Spatial and temporal aspects of neuronal calcium and sodium signals measured with low-affinity fluorescent indicators.

Low-affinity fluorescent indicators for Ca2+ or Na+ allow measuring the dynamics of intracellular concentration of these ions with little perturbation from physiological conditions because they are weak buffers. When using synthetic indicators, which are small molecules with fast kinetics, it is also possible to extract spatial and temporal information on the sources of ion transients, their localization, and their disposition. This review examines these important aspects from the biophysical point of view, and how they have been recently exploited in neurophysiological studies. We first analyze the environment where Ca2+ and Na+ indicators are inserted, highlighting the interpretation of the two different signals. Then, we address the information that can be obtained by analyzing the rising phase and the falling phase of the Ca2+ and Na+ transients evoked by different stimuli, focusing on the kinetics of ionic currents and on the spatial interpretation of these measurements, especially on events in axons and dendritic spines. Finally, we suggest how Ca2+ or Na+ imaging using low-affinity synthetic fluorescent indicators can be exploited in future fundamental or applied research.

Injectable ultrasound-powered bone-adhesive nanocomposite hydrogel for electrically accelerated irregular bone defect healing.

The treatment of critical-size bone defects with irregular shapes remains a major challenge in the field of orthopedics. Bone implants with adaptability to complex morphological bone defects, bone-adhesive properties, and potent osteogenic capacity are necessary. Here, a shape-adaptive, highly bone-adhesive, and ultrasound-powered injectable nanocomposite hydrogel is developed via dynamic covalent crosslinking of amine-modified piezoelectric nanoparticles and biopolymer hydrogel networks for electrically accelerated bone healing. Depending on the inorganic-organic interaction between the amino-modified piezoelectric nanoparticles and the bio-adhesive hydrogel network, the bone adhesive strength of the prepared hydrogel exhibited an approximately 3-fold increase. In response to ultrasound radiation, the nanocomposite hydrogel could generate a controllable electrical output (-41.16 to 61.82 mV) to enhance the osteogenic effect in vitro and in vivo significantly. Rat critical-size calvarial defect repair validates accelerated bone healing. In addition, bioinformatics analysis reveals that the ultrasound-responsive nanocomposite hydrogel enhanced the osteogenic differentiation of bone mesenchymal stem cells by increasing calcium ion influx and up-regulating the PI3K/AKT and MEK/ERK signaling pathways. Overall, the present work reveals a novel wireless ultrasound-powered bone-adhesive nanocomposite hydrogel that broadens the therapeutic horizons for irregular bone defects.

Performance and attitudes toward real-time computer-aided polyp detection during colonoscopy in a large tertiary referral center in the United States.

BACKGROUND AND AIMS: Computer-aided detection (CADe) has been shown to improve polyp detection in clinical trials. Limited data exist on the impact, utilization, and attitudes toward artificial intelligence (AI)-assisted colonoscopy in daily clinical practice. We aimed to evaluate the effectiveness of the first U.S. Food and Drug Administration-approved CADe device for polyp detection in the United States and the attitudes toward its implementation. METHODS: We performed a retrospective analysis of a prospectively maintained database of patients undergoing colonoscopy at a tertiary center in the United States before and after a real-time CADe system was made available. The decision to activate the CADe system was at the discretion of the endoscopist. An anonymous survey was circulated among endoscopy physicians and staff at the beginning and conclusion of the study period regarding their attitudes toward AI-assisted colonoscopy. RESULTS: CADe was activated in 52.1% of cases. Compared with historical control subjects, there was no statistically significant difference in adenomas detected per colonoscopy (1.08 vs 1.04, P = .65), even after excluding diagnostic and therapeutic indications and cases where CADe was not activated (1.27 vs 1.17, P = .45). In addition, there was no statistically significant difference in adenoma detection rate (ADR), median procedure, and withdrawal times. Survey results demonstrated mixed attitudes toward AI-assisted colonoscopy, of which main concerns were high number of false-positive signals (82.4%), high level of distraction (58.8%), and impression it prolonged procedure time (47.1%). CONCLUSIONS: CADe did not improve adenoma detection in daily practice among endoscopists with high baseline ADRs. Despite its availability, AI-assisted colonoscopy was only activated in half of the cases, and multiple concerns were raised by staff and endoscopists. Future studies will help elucidate the patients and endoscopists that would benefit most from AI-assisted colonoscopy.

Biliary Drainage in Hilar and Perihilar Cholangiocarcinoma; 25-Year Experience at a Tertiary Cancer Center.

INTRODUCTION: This study analyzes the optimal biliary stenting strategy for palliation in cholangiocarcinoma (CCA). METHODS: This is a retrospective study of patients with CCA who underwent biliary drainage from 1997-2023. A per-patient analysis of percutaneous biliary drainage (PTBD) rates, the median number of ERCPs, overall survival (OS), and a per-procedure analysis of clinical success (CS), stent-specific Adverse Events (AEs), and mean time to reintervention by stent type and laterality (unilateral(u) & bilateral(b)) is presented. RESULTS: A total of 333 patients underwent 1,050 ERCPs; 85% with plastic stents (PS). PTBD was eventually done in 23% of PS patients, 35% of whom had PS removed prior to PTBD. ERCPs with SEMS/uniSEMS use had higher CS (89%/91%) vs PS/uPS (85% both) and PS within SEMS (PS-SEMS)/uPS-SEMS (71%/74%;p=0.013/p=0.054). Compared to PS, SEMS and PS-SEMS were associated with higher stent-specific AEs (OR SEMS 4.85; 3.23-7.27; PS-SEMS 9.99; 5.33-18.71;p<0.001). Straight PS were associated with more stent-specific AEs compared to double-pigtail stents (OR 6.74; 3.95-11.45;p<0.001). More 7 Fr stents were used in cases with balloon dilation (BD, 109 vs. 88 with no BD; p<0.001). BD had 79% CS rate vs 87% without BD (p<0.001). Cases with pus on ERCP and those with BD had a shorter mean time to reintervention. On regression analyses, higher Bismuth class, PS use, and PS-SEMS use were associated with a shorter mean time to repeat ERCP. 52% of patients in the bSEMS arm died from cholangitis (p=0.005). CONCLUSION: The relatively higher clinical success of SEMS is countered by the higher stent-specific complication rate. PS can be removed and may better facilitate PTBD. Within PS types, DPTs may have fewer stent-specific AEs. Cases requiring balloon dilation and with endoscopic evidence of pus may benefit from earlier reintervention.

Endoscopy for the diagnosis and treatment of gastrointestinal bleeding caused by malignancy.

BACKGROUND AND AIM: The diagnosis and treatment of gastrointestinal (GI) bleeding secondary to malignancy can be challenging. Endoscopy is the gold standard to diagnose and treat gastrointestinal bleeding but clinical characteristics and outcomes of patients with malignancy-related bleeding are not well understood. This study aims to look at clinical characteristics, endoscopic findings, safety and clinical outcomes of endoscopic interventions for GI malignancy-related bleeding. METHODS: We retrospectively reviewed outcomes of patients with confirmed GI malignancies who underwent endoscopy for GI bleeding at MD Anderson Cancer Center between 2010 and 2019. Cox hazard analysis was conducted to identify factors associated with survival. RESULTS: A total of 313 patients were included, with median age of 59 years; 74.8% were male. The stomach (30.0%) was the most common tumor location. Active bleeding was evident endoscopically in 47.3% of patients. Most patients (77.3%) did not receive endoscopic treatment. Of the patients who received endoscopic treatment, 57.7% had hemostasis. No endoscopy-related adverse events were recorded. Endoscopic treatment was associated with hemostasis (P < 0.001), but not decreased recurrent bleeding or mortality. Absence of active bleeding on endoscopy, stable hemodynamic status at presentation, lower cancer stage, and surgical intervention were associated with improved survival. CONCLUSIONS: This study indicates that endoscopy is a safe diagnostic tool in this patient population; while endoscopic treatments may help achieve hemostasis, it may not decrease the risk of recurrent bleeding or improve survival.

How much progress have we made?: a 20-year experience regarding esophageal stents for the palliation of malignant dysphagia.

Esophageal stents are widely used for the palliation of malignant esophageal obstruction. Advances in technology have made esophageal stenting technically feasible and widespread for such obstruction, but complications remain frequent. We present outcomes of a large cohort undergoing esophageal stent placement for malignant esophageal obstruction at a tertiary care cancer center. Patients who underwent placement of esophageal stents for malignancy-related esophageal obstruction between 1 January 2001 and 31 July 2020 were identified. Exclusion criteria included stents placed for benign stricture, fistulae, obstruction of proximal esophagus (proximal to 24 cm from incisors), or post-surgical indications. Patient charts were reviewed for demographics, procedure and stent characteristics, complications, and follow-up. A total of 242 patients underwent stent placement (median age: 64 years, 79.8% male). The majority, 204 (84.3%), had esophageal cancer. During the last two decades, there has been an increasing trend in the number of esophageal stents placed. Though plastic stents were previously used, these are no longer utilized. Complications are frequent and include early complications of pain in 68 (28.1%) and migration in 21 (8.7%) and delayed complications of recurrent symptoms of dysphagia in 46 (19.0%) and migration in 26 (10.7%). Over the study period, there has not been a significant improvement in the rate of complications. During follow-up, 92 (38%) patients required other enteral nutrition modalities after esophageal stent placement. No patient, treatment, or stent characteristics were significantly associated with stent complication or outcome. Esophageal stent placement is an increasingly popular method for palliation of malignant dysphagia. However, complications, particularly pain, migration, and recurrent symptoms of dysphagia are common. Almost 40% of patients may also require other methods of enteral access after esophageal stent placement. Given the high complication rates and suboptimal outcomes, removable stents should be considered as first-line in the case of poor palliative response.

Mechanism of ArcLight derived GEVIs involves electrostatic interactions that can affect proton wires.

The genetically encoded voltage indicators ArcLight and its derivatives mediate voltage-dependent optical signals by intermolecular, electrostatic interactions between neighboring fluorescent proteins (FPs). A random mutagenesis event placed a negative charge on the exterior of the FP, resulting in a greater than 10-fold improvement of the voltage-dependent optical signal. Repositioning this negative charge on the exterior of the FP reversed the polarity of voltage-dependent optical signals, suggesting the presence of "hot spots" capable of interacting with the negative charge on a neighboring FP, thereby changing the fluorescent output. To explore the potential effect on the chromophore state, voltage-clamp fluorometry was performed with alternating excitation at 390 nm followed by excitation at 470 nm, resulting in several mutants exhibiting voltage-dependent, ratiometric optical signals of opposing polarities. However, the kinetics, voltage ranges, and optimal FP fusion sites were different depending on the wavelength of excitation. These results suggest that the FP has external, electrostatic pathways capable of quenching fluorescence that are wavelength specific. One mutation to the FP (E222H) showed a voltage-dependent increase in fluorescence when excited at 390 nm, indicating the ability to affect the proton wire from the protonated chromophore to the H222 position. ArcLight-derived sensors may therefore offer a novel way to map how conditions external to the ß-can structure can affect the fluorescence of the chromophore and transiently affect those pathways via conformational changes mediated by manipulating membrane potential.

Efficacy of uncovered colonic stents for extrinsic versus intrinsic malignant large bowel obstruction.

BACKGROUND: Previous studies evaluating self-expandable metal stents (SEMS) for management of malignant extrinsic colon obstruction have yielded conflicting results. We evaluated the efficacy of uncovered SEMS for extrinsic colon malignancy (ECM) versus intrinsic colon malignancy (ICM). METHODS: Retrospective review of all patients referred for colonic SEMS at a tertiary cancer center between 2007 and 2018 was performed. Primary outcome measures were technical success, clinical success, intervention rate, and overall survival. RESULTS: 138 patients with ECM and 119 patients with ICM underwent attempted stent placement. The rectum and/or sigmoid colon was the most common stricture site. Technical success was lower in the ECM group [86% vs 96% (p = .009)]. Clinical success was lower in the ECM group both at 7 days [82% vs 95% (p = .004)] and at 90 days [60% vs 86% (p < .001)]. Subsequent intervention was required more frequently [44% vs 35%; p = .23] and earlier [median 9 vs 132 days; p < .001] in the ECM group. Median overall survival in the ECM group was 92 vs 167 days. Among predictive variables analyzed, the ECM group had a higher frequency of peritoneal metastasis (87% vs 32%; p < .001), multifocal strictures with requirement for multiple stents (20% vs 6%; p = .002), sharp angulated strictures (39% vs 25%; p = .04) , and radiation therapy (21% vs 10%; p = .02). CONCLUSIONS: Colonic SEMS for ECM is associated with lower technical and clinical success with earlier intervention rates compared with ICM. Our findings can be used to better inform patients and referring providers as well as guide new stent design to enhance efficacy in this population.

Assessment of genetic referrals and outcomes for women with triple negative breast cancer in regional cancer centres in Australia.

BACKGROUND: Guidelines for referral to cancer genetics service for women diagnosed with triple negative breast cancer have changed over time. This study was conducted to assess the changing referral patterns and outcomes for women diagnosed with triple negative breast cancer across three regional cancer centres during the years 2014-2018. METHODS: Following ethical approval, a retrospective electronic medical record review was performed to identify those women diagnosed with triple negative breast cancer, and whether they were referred to a genetics service and if so, the outcome of that genetics assessment and/or genetic testing. RESULTS: There were 2441 women with newly diagnosed breast cancer seen at our cancer services during the years 2014-2018, of whom 237 women were diagnosed with triple negative breast cancer. Based on age of diagnosis criteria alone, 13% (31/237) of our cohort fulfilled criteria for genetic testing, with 81% (25/31) being referred to a cancer genetics service. Of this group 68% (21/31) were referred to genetics services within our regions and went on to have genetic testing with 10 pathogenic variants identified; 5x BRCA1, 4x BRCA2 and × 1 ATM:c.7271 T > G. CONCLUSIONS: Referral pathways for women diagnosed with TNBC to cancer genetics services are performing well across our cancer centres. We identified a group of women who did not meet eligibility criteria for referral at their time of diagnosis, but would now be eligible, as guidelines have changed. The use of cross-discipline retrospective data reviews is a useful tool to identify patients who could benefit from being re-contacted over time for an updated cancer genetics assessment.

Safety and efficacy of metal stents for malignant colonic obstruction in patients treated with bevacizumab.

BACKGROUND AND AIMS: The aim of this study was to examine clinical outcomes and adverse events (AEs) of self-expandable metal stents (SEMSs) in the management of malignant colonic obstruction (MCO). METHODS: Patients with SEMSs for MCO treated at our institution from 2007 to 2016 were included. Clinical success was defined as successful oral intake after the procedure and technical success as stent deployment across the stricture in the desired location. RESULTS: Of 199 patients, the mean age was 58, 54% were men, and 99% had stage IV cancer. MCO etiology was colorectal cancer in 82% and extrinsic compression in 17%. Technical success was achieved in 99.5% and clinical success in 89%. The SEMSs were palliative in 97% and were a bridge to surgery in 4%. MCO occurred in the left side of the colon in 90%, transverse in 4.5%, and ascending colon in 5.5%. SEMSs were placed in curved segments in 30% and straight segments in 70%. Tandem SEMSs were required in 27 patients. Forty-six patients had 48 AEs (24%), including 2% periprocedure, 15% postprocedure, and 83% after 72 hours. Stent-related AEs (n = 25) included persistent obstruction (n = 14), occlusion (n = 10), and failure of expansion (n = 1). Procedural AEs (n = 23) included minor bleeding (n = 2), perforations (n = 4), abdominal pain (n = 12), stent migration (n = 4), and respiratory insufficiency (n = 1). Repeat procedures were performed in 21 of 46 patients. After SEMSs, 48 patients underwent surgery, including resection with primary anastomosis (n = 8), resection with definitive stoma (n = 18), and diverting stoma without resection (n = 19). Mean time to surgery after SEMS placement was 175 days. Postsurgical AEs occurred in those with resections (leak, 2; infection, 2). Of 104 receiving bevacizumab, 22% had AEs, including 1 perforation compared with 3 in the nonbevacizumab group (P = .549). Mean overall survival was 5.6 months. Extrinsic compression and curved strictures were associated with poor clinical success by univariate analysis and etiology (noncolonic with poor outcome) by multivariate analysis. CONCLUSIONS: SEMSs for MCO has high technical but suboptimal clinical success. Curved strictures and extrinsic compression are associated with poor outcomes. The perforation rate was not higher in the bevacizumab compared with the nonbevacizumab group, although this should be further validated in a larger population.

Safety of endoscopy in cancer patients with thrombocytopenia and neutropenia.

BACKGROUND AND AIMS: Cancer patients are prone to thrombocytopenia and neutropenia, which increase the risk of bleeding and infection. We assessed the safety of endoscopic procedures in cancer patients with thrombocytopenia and/or neutropenia. METHODS: We studied consecutive cancer patients with thrombocytopenia and/or neutropenia who underwent endoscopic procedures from 2010 through 2015. Neutropenia was defined as an absolute neutrophil count (ANC) <1000 cells/µL, and thrombocytopenia as a platelet count <100 × 103/µL. Univariate and multivariate generalized estimating equation models were used to assess factors associated with risk of adverse events (AEs) or death. RESULTS: We identified 588 patients who underwent 783 procedures; 608 procedures were performed in the setting of thrombocytopenia and 675 procedures in the setting of neutropenia. Concurrent neutropenia and thrombocytopenia were recorded in 500 endoscopies. Twenty-four patients (4.1%) experienced infectious AEs, whereas 29 (4.9%) experienced bleeding AEs within 1 week of the procedure. On multivariate analysis, platelet count ≤50 × 103/µL was associated with risk of bleeding AEs. In contrast, poor performance status was associated with increased risk of infection AEs (P < .01). No association was observed between low ANC and infectious AEs. Poor performance status (P < .01) and platelet count ≤100 × 103/µL (P < .05) were associated with increased risk of 30-day mortality. A persistent platelet count <20 × 103/µL after the procedure, with a baseline platelet count of ≤20 × 103/µL before the procedure, was associated with significant risk of bleeding AEs compared with a platelet count >20 × 103/µL after the procedure (P < .01); furthermore, if the platelet count increased to >50 × 103/µL after the procedure, the bleeding risk after the procedure was greatly reduced (P < .01). CONCLUSIONS: Endoscopic procedures are relatively safe in cancer patients with platelet count >50 × 103/µL. Nevertheless, a platelet count of ≥20 × 103/µL could be an appropriate threshold for platelet transfusion if 50 × 103/µL is difficult to achieve. The functional status of the patient, in the absence of the need for urgent or necessary endoscopic interventions, should be considered when deciding whether to perform endoscopy. The risk of procedure and the ANC did not seem to affect the outcomes.

Biliary Decompression in Perihilar Cholangiocarcinoma Improves Survival: A Single-Center Retrospective Analysis.

BACKGROUND AND AIMS: The complex biliary strictures of perihilar cholangiocarcinoma present significant challenges for providing adequate and long-lasting biliary drainage. The best approach to relieve obstruction remains controversial. The purpose of this study was to assess stenting outcomes in perihilar cholangiocarcinoma. METHODS: This study was approved by the center's institutional review board. Subjects with a diagnosis of perihilar cholangiocarcinoma who underwent endoscopic retrograde cholangiopancreatography (ERCP) were identified from endoscopic and pathologic databases from 1997 to 2014. Patient characteristics, endoscopic data, and follow-up evaluation data were retrospectively collected via review of available medical records. RESULTS: A total of 199 patients with perihilar cholangiocarcinoma who underwent a total of 504 ERCPs were included in the study. Nine of 504 (1.8%) procedures were technical failures. Among the 495 technically successful procedures, 347 (70.1%) procedures were clinical successes. Clinical success was significantly associated with longer overall survival (HR 0.57; p = 0.002). A higher proportion of patients with bilateral drainage had clinical success, compared with those with unilateral drainage. Cholangitis was not more common in the bilateral group compared to the unilateral group except in the group where a segment was not drained (1.9% vs 1.6% vs 7.1%, respectively). Patients with metal stents were 3.8 times more likely to have clinical success than those with plastic stents. CONCLUSIONS: In conclusion, adequate biliary drainage improves overall survival. Bilateral stenting if anatomy permits with self-expanding metal stents rather than plastic stents appears to provide the optimal chance of clinical success.

Sodium Dynamics in Pyramidal Neuron Dendritic Spines: Synaptically Evoked Entry Predominantly through AMPA Receptors and Removal by Diffusion.

Dendritic spines are key elements underlying synaptic integration and cellular plasticity, but many features of these important structures are not known or are controversial. We examined these properties using newly developed simultaneous sodium and calcium imaging with single-spine resolution in pyramidal neurons in rat hippocampal slices from either sex. Indicators for both ions were loaded through the somatic patch pipette, which also recorded electrical responses. Fluorescence changes were detected with a high-speed, low-noise CCD camera. Following subthreshold electrical stimulation, postsynaptic sodium entry is almost entirely through AMPA receptors with little contribution from entry through NMDA receptors or voltage-gated sodium channels. Sodium removal from the spine head is through rapid diffusion out to the dendrite through the spine neck with a half-removal time of ∼16 ms, which suggests the neck has low resistance. Peak [Na+]i changes during single EPSPs are ∼5 mm Stronger electrical stimulation evoked small plateau potentials that had significant longer-lasting localized [Na+]i increases mediated through NMDA receptors.SIGNIFICANCE STATEMENT Dendritic spines, small structures that are difficult to investigate, are important elements in the fundamental processes of synaptic integration and plasticity. The main tool for examining these structures has been calcium imaging. However, the kinds of information that calcium imaging reveals is limited. We used newly developed, high-speed, simultaneous sodium and calcium imaging to examine ion dynamics in spines in hippocampal pyramidal neurons. We found that following single subthreshold synaptic activation most sodium entry was through AMPA receptors and not through NMDA receptors or through voltage-gated sodium channels and that the spine neck is not a significant resistance barrier. Most spine mechanisms are linear. However, regenerative NMDA conductances can be activated with stronger stimulation.

Pancreatic cancer screening in high-risk individuals with germline genetic mutations.

BACKGROUND AND AIMS: Pancreatic cancer (PC) is a deadly disease that is most commonly diagnosed at an incurable stage. Different high-risk genetic variants and cancer syndromes increase the lifetime risk of developing PC. This study aims to assess the yield of initial PC screening in patients with high-risk germline mutations. METHODS: Asymptomatic adults underwent PC screening by EUS, magnetic resonance imaging, or CT during a 10-year period and were retrospectively identified. High-risk individuals were defined as carrying germline mutations in BRCA1, BRCA2, p53 (Li-Fraumeni), STK11 (Peutz-Jeghers), MSH2 (Lynch), ATM (ataxia-telangiectasia), or APC (familial adenomatous polyposis). Patients without germline mutations were excluded. RESULTS: In total, 86 patients met the study criteria. The median age was 48.5 years (interquartile range, 40-58), 79.1% (68) were women, and 43.0% (37) had a family history of PC. The genetic mutations were BRCA2 (50, 58.1%), BRCA1 (14, 16.3%), p53 (12, 14.0%), STK11 (5, 5.8%), MSH2 (3, 3.5%), ATM (1, 1.2%), and APC (1, 1.2%). Screening detected a pancreatic abnormality (PA) in 26.7% (23/86), including cysts (11, 47.8%), hyperechoic strands and foci (10, 43.5%), and mild pancreatic duct dilation (2, 8.7%). Patients older than 60 years were more likely to have a PA detected (P = .043). EUS detected more PAs than magnetic resonance imaging or CT. No cases of PC were diagnosed by screening or during follow-up (median, 29.8 months; interquartile range, 21.7-43.5). CONCLUSIONS: Unless indicated otherwise by family or personal history, PC screening under the age of 50 is low yield. Linear EUS may be the preferred modality for initial PC screening.

The Health Impacts of Environmental Policy: The North Carolina Clean Smokestacks Act.

The North Carolina Clean Smokestacks Act and related policies led to substantial decreases of emitted air pollutants from coal-fired power plants. Improved air quality was associated with statewide improvements in respiratory, cardiovascular, and cerebrovascular health in North Carolina. The effectiveness of environmental policies can be monitored for impact on both environmental and health outcomes.

Hypocretin/Orexin Peptides Alter Spike Encoding by Serotonergic Dorsal Raphe Neurons through Two Distinct Mechanisms That Increase the Late Afterhyperpolarization.

UNLABELLED: Orexins (hypocretins) are neuropeptides that regulate multiple homeostatic processes, including reward and arousal, in part by exciting serotonergic dorsal raphe neurons, the major source of forebrain serotonin. Here, using mouse brain slices, we found that, instead of simply depolarizing these neurons, orexin-A altered the spike encoding process by increasing the postspike afterhyperpolarization (AHP) via two distinct mechanisms. This orexin-enhanced AHP (oeAHP) was mediated by both OX1 and OX2 receptors, required Ca(2+) influx, reversed near EK, and decayed with two components, the faster of which resulted from enhanced SK channel activation, whereas the slower component decayed like a slow AHP (sAHP), but was not blocked by UCL2077, an antagonist of sAHPs in some neurons. Intracellular phospholipase C inhibition (U73122) blocked the entire oeAHP, but neither component was sensitive to PKC inhibition or altered PKA signaling, unlike classical sAHPs. The enhanced SK current did not depend on IP3-mediated Ca(2+) release but resulted from A-current inhibition and the resultant spike broadening, which increased Ca(2+) influx and Ca(2+)-induced-Ca(2+) release, whereas the slower component was insensitive to these factors. Functionally, the oeAHP slowed and stabilized orexin-induced firing compared with firing produced by a virtual orexin conductance lacking the oeAHP. The oeAHP also reduced steady-state firing rate and firing fidelity in response to stimulation, without affecting the initial rate or fidelity. Collectively, these findings reveal a new orexin action in serotonergic raphe neurons and suggest that, when orexin is released during arousal and reward, it enhances the spike encoding of phasic over tonic inputs, such as those related to sensory, motor, and reward events. SIGNIFICANCE STATEMENT: Orexin peptides are known to excite neurons via slow postsynaptic depolarizations. Here we elucidate a significant new orexin action that increases and prolongs the postspike afterhyperpolarization (AHP) in 5-HT dorsal raphe neurons and other arousal-system neurons. Our mechanistic studies establish involvement of two distinct Ca(2+)-dependent AHP currents dependent on phospholipase C signaling but independent of IP3 or PKC. Our functional studies establish that this action preserves responsiveness to phasic inputs while attenuating responsiveness to tonic inputs. Thus, our findings bring new insight into the actions of an important neuropeptide and indicate that, in addition to producing excitation, orexins can tune postsynaptic excitability to better encode the phasic sensory, motor, and reward signals expected during aroused states.

Transcriptional control of photosynthetic capacity: conservation and divergence from Arabidopsis to rice.

Contents 32 I. 32 II. 33 III. 36 IV. 41 43 References 43 SUMMARY: Photosynthesis is one of the most important biological processes on Earth. It provides the consumable energy upon which almost all organisms are dependent, and modulates the composition of the planet's atmosphere. To carry out photosynthesis, plants require a large cohort of genes. These genes encode proteins that capture light energy, store energy in sugars and build the subcellular structures required to facilitate this energy capture. Although the function of many of these genes is known, little is understood about the transcriptional networks that coordinate their expression. This review places our understanding of the transcriptional regulation of photosynthesis in Arabidopsis thaliana in an evolutionary context, to provide new insight into transcriptional regulatory networks that control photosynthesis gene expression in grasses. The similarities and differences between the rice and Arabidopsis networks are highlighted, revealing substantial disparity between the two systems. In addition, avenues are identified that may be exploited for photosynthesis engineering projects in the future.

Understanding calcium waves and sparks in central neurons.

All cells use changes in intracellular calcium concentration ([Ca(2+)](i)) to regulate cell signalling events. In neurons, with their elaborate dendritic and axonal arborizations, there are clear examples of both localized and widespread Ca(2+) signals. [Ca(2+)](i) changes that are generated by Ca(2+) entry through voltage- and ligand-gated channels are the best characterized. In addition, the release of Ca(2+) from intracellular stores can result in increased [Ca(2+)](i); the signals that trigger this release have been less well-studied, in part because they are not usually associated with specific changes in membrane potential. However, recent experiments have revealed dramatic widespread Ca(2+) waves and localized spark-like events, particularly in dendrites. Here we review emerging data on the nature of these signals and their functions.

An international randomised trial of celecoxib versus celecoxib plus difluoromethylornithine in patients with familial adenomatous polyposis.

BACKGROUND AND AIM: Although Non-steroidal anti-inflammatory drugs reduce colorectal adenoma burden in familial adenomatous polyposis (FAP), the utility of combining chemopreventive agents in FAP is not known. We conducted a randomised trial of celecoxib (CXB) versus CXB+diflouromethylornithine (DFMO) to determine the synergistic effect, if any. METHODS: The primary endpoint was % change in adenoma count in a defined field. Secondary endpoints were adenoma burden (weighted by adenoma diameter) and video review of entire colon/rectal segments. Adverse event (AEs) were monitored by National Cancer Institution toxicity criteria. RESULTS: 112 subjects were randomised: 60 men and 52 women at a mean age of 38 years. For the 89 patients who had landmark-matched polyp counts available at baseline and 6 months, the mean % change in adenoma count over the 6 months of trial was -13.0% for CXB+DFMO and -1.0% for CXB (p=0.69). Mean % change in adenoma burden was -40% (CXB+DFMO) vs -27% (CXB) (p=0.13). Video-based global polyp change was -0.80 for CXB+DFMO vs -0.33 for CXB (p=0.03). Fatigue was the only significant AE, worse on the CXB arm (p=0.02). CONCLUSIONS: CXB combined with DFMO yielded moderate synergy according to a video-based global assessment. No significant difference in adenoma count, the primary endpoint, was seen between the two study arms. No evidence of DFMO-related ototoxicity was seen. There were no adverse cardiovascular outcomes in either trial arm and no significant increase in AEs in the CXB+DFMO arm of the trial. Differences in outcomes between primary and secondary endpoints may relate to sensitivity of the endpoint measures themselves. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov number N01-CN95040.