Cannabinoid type 2 receptor-mediated cell type-specific self-inhibition in hippocampal and cortical neurons

Endogenous cannabinoids are lipid-based ligands of the main cannabinoid receptors type 1 and 2 (CB1R and CB2R). In contrast to the well-studied effects of CB1Rs, much less is known about the physiological role of CB2Rs in the central nervous system (CNS). In fact, CB2Rs were considered as peripheral cannabinoid receptors representing the complementary cannabinoid receptor to the CB1R in the CNS. However recent pharmacological, behavioral and genetic studies have determined the presence of functional CB2Rs in the brain and their involvement in various physiological and pathological conditions. Endocannabinoids are produced in an activity-dependent manner. Next to their well-described role as retrograde modulators of synaptic transmission, endocannabinoids also mediate a cell-autonomous slow self-inhibition (SSI). Action potential-driven endocannabinoid production followed by binding to cannabinoid receptors induces a long-lasting hyperpolarization of the membrane potential, rendering the cell less excitable. Several studies described different endocannabinoid receptors and cellular mechanisms by which SSI is implemented in different cell types and brain areas: While pyramidal cells and interneurons in the somatosensory cortex were reported to mediate SSI via CB1Rs and G protein-coupled inward rectifying K+ (GIRK) channels, hippocampal principal cells were shown to mediate SSI in a CB2R mediated and input-resistance-independent manner. However, the molecular mechanism by which the long-lasting hyperpolarization is implemented was not clear. During my thesis I was part of a team that demonstrated that hippocampal SSI is mediated via activation of the Na+/bicarbonate cotransporter. In order to get further insight in the occurrence of SSI in different classes of neurons, we analyzed the presence of SSI in one type of hippocampal interneuron and showed that oriens-lacunosum moleculare (OLM) interneurons do not express SSI. Further, we investigate SSI in different neuron types of layer 2/3 in the primary somatosensory cortex and show that regular firing cells express SSI in contrast to fast-spiking interneurons. Trains of action potentials induced a long-lasting hyperpolarization that was accompanied by a change in input resistance due to GIRK channel activation. By using cannabinoid receptor-specific pharmacology as well as transgenic mice lacking either CB1Rs or CB2Rs, we demonstrate that this effect is mediated by CB2R activation. Taken together, hippocampal and cortical SSI both represent a CB2R-dependent mechanism; however the underlying mechanism by which the hyperpolarization is implemented differs between the different brain regions. By describing an additional cellular mechanism for SSI induction, these findings add further insights on the physiological role of CB2Rs and expand our knowledge about cell type-specific differential cannabinoid signaling. Moreover, these findings suggest CB2Rs as a promising target for therapeutic approaches

A Universal Footstep Planning Methodology for Continuous Walking in Challenging Terrain Applicable to Different Types of Legged Robots

In recent years, the capabilities of legged locomotion controllers have been significantly advanced enabling them to traverse basic types of uneven terrain without visual perception. However, safely and autonomously traversing longer distances over difficult uneven terrain requires appropriate motion planning using online collected environmental knowledge. In this paper, we present such a novel methodology for generic closed-loop preceding horizon footstep planning that enables legged robots equipped with capable locomotion controllers to autonomously traverse previously unknown terrain while continuously walking long distances. Hereby, our approach addresses the challenge of online terrain perception and soft real-time footstep planning. The proposed new formulation of the search-based planning problem makes no specific assumptions about the robot kinematics (e.g. number of legs) or the used locomotion control schemes. Therefore, it can be applied to a broad range of different types of legged robots. Unlike current methods, the proposed new framework can optionally consider the floating base as part of the state-space. It is possible to configure the complexity of the planner online, from efficiently solving tasks in flat terrain to using non-contiguous contacts in highly challenging terrain. Finally, the presented methodology is successfully applied and evaluated in virtual and real experiments on state of the art bipedal, quadrupedal, and a novel eight-legged robot

Developing effective catalyst for mine methane conversion to hydrogen-containing gas

The Ni/Ce[1-x]La[x]O[y] and Ni/Ce[1-x]La[x]O[y]/Al[2]O[3] catalysts (x=0-1) were prepared for the mine methane conversion to hydrogen-containing gas. The influence of the support composition on the physicochemical characteristics of the catalysts and their activity in autothermal reforming of methane (ATR of CH[4]) was studied. It was shown that the dispersion of the Ni active component was enhanced with growth of molar ratio of La in the support composition that leads to the increase in catalyst stability in ATR of CH[4]. The Ni/Ce[1-x]La[x]O[y]/Al[2]O[3] catalysts in comparison to the Ni/Ce[1]-xLa[x]O[y] provide higher H[2] yield: at 850oC it is equal to 65-75%

Prediction of Physical Frailty in Orthogeriatric Patients Using Sensor Insole–Based Gait Analysis and Machine Learning Algorithms: Cross-sectional Study

Background: Assessment of the physical frailty of older patients is of great importance in many medical disciplines to be able to implement individualized therapies. For physical tests, time is usually used as the only objective measure. To record other objective factors, modern wearables offer great potential for generating valid data and integrating the data into medical decision-making. Objective: The aim of this study was to compare the predictive value of insole data, which were collected during the Timed-Up-and-Go (TUG) test, to the benchmark standard questionnaire for sarcopenia (SARC-F: strength, assistance with walking, rising from a chair, climbing stairs, and falls) and physical assessment (TUG test) for evaluating physical frailty, defined by the Short Physical Performance Battery (SPPB), using machine learning algorithms. Methods: This cross-sectional study included patients aged >60 years with independent ambulation and no mental or neurological impairment. A comprehensive set of parameters associated with physical frailty were assessed, including body composition, questionnaires (European Quality of Life 5-dimension [EQ 5D 5L], SARC-F), and physical performance tests (SPPB, TUG), along with digital sensor insole gait parameters collected during the TUG test. Physical frailty was defined as an SPPB score≤8. Advanced statistics, including random forest (RF) feature selection and machine learning algorithms (K-nearest neighbor [KNN] and RF) were used to compare the diagnostic value of these parameters to identify patients with physical frailty. Results: Classified by the SPPB, 23 of the 57 eligible patients were defined as having physical frailty. Several gait parameters were significantly different between the two groups (with and without physical frailty). The area under the receiver operating characteristic curve (AUROC) of the TUG test was superior to that of the SARC-F (0.862 vs 0.639). The recursive feature elimination algorithm identified 9 parameters, 8 of which were digital insole gait parameters. Both the KNN and RF algorithms trained with these parameters resulted in excellent results (AUROC of 0.801 and 0.919, respectively). Conclusions: A gait analysis based on machine learning algorithms using sensor soles is superior to the SARC-F and the TUG test to identify physical frailty in orthogeriatric patients

Reduced awareness for osteoporosis in hip fracture patients compared to elderly patients undergoing elective hip replacement

Background: Osteoporotic fractures are associated with a loss of quality of life, but only few patients receive an appropriate therapy. Therefore, the present study aims to investigate the awareness of musculoskeletal patients to participate in osteoporosis assessment and to evaluate whether there are significant differences between acute care patients treated for major fractures of the hip compared to elective patients treated for hip joint replacement.; Methods: From May 2015 to December 2016 patients who were undergoing surgical treatment for proximal femur fracture or total hip replacement due to osteoarthritis and were at risk for an underlying osteoporosis (female > 60 and male > 70 years) were included in the study and asked to complete a questionnaire assessing the awareness for an underlying osteoporosis. ASA Score, FRAX Score, and demographic information have also been examined. Results: In total 268 patients (female = 194 (72.0%)/male = 74 (28%)), mean age 77.7 years (±7.7) undergoing hip surgery were included. Of these, 118 were treated for fracture-related etiology and 150 underwent total hip arthroplasty in an elective care setting. Patients were interviewed about their need for osteoporosis examination during hospitalization. Overall, 76 of 150 patients receiving elective care (50.7%) considered that an examination was necessary, whereas in proximal femur fracture patients the awareness was lower, and the disease osteoporosis was assessed as threatening by significantly fewer newly fractured patients. By comparison, patients undergoing trauma surgery had a considerably greater risk of developing another osteoporotic fracture than patients undergoing elective surgery determined by the FRAX(®) Score (p ≤ 0.001).; Conclusions: The patients’ motivation to endure additional osteoporosis diagnostic testing is notoriously low and needs to be increased. Patients who underwent acute care surgery for a fragility proximal femur fracture, although acutely affected by the potential consequences of underlying osteoporosis, showed lower awareness than the elective comparison population that was also on average 6.1 years younger. Although elective patients were younger and at a lower risk, they seemed to be much more willing to undergo further osteoporosis assessment. In order to better identify and care for patients at risk, interventions such as effective screening, early initiation of osteoporosis therapy in the inpatient setting and a fracture liaison service are important measures

Branch point strength controls species-specific CAMK2B alternative splicing and regulates LTP

Regulation and functionality of species-specific alternative splicing has remained enigmatic to the present date. Calcium/calmodulin-dependent protein kinase IIβ (CaMKIIβ) is expressed in several splice variants and plays a key role in learning and memory. Here, we identify and characterize several primate-specific CAMK2B splice isoforms, which show altered kinetic properties and changes in substrate specificity. Furthermore, we demonstrate that primate-specific CAMK2B alternative splicing is achieved through branch point weakening during evolution. We show that reducing branch point and splice site strengths during evolution globally renders constitutive exons alternative, thus providing novel mechanistic insight into cis-directed species-specific alternative splicing regulation. Using CRISPR/Cas9, we introduce a weaker, human branch point sequence into the mouse genome, resulting in strongly altered Camk2b splicing in the brains of mutant mice. We observe a strong impairment of long-term potentiation in CA3-CA1 synapses of mutant mice, thus connecting branch point–controlled CAMK2B alternative splicing with a fundamental function in learning and memory

Quantification of intracellular payload release from polymersome nanoparticles

Polymersome nanoparticles (PMs) are attractive candidates for spatio-temporal controlled delivery of therapeutic agents. Although many studies have addressed cellular uptake of solid nanoparticles, there is very little data available on intracellular release of molecules encapsulated in membranous carriers, such as polymersomes. Here, we addressed this by developing a quantitative assay based on the hydrophilic dye, fluorescein. Fluorescein was encapsulated stably in PMs of mean diameter 85 nm, with minimal leakage after sustained dialysis. No fluorescence was detectable from fluorescein PMs, indicating quenching. Following incubation of L929 cells with fluorescein PMs, there was a gradual increase in intracellular fluorescence, indicating PM disruption and cytosolic release of fluorescein. By combining absorbance measurements with flow cytometry, we quantified the real-time intracellular release of a fluorescein at a single-cell resolution. We found that 173 ± 38 polymersomes released their payload per cell, with significant heterogeneity in uptake, despite controlled synchronisation of cell cycle. This novel method for quantification of the release of compounds from nanoparticles provides fundamental information on cellular uptake of nanoparticle-encapsulated compounds. It also illustrates the stochastic nature of population distribution in homogeneous cell populations, a factor that must be taken into account in clinical use of this technology.</p

Single cell analyses and machine learning define hematopoietic progenitor and HSC-like cells derived from human PSCs

Haematopoietic stem and progenitor cells (HSPCs) develop through distinct waves at various anatomical sites during embryonic development. The in vitro differentiation of human pluripotent stem cells (hPSCs) is able to recapitulate some of these processes, however, it has proven difficult to generate functional haematopoietic stem cells (HSCs). To define the dynamics and heterogeneity of HSPCs that can be generated in vitro from hPSCs, we exploited single cell RNA sequencing (scRNAseq) in combination with single cell protein expression analysis. Bioinformatics analyses and functional validation defined the transcriptomes of naïve progenitors as well as erythroid, megakaryocyte and leukocyte-committed progenitors and we identified CD44, CD326, ICAM2/CD9 and CD18 as markers of these progenitors, respectively. Using an artificial neural network (ANN), that we trained on a scRNAseq derived from human fetal liver, we were able to identify a wide range of hPSCs-derived HPSC phenotypes, including a small group classified as HSCs. This transient HSC-like population decreased as differentiation proceeded and was completely missing in the dataset that had been generated using cells selected on the basis of CD43expression. By comparing the single cell transcriptome of in vitro-generated HSC-like cells with those generated within the fetal liver we identified transcription factors and molecular pathways that can be exploited in the future to improve the in vitro production of HSCs

A boundary exchange influence on deglacial neodymium isotope records from the deep western Indian Ocean

The use of neodymium (Nd) isotopes to reconstruct past water mass mixing relies upon the quasi-conservative behaviour of this tracer, whereas recent studies in the modern oceans have suggested that boundary exchange, involving the addition of Nd from ocean margin sediments, may be an important process in the Nd cycle. Here we suggest that the relative importance of water mass advection versus boundary exchange can be assessed where the deep western boundary current in the Indian Ocean flows past the Madagascan continental margin; a potential source of highly unradiogenic Nd. Foraminiferal coatings and bulk sediment reductive leachates are used to reconstruct bottom water Nd isotopic composition (εNd) in 8 Holocene age coretops, with excellent agreement between the two methods. These data record spatial variability of ∼4 εNd units along the flow path of Circumpolar Deep Water; εNd≈−8.8 in the deep southern inflow upstream of Madagascar, which evolves towards εNd≈−11.5 offshore northern Madagascar, whereas εNd≈−7.3 where deep water re-circulates in the eastern Mascarene Basin. This variability is attributed to boundary exchange and, together with measurements of detrital sediment εNd, an isotope mass balance suggests a deep water residence time for Nd of ≤400 yr along the Madagascan margin. Considering deglacial changes, a core in the deep inflow upstream of Madagascar records εNd changes that agree with previous reconstructions of the Circumpolar Deep Water composition in the Southern Ocean, consistent with a control by water mass advection and perhaps indicating a longer residence time for Nd in the open ocean away from local sediment inputs. In contrast, sites along the Madagascan margin record offset εNd values and reduced glacial–interglacial variability, underlining the importance of detecting boundary exchange before inferring water mass source changes from Nd isotope records. The extent of Madagascan boundary exchange appears to be unchanged between the Holocene and Late Glacial periods, while a consistent shift towards more radiogenic εNd values at all sites in the Late Glacial compared to the Holocene may represent a muted signal of a change in water mass source or composition

Reactivity of neodymium carriers in deep sea sediments: Implications for boundary exchange and paleoceanography

The dissolved neodymium (Nd) isotopic distribution in the deep oceans is determined by continental weathering inputs, water mass advection, and boundary exchange between particulate and dissolved fractions. Reconstructions of past Nd isotopic variability may therefore provide evidence on temporal changes in continental weathering inputs and/or ocean circulation patterns over a range of timescales. However, such an approach is limited by uncertainty in the mechanisms and importance of the boundary exchange process, and the challenge in reliably recovering past seawater Nd isotopic composition (εNd) from deep sea sediments. This study addresses these questions by investigating the processes involved in particulate–solution interactions and their impact on Nd isotopes. A better understanding of boundary exchange also has wider implications for the oceanic cycling and budgets of other particle-reactive elements. Sequential acid-reductive leaching experiments at pH ∼2–5 on deep sea sediments from the western Indian Ocean enable us to investigate natural boundary exchange processes over a timescale appropriate to laboratory experiments. We provide evidence that both the dissolution of solid phases and exchange processes influence the εNd of leachates, which suggests that both processes may contribute to boundary exchange. We use major element and rare earth element (REE) data to investigate the pools of Nd that are accessed and demonstrate that sediment leachate εNd values cannot always be explained by admixture between an authigenic component and the bulk detrital component. For example, in core WIND 24B, acid-reductive leaching generates εNd values between −11 and −6 as a function of solution/solid ratios and leaching times, whereas the authigenic components have εNd ≈ −11 and the bulk detrital component has εNd ≈ −15. We infer that leaching in the Mascarene Basin accesses authigenic components and a minor radiogenic volcanic component that is more reactive than Madagascan-derived clays. The preferential mobilisation of such a minor component demonstrates that the Nd released by boundary exchange could often have a significantly different εNd composition than the bulk detrital sediment. These experiments further demonstrate certain limitations on the use of acid-reductive leaching to extract the εNd composition of the authigenic fraction of bulk deep sea sediments. For example, the detrital component may contain a reactive fraction which is also acid-extractible, while the incongruent nature of this dissolution suggests that it is often inappropriate to use the bulk detrital sediment elemental chemistry and/or εNd composition when assessing possible detrital contamination of leachates. Based on the highly systematic controls observed, and evidence from REE patterns on the phases extracted, we suggest two approaches that lead to the most reliable extraction of the authigenic εNd component and good agreement with foraminiferal-based approaches; either (i) leaching of sediments without a prior decarbonation step, or (ii) the use of short leaching times and low solution/solid ratios throughout