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Caged naloxone reveals opioid signaling deactivation kinetics.

ABSTRACT: The spatiotemporal dynamics of opioid signaling in the brain remain poorly defined. Photoactivatable opioid ligands provide a means to quantitatively measure these dynamics and their underlying mechanisms in brain tissue. Although activation kinetics can be assessed using caged agonists, deactivation kinetics are obscured by slow clearance of agonist in tissue. To reveal deactivation kinetics of opioid signaling we developed a caged competitive antagonist that can be quickly photoreleased in sufficient concentrations to render agonist dissociation effectively irreversible. Carboxynitroveratryl-naloxone (CNV-NLX), a caged analog of the competitive opioid antagonist NLX, was readily synthesized from commercially available NLX in good yield and found to be devoid of antagonist activity at heterologously expressed opioid receptors. Photolysis in slices of rat locus coeruleus produced a rapid inhibition of the ionic currents evoked by multiple agonists of the ?-opioid receptor (MOR), but not of ?-adrenergic receptors, which activate the same pool of ion channels. Using the high-affinity peptide agonist dermorphin, we established conditions under which light-driven deactivation rates are independent of agonist concentration and thus intrinsic to the agonist-receptor complex. Under these conditions, some MOR agonists yielded deactivation rates that are limited by G protein signaling, whereas others appeared limited by agonist dissociation. Therefore, the choice of agonist determines which feature of receptor signaling is unmasked by CNV-NLX photolysis.

SUBMITTER: Banghart MR

PROVIDER: S-EPMC3807075 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Caged naloxone reveals opioid signaling deactivation kinetics.

Banghart Matthew R MR Williams John T JT Shah Ruchir C RC Lavis Luke D LD Sabatini Bernardo L BL

Molecular pharmacology 20130819 5

The spatiotemporal dynamics of opioid signaling in the brain remain poorly defined. Photoactivatable opioid ligands provide a means to quantitatively measure these dynamics and their underlying mechanisms in brain tissue. Although activation kinetics can be assessed using caged agonists, deactivation kinetics are obscured by slow clearance of agonist in tissue. To reveal deactivation kinetics of opioid signaling we developed a caged competitive antagonist that can be quickly photoreleased in suf ...[more]

PMID: 23960100

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Project description:BACKGROUND:North America is in the midst of an unabated opioid overdose epidemic due to the increasing non-medical use of fentanyl and ultra-potent opioids. Naloxone is an effective antidote to opioid toxicity, yet its optimal dosing in the context of fentanyl and ultra-potent opioid overdoses remains unknown. This review aims to determine the relationship between the first empiric dose of naloxone and reversal of toxicity, adverse events, and the total cumulative dose required among patients with undifferentiated opioid overdoses and those with suspected toxicity from ultra-potent opioids. Secondary objectives include evaluating the relationship between the cumulative naloxone dose and toxicity reversal and adverse events, among patients with undifferentiated opioid overdoses and those with suspected toxicity from ultra-potent opioids. METHODS:To identify studies, we will search MEDLINE, Embase, CENTRAL, DARE, CDAG, CINAHL, Science Citation Index, multiple trial registries, and the gray literature. Included studies will evaluate patients with suspected or confirmed opioid toxicity from undifferentiated opioids and ultra-potent opioids, who received an empiric and possibly additional doses of naloxone. The main outcomes of interest are the relationship between naloxone dose and toxicity reversal and adverse events. We will include controlled and non-controlled interventional studies, observational studies, case reports/series, and reports from poison control centers. We will extract data and assess study quality in duplicate with discrepancies resolved by consensus or a third party. We will use the Downs and Black and Cochrane risk of bias tools for observational and randomized controlled studies. If we find sufficient variation in dose, we will fit a random effects one-stage model to estimate a dose-response relationship. We will conduct multiple subgroup analyses, including by type of opioid used and by suspected high and low prevalence of ultra-potent opioid use based on geographic location and time of the original studies. DISCUSSION:Our review will include the most up-to-date available data including ultra-potent opioids to inform the current response to the opioid epidemic, addressing the limitations of recent reviews. We anticipate limitations relating to study heterogeneity. We will disseminate study results widely to update overdose treatment guidelines and naloxone dosing in Take Home Naloxone programs.

Project description:In mammals, the photopigment melanopsin (Opn4) is found in a subset of retinal ganglion cells that serve light detection for circadian photoentrainment and pupil constriction (i.e., mydriasis). For a given species, the efficiency of photoentrainment and length of time that mydriasis occurs is determined by the spectral sensitivity and deactivation kinetics of melanopsin, respectively, and to date, neither of these properties have been described in marine mammals. Previous work has indicated that the absorbance maxima (λmax) of marine mammal rhodopsins (Rh1) have diversified to match the available light spectra at foraging depths. However, similar to the melanopsin λmax of terrestrial mammals (~480 nm), the melanopsins of marine mammals may be conserved, with λmax values tuned to the spectrum of solar irradiance at the water's surface. Here, we investigated the Opn4 pigments of 17 marine mammal species inhabiting diverse photic environments including the Infraorder Cetacea, as well as the Orders Sirenia and Carnivora. Both genomic and cDNA sequences were used to deduce amino acid sequences to identify substitutions most likely involved in spectral tuning and deactivation kinetics of the Opn4 pigments. Our results show that there appears to be no amino acid substitutions in marine mammal Opn4 opsins that would result in any significant change in λmax values relative to their terrestrial counterparts. We also found some marine mammal species to lack several phosphorylation sites in the carboxyl terminal domain of their Opn4 pigments that result in significantly slower deactivation kinetics, and thus longer mydriasis, compared to terrestrial controls. This finding was restricted to cetacean species previously found to lack cone photoreceptor opsins, a condition known as rod monochromacy. These results suggest that the rod monochromat whales rely on extended pupillary constriction to prevent photobleaching of the highly photosensitive all-rod retina when moving between photopic and scotopic conditions.

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Project description:IntroductionEmergency departments (EDs) may be high-yield venues to address opioid deaths with education on both overdose prevention and appropriate actions in a witnessed overdose. In addition, the ED has the potential to equip patients with nasal naloxone kits as part of this effort. We evaluated the feasibility of an ED-based overdose prevention program and described the overdose risk knowledge, opioid use, overdoses, and overdose responses among participants who received overdose education and naloxone rescue kits (OEN) and participants who received overdose education only (OE).MethodsProgram participants were surveyed by telephone after their ED visit about their substance use, overdose risk knowledge, history of witnessed and personal overdoses, and actions in a witnessed overdose including use of naloxone.ResultsA total of 415 ED patients received OE or OEN between January 1, 2011 and February 28, 2012. Among those, 51 (12%) completed the survey; 37 (73%) of those received a naloxone kit, and 14 (27%) received OE only. Past 30-day opioid use was reported by 35% OEN and 36% OE, and an overdose was reported by 19% OEN and 29% OE. Among 53% (27/51) of participants who witnessed another individual experiencing an overdose, 95% OEN and 88% OE stayed with victim, 74% OEN and 38% OE called 911, 26% OEN and 25% OE performed rescue breathing, and 32% OEN (n=6) used a naloxone kit to reverse the overdose. We did not detect statistically significant differences between OEN and OE-only groups in opioid use, overdose or response to a witnessed overdose.ConclusionThis is the first study to demonstrate the feasibility of ED-based opioid overdose prevention education and naloxone distribution to trained laypersons, patients and their social network. The program reached a high-risk population that commonly witnessed overdoses and that called for help and used naloxone, when available, to rescue people. While the study was retrospective with a low response rate, it provides preliminary data for larger, prospective studies of ED-based overdose prevention programs.

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Caged naloxone reveals opioid signaling deactivation kinetics.

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Caged naloxone reveals opioid signaling deactivation kinetics.

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