Effects of (2R,6R)-hydroxynorketamine in assays of acute pain-stimulated and pain-depressed behaviors in mice.

Ketamine has been shown to produce analgesia in various acute and chronic pain states; however, abuse liability concerns have limited its utility. The ketamine metabolite (2R,6R)-hydroxynorketamine (HNK) has been shown to produce antidepressant-like effects similar to ketamine without abuse liability concerns. (2R,6R)-HNK produces sustained analgesia in models of chronic pain, but has yet to be evaluated in models of acute pain. The present study evaluated the efficacy of acute (2R,6R)-HNK administration (one injection) in assays of pain-stimulated (52- and 56-degree hot plate test and acetic acid writhing) and pain-depressed behavior (locomotor activity and rearing) in male and female C57BL/6 mice. In assays of pain-stimulated behaviors, (2R,6R)-HNK (1-32 mg/kg) failed to produce antinociception in the 52- and 56-degree hot plate and acetic acid writhing assays. In assays of pain-depressed behaviors, 0.56% acetic acid produced a robust depression of locomotor activity and rearing that was not blocked by pretreatment of (2R,6R)-HNK (3.2-32 mg/kg). The positive controls morphine (hot plate test) and ketoprofen (acetic acid writhing, locomotor activity, and rearing) blocked pain-stimulated and pain-depressed behaviors. Finally, the effects of intermittent (2R,6R)-HNK administration were evaluated in 52-degree hot plate and pain-depressed locomotor activity and rearing. Intermittent administration of (2R,6R)-HNK also did not produce antinociceptive effects in the hot plate or pain-depressed locomotor activity assays. These results suggest that (2R,6R)-HNK is unlikely to have efficacy in treating acute pain; however, the efficacy of (2R,6R)-HNK in chronic pain states should continue to be evaluated.

Optimizing spinal cord injury in zebrafish larvae: effects of age on the injury response.

Zebrafish are an increasingly popular model to study spinal cord injury (SCI) regeneration. The transparency of larval zebrafish makes them ideal to study cellular processes in real time. Standardized approaches, including age of injury, are not readily available making comparisons of the results with other models challenging. In this study, we systematically examined the response to spinal cord transection of larval zebrafish at three different ages (3-7 days post fertilization or dpf) to determine whether the developmental complexity of the central nervous system affects the overall response to SCI. We then used imaging and behavioral analysis to evaluate whether differences existed based on the age of injury. All ages of larval zebrafish upregulated the required genes for glial bridge formation, ctgfa and gfap, at the site of injury, consistent with studies from adult zebrafish. Though all larval ages upregulated factors required to promote glial bridging, young larval zebrafish (3 dpf) were better able to regenerate axons independent of the glial bridge, unlike older zebrafish (7 dpf). Consistent with this data, locomotor experiments demonstrated that some swimming behavior occurs independent of glial bridge formation, further highlighting the need for standardization of this model and recovery assays. Overall, we found subtle cellular differences based on the age of transection in zebrafish, underlining the importance of considering age when designing experiments aimed at understanding regeneration.

Acyl Peptide Enzyme Hydrolase (APEH) activity is inhibited by lipid metabolites and peroxidation products.

Acyl Peptide Enzyme Hydrolase (APEH) activity is decreased in certain diseases but the mechanism and impact behind this loss in activity is not well understood. We hypothesized that lipid metabolites and lipid peroxidation products produced in inflammatory diseases may bind to and inhibit APEH activity. In vitro studies carried out in mammalian cell lysates, as well as with purified APEH protein, support our hypothesis that cellular lipid metabolites and lipid peroxidation products significantly decrease APEH activity. Enzymatic assays and molecular docking in silico analysis suggest that larger lipid metabolites are the best APEH inhibitors. APEH activity was measured in vivo in mice exposed to chronic e-cigarette vapor, as e-cigarettes are known to increase reactive oxygen species and lipid peroxidation products. In support of our in vitro findings, APEH activity in our mouse model demonstrates decreased APEH activity in the brains of mice exposed to e-cigarette vapor. These results provide a novel mechanism by which APEH activity may be inhibited in disease states. Furthermore, APEH inhibition may contribute to disease development and progression in pathologies associated with redox imbalances and can potentially act as biomarker for oxidative stress in disease.

Nicotine-free vapor inhalation produces behavioral disruptions and anxiety-like behaviors in mice: Effects of puff duration, session length, sex, and flavor.

Electronic-cigarette's (ECIGs) popularity has grown over the last decade and changed the way individuals administer nicotine. Preclinical research is imperative for understanding the addictive properties and health-risks associated with ECIG use; however, there is not a standard dosing regimen used across research laboratories. The main objective was to determine how vapor puff durations, administration session length, and flavored e-liquid alter general and mood-disorder related behaviors while providing a foundation of vapor administration parameters. Adult male and female C57BL/6 mice were exposed to several nicotine-free unflavored vapor puff durations (1, 3, 6, or 10 s) and vapor administration session lengths (10 and 30 min) then measured on the following assays: locomotor activity (LMA), tail suspension test (TST), and light-dark test. The effects of mecamylamine and the time-course of vapor-induced depression of LMA also were assessed. Additionally, mice were exposed to flavored (strawberry and adventurers tobacco blend) vapor inhalation and measured on locomotor activity, tail suspension test, and light-dark test. Following both 10 and 30 min vapor administration session, there was a puff duration-dependent decrease in distance traveled, time in center, and rearing. The vapor-induced depression of LMA was not mediated by nicotine or nicotinic acetylcholine receptor (nAChR) activation and lasted 60-90 min. The 10 s puff duration produced an anxiogenic-like effect in the light-dark test by decreasing the time spent in the light side. Vapor inhalation did not significantly alter TST behavior. No significant effects of sex or flavor were found. The anxiogenic-like effects of nicotine-free vapor inhalation are concerning as many adolescents vape nicotine-free flavored e-liquid, and there is an association between ECIGs and mood disorders. Additionally, these studies demonstrate that vapor puff duration, but not vapor administration session length, is an important variable to consider during research design as it can become a confounding variable and alter baseline behaviors.

Assessment of the rapid and sustained antidepressant-like effects of dextromethorphan in mice.

The glutamatergic system has emerged as a novel pathway for treating major depressive disorder (MDD) with the focus on producing both rapid and sustained antidepressant effects. Dextromethorphan is a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist that has produced antidepressant-like effects in forced swim and tail suspension tests (TST); however, the rapid and sustained antidepressant-like effects of dextromethorphan have not been evaluated. This study evaluated the rapid and sustained (24 h) antidepressant-like effects of dextromethorphan (0-32 mg/kg) in C56BL/6 mice using the novelty-induced hypophagia (NIH) test and TST, respectively. Additionally, we evaluated anxiety-related behavior and locomotor effects of dextromethorphan (0-56.0 mg/kg) using the light-dark and open field tests. Dextromethorphan (32 mg/kg) produced acute (30 min) antidepressant-like effects in TST, but failed to produce antidepressant-like effects 24 h after drug administration. Treatment of dextromethorphan (32 mg/kg) alone or in combination with CYP2D6 enzyme inhibitor Quinidine (32 mg/kg) failed to produce rapid antidepressant-like effects by increasing the latency to drink in the NIH test rather than decreasing the latency to drink. Dextromethorphan (56 mg/kg) produced an anxiogenic-like effect by decreasing the time spent in the light side, number of entries, and latency to enter the light side in the light-dark test. Administration of dextromethorphan (0-56 mg/kg) did not significantly alter locomotor activity. Although dextromethorphan is considered a noncompetitive NMDA receptor antagonist, dextromethorphan binds to several monoaminergic receptors (SERT and NET) and likely produces the antidepressant-like effects through these receptors similar to traditional antidepressant drugs. Additionally, these results suggest that the therapeutic window for dextromethorphan in the clinical population is small as similar doses produce antidepressant-like and anxiogenic-like behaviors.

Repeated nicotine vapor inhalation induces behavioral sensitization in male and female C57BL/6 mice.

Electronic cigarette use has significantly increased over the past decade. However, there is limited preclinical research on the behavioral and abuse-related effects of nicotine vapor inhalation in rodents. The present study evaluates the effects of repeated nicotine vapor inhalation in male and female mice using a nicotine behavioral sensitization model. Male and female C57BL/6 mice were administered vaporized nicotine (0-10.0 mg/ml) or the positive control of intraperitoneally administered nicotine (0.5 mg/kg) once daily for 5 days, and locomotor activity was assessed. Body temperatures were measured before and after nicotine vapor inhalation to assess hypothermia. Nicotine vapor inhalation (1.0-3.0 mg/ml) produced a dose-dependent behavioral sensitization effect and produced hypothermia in male and female mice. Nicotine (0.5 mg/kg) also produced significant behavioral sensitization. No sex differences were found for nicotine behavioral sensitization with either route of administration. Pretreatment with the nonselective nicotinic antagonist mecamylamine blocked the behavioral sensitization produced by 1.0 mg/ml of nicotine vapor inhalation. These results established that nicotine vapor inhalation produces behavioral sensitization in an inverted U-shaped curve that is similar to the effects of injected nicotine across several behavioral models. Additionally, pretreatment with mecamylamine demonstrated that nicotinic receptor activation was responsible for the behavioral sensitization produced by nicotine vapor inhalation and was not a conditioned response to the vapor. The methods used in the present study provide an additional behavioral approach for evaluating the behavioral effects of repeated nicotine vapor inhalation that allows the manipulation of several variables, including e-liquid oil blend, e-liquid flavors, puff duration, etc.