Equipotent Doses Research Articles

Serotonin enhances oxybuprocaine- and proxymetacaine-induced cutaneous analgesia in rats

The aim of the study was to investigate the analgesic effects of adding serotonin to oxybuprocaine or proxymetacaine preparations. We employed a rat model of the cutaneous trunci muscle reflex (CTMR) to conduct the dose-response curves and duration of drugs (oxybuprocaine, proxymetacaine, or serotonin) as an infiltrative anesthetic. The use of isobolographic methods to analyze the drug-drug interactions. We showed that oxybuprocaine and proxymetacaine, as well as serotonin produced dose-dependent skin antinociception. On the basis of 50% effective dose (ED50), the rank order of drug potency was serotonin [7.22 (6.45–8.09) μmol/kg] < oxybuprocaine [1.03 (0.93–1.15) μmol/kg] < proxymetacaine [0.59 (0.53–0.66) μmol/kg] (P < 0.01 for each comparison). The sensory block duration of serotonin was longer (P < 0.01) than that of oxybuprocaine or proxymetacaine at the equipotent doses (ED25, ED50, and ED75). The mixture of serotonin with oxybuprocaine or proxymetacaine produced a better analgesic effect than the drug itself. We have concluded that oxybuprocaine, proxymetacaine, or serotonin displays dose-related cutaneous analgesia. Oxybuprocaine or proxymetacaine is more potent and has a shorter duration of cutaneous analgesia than serotonin. Serotonin produces a synergistic antinociceptive interaction with oxybuprocaine or proxymetacaine.

Therapeutic Targeting of an RNA Splicing Factor Network for the Treatment of Myeloid Neoplasms

RNA-binding proteins (RBPs) regulate many aspects of transcription and translation in a cell- and tissue-specific manner and are frequently dysregulated in malignancy. We systematically evaluated RBPs preferentially required in acute myeloid leukemia (AML) over other forms of cancer or normal hematopoietic precursors using a CRISPR/Cas9 domain-based, loss-of-function screen targeting 490 classical RBPs with 2,900 sgRNAs (Fig. A). This screen was performed in cells lines representing AML, T-cell acute lymphoblastic leukemia (T-ALL), and lung adenocarcinoma (LUAD) and revealed multiple RBPs preferentially required for AML survival, but not for T-ALL or LUAD survival. We identified genes encoding 21 RBPs that were >3-fold depleted in AML cells and significantly overexpressed in AML patient samples versus normal adult CD34+ precursors (p-value < 0.05; Fig. B).Amongst RBPs required and upregulated in AML was RBM39, an RBP described to be involved in a number of cellular processes and to interact with key splicing proteins SF3B1 and U2AF2. Genetic ablation of Rbm39 in mouse MLL-AF9 leukemia cells dramatically delayed AML development and progression (Fig. C). In parallel, it has recently been described that a class of clinically-validated anti-cancer sulfonamide compounds (including indisulam and E7820) mediate RBM39 degradation as their dominant cellular mechanism of action. This occurs via novel interactions with the DCAF15 adapter protein of the CUL4/Ddb1 ubiquitin ligase complex with RBM39 as a neo-substrate. Treatment of MOLM-13 cells xenografted into mice with indisulam conferred significant anti-leukemic effects and improved overall survival (Fig. D).To explore the mechanism of RBM39 dependence in AML, we performed proteomic analyses of RBM39 interacting proteins in MOLM-13 cells as well as transcriptome-wide analysis of RBM39 RNA binding by enhanced UV cross-linking and immunoprecipitation (eCLIP) in the same cells. RBM39 physically interacted with an entire network of RBPs identified by our CRISPR screen as crucial for AML cell survival in addition to interacting with the core SF3b splicing complex. Further, anti-RBM39 eCLIP revealed RBM39 binding to exonic regions and most enriched at exon/intron borders at 5‘ and 3‘ splice sites of pre-mRNA (Fig. E), suggesting a prominent role of RBM39 in regulating splicing. Consistent with this, RNA-sequencing of AML cells following RBM39 deletion revealed significant effects of RBM39 loss on RNA splicing, most prominently causing increased cassette exon skipping (Fig. F).Recent studies suggest that myeloid leukemias with mutations in RNA splicing factors are sensitized to pharmacologic perturbation of RNA splicing. Analysis of the effects of RBM39 degrading compounds over a panel of 18 AML cells revealed that leukemia cells bearing splicing factor mutations or with high DCAF15 expression were the most sensitive to treatment (Fig. G). Genetic introduction of SF3B1, SRSF2, or U2AF1 hotspot mutations in K562 or NALM6 cells resulted in a 20-50% reduction in IC50 in response to sulfonamides.We next performed RNA sequencing of isogenic K562 cells with or without knockin of SF3B1K700E and SRSF2P95H mutations into the endogenous loci, and treated at the IC50 of E7820 or E7107, a small molecule that inhibits the SF3b core spliceosome complex. Treatment with either drug dramatically increased cassette exon skipping events and intron retention relative to DMSO control, with greater effects in splicing mutant cells. However, at equipotent doses, E7820 markedly increased mis-splicing compared with E7107. Furthermore, E7820 treatment resulted in mis-splicing of a number of RBP targets identified in our CRISPR screen as being required for AML survival, including SUPT6H, hnRNPH, and SRSF10, as well as RBM3 and U2AF2, consistent with previous observations (Fig. H).Here through systematic evaluation of RBPs across several cancers, we identify RBPs specifically required in AML. In so doing we identify a network of functionally and physically interacting RBPs upregulated in AML over normal precursors. Genetic or pharmacologic elimination one such RBP, RBM39, led to aberrant splicing of multiple members of this RBP network as well as of transcriptional regulators required for AML survival. These data suggest important clinical potential for anti-cancer sulfonamide treatment in splicing mutant myeloid leukemias. [Display omitted] DisclosuresUehara:Eisai: Employment. Owa:Eisai: Employment.

Anesthetics Have Different Effects on the Electrocorticographic Spectra of Wild-type and Mitochondrial Mutant Mice.

WHAT THIS ARTICLE TELLS US THAT IS NEW: BACKGROUND:: Knockout of the mitochondrial protein Ndufs4 (Ndufs4[KO]) in mice causes hypersensitivity to volatile anesthetics but resistance to ketamine. The authors hypothesized that electrocorticographic changes underlying the responses of Ndufs4(KO) to volatile anesthetics and to ketamine would be similar in mutant and control mice. Electrocorticographic recordings at equipotent volatile anesthetic concentrations were compared between genotypes. In separate studies, control and cell type-specific Ndufs4(KO) mice were anesthetized with intraperitoneal ketamine to determine their ED50s. Ndufs4 (KO) did not differ from controls in baseline electrocorticography (N = 5). Compared to baseline, controls exposed to isoflurane (EC50) lost power (expressed as mean baseline [µV/Hz]; mean isoflurane [µV/Hz]) in delta (2.45; 0.50), theta (1.41; 0.16), alpha (0.23; 0.05), beta (0.066; 0.016), and gamma (0.020; 0.005) frequency bands (N = 5). Compared to baseline, at their isoflurane EC50, Ndufs4(KO) maintained power in delta (1.08; 1.38), theta (0.36; 0.26), and alpha (0.09; 0.069) frequency bands but decreased in beta (0.041; 0.023) and gamma (0.020; 0.0068) frequency bands (N = 5). Similar results were seen for both genotypes in halothane. Vesicular glutamate transporter 2 (VGLUT2)-specific Ndufs4(KO) mice were markedly resistant to ketamine (ED50; 125 mg/kg) compared to control mice (ED50; 75 mg/kg; N = 6). At their respective ED95s for ketamine, mutant (N = 5) electrocorticography spectra showed a decrease in power in the beta (0.040; 0.020) and gamma (0.035; 0.015) frequency bands not seen in controls (N = 7). Significant differences exist between the electrocorticographies of mutant and control mice at equipotent doses for volatile anesthetics and ketamine. The energetic state specifically of excitatory neurons determines the behavioral response to ketamine.

Abstract 067: Mechanism of Action of 8-Aminoguanine on Renal Excretory Function

The endogenous purines 8-aminoguanosine and 8-aminoguanine (8-AG) are K + -sparing natriuretics that increase glucose excretion and attenuate salt-induced hypertension. Most effects of 8-aminoguanosine are not direct, but require conversion in the systemic circulation to 8-AG (i.e., 8-aminoguanosine is a prodrug/prohormone). However, the mechanism of action by which 8-AG affects renal excretory function is unknown and is the subject of this investigation. Because 8-AG has structural similarities with inhibitors of epithelial Na + channels (ENaC), Na + /H + exchangers (NHE), and adenosine A 1 receptors, we examined the effects of 8-AG on amiloride-sensitive ENaC activity in mouse collecting duct cells, on intracellular pH of human renal proximal tubule epithelial cells, and on in vivo (in rats) pharmacological responses to 2-chloro-N 6 -cyclopentyladenosine (A 1 -receptor agonist). 8-AG did not block ENaC, NHE, or A 1 receptors. Because Rac1 enhances activity of mineralocorticoid receptors and some guanosine analogues inhibit Rac1, we examined the effects of 8-AG on Rac1 activity in mouse collecting duct cells. Rac1 activity was significantly ( P =0.024) inhibited by 8-AG (30 μM; from 2.5 ± 0.7 to 1.7 ± 0.7 arbitrary units). Because 8-AG is an inhibitor of purine nucleoside phosphorylase (PNPase; metabolizes inosine to hypoxanthine and guanosine to guanine), we compared the renal effects (in rats) of approximately equipotent intravenous doses of 8-AG (33.5 μmol/kg) vs. 9-deazaguanine (9-DG; 67 μmol/kg; PNPase inhibitor). 8-AG and 9-DG induced similar increases in urinary Na + (μmol/30 min; 8-AG, from 4.4 ± 2.0 to 38 ± 17; 9-DG, from 5.6 ± 3.3 to 34 ± 8.6) and glucose (μg/30 min; 8-AG, from 31 ± 17 to 225 ± 77; 9-DG, from 6.9 ± 3.9 to 144 ± 81) excretion. Intravenous 8-AG increased urinary excretion of guanosine and inosine (PNPase substrates), yet decreased excretion of guanine and hypoxanthine (PNPase products). 8-AG reduced K + excretion (μmol/30 min; 8-AG, from 29 ± 6.6 to 5.4 ± 1.7) whereas 9-DG did not. However, the Rac1 inhibitor Nsc23766 (9.4 μmol/kg) mimicked the effects of 8-AG on K + excretion (μmol/30 min; from 40 ± 7.1 to 18 ± 6.5). Conclusion: Likely 8-AG increases Na + and glucose excretion by blocking PNPase and decreases K + excretion by inhibiting Rac1.

Equipotent dose of levobupivacaine reduce the incidence of instrumental vaginal delivery when compared to ropivacaine during epidural labour analgesia

Background : Instrumental vaginal delivery is a major concern during epidural labour analgesia. In previous studies levobupivacaine 0.1% was associated with increase in instrumental vaginal delivery when compared to 0.1% ropivacaine. Reducing the concentration of levobupivacaine may decrease the incidence of instrumental delivery. Hence, we compared 0.08% levobupivacaine and 0.1% ropivacaine with fentanyl as adjuvant for epidural labour analgesia in terms of mode of delivery. M ethods: This prospective randomised controlled trial was conducted on 70 nulliparous parturients with singleton uncomplicated pregnancy. They were randomized into two groups to receive 0.08% levobupivacaine or 0.1% ropivacaine with 2mg/ml fentanyl as intermittent epidural boluses. The epidural analgesia was initiated with 12ml of study drug solution in active stage of labour and was maintained by repeated demand boluses whenever the Visual Analogue Scale (VAS) score was > 3. Onset, duration and quality of analgesia, degree of motor blockade was analysed. Primary outcome measure was the mode of delivery. Onset and duration of analgesia and Apgar score of baby were the secondary outcomes. Results: Instrumental vaginal delivery was 6.45% in levobupivacaine(L) and 4.54%in ropivacaine(R) group. However,11.4% and 37.1% parturients underwent caesarean section in group L and group R respectively (p=0.012**). Mean onset of analgesia and duration of analgesia was comparable. More than 80% of parturients had excellent pain relief in both the groups with good baby APGAR score. Conclusion: We conclude that 12ml of intermittent epidural boluses of 0.08% levobupivacaine and 0.1% ropivacaine with 2mg/ml fentanyl (equipotent dose) provided equal and effective analgesia during labour with comparable incidence of instrumental vaginal delivery.

Don’t Hang Up Your Lead, Yet

Three botulinum neurotoxin type A (BoNT/A) products, incobotulinumtoxinA, onabotulinumtoxinA, and abobotulinumtoxinA, all manufactured by different methods, are employed in clinical practice. Comparing the three BoNT/A products is difficult because their concentrations and volumes differ and the precise dose equivalence ratio is not known. We aimed to compare the neurotoxic potencies by a systematic analysis of injected volume and dose.The potency of BoNT in inducing hind limb paresis was assessed by analyzing the wheel-running performance of mice. To standardize the volume, the effect of an identical dose of incobotulinumtoxinA dissolved in different volumes of saline (15, 10, 5, and 2 μl) was studied in four groups of mice (n = 13–15). The potencies of the BoNT products were then compared by injecting identical volumes (10 μl) containing different doses into both hind leg muscles.Mice injected with incobotulinumtoxinA showed a volume-dependent reduction in wheel-running, with larger volumes inducing more intense paresis. A standardized volume containing the same number of mouse units of the BoNT/A products produced different degrees of paresis. The conversion ratio of incobotulinumtoxinA and onabotulinumtoxinA is estimated to be between 1:0.75 and 1:0.5. OnabotulinumtoxinA displayed a two-fold greater potency than abobotulinumtoxinA. Doses of onabotulinumtoxinA and abobotulinumtoxinA that produce an identical severity of pareses even result in the same duration of pareses.This wheel-running assay allows one to compare the neurotoxic potency of different volumes and doses of the BoNT products in vivo. Our results argue against common clinical practice because incobotulinumtoxinA and onabotulinumtoxinA are not readily interchangeable and a two-fold dose of abobotulinumtoxinA is needed to induce an effect identical to onabotulinumtoxinA. In addition, this emphasizes that the duration of BoNT-induced effect is the same as long as equipotent doses of BoNT are injected.

Comparison of cardiotoxicity of levobupivacaine and bupivacaine with the brain natriuretic peptide

Introduction: Local anesthetics are also considered to be potentially toxic to the cardiovascular system since they act on ion channels in nerve cell membranes as well as in other tissues that can be stimulated. Aim : In this study we aimed to compare cardiotoxicity levels of both drugs with brain natriuretic peptide (BNP) protein level in patients undergoing combined spinal epidural anesthesia and postoperative analgesia when given bupivacaine and levobupivacaine at equipotent doses. Materyal and Method: In this prospective, randomized, double-blind study a total of 30 patients, aged 25-70 years, who were included in the American Society of Anesthesiologists (ASA) I-III risk class and underwent total knee arthroplasty surgery, were included in the study. Patients were randomized as Group I (n = 15) Levobupivacaine and Group II (n = 15) bupivacaine and combined spinal-epidural anesthesia was administered. The operation follow-up of patients were evaluated with the systolic arterial pressure (SAB), the diastolic arterial pressure (DAB), the mean arterial pressure (MAP) and the heart rate (HR), and postoperative follow-up with SAB, DAB, KH, visual analogue scale (VAS) values were monitored at zero minute, 1st, 6th, 12th and 24th hour. BNP levels were measured at 5th minute after spinal anesthesia, sensory block at thoracal level 10 and post-operative 24th hour. Results: A statistically significant difference was found in the VAS values ​​measured at postoperative 12th hour and 24th hour in favor of levobupivacaine group (p <0.05). In the comparison of BNP levels between the groups, there was no statistically significant difference between the two groups at all measurement times (p> 0.05). Conclusion : Similar results were obtained in intraoperative and postoperative BNP levels when using levobupivacaine and bupivacaine in equal doses for both spinal anesthesia and epidural analgesia.

Correction: Comparison of desflurane and propofol at equipotent doses in combination with constant rate infusion of dexmedetomidine on global and peripheral perfusion and oxygenation in horses

Correction: Comparison of desflurane and propofol at equipotent doses in combination with constant rate infusion of dexmedetomidine on global and peripheral perfusion and oxygenation in horses

High Dose of Pralidoxime Reverses Paraoxon-Induced Respiratory Toxicity in Mice.

The efficiency of pralidoxime in the treatment of human organophosphates poisoning is still unclear. In a rat model, we showed that pralidoxime induced a complete but concentration-dependent reversal of paraoxon-induced respiratory toxicity. The aim of this study was to assess the efficiency of pralidoxime in a species other than rats. A dose of diethylparaoxon corresponding to 50% of the median lethal dose was administered subcutaneously to male F1B6D2 mice. Ascending single pralidoxime doses of 10, 50-100 and 150 mg kg-1 were administered intramuscularly 30 min after diethylparaoxon administration. Ventilation at rest was assessed using whole-body plethysmography and mice temperature was assessed using infrared telemetry. Results are expressed as mean±SE. Statistical analysis used non-parametric tests. From 30 to 150 min post-injection, diethylparaoxon induced clinical symptoms and a decrease in respiratory frequency, which resulted from an increase in expiratory and inspiratory times associated with an increase in the tidal volume. In the 10-, 50- and 100-mg kg-1 pralidoxime groups, there was a trend towards a non-significant improvement of paraoxon-induced respiratory toxicity. The 150 mg kg-1 dose of pralidoxime induced a significant reversal of all respiratory parameters. In the present study, a toxic but non-lethal model of diethylparaoxon in awake, unrestrained mice was observed. By administering an equipotent dose of diethylparaoxon to rats, a 150 mg kg-1 dose of pralidoxime administered alone completely reversed diethylparaoxon-induced respiratory toxicity in mice. The dose dependency of reversal suggests that further studies are needed for assessing plasma concentrations of pralidoxime resulting in reversal of toxicity.

Vascular brain-derived neurotrophic factor pathway in rats with adjuvant-induced arthritis: Effect of anti-rheumatic drugs

Background and aimsIn rheumatoid arthritis, the control of both disease activity and standard cardiovascular (CV) risk factors is expected to attenuate the increased CV risk. Evidence that brain-derived neurotrophic factor (BDNF) plays a role in vascular biology led us to investigate the vascular BDNF pathway in arthritis rats as well as the interaction between endothelial nitric oxide (NO) and BDNF production. MethodsThe aortic BDNF pathway was studied in rats with adjuvant-induced arthritis, (AIA) using Western blot and immunohistochemical analysis. Control of arthritis score was achieved by administration (for 3 weeks) of an equipotent dosage of etanercept, prednisolone, methotrexate, celecoxib or diclofenac. Aortas were exposed to an NO donor or an NO synthase inhibitor and vasoreactivity experiments were performed using LM22A-4 as a TrkB agonist. ResultsVascular BDNF and full length tropomyosin-related kinase B receptor (TrkB-FL) were higher in AIA than in control rats. These changes coincided with decreased endothelial immunoreactivity in BDNF and pTrkBtyr816 and were disconnected from arthritis score. Among anti-rheumatic drugs, only prednisolone and methotrexate prevented AIA-induced vascular BDNF loss. The effect of AIA on aortic BDNF levels was reversed by an NO donor and reproduced by an NOS inhibitor. Finally, LM22A-4 induced both NO-dependent vasodilation and phosphorylation of endothelial NO synthase at serine 1177. ConclusionsOur study identified changes in the BDNF/TrkB pathway as a disease activity-independent component of AIA-associated changes in endothelial phenotype. It provides new perspectives in the understanding and management of the high CV risk reported in rheumatoid arthritis.

Cardiovascular and respiratory effects of two doses of fentanyl in the presence or absence of bradycardia in isoflurane-anesthetized dogs

ObjectiveTo compare the cardiopulmonary effects of low and high doses of fentanyl before and after the correction of bradycardia in isoflurane-anesthetized dogs. Study designProspective, randomized crossover trial. AnimalsEight healthy male Beagle dogs weighing 11.1 ± 1.3 kg [mean ± standard deviation (SD)] and aged approximately 1 year. MethodsThe dogs were anesthetized with isoflurane [1.3 × minimum alveolar concentration (MAC)] on two occasions and fentanyl was administered intravenously; either low-dose fentanyl, loading dose (33 μg kg–1) and infusion (0.2 μg kg–1 minute–1) or a high-dose, loading dose (102 μg kg–1) and infusion (0.8 μg kg–1 minute–1). Cardiopulmonary variables were measured at three time points in equipotent isoflurane concentrations (1.3 MAC): before fentanyl administration (ISO), during fentanyl-induced bradycardia (ISO–F) and after administration of glycopyrrolate normalized heart rate (ISO–FNHR). Data are mean ± SD. ResultsHeart rate and cardiac index (CI) decreased and systemic vascular resistance index (SVRI) increased at ISO–F in both treatments. Bradycardia and vasoconstriction at ISO–F were greater in high than in low-dose fentanyl (42 ± 7 versus 57 ± 15 beats minute–1 and 3457 ± 1108 versus 2528 ± 968 dyne second cm–5 m–2), respectively. Oxygen delivery index (DO2I) decreased only during high-dose fentanyl. CI and DO2I were higher in both treatments at ISO–FNHR than at ISO–F; however, they were higher only during the high-dose fentanyl than at ISO. SVRI was higher at ISO–F than at ISO and ISO–FNHR in both treatments, and was higher at ISO–F in the high than in the low-dose treatment. Conclusions and clinical relevanceAn overall improvement in cardiovascular function of dogs anesthetized with equipotent isoflurane doses (1.3 MAC) was observed after the treatment of bradycardia only with the high-dose fentanyl.

Efficacy of agglutination test in diagnosis of motile Aeromonas septicemia in Nile tilapia (Oreochromis niloticus)

The time-course of the neuromuscular effects of vecuronium (n = 25) and atracurium (n = 25) has been compared at three different levels of maintenance dose in anaesthetized patients. Following intubation with vecuronium 0.1 mg kg-1 or atracurium 0.5 mg kg-1, surgical muscle relaxation was maintained by using increments of equipotent maintenance doses equivalent to 0.5, 1.0 and 1.5 x ED95 for each drug. Repeat doses were administered each time the twitch height, depressed by the previous dose, returned to 25% of its control value. The apparent increase in the duration of action, i.e. the difference between the duration of the last and the first maintenance dose, did not reach statistical significance and approximated 3 +/- 2, 6 +/- 4, 11 +/- 5 and 3 +/- 2, 8 +/- 13, 5 +/- 7 min following the low, medium and high maintenance doses of vecuronium and atracurium, respectively.

Intrathecal pramoxine causes long-lasting spinal sensory and motor block in rats.

The objective of this experiment was to investigate spinal anaesthetic effects of pramoxine and its comparison with bupivacaine, a long-lasting local anaesthetic. After intrathecal injection, three neurobehavioural assessments, which consisted of nociceptive, proprioceptive and motor block, were constructed in rats. The effects of bupivacaine and pramoxine (four doses of each drug) in a dose-related manner were conducted to obtain the ED50 (50% effective dose). Pramoxine potency and duration at provoking spinal nociceptive, proprioceptive and motor block were compared with those of bupivacaine. We manifested that pramoxine provoked dose-relatedly spinal blockades of nociception, proprioception and motor function. Based on the ED50 , the rank potency at producing spinal nociceptive, proprioceptive and motor block was bupivacaine (0.90 (0.82-1.02), 1.00 (0.92-1.08) and 1.16 (1.02-1.34) μmol/kg) greater (P < 0.01 for the differences) than pramoxine (15.47 (14.04-17.05), 16.46 (15.06-17.99), and 17.77 (16.48-19.15) μmol/kg). The spinal block duration created by bupivacaine was not predominantly different (P > 0.05 for the differences) from that created by pramoxine at the equipotent doses (ED75 , ED50 and ED25 ). Our preclinical experiment indicated that pramoxine elicited a dose-related spinal block, was less potent than bupivacaine and had a similar duration of spinal block compared with bupivacaine.

A neurosteroid analogue with T-type calcium channel blocking properties is an effective hypnotic, but is not harmful to neonatal rat brain

BackgroundMore than 4 million children are exposed annually to sedatives and general anaesthetics (GAs) in the USA alone. Recent data suggest that common GAs can be detrimental to brain development causing neurodegeneration and long-term cognitive impairments. Challenged by a recent US Food and Drug Administration (FDA) warning about potentially neurotoxic effects of GAs in children, there is an urgent need to develop safer GAs. MethodsPostnatal Day 7 (P7) rat pups of both sexes were exposed to six (repeated every 2 h) injections of equipotent hypnotic doses of ketamine or the neuroactive steroid (3β,5β,17β)-3-hydroxyandrostane-17-carbonitrile (3β-OH) for 12 h. Loss of righting reflex was used to assess hypnotic properties and therapeutic index; quantitative caspase-3 immunohistochemistry was used to assess developmental neuroapoptosis; patch-clamp recordings in acute brain slices were used to assess the effects of 3β-OH on neuronal excitability and synaptic transmission. Cognitive abilities of rats exposed to ketamine, 3β-OH, or vehicle at P7 were assessed in young adulthood using the radial arm maze. ResultsThe neuroactive steroid 3β-OH has a therapeutic index similar to ketamine, a commonly used clinical GA. We report that 3β-OH is safe and, unlike ketamine, does not cause neuroapoptosis or impair cognitive development when administered to P7 rat pups. Interestingly, 3β-OH blocks T-type calcium channels and presynaptically dampens synaptic transmission at hypnotically-relevant brain concentrations, but it lacks a direct effect on γ-aminobutyric acid A or glutamate-gated ion channels. ConclusionsThe neurosteroid 3β-OH is a relatively safe hypnotic that warrants further consideration for paediatric anaesthesia.

PHARMACOKINETICS OF SINGLE-DOSE BUPRENORPHINE, BUTORPHANOL, AND HYDROMORPHONE IN THE DOMESTIC FERRET (MUSTELA PUTORIUS FURO)

The objective of this study was to establish the clinical pharmacokinetic profile of 4 different opioid drugs (buprenorphine, butorphanol, hydromorphone, and morphine) in the domestic ferret (Mustela putorius furo). Twenty-four, approximately 1-year-old, male neutered purpose-bred domestic ferrets were used for this study. The ferrets were divided into 4 groups of 6, with a different opioid drug used for each group. A preopioid venous blood sample was obtained via cranial vena cava venipuncture. Following the initial blood collection, a single injection of opioid (hydromorphone 0.1mg/kg, buprenorphine 0.04mg/kg, butorphanol 0.3mg/kg, and morphine 1mg/kg) was given to each ferret, dependent on assigned drug group, intramuscularly (buprenorphine) or subcutaneously (hydromorphone, butorphanol, and morphine). Intramuscular injections were administered in the semimembranosis and semitendinosis muscles, whereas the subcutaneous injections were delivered in the intrascapular subcutaneous space. A venous blood sample was obtained at 5, 15, 30, 60, 120, 240, 360, 480, and 720minutes postinjection from the ferrets in the buprenorphine, butorphanol, and hydromorphone groups. Mass spectrometry and liquid chromatography was performed to obtain plasma concentrations of the administered drugs. The mean maximum concentration of buprenorphine was 6.96ng/mL, butorphanol was 48.6ng/mL, and hydromorphone was 17.3ng/mL. Maximum concentrations were achieved at a mean of 9minutes after administration for buprenorphine, 13.3minutes for butorphanol, and 8.33minutes for hydromorphone. The mean half-life of buprenorphine was 219.1minutes, butorphanol was 91.1minutes, and hydromorphone was 24.7minutes. Owing to severe complications arising within the morphine group, including hypersalivation and vomiting, the morphine study was discontinued prior to blood sample collection. Intramuscular injections of buprenorphine and subcutaneous injections of butorphanol or hydromorphone appeared to be well tolerated by all ferrets. The pharmacokinetics of buprenorphine, butorphanol, and hydromorphone of a single equipotent dose of each drug have been established through this research investigation and may be useful for further studies.

Efavirenz pharmacokinetics during pregnancy and infant washout.

Limited data exist on efavirenz pharmacokinetics in HIV-positive pregnant women and neonatal washout. HIV-infected pregnant women receiving 600 mg efavirenz once daily had intensive steady-state 24-h pharmacokinetics profiles during the second trimester (2T), third trimester (3T) and 6-12 weeks postpartum (PP). Maternal and umbilical cord blood samples were drawn at delivery and neonatal washout pharmacokinetics were determined. Therapeutic targets were the estimated 10th percentile efavirenz area under the concentration-time curve (AUC) in non-pregnant historical controls (40.0 μg•h/ml) and a trough concentration (C24 h) of 1 μg/ml. Data were prospectively collected within two trials: IMPAACT P1026s (United States) and PANNA (Europe). Among 42 women studied, 15, 42 and 40 had efavirenz pharmacokinetic data available in 2T, 3T and PP, respectively. Median (range) 3T age 33 (20.7-43.5) years, weight 74 (50-132) kg and gestational age 33.4 (28.4-37.9 weeks). Efavirenz AUC during the 3T (60 μg•h/ml) was similar to that reported in non-pregnant adults (58 μg•h/ml). Exposure in the 2T was lower, but within the 0.80-1.25 range. C24 concentrations during pregnancy were lower compared to historical controls on 600 mg efavirenz, however, they were similar to the C24 concentrations after equally potent dose of 400 mg efavirenz. Cord blood/maternal plasma concentration ratio (range) was 0.67 (0.36-0.95). Among 23 infants with washout data available, median (interquartile range) elimination half-life was 65.6 h (40.6-129). HIV RNA viral loads at delivery were <400 and <50 copies/ml for 96.7% and 86.7% of women, respectively. In 3T and PP, respectively, 8/41 (19%) and 6/40 (15%) had AUC below target; 7/41 (17%) and 3/39 (8%) had C24 below target. Efavirenz exposure was similar during pregnancy compared with PP, C24 was in line with C24 after 400 mg equipotent efavirenz dosing. Efavirenz readily crossed the placenta and infant elimination half-life was over twice that of maternal participants. Clincaltrials.gov identifiers: NCT00825929 and NCT00042289.

Specific inhibition of acetylcholinesterase as an approach to decrease muscarinic side effects during myasthenia gravis treatment

Non-selective inhibitors of cholinesterases (ChEs) are clinically used for treatment of myasthenia gravis (MG). While being generally safe, they cause numerous adverse effects including induction of hyperactivity of urinary bladder and intestines affecting quality of patients life. In this study we have compared two ChEs inhibitors, a newly synthesized compound C547 and clinically used pyridostigmine bromide, by their efficiency to reduce muscle weakness symptoms and ability to activate contractions of urinary bladder in a rat model of autoimmune MG. We found that at dose effectively reducing MG symptoms, C547 did not affect activity of rat urinary bladder. In contrast, at equipotent dose, pyridostigmine caused a significant increase in tonus and force of spontaneous contractions of bladder wall. We also found that this profile of ChEs inhibitors translates into the preparation of human urinary bladder. The difference in action observed for C547 and pyridostigmine we attribute to a high level of pharmacological selectivity of C547 in inhibiting acetylcholinesterase as compared to butyrylcholinesterase. These results raise reasonable hope that selective acetylcholinesterase inhibitors should show efficacy in treating MG in human patients with a significant reduction in adverse effects related to hyperactivation of smooth muscles.

Ceftriaxone concentration at the surgical site following systemic and isolated upper limb injection.

Background and Aims:This study was conducted to find out the equipotent dose of isolated upper limb injection of ceftriaxone in the upper limb (IUL) surgeries under tourniquet that would attain a peak bone marrow concentration (Cmax) similar to systemic (ST) 1 g injection.Material and Methods:Patients were allocated into two groups – ST and IUL. ST group (n = 5) received 1 g of ceftriaxone 20 min before tourniquet inflation, and IUL group received calculated dose (n = 5 in each dosage, i.e., 200, 100, 75, and 50 mg) diluted in 50 mL of normal saline distally after tourniquet inflation. Venous and bone marrow samples were collected at various time intervals intra- and post-operatively. Ceftriaxone concentration was analyzed by high-performance liquid chromatography.Results:There was no significant difference between Cmax following ST 1 g injection and IUL injection with 75 mg (155.8 ± 2.1 vs 158.5 ± 3.1 μg/mL, respectively; P = 0.1). There was significant difference in area under curve (AUC) and t½ between ST 1 g injection and IUL injection with 75 mg of ceftriaxone (AUC 1285 ± 67 vs 784.4 ± 28 μg/mL/h, respectively; P < 0.001), (t½ 5.2 ± 0.5 vs 4.7 ± 0.3 h, respectively; P < 0.001). None of the patients in the ST and IUL groups had post-operative infection up to a period of 1 week duration.Conclusion:IUL injection with 75 mg of ceftriaxone can be equipotent and as effective as ST 1 g injection in upper limb orthopedic surgeries under tourniquet.

Subcutaneous infiltration of doxylamine on cutaneous analgesia in rats.

We aimed to evaluate the effect of doxylamine, a first generation antihistamine, as a local analgesic agent by comparing its effect to bupivacaine. After blocking the cutaneous trunci muscle reflex (CTMR) by subcutaneous injection of doxylamine, we assessed doxylamine's cutaneous analgesic effect in rats. The dose-related effect and duration of doxylamine on infiltrative cutaneous analgesia were compared with that of bupivacaine. We demonstrated that doxylamine, as well as the local anesthetic bupivacaine produced the cutaneous analgesic effects in a dose-related fashion. At the equipotent dose (50% effective doses (ED50)), the relative potency was bupivacaine (0.41 (0.36-0.48) mmol)> doxylamine (7.39 (6.91-7.91)mmol) (p<0.01). On an equipotent basis (ED25, ED50 and ED75), subcutaneous doxylamine resulted in greater duration of action (p<0.01) than bupivacaine at producing cutaneous analgesia. The result of this experiment indicated that doxylamine has the local anesthetic property less potent than bupivacaine, but its nociceptive block duration is longer than that of bupivacaine at an equianalgesic dose.

Synergistic antitumor interaction between valproic acid, capecitabine and radiotherapy in colorectal cancer: critical role of p53

BackgroundRecurrence with distant metastases has become the predominant pattern of failure in locally advanced rectal cancer (LARC), thus the integration of new antineoplastic agents into preoperative fluoropyrimidine-based chemo-radiotherapy represents a clinical challenge to implement an intensified therapeutic strategy.The present study examined the combination of the histone deacetylase inhibitor (HDACi) valproic acid (VPA) with fluoropyrimidine-based chemo-radiotherapy on colorectal cancer (CRC) cells.MethodsHCT-116 (p53-wild type), HCT-116 p53−/− (p53-null), SW620 and HT29 (p53-mutant) CRC cell lines were used to assess the antitumor interaction between VPA and capecitabine metabolite 5′-deoxy-5-fluorouridine (5′-DFUR) in combination with radiotherapy and to evaluate the role of p53 in the combination treatment. Effects on proliferation, clonogenicity and apoptosis were evaluated, along with γH2AX foci formation as an indicator for DNA damage.ResultsCombined treatment with equipotent doses of VPA and 5′-DFUR resulted in synergistic effects in CRC lines expressing p53 (wild-type or mutant). In HCT-116 p53−/− cells we observed antagonist effects. Radiotherapy further potentiated the antiproliferative, pro-apoptotic and DNA damage effects induced by 5′-DFUR/VPA combination in p53 expressing cells.ConclusionsThese results highlighted the role of VPA as valuable candidate to be added to preoperative chemo-radiotherapy in LARC. On these bases we launched the ongoing phase I/II study of VPA and short-course radiotherapy plus capecitabine as preoperative treatment in low-moderate risk rectal cancer (V-shoRT-R3).