Altered Transition Between Agonist-and Antagonist-Favoring States of Μ-Opioid Receptor in Brain Membranes with Modified Microviscosity

In unmodified synaptosomal brain membranes the presence of NaCl inhibited the binding to Μ receptors of the tritiated opioid agonists etorphine, Tyr-D-Ala-Gly-(Me)Phe-Gly-ol, and sufentanil by 53, 43, and 37%, respectively, and increased that of the antagonist [ 3 H]naltrexone by 54%. On the other hand, in membranes whose microviscosity was increased by incorporation of cholesteryl hemi-succinate (CHS) the effects of sodium on opioid agonist and antagonist binding were abolished and strongly reduced, respectively. Furthermore, in the modified membranes the ability of sodium to protect the opioid receptor from inactivation by the sulfhydryl-reactive agent N -ethyl-maleimide (NEM) was diminished. In CHS-treated membranes whose elevated microviscosity was reduced by the incorporation of oleic acid, the effectiveness of sodium in modulating opioid binding and attenuating receptor inactivation by NEM was restored. The results implicate membrane microviscosity in the mechanism by which sodium modulates the conversion between agonist-and antagonist-favoring states of Μ opioid receptor.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65931/1/j.1471-4159.1993.tb03630.x.pd

Altered Microviscosity at Brain Membrane Surface Induces Distinct and Reversible Inhibition of Opioid Receptor Binding

In synaptosomal membranes from rat and monkey brain cortex, the addition of petroselenic (18:1, cis -Δ 6 ) acid, oleic (18:1, m-Δ 9 ) acid, and vaccenic (18:1, cis -Δ 11 ) acid or their corresponding methyl esters at 0.5 Μmol/mg of membrane protein caused a similar 7–10% decrease in the microviscosity of the membrane core, whereas at the membrane surface the microviscosity was reduced 5–7% by the fatty acids but only 1% by their methyl esters. Concomi-tantly, the fatty acids, but not the methyl esters, inhibited the specific binding of the tritiated Μ-, Δ-, and K-opioids Tyr-D-Ala-Gly-(Me)Phe-Gly-ol (DAMGO), [D-Pen 2 ,D-Pen 5 ]-enkephalin (DPDPE), and U69,593, respectively. As shown with oleic acid, the sensitivity of opioid receptor binding toward inhibition by fatty acids was in the order Δ > ΜΚ k , whereby the binding of [ 3 H]DPDPE was abolished, but significant inhibition of [ 3 H]U69,593 binding, determined in membranes from monkey brain, required membrane modification with a twofold higher fatty acid concentration. Except for the unchanged K D of [ 3 H]U69,593, the inhibition by oleic acid involved both the B max and affinity of opioid binding. Cholesteryl hemisuccinate (0.5–3 Μmol/mg of protein), added to membranes previously modified by fatty acids, reversed the fluidization caused by the latter compounds and restored inhibited Μ-, Δ-, and k -opioid binding toward control values. In particular, the B max of [ 3 H]-DPDPE binding completely recovered after being undetectable. The results implicate membrane surface fluidity in the modulation of opioid receptor binding, reveal distinct sensitivity of Δ, Μ, and K receptors toward that modulation, and identify unsaturated fatty acids and cholesterol as possible endogenous regulators of opioid receptor function.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/66380/1/j.1471-4159.1992.tb08432.x.pd

Modulation of opioid signal transduction in SH-SY5Y neural cells by differentiating agents: Concurrent mu receptor upregulation and effector desensitization by phorbol ester

Long-term exposure of SH-SY5Y human neural cells to retinoic acid (RA) increased the binding of [3H]DAMGO and [3H]DPDPE by up to 3-fold compared to membranes of untreated cells. In contrast, incubation of the cells with phorbol ester (TPA) selectively up-regulated the binding of [3H]DAMGO. RA enhanced the maximal inhibition of cAMP formation in SH-SY5Y cells by both DAMGO and DPDPE from 20% for both opioids in control cells to 75% and 50%, respectively. On the other hand, the effect of TPA was limited to a marginal increase in the inhibition of cAMP formation by DAMGO. Parameters of GTP[gamma]35S binding as well as low-Km GTPase activity revealed no deterioration in the structure or function of total G protein in the TPA-treated cells, and initial Western blots showed no difference in the cell content of Go. Ongoing experiments are focusing on the covalent modification of specific G protein subtypes involved in [mu] and [delta] opioid signal transduction in SH-SY5Y cells differentiated by TPA.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/31768/1/0000709.pd

Study protocol for a cluster randomised controlled factorial design trial to assess the effectiveness and feasibility of reactive focal mass drug administration and vector control to reduce malaria transmission in the low endemic setting of Namibia.

INTRODUCTION: To interrupt malaria transmission, strategies must target the parasite reservoir in both humans and mosquitos. Testing of community members linked to an index case, termed reactive case detection (RACD), is commonly implemented in low transmission areas, though its impact may be limited by the sensitivity of current diagnostics. Indoor residual spraying (IRS) before malaria season is a cornerstone of vector control efforts. Despite their implementation in Namibia, a country approaching elimination, these methods have been met with recent plateaus in transmission reduction. This study evaluates the effectiveness and feasibility of two new targeted strategies, reactive focal mass drug administration (rfMDA) and reactive focal vector control (RAVC) in Namibia. METHODS AND ANALYSIS: This is an open-label cluster randomised controlled trial with 2×2 factorial design. The interventions include: rfMDA (presumptive treatment with artemether-lumefantrine (AL)) versus RACD (rapid diagnostic testing and treatment using AL) and RAVC (IRS with Acellic 300CS) versus no RAVC. Factorial design also enables comparison of the combined rfMDA+RAVC intervention to RACD. Participants living in 56 enumeration areas will be randomised to one of four arms: rfMDA, rfMDA+RAVC, RACD or RACD+RAVC. These interventions, triggered by index cases detected at health facilities, will be targeted to individuals residing within 500 m of an index. The primary outcome is cumulative incidence of locally acquired malaria detected at health facilities over 1 year. Secondary outcomes include seroprevalence, infection prevalence, intervention coverage, safety, acceptability, adherence, cost and cost-effectiveness. ETHICS AND DISSEMINATION: Findings will be reported on clinicaltrials.gov, in peer-reviewed publications and through stakeholder meetings with MoHSS and community leaders in Namibia. TRIAL REGISTRATION NUMBER: NCT02610400; Pre-results

Effectiveness of reactive focal mass drug administration and reactive focal vector control to reduce malaria transmission in the low malaria-endemic setting of Namibia: a cluster-randomised controlled, open-label, two-by-two factorial design trial.

BACKGROUND: In low malaria-endemic settings, screening and treatment of individuals in close proximity to index cases, also known as reactive case detection (RACD), is practised for surveillance and response. However, other approaches could be more effective for reducing transmission. We aimed to evaluate the effectiveness of reactive focal mass drug administration (rfMDA) and reactive focal vector control (RAVC) in the low malaria-endemic setting of Zambezi (Namibia). METHODS: We did a cluster-randomised controlled, open-label trial using a two-by-two factorial design of 56 enumeration area clusters in the low malaria-endemic setting of Zambezi (Namibia). We randomly assigned these clusters using restricted randomisation to four groups: RACD only, rfMDA only, RAVC plus RACD, or rfMDA plus RAVC. RACD involved rapid diagnostic testing and treatment with artemether-lumefantrine and single-dose primaquine, rfMDA involved presumptive treatment with artemether-lumefantrine, and RAVC involved indoor residual spraying with pirimiphos-methyl. Interventions were administered within 500 m of index cases. To evaluate the effectiveness of interventions targeting the parasite reservoir in humans (rfMDA vs RACD), in mosquitoes (RAVC vs no RAVC), and in both humans and mosquitoes (rfMDA plus RAVC vs RACD only), an intention-to-treat analysis was done. For each of the three comparisons, the primary outcome was the cumulative incidence of locally acquired malaria cases. This trial is registered with ClinicalTrials.gov, number NCT02610400. FINDINGS: Between Jan 1, 2017, and Dec 31, 2017, 55 enumeration area clusters had 1118 eligible index cases that led to 342 interventions covering 8948 individuals. The cumulative incidence of locally acquired malaria was 30·8 per 1000 person-years (95% CI 12·8-48·7) in the clusters that received rfMDA versus 38·3 per 1000 person-years (23·0-53·6) in the clusters that received RACD; 30·2 per 1000 person-years (15·0-45·5) in the clusters that received RAVC versus 38·9 per 1000 person-years (20·7-57·1) in the clusters that did not receive RAVC; and 25·0 per 1000 person-years (5·2-44·7) in the clusters that received rfMDA plus RAVC versus 41·4 per 1000 person-years (21·5-61·2) in the clusters that received RACD only. After adjusting for imbalances in baseline and implementation factors, the incidence of malaria was lower in clusters receiving rfMDA than in those receiving RACD (adjusted incidence rate ratio 0·52 [95% CI 0·16-0·88], p=0·009), lower in clusters receiving RAVC than in those that did not (0·48 [0·16-0·80], p=0·002), and lower in clusters that received rfMDA plus RAVC than in those receiving RACD only (0·26 [0·10-0·68], p=0·006). No serious adverse events were reported. INTERPRETATION: In a low malaria-endemic setting, rfMDA and RAVC, implemented alone and in combination, reduced malaria transmission and should be considered as alternatives to RACD for elimination of malaria. FUNDING: Novartis Foundation, Bill & Melinda Gates Foundation, and Horchow Family Fund

Opioid receptor binding and receptor-effector coupling in intact neurons and their isolated membranes

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/28467/1/0000258.pd