Microcrystalline identification of selected designer drugs

A microcrystalline test for the detection of 4-methylmethcathinone (mephedrone), benzylpiperazine (BZP) and 5,6-methylenedioxy-2-aminoindane (MDAI) using aqueous solutions of mercury chloride is described. Each of the compounds investigated formed specific drug–reagent crystals within minutes. The uniqueness of the test was confirmed by comparison of the microcrystalline response to that of other psychoactive stimulants and a common cutting agent. The limit of detection and cut-off levels for reference standards were established to 3 g/L and 5 g/L for mephedrone, 0.5 g/L for MDAI and 0.2 g/L and 0.3 g/L for BZP, respectively. Various mixtures of standards of either mephedrone, BZP or MDAI combined with caffeine were investigated for their microcrystalline response. Results showed that simultaneous detection of drug and cutting agent was possible with the concentrations tested but were dependant on the ratio of drug to cutting agent. BZP could be detected alongside caffeine from as low as 20% (v/v), MDAI from 40% (v/v) and mephedrone from 50% (v/v) and higher. Finally, seven samples of online purchased ‘legal highs’ were analysed using the developed test and the findings were compared to FTIR and GC–MS results. It was shown that 6 out of 7 samples did not contain the advertised active ingredient. Five samples consisted of BZP, caffeine and 1-[3-(trifluoromethyl)phenyl]piperazine (3-TFMPP). The microcrystalline tests carried out on these samples showed positive results for both BZP and caffeine without interference from other substances present

Reversing microcrystalline tests: an analytical approach to recycling of microcrystals from drugs of abuse

A combined analysis of microcrystalline tests followed by LC-MS or GC-MS analysis is described. Microcrystalline tests are shown to be non-destructive as addition products formed were easily dissociated after the application of an appropriate solvent. Subsequent analysis of the sample was done to quantify the recovery of the drug. Examples were performed using the date rape drug γ-hydroxybutyrate (GHB) and the synthetic opioid methadone

Increasing Glutathione Levels by a Novel Posttranslational Mechanism Inhibits Neuronal Hyperexcitability

Glutathione (GSH) depletion, and impaired redox homeostasis have been observed in experimental animal models and patients with epilepsy. Pleiotropic strategies that elevate GSH levels via transcriptional regulation have been shown to significantly decrease oxidative stress and seizure frequency, increase seizure threshold, and rescue certain cognitive deficits. Whether elevation of GSH per se alters neuronal hyperexcitability remains unanswered. We previously showed that thiols such as dimercaprol (DMP) elevate GSH via post-translational activation of glutamate cysteine ligase (GCL), the rate limiting GSH biosynthetic enzyme. Here, we asked if elevation of cellular GSH by DMP altered neuronal hyperexcitability in-vitro and in-vivo. Treatment of primary neuronal-glial cerebrocortical cultures with DMP elevated GSH and inhibited a voltage-gated potassium channel blocker (4-aminopyridine, 4AP) induced neuronal hyperexcitability. DMP increased GSH in wildtype (WT) zebrafish larvae and significantly attenuated convulsant pentylenetetrazol (PTZ)-induced acute \u27seizure-like\u27 swim behavior. DMP treatment increased GSH and inhibited convulsive, spontaneous \u27seizure-like\u27 swim behavior in the Dravet Syndrome (DS) zebrafish larvae (scn1Lab). Furthermore, DMP treatment significantly decreased spontaneous electrographic seizures and associated seizure parameters in scn1Lab zebrafish larvae. We investigated the role of the redox-sensitive mammalian target of rapamycin (mTOR) pathway due to the presence of several cysteine-rich proteins and their involvement in regulating neuronal excitability. Treatment of primary neuronal-glial cerebrocortical cultures with 4AP or l-buthionine-(S,R)-sulfoximine (BSO), an irreversible inhibitor of GSH biosynthesis, significantly increased mTOR complex I (mTORC1) activity which was rescued by pre-treatment with DMP. Furthermore, BSO-mediated GSH depletion oxidatively modified the tuberous sclerosis protein complex (TSC) consisting of hamartin (TSC1), tuberin (TSC2), and TBC1 domain family member 7 (TBC1D7) which are critical negative regulators of mTORC1. In summary, our results suggest that DMP-mediated GSH elevation by a novel post-translational mechanism can inhibit neuronal hyperexcitability both in-vitro and in-vivo and a plausible link is the redox sensitive mTORC1 pathway

What determines women's participation in collective action? Evidence from a western Ugandan coffee cooperative

Women smallholders face greater constraints than men in accessing capital and commodity markets in Sub-Saharan Africa. Collective action has been promoted to remedy those disadvantages. Using survey data of 421 women members and 210 nonmembers of a coffee producer cooperative in Western Uganda, this study investigates the determinants of women's participation in cooperatives and women's intensity of participation. The results highlight the importance of access to and control over land for women to join the cooperative in the first place. Participation intensity is measured through women's participation in collective coffee marketing and share capital contributions. It is found that duration of membership, access to extension services, more equal intrahousehold power relations, and joint land ownership positively influence women's ability to commit to collective action. These findings demonstrate the embeddedness of collective action in gender relations and the positive value of women's active participation for agricultural-marketing cooperatives

Neuron-Specific Mitochondrial Oxidative Stress Results in Epilepsy, Glucose Dysregulation and a Striking Astrocyte Response

Mitochondrial superoxide (O2.−) production is implicated in aging, neurodegenerative disease, and most recently epilepsy. Yet the specific contribution of neuronal O2.− to these phenomena is unclear. Here, we selectively deleted superoxide dismutase-2 (SOD2) in neuronal basic helix-loop-helix transcription factor (NEX)-expressing cells restricting deletion to a subset of excitatory principle neurons primarily in the forebrain (cortex and hippocampus). This resulted in nSOD2 KO mice that lived into adulthood (2-3 months) with epilepsy, selective loss of neurons, metabolic rewiring and a marked mitohormetic gene response. Surprisingly, expression of an astrocytic gene, glial fibrillary acidic protein (GFAP) was significantly increased relative to WT. Further studies in rat primary neuron-glial cultures showed that increased mitochondrial O2.−, specifically in neurons, was sufficient to upregulate GFAP. These results suggest that neuron-specific mitochondrial O2.− is sufficient to drive a complex and catastrophic epileptic phenotype and highlights the ability of SOD2 to act in a cell-nonautonomous manner to influence an astrocytic response

Neurofibromin Deficiency Induces Endothelial Cell Proliferation and Retinal Neovascularization

Purpose: Neurofibromatosis type 1 (NF1) is the result of inherited mutations in the NF1 tumor suppressor gene, which encodes the protein neurofibromin. Eye manifestations are common in NF1 with recent reports describing a vascular dysplasia in the retina and choroid. Common features of NF1 retinopathy include tortuous and dilated feeder vessels that terminate in capillary tufts, increased endothelial permeability, and neovascularization. Given the retinal vascular phenotype observed in persons with NF1, we hypothesize that preserving neurofibromin may be a novel strategy to control pathologic retinal neovascularization. Methods: Nf1 expression in human endothelial cells (EC) was reduced using small hairpin (sh) RNA and EC proliferation, migration, and capacity to form vessel-like networks were assessed in response to VEGF and hypoxia. Wild-type (WT), Nf1 heterozygous (Nf1+/-), and Nf1flox/+;Tie2cre pups were subjected to hyperoxia/hypoxia using the oxygen-induced retinopathy model. Retinas were analyzed quantitatively for extent of retinal vessel dropout, neovascularization, and capillary branching. Results: Neurofibromin expression was suppressed in response to VEGF, which corresponded with activation of Mek-Erk and PI3-K-Akt signaling. Neurofibromin-deficient EC exhibited enhanced proliferation and network formation in response to VEGF and hypoxia via an Akt-dependent mechanism. In response to hyperoxia/hypoxia, Nf1+/- retinas exhibited increased vessel dropout and neovascularization when compared with WT retinas. Neovascularization was similar between Nf1+/- and Nf1flox/+;Tie2cre retinas, but capillary drop out in Nf1flox/+;Tie2cre retinas was significantly reduced when compared with Nf1+/- retinas. Conclusions: These data suggest that neurofibromin expression is essential for controlling endothelial cell proliferation and retinal neovascularization and therapies targeting neurofibromin-deficient EC may be beneficial

Geriatric Oncology as an Unmet Workforce Training Need in the United Kingdom—A Narrative Review by the British Oncology Network for Undergraduate Societies (BONUS) and the International Society of Geriatric Oncology (SIOG) UK Country Group

Cancer is a disease associated with ageing. Managing cancer in older adults may prove challenging owing to pre-existing frailty, comorbidity, and wider holistic needs, as well as the unclear benefits and harms of standard treatment options. With the ongoing advances in oncology and the increasing complexity of treating older adults with cancer, the geriatric oncology field must be a priority for healthcare systems in education, research, and clinical practice. However, geriatric oncology is currently not formally taught in undergraduate education or postgraduate training programmes in the United Kingdom (UK). In this commentary, we outline the landscape of geriatric oncology undergraduate education and postgraduate training for UK doctors. We highlight current challenges and opportunities and provide practical recommendations for better preparing the medical workforce to meet the needs of the growing population of older adults with cancer. This includes key outcomes to be considered for inclusion within undergraduate and postgraduate curricula

Neuronal SIRT3 Deletion Predisposes to Female-Specific Alterations in Cellular Metabolism, Memory, and Network Excitability

Mitochondrial dysfunction is an early event in the pathogenesis of neurologic disorders and aging. Sirtuin 3 (SIRT3) regulates mitochondrial function in response to the cellular environment through the reversible deacetylation of proteins involved in metabolism and reactive oxygen species detoxification. As the primary mitochondrial deacetylase, germline, or peripheral tissue-specific deletion of SIRT3 produces mitochondrial hyperacetylation and the accelerated development of age-related diseases. Given the unique metabolic demands of neurons, the role of SIRT3 in the brain is only beginning to emerge. Using mass spectrometry-based acetylomics, high-resolution respirometry, video-EEG, and cognition testing, we report targeted deletion of SIRT3 from select neurons in the cortex and hippocampus produces altered neuronal excitability and metabolic dysfunction in female mice. Targeted deletion of SIRT3 from neuronal helix-loop-helix 1 (NEX)-expressing neurons resulted in mitochondrial hyperacetylation, female-specific superoxide dismutase-2 (SOD2) modification, increased steady-state superoxide levels, metabolic reprogramming, altered neuronal excitability, and working spatial memory deficits. Inducible neuronal deletion of SIRT3 likewise produced female-specific deficits in spatial working memory. Together, the data demonstrate that deletion of SIRT3 from forebrain neurons selectively predisposes female mice to deficits in mitochondrial and cognitive function. SIGNIFICANCE STATEMENT Mitochondrial SIRT3 is an enzyme shown to regulate energy metabolism and antioxidant function, by direct deacetylation of proteins. In this study, we show that neuronal SIRT3 deficiency renders female mice selectively vulnerable to impairment in redox and metabolic function, spatial memory, and neuronal excitability. The observed sex-specific effects on cognition and neuronal excitability in female SIRT3-deficient mice suggest that mitochondrial dysfunction may be one factor underlying comorbid neuronal diseases, such as Alzheimer\u27s disease and epilepsy. Furthermore, the data suggest that SIRT3 dysfunction may predispose females to age-related metabolic and cognitive impairment

Large Eccentricity, Low Mutual Inclination: The Three-dimensional Architecture of a Hierarchical System of Giant Planets

We establish the three-dimensional architecture of the Kepler-419 (previously KOI-1474) system to be eccentric yet with a low mutual inclination. Kepler-419b is a warm Jupiter at semi-major axis ɑ = 0.370^(+0.007)_(-0.006) AU with a large eccentricity (e = 0.85^(+0.08)_(-0.07)) measured via the "photoeccentric effect." It exhibits transit timing variations (TTVs) induced by the non-transiting Kepler-419c, which we uniquely constrain to be a moderately eccentric (e = 0.184 ± 0.002), hierarchically separated (a = 1.68 ± 0.03 AU) giant planet (7.3 ± 0.4 M J_(up)). We combine 16 quarters of Kepler photometry, radial-velocity (RV) measurements from the HIgh Resolution Echelle Spectrometer on Keck, and improved stellar parameters that we derive from spectroscopy and asteroseismology. From the RVs, we measure the mass of the inner planet to be 2.5 ± 0.3 M J_(up) and confirm its photometrically measured eccentricity, refining the value to e = 0.83 ± 0.01. The RV acceleration is consistent with the properties of the outer planet derived from TTVs. We find that despite their sizable eccentricities, the planets are coplanar to within 9^(+8)_(-6) degrees, and therefore the inner planet's large eccentricity and close-in orbit are unlikely to be the result of Kozai migration. Moreover, even over many secular cycles, the inner planet's periapse is most likely never small enough for tidal circularization. Finally, we present and measure a transit time and impact parameter from four simultaneous ground-based light curves from 1 m class telescopes, demonstrating the feasibility of ground-based follow-up of Kepler giant planets exhibiting large TTVs