The Fuzzy Logic of Network Connectivity in Mouse Visual Thalamus.

In an attempt to chart parallel sensory streams passing through the visual thalamus, we acquired a 100-trillion-voxel electron microscopy (EM) dataset and identified cohorts of retinal ganglion cell axons (RGCs) that innervated each of a diverse group of postsynaptic thalamocortical neurons (TCs). Tracing branches of these axons revealed the set of TCs innervated by each RGC cohort. Instead of finding separate sensory pathways, we found a single large network that could not be easily subdivided because individual RGCs innervated different kinds of TCs and different kinds of RGCs co-innervated individual TCs. We did find conspicuous network subdivisions organized on the basis of dendritic rather than neuronal properties. This work argues that, in the thalamus, neural circuits are not based on a canonical set of connections between intrinsically different neuronal types but, rather, may arise by experience-based mixing of different kinds of inputs onto individual postsynaptic cells.

Saturated Reconstruction of a Volume of Neocortex.

We describe automated technologies to probe the structure of neural tissue at nanometer resolution and use them to generate a saturated reconstruction of a sub-volume of mouse neocortex in which all cellular objects (axons, dendrites, and glia) and many sub-cellular components (synapses, synaptic vesicles, spines, spine apparati, postsynaptic densities, and mitochondria) are rendered and itemized in a database. We explore these data to study physical properties of brain tissue. For example, by tracing the trajectories of all excitatory axons and noting their juxtapositions, both synaptic and non-synaptic, with every dendritic spine we refute the idea that physical proximity is sufficient to predict synaptic connectivity (the so-called Peters' rule). This online minable database provides general access to the intrinsic complexity of the neocortex and enables further data-driven inquiries.

Connectomes across development reveal principles of brain maturation.

An animal's nervous system changes as its body grows from birth to adulthood and its behaviours mature1-8. The form and extent of circuit remodelling across the connectome is unknown3,9-15. Here we used serial-section electron microscopy to reconstruct the full brain of eight isogenic Caenorhabditis elegans individuals across postnatal stages to investigate how it changes with age. The overall geometry of the brain is preserved from birth to adulthood, but substantial changes in chemical synaptic connectivity emerge on this consistent scaffold. Comparing connectomes between individuals reveals substantial differences in connectivity that make each brain partly unique. Comparing connectomes across maturation reveals consistent wiring changes between different neurons. These changes alter the strength of existing connections and create new connections. Collective changes in the network alter information processing. During development, the central decision-making circuitry is maintained, whereas sensory and motor pathways substantially remodel. With age, the brain becomes progressively more feedforward and discernibly modular. Thus developmental connectomics reveals principles that underlie brain maturation.

Multi-layered maps of neuropil with segmentation-guided contrastive learning.

Maps of the nervous system that identify individual cells along with their type, subcellular components and connectivity have the potential to elucidate fundamental organizational principles of neural circuits. Nanometer-resolution imaging of brain tissue provides the necessary raw data, but inferring cellular and subcellular annotation layers is challenging. We present segmentation-guided contrastive learning of representations (SegCLR), a self-supervised machine learning technique that produces representations of cells directly from 3D imagery and segmentations. When applied to volumes of human and mouse cortex, SegCLR enables accurate classification of cellular subcompartments and achieves performance equivalent to a supervised approach while requiring 400-fold fewer labeled examples. SegCLR also enables inference of cell types from fragments as small as 10 µm, which enhances the utility of volumes in which many neurites are truncated at boundaries. Finally, SegCLR enables exploration of layer 5 pyramidal cell subtypes and automated large-scale analysis of synaptic partners in mouse visual cortex.

Discovery of novel amino acid production traits by evolution of synthetic co-cultures.

BACKGROUND: Amino acid production features of Corynebacterium glutamicum were extensively studied in the last two decades. Many metabolic pathways, regulatory and transport principles are known, but purely rational approaches often provide only limited progress in production optimization. We recently generated stable synthetic co-cultures, termed Communities of Niche-optimized Strains (CoNoS), that rely on cross-feeding of amino acids for growth. This setup has the potential to evolve strains with improved production by selection of faster growing communities. RESULTS: Here we performed adaptive laboratory evolution (ALE) with a CoNoS to identify mutations that are relevant for amino acid production both in mono- and co-cultures. During ALE with the CoNoS composed of strains auxotrophic for either L-leucine or L-arginine, we obtained a 23% growth rate increase. Via whole-genome sequencing and reverse engineering, we identified several mutations involved in amino acid transport that are beneficial for CoNoS growth. The L-leucine auxotrophic strain carried an expression-promoting mutation in the promoter region of brnQ (cg2537), encoding a branched-chain amino acid transporter in combination with mutations in the genes for the Na+/H+-antiporter Mrp1 (cg0326-cg0321). This suggested an unexpected link of Mrp1 to L-leucine transport. The L-arginine auxotrophic partner evolved expression-promoting mutations near the transcriptional start site of the yet uncharacterized operon argTUV (cg1504-02). By mutation studies and ITC, we characterized ArgTUV as the only L-arginine uptake system of C. glutamicum with an affinity of KD = 30 nM. Finally, deletion of argTUV in an L-arginine producer strain resulted in a faster and 24% higher L-arginine production in comparison to the parental strain. CONCLUSION: Our work demonstrates the power of the CoNoS-approach for evolution-guided identification of non-obvious production traits, which can also advance amino acid production in monocultures. Further rounds of evolution with import-optimized strains can potentially reveal beneficial mutations also in metabolic pathway enzymes. The approach can easily be extended to all kinds of metabolite cross-feeding pairings of different organisms or different strains of the same organism, thereby enabling the identification of relevant transport systems and other favorable mutations.

Cell diversity and network dynamics in photosensitive human brain organoids.

In vitro models of the developing brain such as three-dimensional brain organoids offer an unprecedented opportunity to study aspects of human brain development and disease. However, the cells generated within organoids and the extent to which they recapitulate the regional complexity, cellular diversity and circuit functionality of the brain remain undefined. Here we analyse gene expression in over 80,000 individual cells isolated from 31 human brain organoids. We find that organoids can generate a broad diversity of cells, which are related to endogenous classes, including cells from the cerebral cortex and the retina. Organoids could be developed over extended periods (more than 9 months), allowing for the establishment of relatively mature features, including the formation of dendritic spines and spontaneously active neuronal networks. Finally, neuronal activity within organoids could be controlled using light stimulation of photosensitive cells, which may offer a way to probe the functionality of human neuronal circuits using physiological sensory stimuli.

Connectome 2.0: Developing the next-generation ultra-high gradient strength human MRI scanner for bridging studies of the micro-, meso- and macro-connectome.

The first phase of the Human Connectome Project pioneered advances in MRI technology for mapping the macroscopic structural connections of the living human brain through the engineering of a whole-body human MRI scanner equipped with maximum gradient strength of 300 mT/m, the highest ever achieved for human imaging. While this instrument has made important contributions to the understanding of macroscale connectional topology, it has also demonstrated the potential of dedicated high-gradient performance scanners to provide unparalleled in vivo assessment of neural tissue microstructure. Building on the initial groundwork laid by the original Connectome scanner, we have now embarked on an international, multi-site effort to build the next-generation human 3T Connectome scanner (Connectome 2.0) optimized for the study of neural tissue microstructure and connectional anatomy across multiple length scales. In order to maximize the resolution of this in vivo microscope for studies of the living human brain, we will push the diffusion resolution limit to unprecedented levels by (1) nearly doubling the current maximum gradient strength from 300 mT/m to 500 mT/m and tripling the maximum slew rate from 200 T/m/s to 600 T/m/s through the design of a one-of-a-kind head gradient coil optimized to minimize peripheral nerve stimulation; (2) developing high-sensitivity multi-channel radiofrequency receive coils for in vivo and ex vivo human brain imaging; (3) incorporating dynamic field monitoring to minimize image distortions and artifacts; (4) developing new pulse sequences to integrate the strongest diffusion encoding and highest spatial resolution ever achieved in the living human brain; and (5) calibrating the measurements obtained from this next-generation instrument through systematic validation of diffusion microstructural metrics in high-fidelity phantoms and ex vivo brain tissue at progressively finer scales with accompanying diffusion simulations in histology-based micro-geometries. We envision creating the ultimate diffusion MRI instrument capable of capturing the complex multi-scale organization of the living human brain - from the microscopic scale needed to probe cellular geometry, heterogeneity and plasticity, to the mesoscopic scale for quantifying the distinctions in cortical structure and connectivity that define cyto- and myeloarchitectonic boundaries, to improvements in estimates of macroscopic connectivity.

Nanobody immunostaining for correlated light and electron microscopy with preservation of ultrastructure.

Morphological and molecular characteristics determine the function of biological tissues. Attempts to combine immunofluorescence and electron microscopy invariably compromise the quality of the ultrastructure of tissue sections. We developed NATIVE, a correlated light and electron microscopy approach that preserves ultrastructure while showing the locations of multiple molecular moieties, even deep within tissues. This technique allowed the large-scale 3D reconstruction of a volume of mouse hippocampal CA3 tissue at nanometer resolution.

Deciphering the molecular effects of romidepsin on germ cell tumours: DHRS2 is involved in cell cycle arrest but not apoptosis or induction of romidepsin effectors.

Testicular germ cell tumours (GCTs) mostly affect young men at age 17-40. Although high cure rates can be achieved by orchiectomy and chemotherapy, GCTs can still be a lethal threat to young patients with metastases or therapy resistance. Thus, alternative treatment options are needed. Based on studies utilising GCT cell lines, the histone deacetylase inhibitor romidepsin is a promising therapeutic option, showing high toxicity at very low doses towards cisplatin-resistant GCT cells, but not fibroblasts or Sertoli cells. In this study, we extended our analysis of the molecular effects of romidepsin to deepen our understanding of the underlying mechanisms. Patients will benefit from these analyses, since detailed knowledge of the romidepsin effects allows for a better risk and side-effect assessment. We screened for changes in histone acetylation of specific lysine residues and analysed changes in the DNA methylation landscape after romidepsin treatment of the GCT cell lines TCam-2, 2102EP, NCCIT and JAR, while human fibroblasts were used as controls. In addition, we focused on the role of the dehydrogenase/reductase DHRS2, which was strongly up-regulated in romidepsin treated cells, by generating DHRS2-deficient TCam-2 cells using CRISPR/Cas9 gene editing. We show that DHRS2 is dispensable for up-regulation of romidepsin effectors (GADD45B, DUSP1, ZFP36, ATF3, FOS, CDKN1A, ID2) but contributes to induction of cell cycle arrest. Finally, we show that a combinatory treatment of romidepsin plus the gluccocorticoid dexamethasone further boosts expression of the romidepsin effectors and reduces viability of GCT cells more strongly than under single agent treatment. Thus, romidepsin and dexamethasone might represent a new combinatorial approach for treatment of GCT.

Mechanism of the Water-Gas Shift Reaction Catalyzed by Efficient Ruthenium-Based Catalysts: A Computational and Experimental Study.

Supported ionic liquid phase (SILP) catalysis enables a highly efficient, Ru-based, homogeneously catalyzed water-gas shift reaction (WGSR) between 100 °C and 150 °C. The active Ru-complexes have been found to exist in imidazolium chloride melts under operating conditions in a dynamic equilibrium, which is dominated by the [Ru(CO)3 Cl3 ]- complex. Herein we present state-of-the-art theoretical calculations to elucidate the reaction mechanism in more detail. We show that the mechanism includes the intermediate formation and degradation of hydrogen chloride, which effectively reduces the high barrier for the formation of the requisite dihydrogen complex. The hypothesis that the rate-limiting step involves water is supported by using D2 O in continuous catalytic WGSR experiments. The resulting mechanism constitutes a highly competitive alternative to earlier reported generic routes involving nucleophilic addition of hydroxide in the gas phase and in solution.

Patients with small and diminutive proximal hyperplastic polyps have higher rates of synchronous advanced neoplasia compared with patients without serrated lesions.

BACKGROUND AND AIMS: The association of proximal small and diminutive hyperplastic polyps (HPs) with synchronous advanced neoplasia is not well-defined. However, sessile serrated polyps (SSPs), even when small, are known to portend a risk of synchronous neoplasia. Currently, the U.S. Multi-Society Task Force on Colorectal Cancer does not recommend a change in the surveillance interval when proximal small HPs are detected. We aimed to compare the rates of synchronous advanced neoplasia in a screening colonoscopy cohort of patients with small and then diminutive proximal HPs in comparison, first to a cohort absent any serrated or proximal HPs and then in comparison with a cohort with small proximal SSPs. METHODS: Consecutive screening colonoscopies were recorded between 2005 and 2010 at an academic medical center. Patients were divided into 3 mutually exclusive groups. Group 1 consisted of patients with at least 1 HP that was proximal to the sigmoid colon, <1 cm in endoscopic size, and up to 3 total HPs in number. Group 2 included patients without any proximal HPs or SSPs. Group 3 consisted of patients with 1 to 2 SSPs, with at least 1 being proximal to the sigmoid colon, that were <1 cm in endoscopic size and without dysplasia. Rates of synchronous advanced neoplasia in patients with small (<1 cm) and diminutive (≤5 mm) proximal HPs were compared with the rates for the other 2 groups. RESULTS: There were 482 of 2569 patients (18.8%) with a small proximal HP who met the criteria for Group 1. The rate of synchronous advanced neoplasia in patients with a small proximal HP (61/482, 12.7%) was significantly greater compared with the average risk in the non-serrated cohort (Group 2, 133/1878, 7.1%; P < .001). There was no significant difference in the rate of synchronous advanced neoplasia when the small proximal HP group was subdivided by size (≤5 mm, 51/404, 12.6% vs 6-9 mm, 10/78, 12.8%; P = 1.00). The rate of synchronous advanced neoplasia in patients with diminutive (≤5 mm) proximal HPs (51/404, 12.6%) was not significantly different from the rate observed with proximal SSPs of similar size (17/113, 15.0%; P = .529). CONCLUSION: Patients with small and diminutive proximal HPs tend to harbor higher rates of synchronous advanced neoplasia compared with those without any serrated lesions detected on screening colonoscopy. Surveillance outcomes for metachronous advanced neoplasia for patients with small proximal HPs deserves further study. The synchronous advanced neoplasia rate in patients with proximal diminutive HPs is similar to that of proximal diminutive SSPs and could have implications in a resect and discard strategy.

Adenoma detection rate metrics in colorectal cancer surveillance colonoscopy.

BACKGROUND: A target goal for screening adenoma detection rate (S-ADR) of ≥ 25% has been set to define high-quality colonoscopy performance. However, there is no current accepted target goal for ADR in colorectal cancer (CRC) surveillance. This makes quality assessment challenging when physicians perform cancer surveillance colonoscopy but minimal screening procedures. METHODS: In this cohort study, consecutive colonoscopies performed at either Rush University Medical Center or Rush Oak Park Hospital by a gastroenterologist or colorectal surgeon in average risk screening population and CRC surveillance population were reviewed retrospectively from 2006 to 2012 and prospectively from 2013 to 2016. ADR in first surveillance colonoscopy following surgical resection of CRC (CRC-ADR) was reported in high-quality detectors (HQD) or low-quality detectors (LQD) based on achievement of 25% ADR in consecutive screening colonoscopy in average risk patients. Pearson's correlation was used to describe the association between individual S-ADR and CRC-ADR for colonoscopists. RESULTS: There was a very strong positive correlation (r = 0.88, p = 0.002) between ADR in average risk screening and first time CRC surveillance. For HQD as defined by S-ADR ≥ 25% (n = 10 colonoscopists), the CRC-ADR was 37.7% (78/207, SD 8%) which was very similar to their respective S-ADR of 33.4% (816/2440, p = 0.22). For LQD (n = 5 colonoscopists), the CRC-ADR was 20.2% (40/198) which was similar to their respective S-ADR of 20.1% (119/591, p = 0.99). The CRC-ADR was significantly higher for HQD than for LQD (37.7 vs. 20.2%, p < 0.0001). CONCLUSIONS: The major finding of this study is a defined CRC-ADR for HQD based on the ability to achieve S-ADR ≥ 25%. S-ADR strongly correlates with CRC-ADR. CRC-ADR is quite similar to the colonoscopists' respective S-ADR for both HQD and LQD. For colonoscopists who perform limited screening colonoscopies but do perform CRC surveillance colonoscopies, ADR metrics similar to S-ADR to assess quality in colonoscopy could be considered.

Maintaining low non-neoplastic polypectomy rates in high-quality screening colonoscopy.

BACKGROUND AND AIMS: Non-neoplastic polypectomies (NNPs) add pathology and procedural costs but do not reduce cancer risk and should be minimized. We sought to define the minimal non-neoplastic polypectomy rate (NNPR) for those colonoscopists achieving high-quality colorectal cancer screening based on adenoma detection rates (ADRs). METHODS: NNPRs for colonoscopists achieving high-quality adenoma detection rates were reported to determine minimal NNPR goals. Two approaches to tracking NNPR monitoring were compared: (1) total NNPR, an NNPR inclusive of all non-neoplastic specimens with exclusion of only hyperplastic polyp, sessile serrated polyp, and adenoma; and (2) normal tissue-only NNPR, an NNPR inclusive of those specimens with only normal colonic mucosa or lymphoid follicles. RESULTS: For those performing colonoscopy with high-quality ADRs (≥25%), half (6/12) of the colonoscopists had a total NNPR of ≤8.5% and 2 gastroenterologists had a total NNPR of ≤3.4%. The mean total NNPR of the cohort was 8.7% versus the normal tissue only NNPR, which was 7.5% (mean difference of 1.2%, standard deviation ± 0.97). The widest variation between total NNPR versus normal tissue only NNPR for any colonoscopist was 2.9%. The total NNPR ranged between 2.6% and 21.3% among 14 colonoscopists. CONCLUSIONS: Colonoscopy with a high-quality ADR can be achieved while maintaining a low total NNPR. A total NNPR, inclusive of all non-neoplastic specimens as an alternative to an approach in which all specimens require individual review in order to select out only normal tissue can be considered for monitoring of NNPR.

Time-to-event analysis of surgically treated posthemorrhagic hydrocephalus in preterm infants: a single-institution retrospective study.

PURPOSE: The purpose of this study is to report time points relevant to the neurosurgical management of posthemorrhagic hydrocephalus (PHH). METHODS: Data were collected retrospectively on 104 preterm infants with intraventricular hemorrhage (IVH) who received neurosurgical intervention for PHH at St. Louis Children's Hospital from 1994 to 2016. Kaplan-Meier curves were constructed for various endpoints. RESULTS: IVH grade on head ultrasound obtained through routine clinical care was II, III, and IV in 5 (4.8%), 33 (31.7%), and 66 (63.5%) of the patients, respectively. Neither IVH size nor location appeared to affect development of PHH. Days from birth to IVH, ventriculomegaly, temporizing neurosurgical procedure (TNP), and permanent neurosurgical intervention were 2.0 (95% CI 1.7-2.3), 3.0 (2.5-3.5), 24.0 (22.2-25.8), and 101.0 (90.4-111.6), respectively. Grades III and IV IVH did not differ in age at IVH diagnosis (Χ 2 (1 d.f.) = 1.32, p = 0.25), ventriculomegaly (Χ 2 = 0.73, p = 0.40), TNP (Χ 2 = 0.61, p = 0.43), or permanent intervention (Χ 2 = 2.48, p = 0.17). Ventricular reservoirs and ventriculosubgaleal shunts were used in 71 (68.3%) and 30 (28.8%), respectively. Eighty (76.9%) of the patients ultimately received a VPS. Five (4.8%) underwent a primary endoscopic third ventriculostomy (ETV), and two (1.9%) had ETV for a revision procedure. Four of the seven ETVs had choroid plexus cauterization. CONCLUSIONS: Although most infants who develop IVH and ventriculomegaly will do so within a few days of birth, at-risk infants should be observed for at least 4 weeks with serial head ultrasounds to monitor for PHH requiring surgery.

Cerebrospinal Fluid Biomarkers of Pediatric Hydrocephalus.

Hydrocephalus (HC) is a common, debilitating neurological condition that requires urgent clinical decision-making. At present, neurosurgeons rely heavily on a patient's history, physical examination findings, neuroimaging, and clinical judgment to make the diagnosis of HC or treatment failure (e.g., shunt malfunction). Unfortunately, these tools, even in combination, do not eliminate subjectivity in clinical decisions. In order to improve the management of infants and children with HC, there is an urgent need for new biomarkers to complement currently available tools and enable clinicians to confidently establish the diagnosis of HC, assess therapeutic efficacy/treatment failure, and evaluate current and future developmental challenges, so that every child has access to the resources they need to optimize their outcome and quality of life.

Formation Mechanism of the First Carbon-Carbon Bond and the First Olefin in the Methanol Conversion into Hydrocarbons.

The elementary reactions leading to the formation of the first carbon-carbon bond during early stages of the zeolite-catalyzed methanol conversion into hydrocarbons were identified by combining kinetics, spectroscopy, and DFT calculations. The first intermediates containing a C-C bond are acetic acid and methyl acetate, which are formed through carbonylation of methanol or dimethyl ether even in presence of water. A series of acid-catalyzed reactions including acetylation, decarboxylation, aldol condensation, and cracking convert those intermediates into a mixture of surface bounded hydrocarbons, the hydrocarbon pool, as well as into the first olefin leaving the catalyst. This carbonylation based mechanism has an energy barrier of 80 kJ mol(-1) for the formation of the first C-C bond, in line with a broad range of experiments, and significantly lower than the barriers associated with earlier proposed mechanisms.

Embedded-cluster calculations in a numeric atomic orbital density-functional theory framework.

We integrate the all-electron electronic structure code FHI-aims into the general ChemShell package for solid-state embedding quantum and molecular mechanical (QM/MM) calculations. A major undertaking in this integration is the implementation of pseudopotential functionality into FHI-aims to describe cations at the QM/MM boundary through effective core potentials and therewith prevent spurious overpolarization of the electronic density. Based on numeric atomic orbital basis sets, FHI-aims offers particularly efficient access to exact exchange and second order perturbation theory, rendering the established QM/MM setup an ideal tool for hybrid and double-hybrid level density functional theory calculations of solid systems. We illustrate this capability by calculating the reduction potential of Fe in the Fe-substituted ZSM-5 zeolitic framework and the reaction energy profile for (photo-)catalytic water oxidation at TiO2(110).

When 'neutral' evidence still has probative value (with implications from the Barry George Case).

The likelihood ratio (LR) is a probabilistic method that has been championed as a 'simple rule' for evaluating the probative value of forensic evidence in court. Intuitively, if the LR is greater than one then the evidence supports the prosecution hypothesis; if the LR is less than one it supports the defence hypothesis, and if the LR is equal to one then the evidence favours neither (and so is considered 'neutral'-having no probative value). It can be shown by Bayes' theorem that this simple relationship only applies to pairs of hypotheses for which one is the negation of the other (i.e. to mutually exclusive and exhaustive hypotheses) and is not applicable otherwise. We show how easy it can be - even for evidence experts - to use pairs of hypotheses that they assume are mutually exclusive and exhaustive but are not, and hence to arrive at erroneous conclusions about the value of evidence using the LR. Furthermore, even when mutually exclusive and exhaustive hypotheses are used there are extreme restrictions as to what can be concluded about the probative value of evidence just from a LR. Most importantly, while the distinction between source-level hypotheses (such as defendant was/was not at the crime scene) and offence-level hypotheses (defendant is/is not guilty) is well known, it is not widely understood that a LR for evidence about the former generally has no bearing on the LR of the latter. We show for the first time (using Bayesian networks) the full impact of this problem, and conclude that it is only the LR of the offence level hypotheses that genuinely determines the probative value of the evidence. We investigate common scenarios in which evidence has a LR of one but still has significant probative value (i.e. is not neutral as is commonly assumed). As illustration we consider the ramifications of these points for the case of Barry George. The successful appeal against his conviction for the murder of Jill Dando was based primarily on the argument that the firearm discharge residue (FDR) evidence, assumed to support the prosecution hypothesis at the original trial, actually had a LR equal to one and hence was 'neutral'. However, our review of the appeal transcript shows numerous examples of the problems with the use of hypotheses identified above. We show that if one were to follow the arguments recorded in the Appeal judgement verbatim, then contrary to the Appeal conclusion, the probative value of the FDR evidence may not have been neutral as was concluded.

Introduction to Tobacco Cessation and Motivational Interviewing: Evaluation of a Lecture and Case-Based Learning Activity for Medical Students.

Introduction Despite the fact that tobacco use continues to have significant public health impacts, most healthcare providers are not adequately trained to counsel patients on their tobacco use or to provide cessation resources. Although all healthcare providers have a role in providing tobacco cessation assistance, physicians and advanced practice providers are generally the only practitioners able to furnish tobacco cessation medications and bill insurance for their cessation services. Therefore, ensuring these practitioners are properly trained to offer tobacco cessation to their patients is critical to addressing this public health threat. In line with this goal, this study outlines the curriculum evaluation for an innovative student-facilitated tobacco cessation activity for medical students. Methods A lecture and case-based learning activity was created and piloted with a class of first-year medical students. The activity was facilitated by fourth-year medical students. Students took a pre-session survey to establish baseline experience and beliefs and a post-session survey to ascertain their confidence in applying what was covered in the session. Descriptive statistics were utilized to analyze the data. Results One hundred and twenty-eight students completed both surveys. Prior to the activity, students reported low levels of confidence in their ability to counsel patients and knowledge of cessation resources. Following the activity, more than 90% reported improvement in their ability to assess a patient's willingness to quit and counsel those ready to quit. Greater than 80% reported an improvement in their ability to counsel patients not ready to quit and to establish a quit plan. More than 90% of students reported that the session increased their self-efficacy in helping patients quit and that it was worth their time, with 96% committing to increasing their tobacco cessation efforts with their patients. Discussion Students valued the training and almost all reported that it increased their ability to help patients quit smoking. The use of student-facilitated case-based learning provided both an opportunity for students to practice cessation techniques and a low-stakes introduction to the OSCE format without the need for extensive faculty resources. Although this session was run with first-year medical students, the curriculum presented can be used for residents, nurses, and other healthcare professionals.

Extensive soma-soma plate-like contact sites (ephapses) connect suprachiasmatic nucleus neurons.

The hypothalamic suprachiasmatic nucleus (SCN) is the central pacemaker for mammalian circadian rhythms. As such, this ensemble of cell-autonomous neuronal oscillators with divergent periods must maintain coordinated oscillations. To investigate ultrastructural features enabling such synchronization, 805 coronal ultrathin sections of mouse SCN tissue were imaged with electron microscopy and aligned into a volumetric stack, from which selected neurons within the SCN core were reconstructed in silico. We found that clustered SCN core neurons were physically connected to each other via multiple large soma-to-soma plate-like contacts. In some cases, a sliver of a glial process was interleaved. These contacts were large, covering on average ∼21% of apposing neuronal somata. It is possible that contacts may be the electrophysiological substrate for synchronization between SCN neurons. Such plate-like contacts may explain why the synchronization of SCN neurons is maintained even when chemical synaptic transmission or electrical synaptic transmission via gap junctions is blocked. Such ephaptic contact-mediated synchronization among nearby neurons may therefore contribute to the wave-like oscillations of circadian core clock genes and calcium signals observed in the SCN.

Three­dimensional reconstruction of SCN tissue via serial electron microscopy revealed a novel structural feature of SCN neurons that may account for interneuronal synchronization that persists even when the predominant mechanisms of neuronal communication are blocked. We found that SCN core neurons are connected by multiple soma­soma contact specializations, ultrastructural elements that could enable synchronization of tightly packed neurons organized in clustered networks. This extensive network of plate­like soma­soma contacts among clustered SCN neurons may provide insight into how ∼20,000 autonomous neuronal oscillators with a broad range of intrinsic periods remain synchronized in the absence of ordinary communication modalities, thereby conferring the resilience required for the SCN to function as the mammalian circadian pacemaker.