Reduced ATGL-mediated lipolysis attenuates ß-adrenergic-induced AMPK signaling, but not the induction of PKA-targeted genes, in adipocytes and adipose tissue.

5'-AMP-activated protein kinase (AMPK) is activated as a consequence of lipolysis and has been shown to play a role in regulation of adipose tissue mitochondrial content. Conversely, the inhibition of lipolysis has been reported to potentiate the induction of protein kinase A (PKA)-targeted genes involved in the regulation of oxidative metabolism. The purpose of the current study was to address these apparent discrepancies and to more fully examine the relationship between lipolysis, AMPK, and the ß-adrenergic-mediated regulation of gene expression. In 3T3-L1 adipocytes, the adipose tissue triglyceride lipase (ATGL) inhibitor ATGListatin attenuated the Thr(172) phosphorylation of AMPK by a ß3-adrenergic agonist (CL 316,243) independent of changes in PKA signaling. Similarly, CL 316,243-induced increases in the Thr(172) phosphorylation of AMPK were reduced in adipose tissue from whole body ATGL-deficient mice. Despite reductions in the activation of AMPK, the induction of PKA-targeted genes was intact or, in some cases, increased. Similarly, markers of mitochondrial content and respiration were increased in adipose tissue from ATGL knockout mice independent of changes in the Thr(172) phosphorylation of AMPK. Taken together, our data provide evidence that AMPK is not required for the regulation of adipose tissue oxidative capacity in conditions of reduced fatty acid release.

Voluntary wheel running attenuates lipopolysaccharide-induced liver inflammation in mice.

Sepsis induces an acute inflammatory response in the liver, which can lead to organ failure and death. Given the anti-inflammatory effects of exercise, we hypothesized that habitual physical activity could protect against acute sepsis-induced liver inflammation via mechanisms, including heat shock protein (HSP) 70/72. Male C57BL/6J mice (n = 80, ∼8 wk of age) engaged in physical activity via voluntary wheel running (VWR) or cage control (SED) for 10 wk. To induce sepsis, we injected (2 mg/kg ip) LPS or sterile saline (SAL), and liver was harvested 6 or 12 h later. VWR attenuated increases in body and epididymal adipose tissue mass, improved glucose tolerance, and increased liver protein content of PEPCK (P < 0.05). VWR attenuated increases in LPS-induced IL-6 signaling and mRNA expression of other inflammatory markers (TNF-α, chemokine C-C motif ligand 2, inducible nitric oxide synthase, IL-10, IL-1ß) in the liver; however, this was not reflected at the whole body level, as systemic markers of inflammation were similar between SED and VWR. Insulin tolerance was greater in VWR compared with SED at 6 but not 12 h after LPS. The protective effect of VWR occurred in parallel with increases in the liver protein content of HSP70/72, a molecular chaperone that can protect against inflammatory challenges. This study provides novel evidence that physical activity protects against the inflammatory cascade induced by LPS in the liver and that these effects may be mediated via HSP70/72.

Structure-Activity Relationships of Dopamine Transporter Pharmacological Chaperones.

Mutations in the dopamine transporter gene (SLC6A3) have been implicated in many human diseases. Among these is the infantile parkinsonism-dystonia known as Dopamine Transporter Deficiency Syndrome (DTDS). Afflicted individuals have minimal to no functional dopamine transporter protein. This is primarily due to retention of misfolded disease-causing dopamine transporter variants. This results in a variety of severe motor symptoms in patients and the disease ultimately leads to death in adolescence or young adulthood. Though no treatment is currently available, pharmacological chaperones targeting the dopamine transporter have been shown to rescue select DTDS disease-causing variants. Previous work has identified two DAT pharmacological chaperones with moderate potency and efficacy: bupropion and ibogaine. In this study, we carried out structure-activity relationships (SARs) for bupropion and ibogaine with the goal of identifying the chemical features required for pharmacological chaperone activity. Our results show that the isoquinuclidine substituent of ibogaine and its analogs is an important feature for pharmacological chaperone efficacy. For bupropion, the secondary amine group is essential for pharmacological chaperone activity. Lastly, we describe additional ibogaine and bupropion analogs with varying chemical modifications and variable pharmacological chaperone efficacies at the dopamine transporter. Our results contribute to the design and refinement of future dopamine transporter pharmacological chaperones with improved efficacies and potencies.

The IDEAL household energy dataset, electricity, gas, contextual sensor data and survey data for 255 UK homes.

The IDEAL household energy dataset described here comprises electricity, gas and contextual data from 255 UK homes over a 23-month period ending in June 2018, with a mean participation duration of 286 days. Sensors gathered 1-second electricity data, pulse-level gas data, 12-second temperature, humidity and light data for each room, and 12-second temperature data from boiler pipes for central heating and hot water. 39 homes also included plug-level monitoring of selected electrical appliances, real-power measurement of mains electricity and key sub-circuits, and more detailed temperature monitoring of gas- and heat-using equipment, including radiators and taps. Survey data included occupant demographics, values, attitudes and self-reported energy awareness, household income, energy tariffs, and building, room and appliance characteristics. Linked secondary data comprises weather and level of urbanisation. The data is provided in comma-separated format with a custom-built API to facilitate usage, and has been cleaned and documented. The data has a wide range of applications, including investigating energy demand patterns and drivers, modelling building performance, and undertaking Non-Intrusive Load Monitoring research.

Technical and nidus-specific factors associated with adequacy of intraprocedural biopsy samples preceding radiofrequency ablation of osteoid osteoma.

PURPOSE: To examine the diagnostic yield of intraprocedural percutaneous biopsy performed at the time of radiofrequency ablation of suspected Osteoid Osteoma (OO) and identify technical and nidus-specific factors associated with diagnostic adequacy. MATERIALS AND METHODS: Following IRB approval, a total of 42 patients (male: 28, female: 14; mean age: 29 years) who underwent intraprocedural biopsy immediately prior to RFA between June 2010 and June 2017 were retrospectively identified. The nidi were located in various locations. The nidi had a mean size of 6.3 mm (range: 3-11 mm, Standard deviation (SD): 2.26). Core biopsies were performed by one of 15 operators. Biopsies were performed with two needle types ranging from 11-G to 15-G with a mean number of samples of 1.8 (range: 1-5, SD: 1.01). Electronic records and imaging were reviewed for demographics, nidus characteristics, biopsy details and diagnostic yield. Multivariate logistic regression of nidus-specific and biopsy-specific factors was performed. RESULTS: A total of 22/42 (52.3%) of the biopsies were adequate for histological diagnosis of OO. For the two experienced operators, the diagnostic yield was 67% (6/9) and 80% (8/10). Biopsy adequacy was significantly correlated with presence of an osteoid matrix (p = 0.03), obtaining >1 core sample (p = 0.03), the needle track passing through the nidus (p = 0.0003) and thinner (2.5 mm) intraprocedural CT slices (p = 0.03). On multivariate analysis, use of thinner intraprocedural CT slices was found to be associated with adequate biopsy (p = 0.02). CONCLUSION: Intraprocedural percutaneous biopsy samples of nidi highly-suspected to be OO at the time of RFA were diagnostic in 52% of patients. Multivariate analysis shows thinner intraprocedural CT slices to be a significantly associated with biopsy adequacy.

Analysis of the Chemotherapy-Free Interval following Image-Guided Ablation in Sarcoma Patients.

One way to enhance quality of life for patients with metastatic sarcoma is to maximize time off chemotherapy-a chemotherapy-free interval. While image-guided ablation of sarcoma metastases may reduce the need for chemotherapy, it remains unknown how long ablation could extend the chemotherapy-free interval. The purpose of our study was to determine the chemotherapy-free interval in comparison to overall survival and progression-free survival in sarcoma patients who undergo ablation procedures. An IRB-approved, single institution, HIPAA compliant database was queried for sarcoma patients who underwent image-guided ablation procedures between 2007 and 2018. Patient demographics, histologic subtype, and other clinical characteristics were recorded. Kaplan-Meier analysis was performed to compute median overall survival, median progression-free survival (local and distant), and the median chemotherapy-free interval (systemic and cytotoxic) after ablation. Univariate and multivariate analyses were performed using the log-rank test and Cox proportional-hazards model, respectively. A total of 100 sarcoma patients were included in the analysis. The most common histologic subtype was leiomyosarcoma (38%). Median overall survival after ablation of sarcoma metastases was 52.4 months (95% CI: 46.9-64.0 months). The median systemic chemotherapy-free interval following ablation of sarcoma metastases was 14.7 months (95% CI: 8.6-34.3 months). The median cytotoxic chemotherapy-free interval following ablation of sarcoma metastases was 81.3 months (95% CI: 34.3-median not reached). In conclusion, ablation of sarcoma metastases can provide an extended systemic chemotherapy-free interval of greater than 1 year. Ablation of sarcoma metastases may improve patient quality of life by extending the chemotherapy-free interval.

Regulation of Hepatic Follistatin Expression at Rest and during Exercise in Mice.

INTRODUCTION: Follistatin (FST) is a protein with numerous biological roles and was recently identified as an exercise-inducible hepatokine; however, the signals that regulate this are not well understood. The purpose of this study was to delineate potential endocrine factors that may regulate hepatic FST at rest and during exercise. METHODS: This study used four experiments. First, male and female C57BL/6J mice remained sedentary or were subjected to a single bout of exercise at moderate or exhaustive intensity with liver collected immediately post. Second, mice were injected with glucagon (1 mg·kg, 60 min), epinephrine (2 mg·kg, 30 min), glucagon then epinephrine, or saline. Third, mice were pretreated with propranolol (20-60 mg·kg, 30 min) before epinephrine injection. Fourth, glucagon receptor wild type (Gcgr) or knockout (Gcgr) mice were pretreated with saline or propranolol (20 mg·kg, 30 min) and were subjected to a single bout of exhaustive exercise with liver collected immediately post or after 2 h recovery. In all experiments liver FST mRNA expression was measured, and in experiment four FST protein content was measured. RESULTS: A single bout of treadmill exercise performed at an exhaustive but not moderate-intensity increased FST expression, as did injection of glucagon or epinephrine alone and when combined. Pretreatment of mice with propranolol attenuated the epinephrine-induced increase in FST expression. The exercise-induced increase in FST expression was attenuated in Gcgr mice, with no effect of propranolol. Gcgr mice had higher protein content of FST, but there was no effect of exercise or propranolol. CONCLUSIONS: These data suggest that both glucagon and epinephrine regulate hepatic FST expression at rest; however, only glucagon is required for the exercise-induced increase.

Glucagon receptor knockout mice are protected against acute olanzapine-induced hyperglycemia.

OBJECTIVES: To determine if glucagon is involved in mediating the increase in blood glucose levels caused by the second-generation antipsychotic drug olanzapine. MATERIALS AND METHODS: Whole body glucagon receptor deficient mice (Gcgr-/-) or WT littermate controls were injected with olanzapine (5mg/kg BW IP) and changes in blood glucose measured over the following 120min. Separate cohorts of mice were treated with olanzapine and changes in pyruvate tolerance, insulin tolerance and whole body substrate oxidation were determined. RESULTS: Olanzapine treatment increased serum glucagon and lead to rapid increases in blood glucose concentrations in WT mice. Gcgr-/- mice were protected against olanzapine-induced increases in blood glucose but this was not explained by differences in terminal serum insulin concentrations, enhanced AKT phosphorylation in skeletal muscle, adipose tissue or liver or differences in RER. In both genotypes olanzapine induced an equivalent degree of insulin resistance as measured using an insulin tolerance test. Olanzapine treatment led to an exaggerated glucose response to a pyruvate challenge in WT but not Gcgr-/- mice and this was paralleled by reductions in the protein content of PEPCK and G6Pase in livers from Gcgr-/- mice. CONCLUSIONS: Gcgr-/- mice are protected against olanzapine-induced increases in blood glucose. This is likely a result of reductions in liver glucose output, perhaps secondary to decreases in PEPCK and G6Pase protein content. Our findings highlight the central role of the liver in mediating olanzapine-induced disturbances in glucose homeostasis.

Imaging Neurons within Thick Brain Sections Using the Golgi-Cox Method.

The Golgi-Cox method of neuron staining has been employed for more than two hundred years to advance our understanding of neuron morphology within histological brain samples. While it is preferable from a practical perspective to prepare brain sections at the greatest thickness possible, in order to increase the probability of identifying stained neurons that are fully contained within single sections, this approach is limited from a technical perspective by the working distance of high-magnification microscope objectives. We report here a protocol to stain neurons using the Golgi-Cox method in mouse brain sections that are cut at 500 µm thickness, and to visualize neurons throughout the depth of these sections using an upright microscope fitted with a high-resolution 30X 1.05 N.A. silicone oil-immersion objective that has an 800 µm working distance. We also report two useful variants of this protocol that may be employed to counterstain the surface of mounted brain sections with the cresyl violet Nissl stain, or to freeze whole brains for long-term storage prior to sectioning and final processing. The main protocol and its two variants produce stained thick brain sections, throughout which full neuron dendritic trees and dendrite spines may be reliably visualized and quantified.