A Novel Role for the Histone Demethylase JMJD3 in Mediating Heroin-induced Relapse-Like Behaviors.

BACKGROUND: Epigenetic changes, leading to long-term neuroadaptations following opioid exposure are not well understood. We examined how histone demethylase JMJD3 in the nucleus accumbens (NAc) influences heroin seeking after abstinence from self-administration. METHODS: Male Sprague-Dawley rats were trained to self-administer heroin. Western blotting and qPCR were performed to quantify JMJD3 and bone morphogenetic protein (BMP) pathway expression in the NAc (n = 7-11/group). Pharmacological inhibitors or viral expression vectors were microinfused into the NAc to manipulate JMJD3 or the BMP pathway member SMAD1 (n = 9-11/group). The RiboTag capture method (n = 3-5/group) and viral vectors (n = 7-8/group) were used in male transgenic rats to identify the contributions of D1- and D2-type medium spiny neurons (MSN) in the NAc. Drug-seeking was tested by cue-induced response previously paired with drug infusion. RESULTS: Levels of JMJD3 and phosphorylated SMAD1/5 in the NAc were increased after 14 days of abstinence from heroin self-administration. Pharmacological and virus-mediated inhibition of JMJD3 or the BMP pathway attenuated cue-induced seeking. Pharmacological inhibition of BMP signaling reduced JMJD3 expression and histone 3 lysine 27 trimethylation (H3K27me3) levels. JMJD3 bidirectionally affected seeking: expression of the wild type increased whereas expression of a catalytic dead mutant decreased cue-induced seeking. JMJD3 expression was increased in D2+ but not D1+ MSNs. Expression of the mutant JMJD3 in D2+ neurons was sufficient to decrease cue-induced heroin seeking. CONCLUSIONS: JMJD3 mediates persistent cellular and behavioral adaptations underlying heroin relapse and this activity is regulated by the BMP pathway.

EGR3 regulates opioid-related nociception and motivation in male rats.

Chronic pain can be a debilitating condition, leading to profound changes in nearly every aspect of life. However, the reliance on opioids such as oxycodone for pain management is thought to initiate dependence and addiction liability. The neurobiological intersection at which opioids relieve pain and possibly transition to addiction is poorly understood. Using RNA sequencing pathway analysis in rats with complete Freund's adjuvant (CFA)-induced chronic inflammation, we found that the transcriptional signatures in the medial prefrontal cortex (mPFC; a brain region where pain and reward signals integrate) elicited by CFA in combination with oxycodone differed from those elicited by CFA or oxycodone alone. However, the expression of Egr3 was augmented in all animals receiving oxycodone. Furthermore, virus-mediated overexpression of EGR3 in the mPFC increased mechanical pain relief but not the affective aspect of pain in animals receiving oxycodone, whereas pharmacological inhibition of EGR3 via NFAT attenuated mechanical pain relief. Egr3 overexpression also increased the motivation to obtain oxycodone infusions in a progressive ratio test without altering the acquisition or maintenance of oxycodone self-administration. Taken together, these data suggest that EGR3 in the mPFC is at the intersection of nociceptive and addictive-like behaviors.

Effects of alcohol on brain oxygenation and brain hypoxia induced by intravenous heroin.

Alcohol is the most commonly used psychoactive drug, often taken in conjunction with opioid drugs. Since both alcohol and opioids can induce CNS depression, it is often assumed that alcohol potentiates the known hypoxic effects of opioid drugs. To address this supposition, we used oxygen sensors to examine the effects of alcohol on brain oxygenation and hypoxic responses induced by intravenous heroin in awake, freely moving rats. To eliminate robust sensory effects of alcohol following its oral or intraperitoneal delivery, alcohol was administered directly into the stomach via chronically implanted intragastric catheters at human relevant doses. Alcohol delivered at a 0.5 g/kg dose did not affect brain oxygen levels, except for a weak transient increase during drug delivery. This phasic oxygen increase was stronger at a 2.0 g/kg alcohol dose and followed by a weaker tonic increase. Since alcohol absorption from intragastric delivery is much slower and more prolonged than with intraperitoneal or intravenous injections, the rapid rise of brain oxygen levels suggests that alcohol has a direct action on sensory afferents in the stomach well before the drug physically reaches brain tissue via circulation. Despite slow tonic increases in brain oxygen, alcohol at the 2.0 g/kg dose strongly potentiates heroin-induced oxygen responses, increasing both the magnitude and duration of oxygen decrease. Therefore, under the influence of alcohol, the use of opioid drugs becomes much more dangerous, increasing brain hypoxia and enhancing the probability of serious health complications, including coma and death.

Relationships between oxygen changes in the brain and periphery following physiological activation and the actions of heroin and cocaine.

Using two-sensor electrochemical recordings in freely moving rats, we examined the relationship between physiological and drug-induced oxygen fluctuations in the brain and periphery. Animals chronically implanted with oxygen sensors in the nucleus accumbens (NAc) and subcutaneous (SC) space were subjected to several mildly arousing stimuli (sound, tail-pinch and social interaction) and intravenous injections of cocaine and heroin. Arousing stimuli induced rapid increases in NAc oxygen levels followed by and correlated with oxygen decreases in the SC space. Therefore, cerebral vasodilation that increases cerebral blood flow and oxygen entry into brain tissue results from both direct neuronal activation and peripheral vasoconstriction, which redistributes arterial blood from periphery to the brain. The latter factor could also explain a similar pattern of oxygen responses found in the substantia nigra reticulata, suggesting hyperoxia as a global phenomenon with minor structural differences during early time intervals following the stimulus onset. While arousing stimuli and cocaine induced similar oxygen responses in the brain and SC space, heroin induced a biphasic down-up brain oxygen fluctuation associated with a monophasic oxygen decrease in the SC space. Oxygen decreases occurred more rapidly and stronger in the SC space, reflecting a drop in blood oxygen levels due to respiratory depression.

A role for the endocannabinoid enzymes monoacylglycerol and diacylglycerol lipases in cue-induced cocaine craving following prolonged abstinence.

Following exposure to drugs of abuse, long-term neuroadaptations underlie persistent risk to relapse. Endocannabinoid signaling has been associated with drug-induced neuroadaptations, but the role of lipases that mediate endocannabinoid biosynthesis and metabolism in regulating relapse behaviors following prolonged periods of drug abstinence has not been examined. Here, we investigated how pharmacological manipulation of lipases involved in regulating the expression of the endocannabinoid 2-AG in the nucleus accumbens (NAc) influence cocaine relapse via discrete neuroadaptations. At prolonged abstinence (30 days) from cocaine self-administration, there is an increase in the NAc levels of diacylglycerol lipase (DAGL), the enzyme responsible for the synthesis of the endocannabinoid 2-AG, along with decreased levels of monoacylglycerol lipase (MAGL), which hydrolyzes 2-AG. Since endocannabinoid-mediated behavioral plasticity involves phosphatase dysregulation, we examined the phosphatase calcineurin after 30 days of abstinence and found decreased expression in the NAc, which we demonstrate is regulated through the transcription factor EGR1. Intra-NAc pharmacological manipulation of DAGL and MAGL with inhibitors DO-34 and URB-602, respectively, bidirectionally regulated cue-induced cocaine seeking and altered the phosphostatus of translational initiation factor, eIF2α. Finally, we found that cocaine seeking 30 days after abstinence leads to decreased phosphorylation of eIF2α and reduced expression of its downstream target NPAS4, a protein involved in experience-dependent neuronal plasticity. Together, our findings demonstrate that lipases that regulate 2-AG expression influence transcriptional and translational changes in the NAc related to drug relapse vulnerability.

Cocaine added to heroin fails to affect heroin-induced brain hypoxia.

Heroin and cocaine are both highly addictive drugs that cause unique physiological and behavioral effects. These drugs are often co-administered and cocaine has been found in ~20% of cases of opioid overdose death. Respiratory depression followed by brain hypoxia is the most dangerous effect of high-dose opioids that could result in coma and even death. Conversely, cocaine at optimal self-administering doses increases brain oxygen levels. Considering these differences, it is unclear what pattern of oxygen changes will occur when these drugs are co-administered. Here, we used high-speed amperometry with oxygen sensors to examine changes in oxygen concentrations in the nucleus accumbens (NAc) induced by intravenous (iv) cocaine, heroin, and their mixtures in freely-moving rats. Cocaine delivered at a range of doses, both below (0.25 mg/kg) and within the optimal range of self-administration (0.5 and 1.0 mg/kg) modestly increased NAc oxygen levels. In contrast, heroin increased oxygen levels at a low reinforcing dose (0.05 mg/kg), but induced a biphasic down-up change at higher reinforcing doses (0.1 and 0.2 mg/kg), and caused a strong monophasic oxygen decrease during overdose (0.6 mg/kg). When combined at moderate doses, cocaine (0.25, 0.5 mg/kg) slightly increased and prolonged oxygen increases induced by heroin alone (0.5 and 0.1 mg/kg), but oxygen decreases were identical when cocaine (1 mg/kg) was combined with heroin at large doses (0.2 and 0.6 mg/kg). Therefore, health dangers of speedball may result from de-compensation of vital functions due to diminished intra-brain oxygen inflow induced by high-dose heroin coupled with enhanced oxygen use induced by cocaine.

Role of videolaryngoscope in the management of difficult airway in adults: A survey.

BACKGROUND AND AIMS: A number of videolaryngoscopes (VLs) have flooded the Indian market. As per All India Difficult Airway Association 2016 guidelines, all anaesthesiologists should have access to a VL and must be trained to use it. We conducted an electronic survey to know the perception of Indian anaesthesiologists, who are members of the Indian Society of Anaesthesiologists (Karnataka State Chapter) towards the role of VL in the management of difficult airway (DA) and factors governing their use. METHODS: An electronic survey was sent to 2580 ISA members to know the availability, use and attitude towards VLs in the management of DA in adults. The survey was open for a period of 2 months and responses analysed. RESULTS: The response rate was 25.8% (666 out of 2580). A total of 280 (42%) respondents had access to VL. The respondents rated VL as 4th preference for anticipated DA and 1st for unanticipated DA (if available). The most widely used VLs were C-MAC, Airtraq, and Kingvision. As per 133 respondents (20%), access to VL in institutes was restricted only to consultants and the main reason being cost. The clarity of the image was the most important factor the respondents expected in a VL. CONCLUSIONS: Less than half of respondents had access to VLs. Most of them having access to it worked in corporate hospitals. The high cost of the device and steep learning curve are still barriers against its widespread use. We conclude that low-cost devices, with increased clarity may make usage of VLs frequent and available to residents.

Fecal microbiota analysis of polycystic kidney disease patients according to renal function: A pilot study.

IMPACT STATEMENT: The heterogeneity of the renal disease, therapeutic interventions, and the original cause of the renal failure, all directly affect the microbiota. We delineate in this report the direct effect of decreased renal function on the bacterial composition following stringent criteria to eliminate the possibilities of other confounding factors and dissect the direct effects of the uremic milieu. We analyzed the microbiome following three different approaches to further evaluate the effects of mild, moderate and advanced renal insufficiency on the microbiome. We also present here a detailed functional analysis of the projected altered pathways secondary to changes in the microbiome composition.

E3 Ubiquitin-Protein Ligase SMURF1 in the Nucleus Accumbens Mediates Cocaine Seeking.

BACKGROUND: Substance use disorder is a neurobiological disease characterized by episodes of relapse despite periods of withdrawal. It is thought that neuroadaptations in discrete brain areas of the reward pathway, including the nucleus accumbens, underlie these aberrant behaviors. The ubiquitin-proteasome system degrades proteins and has been shown to be involved in cocaine-induced plasticity, but the role of E3 ubiquitin ligases, which conjugate ubiquitin to substrates, is unknown. Here, we examined E3 ubiquitin-protein ligase SMURF1 (SMURF1) in neuroadaptations and relapse behavior during withdrawal following cocaine self-administration. METHODS: SMURF1 and downstream targets ras homolog gene family, member A (RhoA), SMAD1/5, and Runt-related transcript factor 2 were examined using Western blotting (n = 9-11/group), quantitative polymerase chain reaction (n = 6-9/group), co-immunoprecipitation (n = 9-11/group), tandem ubiquitin binding entities affinity purification (n = 5-6/group), and quantitative chromatin immunoprecipitation (n = 3-6/group) (2 rats/sample). Viral-mediated gene transfer (n = 7-12/group) and intra-accumbal microinjections (n = 9-10/group) were used to examine causal roles of SMURF1 and substrate RhoA, respectively, in cue-induced cocaine seeking. RESULTS: SMURF1 protein expression was decreased, while SMURF1 substrates RhoA and SMAD1/5 were increased, in the nucleus accumbens on withdrawal day 7, but not on withdrawal day 1, following cocaine self-administration. Viral-mediated gene transfer of Smurf1 or constitutive activation of RhoA attenuated cue-induced cocaine seeking, while catalytically inactive Smurf1 enhanced cocaine seeking. Furthermore, SMURF1-regulated, SMAD1/5-associated transcription factor Runt-related transcript factor 2 displayed increased binding at promoter regions of genes previously associated with cocaine-induced plasticity. CONCLUSIONS: SMURF1 is a key mediator of neuroadaptations in the nucleus accumbens following cocaine exposure and mediates cue-induced cocaine seeking during withdrawal.

Chronic kidney disease, uremic milieu, and its effects on gut bacterial microbiota dysbiosis.

Several lines of evidence suggest that gut bacterial microbiota is altered in patients with chronic kidney disease (CKD), though the mechanism of which this dysbiosis takes place is not well understood. Recent studies delineated changes in gut microbiota in both CKD patients and experimental animal models using microarray chips. We present 16S ribosomal RNA gene sequencing of both stool pellets and small bowel contents of C57BL/6J mice that underwent a remnant kidney model and establish that changes in microbiota take place in the early gastrointestinal tract. Increased intestinal urea concentration has been hypothesized as a leading contributor to dysbiotic changes in CKD. We show that urea transporters (UT)-A and UT-B mRNA are both expressed throughout the whole gastrointestinal tract. The noted increase in intestinal urea concentration appears to be independent of UTs' expression. Urea supplementation in drinking water resulted in alteration in bacterial gut microbiota that is quite different than that seen in CKD. This indicates that increased intestinal urea concentration might not fully explain the CKD- associated dysbiosis.

A Novel Role for Oligodendrocyte Precursor Cells (OPCs) and Sox10 in Mediating Cellular and Behavioral Responses to Heroin.

Opiate abuse and addiction have become a worldwide epidemic with great societal and financial burdens, highlighting a critical need to understand the neurobiology of opiate addiction. Although several studies have focused on drug-dependent changes in neurons, the role of glia in opiate addiction remains largely unstudied. RNA sequencing pathway analysis from the prefrontal cortex (PFC) of male rats revealed changes in several genes associated with oligodendrocyte differentiation and maturation following heroin self-administration. Among these genes changed was Sox10, which is regulated, in part, by the chromatin remodeler BRG1/SMARCA4. To directly test the functional role of Sox10 in mediating heroin-induced behavioral plasticity, we selectively overexpressed Sox10 and BRG1 in the PFC. Overexpression of either Sox10 or BRG1 decreased the motivation to obtain heroin infusions in a progressive ratio test without altering the acquisition or maintenance of heroin self-administration. These data demonstrate a critical, and perhaps compensatory, role of Sox10 and BRG1 in oligodendrocytes in regulating the motivation for heroin.

Advanced glycation end products dietary restriction effects on bacterial gut microbiota in peritoneal dialysis patients; a randomized open label controlled trial.

The modern Western diet is rich in advanced glycation end products (AGEs). We have previously shown an association between dietary AGEs and markers of inflammation and oxidative stress in a population of end stage renal disease (ESRD) patients undergoing peritoneal dialysis (PD). In the current pilot study we explored the effects of dietary AGEs on the gut bacterial microbiota composition in similar patients. AGEs play an important role in the development and progression of cardiovascular (CVD) disease. Plasma concentrations of different bacterial products have been shown to predict the risk of incident major adverse CVD events independently of traditional CVD risk factors, and experimental animal models indicates a possible role AGEs might have on the gut microbiota population. In this pilot randomized open label controlled trial, twenty PD patients habitually consuming a high AGE diet were recruited and randomized into either continuing the same diet (HAGE, n = 10) or a one-month dietary AGE restriction (LAGE, n = 10). Blood and stool samples were collected at baseline and after intervention. Variable regions V3-V4 of 16s rDNA were sequenced and taxa was identified on the phyla, genus, and species levels. Dietary AGE restriction resulted in a significant decrease in serum Nε-(carboxymethyl) lysine (CML) and methylglyoxal-derivatives (MG). At baseline, our total cohort exhibited a lower relative abundance of Bacteroides and Alistipes genus and a higher abundance of Prevotella genus when compared to the published data of healthy population. Dietary AGE restriction altered the bacterial gut microbiota with a significant reduction in Prevotella copri and Bifidobacterium animalis relative abundance and increased Alistipes indistinctus, Clostridium citroniae, Clostridium hathewayi, and Ruminococcus gauvreauii relative abundance. We show in this pilot study significant microbiota differences in peritoneal dialysis patients' population, as well as the effects of dietary AGEs on gut microbiota, which might play a role in the increased cardiovascular events in this population and warrants further studies.

Effect of newer antioxidants on the bond strength of composite on bleached enamel.

AIM: The study aims to evaluate the effect of the application of two antioxidants on the bond strength of composite resin to bleached enamel. MATERIALS AND METHODS: Eighty enamel surfaces were obtained from forty human extracted premolars. Specimens were randomly divided into four groups (n = 20). Group 1: No bleaching (control); Group 2a: Bleaching with 15% carbamide peroxide gel; Group 2b: Bleaching, followed by application of 10% sodium ascorbate gel; Group 2c: Bleaching, followed by application of 5% proanthocyanidin agent. Surfaces were etched followed by application of total etch bonding system, and composite resin cylinders were bonded. Specimens were tested for shear bond strength. STATISTICAL ANALYSIS USED: One-way analysis of variance was used for multiple group comparison and post hoc Tukey's test for individual group-wise comparison. RESULTS: Significantly higher shear bond strength values were observed in Group 2c and 2b as compared with Group 1 and 2a (P < 0.05). Among the antioxidants, Group 2c showed significantly higher shear bond strength values than Group 2b (P < 0.05). CONCLUSION: It can be concluded that the use of antioxidant before bonding procedures on bleached enamel completely neutralizes the deleterious effects of bleaching and increases the bond strength significantly.

THE EFFECT OF A SINGLE DOSE OF MAGNESIUM SULPHATE AS AN ADJUVANT TO EPIDURAL BUPIVACAINE FOR INFRAUMBILICAL SURGERIES: A PROSPECTIVE DOUBLE-BLIND, RANDOMIZED CONTROL TRIAL.

CONTEXT: Epidural anesthesia provides the advantage of segmental blockade and many adjuvants have been added to shorten the onset of action, improve the quality of analgesia and prolong the duration of analgesia. Magnesium sulphate(MgSO4) by virtue of its anti-iociceptive property has been administered by various routes. AIM: To assess the effect of MgSO4 on the duration of onset of action of injection bupivacaine for epidural anesthesia in infraumbilical surgeries. MATERIALS AND METHODS: A prospective, double-blind, randomized control study was conducted in 40 patients. Group M received 15 ml of bupivacaine 0.5% + 1 ml of 50 mg MgSO4 and Group C received 15 ml of bupivacaine 0.5% + 1 ml of normal saline via epidural route. Onset time of the sensory and motor blockade were the primary outcomes studied. Highest level of sensory block, time for two segment regression, hemodynamic parameters, side effects were the secondary parameters. RESULTS: There was a significant difference between the groups in the mean onset time of sensory blockade at T8, 12.85 ± 2.32 min in Group M and 16.75 ± 1.74 min in Group C. Median level of sensory blockade was comparable. Mean onset time of motor blockade was 13.85 ± 3.28 min in Group M and 23.25 ± 3.35 min in Group C which was clinically and statistically significant. Time for two segment regression of sensory blockade was 95.75 ± 11.84 min in Group M and 55.5 ± 8.57 min in Group C which was significant. Hemodynamic parameters and side effects were comparable. CONCLUSION: Magnesium sulphate as an adjuvant provides rapid onset of epidural anesthesia and prolongs the duration of analgesia with minimal side effects.

EFFECT OF DEXMEDETOMIDINE ON HEMODYNAMIC PARAMETERS DURING EXTUBATION. A PROSPECTIVE RANDOMIZED DOUBLE BLIND STUDY.

BACKGROUND: Extubation is known to produce significant hemodynamic disturbances. There is a need to avoid increase in heart rate and blood pressure in hypertensive and cardiac patients and in vascular, neuro and intraocular surgeries. AIMS: To study the ability of dexmedetomidine to attenuate the hemodynamic responses during extubation. MATERIALS AND METHODS: 80 patients of ASA Grade I-II aged 18-50 years received standard anesthesia. At the closure of skin incision, patients were randomly allocated to receive either dexmedetomidine 0.5 µg/kg (Group D) or saline placebo (Group C) intravenously over 10 minutes in a double-blind design. Heart rate (HR), systolic, diastolic and mean arterial pressures (SBP, DBP, MAP) were assessed before, during- and after extubation. Time to eye opening and extubation, sedation, complications such as coughing, laryngospasm, bronchospasm and desaturation were recorded. RESULTS: HR, SBP, DBP and MAP were comparable to basal values in group D at extubation and lower than baseline values post-extubation but significant increase was noted in group C (P <0.001). Time to extubation and eye opening were prolonged in Group D (P <0.001). Incidence of hypotension was more in group D (22%) but was transient. Incidence of coughing was lower in Group D than in group C (P <0.001). Patients in group D were more sedated for 30 minutes post extubation. CONCLUSION: Dexmedetomidine 0.5 µg/kg given before extubation attenuates hemodynamic reflexes during emergence from anesthesia without causing undue sedation, but prolongs time to extubation.

Comparative evaluation of microleakage of conventional and modifications of glass ionomer cement in primary teeth: An in vitro study.

AIMS: The aim of the present study was to evaluate the microleakage among conventional, resin modified glass ionomer cements (GIC), and compomer cements in primary teeth. MATERIALS AND METHODS: Forty-five over retained non carious primary molars beyond exfoliation time were collected and randomly divided into three groups (n = 15). Group A: GC Fuji II; Group B: Vitremer; Group C: Compoglass F. A standard Class V cavity was prepared on the buccal surface of each tooth with no mechanical retention and restored accordingly. Then all the samples were subjected to thermocycling for 250 cycles at different temperatures and covered with nail varnish. Later, samples were immersed in 0.5% methylene blue dye for 24 h. Teeth were sectioned buccolingually through the center of the restoration and studied under a stereomicroscope for dye penetration. Data obtained were analyzed using Kruskal-Wallis ANOVA and Mann-Whitney U-test. RESULTS: Samples restored with vitremer showed comparatively higher microleakage than the samples in other groups. However, overall there were no significant difference between the microleakage scores of the samples in all three groups (P > 0.05). CONCLUSION: It can be concluded that none of the three GICs was free from microleakage. Hence, further research is required to compare microleakage of the newer material.

Chilaiditi's syndrome.

Chilaidit's syndrome is a rare condition characterised by the interposition of the colon between the liver and the right hemidiaphragm. We present a case of 20-year-old male who reported with breathlessness and epigastric pain, and he was diagnosed radiologically to have Chilaiditi's syndrome.

Synthesis, in vitro antioxidant, anthelmintic and molecular docking studies of novel dichloro substituted benzoxazole-triazolo-thione derivatives.

A novel 6,8-dichloro [1,2,4]triazolo [3,4-b] [1,3]benzoxazole-3(2H)-thione 4 and its derivatives 5a and 5b are synthesized from 5,7-dichloro-2-hydrazinyl-1,3-benzoxazole 3, obtained by reaction of hydrazine hydrate with ethyl [(5,7-dichloro-1,3-benzoxazol-2-yl)sulfanyl]acetate 2. The newly synthesized compounds are characterized by analytical (1)H NMR, (13)C NMR, LC-MS mass spectrometry and elemental analysis. All synthesized compounds are screened for in vitro antioxidant and anthelmintic activities. In correlation to anthelmintic activity, compounds are subjected to molecular docking studies for the binding to ß-Tubulin, target protein elite to the parasites. Compounds 3, 4 and 5a exhibited potential radical scavenging capacity with good anthelmintic activity. In molecular docking study also, compounds showed minimum binding energy and have good affinity toward the active pocket thus, they may be considered as good inhibitor of ß-Tubulin.