Motor neurons generate pose-targeted movements via proprioceptive sculpting.

Motor neurons are the final common pathway1 through which the brain controls movement of the body, forming the basic elements from which all movement is composed. Yet how a single motor neuron contributes to control during natural movement remains unclear. Here we anatomically and functionally characterize the individual roles of the motor neurons that control head movement in the fly, Drosophila melanogaster. Counterintuitively, we find that activity in a single motor neuron rotates the head in different directions, depending on the starting posture of the head, such that the head converges towards a pose determined by the identity of the stimulated motor neuron. A feedback model predicts that this convergent behaviour results from motor neuron drive interacting with proprioceptive feedback. We identify and genetically2 suppress a single class of proprioceptive neuron3 that changes the motor neuron-induced convergence as predicted by the feedback model. These data suggest a framework for how the brain controls movements: instead of directly generating movement in a given direction by activating a fixed set of motor neurons, the brain controls movements by adding bias to a continuing proprioceptive-motor loop.

Synaptic gradients transform object location to action.

To survive, animals must convert sensory information into appropriate behaviours1,2. Vision is a common sense for locating ethologically relevant stimuli and guiding motor responses3-5. How circuitry converts object location in retinal coordinates to movement direction in body coordinates remains largely unknown. Here we show through behaviour, physiology, anatomy and connectomics in Drosophila that visuomotor transformation occurs by conversion of topographic maps formed by the dendrites of feature-detecting visual projection neurons (VPNs)6,7 into synaptic weight gradients of VPN outputs onto central brain neurons. We demonstrate how this gradient motif transforms the anteroposterior location of a visual looming stimulus into the fly's directional escape. Specifically, we discover that two neurons postsynaptic to a looming-responsive VPN type promote opposite takeoff directions. Opposite synaptic weight gradients onto these neurons from looming VPNs in different visual field regions convert localized looming threats into correctly oriented escapes. For a second looming-responsive VPN type, we demonstrate graded responses along the dorsoventral axis. We show that this synaptic gradient motif generalizes across all 20 primary VPN cell types and most often arises without VPN axon topography. Synaptic gradients may thus be a general mechanism for conveying spatial features of sensory information into directed motor outputs.

Base-Free Dynamic Kinetic Resolution of Secondary Alcohols with a Ruthenium-Lipase Couple.

We report the dynamic kinetic resolution (DKR) of various secondary alcohols by the combination of a ruthenium catalyst and an anionic surfactant-activated lipoprotein lipase. The DKR reactions performed under totally base-free conditions at room temperature provided the products of excellent enantiopurities (91-99% ee or greater) in high yields (92-99%). More importantly, the DKR of α-arylallyl alcohols was achieved for the first time with high yields (87-91%).

Methyllucidone inhibits STAT3 activity by regulating the expression of the protein tyrosine phosphatase MEG2 in DU145 prostate carcinoma cells.

During the search for signal transducer and activator of transcription 3 (STAT3) inhibitors from natural products, methyllucidone, isolated from Lindera species (Lauraceae), was identified as a STAT3 inhibitor. Methyllucidone inhibited STAT3 phosphorylation at tyrosine 705 in a dose- and time dependent manner in DU145 prostate cancer cells and suppressed IL-6-induced STAT3 phosphorylation at Tyr-705 in LNCaP cells. Methyllucidone decreased the expression levels of STAT3 target genes, such as cyclin D1, cyclin A, Bcl-2, Mcl-1, and survivin. Methyllucidone inhibited DU145 cell growth and induced apoptosis by arresting the cell cycle at G1 phase. Notably, knockdown of the MEG2 gene by small interfering RNA suppressed the ability of methyllucidone to inhibit STAT3 activation. Methyllucidone regulates STAT3 activity by modulating MEG2 expression, and our results suggest that this compound is a novel inhibitor of the STAT3 pathway and may be a useful lead molecule for the development of a therapeutic STAT3 inhibitor.

Ca2+ Regulation of Cav3.3 T-type Ca2+ Channel Is Mediated by Calmodulin.

Calcium-dependent inactivation of high voltage-activated Ca2+ channels plays a crucial role in limiting rises in intracellular calcium (Ca2+i). A key mediator of these effects is calmodulin, which has been found to bind the C-terminus of the pore-forming α subunit. In contrast, little is known about how Ca2+i can regulate low voltage-activated T-type Ca2+ channels. Using whole cell patch clamp, we examined the biophysical properties of Ca2+ current through the three T-type Ca2+ channel isoforms, Cav3.1, Cav3.2, or Cav3.3, comparing internal solutions containing 27 nM and l µM free Ca2+ Both activation and inactivation kinetics of Cav3.3 current in l µM Ca2+i solution were more rapid than those in 27 nM Ca2+i solution. In addition, both activation and steady-state inactivation curves of Cav3.3 were negatively shifted in the higher Ca2+i solution. In contrast, the biophysical properties of Cav3.1 and Cav3.2 isoforms were not significantly different between the two internal solutions. Overexpression of CaM1234 (a calmodulin mutant that doesn't bind Ca2+) occluded the effects of l µM Ca2+i on Cav3.3, implying that CaM is involved in the Ca2+i regulation effects on Cav3.3. Yeast two-hybrid screening and co-immunoprecipitation experiments revealed a direct interaction of CaM with the carboxyl terminus of Cav3.3. Taken together, our results suggest that Cav3.3 T-type channel is potently regulated by Ca2+i via interaction of Ca2+/CaM with the carboxyl terminus of Cav3.3.

Cytokine Response to Diet and Exercise Affects Atheromatous Matrix Metalloproteinase-2/9 Activity in Mice.

BACKGROUND: The aim of this study is to identify the principal circulating factors that modulate atheromatous matrix metalloproteinase (MMP) activity in response to diet and exercise.Methods and Results:Apolipoprotein-E knock-out (ApoE-/-) mice (n=56) with pre-existing plaque, fed either a Western diet (WD) or normal diet (ND), underwent either 10 weeks of treadmill exercise or had no treatment. Atheromatous MMP activity was visualized using molecular imaging with a MMP-2/9 activatable near-infrared fluorescent (NIRF) probe. Exercise did not significantly reduce body weight, visceral fat, and plaque size in either WD-fed animals or ND-fed animals. However, atheromatous MMP-activity was different; ND animals that did or did not exercise had similarly low MMP activities, WD animals that did not exercise had high MMP activity, and WD animals that did exercise had reduced levels of MMP activity, close to the levels of ND animals. Factor analysis and path analysis showed that soluble vascular cell adhesion molecule (sVCAM)-1 was directly positively correlated to atheromatous MMP activity. Adiponectin was indirectly negatively related to atheromatous MMP activity by way of sVCAM-1. Resistin was indirectly positively related to atheromatous MMP activity by way of sVCAM-1. Visceral fat amount was indirectly positively associated with atheromatous MMP activity, by way of adiponectin reduction and resistin elevation. MMP-2/9 imaging of additional mice (n=18) supported the diet/exercise-related anti-atherosclerotic roles for sVCAM-1. CONCLUSIONS: Diet and exercise affect atheromatous MMP activity by modulating the systemic inflammatory milieu, with sVCAM-1, resistin, and adiponectin closely interacting with each other and with visceral fat.

Ionotropic glutamate receptors IR64a and IR8a form a functional odorant receptor complex in vivo in Drosophila.

Drosophila olfactory sensory neurons express either odorant receptors or ionotropic glutamate receptors (IRs). The sensory neurons that express IR64a, a member of the IR family, send axonal projections to either the DC4 or DP1m glomeruli in the antennal lobe. DC4 neurons respond specifically to acids/protons, whereas DP1m neurons respond to a broad spectrum of odorants. The molecular composition of IR64a-containing receptor complexes in either DC4 or DP1m neurons is not known, however. Here, we immunoprecipitated the IR64a protein from lysates of fly antennal tissue and identified IR8a as a receptor subunit physically associated with IR64a by mass spectrometry. IR8a mutants and flies in which IR8a was knocked down by RNAi in IR64a+ neurons exhibited defects in acid-evoked physiological and behavioral responses. Furthermore, we found that the loss of IR8a caused a significant reduction in IR64a protein levels. When expressed in Xenopus oocytes, IR64a and IR8a formed a functional ion channel that allowed ligand-evoked cation currents. These findings provide direct evidence that IR8a is a subunit that forms a functional olfactory receptor with IR64a in vivo to mediate odor detection.

A new micro-computed tomography-based high-resolution blood-brain barrier imaging technique to study ischemic stroke.

BACKGROUND AND PURPOSE: Micro-computed tomography (mCT) offers high-resolution images, but it suffers from low contrast sensitivity and poor soft tissue contrast. We introduce a new mCT imaging technique with improved sensitivity for the dynamic spatial and temporal characterization of poststroke blood-brain barrier (BBB) dysfunction in small animals in vivo. METHODS: Transient middle cerebral artery occlusion was induced for 1 hour in 10- to 12-week-old C57BL/6 mice (n=35). At 4, 24, and 48 hours after ischemic stroke, serial in vivo mCT imaging was performed 5 minutes after intravenous infusion (n=3) or intracarotid infusion of iopromide (240 µL) for 5 minutes (n=32). After intravenous injection of 2% Evans blue, we performed ex vivo near-infrared fluorescent imaging of parenchymal Evans blue leakage, visual assessment of poststroke parenchymal hematoma, triphenyltetrazolium chloride staining of the brain tissue, and quantitative mapping of stroke-related brain lesions. RESULTS: Infarct-related BBB dysfunction could be demonstrated with intra-arterial but not with intravenous infusion of iopromide. Iopromide leakage across the dysfunctional BBB showed a monophasic (not biphasic) course for 48 hours after ischemic insult in both the parenchymal hematoma (n=5) and the non-parenchymal hematoma (n=24) groups, with relatively severe leakiness and greater hemispheric midline shift in animals with hemorrhage. Parenchymal staining on in vivo mCT overlapped with ex vivo fluorescent staining because of Evans blue. Multivariable analyses showed that midline shift and the amount of iopromide leakage at each of the 3 time points predicted the final infarct size at 48 hours. CONCLUSIONS: The new mCT BBB imaging technique, based on the intra-arterial infusion of clinically available iopromide, allows serial quantitative visualization of poststroke BBB dysfunction in mice, with high resolution and in a sensitive manner.

Photodynamic therapy using a protease-mediated theranostic agent reduces cathepsin-B activity in mouse atheromata in vivo.

OBJECTIVE: To investigate whether an intravenously injected cathepsin-B activatable theranostic agent (L-SR15) would be cleaved in and release a fluorescent agent (chlorin-e6) in mouse atheromata, allowing both the diagnostic visualization and therapeutic application of these fluorophores as photosensitizers during photodynamic therapy to attenuate plaque-destabilizing cathepsin-B activity by selectively eliminating macrophages. APPROACH AND RESULTS: Thirty-week-old apolipoprotein E knock-out mice (n=15) received intravenous injection of L-SR15 theranostic agent, control agent D-SR16, or saline 3× (D0, D7, D14). Twenty-four hours after each injection, the bilateral carotid arteries were exposed, and Cy5.5 near-infrared fluorescent imaging was performed. Fluorescent signal progressively accumulated in the atheromata of the L-SR15 group animals only, indicating that photosensitizers had been released from the theranostic agent and were accumulating in the plaque. After each imaging session, photodynamic therapy was applied with a continuous-wave diode-laser. Additional near-infrared fluorescent imaging at a longer wavelength (Cy7) with a cathepsin-B-sensing activatable molecular imaging agent showed attenuation of cathepsin-B-related signal in the L-SR15 group. Histological studies demonstrated that L-SR15-based photodynamic therapy decreased macrophage infiltration by inducing apoptosis without significantly affecting plaque size or smooth muscle cell numbers. Toxicity studies (n=24) showed that marked erythematous skin lesion was generated in C57/BL6 mice at 24 hours after intravenous injection of free chlorin-e6 and ultraviolet light irradiation; however, L-SR15 or saline did not cause cutaneous phototoxicity beyond that expected of ultraviolet irradiation alone, neither did we observe systemic toxicity or neurobehavioral changes. CONCLUSIONS: This is the first study showing that macrophage-secreted cathepsin-B activity in atheromata could be attenuated by photodynamic therapy using a protease-mediated theranostic agent.

Synergistic actions of metabotropic acetylcholine and glutamate receptors on the excitability of hippocampal CA1 pyramidal neurons.

A variety of neurotransmitters are responsible for regulating neural activity during different behavioral states. Unique responses to combinations of neurotransmitters provide a powerful mechanism by which neural networks could be differentially activated during a broad range of behaviors. Here, we show, using whole-cell recordings in rat hippocampal slices, that group I metabotropic glutamate receptors (mGluRs) and muscarinic acetylcholine receptors (mAChRs) synergistically increase the excitability of hippocampal CA1 pyramidal neurons by converting the post-burst afterhyperpolarization to an afterdepolarization via a rapidly reversible upregulation of Ca(v)2.3 R-type calcium channels. Coactivation of mAChRs and mGluRs also induced a long-lasting enhancement of the responses mediated by each receptor type. These results suggest that cooperative signaling via mAChRs and group I mGluRs could provide a mechanism by which cognitive processes may be modulated by conjoint activation of two separate neurotransmitter systems.

A post-burst after depolarization is mediated by group i metabotropic glutamate receptor-dependent upregulation of Ca(v)2.3 R-type calcium channels in CA1 pyramidal neurons.

Activation of group I metabotropic glutamate receptors (subtypes mGluR1 and mGluR5) regulates neural activity in a variety of ways. In CA1 pyramidal neurons, activation of group I mGluRs eliminates the post-burst afterhyperpolarization (AHP) and produces an afterdepolarization (ADP) in its place. Here we show that upregulation of Ca(v)2.3 R-type calcium channels is responsible for a component of the ADP lasting several hundred milliseconds. This medium-duration ADP is rapidly and reversibly induced by activation of mGluR5 and requires activation of phospholipase C (PLC) and release of calcium from internal stores. Effects of mGluR activation on subthreshold membrane potential changes are negligible but are large following action potential firing. Furthermore, the medium ADP exhibits a biphasic activity dependence consisting of short-term facilitation and longer-term inhibition. These findings suggest that mGluRs may dramatically alter the firing of CA1 pyramidal neurons via a complex, activity-dependent modulation of Ca(v)2.3 R-type channels that are activated during spiking at physiologically relevant rates and patterns.

Role of Titanium Dioxide-Immobilized PES Beads in a Combined Water Treatment System of Tubular Alumina Microfiltration and PES Beads.

The membrane process has a limit to the decay of various pollutants in water. To improve the problem, the roles of backwashing media and titanium dioxide (TiO2) photocatalyst-immobilized-polyethersulfone (PES) beads' concentration were investigated in a combined system of tubular alumina MF and the PES beads for advanced drinking water treatment. The space between the outside of the MF membrane and the module inside was filled with the PES beads. UV at a wavelength of 352 nm was irradiated from outside of the acryl module. A quantity of humic acid and kaolin was dissolved in distilled water for synthetic water. Water or air intermittent backwashing was performed outside to inside. The membrane fouling resistance after 3 h process (Rf,180) was minimum at 30 g/L of the PES beads for water backwashing, and at 40 g/L for air backwashing when increasing the PES beads from 0 to 50 g/L. The irreversible membrane fouling resistance after physical cleaning (Rif) was at the bottom at 5 g/L of the PES beads for water backwashing, which was 3.43 times higher than minimal at 40 g/L of the PES beads for air backwashing. The treatment effectiveness of turbidity increased when increasing the PES beads' concentration from 0 to 50 g/L; however, it reached a maximum at 98.1% at 40 g/L and 99.2% at 50 g/L for water and air backwashing, respectively. The treatment effectiveness of UV254 absorbance, which was dissolved organic matter (DOM), increased dramatically when increasing the PES beads; however, it reached a peak of 83.0% at 40 g/L and 86.0% at 50 g/L for water and air backwashing, respectively. Finally, the best PES beads' concentration was 20~30 g/L to minimize the membrane fouling; however, it was 50 g/L to remove pollutants effectively. The water backwashing was better than the air at treating DOM; however, the air backwashing was more effective than the water at removing turbid matter and reducing membrane fouling.

Sources and sequestration rate of organic carbon in sediments of the bare tidal flat ecosystems: A model approach.

Humans have been contributing adversely to greenhouse gas emissions by generating a vast amount of CO2, primarily causing climate change. Nature-based climate solutions, consisting of both terrestrial and marine ecosystems, are tremendous potential for sequestering and storing significant amounts of carbon, which can help to slow the progression of climate change. In this study, we use a carbon balance model to simulate the carbon sequestration rate and carbon stored in bare tidal flat (BTF) areas along Korea's west and south coasts from 2018 to 2050. Furthermore, the percentage of potential carbon sources deposited at BTF sites was calculated using a two-terminal mixing model and δ13C data. The carbon deposited on the BTF areas is the result of lateral carbon transport from upslope terrestrial regions as well as marine sources. Based on the δ13C isotope, this study classified potential carbon sources in BTFs sediment into two categories: terrestrial and marine. The results indicate that the proportion of organic carbon contribution from terrestrial sources ranged from 7.63% to 49% in the BTF studied areas. We discuss the validity of projection which was investigated over three years, from 2018 to 2020. A preliminary conclusion is that future carbon storage at BTF sites will increase significantly. Carbon accumulation increases linearly over time in nearly all areas studied, with carbon sequestration rates ranging from 0.053 to 0.623 (MgC ha-1 yr-1). This study found that a significant amount of carbon is sequestered for a long time in the BTF regions based on model simulation results. In addition, it also contributes to projects that seek to promote and conserve these climate benefits by providing estimates of carbon storage in coastal BTFs that can be included in NDCs for the Paris Agreement.

Molecular cloning and characterization of a hamster Cav1.3 Ca2+ channel variant with a long carboxyl terminus.

We have cloned a hamster Cav1.3 variant with a long carboxyl terminus. This differs from the first hamster Cav1.3 clone which has a short carboxyl terminus. When relative expression levels of the two variants were examined using quantitative RT-PCR, the long Cav1.3 transcripts were detected abundantly in the brain and testis, moderately in the heart, pancreas, and kidney, and weakly in the lung. Comparatively, the short Cav1.3 transcripts were detected less abundantly in most of the tissues. The two Cav1.3 variants were reconstituted in Xenopus oocytes and their electrophysiological properties were characterized using a two-electrode voltage clamping method. The long Cav1.3 variant was ~5-fold better expressed than the short Cav1.3 variant. When Ca2+ was used as a charge carrier, the long Cav1.3 variant containing an IQ (Ile-Gln) motif displayed strong calcium-dependent inactivation, while the short variant that was deficient of an IQ motif showed little calcium-dependent inactivation. Examination of other biophysical properties revealed that potentials for activation threshold, peak current, and half-activation and inactivation of the long Cav1.3 were significantly lower than those of the short Cav1.3. These findings suggest that the long carboxyl tail plays crucial roles in not only facilitating calcium-dependent inactivation, but also improving expression and negative shifting of the activation and inactivation properties.

Optimal Water Backwashing Condition in Combined Water Treatment of Alumina Microfiltration and PP Beads.

Membrane fouling is a dominant limit of the membrane separation process. In this research, the optimal water backwashing to solve the membrane fouling problem was investigated in the combined water treatment process of alumina MF and pure polypropylene (PP) beads. Additionally, the influence of membrane shape (tubular or seven channel) was examined, depending on the water backwashing period. The optimal backwashing time (BT) could be 20 s in the combined water treatment process, because of the highest total treated volume (VT) in our BT 6-30 s conditions. The optimal backwashing period (BP) could be 6 min, because of the minimum membrane fouling and the maximum VT in the combined process of tubular alumina MF and PP beads. The resistance of reversible membrane fouling (Rrf) showed a major resistance of total membrane fouling, and that of irreversible membrane fouling (Rif) was a minor one, in the combined process using tubular or seven channel MF. The Rif showed a decreasing trend obviously, as decreasing BT from NBW to 2 min for seven channel MF. It means that the more frequent water backwashing could be more effective to control the membrane fouling, especially irreversible fouling, for seven channel membranes than tubular membranes.

Healing Ion-Implanted Semiconductors by Hybrid Microwave Annealing: Activation of Nitrogen-Implanted TiO2.

In order to recover the damaged structure of a nitrogen-implanted TiO2 (N-I-TiO2) photoanode, hybrid microwave annealing (HMA) is proposed as an alternative postannealing process instead of conventional thermal annealing (CTA). Compared to CTA, HMA provides distinctive advantages: (i) facile transformation of the interstitial N-N states into substitutional N-Ti states, (ii) better preservation of the ion-implanted nitrogen in TiO2, and (iii) effective alleviation of lattice strain and reconstruction of the broken bonds. As a result, the HMA-activated photoanode improves the photocurrent density by a factor of ∼3.2 from 0.29 to 0.93 mA cm-2 at 1.23 VRHE and the incident photon-to-current conversion efficiency (IPCE) from ∼2.9% to ∼10.5% at 430 nm relative to those of the as-prepared N-I-TiO2 photoanode in photoelectrochemical water oxidation, which are much better than those of the CTA-activated photoanode (0.58 mA cm-2 at 1.23 VRHE and IPCE of 5.7% at 430 nm), especially in the visible light region (≥420 nm).

The Function of Adsorption, Photo-Oxidation, and Humic Acid Using Air Backwashing in Integrated Water Treatment of Multichannel Ceramic MF and PP Particles.

For advanced water treatment, function of microfiltration (MF), adsorption, photo-oxidation, humic acid (HA), and polypropylene (PP) particles on membrane fouling and decay effectiveness were investigated in an integrated water treatment, of multichannel ceramic MF and PP particles, using UV radiation and air backwashing. The synthetic feed was organized with HA and kaolin. The membrane fouling resistance (Rf) of the (MF + PP) system presented the lowermost, and amplified intensely from the (MF + UV) to MF system. The percentages of MF and adsorption by PP particles for turbidity treatment were 87.6% and 3.8%, individually; however, the percentages of MF and adsorption by PP particles for dissolved organic matters (DOM) treatment were 27.9% and 5.0%, respectively. The decay effectiveness of turbidity presented the greatest 95.4% at HA of 10 mg/L; however, that of DOM increased as HA concentration ascended. The ultimate Rf after 180 min procedure showed the maximum at 30 g/L of PP particles concentration, and improved dramatically, as PP particles decreased. Finally, the maximum VT was acquired at 30 and 50 g/L of PP particles, because flux preserved greater throughout the procedure. The decay effectiveness of turbidity and DOM showed the maximal 95.4% and 56.8% at 40 and 50 g/L of PP particles, respectively.

Concurrent Formation of N-H Imines and Carbonyl Compounds by Ruthenium-Catalyzed C-C Bond Cleavage of ß-Hydroxy Azides.

A commercial cyclopentadienylrutenium dicarbonyl dimer ([CpRu(CO)2]2) efficiently catalyzes the formation of N-H imines and carbonyl compounds simultaneously from ß-hydroxy azides via C-C bond cleavage under visible light. Density functional theory calculations for the cleavage reaction support the mechanism involving chelation of alkoxy azide species and liberation of nitrogen as the driving force. The synthetic utility of the reaction was demonstrated by a new amine synthesis promoted by chemoselective allylation of imine and synthesis of isoquinoline.

A Preclinical Trial and Molecularly Annotated Patient Cohort Identify Predictive Biomarkers in Homologous Recombination-deficient Pancreatic Cancer.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) arising in patients with a germline BRCA1 or BRCA2 (gBRCA) mutation may be sensitive to platinum and PARP inhibitors (PARPi). However, treatment stratification based on gBRCA mutational status alone is associated with heterogeneous responses. EXPERIMENTAL DESIGN: We performed a seven-arm preclinical trial consisting of 471 mice, representing 12 unique PDAC patient-derived xenografts, of which nine were gBRCA mutated. From 179 patients whose PDAC was whole-genome and transcriptome sequenced, we identified 21 cases with homologous recombination deficiency (HRD), and investigated prognostic biomarkers. RESULTS: We found that biallelic inactivation of BRCA1/BRCA2 is associated with genomic hallmarks of HRD and required for cisplatin and talazoparib (PARPi) sensitivity. However, HRD genomic hallmarks persisted in xenografts despite the emergence of therapy resistance, indicating the presence of a genomic scar. We identified tumor polyploidy and a low Ki67 index as predictors of poor cisplatin and talazoparib response. In patients with HRD PDAC, tumor polyploidy and a basal-like transcriptomic subtype were independent predictors of shorter survival. To facilitate clinical assignment of transcriptomic subtype, we developed a novel pragmatic two-marker assay (GATA6:KRT17). CONCLUSIONS: In summary, we propose a predictive and prognostic model of gBRCA-mutated PDAC on the basis of HRD genomic hallmarks, Ki67 index, tumor ploidy, and transcriptomic subtype.