Nested partnerships and interdisciplinary science: from the National Medical Cyclotron to the research cyclotron of the National Imaging Facility

In Australia, the routine use of medical isotopes produced by a prototype cyclotron for diagnostic imaging commenced in the early 1990s. Since then, the mainly clinically focused imaging in nuclear medicine has become a broader and more interdisciplinary endeavour. As 'molecular imaging', it has become a field that supports a wide range of basic, translational and clinical research and draws in skills from many areas, including physics, chemistry, engineering, biology and medicine. Such growth has been accompanied by the emergence of scientific collaborations well beyond individual institutions. This paper provides the historical context to the former National Medical Cyclotron (NMC) facility (1992-2009) at Camperdown, Sydney and the subsequent partnerships that led to its refurbishment as the new site of the National Imaging Facility (NIF) Cyclotron, a flagship research facility enabled by the National Collaborative Research Infrastructure Strategy (NCRIS). It is now the centrepiece of a physical research infrastructure as well as a growing network of collaborations that open up access to medical isotopes for research and clinical applications across Australia to new users and applications. It is also a contemporary example of how science has moved from individual scholarly endeavour to highly networked activity. The funding model initiated through NCRIS included shared funding, funding leveraging and in-kind contributions primarily for the establishment of the large instrument and laboratory infrastructure rather than their operational costs. Here, we illustrate how partnership arrangements emerged at institutional, state and national level and how they address the task of providing open access to, and sustainable operation of, a major piece of research infrastructure that spans multiple institutions. © The Royal Society of NSW

Testicular translocator protein expression is differentially altered by synthetic cannabinoid HU210 in adult and adolescent Rats

Objective: The translocator protein (TSPO) has been implicated in numerous functions including steroid production and regulation of stress and anxiety. Cannabinoids have been shown to reduce plasma testosterone levels and alter anxiety levels. The aim of the present study was to determine whether the synthetic cannabinoid HU210 is able to regulate TSPO expression in several peripheral organs. Methods: HU210 (100 μg/kg) was administered intraperitoneally to both adult and adolescent male ratsfor 14 days. TSPO receptor expression in several organs, including the liver, spleen, kidneys and testes, was quantified by membrane receptor binding using the selective radiolig and, PK11195. In cases where receptor binding data indicated significant cannabinoid-induced differences, further RT-qPCR was carried out to determine the transcriptional regulation of the TSPO gene. Additionally, film-autography was used to identify potential changes in the spatial distribution of the TSPO tissue binding sites. Results: Results indicate that HU210 induces significant reductions in testicular TSPO expression in adult but not adolescent rats. No changes were found in other organs examined. These results are consistent with the previously observed effects of cannabinoids on testosterone production and a presumed role for TSPO in steroidogenesis. Conclusions: Overall, these results suggest that cannabinoids may alter testosterone production by altering the expression of testicular TSPO and that the alteration of TSPO occurs in an age-dependent manner.© 2014 Chan RHY, et al

Intrinsic synergistic-topological mechanism versus synergistic-topological matrix in microtubule self-organization

Background In this body of work we investigate the synergistic-topological relationship during self-organization of the microtubule fiber in vitro, which is composed of straight, axially shifted and non-shifted, acentrosomal microtubules under crowded conditions. Methods We used electron microscopy to observe morphological details of ordered straight microtubules. This included the observation of the differences in length distribution between microtubules in ordered and non-ordered phases followed by the observation of the formation of interface gaps between axially shifted and ordered microtubules. We performed calculations to confirm that the principle of summation of pairwise electrostatic forces act between neighboring microtubules all their entire length. Results We have shown that the self-organization of a microtubule fiber imposes a variety of topological restrictions onto its constituting components: (a) tips of axially shifted neighboring microtubules are not in direct contact but rather create an ‘interface gap’; (b) fibers are always composed of a restricted number of microtubules at given solution conditions; (c) the average length of microtubules that constitute a fiber is always shorter than that of microtubules outside a fiber; (d) the length distribution of microtubules that constitute a fiber is narrower than that of microtubules outside a fiber and this effect is more pronounced at higher GTP-tubulin concentrations; (e) a cooperative motion of fiber microtubules due to actualization of the summation principle of pairwise electrostatic forces; (f) appearance of local GTP-tubulin depletion immediately in front of the tips of fiber microtubules. Conclusion Overall our data indicate that under crowded conditions in vitro, the self-organization of a microtubule fiber is governed by an intrinsic synergistic-topological mechanism, which in conjunction with the topological changes, GTP-tubulin depletion, and cooperative motion of fiber constituting microtubules, may generate and maintain a ‘synergistic-topological matrix’. Failure of the mechanism to form biologically feasible microtubule synergistic-topological matrix may, per se, precondition tumorigenesis. © 2014 BioMed Central Lt

The 18 kDa translocator protein, microglia and neuroinflammation

The 18 kDa translocator protein (TSPO), previously known as the peripheral benzodiazepine receptor, is expressed in the injured brain. It has become known as an imaging marker of “neuroinflammation” indicating active disease, and is best interpreted as a nondiagnostic biomarker and disease staging tool that refers to histopathology rather than disease etiology. The therapeutic potential of TSPO as a drug target is mostly based on the understanding that it is an outer mitochondrial membrane protein required for the translocation of cholesterol, which thus regulates the rate of steroid synthesis. This pivotal role together with the evolutionary conservation of TSPO has underpinned the belief that any loss or mutation of TSPO should be associated with significant physiological deficits or be outright incompatible with life. However, against prediction, full Tspo knockout mice are viable and across their lifespan do not show the phenotype expected if cholesterol transport and steroid synthesis were significantly impaired. Thus, the “translocation” function of TSPO remains to be better substantiated. Here, we discuss the literature before and after the introduction of the new nomenclature for TSPO and review some of the newer findings. In light of the controversy surrounding the function of TSPO, we emphasize the continued importance of identifying compounds with confirmed selectivity and suggest that TSPO expression is analyzed within specific disease contexts rather than merely equated with the reified concept of “neuroinflammation.” © 2014 The Authors© 2014 The Authors. Brain Pathology published by John Wiley & Sons Ltd on behalf of International Society of Neuropathology. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited

COX-2, CB2 and P2X7-immunoreactivities are increased in activated microglial cells/macrophages of multiple sclerosis and amyotrophic lateral sclerosis spinal cord

BACKGROUND: While multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS) are primarily inflammatory and degenerative disorders respectively, there is increasing evidence for shared cellular mechanisms that may affect disease progression, particularly glial responses. Cyclooxygenase 2 (COX-2) inhibition prolongs survival and cannabinoids ameliorate progression of clinical disease in animal models of ALS and MS respectively, but the mechanism is uncertain. Therefore, three key molecules known to be expressed in activated microglial cells/macrophages, COX-2, CB2 and P2X7, which plays a role in inflammatory cascades, were studied in MS and ALS post-mortem human spinal cord. METHODS: Frozen human post mortem spinal cord specimens, controls (n = 12), ALS (n = 9) and MS (n = 19), were available for study by immunocytochemistry and Western blotting, using specific antibodies to COX-2, CB2 and P2X7, and markers of microglial cells/macrophages (CD 68, ferritin). In addition, autoradiography for peripheral benzodiazepine binding sites was performed on some spinal cord sections using [3H] (R)-PK11195, a marker of activated microglial cells/macrophages. Results of immunostaining and Western blotting were quantified by computerized image and optical density analysis respectively. RESULTS: In control spinal cord, few small microglial cells/macrophages-like COX-2-immunoreactive cells, mostly bipolar with short processes, were scattered throughout the tissue, whilst MS and ALS specimens had significantly greater density of such cells with longer processes in affected regions, by image analysis. Inflammatory cell marker CD68-immunoreactivity, [3H] (R)-PK11195 autoradiography, and double-staining against ferritin confirmed increased production of COX-2 by activated microglial cells/macrophages. An expected 70-kDa band was seen by Western blotting which was significantly increased in MS spinal cord. There was good correlation between the COX-2 immunostaining and optical density of the COX-2 70-kDa band in the MS group (r = 0.89, P = 0.0011, n = 10). MS and ALS specimens also had significantly greater density of P2X7 and CB2-immunoreactive microglial cells/macrophages in affected regions. CONCLUSION: It is hypothesized that the known increase of lesion-associated extracellular ATP contributes via P2X7 activation to release IL-1 beta which in turn induces COX-2 and downstream pathogenic mediators. Selective CNS-penetrant COX-2 and P2X7 inhibitors and CB2 specific agonists deserve evaluation in the progression of MS and ALS

An Alteration in the Lateral Geniculate Nucleus of Experimental Glaucoma Monkeys: In vivo Positron Emission Tomography Imaging of Glial Activation

We examined lateral geniculate nucleus (LGN) degeneration as an indicator for possible diagnosis of glaucoma in experimental glaucoma monkeys using positron emission tomography (PET). Chronic intraocular pressure (IOP) elevation was induced by laser trabeculoplasty in the left eyes of 5 cynomolgus monkeys. Glial cell activation was detected by PET imaging with [11C]PK11195, a PET ligand for peripheral-type benzodiazepine receptor (PBR), before and at 4 weeks after laser treatment (moderate glaucoma stage). At mild, moderate, and advanced experimental glaucoma stages (classified by histological changes based on the extent of axonal loss), brains were stained with cresyl violet, or antibodies against PBR, Iba-1 (a microglial marker), and GFAP (an activated astrocyte marker). In laser-treated eyes, IOP was persistently elevated throughout all observation periods. PET imaging showed increased [11C]PK11195 binding potential in the bilateral LGN at 4 weeks after laser treatment; the increase in the ipsilateral LGN was statistically significant (P<0.05, n = 4). Immunostaining showed bilateral activations of microglia and astrocytes in LGN layers receiving input from the laser-treated eye. PBR-positive cells were observed in LGN layers receiving input from laser-treated eye at all experimental glaucoma stages including the mild glaucoma stage and their localization coincided with Iba-1 positive microglia and GFAP-positive astrocytes. These data suggest that glial activation occurs in the LGN at a mild glaucoma stage, and that the LGN degeneration could be detected by a PET imaging with [11C]PK11195 during the moderate experimental glaucoma stage after unilateral ocular hypertension. Therefore, activated glial markers such as PBR in the LGN may be useful in noninvasive molecular imaging for diagnosis of glaucoma

The PPAR-gamma agonist pioglitazone protects cortical neurons from inflammatory mediators via improvement in peroxisomal function

<p>Abstract</p> <p>Background</p> <p>Inflammation is known to play a pivotal role in mediating neuronal damage and axonal injury in a variety of neurodegenerative disorders. Among the range of inflammatory mediators, nitric oxide and hydrogen peroxide are potent neurotoxic agents. Recent evidence has suggested that oligodendrocyte peroxisomes may play an important role in protecting neurons from inflammatory damage.</p> <p>Methods</p> <p>To assess the influence of peroxisomal activation on nitric oxide mediated neurotoxicity, we investigated the effects of the peroxisomal proliferator activated receptor (PPAR) gamma agonist, pioglitazone in primary cortical neurons that were either exposed to a nitric oxide donor or co-cultured with activated microglia.</p> <p>Results</p> <p>Pioglitazone protected neurons and axons against both nitric-oxide donor-induced and microglia-derived nitric oxide-induced toxicity. Moreover, cortical neurons treated with this compound showed a significant increase in the protein and gene expression of PPAR-gamma, which was associated with a concomitant increase in the enzymatic activity of catalase. In addition, the protection of neurons and axons against hydrogen peroxide-induced toxicity afforded by pioglitazone appeared to be dependent on catalase.</p> <p>Conclusions</p> <p>Collectively, these observations provide evidence that modulation of PPAR-gamma activity and peroxisomal function by pioglitazone attenuates both NO and hydrogen peroxide-mediated neuronal and axonal damage suggesting a new therapeutic approach to protect against neurodegenerative changes associated with neuroinflammation.</p

Different patterns of Ca2+ signals are induced by low compared to high concentrations of P2Y agonists in microglia

Brain-resident macrophages (microglia) are key cellular elements in the preservation of tissue integrity. On the other hand, they can also contribute to the development of pathological events by causing an extensive and inappropriate inflammatory response. A growing number of reports indicate the involvement of nucleotides in the control of microglial functions. With this study on P2Y receptors in rat microglia, we want to contribute to the definition of their expression profile and to the characterisation of their signalling mechanisms leading to Ca2+ movements. Endogenous nucleotides, when applied at a concentration of 100 μM, elicited robust Ca2+ transients, thanks to a panel of metabotropic receptors comprising mainly P2Y2, P2Y6 and P2Y12 subtypes. The involvement of P2Y12 receptors in Ca2+ responses induced by adenine nucleotides was confirmed by the pharmacological and pertussis toxin sensitivity of the response induced by adenosine diphosphate (ADP). Beside the G protein involved, Gi and Gq respectively, adenine and uracil nucleotides differed also for induction by the latter of a capacitative Ca2+ plateau. Moreover, when applied at low (sub-micromolar) concentrations with a long-lasting challenge, uracil nucleotides elicited oscillatory Ca2+ changes with low frequency of occurrence (≤ 1 min−), sometimes superimposed to an extracellular Ca2+-dependent sustained Ca2+ rise. We conclude that different patterns of Ca2+ transients are induced by low (i.e., oscillatory Ca2+ activity) compared to high (i.e., fast release followed by sustained raise) concentrations of nucleotides, which can suggest different roles played by receptor stimulation depending not only on the type but also on the concentration of nucleotides