. / Dissertation/Thesis / Ph.D. Biology 2012

Une dysfonction olfactive et une altération de la neurogenèse sont observées dans plusieurs maladies neurodégénératives, y compris la maladie de Huntington (MH). Ce déficit est un symptôme précoce de la MH et est en corrélation avec le déclin de la performance cognitive globale, la dépression et la dégénérescence des régions olfactives dans le cerveau. La dysfonction olfactive dans les maladies neurodégénératives est souvent accompagnée par des anomalies structurelles de l’épithélium olfactif, du bulbe olfactif (BO) et du cortex olfactif chez l’humain. En dépit de preuves claires démontrant la dysfonction olfactive chez les patients de la MH l'information disponible est limitée dans des modèles murins et les mécanismes sous-jacents ne sont pas connus. Une diminution du volume et du compte neuronale dans le PC est observée dans les souris YAC128 vs le type sauvage (WT) âgée (12 mois). Nous avons également examiné les comportements lors d'exposition à des odeurs sociales et non sociales chez des souris en utilisant le test habituation. Une habituation aux odeurs tend à être observé dans les souris YAC128 de 1 mois vs WT se traduisant par une tendance de l’augmentation de la durée d’exploration des YAC128 vs WT lors de leurs 2e et 3e expositions à une odeur. Dans les couches glomérulaire et plexiforme externe, l’intensité réciproque du marquage de TH et de GFAP, marqueur de cellules gliales présente une tendance pour une augmentation dans les YAC128 comparativement au WT. Une tendance pour une diminution de Iba-1, marqueur de neuroinflammation, a aussi été observé dans la couche granulaire du BO de YAC128 âgée vs WT. Malgré une diminution de l’expression en ARNm de caspase-3 et -8, une augmentation de l'expression protéique de la proforme de caspase-8 et des formes actives de la caspase-8, -6 et -9 a été observés au stade présymptomatique dans des BO de YAC128 vs WT. De façon similaire aux YAC128, une atrophie du BO à 6 mois est remarqué dans le modèle de rat BACHD (lignées TG5 et TG9). Globalement, un niveau d’expression de la protéine huntington mutante (httm) est plus élevé dans les TG5. L’expression protéique dans les BO de la proforme des caspase-3, -6 et -8 sont supérieurs dans les TG9 lorsque comparés au TG5 et au WT. L’identification de marqueurs précoces pour la MH contribuera aux approches thérapeutiques et permettra de clarifier l'utilité des tests de la fonction olfactive dans les individus à risques de la MH.

Philosophiae Doctor - PhD / Tsetse flies are the sole vectors of trypanosomes that cause nagana and sleeping sickness in animals and humans respectively in tropical Africa. Tsetse are unique: both sexes adults are exclusive blood-feeders, females are mated young and give birth to a single mature larva in sheltered habitats per pregnancy. Tsetse use chemoreception to detect and respond to chemical stimuli, helping them to locate hosts, mates, larviposition and resting sites. The detection is facilitated by chemoreceptors expressed on sensory neurons to cause specific responses. Specific molecular factors that mediate these responses are poorly understood in tsetse flies. This study aimed to identify and characterize genes that potentially mediate chemoreception in Glossina morsitans morsitans tsetse flies. These genes included sensory odorant (OR), gustatory (GR), ionotropic (IR), and related genes for odorant-binding (OBP), chemosensory (CSP) and sensory neuron membrane (SNMP) proteins. Synaptic transmission in higher brain sites may involve ionotropic glutamate-gated (iGluR) and metabotropic glutamate-gated (mGluR) receptors. The genes were annotated in G. m. morsitans genome scaffold assembly GMOY1.1 Yale strain using orthologs from D. melanogaster as query via TBLASTX algorithm at e-value below 1e-03. Positive blast hits were seeded as gene constructs in their respective scaffolds, and used as genomic reference onto which female fly-derived RNA sequence reads were mapped using CLC Genomics workbench suite. Seeded gene models were modified using RNA-Seq reads then viewed and re-edited using Artemis genome viewer tool. The genome was iteratively searched using the G. m. morsitans gene model sequences to recover additional similar hit sequences. The gene models were confirmed through comparisons against the NCBI conserved domains database (CDD) and non-redundant Swiss-Prot database. Trans-membrane domains and secretory peptides were predicted using TMHMM and SignalP tools respectively. Putative functions of the genes were confirmed via Blast2GO searches against gene ontology database. Evolutionary relationships amongst and between the genes were established using maximum likelihood estimates using best fitting amino acid model test in MEGA5 suite and PhyML tool. Expression profiles of genes were estimated using the RNA-seq data via CLCGenomics RNA-sequences analysis pipeline. Overall, 46 ORs, 14 GRs, and 19 IRs were identified, of which 21, 6 and 4 were manually identified for ORs, GRs, and IRs respectively. Additionally, 15 iGluRs, 6 mGluRs, 5 CSPs, 15 CD36-like, and 32 OBPs were identified. Six copies of OR genes (GmmOR41-46) were homologous to DmelOr67d, a single copy cis vacenyl acetate (cVA) receptor . Genes whose receptor homologs are associated with responses to CO2, GmmGR1-4, had higher expression profiles from amongst glossina GR genes. Known core-receptor homologs OR1, IR8a, IR25a and IR64a were conserved, and three species-specific divergent IRs (IR10a, IR56b and IR56d) were identified. Homologs of GluRIID, IR93a, and sweet taste receptors (Gr5a and Gr64a) were not identified in the genome. Homolog for LUSH protein, GmmOBP26, and sensory neuron membrane receptors SNMP1 and SNMP2 were conserved in the genome. Results indicate reduced repertoire of the chemosensory genes, and suggest reduced host range of the tsetse flies compared to other Diptera. Genes in multiple copies suggest their prioritization in chemoreception, which in turn may be tied to high specificity in host selection. Genes with high sequence conservation and expression profiles probably relate to their broad expression and utility within the fly nervous system. These results lay foundation for future comparative studies with other insects, provide opportunities for functional studies, and form the basis for re-examining new approaches for improving tsetse control tools and possible drug targets based on chemoreception.

Artificial olfaction has the potential to revolutionise medical diagnosis, speed up threat detection, and provide long-term data collection on atmospheric pollution. Current technology provides a means to detect some compounds, but detection speed is slow, and accuracy is often balanced against cost and reusability of devices. Insects have long been the inspiration for artificial olfaction and are able to detect low concentrations of compounds over vast areas and navigate towards these targets. A deeper understanding of how this is achieved could inform the design of better artificial olfactory devices. What features of the insect olfactory system allow the rapid detection of miniscule concentrations of a wide variety of compounds? How does an insect discriminate odours using a limited number of olfactory receptor types?Using UAS/GAL4 technology, Drosophila melanogaster larvae expressing only a single functional olfactory sensory neuron (OSN), of their full repertoire of 21, were used to explore peripheral olfactory responses. Three different fly lines, which each expressed different olfactory receptor (OR) types, were used to record the electrophysiological responses of the peripheral OSNs to a panel of biologically significant odours, at differing concentrations. These responses were compared to those from a leading electronic nose (metal oxide sensors) for the same odour conditions, and the features of the responses were characterised. A novel odour delivery system was also developed to accurately deliver these odours and concentrations repeatably, and was validated using a commercially available photoionisation detector. The ability to correctly select the odour from which a response was recorded, out of a choice of five odours, at two concentrations each, was scrutinised using a classifier algorithm. The three OSN types achieved 53%, 62% and 75% accuracy, respectively. This is the first instance that Drosophila larvae have been conclusively shown to be able to discriminate odour concentrations using only a single peripheral OSN. The two metal oxide sensors achieved 92% and 95% accuracy under the same conditions. Whilst the features of the responses used to discriminate odours differed between the biological and electronic systems, the time frame required for correct classification of sensor data was only, in some cases, three seconds longer than in OSNs (~0.5 seconds), indicating that metal oxide sensors may have a useful role to play in biologically inspired artificial olfaction.

The study of feeding behavior, and in particular the study of subjective hedonic experience and objective measures of motivation, are central to understanding how appetite regulation can be compromised in certain individuals. Furthermore, with an integrated picture of physiological and behavioral changes that can occur as a result of energy deprivation what emerges is a better understanding of how palatable food can disrupt attempts at regulating body weight at lower levels of body energy stores. In Article I, the genetic association study examining a potential role for a dopamine-related polymorphism in weight loss, it was shown that contrary to the main hypothesis there was no association between TaqIA polymorphism and the amount of body weight loss. In Article II, it was shown that palatability and olfaction ratings increased as a result of a 24 hour fast and females demonstrated larger improvements in overall olfactory performance. Initial body weight was positively related to improved odor detection threshold and total odour score (TDI). Using the same population sample as Article II, Article III highlights that higher sensitivity to reward and disinhibition scores correlated with responding for palatable snack food stimuli in the relative-reinforcing value of food (RRV) task, further indicating that RRV has strong ties with impulsivity. There was a demonstrable lack of negative alliesthesia under the fasted condition where, after a 75% increase in ad libitum energy intake (EI) relative to the fed condition, this greater amount of food consumed was still rated as being more palatable than the lesser amounts consumed under the fed condition. In Article IV it was shown that an equicaloric (-25%) energy deficit by diet alone was a greater challenge to appetite regulation and resulted in greater compensatory increases in EI than deprivation by exercise alone. Independent of deprivation modality there were significant improvements in odour threshold scores. TDI score increased only under diet alone; furthermore, the noted increase in mean TDI score was positively related to increased ad libitum EI. The picture that emerges is that, acutely, a complete fast has more pronounced effects on appetite and ad libitum EI than dieting alone, which in turn had greater effects than exercise alone or controls. Also, TDI improved under all three methods of energy deprivation, but moreso under conditions of deprivation by diet alone.