Voltage-induced calcium release in Caenorhabditis elegans body muscles.

Type 1 voltage-activated calcium channels (CaV1) in the plasma membrane trigger calcium release from the sarcoplasmic reticulum (SR) by two mechanisms. In voltage-induced calcium release (VICR), CaV1 voltage sensing domains are directly coupled to ryanodine receptors (RYRs), an SR calcium channel. In calcium-induced calcium release (CICR), calcium ions flowing through activated CaV1 channels bind and activate RYR channels. VICR is thought to occur exclusively in vertebrate skeletal muscle while CICR occurs in all other muscles (including all invertebrate muscles). Here, we use calcium-activated SLO-2 potassium channels to analyze CaV1-SR coupling in Caenorhabditis elegans body muscles. SLO-2 channels were activated by both VICR and external calcium. VICR-mediated SLO-2 activation requires two SR calcium channels (RYRs and IP3 Receptors), JPH-1/Junctophilin, a PDZ (PSD95, Dlg1, ZO-1 domain) binding domain (PBD) at EGL-19/CaV1's carboxy-terminus, and SHN-1/Shank (a scaffolding protein that binds EGL-19's PBD). Thus, VICR occurs in invertebrate muscles.

Post-synaptic GABAA receptors potentiate transmission by recruiting CaV2 channels to their inputs.

We describe a retrograde synaptic signal at the C. elegans GABAergic neuromuscular junction. At this synapse, GABA release is controlled by two voltage-activated calcium channels (UNC-2/CaV2 and EGL-19/CaV1), and muscle responses are mediated by a single GABA receptor (UNC-49/GABAA). Mutations inactivating UNC-49 or those preventing UNC-49 synaptic clustering cause retrograde defects in GABAergic motor neurons, whereby UNC-2/CaV2 levels at active zones, UNC-2 current, and pre-synaptic GABA release are decreased. Inactivating post-synaptic GABAA receptors has no effect on GABA neuron EGL-19/CaV1 levels nor on several other pre-synaptic markers. The effect of GABAA receptors on pre-synaptic strength is not a consequence of decreased GABA transmission and is input selective. Finally, pre-synaptic UNC-2/CaV2 levels are increased when post-synaptic GABAA receptors are increased but are unaffected by increased extra-synaptic receptors. Collectively, these results suggest that clustered post-synaptic GABAA receptors adjust the strength of their inputs by recruiting CaV2 to contacting active zones.

Cannabidiol and cannabis-inspired terpene blends have acute prosocial effects in the BTBR mouse model of autism spectrum disorder.

Introduction: Cannabidiol (CBD) is a non-intoxicating phytocannabinoid with increasing popularity due to its purported therapeutic efficacy for numerous off-label conditions including anxiety and autism spectrum disorder (ASD). Those with ASD are commonly deficient in endogenous cannabinoid signaling and GABAergic tone. CBD has a complex pharmacodynamic profile that includes enhancing GABA and endocannabinoid signaling. Thus, there is mechanistic justification for investigating CBD's potential to improve social interaction and related symptoms in ASD. Recent clinical trials in children with ASD support CBD's beneficial effects in numerous comorbid symptoms, but its impact on social behavior is understudied. Methods: Here, we tested the prosocial and general anxiolytic efficacy of a commercially available CBD-rich broad spectrum hemp oil delivered by repeated puff vaporization and consumed via passive inhalation in the female cohort of the BTBR strain, a common inbred mouse line for preclinical assessment of ASD-like behaviors. Results: We observed that CBD enhanced prosocial behaviors using the 3-Chamber Test with a different vapor dose-response relationship between prosocial behavior and anxiety-related behavior on the elevated plus maze. We also identified that inhalation of a vaporized terpene blend from the popular OG Kush cannabis strain increased prosocial behavior independently of CBD and acted together with CBD to promote a robust prosocial effect. We observed similar prosocial effects with two additional cannabis terpene blends from the Do-Si-Dos and Blue Dream strains, and further reveal that these prosocial benefits rely on the combination of multiple terpenes that comprise the blends. Discussion: Our results illustrate the added benefit of cannabis terpene blends for CBD-based treatment of ASD.

Zinc activation of OTOP proton channels identifies structural elements of the gating apparatus.

Otopetrin proteins (OTOPs) form proton-selective ion channels that are expressed in diverse cell types where they mediate detection of acids or regulation of pH. In vertebrates there are three family members: OTOP1 is required for formation of otoconia in the vestibular system and it forms the receptor for sour taste, while the functions of OTOP2 and OTOP3 are not yet known. Importantly, the gating mechanisms of any of the OTOP channels are not well understood. Here, we show that zinc (Zn2+), as well as other transition metals including copper (Cu2+), potently activates murine OTOP3 (mOTOP3). Zn2+ pre-exposure increases the magnitude of mOTOP3 currents to a subsequent acid stimulus by as much as 10-fold. In contrast, mOTOP2 currents are insensitive to activation by Zn2+. Swapping the extracellular tm 11-12 linker between mOTOP3 and mOTOP2 was sufficient to eliminate Zn2+ activation of mOTOP3 and confer Zn2+ activation on mOTOP2. Mutation to alanine of H531 and E535 within the tm 11-12 linker and H234 and E238 within the 5-6 linker reduced or eliminated activation of mOTOP3 by Zn2+, indicating that these residues likely contribute to the Zn2+ activating site. Kinetic modeling of the data is consistent with Zn2+ stabilizing the opn2+en state of the channel, competing with H+ for activation of the channels. These results establish the tm 11-12 and tm 5-6 linkers as part of the gating apparatus of OTOP channels and a target for drug discovery. Zn2+ is an essential micronutrient and its activation of OTOP channels will undoubtedly have important physiological sequelae.

The serine hydrolase ABHD6 controls survival and thermally induced seizures in a mouse model of Dravet syndrome.

Evidence suggests that inhibition of α/ß hydrolase-domain containing 6 (ABHD6) reduces seizures; however, the molecular mechanism of this therapeutic response remains unknown. We discovered that heterozygous expression of Abhd6 (Abhd6+/-) significantly reduced the premature lethality of Scn1a+/- mouse pups, a genetic mouse model of Dravet Syndrome (DS). Both Abhd6+/- mutation and pharmacological inhibition of ABHD6 reduced the duration and incidence of thermally induced seizures in Scn1a+/- pups. Mechanistically, the in vivo anti-seizure response resulting from ABHD6 inhibition is mediated by potentiation of gamma-aminobutyric acid receptors Type-A (GABAAR). Brain slice electrophysiology showed that blocking ABHD6 potentiates extrasynaptic (tonic) GABAAR currents that reduce dentate granule cell excitatory output without affecting synaptic (phasic) GABAAR currents. Our results unravel an unexpected mechanistic link between ABHD6 activity and extrasynaptic GABAAR currents that controls hippocampal hyperexcitability in a genetic mouse model of DS. BRIEF SUMMARY: This study provides the first evidence for a mechanistic link between ABHD6 activity and the control of extrasynaptic GABAAR currents that controls hippocampal hyperexcitability in a genetic mouse model of Dravet Syndrome and can be targeted to dampened seizures.

Cannabidiol's neuroprotective properties and potential treatment of traumatic brain injuries.

Cannabidiol (CBD) has numerous pharmacological targets that initiate anti-inflammatory, antioxidative, and antiepileptic properties. These neuroprotective benefits have generated interest in CBD's therapeutic potential against the secondary injury cascade from traumatic brain injury (TBI). There are currently no effective broad treatment strategies for combating the damaging mechanisms that follow the primary injury and lead to lasting neurological consequences or death. However, CBD's effects on different neurotransmitter systems, the blood brain barrier, oxidative stress mechanisms, and the inflammatory response provides mechanistic support for CBD's clinical utility in TBI. This review describes the cascades of damage caused by TBI and CBD's neuroprotective mechanisms to counter them. We also present challenges in the clinical treatment of TBI and discuss important future clinical research directions for integrating CBD in treatment protocols. The mechanistic evidence provided by pre-clinical research shows great potential for CBD as a much-needed improvement in the clinical treatment of TBI. Upcoming clinical trials sponsored by major professional sport leagues are the first attempts to test the efficacy of CBD in head injury treatment protocols and highlight the need for further clinical research.

The ADRENAL score: A comprehensive scoring system for standardized evaluation of adrenal tumor.

Objectives: To propose an original and standardized scoring system to quantify the functional and anatomical characteristics of adrenal tumor. Materials and methods: Four groups of consecutive adrenalectomies (n = 458) with heterogeneity in tumor characteristics and surgical approaches, including 212 laparoscopic cases (Group 1) and 105 robotic cases (Group 2) from The First Affiliated Hospital of Nanchang University, 28 robotic cases from Temple University Hospital (Group 3) and 113 laparoscopic cases from The First Affiliated Hospital of Guangxi Medical University (Group 4). All patients were followed up for 4.5 to 5.5 years. Six parameters including functional status or suspicion of malignancy, tumor size, relationship to adjacent organs, intratumoral enhancement on CT, nearness of the tumor to major vessels and body mass index were assessed and scored on a 0, 1 and 2 points scale. Correlation between the sum of the 6 scores and tumor laterality (ADRENAL score) verse operative time (OT), estimated blood loss (EBL), perioperative complications, transfusion, conversion and length of hospital stay was analyzed. Results: ADRENAL score was a strong predictor of both OT and EBL in all four groups (p < 0.05 for all tests). In Group 2 and 4, higher ADRENAL score seemed to correlate with longer hospital stay. No statistically significant correlation between ADRENAL score and complication, transfusion or conversion was noted yet. Conclusions: ADRENAL score appears to be a valid predictor of surgical outcomes. It may provide a common reference for adrenal surgery training program, preoperative risk assessment and stratified comparative analysis of adrenal surgeries via different techniques and approaches.

CASK and FARP localize two classes of post-synaptic ACh receptors thereby promoting cholinergic transmission.

Changes in neurotransmitter receptor abundance at post-synaptic elements play a pivotal role in regulating synaptic strength. For this reason, there is significant interest in identifying and characterizing the scaffolds required for receptor localization at different synapses. Here we analyze the role of two C. elegans post-synaptic scaffolding proteins (LIN-2/CASK and FRM-3/FARP) at cholinergic neuromuscular junctions. Constitutive knockouts or muscle specific inactivation of lin-2 and frm-3 dramatically reduced spontaneous and evoked post-synaptic currents. These synaptic defects resulted from the decreased abundance of two classes of post-synaptic ionotropic acetylcholine receptors (ACR-16/CHRNA7 and levamisole-activated AChRs). LIN-2's AChR scaffolding function is mediated by its SH3 and PDZ domains, which interact with AChRs and FRM-3/FARP, respectively. Thus, our findings show that post-synaptic LIN-2/FRM-3 complexes promote cholinergic synaptic transmission by recruiting AChRs to post-synaptic elements.

Structural motifs for subtype-specific pH-sensitive gating of vertebrate otopetrin proton channels.

Otopetrin (OTOP) channels are proton-selective ion channels conserved among vertebrates and invertebrates, with no structural similarity to other ion channels. There are three vertebrate OTOP channels (OTOP1, OTOP2, and OTOP3), of which one (OTOP1) functions as a sour taste receptor. Whether extracellular protons gate OTOP channels, in addition to permeating them, was not known. Here, we compare the functional properties of the three murine OTOP channels using patch-clamp recording and cytosolic pH microfluorimetry. We find that OTOP1 and OTOP3 are both steeply activated by extracellular protons, with thresholds of pHo <6.0 and 5.5, respectively, and kinetics that are pH-dependent. In contrast, OTOP2 channels are broadly active over a large pH range (pH 5 pH 10) and carry outward currents in response to extracellular alkalinization (>pH 9.0). Strikingly, we could change the pH-sensitive gating of OTOP2 and OTOP3 channels by swapping extracellular linkers that connect transmembrane domains. Swaps of extracellular linkers in the N domain, comprising transmembrane domains 1-6, tended to change the relative conductance at alkaline pH of chimeric channels, while swaps within the C domain, containing transmembrane domains 7-12, tended to change the rates of OTOP3 current activation. We conclude that members of the OTOP channel family are proton-gated (acid-sensitive) proton channels and that the gating apparatus is distributed across multiple extracellular regions within both the N and C domains of the channels. In addition to the taste system, OTOP channels are expressed in the vertebrate vestibular and digestive systems. The distinct gating properties we describe may allow them to subserve varying cell-type specific functions in these and other biological systems.

Stereotyped behavioral maturation and rhythmic quiescence in C. elegans embryos.

Systematic analysis of rich behavioral recordings is being used to uncover how circuits encode complex behaviors. Here, we apply this approach to embryos. What are the first embryonic behaviors and how do they evolve as early neurodevelopment ensues? To address these questions, we present a systematic description of behavioral maturation for Caenorhabditis elegans embryos. Posture libraries were built using a genetically encoded motion capture suit imaged with light-sheet microscopy and annotated using custom tracking software. Analysis of cell trajectories, postures, and behavioral motifs revealed a stereotyped developmental progression. Early movement is dominated by flipping between dorsal and ventral coiling, which gradually slows into a period of reduced motility. Late-stage embryos exhibit sinusoidal waves of dorsoventral bends, prolonged bouts of directed motion, and a rhythmic pattern of pausing, which we designate slow wave twitch (SWT). Synaptic transmission is required for late-stage motion but not for early flipping nor the intervening inactive phase. A high-throughput behavioral assay and calcium imaging revealed that SWT is elicited by the rhythmic activity of a quiescence-promoting neuron (RIS). Similar periodic quiescent states are seen prenatally in diverse animals and may play an important role in promoting normal developmental outcomes.

Functional Status and Engagement in Physical Activity Among Maintenance Dialysis Patients: A Mixed Methods Study.

Rationale & Objective: People receiving maintenance hemodialysis (HD) experience significant activity barriers but desire the ability to do more and remain independent. To learn about how to help people who require dialysis stay active, a mixed methods study was designed to assess functional status and explore participants' lived activity experiences. Study Design: A concurrent mixed methods design was chosen to increase understanding of the real-life activity experiences of people who require dialysis through in-depth interviews paired with functional status measures. The qualitative findings were fully integrated with the quantitative results to link characteristics associated with different physical activity levels. Setting & Participants: A purposive sample of 15 adult patients receiving maintenance HD for at least 3 months was recruited from 7 dialysis centers in Newark, New Jersey. Analytical Approach: Thematic analysis using principles of interpretive phenomenology. Fully integrated quantitative and qualitative data with joint displays and conversion mixed methods. Results: Participants had a median age of 58 years and were predominantly African American (83%) and men (67%). Three descriptive categories were generated about the participants. They described physical activity as a routine daily activity rather than structured exercise. All participants experienced substantial hardship in addition to chronic kidney disease and expressed that family, friends, and faith were essential to their ability to be active. An overarching theme was generated for participants' mindsets about physical activity. Within the mindset theme, we discerned 3 subthemes comprising characteristics of participants' mindsets by levels of engagement in physical activity. Limitations: While code saturation and trends in functional status measures were achieved with 15 participants, a larger sample size would allow for deeper meaning saturation and statistical inference. Conclusions: Patients receiving maintenance HD with an engaged mindset exhibited more adaptive coping skills, moved more, wanted to help others, and had a normal body weight habitus. These participants employed adaptive coping skills to carry out daily life activities of importance, highlighting the value of adaptive coping to help overcome the challenges of being physically active.

Shank promotes action potential repolarization by recruiting BK channels to calcium microdomains.

Mutations altering the scaffolding protein Shank are linked to several psychiatric disorders, and to synaptic and behavioral defects in mice. Among its many binding partners, Shank directly binds CaV1 voltage activated calcium channels. Here, we show that the Caenorhabditis elegans SHN-1/Shank promotes CaV1 coupling to calcium activated potassium channels. Mutations inactivating SHN-1, and those preventing SHN-1 binding to EGL-19/CaV1 all increase action potential durations in body muscles. Action potential repolarization is mediated by two classes of potassium channels: SHK-1/KCNA and SLO-1 and SLO-2 BK channels. BK channels are calcium-dependent, and their activation requires tight coupling to EGL-19/CaV1 channels. SHN-1's effects on AP duration are mediated by changes in BK channels. In shn-1 mutants, SLO-2 currents and channel clustering are significantly decreased in both body muscles and neurons. Finally, increased and decreased shn-1 gene copy number produce similar changes in AP width and SLO-2 current. Collectively, these results suggest that an important function of Shank is to promote microdomain coupling of BK with CaV1.

Generalizable Compositional Features Influencing the Proteostatic Fates of Polar Low-Complexity Domains.

Protein aggregation is associated with a growing list of human diseases. A substantial fraction of proteins in eukaryotic proteomes constitutes a proteostasis network-a collection of proteins that work together to maintain properly folded proteins. One of the overarching functions of the proteostasis network is the prevention or reversal of protein aggregation. How proteins aggregate in spite of the anti-aggregation activity of the proteostasis machinery is incompletely understood. Exposed hydrophobic patches can trigger degradation by the ubiquitin-proteasome system, a key branch of the proteostasis network. However, in a recent study, we found that model glycine (G)-rich or glutamine/asparagine (Q/N)-rich prion-like domains differ in their susceptibility to detection and degradation by this system. Here, we expand upon this work by examining whether the features controlling the degradation of our model prion-like domains generalize broadly to G-rich and Q/N-rich domains. Experimentally, native yeast G-rich domains in isolation are sensitive to the degradation-promoting effects of hydrophobic residues, whereas native Q/N-rich domains completely resist these effects and tend to aggregate instead. Bioinformatic analyses indicate that native G-rich domains from yeast and humans tend to avoid degradation-promoting features, suggesting that the proteostasis network may act as a form of selection at the molecular level that constrains the sequence space accessible to G-rich domains. However, the sensitivity or resistance of G-rich and Q/N-rich domains, respectively, was not always preserved in their native protein contexts, highlighting that proteins can evolve other sequence features to overcome the intrinsic sensitivity of some LCDs to degradation.

Presynaptic Gαo (GOA-1) signals to depress command neuron excitability and allow stretch-dependent modulation of egg laying in Caenorhabditis elegans.

Egg laying in the nematode worm Caenorhabditis elegans is a two-state behavior modulated by internal and external sensory input. We have previously shown that homeostatic feedback of embryo accumulation in the uterus regulates bursting activity of the serotonergic HSN command neurons that sustains the egg-laying active state. How sensory feedback of egg release signals to terminate the egg-laying active state is less understood. We find that Gαo, a conserved Pertussis Toxin-sensitive G protein, signals within HSN to inhibit egg-laying circuit activity and prevent entry into the active state. Gαo signaling hyperpolarizes HSN, reducing HSN Ca2+ activity and input onto the postsynaptic vulval muscles. Loss of inhibitory Gαo signaling uncouples presynaptic HSN activity from a postsynaptic, stretch-dependent homeostat, causing precocious entry into the egg-laying active state when only a few eggs are present in the uterus. Feedback of vulval opening and egg release activates the uv1 neuroendocrine cells which release NLP-7 neuropeptides which signal to inhibit egg laying through Gαo-independent mechanisms in the HSNs and Gαo-dependent mechanisms in cells other than the HSNs. Thus, neuropeptide and inhibitory Gαo signaling maintain a bi-stable state of electrical excitability that dynamically controls circuit activity in response to both external and internal sensory input to drive a two-state behavior output.

Efficacy and safety of voclosporin versus placebo for lupus nephritis (AURORA 1): a double-blind, randomised, multicentre, placebo-controlled, phase 3 trial.

BACKGROUND: Voclosporin, a novel calcineurin inhibitor approved for the treatment of adults with lupus nephritis, improved complete renal response rates in patients with lupus nephritis in a phase 2 trial. This study aimed to evaluate the efficacy and safety of voclosporin for the treatment of lupus nephritis. METHODS: This multicentre, double-blind, randomised phase 3 trial was done in 142 hospitals and clinics across 27 countries. Patients with a diagnosis of systemic lupus erythematosus with lupus nephritis according to the American College of Rheumatology criteria, and a kidney biopsy within 2 years that showed class III, IV, or V (alone or in combination with class III or IV) were eligible. Patients were randomly assigned (1:1) to oral voclosporin (23·7 mg twice daily) or placebo, on a background of mycophenolate mofetil (1 g twice daily) and rapidly tapered low-dose oral steroids, by use of an interactive web response system. The primary endpoint was complete renal response at 52 weeks defined as a composite of urine protein creatinine ratio of 0·5 mg/mg or less, stable renal function (defined as estimated glomerular filtration rate [eGFR] ≥60 mL/min/1·73 m2 or no confirmed decrease from baseline in eGFR of >20%), no administration of rescue medication, and no more than 10 mg prednisone equivalent per day for 3 or more consecutive days or for 7 or more days during weeks 44 through 52, just before the primary endpoint assessment. Safety was also assessed. Efficacy analysis was by intention-to-treat and safety analysis by randomised patients receiving at least one dose of study treatment. The trial is registered with ClinicalTrials.gov, NCT03021499. FINDINGS: Between April 13, 2017, and Oct 10, 2019, 179 patients were assigned to the voclosporin group and 178 to the placebo group. The primary endpoint of complete renal response at week 52 was achieved in significantly more patients in the voclosporin group than in the placebo group (73 [41%] of 179 patients vs 40 [23%] of 178 patients; odds ratio 2·65; 95% CI 1·64-4·27; p<0·0001). The adverse event profile was balanced between the two groups; serious adverse events occurred in 37 (21%) of 178 in the voclosporin group and 38 (21%) of 178 patients in the placebo group. The most frequent serious adverse event involving infection was pneumonia, occurring in 7 (4%) patients in the voclosporin group and in 8 (4%) patients in the placebo group. A total of six patients died during the study or study follow-up period (one [<1%] patient in the voclosporin group and five [3%] patients in the placebo group). None of the events leading to death were considered by the investigators to be related to the study treatments. INTERPRETATION: Voclosporin in combination with MMF and low-dose steroids led to a clinically and statistically superior complete renal response rate versus MMF and low-dose steroids alone, with a comparable safety profile. This finding is an important advancement in the treatment of patients with active lupus nephritis. FUNDING: Aurinia Pharmaceuticals.

Caenorhabditis elegans junctophilin has tissue-specific functions and regulates neurotransmission with extended-synaptotagmin.

The junctophilin family of proteins tether together plasma membrane (PM) and endoplasmic reticulum (ER) membranes, and couple PM- and ER-localized calcium channels. Understanding in vivo functions of junctophilins is of great interest for dissecting the physiological roles of ER-PM contact sites. Here, we show that the sole Caenorhabditis elegans junctophilin JPH-1 localizes to discrete membrane contact sites in neurons and muscles and has important tissue-specific functions. jph-1 null mutants display slow growth and development due to weaker contraction of pharyngeal muscles, leading to reduced feeding. In the body wall muscle, JPH-1 colocalizes with the PM-localized EGL-19 voltage-gated calcium channel and ER-localized UNC-68 RyR calcium channel, and is required for animal movement. In neurons, JPH-1 colocalizes with the membrane contact site protein Extended-SYnaptoTagmin 2 (ESYT-2) in the soma, and is present near presynaptic release sites. Interestingly, jph-1 and esyt-2 null mutants display mutual suppression in their response to aldicarb, suggesting that JPH-1 and ESYT-2 have antagonistic roles in neuromuscular synaptic transmission. Additionally, we find an unexpected cell nonautonomous effect of jph-1 in axon regrowth after injury. Genetic double mutant analysis suggests that jph-1 functions in overlapping pathways with two PM-localized voltage-gated calcium channels, egl-19 and unc-2, and with unc-68 for animal health and development. Finally, we show that jph-1 regulates the colocalization of EGL-19 and UNC-68 and that unc-68 is required for JPH-1 localization to ER-PM puncta. Our data demonstrate important roles for junctophilin in cellular physiology, and also provide insights into how junctophilin functions together with other calcium channels in vivo.

Male pheromones modulate synaptic transmission at the C. elegans neuromuscular junction in a sexually dimorphic manner.

The development of functional synapses in the nervous system is important for animal physiology and behaviors, and its disturbance has been linked with many neurodevelopmental disorders. The synaptic transmission efficacy can be modulated by the environment to accommodate external changes, which is crucial for animal reproduction and survival. However, the underlying plasticity of synaptic transmission remains poorly understood. Here we show that in Caenorhabditis elegans, the male environment increases the hermaphrodite cholinergic transmission at the neuromuscular junction (NMJ), which alters hermaphrodites' locomotion velocity and mating efficiency. We identify that the male-specific pheromones mediate this synaptic transmission modulation effect in a developmental stage-dependent manner. Dissection of the sensory circuits reveals that the AWB chemosensory neurons sense those male pheromones and further transduce the information to NMJ using cGMP signaling. Exposure of hermaphrodites to the male pheromones specifically increases the accumulation of presynaptic CaV2 calcium channels and clustering of postsynaptic acetylcholine receptors at cholinergic synapses of NMJ, which potentiates cholinergic synaptic transmission. Thus, our study demonstrates a circuit mechanism for synaptic modulation and behavioral flexibility by sexual dimorphic pheromones.

MRI demonstration of gadolinium deposition in bone after monthly triple-dose gadopentetate dimeglumine and correlation with frequency of hypophosphatemia.

OBJECTIVES: We retrospectively analyzed data of the BECOME trial to investigate whether serial administration of triple-dose (3-dose) gadopentetate dimeglumine would result in the development of T1 signal-to-noise (S/N) changes in the cranial diploic space and whether S/N changes correlated with on-study hypophosphatemia. METHODS: Signal intensity analysis was performed on the first year's data of the BECOME trial using 3-dose Gd (14 months, maximum number of doses, 39, mean: 36). Routine blood and urine tests were obtained each month for safety monitoring. Linear mixed regression modeling with random intercept was used to analyze monthly signal-to-noise ratio (S/N = Bone/Air) using an ROI of the diploic space created from T2W images and overlaid on T1FS (T1 fat-saturated) images. Incidence of phosphate abnormalities was analyzed using the general estimation equation; correlation of phosphate and S/N change was achieved with type 3 test of fixed effects. RESULTS: Cranial diploic space T1FS S/N increased over 14 months: S/N = 0.039 mean monthly increase (S.E. 0.008; p < 0.0001). Subjects with consistently normal phosphate levels (n = 32) experienced more of a S/N increase than patients with at least one episode of hypophosphatemia (n = 35) (0.057 vs. 0.023, respectively, p = 0.037). Those with moderate hypophosphatemia demonstrated no significant S/N increase. CONCLUSION: Monthly administration of 3-dose gadopentetate dimeglumine is associated with development of increased S/N on T1FS imaging in the cranial diploic space, suggesting Gd retention in bone. Our data suggests MRI could be used as a noninvasive method of tracking Gd retention in bone, which was more pronounced in patients with normal phosphate levels.

A Hyperactive Form of unc-13 Enhances Ca2+ Sensitivity and Synaptic Vesicle Release Probability in C. elegans.

Munc13 proteins play several roles in regulating short-term synaptic plasticity. However, the underlying molecular mechanisms remain largely unclear. Here we report that C. elegans UNC-13L, a Munc13-1 ortholog, has three domains that inhibit synaptic vesicle (SV) exocytosis. These include the X (sequence between C2A and C1), C1, and C2B domains. Deleting all three inhibitory domains produces a hyperactive UNC-13 (sUNC-13) that exhibits dramatically increased neurotransmitter release, Ca2+ sensitivity of release, and release probability. The vesicular pool in unc-13 mutants rescued by sUNC-13 exhibits a faster synaptic recovery and replenishment rate, demonstrating an important role of sUNC-13 in regulating synaptic plasticity. Analysis of double mutants suggests that sUNC-13 enhances tonic release by increasing the open probability of UNC-64/syntaxin-1A, whereas its effects on evoked release appear to be mediated by additional functions, presumably by further regulating the activity of the assembled soluble N-ethylmaleimide-sensitive factor activating protein receptor (SNARE) complex.