Cellular and nuclear morphology…and calcium signaling: revealing the interplay between structure and function

Poster presentation: Calcium plays a pivotal role in relaying electrical signals of the cell to subcellular compartments, such as the nucleus. Since this one ion type is used by the cell for many processes a neuron needs to establish finely tuned calcium pathways in order to be able to differentiate multiple tasks, [1-3]. While it is known that neurons can actively change their shape upon neuronal activity, [4-7], we here present novel findings of activity-regulated nuclear morphology, [8,9]. With the help of an experimental and computational modeling approach, we show that hippocampal neurons can change the previously spherical shape of their nuclei to complex and infolded morphologies. This morphology regulation is demonstrated to be regulated by NMDA-receptor gated calcium, while synaptic and extra-synaptic NMDA-receptors elicit opposing effects on nuclear morphology, [8]. The structural alterations of the cell nucleus have significant effects on nuclear calcium dynamics. Compartmentalization of the nucleus, due to membrane infoldings, changes calcium frequencies, amplitudes and spatial distributions, [8,10]. Since these parameters have been shown to control downstream events towards gene transcription, [11,12], the results elucidate the cellular control of nuclear function with the help of morphology modulation. With respect to processes downstream of calcium, we show that histone H3 phosphorylation is closely linked to nuclear morphology. Investigating the nuclear morphologies of hippocampal neurons, two major classes were identified [9,10]. One class contains non-infolded nuclei that have the function of calcium signal integrators, while the other class contains highly infolded nuclei, which function as frequency detectors of nuclear calcium, [10]. Extending this interdisciplinary approach of investigating structure/function relationships in neurons, the effects of cellular morphology – as well as the morphology of the endoplasmic reticulum and other organelles – on neuronal calcium signals is currently being investigated. This endeavor makes use of highly detailed, three-dimensional models of neuronal calcium dynamics, including the three-dimensional morphology of the cell and its organelles

In Vivo Imaging of Transplanted Islets with ^(64)Cu-DO3A-VS-Cys^(40)-Exendin-4 by Targeting GLP-1 Receptor

Glucagon-like peptide 1 receptor (GLP-1R) is highly expressed in pancreatic islets, especially on β-cells. Therefore, a properly labeled ligand that binds to GLP-1R could be used for in vivo pancreatic islet imaging. Because native GLP-1 is degraded rapidly by dipeptidyl peptidase-IV (DPP-IV), a more stable agonist of GLP-1 such as Exendin-4 is a preferred imaging agent. In this study, DO3A-VS-Cys^(40)-Exendin-4 was prepared through the conjugation of DO3A-VS with Cys^(40)-Exendin-4. The in vitro binding affinity of DO3A-VS-Cys^(40)-Exendin-4 was evaluated in INS-1 cells, which overexpress GLP-1R. After ^(64)Cu labeling, biodistribution studies and microPET imaging of ^(64)Cu-DO3A-VS-Cys^(40)-Exendin-4 were performed on both subcutaneous INS-1 tumors and islet transplantation models. The subcutaneous INS-1 tumor was clearly visualized with microPET imaging after the injection of ^(64)Cu-DO3A-VS-Cys^(40)-Exendin-4. GLP-1R positive organs, such as pancreas and lung, showed high uptake. Tumor uptake was saturable, reduced dramatically by a 20-fold excess of unlabeled Exendin-4. In the intraportal islet transplantation models, ^(64)Cu-DO3A-VS-Cys^(40)-Exendin-4 demonstrated almost two times higher uptake compared with normal mice. ^(64)Cu-DO3A-VS-Cys^(40)-Exendin-4 demonstrated persistent and specific uptake in the mouse pancreas, the subcutaneous insulinoma mouse model, and the intraportal human islet transplantation mouse model. This novel PET probe may be suitable for in vivo pancreatic islets imaging in the human

Food availability drives plastic self-repair response in a basal metazoan- case study on the ctenophore Mnemiopsis leidyi A. Agassiz 1865

Many marine invertebrates including ctenophores are capable of extensive body regeneration when injured. However, as for the invasive ctenophore Mnemiopsis leidyi, there is a constant subportion of individuals not undergoing whole body regeneration but forming functionally stable half-animals instead. Yet, the driving factors of this phenomenon have not been addressed so far. This study sheds new light on how differences in food availability affect self-repair choice and regeneration success in cydippid larvae of M. leidyi. As expected, high food availability favored whole-body regeneration. However, under low food conditions half-animals became the preferential self-repair mode. Remarkably, both regenerating and half-animals showed very similar survival chances under respective food quantities. As a consequence of impaired food uptake after injury, degeneration of the digestive system would often occur indicating limited energy storage capacities. Taken together, this indicates that half-animals may represent an alternative energy-saving trajectory which implies self-repair plasticity as an adaptive trade-off between high regeneration costs and low energy storage capacities. We conclude that self-repair plasticity could lead to higher population fitness of ctenophores under adverse conditions such as in ships’ ballast water tanks which is postulated to be the major vector source for the species’ spreading around the globe

Phytochemical characterization of Tabernanthe iboga root bark and its effects on dysfunctional metabolism and cognitive performance in high-fat-fed C57BL/6J mice

Preparations of the root bark of Tabernanthe iboga have long been used in Central and West African traditional medicine to combat fatigue, as a neuro-stimulant in rituals, and for treatment of diabetes. The principal alkaloid of T. iboga, ibogaine, has attracted attention in many countries around the world for providing relief for opioid craving in drug addicts. Using a plant metabolomics approach, we detected five phenolic compounds, including 3- O-caffeoylquinic acid, and 30 alkaloids, seven of which were previously reported from T. iboga root bark. Following a report that iboga extracts contain insulinotropic agents, we aimed to determine the potential alleviating effects of the water extract of iboga root bark on high-fat diet (HFD)-induced hyperglycemia as well as its effects on cognitive function in male C57BL/6J mice. Feeding a HFD to mice for 10 weeks produced manifestations of metabolic syndrome such as increased body weight and increased plasma levels of glucose, triacylglycerols, total cholesterol, LDL-cholesterol, insulin, leptin, and pro-inflammatory mediators (IL-6, MCP-1, ICAM-1), as compared to mice fed a low-fat diet (LFD). Supplementation of HFD with iboga extract at ibogaine doses of 0.83 (low) and 2.07 (high) mg/kg/day did not improve these HFD-induced metabolic effects except for a reduction of plasma MCP-1 in the low dose group, indicative of an anti-inflammatory effect. When the HFD mice were tested in the water maze, the high-dose iboga extract caused hippocampus-dependent impairments in spatial learning and memory, as compared to mice receiving only a HFD.Peer reviewedFinal Published versio

Muscarinic M1 receptors modulate endotoxemia-induced loss of synaptic plasticity

Septic encephalopathy is associated with rapid deterioration of cortical functions. Using magnetic resonance imaging (MRI) we detected functional abnormalities in the hippocampal formation of patients with septic delirium. Hippocampal dysfunction was further investigated in an animal model for sepsis using lipopolysaccharide (LPS) injections to induce endotoxemia in rats, followed by electrophysiological recordings in brain slices. Endotoxemia induced a deficit in long term potentiation which was completely reversed by apamin, a blocker of small conductance calcium-activated potassium (SK) channels, and partly restored by treatment with physostigmine (eserine), an acetylcholinesterase inhibitor, or TBPB, a selective M1 muscarinic acetylcholine receptor agonist. These results suggest a novel role for SK channels in the etiology of endotoxemia and explain why boosting cholinergic function restores deficits in synaptic plasticity. Drugs which enhance cholinergic or M1 activity in the brain may prove beneficial in treatment of septic delirium in the intensive care unit

Quantitative, Simultaneous PET/MRI for Intratumoral Imaging with an MRI-Compatible PET Scanner

Noninvasive methods are needed to explore the heterogeneous tumor microenvironment and its modulation by therapy. Hybrid PET/MRI systems are being developed for small-animal and clinical use. The advantage of these integrated systems depends on their ability to provide MR images that are spatially coincident with simultaneously acquired PET images, allowing combined functional MRI and PET studies of intratissue heterogeneity. Although much effort has been devoted to developing this new technology, the issue of quantitative and spatial fidelity of PET images from hybrid PET/MRI systems to the tissues imaged has received little attention. Here, we evaluated the ability of a first-generation, small-animal MRI-compatible PET scanner to accurately depict heterogeneous patterns of radiotracer uptake in tumors. Methods: Quantitative imaging characteristics of the MRI-compatible PET (PET/MRI) scanner were evaluated with phantoms using calibration coefficients derived from a mouse-sized linearity phantom. PET performance was compared with a commercial small-animal PET system and autoradiography in tumor-bearing mice. Pixel and structure-based similarity metrics were used to evaluate image concordance among modalities. Feasibility of simultaneous PET/MRI functional imaging of tumors was explored by following ^(64)Cu-labeled antibody uptake in relation to diffusion MRI using cooccurrence matrix analysis. Results: The PET/MRI scanner showed stable and linear response. Activity concentration recovery values (measured and true activity concentration) calculated for 4-mm-diameter rods within linearity and uniform activity rod phantoms were near unity (0.97 ± 0.06 and 1.03 ± 0.03, respectively). Intratumoral uptake patterns for both ^(18)F-FDG and a ^(64)Cu-antibody acquired using the PET/MRI scanner and small-animal PET were highly correlated with autoradiography (r > 0.99) and with each other (r = 0.97 ± 0.01). On the basis of these data, we performed a preliminary study comparing diffusion MRI and radiolabeled antibody uptake patterns over time and visualized movement of antibodies from the vascular space into the tumor mass. Conclusion: The MRI-compatible PET scanner provided tumor images that were quantitatively accurate and spatially concordant with autoradiography and the small-animal PET examination. Cooccurrence matrix approaches enabled effective analysis of multimodal image sets. These observations confirm the ability of the current simultaneous PET/MRI system to provide accurate observations of intratumoral function and serve as a benchmark for future evaluations of hybrid instrumentation

Phytochemical Characterisation and Effects of the Native Plant Tabernanthe iboga in Models of Diabetes

Preparations of the root bark of Gabonese plants, T. iboga and G. tessmannii have long been used in Central and West African traditional medicine for the treatment of diabetes. In this thesis phytochemical characterisation of T. iboga aqueous extract revealed the presence of 23 unknown alkaloids and 7 already known alkaloids, including ibogaine, the main indole alkaloid found in T. iboga. For the first time, phenolic compounds have been identified in significant quantity in T. Iboga, the most prevalent being 3-O-CQA. The therapeutic potential of these constituents in diabetes remains to be determined. The aqueous extracts of these Gabonese plants for the potential to improve insulin release was examined. T. iboga (1 μg/ml) and G. tessmannii (70 μg/ml) demonstrated insulinotropic effects in isolated rat pancreatic islets at non-stimulatory and stimulatory glucose concentrations (2.8 and 11.1 mM, for T. iboga and 2.8 to 16.7 mM, for G. tessmannii), similar to the insulin secretagogue, tolbutamide (200 μM). An additive effect on glucose-induced insulin release was observed for both plant aqueous extracts in the presence of tolbutamide (200 μM). The mechanism of action of the insulinotropic effect of T. iboga (1 μg/ml) was explored through the use of a σ2 receptor antagonist, SM-21 (1 and 10 μM) which significantly blocked T. iboga (1 μg/ml) insulin potentiation at non-stimulatory and stimulatory glucose concentrations (2.8 and 11 mM) indicating these effects were seen to be partly through σ2 receptors. T. iboga (50, 100 and 200 mg/kg) when administered daily over 28 days demonstrated no observable toxicity and exerted hypoglycaemic activity on glucose tolerance in healthy rats. Feeding a 10% fructose solution over 2 weeks combined with a low intraperitoneal dose of streptozotocin (STZ, 40 mg/kg) to rats led to the development of a T2D compared with T1D rats over 4 weeks. T. iboga at the lowest daily dose of 50 mg/kg significantly improved hyperglycaemia after 3 and 4 weeks of treatment compared with glibenclamide. After 2 and 4 weeks daily administration of T. Iboga (50 and 200 mg/kg) glucose tolerance over a 2 hour fasted glucose load was improved compared with T2D rats treated with vehicle alone. Feeding a high fat diet (HFD) to mice for 10 weeks produced manifestations of metabolic syndrome and T2D, as compared to mice fed a low-fat diet (LFD). Supplementation of HFD with T. iboga aqueous extract at ibogaine doses of 0.83 (low) and 2.07 (high) mg/kg/day did not improve these HFD-induced metabolic effects except for a reduction of plasma MCP-1 in the low dose group, indicative of an anti-inflammatory effect. Overall, T. iboga and G. tessmannii aqueous extracts have demonstrated insulin potentiation, hypoglycaemic and anti-hyperglycaemic activities. However, further investigations are needed to validate their safe use for the management of DM in Gabon

Image informatics strategies for deciphering neuronal network connectivity

Brain function relies on an intricate network of highly dynamic neuronal connections that rewires dramatically under the impulse of various external cues and pathological conditions. Among the neuronal structures that show morphologi- cal plasticity are neurites, synapses, dendritic spines and even nuclei. This structural remodelling is directly connected with functional changes such as intercellular com- munication and the associated calcium-bursting behaviour. In vitro cultured neu- ronal networks are valuable models for studying these morpho-functional changes. Owing to the automation and standardisation of both image acquisition and image analysis, it has become possible to extract statistically relevant readout from such networks. Here, we focus on the current state-of-the-art in image informatics that enables quantitative microscopic interrogation of neuronal networks. We describe the major correlates of neuronal connectivity and present workflows for analysing them. Finally, we provide an outlook on the challenges that remain to be addressed, and discuss how imaging algorithms can be extended beyond in vitro imaging studies