Search

Multiple-dose pharmacokinetics (PK) and safety were investigated in this phase 1 study of PF-06372865, a positive allosteric modulator of α2/3/5 subunit-containing γ-aminobutyric acid A receptors (NCT03351751). In 2 cohorts (7-8 PF-06372865 and 2 placebo in each cohort), healthy adult subjects received twice-daily oral doses of PF-06372865 for 21 days, which included titration in the first 7 days, followed by a maintenance dose of 25 mg twice daily (Cohort 1) and 42.5 mg twice daily (Cohort 2) for 14 days. Serial PK samples were collected on days 1 and 21. Nineteen subjects were assigned to study treatments; 18 completed the study. Approximate dose-proportional increases in maximum plasma concentratin and area under the plasma concentration-time curve over the dosing interval were observed. PF-06372865 was rapidly absorbed with a median time to maximum concentration of 1 to 2 hours following both single- and multiple-dose administration. Mean terminal elimination half-life on day 21 was approximately 11 hours in both cohorts. All adverse events were mild; the most frequently reported was dizziness. After titration, there were no reports of somnolence. There were no clinically significant safety findings, including a lack of withdrawal symptoms on discontinuation of treatment. These results demonstrate that PF-06372865 is safe and well tolerated at doses estimated to achieve high receptor occupancy (>80%), a profile differentiated from nonselective benzodiazepines., (© 2021 Cerevel Therapeutics. Clinical Pharmacology in Drug Development published by Wiley Periodicals LLC on behalf of American College of Clinical Pharmacology.)

For radioligands without a reference region, the Lassen plot can be used to estimate receptor occupancy by an exogenous drug ( O Drug ). However, the Lassen plot is not well-suited for spatial variation in O Drug . To overcome this limitation, we introduce a Lassen plot filter, i.e. a Lassen plot applied to local neighborhoods in PET images. Image data were simulated with regional variation in V N D , O Drug , both, or neither and analyzed using the change in binding potential ( Δ B P N D ), the conventional Lassen plot, and the Lassen plot filter at the region of interest (ROI) and voxel level. All methods were also applied to a human [ 11 C]flumazenil occupancy study using PF-06372865. This combination of a non-selective radioligand and selective drug should lead to varying O Drug   provided the distribution of subtypes varies spatially. In contrast with Δ B P N D and the conventional Lassen plot, ROI-level and voxel-level Lassen plot filter estimates remained unbiased in the presence of regional variation in V N D or O Drug . In the [ 11 C]flumazenil data-set, O Drug was shown to vary regionally in accordance with the distribution of binding sites for [ 11 C]flumazenil and PF-06372865. We demonstrate that a local-neighborhood Lassen plot filter provides robust and unbiased estimates of O Drug and V N D without the need for any user intervention.

Rationale: Previous research suggests that sleep polysomnography and EEG endpoints can be used to assess GABAergic activity; however, the impact of GABA B receptor positive allosteric modulators on sleep endpoints remains unclear., Objectives: This phase 1 study compared a single dose of ASP8062 (35 mg or 70 mg), a GABA B receptor positive allosteric modulator, with placebo and paroxetine (40 mg)., Methods: Healthy adult volunteers were randomized to four treatments (35 mg ASP8062, 70 mg ASP8062, paroxetine 40 mg, or matching placebo), each separated by a 14-day washout. Primary endpoints obtained by polysomnography were time in stage N3 or SWS and time in rapid eye movement (REM) sleep. Secondary endpoints included impact on sleep stages and electroencephalography parameters, pharmacokinetics, nighttime growth hormone (GH), and safety/tolerability., Results: In 20 randomized volunteers, ASP8062 led to a significant and seemingly dose-dependent increase in SWS over the entire night; this increase was mainly observed during the first third of the night. ASP8062 did not impact time in REM sleep. Paroxetine had no effect on SWS but produced a significant reduction in time spent in REM sleep. A dose-dependent trend in increased GH release was also observed with ASP8062. Headache and nausea were the most commonly reported treatment-emergent adverse events (TEAEs) for ASP8062; most TEAEs were mild in severity., Conclusions: Single-dose ASP8062 (35 and 70 mg) appeared to result in CNS penetration and enhanced GABAergic activity as measured by increases in slow-wave sleep and growth hormone release.

The use of hybrid PET/MR imaging facilitates the simultaneous investigation of challenge-related changes in ligand binding to neuroreceptors using PET, while concurrently measuring neuroactivation or blood flow with MRI. Having attained a steady state of the PET radiotracer using a bolus-infusion protocol, it is possible to observe alterations in ligand neuroreceptor binding through changes in distribution volumes. Here, we present an iterative procedure for establishing an administration scheme to obtain steady state [ 11 C]flumazenil concentrations in grey matter in the human brain. In order to achieve a steady state in the shortest possible time, the bolus infusion ratio from a previous examination was adapted to fit the subsequent examination. 17 male volunteers were included in the study. Boli and infusions with different weightings were given to the subjects and were characterised by kbol values from 74 ​min down to 42 ​min. Metabolite analysis was used to ascertain the value of unmetabolised flumazenil in the plasma, and PET imaging was used to assess its binding in the grey matter. The flumazenil time-activity curves (TACs) in the brain were decomposed into activity contributions from pure grey and white matter and analysed for 12 ​vol of interest (VOIs). The curves highlighted a large variability in metabolic rates between the subjects, with kbol ​= ​54.3 ​min being a reliable value to provide flumazenil equilibrium conditions in the majority of the VOIs and cases. The distribution volume of flumazenil in all 12 VOIs was determined., Competing Interests: Declaration of competing interest There are no known conflicts of interest., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Psychosis and dyskinesia significantly diminish the quality of life of patients with advanced Parkinson's disease (PD). Available treatment options are unfortunately few and their use is limited by adverse effects. We have recently shown that activation of metabotropic glutamate 2 and 3 (mGlu2/3) receptors produced significant relief of L-3,4-dihydroxyphenylalanine (L-DOPA)-induced psychosis-like behaviours (PLBs) and dyskinesia in experimental models of PD. Here, using the highly-selective mGlu2 positive allosteric modulator (PAM) LY-487,379, we seek to determine the contribution of selective mGlu2 activation on both L-DOPA-induced PLBs and dyskinesia, in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned primate. We first determined the pharmacokinetic (PK) profile of LY-487,379 in the common marmoset, following which we administered it (0.1, 1 and 10 mg/kg) or its vehicle to 6 MPTP-lesioned marmosets previously exposed to L-DOPA to elicit stable PLBs and dyskinesia. We found that LY-487,379 provided a ≈45% reduction of the global PLBs observed and reduced global dyskinesia score by ≈ 55%. Moreover, LY-487,379 enhanced the anti-parkinsonian effect of L-DOPA, by reducing global parkinsonian score by ≈ 15%. Our data suggest that selective mGlu2 positive allosteric modulation with LY-487,379 may represent a potential therapeutic approach to alleviate both L-DOPA-induced PLBs and dyskinesia in PD., (Copyright © 2020 Elsevier B.V. All rights reserved.)

ASP8062 is an orally active γ-amino-butyric acid type B (GABA B ) receptor positive allosteric modulator currently in phase 2 development. Safety and pharmacokinetic (PK) profiles of ASP8062 were evaluated in 2 studies in healthy subjects. The first study (a first-in-human study) evaluated single ascending doses (SAD) of ASP8062. The second study was composed of 2 parts: part 1 evaluated multiple ascending doses (MAD) of ASP8062 for 14 days, and part 2 was a single-dose arm to assess the PK of ASP8062 in cerebrospinal fluid (CSF). Fifty-six men (SAD) and 56 subjects (24 women and 32 men; MAD) were enrolled. Across the SAD dosing range, area under the concentration-time curve was dose proportional; increases in maximum plasma concentration appeared linear but were slightly less than dose proportional. Time to maximal concentration and half-life were 1-4 hours and ∼40-50 hours, respectively; no food effect was observed. ASP8062 PK properties at steady state were similar to those following a single dose. Steady state was achieved by ∼day 9 with ∼2-fold accumulation, and ASP8062 was detected in CSF. ASP8062 was well tolerated; no clear evidence of ASP8062's effects on safety, cognition, drug withdrawal, or suicidal ideation/behavior was observed. These data support the development of ASP8062 in indications where the GABA B receptor is a target., (© 2020, The American College of Clinical Pharmacology.)

Hydrogen sulfide (H 2 S) is a colorless, highly neurotoxic gas. It is not only an occupational and environmental hazard but also of concern to the Department of Homeland Security for potential nefarious use. Acute high-dose H 2 S exposure causes death, while survivors may develop neurological sequelae. Currently, there is no suitable antidote for treatment of acute H 2 S-induced neurotoxicity. Midazolam (MDZ), an anti-convulsant drug recommended for treatment of nerve agent intoxications, could also be of value in treating acute H 2 S intoxication. In this study, we tested the hypothesis that MDZ is effective in preventing/treating acute H 2 S-induced neurotoxicity. This proof-of-concept study had two objectives: to determine whether MDZ prevents/reduces H 2 S-induced mortality and to test whether MDZ prevents H 2 S-induced neurological sequelae. MDZ (4 mg/kg) was administered IM in mice, 5 min pre-exposure to a high concentration of H 2 S at 1000 ppm or 12 min post-exposure to 1000 ppm H 2 S followed by 30 min of continuous exposure. A separate experiment tested whether MDZ pre-treatment prevented neurological sequelae. Endpoints monitored included assessment of clinical signs, mortality, behavioral changes, and brain histopathological changes. MDZ significantly reduced H 2 S-induced lethality, seizures, knockdown, and behavioral deficits (p < 0.01). MDZ also significantly prevented H 2 S-induced neurological sequelae, including weight loss, behavior deficits, neuroinflammation, and histopathologic lesions (p < 0.01). Overall, our findings show that MDZ is a promising drug for reducing H 2 S-induced acute mortality, neurotoxicity, and neurological sequelae.

Medication during pregnancy is often a necessity for women to treat their acute or chronic diseases. The goal of this study is to evaluate the potential of micelle-like nanoparticles (MNP) for providing safe drug usage in pregnancy and protect both foetus and mother from medication side effects. Clonazepam-loaded MNP were prepared from copolymers [polystyrene-poly(acrylic acid) (PS-PAA), poly(ethylene glycol)-b-poly(lactic acid) (PEG-PLA) and distearyl-sn-glycero-3-phosphoethanolamine-N-[methoxy-poly(ethylene glycol) (PEG-DSPE)] with varying monomer ratios and their drug-loading efficiency, drug release ratio, particle size, surface charge and morphology were characterised. The cellular transport and cytotoxicity experiments were conducted on clonazepam and MNP formulations using placenta-choriocarcinoma-BeWo and brain-endothelial-bEnd3 cells. Clonazepam-loaded PEG 5000 -PLA 4500 MNP reduced the drug transport through BeWo cells demonstrating that MNP may lower foetal drug exposure, thus reduce the drug side effects. However, lipofectamine modified MNP improved the transport of clonazepam and found to be promising for brain and in-utero-specific drug treatment.

Chronic kidney disease (CKD), which affects, not only renal clearance, but also non-renal clearance, is accompanied by a decline in renal function. Although it has been suggested that humoral factors, such as uremic toxins that accumulate in the body under CKD conditions, could be involved in the changes associated with non-renal drug clearance, the overall process is not completely understood. In this study, we report on the role of parathyroid hormone (PTH), a middle molecule uremic toxin, on the expression of drug metabolizing or transporting proteins using rats with secondary hyperparathyroidism (SHPT) as models. In SHPT rats, hepatic and intestinal CYP3A expression was suppressed, but the changes were recovered by the administration of the calcimimetic cinacalcet, a PTH suppressor. Under the same experimental conditions, a pharmacokinetic study using orally administered midazolam, a substrate for CYP3A, showed that the AUC was increased by 5 times in SHPT rats, but that was partially recovered by a cinacalcet treatment. This was directly tested in rat primary hepatocytes and intestinal Caco-2 cells where the expression of the CYP3A protein was down-regulated by PTH (1-34). In Caco-2 cells, PTH (1-34) down-regulated the expression of CYP3A mRNA, but an inactive PTH derivative (13-34) had no effect. 8-Bromo-cyclic adenosine monophosphate, a membrane-permeable cAMP analog, reduced mRNA expression of CYP3A whereas the inhibitors of PI3K, NF-κB, PKC and PKA reversed the PTH-induced CYP3A down-regulation. These results suggest that PTH down-regulates CYP3A through multiple signaling pathways, including the PI3K/PKC/PKA/NF-κB pathway after the elevation of intracellular cAMP, and the effect of PTH can be prevented by cinacalcet treatment., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Objective: To investigate GABA-ergic receptor density and associated brain functional and grey matter changes in focal hand dystonia (FHD)., Methods: 18 patients with FHD of the right hand and 18 age and gender matched healthy volunteers (HV) participated in this study. We measured the density of GABA-A receptors using [ 11 C] Flumazenil and perfusion using [ 15 O] H 2 O. Anatomical images were also used to measure grey matter volume with voxel-based morphometry (VBM)., Results: In FHD patients compared to HV, the vermis VI of the right cerebellum and the left sensorimotor cortex had a decrease of Flumazenil binding potential (FMZ-BP), whereas the striatum and the lateral cerebellum did not show significant change. Bilateral inferior prefrontal cortex had increased FMZ-BP and an increase of perfusion, which correlated negatively with disease duration. Only the left sensorimotor cortex showed a decrease of grey matter volume., Interpretation: Impairments of GABAergic neurotransmission in the cerebellum and the sensorimotor cortical areas could explain different aspects of loss of inhibitory control in FHD, the former being involved in maladaptive plasticity, the latter in surround inhibition. Reorganization of the inferior prefrontal cortices, part of the associative network, might be compensatory for the loss of inhibitory control in sensorimotor circuits. These findings suggest that cerebellar and cerebral GABAergic abnormalities could play a role in the functional imbalance of striato-cerebello-cortical loops in dystonia.

Objectives: Toxicodynetics aims at defining the time-course of major clinical events in drug overdose. We report the toxicodynetics in mono-intoxications with oxazepam and nordiazepam., Methods: Cases of oxazepam or nordiazepam overdoses collected at the Paris poison control centre from 1999 to 2014 on the basis of self-report. A particular attention was paid to eliminate the concomitant alcohol or psychotropic co-ingestions. The toxicodynetic parameters were assessed as previously described. Results are expressed using 10-90 percentiles. In adults, the dose was normalized (TI, toxic Index) by dividing the supposed ingested dose by the maximal recommended dose., Results: Two hundred and fifty-one and 74 cases of oxazepam and nordiazepam poisonings were included, respectively. The Emax for oxazepam and nordiazepam were sleepiness or obtundation in 106 and 36 cases, respectively. Coma was used to qualify only one oxazepam overdose. The median delay in onset of the Emax was 1.5h (0.33-15) in nordiazepam and 4h (0.5-15) in oxazepam overdose. In both overdoses, the onset of Emax occurred on an "on-off" mode. In adults, the greatest TIs in nordiazepam and oxazepam overdoses were 45 and 26.7, respectively. The TI in the oxazepam-induced coma was 26.7, the largest dose., Conclusion: Data collected in PCC allow determining a number of toxicodynetic parameters. Toxicodynetics showed that nordiazepam is not a cause of coma even in large overdose while oxazepam causes coma only at a very high dose. Deep coma in nordiazepam overdose whatever the dose and deep coma in overdose with oxazepam involving TI less than 20 result from unrecognized drug-drug interaction., (Copyright © 2017. Published by Elsevier Masson SAS.)

Background: Epilepsy and depression are two of the common diseases seriously threatening life and health of human. A shared neurobiological substrate led to the bidirectional relationship and high comorbid occurrence of the two disorders. Recently, an increasing number of patients with epilepsy (PWE) require some form of antidepressant medication. However, most of the available antidepressants are inadequate for PWE for some reasons. So, the search for novel and increasingly effective drugs with anticonvulsant and antidepressant activities is necessary., Methods: A series of 2-substituted-6-(4H-1,2,4-triazol-4-yl)benzo[d]oxazoles (5a-p) were designed and synthesized. Their anticonvulsant activities were evaluated using maximal electroshock shock (MES) and subcutaneous pentylenetetrazole (scPTZ) seizure models in mice. Their antidepressant activities were screened with the forced swimming test (FST)., Results: All the compounds showed anti-MES activities in different degree, among which 5g and 5j were the most promising one with ED50 value of 31.7 and 12.7 mg/kg, respectively. What's more, 5g and 5j also exhibited nice anti-scPTZ activities and low neurotoxicity. Interestingly, these compounds also showed good antidepressant activities in FST. And the efficacy of 5g were also confirmed by a tail suspension test and a open field test. The pretreatment of thiosemicarbazide (an inhibitor of γ- aminobutyric acid synthesis enzyme) significantly increased the ED50 of 5g in MES and reversed the reductions in the immobility time of 5g in FST., Conclusion: Triazole-containing benzo[d]oxazole is a good skeleton to develop compounds with both anticonvulsant and antidepressant activities. We have got the compound 5g, which display remarkable antidepressant and anticonvulsant activities, and the GABAergic system was involved in the action mechanism of 5g., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Studies in rodents suggest that flumazenil is a P-glycoprotein substrate at the blood-brain barrier. This study aimed to assess whether [ 11 C]flumazenil is a P-glycoprotein substrate in humans and to what extent increased P-glycoprotein function in epilepsy may confound interpretation of clinical [ 11 C]flumazenil studies used to assess gamma-aminobutyric acid A receptors. Nine drug-resistant patients with epilepsy and mesial temporal sclerosis were scanned twice using [ 11 C]flumazenil before and after partial P-glycoprotein blockade with tariquidar. Volume of distribution, nondisplaceable binding potential, and the ratio of rate constants of [ 11 C]flumazenil transport across the blood-brain barrier (K 1 /k 2 ) were derived for whole brain and several regions. All parameters were compared between pre- and post-tariquidar scans. Regional results were compared between mesial temporal sclerosis and contralateral sides. Tariquidar significantly increased global K 1 /k 2 (+23%) and volume of distribution (+10%), but not nondisplaceable binding potential. At the mesial temporal sclerosis side volume of distribution and nondisplaceable binding potential were lower in hippocampus (both ∼-19%) and amygdala (both ∼-16%), but K 1 /k 2 did not differ, suggesting that only regional gamma-aminobutyric acid A receptor density is altered in epilepsy. In conclusion, although [ 11 C]flumazenil appears to be a (weak) P-glycoprotein substrate in humans, this does not seem to affect its role as a tracer for assessing gamma-aminobutyric acid A receptor density., (© The Author(s) 2015.)

Objective: To provide a comprehensive investigation of the γ-aminobutyric acid (GABA) system in patients with neurofibromatosis type 1 (NF1) that allows understanding the nature of the GABA imbalance in humans at pre- and postsynaptic levels., Methods: In this cross-sectional study, we employed multimodal imaging and spectroscopy measures to investigate GABA type A (GABAA) receptor binding, using [(11)C]-flumazenil PET, and GABA concentration, using magnetic resonance spectroscopy (MRS). Fourteen adult patients with NF1 and 13 matched controls were included in the study. MRS was performed in the occipital cortex and in a frontal region centered in the functionally localized frontal eye fields. PET and MRS acquisitions were performed in the same day., Results: Patients with NF1 have reduced concentration of GABA+ in the occipital cortex (p = 0.004) and frontal eye fields (p = 0.026). PET results showed decreased binding of GABAA receptors in patients in the parieto-occipital cortex, midbrain, and thalamus, which are not explained by decreased gray matter levels., Conclusions: Abnormalities in the GABA system in NF1 involve both GABA concentration and GABAA receptor density suggestive of neurodevelopmental synaptopathy with both pre- and postsynaptic involvement., (© 2016 American Academy of Neurology.)

Hepatocellular carcinoma (HCC) accompanied by severe liver dysfunction is a serious disease, which results in altered hepatic clearance. Generally, maintenance doses depend upon drug clearance, so individual dosage regimens should be customized for HCC patients based on the condition of patients. Based on clearance of CYP isoform-specific substrates at the microsomal level (CLM), microsomal protein per gram of liver (MPPGL), liver weight, hepatic blood flow, hepatic clearance values (CLH) for 10 CYPs in HCC patients (n=102) were extrapolated using a predictive bottom-up pharmacokinetic model. Compared with controls, the CLM values for CYP2C9, 2D6, 2E1 were significantly increased in HCC patients. Additionally, CYP1A2, 2C8, 2C19 CLM values decreased while the values for CYP2A6, 2B6, 3A4/5 were unchanged. The MPPGL values in HCC tissues were significantly reduced. CLH values of HCC patients for CYP1A2, 2A6, 2B6, 2C8, 2C19, and 3A4/5 were significantly reduced, while this for CYP2E1 were markedly increased and those for CYP2C9 and 2D6 did not change. Moreover, disease (fibrosis and cirrhosis) and polymorphisms of the CYP genes have influenced the CLH for some CYPs. Prediction of the effects of HCC on drug clearance may be helpful for the design of clinical studies and the clinical management of drugs in HCC patients., Competing Interests: The authors who have taken part in this study declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.

Knowledge about the neural underpinnings of the negative blood oxygen level dependent (BOLD) responses in functional magnetic resonance imaging (fMRI) is still limited. We hypothesized that pharmacological GABAergic modulation attenuates BOLD responses, and that blood concentrations of a positive allosteric modulator of GABA correlate inversely with BOLD responses in the cingulate cortex. We investigated whether or not pure task-related negative BOLD responses were co-localized with pharmacologically modulated BOLD responses. Twenty healthy adults received either 5 mg diazepam or placebo in a double blind, randomized design. During fMRI the subjects performed a working memory task. Results showed that BOLD responses in the cingulate cortex were inversely correlated with diazepam blood concentrations; that is, the higher the blood diazepam concentration, the lower the BOLD response. This inverse correlation was most pronounced in the pregenual anterior cingulate cortex and the anterior mid-cingulate cortex. For subjects with diazepam plasma concentration > 0.1 mg/L we observed negative BOLD responses with respect to fixation baseline. There was minor overlap between cingulate regions with task-related negative BOLD responses and regions where the BOLD responses were inversely correlated with diazepam concentration. We interpret that the inverse correlation between the BOLD response and diazepam was caused by GABA-related neural inhibition. Thus, this study supports the hypothesis that GABA attenuates BOLD responses in fMRI. The minimal overlap between task-related negative BOLD responses and responses attenuated by diazepam suggests that these responses might be caused by different mechanisms.

Neurotoxic effects of compounds can be tested in vitro using cell systems. One example is aggregating rat brain cell cultures. For the extrapolation of in vitro data to the in vivo situation, it is important to take the biokinetics of the test compound into account. In addition, the exposure in vivo is often for a longer period of time; therefore, it is crucial to incorporate this into in vitro assays as well. In this study, aggregating rat brain cell cultures were exposed to chlorpromazine (CPZ) and diazepam (DZP) for 12-days with repeated exposure. Samples were taken from the stocks, test media, cell culture media and cells at specific time points on the first and last exposure day. These samples were analysed by HPLC-UV. The amount of CPZ in the medium decreased over time, whereas the amount in the cells showed an increase. Accumulation of CPZ in the cells was seen over the 12-day repeated exposure. The amount of DZP in the medium remained stable over time and only up to 2% of DZP added was found in the cells. Different biokinetic behaviour was found for CPZ and DZP. Possible explanations are differences in uptake into the cells or efflux out of the cells. The decrease of CPZ in the medium versus the stable amount of DZP results in differences in exposure concentrations over time, which should be taken into account when interpreting in vitro effect data., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The therapeutic potential of selective mGlu1 activation is vastly unexplored relative to the other group I mGlu receptor, mGlu5; therefore, our lab has focused considerable effort toward developing mGlu1 positive allosteric modulators (PAMs) suitable as in vivo proof of concept tool compounds. Optimization of a series of mGlu1 PAMs based on an N-(3-chloro-4-(1,3-dioxoisoindolin-2-yl)phenyl)-3-methylfuran-2-carboxamide scaffold provided 17e, a potent (mGlu1 EC50 = 31.8 nM) and highly CNS penetrant (brain to plasma ratio (Kp) of 1.02) mGlu1 PAM tool compound, that potentiated not only wild-type human mGlu1 but also mutant mGlu1 receptors derived from deleterious GRM1 mutations found in schizophrenic patients. Moreover, both electrophysiological and in vivo studies indicate the mGlu1 ago-PAMs/PAMs do not possess the same epileptiform adverse effect liability as mGlu5 ago-PAMs/PAMs and maintain temporal activity suggesting a broader therapeutic window.

Positive allosteric modulators of GABAA receptors (GAMs) acting at specific subtypes of GABAA receptors effectively restore compromised spinal pain control in rodents. Studies addressing a similar antihyperalgesic effect in humans are sparse and are hampered by sedative effects of nonselective GAMs available for use in humans. We present results from a randomized controlled double-blind crossover study in 25 healthy volunteers, which addressed potential antihyperalgesic actions of clobazam (CBZ) and clonazepam (CLN) at mildly sedating equianticonvulsive doses. Clobazam was chosen because of its relatively low sedative properties and CLN because of its use in neuropathic pain. Tolterodine (TLT) was used as an active placebo. The primary outcome parameter was a change in the area of cutaneous UVB irradiation-induced secondary hyperalgesia (ASH), which was monitored for 8 hours after drug application. Sedative effects were assessed in parallel to antihyperalgesia. Compared with TLT, recovery from hyperalgesia was significantly faster in the CBZ and CLN groups (P = 0.009). At the time point of maximum effect, the rate of recovery from hyperalgesia was accelerated by CBZ and CLN, relative to placebo by 15.7% (95% confidence interval [CI] 0.8-30.5), P = 0.040, and 28.6% (95% CI 4.5-52.6), P = 0.022, respectively. Active compounds induced stronger sedation than placebo, but these differences disappeared 8 hours after drug application. We demonstrate here that GAMs effectively reduce central sensitization in healthy volunteers. These results provide proof-of-principle evidence supporting efficacy of GAMs as antihyperalgesic agents in humans and should stimulate further research on compounds with improved subtype specificity.

Background and Purpose: The GABA(B) receptor agonist, baclofen, has shown promising effects in patients suffering from pain, post-traumatic stress disorder, alcoholism, overactive bladder and gastroesophageal reflux disease. However, baclofen's short duration of action and side effects limit its wider use. Here we characterized a novel, GABA(B) receptor positive allosteric modulator (PAM) ADX71943., Experimental Approach: In vitro, ADX71943 was assessed for pharmacological activity and selectivity using recombinant and native GABA(B) receptors. In vivo ADX71943 was assessed in the acetic acid-induced writhing (AAW) test in mice and formalin tests (FTs) in mice and rats. Marble burying (MB) and elevated plus maze (EPM) tests, rotarod, spontaneous locomotor activity (sLMA) and body temperature (BT) tests in mice and rats were used to investigate centrally-mediated effects., Key Results: In vitro, in the presence of GABA, ADX71943 increased the potency and efficacy of agonists and showed selectivity at the GABA(B) receptor. ADX71943 reduced pain-associated behaviours in AAW; an effect blocked by GABA(B) receptor antagonist CGP63360. ADX71943 reduced pain in the FT in mice and rats, but was inactive in the MB and EPM despite reaching high concentrations in plasma. ADX71943 had no effect on BT, rotarod and sLMA., Conclusions and Implications: ADX71943 showed consistent and target-related efficacy in tests of disorders that have a significant peripheral component (acute and chronic pain), while having no effect in those associated with centrally-mediated anxiety-like reactivity and side effects. Thus, ADX71943 is a useful pharmacological tool for delineation of peripherally- versus centrally-mediated effects of GABA(B) receptor activation., (© 2014 The British Pharmacological Society.)

What Is Known and Objective: AZD7325 is a selective gamma-amino-butyric acid (GABAA )α2, 3 receptor modulator. The aims of this analysis were to develop population pharmacokinetic (PPK) models of AZD7325 and midazolam and to assess the induction effect of AZD7325 on CYP3A4 with midazolam as a substrate., Methods: Drug-drug interaction data of AZD7325 and midazolam from 24 healthy subjects were available for model development. PPK models were developed in a sequential manner using NONMEM. Both AZD7325 and midazolam pharmacokinetics were described by two-compartment models, and a transit compartment absorption model and a first-order absorption model were applied for the absorption of AZD7325 and midazolam, respectively. The induction of CYP3A by AZD7325 was described by a transit enzyme model, where the elimination of midazolam was proportionally linked to the enzyme amount. Simulations were performed to predict dosing regimens to account for the induction of CYP3A4., Results and Discussion: The population estimates for AZD7325 clearance, intercompartmental clearance, central and peripheral volume were 36, 29·2 L/h, 169 and 392 L, respectively, with interindividual variability (IIV) of 35% and 24% for clearance and central volume. Midazolam clearance, intercompartmental clearance, central and peripheral volume were estimated to be 62·7, 34·7 L/h, 133 and 146 L, respectively, with 43% IIV for clearance. The estimated mean transit time for induction of the CYP3A4 enzyme was 197 h, with 57% IIV., What Is New and Conclusion: The PPK models developed adequately described the clinical observation of AZD7325-mediated CYP3A4 enzyme induction with midazolam as a probe. The model could provide basis for the rational dosing of AZD7325 in clinical practice., (© 2014 John Wiley & Sons Ltd.)

Physical exercise has become a potentially beneficial therapy for reducing neurodegeneration symptoms in Alzheimer's disease. Previous studies have shown that cognitive deterioration, anxiety and the startle response observed in 7-month-old 3xTg-AD mice were ameliorated after 6 months of free access to a running wheel. Also, alterations in synaptic response to paired-pulse stimulation were improved. The present study further investigated some molecular mechanisms underlying the beneficial effects of 6 months of voluntary exercise on synaptic plasticity in 7-month-old 3xTg-AD mice. Changes in binding parameters of [(3)H]-flunitrazepam to GABAA receptor and of [(3)H]-MK-801 to NMDA receptor in cerebral cortex of 3xTgAD mice were restored by voluntary exercise. In addition, reduced expression levels of NMDA receptor NR2B subunit were reestablished. The synaptic proteins synaptophysin and PSD-95 and the neuroprotective proteins GDNF and SIRT1 were downregulated in 3xTgAD mice and were recovered by exercise treatment. Overall, in this paper we highlight the fact that different interrelated mechanisms are involved in the beneficial effects of exercise on synaptic plasticity alterations in the 3xTg-AD mouse model., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Enormous progress in understanding the role of four populations of benzodiazepine-sensitive GABAA receptors was paralleled by the puzzling findings suggesting that substantial separation of behavioral effects may be accomplished by apparently non-selective modulators. We report on SH-I-048A, a newly synthesized chiral positive modulator of GABAA receptors characterized by exceptional subnanomolar affinity, high efficacy and non-selectivity. Its influence on behavior was assessed in Wistar rats and contrasted to that obtained with 2mg/kg diazepam. SH-I-048A reached micromolar concentrations in brain tissue, while the unbound fraction in brain homogenate was around 1.5%. The approximated electrophysiological responses, which estimated free concentrations of SH-I-048A or diazepam are able to elicit, suggested a similarity between the 10mg/kg dose of the novel ligand and 2mg/kg diazepam; however, SH-I-048A was relatively more active at α1- and α5-containing GABAA receptors. Behaviorally, SH-I-048A induced sedative, muscle relaxant and ataxic effects, reversed mechanical hyperalgesia 24h after injury, while it was devoid of clear anxiolytic actions and did not affect water-maze performance. While lack of clear anxiolytic actions may be connected with an enhanced potentiation at α1-containing GABAA receptors, the observed behavior in the rotarod, water maze and peripheral nerve injury tests was possibly affected by its prominent action at receptors containing the α5 subunit. The current results encourage further innovative approaches aimed at linking in vitro and in vivo data in order to help define fine-tuning mechanisms at four sensitive receptor populations that underlie subtle differences in behavioral profiles of benzodiazepine site ligands., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Persistent activation of GABAA receptors triggers compensatory changes in receptor function that are relevant to physiological, pathological and pharmacological conditions. Chronic treatment of cultured neurons with GABA for 48h has been shown to produce a down-regulation of receptor number and an uncoupling of GABA/benzodiazepine site interactions with a half-time of 24-25h. Down-regulation is the result of a transcriptional repression of GABAA receptor subunit genes and depends on activation of L-type voltage-gated calcium channels. The mechanism of this uncoupling is currently unknown. We have previously demonstrated that a single brief exposure of rat primary neocortical cultures to GABA for 5-10min (t½=3min) initiates a process that results in uncoupling hours later (t½=12h) without a change in receptor number. Uncoupling is contingent upon GABAA receptor activation and independent of voltage-gated calcium influx. This process is accompanied by a selective decrease in subunit mRNA levels. Here, we report that the brief GABA exposure induces a decrease in the percentage of α3-containing receptors, a receptor subtype that exhibits a high degree of coupling between GABA and benzodiazepine binding sites. Initiation of GABA-induced uncoupling is prevented by co-incubation of GABA with high concentrations of sucrose suggesting that it is dependent on a receptor internalization step. Moreover, results from immunocytochemical and biochemical experiments indicate that GABA exposure causes an increase in GABAA receptor endocytosis. Together, these data suggest that the uncoupling mechanism involves an initial increase in receptor internalization followed by activation of a signaling cascade that leads to selective changes in receptor subunit levels. These changes might result in the assembly of receptors with altered subunit compositions that display a lower degree of coupling between GABA and benzodiazepine sites. Uncoupling might represent a homeostatic mechanism that negatively regulates GABAergic transmission under physiological conditions in which synaptic GABAA receptors are transiently activated for several minutes., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

The duration of action of a drug is commonly estimated using plasma concentration, which is not always practical to obtain or an accurate estimate of functional half life. For example, flumazenil is used clinically to reverse the effects of benzodiazepines like midazolam; however, its elimination can be altered by other drugs, including some benzodiazepines, thereby altering its half life. This study used Schild analyses to characterize antagonism of midazolam by flumazenil and determine the functional half life of flumazenil. Four monkeys discriminated 0.178mg/kg midazolam while responding under a fixed-ratio 10 schedule of stimulus-shock termination; flumazenil was given at various times before determination of a midazolam dose-effect curve. There was a time-related decrease in the magnitude of shift of the midazolam dose-effect curve as the interval between flumazenil and midazolam increased. The potency of flumazenil, estimated by apparent pA2 values (95% CI), was 7.30 (7.12, 7.49), 7.17 (7.03, 7.31), 6.91 (6.72, 7.10) and 6.80 (6.67, 6.92) at 15, 30, 60 and 120min after flumazenil administration, respectively. The functional half life of flumazenil, derived from potency estimates, was 57±13min. Thus, increasing the interval between flumazenil and midazolam causes orderly decreases in flumazenil potency; however, across a broad range of conditions, the qualitative nature of the interaction does not change, as indicated by slopes of Schild plots at all time points that are not different from unity. Differences in potency of flumazenil are therefore due to elimination of flumazenil and not due to pharmacodynamic changes over time., (© 2013 Published by Elsevier B.V.)

Objective: We conducted a pharmacokinetic (PK) study and a pharmacodynamic (PD) study to assess whether Roux-en-Y gastric bypass (RYGB) surgery is associated with significant changes to PK and PD of oral medications., Background: The effect of RYGB on oral drug disposition is not well understood., Methods: An oral cocktail of probe drugs for major drug-metabolizing enzymes (caffeine, tolbutamide, omeprazole, dextromethorphan, and oral and intravenous midazolam) was administered to 18 RYGB recipients and 18 controls. Timed blood and urine samples were obtained for PK analyses. Forty mg of oral furosemide was administered to 13 RYGB recipients and 14 controls, and urine and blood samples were collected for assessing furosemidePK, and urine volume and urine sodium excretion for PD analyses., Results: Compared with controls, the RYGB group had significantly lower time to maximum plasma concentration (tmax) for caffeine (0.58 ± 0.5 vs 2.1 ± 2.2 hours, P < 0.0001), tolbutamide (1.4 ± 1.8 vs 2.1 ± 2.2 hours, P = 0.0001), omeprazole (1.1 ± 1.1 vs 4.4 ± 1.3 hours, P < 0.0001), and oral midazolam (0.5 ± 0.2 vs 0.7 ± 0.4 hours, P < 0.01). However, maximum plasma concentration, half-life, area under the curve, and oral bioavailability were not different. Compared with controls, the RYGB group had brisk natriuresis, with significantly lower tmax for urine sodium (1.3 ± 0.5 vs 3.1 ± 2.3 hours, P < 0.02) and correspondingly lower tmax for furosemide (1.8 ± 0.3 vs 4.2 ± 1.2 hours, P = 0.006). However, 6-hour urine sodium and 6-hour urine volume were not different between the two groups., Conclusions: RYGB recipients have significantly shorter tmax for the studied orally administered medications, but otherwise no other significant changes in PK were reported.

Flumazenil is a specific, reversibly bound antagonist at benzodiazepine binding sites of gamma-aminobutyric acid A receptors; these sites can be imaged using positron emission tomography with 11C-flumazenil. We reported an exponential decline of flumazenil volume of distribution (proportional to receptor binding) of gamma-aminobutyric acid A receptors in children 2 to 17 years. Six newborns (33.3-46.7 weeks' postconception) were studied. All had experienced epileptic seizures and undergone 60-minute dynamic 11C-flumazenil-positron emission tomography imaging after injection of 0.4 mCi/kg of 11C-flumazenil. All newborns were scanned during their natural sleep. Binding potential (indicating flumazenil receptor binding) was calculated using Logan-plot analysis. Visual and quantitative analyses showed highest receptor binding in the amygdala-hippocampus region, sensory-motor cortex, thalamus, brainstem and basal ganglia, in that order. Cerebellum and most of the cerebral cortex showed relatively low binding. This is the first demonstration of gamma-aminobutyric acid A receptor binding in human neonates and is strikingly different from that in older children/adults, showing a programmed pattern of expression. The ontogeny data of flumazenil receptor binding from children may contribute to understanding regional differences in synaptic plasticity and improve rational therapeutic use of drugs acting at the gamma-aminobutyric acid A receptor in the pediatric population., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Autoinduction in drug metabolism is a known phenomenon observed when a drug induces the enzymes responsible for its own metabolism. The potency, rate and extent of autoinduction following a given treatment paradigm may have therapeutic implications in clinic as well as for in vivo pharmacological assessments in animals. RO4938581, an imidazo-triazolo-benzodiazepine, is a novel GABAAα5 negative modulator recently pursued for the treatment of cognitive dysfunctions. As circulating plasma levels of RO4938581 were shown to decrease rapidly after repeated dosing in rats, with CYP1A2 being involved in the metabolism of the compound, we examined the potential role of RO4938581-mediated autoinduction of CYP1A2. Incubation of rat hepatocytes with RO4938581 revealed potent CYP1A2 induction with significant increase in enzymatic activity at concentrations of 0.1nM and RO4938581 was shown to be 700-fold more potent than β-napththoflavone. Ex vivo studies revealed a 7-fold increase in metabolic CYP1A2 activity in liver microsomes prepared from rats administered with 0.1mg/kg of RO4938581 24h before. This induction profile was reflected in vivo in pharmacokinetic studies in rats where an 8-fold reduction in plasma exposure was observed after a second dose. The reduction in plasma exposures due to CYP1A2 autoinduction were confirmed functionally in contextual fear conditioning paradigm in rats, where a positive pharmacological effect observed after acute drug administration disappeared completely after sub-chronic dosing. Together, these findings suggest that RO4938581 possesses potent CYP1A2 autoinductive properties in rats and may serve as a tool for mechanistic metabolism or drug-drug interaction studies encircling this enzyme in rats., (Copyright © 2013 Elsevier Inc. All rights reserved.)

What Is Already Known About This Subject: • Tofacitinib (CP-690,550) is a novel, oral Janus kinase inhibitor being investigated as a targeted immunomodulator and disease-modifying therapy in rheumatoid arthritis. • Non-renal elimination accounts for 70% of the total clearance of tofacitinib and the metabolism is primarily mediated by cytochrome P450 (CYP) 3A4. • This study was required to determine the effect of tofacitinib on the in vivo pharmacokinetics of a sensitive CYP3A4 substrate., What This Study Adds: • The pharmacokinetics of midazolam, a sensitive CYP3A4 substrate, are not altered when co-administered with tofacitinib in healthy subjects. • Tofacitinib is unlikely to affect the clearance of drugs metabolized by CYP enzymes. • There is no need for dose adjustments of CYP substrates when co-administered with tofacitinib., Aims: To investigate inhibitive and inductive effects of tofacitinib (CP-690,550), a Janus kinase inhibitor, on CYP3A4 function via in vitro and in vivo studies., Methods: In vitro experiments were conducted to assess the inhibition and induction potential of tofacitinib for major drug metabolizing enzymes (CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP3A4). A phase 1, randomized, open-label, two-way crossover study (NCT00902460) was conducted to confirm the lack of inhibitive/inductive effect on a sensitive CYP3A4 substrate, midazolam, in healthy subjects. Midazolam pharmacokinetics were assessed over 24 h following single dose 2 mg administration prior to administering tofacitinib and after twice daily dosing of tofacitinib 30 mg for 6 days. The primary endpoint was midazolam area under the concentration-time profile, from time 0 to infinity (AUC(0,∞))., Results: In vitro studies demonstrated low potential for CYP inhibition (IC(50) estimates tofacitinib > 30 µm), CYP3A4 mRNA induction (observed at tofacitinib concentrations ≥ 25 µm) and no effect on enzymatic activity of CYP substrates. In the human study, AUC(0,∞) adjusted geometric mean ratio for midazolam plus tofacitinib to midazolam alone was 103.97% [90% confidence interval (CI) 95.57, 113.12], wholly within the pre-specified acceptance region (80, 125). The 90% CI for the ratio of adjusted geometric means of maximum plasma concentration (C(max) ) (95.98, 108.87) was also wholly within this acceptance region., Conclusions: These data confirm a lack of an inhibitive or inductive effect of tofacitinib on CYP3A activity in humans and, in conjunction with in vitro data, support the conclusion that tofacitinib is unlikely to influence the CYP enzyme system as a whole., (© 2012 Pfizer Inc. British Journal of Clinical Pharmacology © 2012 The British Pharmacological Society.)

Rationale: Quantitative analysis of electroencephalographic signals (EEG) and their interpretation constitute a helpful tool in the assessment of the bioavailability of psychoactive drugs in the brain. Furthermore, psychotropic drug groups have typical signatures which relate biochemical mechanisms with specific EEG changes., Objectives: To analyze the pharmacological effect of a dose of alprazolam on the connectivity of the brain during wakefulness by means of linear and nonlinear approaches., Methods: EEG signals were recorded after alprazolam administration in a placebo-controlled crossover clinical trial. Nonlinear couplings assessed by means of corrected cross-conditional entropy were compared to linear couplings measured with the classical magnitude squared coherence., Results: Linear variables evidenced a statistically significant drug-induced decrease, whereas nonlinear variables showed significant increases. All changes were highly correlated to drug plasma concentrations. The spatial distribution of the observed connectivity changes clearly differed from a previous study: changes before and after the maximum drug effect were mainly observed over the anterior half of the scalp. Additionally, a new variable with very low computational cost was defined to evaluate nonlinear coupling. This is particularly interesting when all pairs of EEG channels are assessed as in this study., Conclusions: Results showed that alprazolam induced changes in terms of uncoupling between regions of the scalp, with opposite trends depending on the variables: decrease in linear ones and increase in nonlinear features. Maps provided consistent information about the way brain changed in terms of connectivity being definitely necessary to evaluate separately linear and nonlinear interactions.

Objective: This [11C]flumazenil (FMZ) study evaluates the performance of various parametric analysis methods and their ability to detect statistically significant group differences., Methods: Dynamic 60-min FMZ scans were performed in eight healthy and nine individuals with major depressive disorder. Parametric volume of distribution (VT) images were generated using a basis function method (BFM) implementation of the single tissue compartment model (1T) and Logan plot analysis, both with a metabolite-corrected arterial plasma input function. Parametric binding potential (BP ND) images were generated using multilinear reference tissue methods (MRTM0-4), reference Logan and receptor parametric mapping (RPM1-2), with pons as a reference region. Standardized uptake value (SUV) and SUV ratio-to-pons (SUVr) images were calculated over the time interval 30-40 and 20-60 min postinjection. The resulting VT, BP ND, SUV and SUVr values were compared with nonlinear regression values, using both the 1T model and the simplified reference tissue model. Statistical parametric mapping (SPM5) was used to detect group differences, with an emphasis on the bilateral parahippocampal gyri., Results: BFM was more accurate than Logan, but showed more variability. Both RPM methods and MRTM2 showed the best average correlation with the simplified reference tissue model. In using SPM, SUV and SUVr images provided the best contrast between groups in the parahippocampal gyri, but provided large underestimation and overestimation in quantitative comparisons. BFM and RPM methods allowed for the determination of perfusion effects., Conclusion: Parametric Logan VT, MRTM2 and RPM1-2 BP ND methods allow the best quantitative comparison of FMZ binding between groups and show good discriminating performance in SPM analysis.

Objectives: Two investigations aimed to define the pharmacokinetic profile of a modified-release preparation of zaleplon (SKP-1041)., Methods: Protocol SOM001 was a 5-way crossover, double-blind, randomized trial comparing three novel modified-release formulations of zaleplon 15 mg (SKP-1041A, SKP-1041B, SKP-1041C) to placebo and immediate-release zaleplon 10 mg. Protocol SOM002 was a randomized, crossover, open-label trial to compare the pharmacokinetics of SKP-1041B after day and night administration. In SOM001, study drug was administered at 9:00 a.m. (fasted); blood samples were obtained beginning 1 h predose through 12 h postdose. In study SOM002, study drug was administered at 9:00 a.m. or 10:30 p.m.; blood samples were obtained beginning 1 h predose through 12 h postdose. Subjects were 19 (SOM001) and 23 (SOM002) healthy adults between ages 20-46., Results: Dose-normalized total AUCs for modified-release preparations A, B, C and immediate-release zaleplon were not significantly different; peak plasma concentrations were similar for A and B, and both were significantly higher than C. Time to peak plasma concentration for A, B, and C were 4-5 h compared to 1.5 h for immediate-release zaleplon; mean terminal phase half-life was in the range 1-2 h for A, B and immediate-release zaleplon. No significant differences were noted between day and night administration in the SOM002 study., Conclusions: Zaleplon, 15 mg, in a novel, modified-release formulation (SKP-1041) had a time to peak plasma concentrations at 4-5 h postdose compared to 1.5 h for immediate-release zaleplon, 10 mg. The pharmacokinetic profile suggests this formulation may be useful for treating middle-of-the-night awakening., (Copyright © 2011 John Wiley & Sons, Ltd.)

Drug-facilitated sexual assault (DFSA) is a serious and troubling crime. It is important to know if and how different drugs might be used to facilitate assault in order to deter such crime. There are a number of ways in which drugs that are used for DFSA might not be detected by routine screens. The purpose of this analysis was to draw reasonable inferences regarding drugs with a high likelihood of being used for DFSA and not being detected by routine screens. National data from poison control centres, hospital emergency rooms, and law enforcement seizures were used to evaluate the relative magnitude of problems and illicit availability associated with different classes of drugs. General drug classes were examined to include additional drugs that might be used for DFSA on the basis of their amnesic effects, widespread availability, and pharmacokinetics (i.e. short half-life). The benzodiazepine-site ligands zolpidem and eszopiclone, 'club drugs' GHB and ketamine, muscle relaxants such as carisoprodol, and antihistamines such as diphenhydramine were identified as drugs that might be used for DFSA and remain undetected by routine screens. Future studies that are designed to examine the role of these drugs in DFSA cases could provide better estimates of their use for DFSA. A better understanding of what is being missed in DFSA cases might help prioritize the development of new assays, provide rationale for the availability of particular assays for routine testing, and inform practitioners and the general public of the potential DFSA risks of certain drugs., (Copyright © 2010 John Wiley & Sons, Ltd.)

Background: Preclinical and clinical evidence suggests that impaired gamma-aminobutyric (GABA) control, leading to disinhibition within the sensorimotor system, might play a role in dystonia. Aim of this study is the in vivo assessment of the GABAergic system in dystonia using positron emission tomography (PET) and (11) C-flumazenil, a selective GABA(A) receptor ligand., Methods: Fourteen subjects with primary dystonia (9 carriers of the DYT1 mutation and 5 sporadic cases) were compared to 11 controls, using a simplified reference tissue model to measure binding potential., Results: Voxel-based analyses showed a reduction in GABA(A) receptor expression/affinity both in DYT1 carriers and sporadic patients in primary motor and premotor cortex, primary and secondary somatosensory cortex, and in the motor component of the cingulate gyrus., Conclusions: Dysfunction of GABA(A) receptors in sensorimotor systems in primary (genetic and sporadic) dystonia supports the view that lack of GABAergic control may be associated with the generation of dystonic movements., (Copyright © 2011 Movement Disorder Society.)

Objective: The objective was to determine the effects of the SLCO2B1 c.1457C> T polymorphism and apple juice on the pharmacokinetics of fexofenadine and midazolam in humans., Methods: Individuals were divided based on the genotype of SLCO2B1 c.1457C> T (n = 14, c.[1457C]+ c.[= ] 5,c.[1457C]+ c.[1457C> T] 5, and c.[1457C> T]+c.[1457C> T] 4). The oral pharmacokinetics of 60 mg fexofenadine and 5mg midazolam were assessed with water or apple juice (1200 ml/day) in a randomized crossover study. OATP2B1-mediated uptake of fexofenadine and midazolam was evaluated with Xenopus laevis oocyte gene-expression system., Results: When fexofenadine was administered with water, subjects with c.[1457C> T] allele showed a significant decrease in fexofenadine in the area under the plasma concentration-time curve (AUC) compared with c.[1457C] + c[= ] subjects (1110 ± 347 vs. 1762 ± 542 ng . h/ml, P< 0.05). When administered with apple juice, a significant decrease in the fexofenadine AUC was observed compared with water (1342 ± 519 vs. 284 ± 79.2 ng . h/ml, P < 0.05). The apple juice induced decrease in fexofenadine AUC was significantly lower in subjects carrying the c.[1457C> T] allele. Neither the genotype nor the apple juice showed significant effects on the pharmacokinetics of midazolam except for a marginally significant decrease in Cmax after administration with apple juice. The uptake of fexofenadine by OATP2B1 cRNA-injected oocytes was significantly higher than that by water-injected oocytes. Apple juice, but not midazolam, significantly decreased the uptake of fexofenadine by OATP2B1 cRNA-injected oocytes., Conclusion: The results suggest that fexofenadine is a substrate of OATP2B1, and the transport function of OATP2B1 is subject to the genotype of SLCO2B1 c.1457C> T and apple juice. It is likely that apple juice has little effect on CYP3A.

Objectives: To clarify whether alterations in midazolam pharmacokinetics resulting from changes in cytochrome P450 3A (CYP3A) activity lead to changes in its pharmacodynamic effects, benzodiazepine receptor occupancy was measured in the brain of rats after oral administration of midazolam., Methods: Receptor occupancy was measured by radioligand binding assay in rats pretreated with ursodeoxycholic acid (UDCA), ketoconazole and dexamethasone, and the plasma concentration of midazolam was simultaneously determined., Key Findings: There was a significant increase in the apparent dissociation constant and decrease in the maximum number of binding sites for specific [(3) H]flunitrazepam binding after oral administration of midazolam at pharmacologically relevant doses, suggesting that midazolam binds significantly to brain benzodiazepine receptors. Pretreatment with UDCA significantly enhanced the binding. This correlated well with significant enhancement by UDCA of the plasma midazolam concentration. The brain benzodiazepine receptor binding of oral midazolam was significantly enhanced by pretreatment with ketoconazole, a potent inhibitor of CYP3A, whereas it was significantly reduced by treatment with dexamethasone, an inducer of this enzyme. These effects paralleled changes in the plasma concentration of midazolam., Conclusions: The results indicate that pharmacokinetic changes such as altered CYP3A activity significantly influence the pharmacodynamic effect of midazolam by affecting occupancy of benzodiazepine receptors in the brain. They also suggest in-vivo or ex-vivo time-dependent measurements of receptor occupancy by radioligand binding assay to be a tool for elucidating the pharmacokinetic interaction of benzodiazepines with other agents in pre-clinical and clinical evaluations., (© 2010 The Authors. JPP © 2010 Royal Pharmaceutical Society.)

Traditional Chinese herbal medicines are frequently prescribed in pharmacotherapy in Japan. In the present study, we evaluated the possible interaction of several herbal extracts including Rhei Rhizoma extract with cytochrome P450 (CYP) 3A and efflux transporters such as P-glycoprotein and multidrug resistance-associated protein (MRP) 2. Rhei Rhizoma extract (100 microg/ml) significantly suppressed the CYP3A-mediated 6beta-hydroxylation of testosterone in hepatic microsomes, and increased the extent of bioavailability of midazolam, a typical CYP3A substrate, in rats. Also, Rhei Rhizoma extract (300 microg/ml) significantly suppressed P-glycoprotein-mediated efflux transport of rhodamine 123 (Rho123) in rat everted intestine. In an in-vivo study, Rhei Rhizoma extract added to intestinal perfusate at a concentration of 300 microg/ml significantly suppressed the intestinal exsorption of Rho123, though it exerted no effect on the biliary excretion of Rho123. Furthermore, the in-vitro and in-vivo MRP2-mediated intestinal efflux of 2,4-dinitrophenyl-S-glutathione was significantly suppressed by Rhei Rhizoma extract (1000 microg/ml). In conclusion, Rhei Rhizoma extract, which is taken orally at doses of 0.5-1 g each or 1-3 g daily in clinical practice, may cause pharmacokinetic herb-drug interactions in the process of the intestinal and/or hepatic CYP3A-mediated drug metabolism and P-glycoprotein- and/or MRP2-mediated efflux transport in the intestine.

GABA(A) receptor (R) positive allosteric modulators that selectively modulate GABA(A)Rs containing beta(2)- and/or beta(3)- over beta(1)-subunits have been reported across diverse chemotypes. Examples include loreclezole, mefenamic acid, tracazolate, and etifoxine. In general,"beta(2/3)-selective" GABA(A)R positive allosteric modulators are nonbenzodiazepines (nonBZs), do not show alpha-subunit isoform selectivity, yet have anxiolytic efficacy with reduced ataxic/sedative effects in animal models and humans. Here, we report on an enantiomeric pair of nonBZ GABA(A)R positive allosteric modulators that demonstrate differential beta-subunit isoform selectivity. We have tested this enantiomeric pair along with a series of other beta(2/3)-subunit selective, alpha-subunit isoform-selective, BZ and nonBZ GABA(A) positive allosteric modulators using electrophysiological, pharmacokinetic, and behavioral assays to test the hypothesis that ataxia may be correlated with the extent of modulation at beta(1)-subunit-containing GABA(A)Rs. Our findings provide an alternative strategy for designing anxioselective allosteric modulators of the GABA(A)R with BZ-like anxiolytic efficacy by reducing or eliminating activity at beta(1)-subunit-containing GABA(A)Rs.

Ketoconazole inhibits the Cytochrome P450 3A12 (CYP3A12) metabolizing enzyme as well as the p-glycoprotein efflux pump. The extent and clinical consequence of these effects are poorly understood in dogs. The objective was to assess the pharmacokinetics of ketoconazole after single and multiple doses and the effect of multiple doses of ketoconazole on midazolam (a known CYP3A12 substrate) and the opioid fentanyl. Six greyhound dogs were studied. The study consisted of three phases. Phase 1 consisted of i.v. midazolam (0.23 mg/kg base) and fentanyl (15.71 microg/kg base). Phase 2 consisted of a single oral dose of ketoconazole (mean dose 12.34 mg/kg). Phase 3 consisted of i.v. midazolam (0.23 mg/kg) and fentanyl (10 microg/kg) after 5 days of oral ketoconazole (12.25 mg/kg/day). Ketoconazole significantly inhibited its own elimination with the mean residence time (MRT) increasing from 6.24 h in Phase 1 to 12.54 h in Phase 3. Ketoconazole significantly decreased the elimination of midazolam, as expected, with the MRT increasing from 0.81 to 1.49 h. The elimination of fentanyl was not significantly altered by co-administration of ketoconazole with the MRT being 3.90 and 6.35 h. The MRT was the most robust estimate of decreased drug elimination.

A physiologically based pharmacokinetic (PBPK) model was developed for midazolam in the chicken and extended to three other species. Physiological parameters included organ weights obtained from 10 birds of each species and blood flows obtained from the literature. Partition coefficients for midazolam in tissues vs. plasma were estimated from drug residue data obtained at slaughter. The avian models include separate compartments for venous plasma, liver, kidney, muscle, fat and all other tissues. An estimate of total body clearance from an earlier in vitro study was used as a starting value in the model, assuming almost complete removal of the parent compound by liver metabolism. The model was optimized for the chicken with plasma and tissue data from a pharmacokinetic study after intravenous midazolam (5 mg/kg) dose. To determine which parameters had the most influence on the goodness of fit, a sensitivity analysis was performed. The optimized chicken model was then modified for the turkey, pheasant and quail. The models were validated with midazolam plasma and tissue residue data in the turkey, pheasant and quail. The PBPK models in the turkey, pheasant and quail provided good predictions of the observed tissue residues in each species, in particular for liver and kidney.

Benzodiazepines are effective short-term treatments for anxiety disorders, but their use is limited by undesirable side effects related to Central Nervous System impairment and tolerance development. SL65.1498 is a new compound that acts in vitro as a full agonist at the gamma-aminobutyric acid(A) 2 and 3 receptor and as a partial agonist at the 1 and 5 receptor subtypes. It is thought that the compound could be anxiolytic by its activation at the alpha2 and alpha3 receptor subtypes, without causing unfavourable side effects, which are believed to be mediated by the alpha1 and alpha5 subtypes. This study was a double-blind, five-way cross-over study to investigate the effects of three doses of SL65.1498 in comparison with placebo and lorazepam 2 mg in healthy volunteers. The objective was to select a dose level (expected to be therapeutically active), free of any significant deleterious effect. Psychomotor and cognitive effects were measured using a validated battery of measurements, including eye movements, body sway, memory tests, reaction-time assessments, and visual analogue scales. The highest dose of SL65.1498 showed slight effects on saccadic peak velocity and smooth pursuit performance, although to a much lesser extent than lorazepam. In contrast to lorazepam, none of the SL65.1498 doses affected body sway, visual analogue scale alertness, attention, or memory tests. This study showed that the three doses of SL65.1498 were well tolerated and induced no impairments on memory, sedation, psychomotor, and cognitive functions.

Rationale: Gamma-butyrolactone (GBL) and 1,4-butanediol (1,4-BD) are prodrugs for gamma-hydroxybutyrate (GHB). Like GHB, GBL and 1,4-BD are drugs of abuse, but their behavioral effects may differ from GHB under some conditions., Objectives: The first study compared the behavioral effects of GBL (32-240 mg/kg) and 1,4-BD (32-240 mg/kg) with each other and to effects previously reported for GHB (32-420 mg/kg). A second study determined GHB pharmacokinetics following intragastric administration of GHB, GBL, and 1,4-BD., Methods: Operant responding for food, observed behavioral effects, and a fine-motor task occurred at multiple time intervals after administration of drug or vehicle. In a separate pharmacokinetics study, blood samples were collected across multiple time points after administration of GHB, GBL, and 1,4-BD., Results: Like GHB, GBL, and 1,4-BD impaired performance on the fine-motor task, but the onset of motor impairment differed across drugs. GBL and 1,4-BD dose dependently decreased the number of food pellets earned, but at lower doses than previously observed for GHB. Similar to GHB, both GBL and 1,4-BD produced sedation, muscle relaxation, gastrointestinal symptoms, and tremors/jerks. Administration of GBL and 1,4-BD produced higher maximum concentrations of GHB with shorter times to maximum concentrations of GHB in plasma when compared to GHB administration., Conclusions: GBL and 1,4-BD produced behavioral effects similar to those previously reported with GHB and the time course of effects were related to blood levels of GHB. Given their higher potency and faster onset of effects, the abuse liability of GBL and 1,4-BD may be greater than GHB.

Unlabelled: In the management of patients who require intravenous sedation with midazolam care must be taken if the patient is taking herbal drugs. There is a potential for harmful drug interactions between herbal drugs and midazolam.Assessment of which herbal drugs a patient is taking should be part of the treatment planning stage so interactions can be avoided or anticipated., Clinical Relevance: Growing numbers of patients are taking herbal or Chinese medication and may not disclose this to the sedation team. Such drugs have the potential for interactions and can affect induction and recovery from sedation.

Thiopental is a barbiturate used in traumatic brain injuries (TBIs) to reduce intracranial pressure (ICP) and to manage cerebral ischemia. As thiopental follows Michaelis-Menten kinetics, therapeutic drug monitoring (TDM) has been used in practice to improve efficacy and reduce adverse effects. However, its role is still debatable, and TDM is not widely practiced. Current evidence suggests that thiopental therapy may improve mortality and functional outcome in a subpopulation of patients with severe TBI with elevated ICP refractory to conventional medical therapy. Several analytical methods are available to quantify thiopental concentrations. This review uses a previously published 9-step decision-making algorithm to determine whether TDM of thiopental in TBI is warranted. There seems to be poor correlation between thiopental concentration and pharmacological response in terms of neurological response, ICP, electroencephalography, and drug toxicity. There is no established therapeutic range for thiopental continuous infusion due to a wide range of plasma concentrations corresponding to efficacy (25-50 mg/L) and toxicity (30-70 mg/L) and the resulting overlap between the 2. Thiopental exhibits intrapatient and interpatient variability due to age, obesity, renal and hepatic dysfunction, Michaelis-Menten kinetics, and hepatic enzyme autoinduction. Available evidence suggests that TDM of thiopental continuous infusion is not beneficial in improving efficacy or avoiding toxicity. There are however 2 possible scenarios in which TDM may provide additional information to sound clinical judgment. The first is providing patient-specific plasma target concentration to guide titration of therapy. The second scenario is differentiating between brain death and barbiturate-induced coma.

Unlabelled: Phenotyping intestinal and hepatic cytochrome P450 (CYP) 3A activity with oral midazolam can be limited by midazolam-induced central nervous system (CNS) side effects. Determining methods to minimize CNS side effects optimizes use of midazolam as a CYP3A probe., Objective: The objective of this study was to determine the effect of intravenous (i.v.) flumazenil on midazolam apparent oral clearance (a surrogate marker of CYP3A activity). Midazolam pharmacodynamics were also evaluated., Methods: This was a randomized, double-blind, placebo-controlled, single-dose, two-way crossover study. 16 healthy volunteers (8 women) were concomitantly administered i.v. flumazenil 0.005 mg/kg or i.v. placebo and oral midazolam 0.075 mg/kg. Blood samples were obtained to determine midazolam and flumazenil plasma concentrations. Bioequivalence was assessed by determining geometric mean ratios (GMR) and 90% confidence intervals (90% CI). Baseline and post dose digit symbol substitution tests (DSST), Groton maze learning tests (GMLT), and Stanford sleepiness scales (SSS) were conducted., Results: Apparent oral clearance was 2,030 +/- 651 and 1,939 +/- 658 ml/min for the midazolam plus flumazenil and midazolam plus placebo groups. Equivalence in midazolam apparent oral clearance was observed (%GMR flumazenil/placebo, 90% CI 104.8, 94 - 116.6%). Flumazenil partially attenuated oral midazolam pharmacodynamics. Exploratory post hoc analyses revealed that midazolam exposure was 1.9-fold higher in men compared to women., Conclusion: i.v. flumazenil can be used in conjunction with oral midazolam for CYP3A phenotyping.

Typically, the dose-occupancy curves for GABA-A receptor ligands are determined using in vivo binding of [3H]flumazenil. This study describes in vivo binding experiments without the use of tracer ligands. Bound and free fractions were measured directly using a highly sensitive LC/MS/MS detection method after in vivo administration of the GABA-A ligands zolpidem, (RS)-zopiclone, L-838417 and flumazenil, to demonstrate affinity and saturation of the filter-retained, membrane-bound fraction. The in vivo binding of flumazenil and L-838417 both saturated around 200 nM, at a similar level to the specific binding of (S)-zopiclone after doses of the racemic zopiclone, using (R)-zopiclone to estimate non-specific binding. This saturable component represented an estimate of benzodiazepine binding sites available on GABA-A receptors in vivo (200 nM). Dose-occupancy curves were constructed to estimate the dose required to achieve 50% occupancy and matched estimates obtained with tracer methods. In contrast to tracer methods, this method is uniquely suitable to the demonstration of stereoselective binding of the (S)-isomer in vivo after doses of racemic zopiclone. These results demonstrate that the LC/MS/MS measurements of total drug concentrations typically used in early drug development can be adapted to provide information about receptor occupancy in vivo.