Contextual Fear Conditioning Alter Microglia Number and Morphology in the Rat Dorsal Hippocampus

Contextual fear conditioning is a Pavlovian conditioning paradigm capable of rapidly creating fear memories to contexts, such as rooms or chambers. Contextual fear conditioning protocols have long been utilized to evaluate how fear memories are consolidated, maintained, expressed, recalled, and extinguished within the brain. These studies have identified the lateral portion of the amygdala and the dorsal portion of the hippocampus as essential for contextual fear memory consolidation. The current study was designed to evaluate how two different contextual fear memories alter amygdala and hippocampus microglia, brain derived neurotrophic factor (BDNF), and phosphorylated cyclic-AMP response element binding (pCREB). We find rats provided with standard contextual fear conditioning to have more microglia and more cells expressing BDNF in the dentate gyrus as compared to a context only control group. Additionally, standard contextual fear conditioning altered microglia morphology to become amoeboid in shape – a common response to central nervous system insult, such as traumatic brain injury, infection, ischemia, and more. The unpaired fear conditioning procedure (whereby non-reinforced and non-overlapping auditory tones were provided at random intervals during conditioning), despite producing equivalent levels of fear as the standard procedure, did not alter microglia, BDNF or pCREB number in any dorsal hippocampus or lateral amygdala brain regions. Despite this, the unpaired fear conditioning protocol produced some alterations in microglia morphology, but less compared to rats provided with standard contextual fear conditioning. Results from this study demonstrate that contextual fear conditioning is capable of producing large alterations to dentate gyrus plasticity and microglia, whereas unpaired fear conditioning only produces minor changes to microglia morphology. These data show, for the first time, that Pavlovian fear conditioning protocols can induce similar responses as trauma, infection or other insults within the central nervous system

A novel method using intranasal delivery of EdU demonstrates that accessory olfactory ensheathing cells respond to injury by proliferation

Olfactory ensheathing cells (OECs) play an important role in the continuous regeneration of the primary olfactory nervous system throughout life and for regeneration of olfactory neurons after injury. While it is known that several individual OEC subpopulations with distinct properties exist in different anatomical locations, it remains unclear how these different subpopulations respond to a major injury. We have examined the proliferation of OECs from one distinct location, the peripheral accessory olfactory nervous system, following large-scale injury (bulbectomy) in mice. We used crosses of two transgenic reporter mouse lines, S100ß-DsRed and OMP-ZsGreen, to visualise OECs, and main/accessory olfactory neurons, respectively. We surgically removed one olfactory bulb including the accessory olfactory bulb to induce degeneration, and found that accessory OECs in the nerve bundles that terminate in the accessory olfactory bulb responded by increased proliferation with a peak occurring 2 days after the injury. To label proliferating cells we used the thymidine analogue ethynyl deoxyuridine (EdU) using intranasal delivery instead of intraperitoneal injection. We compared and quantified the number of proliferating cells at different regions at one and four days after EdU labelling by the two different methods and found that intranasal delivery method was as effective as intrapeitoneal injection. We demonstrated that accessory OECs actively respond to widespread degeneration of accessory olfactory axons by proliferating. These results have important implications for selecting the source of OECs for neural regeneration therapies and show that intranasal delivery of EdU is an efficient and reliable method for assessing proliferation of olfactory glia

The sorting behaviour of olfactory and vomeronasal axons during regeneration

In order to begin to understand how primary olfactory and vomeronasal organ (VNO) axons target specific regions of the olfactory bulb, we examined the sorting behaviour of these axons following neonatal unilateral olfactory bulbectomy. Bulbectomy induced widespread ipsilateral death of the primary olfactory and VNO neurons. After 4 weeks, many new sensory axons had re-grown into the cranial cavity and established a prominent plexus with evidence of dense tufts that were similar in gross appearance to glomeruli. Axons expressing the cell adhesion molecule OCAM, which normally innervate the ventrolateral and rostral halves of the main and accessory olfactory bulbs, respectively, sorted out and segregated from those axons not expressing this molecule within the plexus. In addition, VNO axons formed large discrete bundles that segregated from main olfactory axons within the plexus. Thus, VNO and primary olfactory axons as well as discrete subpopulations of both are able to sort out and remain segregated in the absence of the olfactory bulb. Sorting and convergence of axons therefore occur independently of the olfactory bulb and are probably attributable either to inherent properties of the axons themselves or to interactions between the axons and accompanying glial ensheathing cells.</p

Factors that modulate olfactory dysfunction

The olfactory system is one of a few areas in the nervous system which is capable of regeneration throughout the life. Olfactory sensory neurons reside in the nasal cavity are continuously replenished with new neurons arising from stem cells. Some factors such as aging, neurodegenerative diseases, head trauma, brain tumor extraction and infection cause olfactory dysfunction which significantly influences physical wellbeing, quality of life, mental health, nutritional status, memory processes, identifying danger and is associated with increased mortality. Therefore, finding a treatment to improve olfactory dysfunction is needed. Recent research efforts in the field have shown some very promising new approaches to treat olfactory dysfunction. This review explores the current studies that have addressed therapeutic approaches to improve olfactory neuron regeneration based on cell transplantation therapy, modulation of physiological olfactory dysfunction and drug treatments

Implantation of a scaffold following bulbectomy induces laminar organization of regenerating olfactory axons

Primary olfactory axons expressing different odorant receptors are interspersed within the olfactory nerve. However, upon reaching the outer nerve fiber layer of the olfactory bulb they defasciculate, sort out, and refasciculate prior to targeting glomeruli in fixed topographic positions. While odorant receptors are crucial for the final targeting of axons to glomeruli, it is unclear what directs the formation of the nerve fiber and glomerular layers of the olfactory bulb. While the olfactory bulb itself may provide instructive cues for the development of these layers, it is also possible that the incoming axons may simply require the presence of a physical scaffold to establish the outer laminar cytoarchitecture. In order to begin to understand the underlying role of the olfactory bulb in development of the outer layers of the olfactory bulb, we physically ablated the olfactory bulbs in OMP-IRES-LacZ and P2-IRES-tau-LacZ neonatal mice and replaced them with artificial biological scaffolds molded into the shape of an olfactory bulb. Regenerating axons projected around the edge of the cranial cavity at the periphery of the artificial scaffold and were able to form an olfactory nerve fiber layer and, to some extent, a glomerular layer. Our results reveal that olfactory axons are able to form rudimentary cytoarchitectonic layers if they are provided with an appropriately shaped biological scaffold. Thus, the olfactory bulb does not appear to provide any tropic substance that either attracts regenerating olfactory axons into the cranial cavity or induces these axons to form a plexus around its outer surface. (c) 2006 Elsevier B.V. All rights reserved

The limitations of investigating appetite through circuit manipulations: are we biting off more than we can chew?

Disordered eating can underpin a number of debilitating and prevalent chronic diseases, such as obesity. Broader advances in psychopharmacology and biology have motivated some neuroscientists to address diet-induced obesity through reductionist, pre-clinical eating investigations on the rodent brain. Specifically, chemogenetic and optogenetic methods developed in the 21st century allow neuroscientists to perform in vivo, region-specific/projection-specific/promoter-specific circuit manipulations and immediately assess the impact of these manipulations on rodent feeding. These studies are able to rigorously conclude whether a specific neuronal population regulates feeding behaviour in the hope of eventually developing a mechanistic neuroanatomical map of appetite regulation. However, an artificially stimulated/inhibited rodent neuronal population that changes feeding behaviour does not necessarily represent a pharmacological target for treating eating disorders in humans. Chemogenetic/optogenetic findings must therefore be triangulated with the array of theories that contribute to our understanding of appetite. The objective of this review is to provide a wide-ranging discussion of the limitations of chemogenetic/optogenetic circuit manipulation experiments in rodents that are used to investigate appetite. Stepping into and outside of medical science epistemologies, this paper draws on philosophy of science, nutrition, addiction biology and neurophilosophy to prompt more integrative, transdisciplinary interpretations of chemogenetic/optogenetic appetite data. Through discussing the various technical and epistemological limitations of these data, we provide both an overview of chemogenetics and optogenetics accessible to non-neuroscientist obesity researchers, as well as a resource for neuroscientists to expand the number of lenses through which they interpret their circuit manipulation findings

Does neuroscience research change behaviour? A scoping review and case study in obesity neuroscience

The language employed by researchers to define and discuss diseases can itself be a determinant of health. Despite this, the framing of diseases in medical research literature is largely unexplored. This scoping review examines a prevalent medical issue with social determinants influenced by the framing of its pathogenesis: obesity. Specifically, we compare the currently dominant framing of obesity as an addiction to food with the emerging frame of obesity developing from neuroinflammation. We triangulate both corpus linguistic and bibliometric analysis of the top 200 most engaging neuroscience journal articles discussing obesity that were published open access in the past 10 years. The constructed Neurobesity Corpus is available for public use. The scoping review analysis confirmed that neuroinflammation is an emerging way for obesity to be framed in medical research. Importantly, the articles analysed that discussed neuroinflammation were less likely to use crisis terminology, such as referring to an obesity “epidemic”. We highlight a potential relationship between the adoption of addiction frames and the use of stigmatising language in medical research

The Grueneberg olfactory organ neuroepithelium recovers after injury

The Grueneberg organ (also termed Grueneberg ganglion) is an olfactory subsystem at the rostral nasal septum of rodents, and has been suggested to exist also in humans. Grueneberg organ neurons respond to coldness and alarm pheromones, but the anatomical arrangement and regenerative capacity are not fully characterised. We examined the relationship between the glia and the neurons using crosses of two transgenic mouse lines, S100ß-DsRed and OMP-ZsGreen, to visualise olfactory ensheathing cells (OECs) and Grueneberg olfactory neurons, respectively. Within the epithelium, Grueneberg organ OECs were in direct contact with Grueneberg organ neuron cell bodies. Individual axons from the neurons initially grew over the surface of the OECs before forming larger fascicles consisting of numerous axons and OECs. Considering the location of the Grueneberg organ so close to the external environment, it may be that the Grueneberg neurons are likely to be subject to damage suggesting that as in other olfactory regions there is a capacity for recovery after injury. Here, we used a well characterised model of olfactory nervous system injury, unilateral bulbectomy, to determine whether Grueneberg organ neurons degenerate after injury. We found that Grueneberg organ neurons degenerated in response to the axotomy, yet by 11 days post injury neurons and/or axons were detected again within the epithelium. Our results demonstrate that while Grueneberg organ neurons and glia have a distinct relationship in the epithelium, they have largely similar characteristics to that of the main olfactory neurons and glia

The neuroinflammation biomarker translocator protein (TSPO) plays a role in sucrose overconsumption in mice

Sugar overconsumption is a major cause of obesity. Existing pharmacotherapeutics for obesity have failed to stop the growing prevalence of this disease. Emerging research suggests that neuroinflammation mediates the pathogenesis of diet-induced obesity. Neuroinflammation has been implicated in many neuropathologies by measuring the expression of a neuroinflammatory biomarker – the translocator protein (TSPO). Etifoxine, a TSPO partial agonist and clinically used anxiolytic, is one of the few antipsychotic medications that does not cause weight gain. Given the proposed role of TSPO in anorexia and obesity, we hypothesized that etifoxine may have therapeutic potential in the treatment of sugar overconsumption. In this study, C57/BL6 mice consumed a 25% sucrose solution for 12 weeks starting at 6 weeks of age following common a common addiction consumption paradigm. After 6 and 12 weeks of sucrose consumption, two groups of 12 mice were treated with either intraperitoneal injections of etifoxine (20 mg/kg or 50 mg/kg) or the TSPO specific antagonist PK11195 (1mg/kg or 10 mg/kg). A single injection of 50 mg/kg etifoxine reduced sucrose consumption both 30 mins and 2 hrs into the drinking session, after 4 and 12 weeks of sucrose consumption. In contrast, PK11195 treatment did not alter sucrose consumption at either dose after 4 or 12 weeks of sucrose consumption. To assess if the effect seen from etifoxine is TSPO specific, a third group of 12 mice were pre-treated with 10 mg/kg PK11195 30 minutes prior to receiving a 50 mg/kg etifoxine injection. Pre-treatment with PK11195 blocked the consumption-reducing effect of etifoxine. Together, these results demonstrate that etifoxine acts through TSPO to reduce sucrose consumption in sucrose overconsuming mice. Given the well-documented safety profile of etifoxine, this study provides preliminary supporting evidence for its potential repurposing as an anti-obesity medication

EdU, a new thymidine analogue for labelling proliferating cells in the nervous system

The standard method of labelling proliferating cells uses the thymidine analogue, bromodeoxyuridine (BrdU), which incorporates into the DNA during S-phase of the cell cycle. A disadvantage of this method is that the immunochemical processing requires pre-treatment of the cells and tissue with heat or acid to reveal the antigen. This pre-treatment reduces reliability of the method and degrades the specimen, reducing the ability for multiple immuno-fluorescence labelling at high resolution. We report here the utility of a novel thymidine analogue, ethynyl deoxyuridine (EdU), detected with a fluorescent azide via the “click” chemistry reaction (the Huisgen 1,3-dipolar cycloaddition reaction of an organic azide to a terminal acetylene). The detection of EdU requires no heat or acid treatment and the incorporated EdU is covalently conjugated to fluorescent probe. The reaction is quick and compatible with fluorescence immunochemistry and other fluorescent probes. We show here that EdU is non-toxic in vitro and in vivo and can be used in place of BrdU to label cells during neurogenesis and the progeny identified at least 30 days later. The fluorescent labelling of EdU, markedly improves the detection of proliferating cells and allows concurrent high resolution fluorescence immunochemistry