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Neurotrophins prevent spiral ganglion neuron (SGN) degeneration in animal models of ototoxin-induced deafness and may be used in the future to improve the hearing of cochlear implant patients. It is increasingly common for patients with residual hearing to undergo cochlear implantation. However, the effect of neurotrophin treatment on acoustic hearing is not known. In this study, brain-derived neurotrophic factor (BDNF) was applied to the round window membrane of adult guinea pigs for 4 weeks using a cannula attached to a mini-osmotic pump. SGN survival was first assessed in ototoxically deafened guinea pigs to establish that the delivery method was effective. Increased survival of SGNs was observed in the basal and middle cochlear turns of deafened guinea pigs treated with BDNF, confirming that delivery to the cochlea was successful. The effects of BDNF treatment in animals with normal hearing were then assessed using distortion product otoacoustic emissions (DPOAEs), pure tone, and click-evoked auditory brainstem responses (ABRs). DPOAE assessment indicated a mild deficit of 5 dB SPL in treated and control groups at 1 and 4 weeks after cannula placement. In contrast, ABR evaluation showed that BDNF lowered thresholds at specific frequencies (8 and 16 kHz) after 1 and 4 weeks posttreatment when compared to the control cohort receiving Ringer's solution. Longer treatment for 4 weeks not only widened the range of frequencies ameliorated from 2 to 32 kHz but also lowered the threshold by at least 28 dB SPL at frequencies ≥16 kHz. BDNF treatment for 4 weeks also increased the amplitude of the ABR response when compared to either the control cohort or prior to treatment. We show that BDNF applied to the round window reduces auditory thresholds and could potentially be used clinically to protect residual hearing following cochlear implantation. [ABSTRACT FROM AUTHOR]

Mitragynine, an alkaloid present in the leaves of Mitragyna speciosa (kratom), has a complex pharmacology that includes low efficacy agonism at μ-opioid receptors (MORs). This study examined the activity of mitragynine at adrenergic α 2 receptors (Aα 2 Rs) in vitro and in vivo. Mitragynine displaced a radiolabeled Aα 2 R antagonist ([ 3 H]RX821002) from human Aα 2A Rs in vitro with lower affinity (K i  = 1260 nM) than the agonists (-)-epinephrine (K i  = 263 nM) or lofexidine (K i  = 7.42 nM). Mitragynine did not significantly stimulate [ 35 S]GTPγS binding at Aα 2A Rs in vitro, but in rats trained to discriminate 32 mg/kg mitragynine from vehicle (intraperitoneally administered; i.p.), mitragynine exerted an Aα 2 R agonist-like effect. Both α 2 R antagonists (atipamezole and yohimbine) and MOR antagonists (naloxone and naltrexone) produced rightward shifts in mitragynine discrimination dose-effect function and Aα 2 R agonists lofexidine and clonidine produced leftward shifts. In the mitragynine trained rats, Aα 2 R agonists also produced leftward shifts in discrimination dose-effect functions for morphine and fentanyl. In a separate rat cohort trained to discriminate 3.2 mg/kg i.p. morphine from vehicle, naltrexone produced a rightward shift, but neither an Aα 2 R agonist or antagonist affected morphine discrimination. In a hypothermia assay, both lofexidine and clonidine produced marked effects antagonized by yohimbine. Mitragynine did not produce hypothermia. Together, these data demonstrate that mitragynine acts in vivo like an Aα 2 R agonist, although its failure to induce hypothermia or stimulate [ 35 S]GTPγS binding in vitro, suggests that mitragynine maybe a low efficacy Aα 2 R agonist., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

This study reports the steady-state pharmacokinetic parameters for mitragynine and characterizes its elimination in male and female rats. Four male and female rats were dosed q12h with 40 mg/kg, and orally administered mitragynine for 5 and 6 days, respectively. Using a validated ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method, the plasma concentrations of mitragynine, its metabolites (7-hydroxymitragynine, 9-hydroxycorynantheidine, and mitragynine acid), and a non-CYP oxidation product (3-dehydromitragynine) were determined at various time points. Sex differences in pharmacokinetics were observed, with females demonstrating significantly higher systemic exposure of mitragynine than males. The mitragynine area under the curve normalized by the dose interval (AUC/τ) was 6741.6 ± 869.5 h*ng/mL in female rats and 1808.9 ± 191.3 h*ng/mL in males ( p < 0.05). Both sexes produced similar metabolite profiles; the major metabolites were mitragynine acid and 9-hydroxycorynantheidine. 7-Hydroxymitragynine was a minor metabolite. However, higher exposure (AUCs) and the maximum plasma concentrations ( C max ) of active metabolites, 7-hydroxymitragynine and 9-hydroxycorynantheidine, were observed in female rats and exhibited substantial sex differences. Renal clearance of mitragynine (CL r ) was low (0.64 ± 0.3 mL/h in males and 0.98 ± 0.4 mL/h in females), and unchanged mitragynine accounted for <1% of the dose excreted in feces (both sexes). The clinical chemistry, complete blood count, and hematological test results reported no abnormal hematological findings after multiple dosing in either sex., Competing Interests: The authors declare no competing financial interest., (© 2024 American Chemical Society.)

Introduction: Intraoperative trauma leading to bleeding during cochlear implantation negatively impacts residual hearing of cochlear implant recipients. There are no clinical protocols for the removal of blood during implantation, to reduce the consequential effects such as inflammation and fibrosis which adversely affect cochlear health and residual hearing. This preclinical study investigated the implementation of an intra-cochlear flushing protocol for the removal of blood., Methods: Three groups of guinea pigs were studied for 28 days after cochlear implantation; cochlear implant-only (control group); cochlear implant with blood injected into the cochlea (blood group); and cochlear implant, blood injection, and flushing of the blood from the cochlea intraoperatively (flush group). Auditory brainstem responses (ABRs) in addition to tissue response volumes were analyzed and compared between groups., Results: After implantation, the blood group exhibited the highest ABR thresholds when compared to the control and flush group, particularly in the high frequencies. On the final day, the control and blood group had similar ABR thresholds across all frequencies tested, whereas the flush group had the lowest thresholds, significantly lower at 24 kHz than the blood and control group. Analysis of the tissue response showed the flush group had significantly lower tissue responses in the basal half of the array when compared with the blood and control group., Conclusions: Flushing intra-cochlear blood during surgery resulted in better auditory function and reduced subsequent fibrosis in the basal region of the cochlea. This finding prompts the implementation of a flushing protocol in clinical cochlear implantation., Level of Evidence: N/A Laryngoscope, 134:1410-1416, 2024., (© 2023 The Authors. The Laryngoscope published by Wiley Periodicals LLC on behalf of The American Laryngological, Rhinological and Otological Society, Inc.)

Background: Sleep disturbance is commonly reported among individuals meeting criteria for cannabis use disorder (CUD), and people who use cannabis frequently report sleep disturbance as a contributor to failed quit attempts. The purpose of this study was to measure sleep in individuals enrolled in treatment for CUD, and to determine whether use of hypnotic medication during treatment increased abstinence rates., Method: The study enrolled 127 adults seeking treatment for CUD in a 12-week clinical trial and randomized to receive extended-release zolpidem (zolpidem-XR) or placebo. All participants received computerized behavioral therapy and abstinence-based contingency management. The study conducted in-home ambulatory polysomnography (PSG) assessments at baseline and during treatment to objectively measure sleep. Self-report measures of recent sleep, Insomnia Severity Index (ISI), and drug use (Timeline Follow-Back) were collected at each study visit, and the study confirmed self-reported abstinence via quantitative urine drug testing., Result: Participants randomized to placebo, but not zolpidem-XR exhibited significant sleep disturbance during week 1 of treatment. Sleep disturbance emerged in the zolpidem-XR group after study medication was stopped at the end of treatment. Though participants assigned to the zolpidem-XR condition had qualitatively greater rates of abstinence compared with placebo (27 % versus 15 % negative at end of treatment), the difference was not statistically significant. Treatment retention was poor (about 50 % drop out in both groups) and medication adherence was a challenge without the use of contingent incentives., Conclusion: Results from this randomized controlled trial suggest that zolpidem-XR can attenuate abstinence-induced sleep disturbance early in treatment for CUD, but that sleep problems are likely to emerge after the medication is stopped. Further research should identify alternative pharmacotherapies and behavioral treatments for CUD and elucidate the role of sleep disturbance in the development and maintenance of CUD., (Copyright © 2023. Published by Elsevier Inc.)

Importance: Buprenorphine treatment for opioid use disorder (OUD) has more than doubled since 2009. However, current US Food and Drug Administration buprenorphine dosing guidelines are based on studies among people using heroin, prior to the emergence of fentanyl in the illicit drug supply., Objective: To estimate the association between buprenorphine dose and time to treatment discontinuation during a period of widespread fentanyl availability., Design, Setting, and Participants: This retrospective cohort study used statewide Rhode Island Prescription Drug Monitoring Program data. Participants were Rhode Island residents initiating buprenorphine treatment for OUD between October 1, 2016, and September 30, 2020. Data analysis was performed from December 9, 2022, to August 10, 2023., Exposure: Daily dose of buprenorphine (16 mg and 24 mg) defined starting on the day of initiation based on total quantity and days' supply dispensed. Patients were censored on any dose change., Main Outcomes and Measures: Buprenorphine treatment discontinuation in the 180 days following initiation, defined as a gap in treatment of more than 27 days based on prescription fill dates and days' supply. Kaplan-Meier and Cox regression survival analyses were conducted to estimate the association between buprenorphine dose and time to treatment discontinuation, controlling for potential informative censoring and measured potential confounders., Results: Among 6499 patients initiating buprenorphine treatment for OUD, most were aged 25 to 44 years (57%; n = 3682), were male (61%; n = 3950), and had private (47%; n = 3025) or Medicaid (33%; n = 2153) insurance. More than half of patients were prescribed a daily dose of interest at initiation (16 mg: 50%; n = 3264; 24 mg: 10%; n = 668). In Kaplan-Meier analyses, 58% of patients discontinued buprenorphine treatment within 180 days (16 mg: 59% vs 24 mg: 53%; log-rank test P = .005). In Cox regression analyses, patients prescribed a dose of 16 mg had a greater risk of treatment discontinuation than those prescribed 24 mg (adjusted hazard ratio, 1.20; 95% CI, 1.06-1.37)., Conclusions and Relevance: In this cohort study of patients initiating buprenorphine treatment from 2016 to 2020, patients prescribed a 24 mg dose of buprenorphine remained in treatment longer than those prescribed 16 mg. The value of higher buprenorphine doses than currently recommended needs to be considered for improving retention in treatment.

Background: Systemically administered steroids are widely utilised for hearing preservation therapies. More recently, steroids have been administered to achieve hearing protection after cochlear implant surgery. Currently there is a lack of understanding as to which administration route offers most therapeutic efficacy, local or systemic administration. Paramount to this are observations in animal studies that systemic administration following implantation offers hearing protection and reduced cochlear fibrosis, despite observations that perilymphatic levels are up to 10-fold higher after local administration in non-implanted cochleae., Aims/objectives: This paper explores the impact that cochlear implantation and associated acute inflammation has on steroid distribution and uptake following systemic administration of dexamethasone., Material and Methods: Eight guinea pigs received systemic dexamethasone 60 min prior to cochlear implantation. Implanted and contralateral non-implanted cochlea were harvested for tissue immunohistochemistry and detection of dexamethasone., Results: Cochleostomy with scala tympani implantation resulted in a significant increase in cochlear dexamethasone signal. This was most notable at the organ of Corti, stria vascularis, and blood product in the scala tympani., Conclusions and Significance: This study demonstrates that the inner ear distribution of systemically administered steroids is enhanced following surgery for cochlear implantation and provides rationale for systemic perioperative steroids in hearing preservation surgery.

Background: Endolymphatic hydrops (EH) has been observed in both animal and human cochleae following cochlear implant (CI) surgery. We tested whether EH could be eliminated by administration of mineralocorticoid steroid antagonist spironolactone and explored the electrophysiological consequences of this., Methods: Sixty-four adult guinea pigs underwent cochlear implantation with a dummy electrode. Animals then survived either 2, 7, or 28 days. Auditory function was monitored by recording electrocochleography from the round window membrane preimplantation, and on the last day of the experiment. Spironolactone or control solution was added to animals' feed for 7 days (if they survived that long) beginning immediately prior to surgery. The presence of EH was determined using thin-sheet laser imaging microscopy., Results: Treatment with spironolactone resulted in significant reduction in EH in the second cochlear turn 7 days postimplantation. In all animals, the compound action potential (CAP) threshold was elevated 2 days postimplantation, but for most frequencies had recovered substantially by 28 days. There was no treatment effect on CAP thresholds. SP/AP ratios were elevated at day 2. The amplitude growth of the CAP did not differ between test and control groups at any time after implantation., Conclusions: EH can be suppressed by antagonism of mineralocorticoid receptors in the week after cochlear implantation. Reduction in EH did not lead to any change in hearing, and there was no indication of synaptopathy signalled by reduced CAP amplitude at high sound intensities. We found no electrophysiological evidence that EH early after implantation impacts negatively upon preservation of residual hearing., Competing Interests: The authors disclose no conflict of interest., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of Otology & Neurotology, Inc.)

Kratom alkaloids have mostly been evaluated for their opioid activity but less at other targets that could contribute to their physiological effects. Here, we investigated the in vitro and in vivo activity of kratom alkaloids at serotonin receptors (5-HTRs). Paynantheine and speciogynine exhibited high affinity for 5-HT 1A Rs and 5-HT 2B Rs, unlike the principal kratom alkaloid mitragynine. Both alkaloids produced antinociceptive properties in rats via an opioid receptor-independent mechanism, and neither activated 5-HT 2B Rs in vitro. Paynantheine, speciogynine, and mitragynine induced lower lip retraction and antinociception in rats, effects blocked by a selective 5-HT 1A R antagonist. In vitro functional assays revealed that the in vivo 5-HT 1A R agonistic effects may be due to the metabolites 9- O -desmethylspeciogynine and 9- O -desmethylpaynantheine and not the parent compounds. Both metabolites did not activate 5-HT 2B R, suggesting low inherent risk of causing valvulopathy. The 5-HT 1A R agonism by kratom alkaloids may contribute to the mood-enhancing effects associated with kratom use.

Ten indole and oxindole alkaloids ( 1 - 10 ) were isolated from the freshly collected leaves of Malaysian Mitragyna speciosa (Kratom). The chemical structures of these compounds were established on the basis of extensive 1D and 2D NMR and HRMS data analysis. The spectroscopic data of mitragynine oxindole B ( 4 ) are reported herein for the first time. The spatial configuration of mitragynine oxindole B ( 4 ) was confirmed by single-crystal X-ray diffraction. Simultaneous quantification of the isolated alkaloids in the M. speciosa leaf specimens collected from different locations in the northern region of Peninsular Malaysia was also performed using UPLC-MS/MS. The oxindole alkaloids ( 1 - 4 ) and the indole alkaloid ( 10 ) were assessed for binding affinity at opioid receptors. Corynoxine ( 1 ) showed high binding affinity to μ-opioid receptors with a K i value of 16.4 nM. Further, corynoxine ( 1 ) was 1.8-fold more potent than morphine in rats subjected to a nociceptive hot plate assay. These findings have important implications for evaluating the combined effects of the minor oxindole alkaloids in the overall therapeutic activity of M. speciosa .

Adherence monitoring is a vital component of clinical efficacy trials, as the regularity of medication consumption affects both efficacy and adverse effect profiles. Pill-counts do not confirm consumption, and invasive plasma assessments can only assist post hoc assessments. We previously reported on the pharmacokinetics of a potential adherence marker to noninvasively monitor dosage consumption during a trial without breaking a blind. We reported that consumption cessation of subtherapeutic 15 mg acetazolamide (ACZ) doses showed a predictable urinary excretion decay that was quantifiable for an extended period. The current study describes the clinical implementation of 15 mg ACZ doses as an adherence marker excipient in distinct cohorts taking ACZ for different "adherence" durations. We confirm that ACZ output did not change (accumulate) during 18-20 days of adherence, and developed and assessed urinary cutoffs as nonadherence indicators. We demonstrate that whereas an absolute concentration cutoff (989 ng/mL) lacked sensitivity, a creatinine normalized equivalent (1,376 ng/mg ACZ) was highly accurate at detecting nonadherence. We also demonstrate that during nonadherent phases of three trials, creatinine-normalized urinary ACZ elimination was reproducible within and across trials with low variability. Excretion was first order, with a decay half-life averaging ~ 2.0 days. Further, excretion remained quantifiable for 14 days, providing a long period during which the date of last consumption might be determined. We conclude that inclusion of 15 mg ACZ as a dosage form adherence marker excipient, provides a reliable and sensitive mechanism to confirm medication consumption and detect nonadherence during clinical efficacy trials., (Published 2020. This article is a U.S. Government work and is in the public domain in the USA.)

Kratom is widely consumed in the United States for self-treatment of pain and opioid withdrawal symptoms. Mitragynine is the most abundant alkaloid in kratom and is a μ-opioid receptor agonist. 7-Hydroxymitragynine (7-HMG) is a mitragynine metabolite that is a more potent and efficacious opioid than its parent mitragynine. 7-HMG contributes to mitragynine's antinociceptive effects in mice, but evidence suggests it may also have a higher abuse potential. This in vitro study demonstrates that 7-HMG is stable in rodent and monkey plasma but is unstable in human plasma. Surprisingly, in human plasma 7-HMG is converted to mitragynine pseudoindoxyl, an opioid that is even more potent than either mitragynine or 7-HMG. This novel metabolite is formed in human plasma to a much greater extent than in the preclinical species tested (mouse, rat, dog, and cynomolgus monkey) and due to its μ-opioid potency may substantially contribute to the pharmacology of kratom in humans to a greater extent than in other tested species., Competing Interests: The authors declare no competing financial interest.

Visualisation of cochlear histopathology in three-dimensions has been long desired in the field of hearing research. This paper outlines a technique that has made this possible and shows a research application in the field of hearing protection after cochlear implantation. The technique utilises robust immunofluorescent labelling followed by effective tissue clearing and fast image acquisition using Light Sheet Microscopy. We can access the health of individual components by immunofluorescent detection of proteins such as myosin VIIa to look at cochlear hair cells, NaKATPase alpha 3 to look at spiral ganglion neurons, and IBA1 to look at macrophages within a single cochlea, whilst maintaining the integrity of fine membranous structures and keeping the cochlear implant in place. This allows the tissue response to cochlear implantation to be studied in detail, including the immune reaction to the implant and the impact on the structure and health of neural components such as hair cells. This technique reduces time and labour required for sectioning of cochleae and can allow visualisation of cellular detail. Use of image analysis software allows conversion of high-resolution image stacks into three-dimensional interactive data sets so volumes and numbers of surfaces can be measured. Immunofluorescent whole cochlea labelling and Light Sheet Microscopy have the capacity to be applied to many questions in hearing research of both the cochlea and vestibular system., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Cochlear implantation leads to many structural changes within the cochlea which can impair residual hearing. In patients with preserved low-frequency hearing, a delayed hearing loss can occur weeks-to-years post-implantation. We explore whether stiffening of the basilar membrane (BM) may be a contributory factor in an animal model. Our objective is to map changes in morphology and Young's modulus of basal and apical areas of the BM after cochlear implantation, using quantitative nanomechanical atomic force microscopy (QNM-AFM) after cochlear implant surgery. Cochlear implantation was undertaken in the guinea pig, and the BM was harvested at four time-points: 1 day, 14 days, 28 days and 84 days post-implantation for QNM-AFM analysis. Auditory brainstem response thresholds were determined prior to implantation and termination. BM tissue showed altered morphology and a progressive increase in Young's modulus, mainly in the apex, over time after implantation. BM tissue from the cochlear base demonstrated areas of extreme stiffness which are likely due to micro-calcification on the BM. In conclusion, stiffening of the BM after cochlear implantation occurs over time, even at sites far apical to a cochlear implant., (Copyright © 2019 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Experiments show that the extent of ongoing fibrotic change within the cochlea can be determined by the volume and pattern of bleeding within the first 24 h following cochlear implantation. Tissue-type plasminogen activator (tPA) is effective at reducing thrombus volume when administered both within and external to the systemic circulation., Aims/objectives: To determine if tPA delivered into the scala tympani immediately following implantation will reduce thrombus volume within the lower basal turn of the cochlea., Materials and Methods: Guinea pigs were implanted with either 'soft' or 'hard' arrays and administered tPA or saline via an intra-cochlear infusion immediately after implantation. Hearing was checked prior to, and 2 weeks after implantation. Cochleae were then harvested and imaged., Results: Animals implanted with 'soft' arrays had 4.2% less tissue response compared with animals implanted with 'hard' arrays. In animals receiving 'soft' arrays, tPA reduced the volume of tissue response (measured by the percentage of the lower basal turn of the scala tympani occupied by tissue response) compared with saline., Conclusions and Significance: tPA may be effective in reducing the overall volume of tissue response in routine 'soft' cochlear implantation and may have a greater effect in the event of significant surgical trauma.

Glucocorticoids have direct anti-inflammatory, anti-oxidant and anti-apoptotic effects on cochlear hair cells. Cochlear glucocorticoid therapy has gained particular attention for its ability to enhance the protection of residual hearing following hearing preservation cochlear implantation. Local drug delivery methods achieve high drug concentrations within the inner ear fluids but are reliant upon diffusion across the round window membrane. Diffusion has been shown to demonstrate large individual variability. This study explores the role of "adjuvant agents", which when administered with glucocorticoids, enhance inner ear absorption and distribution. Guinea pig cochleae were administered either dexamethasone alone or in combination with hyaluronic acid, histamine, or combination histamine and hyaluronic acid, targeted at the round window membrane. Control subjects received saline. Perilymph was sampled from the cochlear apex, and basal to apical dexamethasone concentrations recorded with mass spectroscopy. Cochleae were harvested, and immunohistochemistry employed to explore dexamethasone tissue penetration and distribution. Basal to apical gradients were observed along the scala tympani, with higher dexamethasone concentrations observed at the cochlear base. Gradients were more pronounced and uniform when administered on a hyaluronic acid sponge, while histamine increased absolute concentrations reaching the inner ear. Tissue penetration correlated with perilymph concentration. Our results demonstrate that adjuvant agents can be employed to enhance dexamethasone absorption and distribution in the inner ear, thus proposing therapeutic strategies that may enhance steroid facilitated hearing protection., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Local and systemically delivered glucocorticoids are commonly administered to protect the cochlea against damage associated with a variety of insults. There is reason to believe that dexamethasone administered by these routes may arrive at cochlear target sites via different pathways. Clinically, there is a lack of clarity as to which route is more effective in any specific circumstance. This study explores dexamethasone distribution within the guinea pig cochlea following local and systemic delivery methods. A combination of mass spectroscopy and immunohistochemistry were employed to compare both perilymph distribution, tissue uptake and receptor activation. Local administration of dexamethasone to the round window membrane resulted in greater perilymph concentrations, with a basal to apical gradient that favours the cochlear base. Tissue immunofluorescence was intimately related to perilymph concentration following local administration. Systemic administration resulted in much lower perilymph concentrations, with an inverse basal to apical gradient favouring the cochlear apex. Lower perilymph concentrations following systemic administration were associated with minimal tissue immunofluorescence. Despite this, GR activation of the SGNs was equivalent in both administration regimes. These results bring into question the efficacy of measuring perilymph concentrations alone as a surrogacy for dexamethasone distribution and activity in the cochlea, suggesting that the steroid ligand may arrive at its target receptor via alternative pathways. Our results suggest an equivalence in efficacy between local and systemic administration routes early after drug delivery, when the ultimate outcome of GR activation is the goal., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Sustained local delivery of drugs to the inner ear may be required for future regenerative and protective strategies. The round window is surgically accessible and a promising delivery route. To be viable, a delivery system should not cause hearing loss. This study determined the effect on hearing of placing a drug-delivery microcatheter on to the round window, and delivering either artificial perilymph (AP) or brain-derived neurotrophic factor (BDNF) via this catheter with a mini-osmotic pump. Auditory brainstem responses (ABRs) were monitored for 4 months after surgery, while the AP or BDNF was administered for the first month. The presence of the microcatheter - whether dry or when delivering AP or BDNF for 4 weeks - was associated with an increase in ABR thresholds of up to 15 dB, 16 weeks after implantation. This threshold shift was, in part, delayed by the delivery of BDNF. We conclude that the chronic presence of a microcatheter in the round window niche causes hearing loss, and that this is exacerbated by delivery of AP, and ameliorated temporarily by delivery of BDNF., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Cannabidiol (CBD) is a cannabinoid component of marijuana that has no significant activity at cannabinoid receptors or psychoactive effects. There is considerable interest in CBD as a therapy for epilepsy. Almost a third of epilepsy patients are not adequately controlled by clinically available anti-seizure drugs (ASDs). Initial studies appear to demonstrate that CBD preparations may be a useful treatment for pharmacoresistant epilepsy. The National Institute of Neurological Disorders and Stroke (NINDS) funded Epilepsy Therapy Screening Program (ETSP) investigated CBD in a battery of seizure models using a refocused screening protocol aimed at identifying pharmacotherapies to address the unmet need in pharmacoresistant epilepsy. Applying this new screening workflow, CBD was investigated in mouse 6 Hz 44 mA, maximal electroshock (MES), corneal kindling models and rat MES and lamotrigine-resistant amygdala kindling models. Following intraperitoneal (i.p.) pretreatment, CBD produced dose-dependent protection in the acute seizure models; mouse 6 Hz 44 mA (ED 50 164 mg/kg), mouse MES (ED 50 83.5 mg/kg) and rat MES (ED 50 88.9 mg/kg). In chronic models, CBD produced dose-dependent protection in the corneal kindled mouse (ED 50 119 mg/kg) but CBD (up to 300 mg/kg) was not protective in the lamotrigine-resistant amygdala kindled rat. Motor impairment assessed in conjunction with the acute seizure models showed that CBD exerted seizure protection at non-impairing doses. The ETSP investigation demonstrates that CBD exhibits anti-seizure properties in acute seizure models and the corneal kindled mouse. However, further preclinical and clinical studies are needed to determine the potential for CBD to address the unmet needs in pharmacoresistant epilepsy.

Cannabis enables and enhances the subjective sense of well-being by stimulating the endocannabinoid system (ECS), which plays a key role in modulating the response to stress, reward, and their interactions. However, over time, repeated activation of the ECS by cannabis can trigger neuroadaptations that may impair the sensitivity to stress and reward. This effect, in vulnerable individuals, can lead to addiction and other adverse consequences. The recent shift toward legalization of medical or recreational cannabis has renewed interest in investigating the physiological role of the ECS as well as the potential health effects, both adverse and beneficial, of cannabis. Here we review our current understanding of the ECS and its complex physiological roles. We discuss the implications of this understanding vis-á-vis the ECS's modulation of stress and reward and its relevance to mental disorders in which these processes are disrupted (i.e., addiction, depression, posttraumatic stress disorder, schizophrenia), along with the therapeutic potential of strategies to manipulate the ECS for these conditions.

Rationale: Accurate assessment of medication adherence is critical for determination of medication efficacy in clinical trials, but most current methods have significant limitations. This study tests a subtherapeutic (microdose) of acetazolamide as a medication ingestion marker because acetazolamide is rapidly absorbed and excreted without metabolism in urine and can be noninvasively sampled., Methods: In a double-blind, placebo-controlled, residential study, 10 volunteers received 15 mg oral acetazolamide for 4 consecutive days. Acetazolamide pharmacokinetics were assessed on day 3, and its pharmacokinetic and pharmacodynamic interactions with a model medication (30 mg oxycodone) were examined on day 4. The rate of acetazolamide elimination into urine was followed for several days after dosing cessation., Results: Erythrocyte sequestration (half-life = 50.2 ± 18.5 h, mean ± SD, n = 6), resulted in the acetazolamide microdose exhibiting a substantially longer plasma half-life (24.5 ± 5.6 hours, n = 10) than previously reported for therapeutic doses (3-6 hours). After cessation of dosing, the rate of urinary elimination decreased significantly (F3,23 = 247: P < 0.05, n = 6) in a predictable manner with low intersubject variability and a half-life of 16.1 ± 3.8 h (n = 10). For each of 4 consecutive mornings after dosing cessation, the rates of urinary acetazolamide elimination remained quantifiable.There was no overall effect of acetazolamide on the pharmacodynamics, Cmax, Tmax, or elimination half-life of the model medication tested. Acetazolamide may have modestly increased overall oxycodone exposure (20%, P < 0.05) compared with one of the 2 days when oxycodone was given alone, but there were no observed effects of acetazolamide on oxycodone pharmacodynamic responses., Conclusions: Coformulation of a once-daily trial medication with an acetazolamide microdose may allow estimation of the last time of medication consumption for up to 96 hours postdose. Inclusion of acetazolamide may therefore provide an inexpensive new method to improve estimates of medication adherence in clinical trials.

Effective strategies to monitor pharmacotherapy adherence are necessary, and sensitive biological markers are lacking. This study examined a subtherapeutic dose of quinine as a potential adherence tracer. Primary aims included examination of the plasma and urinary pharmacokinetic profile of once-daily quinine; secondary aims assessed pharmacokinetic/pharmacodynamic interactions with oxycodone (a CYP3A and CYP2D substrate). Healthy, nondependent opioid users (n = 9) were enrolled in this within-subject, double-blind, placebo-controlled inpatient study. Participants received the following oral doses: day 1, oxycodone (30 mg); days 2-4, quinine (80 mg); day 5, quinine and oxycodone (2 hours postquinine). Blood and 24-hour urine samples were collected throughout the study, and pharmacodynamic outcomes were assessed during experimental sessions (days 1, 4, 5). Quinine displayed a plasma Tmax ∼2 hours and t1/2 ∼10 hours. Oxycodone and noroxycodone parameters (Tmax , Cmax , t1/2 ) were similar with or without quinine present, although drug exposure (AUC) was slightly greater when combined with quinine. No pharmacodynamic interactions were detected, and doses were safely tolerated. During washout, quinine urinary concentrations steadily declined (elimination t1/2 ∼16 hours), with a 94% decrease observed 72 hours postdose. Overall, low-dose quinine appears to be a good candidate for a medication additive to monitor adherence for detection of missed medication., (© 2015, The American College of Clinical Pharmacology.)

Aim: To explore morphological or electrophysiological evidence for the presence of endolymphatic hydrops (EH) in guinea pig cochleae in the first 3 months after cochlear implantation., Methods: Dummy silastic electrodes were implanted atraumatically into the basal turn of scala tympani via a cochleostomy. Round window electrocochleography (ECochG) was undertaken prior to and after implantation. Animals survived for 1, 7, 28 or 72 days prior to a terminal experiment, when ECochG was repeated. The cochleae were imaged using micro-CT after post-fixing with osmium tetroxide to reveal the inner ear soft tissue structure. EH was assessed by visual inspection at a series of frequency specific places along the length of the cochlea, and the extent to which Reissner's membrane departed from its neutral position was quantified. Tissue response volumes were calculated. Using ECochG, the ratio of the summating potential to the action potential (SP/AP ratio) was calculated in response to frequencies between 2 and 32 kHz., Results: There was minimal evidence of electrode trauma from cochlear implantation on micro-CT imaging. Tissue response volumes did not change over time. EH was most prevalent 7 days after surgery in implanted ears, as determined by visual inspection. Scala media areas were increased, as expected in cases of EH, over the first month after cochlear implantation. SP/AP ratios decreased immediately after surgery, but were elevated 1 and 7 days after implantation., Conclusions: EH is prevalent in the first weeks after implant surgery, even in the absence of significant electrode insertion trauma., (Copyright © 2015 Elsevier B.V. All rights reserved.)

12-hydroxyeicosatetraenoic acid (12-HETE) is a neuromodulator that is synthesized during ischemia. Its neuronal effects include attenuation of calcium influx and glutamate release as well as inhibition of AMPA receptor (AMPA-R) activation. Because 12-HETE reduces ischemic injury in the heart, we examined whether it can also reduce neuronal excitotoxicity. When treated with 12-(S)HETE, cortical neuron cultures subjected to AMPA-R-mediated glutamate toxicity suffered up to 40% less damage than untreated cultures. The protective effect of 12-(S)HETE was concentration-dependent (EC50 = 88 nm) and stereostructurally selective. Maximal protection was conferred by 300 nm 12-(S)HETE; 300 nm 15-(S)HETE was similarly protective, but 300 nm 5-(S)HETE was less effective. The chiral isomer 12-(R)HETE offered no protection; neither did arachidonic acid or 12-(S)hydroperoxyeicosatetraenoic acid. Excitotoxicity was calcium-dependent, and 12-(S)HETE was demonstrated to protect by inactivating N and L (but not P) calcium channels via a pertussis toxin-sensitive mechanism. Calcium imaging demonstrated that 12-(S)HETE also attenuates glutamate-induced calcium influx into neurons via a pertussis toxin-sensitive mechanism, suggesting that it acts via a G-protein-coupled receptor. In addition, 12-(S)HETE stimulates GTPgammaS binding (indicating G-protein activation) and inhibits adenylate cyclase in forskolin-stimulated cultures over the same concentration range as it exerts its anti-excitotoxic and calcium-influx attenuating effects. These studies demonstrate that 12-(S)HETE can protect neurons from excitotoxicity by activating a G(i/o)-protein-coupled receptor, which limits calcium influx through voltage-gated channels.

Cannabinoid receptor activation in vivo reduces ischemic injury, a phenomenon that has not been successfully reproduced in vitro. Because cyclic adenosine monophosphate (cAMP) levels are radically elevated during ischemic reperfusion, but cannabinoid receptor activation reduces cAMP levels, we hypothesized that cannabinoids might prevent in vitro glutamate toxicity if reperfusion was simulated by cAMP supplementation after glutamate removal. Although neuronal cultures were unaffected by the single addition of either cannabinoid or dibutyryl cAMP (dbcAMP), glutamate toxicity was reduced by 20% when cannabinoid was present during glutamate exposure and either dbcAMP or forskolin was added after glutamate removal. Further studies revealed that cannabinoid receptor activation reduces glutamate toxicity by attenuating calcium influx through N- and P/Q-type calcium channels. The effect of glutamate exposure on neuronal cAMP levels was also examined. Glutamate exposure significantly reduced neuronal cAMP levels, although suppression was even greater when cannabinoid was present. Because neurological outcome after ischemia is poor when cAMP levels during reperfusion are low, it is hypothesized that cAMP elevation after glutamate exposure may offset excitotoxic and/or cannabinoid receptor-induced cAMP depletion. Cannabinoids protect against ischemic injury in vivo, but only reduce toxicity in vitro when cAMP levels are elevated, possibly suggesting that cAMP elevation during reperfusion reduces brain injury by off-setting the effect of Gi/o protein-coupled systems on adenylate cyclase.

Cannabidiol and other cannabinoids were examined as neuroprotectants in rat cortical neuron cultures exposed to toxic levels of the neurotransmitter, glutamate. The psychotropic cannabinoid receptor agonist delta 9-tetrahydrocannabinol (THC) and cannabidiol, (a non-psychoactive constituent of marijuana), both reduced NMDA, AMPA and kainate receptor mediated neurotoxicities. Neuroprotection was not affected by cannabinoid receptor antagonist, indicating a (cannabinoid) receptor-independent mechanism of action. Glutamate toxicity can be reduced by antioxidants. Using cyclic voltametry and a fenton reaction based system, it was demonstrated that Cannabidiol, THC and other cannabinoids are potent antioxidants. As evidence that cannabinoids can act as an antioxidants in neuronal cultures, cannabidiol was demonstrated to reduce hydroperoxide toxicity in neurons. In a head to head trial of the abilities of various antioxidants to prevent glutamate toxicity, cannabidiol was superior to both alpha-tocopherol and ascorbate in protective capacity. Recent preliminary studies in a rat model of focal cerebral ischemia suggest that cannabidiol may be at least as effective in vivo as seen in these in vitro studies.

The neuroprotective actions of cannabidiol and other cannabinoids were examined in rat cortical neuron cultures exposed to toxic levels of the excitatory neurotransmitter glutamate. Glutamate toxicity was reduced by both cannabidiol, a nonpsychoactive constituent of marijuana, and the psychotropic cannabinoid (-)Delta9-tetrahydrocannabinol (THC). Cannabinoids protected equally well against neurotoxicity mediated by N-methyl-D-aspartate receptors, 2-amino-3-(4-butyl-3-hydroxyisoxazol-5-yl)propionic acid receptors, or kainate receptors. N-methyl-D-aspartate receptor-induced toxicity has been shown to be calcium dependent; this study demonstrates that 2-amino-3-(4-butyl-3-hydroxyisoxazol-5-yl)propionic acid/kainate receptor-type neurotoxicity is also calcium-dependent, partly mediated by voltage sensitive calcium channels. The neuroprotection observed with cannabidiol and THC was unaffected by cannabinoid receptor antagonist, indicating it to be cannabinoid receptor independent. Previous studies have shown that glutamate toxicity may be prevented by antioxidants. Cannabidiol, THC and several synthetic cannabinoids all were demonstrated to be antioxidants by cyclic voltametry. Cannabidiol and THC also were shown to prevent hydroperoxide-induced oxidative damage as well as or better than other antioxidants in a chemical (Fenton reaction) system and neuronal cultures. Cannabidiol was more protective against glutamate neurotoxicity than either ascorbate or alpha-tocopherol, indicating it to be a potent antioxidant. These data also suggest that the naturally occurring, nonpsychotropic cannabinoid, cannabidiol, may be a potentially useful therapeutic agent for the treatment of oxidative neurological disorders such as cerebral ischemia.

Unlabelled: A naturally occurring brain lipid, cis-9,10-octadeceamide--oleamide (OA), is found in increased concentrations in the cerebrospinal fluid of sleep-deprived cats, which suggests that it may be an endogenous sleep-inducing substance. We studied the effects of this fatty-acid derivative on the function of cloned gamma-aminobutyric acid (GABA(A)) receptors expressed in Xenopus oocytes. Oocytes were injected with cRNA synthesized in vitro to express simple GABA(A) receptors (alpha1beta1, alpha3beta1, alpha5beta1, and alpha1beta2 subunit combinations) and receptors in which the GABA-induced chloride currents were potentiated in the presence of benzodiazepines (alpha1beta1gamma2s and alpha1beta2gamma2s subunit combinations). OA only produced significant potentiation of the peak Cl- current when applied with GABA to benzodiazepine-sensitive GABA(A) receptors. The peak currents of the simple GABA(A) receptors in the presence of OA were either unaffected or slightly inhibited by OA, but the overall mean currents were not significantly altered. Oleic acid was also capable of potentiating benzodiazepine-sensitive GABA(A) receptor function. The function of other ligand-gated ion channels, such as the N-methyl-D-aspartate receptor (NR1 + NR2A or 2C) and the 5-HT3 receptor expressed in Xenopus oocytes, were unaffected by OA. Sprague-Dawley rats receiving intraperitoneal injections of oleamide (10, 20, or 100 mg/kg) showed no change in the minimum alveolar anesthetic concentration (MAC) of desflurane required to abolish movement in response to noxious (tail clamp) stimulation (control MAC 6.48% +/- 1.28% atm; 100 mg/kg OA MAC 7.05% +/- 0.42% atm). These results reinforce the view that oleyl compounds may be natural modulators of inhibitory ion channel function, but that these effects contribute little to the central nervous system depression produced by volatile anesthetics as measured by MAC., Implications: The putative sleep-inducing substance, oleamide, potentiates benzodiazepine-sensitive gamma-aminobutyric acid receptor function but does not alter desflurane minimum alveolar anesthetic concentration in rats.

Anandamide is an endogenous ligand of cannabinoid receptors that induces pharmacological responses in animals similar to those of cannabinoids such as delta9-tetrahydrocannabinol (THC). Typical pharmacological effects of cannabinoids include disruption of pain, memory formation, and motor coordination, systems that all depend on NMDA receptor mediated neurotransmission. We investigated whether anandamide can influence NMDA receptor activity by examining NMDA-induced calcium flux (deltaCa2+NMDA) in rat brain slices. The presence of anandamide reduced deltaCa2+NMDA and the inhibition was disrupted by cannabinoid receptor antagonist, pertussis toxin treatment, and agatoxin (a calcium channel inhibitor). Whereas these treatments prevented anandamide inhibiting deltaCa2+NMDA, they also revealed another, underlying mechanism by which anandamide influences deltaCa2+NMDA. In the presence of cannabinoid receptor antagonist, anandamide potentiated deltaCa2+NMDA in cortical, cerebellar, and hippocampal slices. Anandamide (but not THC) also augmented NMDA-stimulated currents in Xenopus oocytes expressing cloned NMDA receptors, suggesting a capacity to directly modulate NMDA receptor activity. In a similar manner, anandamide enhanced neurotransmission across NMDA receptor-dependent synapses in hippocampus in a manner that was not mimicked by THC and was unaffected by cannabinoid receptor antagonist. These data demonstrate that anandamide can modulate NMDA receptor activity in addition to its role as a cannabinoid receptor ligand.

Anandamide (arachidonyl ethanolamide) is a compound that was identified from porcine brain lipids by its ability to bind to the brain cannabinoid receptor. This study assessed anandamide as a substrate for a brain lipoxygenase and characterised the brain metabolite 12-hydroxyanandamide. Anandamide was also compared with arachidonic acid as a lipoxygenase substrate by examining enzyme kinetics in the presence of either of the two compounds. In addition, a non-mammalian enzyme was used to generate 11- and 15-hydroxy-anandamide in order to compare the cannabinomimetic properties of a range of anandamide derivatives. A ligand displacement assay indicated a large variation in the affinity of anandamide metabolites for the brain cannabinoid receptor. The brain metabolite, 12-hydroxyanandamide had an affinity twice that of anandamide, although the 11- and 15- hydroxy-metabolites were considerably poorer ligands of this receptor. Consistent with the receptor binding data, 12-hydroxyanandamide (unlike 15-hydroxyanandamide) inhibited forskolin-stimulated cAMP synthesis, indicating it to be a functional agonist at the brain cannabinoid receptor. Pharmacological studies of the capacity of anandamide and its metabolites to inhibit the murine vas deferens twitch response indicated the 12-hydroxy-metabolite to be less active than the parent compound, but a better cannabinomimetic than 15-hydroxyanandamide.

Blood cells from the crab, Carcinus maenas, stimulated with calcium ionophore A23187, in the presence of exogenous fatty acid, produced cyclooxygenase, lipoxygenase and monooxygenase derivatives of eicosatetraenoic (20:4(n - 6)) and eicosapentaenoic (20:5(n - 3)) acids. Isolation, identification and quantification of these products was achieved using chiral and reverse phase-high performance liquid chromatography, gas-chromatography, radioimmunoassay and gas chromatography-mass spectrometry. The principle metabolites observed were 8-hydroxy fatty acids and 'E' series prostaglandins. Smaller amounts of thromboxane B2, 6-keto-prostaglandin F1 alpha and 5-, 9-, 11-, 12- and 15-hydroxy-eicosatetraenoic acids were also synthesised. Lipoxygenase, cyclooxygenase and cytochrome P-450 inhibitors were used to investigate the mode of product formation. Mixtures of hydroxy-fatty acid enantiomers were produced and the dominant chiral form varied with the position of the hydroxyl group. No leukotrienes or lipoxins were detected.