Unique pharmacodynamic properties and low abuse liability of the µ-opioid receptor ligand (S)-methadone.

(R,S)-methadone ((R,S)-MTD) is a µ-opioid receptor (MOR) agonist comprised of (R)-MTD and (S)-MTD enantiomers. (S)-MTD is being developed as an antidepressant and is considered an N-methyl-D-aspartate receptor (NMDAR) antagonist. We compared the pharmacology of (R)-MTD and (S)-MTD and found they bind to MORs, but not NMDARs, and induce full analgesia. Unlike (R)-MTD, (S)-MTD was a weak reinforcer that failed to affect extracellular dopamine or induce locomotor stimulation. Furthermore, (S)-MTD antagonized motor and dopamine releasing effects of (R)-MTD. (S)-MTD acted as a partial agonist at MOR, with complete loss of efficacy at the MOR-galanin Gal1 receptor (Gal1R) heteromer, a key mediator of the dopaminergic effects of opioids. In sum, we report novel and unique pharmacodynamic properties of (S)-MTD that are relevant to its potential mechanism of action and therapeutic use. One-sentence summary: (S)-MTD, like (R)-MTD, binds to and activates MORs in vitro, but (S)-MTD antagonizes the MOR-Gal1R heteromer, decreasing its abuse liability.

Identifying key risks to the performance of privately protected areas (PPAs) through theory of change (ToC).

Privately protected areas (PPAs) are internationally considered to be important policy implementation instruments to augment and strengthen protected area networks. However, there has been limited reflection on the performance of PPAs over time. This paper aims to identify key risks to the performance of PPAs as policy implementation instruments through the application of Theory of Change (ToC). Identifying and understanding these risks are important to allow for the evaluation and monitoring of PPA performance. The ToC method was applied to a specific PPA policy instrument namely, private nature reserves (PNRs) in the South African context. The research results produced 29 key assumptions translated into 29 key risks. These risk are critically discussed against existing South African and international literature. To test and refine the risks further it is recommended that they be applied to PPA case studies in different contexts.

Multiple Sclerosis Performance Test: validation of self-administered neuroperformance modules.

BACKGROUND AND PURPOSE: The purpose was to determine the test-retest reliability, practice effects, convergent validity and sensitivity to multiple sclerosis (MS) disability of neuroperformance subtests from the patient self-administered Multiple Sclerosis Performance Test (MSPT) designed to assess low contrast vision (Contrast Sensitivity Test, CST), upper extremity motor function (Manual Dexterity Test, MDT) and lower extremity motor function (Walking Speed Test, WST) and to introduce the concept of regression-based norms to aid clinical interpretation of performance scores using the MSPT cognition test (Processing Speed Test, PST) as an example. METHODS: Substudy 1 assessed test-retest reliability, practice effects and convergent validity of the CST, MDT and WST in 30 MS patients and 30 healthy controls. Substudy 2 examined sensitivity to MS disability in over 600 MS patients as part of their routine clinic assessment. Substudy 3 compared performance on the PST in research volunteers and clinical samples. RESULTS: The CST, MDT and WST were shown to be reliable, valid and sensitive to MS outcomes. Performance was comparable to technician-administered testing. PST performance was poorer in the clinical sample compared with the research volunteer sample. CONCLUSIONS: The self-administered MSPT neuroperformance modules produce reliable, objective metrics that can be used in clinical practice and support outcomes research. Published studies which require patient voluntary consent may underestimate the rate of cognitive dysfunction observed in a clinical setting.

Molecular stress-inducing compounds increase osteoclast formation in a heat shock factor 1 protein-dependent manner.

Many anticancer therapeutic agents cause bone loss, which increases the risk of fractures that severely reduce quality of life. Thus, in drug development, it is critical to identify and understand such effects. Anticancer therapeutic and HSP90 inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) causes bone loss by increasing osteoclast formation, but the mechanism underlying this is not understood. 17-AAG activates heat shock factor 1 (Hsf1), the master transcriptional regulator of heat shock/cell stress responses, which may be involved in this negative action of 17-AAG upon bone. Using mouse bone marrow and RAW264.7 osteoclast differentiation models we found that HSP90 inhibitors that induced a heat shock response also enhanced osteoclast formation, whereas HSP90 inhibitors that did not (including coumermycin A1 and novobiocin) did not affect osteoclast formation. Pharmacological inhibition or shRNAmir knockdown of Hsf1 in RAW264.7 cells as well as the use of Hsf1 null mouse bone marrow cells demonstrated that 17-AAG-enhanced osteoclast formation was Hsf1-dependent. Moreover, ectopic overexpression of Hsf1 enhanced 17-AAG effects upon osteoclast formation. Consistent with these findings, protein levels of the essential osteoclast transcription factor microphthalmia-associated transcription factor were increased by 17-AAG in an Hsf1-dependent manner. In addition to HSP90 inhibitors, we also identified that other agents that induced cellular stress, such as ethanol, doxorubicin, and methotrexate, also directly increased osteoclast formation, potentially in an Hsf1-dependent manner. These results, therefore, indicate that cellular stress can enhance osteoclast differentiation via Hsf1-dependent mechanisms and may significantly contribute to pathological and therapeutic related bone loss.

Context-dependent role of Grb7 in HER2+ve and triple-negative breast cancer cell lines.

Grb7 is an adapter protein, aberrantly co-overexpressed with HER2 and identified as an independent prognostic marker in breast cancer. It has been established that Grb7 exacerbates the cellular growth and migratory behaviour of HER2+ve breast cancer cells. Less is known about Grb7's role in the context of HER2-ve cells. Here we directly compare the effect of stable Grb7 knockdown in oestrogen sensitive (T47D), HER2+ve (SKBR3) and triple-negative (MDA-MB-468 and MDA-MB-231) breast cancer cell lines on anchorage dependent and independent cell growth, wound healing and chemotaxis. All cell lines showed reduced ability to migrate upon Grb7 knockdown, despite their greatly varied endogenous levels of Grb7. Decreased cell proliferation was not observed in any of the cell lines upon Grb7 knockdown; however, decreased ability to form colonies was observed for all but the oestrogen sensitive cell line, depending upon the stringency of the growth conditions. The data reveal that Grb7 plays an important role in breast cancer progression, beyond the context of HER2+ve cell types.

Sex dependence of opioid-mediated responses to subanesthetic ketamine in rats.

Subanesthetic ketamine is increasingly used for the treatment of varied psychiatric conditions, both on- and off-label. While it is commonly classified as an N-methyl D-aspartate receptor (NMDAR) antagonist, our picture of ketamine's mechanistic underpinnings is incomplete. Recent clinical evidence has indicated, controversially, that a component of the efficacy of subanesthetic ketamine may be opioid dependent. Using pharmacological functional ultrasound imaging in rats, we found that blocking opioid receptors suppressed neurophysiologic changes evoked by ketamine, but not by a more selective NMDAR antagonist, in limbic regions implicated in the pathophysiology of depression and in reward processing. Importantly, this opioid-dependent response was strongly sex-dependent, as it was not evident in female subjects and was fully reversed by surgical removal of the male gonads. We observed similar sex-dependent effects of opioid blockade affecting ketamine-evoked postsynaptic density and behavioral sensitization, as well as in opioid blockade-induced changes in opioid receptor density. Together, these results underscore the potential for ketamine to induce its affective responses via opioid signaling, and indicate that this opioid dependence may be strongly influenced by subject sex. These factors should be more directly assessed in future clinical trials.

Mu Opioid Receptor Activation Mediates (S)-ketamine Reinforcement in Rats: Implications for Abuse Liability.

BACKGROUND: (S)-ketamine is an NMDA receptor antagonist, but it also binds to and activates mu opioid receptors (MORs) and kappa opioid receptors in vitro. However, the extent to which these receptors contribute to (S)-ketamine's in vivo pharmacology is unknown. METHODS: We investigated the extent to which (S)-ketamine interacts with opioid receptors in rats by combining in vitro and in vivo pharmacological approaches, in vivo molecular and functional imaging, and behavioral procedures relevant to human abuse liability. RESULTS: We found that the preferential opioid receptor antagonist naltrexone decreased (S)-ketamine self-administration and (S)-ketamine-induced activation of the nucleus accumbens, a key brain reward region. A single reinforcing dose of (S)-ketamine occupied brain MORs in vivo, and repeated doses decreased MOR density and activity and decreased heroin reinforcement without producing changes in NMDA receptor or kappa opioid receptor density. CONCLUSIONS: These results suggest that (S)-ketamine's abuse liability in humans is mediated in part by brain MORs.

6-O-(2-[18F]Fluoroethyl)-6-O-Desmethyl-Diprenorphine ([18F]FE-DPN) Preferentially Binds to Mu Opioid Receptors In Vivo.

PURPOSE: 6-O-(2-[18F]Fluoroethyl)-6-O-desmethyl-diprenorphine ([18F]FE-DPN) is regarded as a non-selective opioid receptor radiotracer. PROCEDURE: Here, we report the first characterization of [18F]FE-DPN synthesized from the novel precursor, 6-O-(2-tosyloxyethoxy)-6-O-desmethyl-3-O-trityl-diprenorphine (TE-TDDPN), using a one-pot, two-step nucleophilic radiosynthesis to image opioid receptors in rats and mice using positron emission tomography. RESULTS: We also show that [18F]FE-DPN and [3H]DPN exhibit negligible brain uptake in mu opioid receptor (MOR) knockout mice. CONCLUSIONS: Taken together with prior findings, our results suggest that [18F]FE-DPN and [3H]DPN preferentially bind to MOR in rodents in vivo.

Benzopyrazine derivatives: A novel class of growth factor receptor bound protein 7 antagonists.

Growth factor receptor bound protein 7 (Grb7) is an adapter protein that functions as a downstream effector of growth factor mediated signal transduction. Over-expression of Grb7 has been implicated in a variety of cancers such as breast, blood, pancreatic, esophageal, and gastric carcinomas. Inhibition of Grb7 has been shown to reduce the migratory and proliferative potential of these cancers, making it an attractive therapeutic target. Starting with a known peptide antagonist, the present work reports the application of a succession of computational ligand design tools comprising a ligand shape based similarity search, molecular docking and a 2D-similarity search to identify small molecular antagonists of the Grb7-SH2 domain from the NCI chemical database. Binding to the Grb7-SH2 domain was then experimentally tested using melting point shift assays and isothermal titration calorimetry. Overall, a total of 11 benzopyrazine based small molecular antagonists were identified with affinity for the Grb7-SH2 domain. Representative compounds tested using ITC were revealed to possess moderate binding affinity in the low micromolar range. Finally, the lead compound (NSC642056) was found to reduce the growth of a Grb7-expressing breast cancer cell line with an IC(50) of 86µM. It is expected that the identified antagonists will be useful additions to further explore the function of Grb7 and for the development of inhibitors with therapeutic potential.

Nb3Sn multicell cavity coating system at Jefferson Lab.

Superconducting radio frequency niobium cavities are the building blocks of modern accelerators for scientific applications. Lower surface resistance, higher fields, and high operating temperatures advance the reach of the future accelerators for scientific discovery as well as potentially enabling cost-effective industrial solutions. We describe the design and performance of an Nb3Sn coating system that converts the inner surface of niobium cavities to an Nb3Sn film. The niobium surface, heated by radiation from the niobium retort, is exposed to Sn and SnCl2 vapor during the heat cycle, which results in about 2 µm Nb3Sn film on the niobium surface. Film composition and structure as well as radio frequency properties with 1-cell R&D cavities and 5-cell practical accelerator cavities are presented.

Comparison of fibrin glue with sutures for pterygium excision surgery with conjunctival autografts.

PURPOSE: To compare pterygium excision conjunctival autograft surgery using Tisseel fibrin glue versus Vicryl sutures. METHOD: Prospective randomized clinical trial. Fifty patients were randomized into either sutured graft or glued graft groups. Twenty-five received standard conjunctival autograft sutured with Vicryl and 25 received surgery with the autograft placed with Tisseel fibrin glue. Outcome measures include surgical time, patient discomfort, pathology, complications and recurrence rates at 3, 6 and 12 months. Patients were followed up at days 1, 7, 14, 30, 90, 180 and 365. RESULTS: The mean surgical time for the glue group was significantly shorter at 12 min compared with the suture group at 26 min (P < 0.001). Postoperative pain was significantly less at day 1 (P < 0.001) and day 2 (P < 0.05) but was not significantly different following these visits. Complications in the glue group included one patient with an absent graft at week 1 that required revision of the graft. At 3 months there were no recurrences in the glue group and two recurrences in the suture group. The 6- and 12-month recurrence rate was unchanged and not significantly different between the glue and suture groups (both 0/24 and 2/23, respectively). CONCLUSION: Both glued and sutured conjunctival autografting procedures are safe and effective methods for pterygium surgery. The glued autograft recurrence rate at 12 months was similar to that of sutured grafts. Conjunctival autograft with fibrin glue in pterygium surgery decreased surgical time and resulted in less postoperative pain in the first 48 hours but had a higher complication rate.

Juvenile multiple sclerosis-like episodes associated with a defect of mitochondrial beta oxidation.

We describe a young girl who presented with recurrent episodes of central nervous system (CNS) demyelination mimicking multiple sclerosis. Metabolic evaluations and decreased oxidation of [9,10(n)-3H] palmitate demonstrated defective mitochondrial beta oxidation, but complementation studies of the patient's cells, fused with cell lines with known defects of beta oxidation, failed to identify a known disorder. While progressive CNS demyelination has occurred in patients with defective peroxisomal very long-chain fatty acid oxidation, this is the 1st time it has occurred with defective mitochondrial beta oxidation. This patient appears to represent a novel disorder of beta oxidation producing intermittent demyelination with profound CNS symptoms. Recognition of the defect led to appropriate therapy, which caused marked clinical improvement.

Bremsstrahlung imaging using the gamma camera: factors affecting attenuation.

Quantitative imaging of bremsstrahlung from pure beta emitters is proposed as a means for in vivo management of antibody therapy. The method involves the use of high-energy collimation, an empirically selected broad photon energy window to enhance detector sensitivity, and a Wiener restoration filter to compensate for system blur. The measured and filtered data were obtained for an idealized scattering medium and isolated spherical sources. An effective linear attenuation coefficient of about 0.13 cm-1 was determined from the raw image data of 32P. A coefficient of 0.14 cm-1 was determined after the images were restored using the Wiener filter. The measured attenuation was not significantly dependent on the size of the region of interest or the size of the source. Its variation was within the experimental error of measurement (+/- 5%). The measured sensitivity (6 x 10(-6) cps/Bq) was sufficient for imaging therapy doses of 32P or 90Y.

Aerobic versus strength training for risk factor intervention in middle-aged men at high risk for coronary heart disease.

To compare the effects of strength training (ST) to those of aerobic training (AT) for coronary heart disease (CHD) risk factor intervention, we studied 37 previously untrained males (aged 50 +/- 9 years, mean +/- SD) before and after 20 weeks of either ST (N = 14), AT (walk/jog, N = 13), or no exercise (inactive controls, N = 10). Lipoprotein and lipid profiles, blood pressure, and glucose and insulin responses to an oral glucose tolerance test (OGTT) were assessed before and after the training period in all three groups. The ST program produced significant reductions in plasma glucose levels at 60, 90, and 120 minutes (P < .05) after glucose ingestion, whereas the AT program resulted in significant reductions only at 90 and 120 minutes (P < .05). ST also decreased insulin levels during fasting (P < .05) and at 90 and 120 minutes (P < .01) after glucose ingestion. AT decreased insulin levels at 90 and 120 minutes (P < .01) after glucose ingestion. Both training programs reduced the total area under the glucose tolerance curve for glucose (both P < .05) and insulin (both P < .05), but there were no significant differences in these changes between the two groups. None of the glucose or insulin values were significantly altered in the control group. There were no significant changes in lipoprotein and lipid profiles or blood pressure in any of the three groups. These results suggest that ST and AT have comparable effects on risk factors for CHD.(ABSTRACT TRUNCATED AT 250 WORDS)

Phleborheography of the upper extremity.

Phleborheography is a well-established technique for the noninvasive diagnosis of deep venous thrombosis of the lower extremity. For ten years we have used phleborheography in the diagnosis of axillary and subclavian venous thrombosis. We developed modifications in technique and interpretation that are necessary for accurate application of phleborheography to the upper extremity. When compared with venography, the sensitivity and specificity of this technique approached 90%. Therefore, phleborheography appears to be a useful screening examination for deep venous thrombosis of the upper extremity.

Inhibitory effect of lichen constituents on mutagenicity induced by heterocyclic amines.

Physodic acid, one of the main constituents of Hypogymnia enteromorpha, inhibited the mutagenicity of indirect mutagens, including benzo[a]pyrene and heterocyclic amines in Salmonella typhimurium TA 98. In contrast, it was not effective against direct mutagens such as 6-nitropiperonal and adriamycin. Its antimutagenicity was not associated with free-radical scavenging or antioxidative activities. Physodic acid seemed to inhibit the formation of reactive metabolites, such as N-hydroxy-Trp-P-2, by blocking the hepatic microsomal oxidation systems. Another component of H. enteromorpha, physodalic acid, also inhibited mutagenicity of a heterocyclic amine, Trp-P-2, in S. typhimurium TA 98, even though it was reportedly mutagenic in S. typhimurium TA 100.

Effect of butyl benzyl phthalate on reproduction and zinc metabolism.

Butyl benzyl phthalate (BBP) has been shown to be teratogenic. One mechanism contributing to the teratogenicity of several developmental toxicants, is chemical-induced changes in maternal zinc (Zn) metabolism which result in an increased synthesis of maternal liver metallothionein (Mt), and a subsequent reduction in Zn delivery to the conceptus. We investigated the effects of maternal BBP exposure on maternal-fetal Zn metabolism in Wistar rats. In study I, dams were gavaged with BBP (0,250,1000,1500 or 2000 mg/kg) on gestation days (GD) 11 through 13, and killed on GD 20. Maternal toxicity was evident in the three highest dose groups. Embryo/fetal death and small pup weights and lengths were noted in the 2000 mg BBP/kg group. Fetuses in the 1500 and 2000 mg/kg groups were characterized by poor skeletal ossification, and a high frequency of cleft palate. Rib anomalies were observed in the three highest dose groups. Maternal liver Mt concentrations were only slightly elevated in the 1500 and 2000 mg/kg groups. In study II, dams treated as above, were gavaged with 65Zn and killed 18 h later. While the 2000 mg/kg group had high percentages of 65Zn in some maternal tissues, sequestration of 65Zn in maternal liver was not evident. Thus, BBP is not a strong inducer of Mt, and the teratogenicity of BBP does not appear to be due to alterations in maternal and/or embryonic Zn metabolism.

Strength training does not improve lipoprotein-lipid profiles in men at risk for CHD.

Sixteen untrained males, age 46 +/- 11 yr (mean +/- SD), were studied to determine the effects of 20 wk of strength training on lipoprotein-lipid profiles and post-heparin lipase activities. All subjects had abnormal lipoprotein-lipid profiles and at least two other risk factors for coronary heart disease (CHD). To control for day-to-day variations in blood lipoprotein levels, baseline values were established by taking at least two blood samples on separate days from the training and control groups. The training program resulted in a 50% increase in upper body strength (P less than 0.001) and a 37% increase in lower body strength (P less than 0.001) as measured by the one repetition maximum test (1-RM). No changes in the 1-RM test were observed in the control group. There were no significant changes in VO2max (34.5 +/- 6.4 vs 36.2 +/- 7.4 ml.kg-1.min-1 or percent fat (25.4 +/- 4.2 vs 24.9 +/- 4.1%) with training. There were also no significant changes in plasma concentrations of triglyceride (193 +/- 96 vs 171 +/- 101 mg.dl-1), total cholesterol (231 +/- 22 vs 210 +/- 22 mg.dl-1), and HDL-C (35 +/- 6 vs 36 +/- 8 mg.dl-1), or LDL-C (139 +/- 16 vs 139 +/- 21 mg.dl-1). Furthermore, the activities of post-heparin lipoprotein lipase (9 +/- 4 vs 13 +/- 5 moles.ml-1.h-1) and hepatic lipase (35 +/- 10 vs 35 +/- 9 moles.ml-1.h-1) did not change with training.(ABSTRACT TRUNCATED AT 250 WORDS)

A case of glutaric acidemia type II (severe multiple acyl-CoA dehydrogenation disorder) with subsequent prenatal exclusion in a sibling.

A case of severe multiple acyl-CoA dehydrogenation disorder is described. This is the second such case reported to have had an elevated maternal serum alpha-fetoprotein and normal amniotic alpha-fetoprotein. The child's 3-day extrauterine life was characterized by intractable acidosis, respiratory distress, and ventricular fibrillation. The characteristic biochemical, morphologic, and microscopic findings of this condition are reviewed. A subsequent pregnancy was evaluated using chorionic villus sampling and analysis of cultured trophoblasts. The trophoblasts were biochemically normal, and a normal child was subsequently delivered. Since the manifestations of this disorder developed in utero, prenatal diagnosis and therapy offer the only hope for a more prolonged survival.

Evaluating the effects of elevated levels of atmospheric trace gases on herbs and shrubs: a prototype dual array field exposure system.

In the context of global climate change, an understanding of the long-term effects of increasing concentrations of atmospheric trace gases (carbon dioxide, CO(2), ozone, O(3), oxides of nitrogen, NO(x) etc.) on both cultivated and native vegetation is of utmost importance. Over the years, under field conditions, various trace gas-vegetation exposure methodologies with differing advantages and disadvantages have been used. Because of these variable criteria, with elevated O(3) or CO(2) levels, at the present time the approach of free-air experimental-release of the gas into study plots is attracting much attention. However, in the case of CO(2), this approach (using 15 m diameter study plot with a single circular array of vent pipes) has proven to be cost prohibitive (about 59000-98000 dollars/year/replicate) due to the consumption of significant quantities of the gas to perform the experiment (CO(2) level elevated to 400 ppm above the ambient). Therefore, in this paper, we present a new approach consisting of a dual, concentric exposure array of vertical risers or vent pipes. The purpose of the outer array (17 m diameter) is to vent ambient air outward and toward the incoming wind, thus providing an air curtain to reduce the velocity of that incoming wind to simulate the mode or the most frequently occurring wind speed at the study site. The inner array (15 m diameter) vents the required elevated levels of trace gases (CO(2), O(3), etc.) into the study plot. This dual array system is designed to provide spatial homogeneity (shown through diffusion modeling) of the desired trace-gas levels within the study plot and to also reduce its consumption. As an example, while in the single-array free-air CO(2)-release system the consumption of CO(2) to elevate its ambient concentration by 400 ppm is calculated to be about 980 tons/year/replicate, it is estimated that in the dual array system it would be approximately 590 tons/year/replicate. Thus, the dual array system may provide substantial cost savings (24000-39000 dollars/year/replicate) in the CO(2) consumption (60-100 dollars/ton of CO(2)) alone. Similarly, benefits in the requirements of other trace gases (O(3), NO(x), etc.) are expected, in future multivariate studies on global climate change.