Correction: Sensitivity of self-reported opioid use in case-control studies: Healthy individuals versus hospitalized patients.

[This corrects the article DOI: 10.1371/journal.pone.0183017.].

Effervescent tablet-assisted demulsified dispersive liquid-liquid microextraction based on solidification of floating organic droplet for determination of methadone in water and biological samples prior to GC-flame ionization and GC-MS.

A novel effervescent tablet-assisted demulsified dispersive liquid-liquid microextraction based on the solidification of floating organic droplet was developed to determine methadone prior to gas chromatography with flame ionization detection and gas chromatography with mass spectrometry. In this method, a tablet composed of citric acid, sodium carbonate, and 1-undecanol was utilized. The resulting effervescent tablet generated carbon dioxide in situ to disperse 1-undecanol in the sample. Thus, the dispersive and extraction processes were performed in one synchronous step. An aliquot of acetonitrile as the demulsifier solvent was used for the separation of two phases instead of centrifugation. Under optimal conditions, the developed method was linear up to 50 000 µg/L with correlation coefficients higher than 0.99. Moreover, limits of detection and limits of the quantification were in the range of 3-10  and 7-30 µg/L in water and biological samples, respectively. Intra- and interday precisions (n = 6) of the spiked methadone at a concentration level of 50 µg/L were over ranges of 5.1-6.8% and 5.7-7.1%, respectively. The preconcentration factors and recovery values were obtained in the range of 140-145 and 98.1 to 101.6% in real samples, respectively.

Evaluation of dynorphin and kappa-opioid receptor level in the human blood lymphocytes and plasma: Possible role as a biomarker in severe opioid use disorder.

BACKGROUND: The dynorphin (DYN)/kappa opioid receptor (KOR) system plays an important role in the development of addiction, and dysregulation of this system could lead to abnormal activity in the reward pathway. It has been reported that the expression state of the neurotransmitters and their receptors in the brain is reflected in peripheral blood lymphocytes (PBLs). METHODS: We have evaluated the PBLs and plasma samples of four groups: 1) subjects with severe opioid use disorder (SOD), 2) methadone-maintenance treated (MMT) individuals, 3) long-term abstinent subjects having former SOD, and 4) healthy control subjects (n = 20 in each group). The mRNA expression level of preprodynorphin (pPDYN) and KOR in PBLs has been evaluated by real-time PCR. Peptide expression of PDYN in PBLs has been studied by western blot, and DYN concentration in plasma has been measured by ELISA. RESULTS: The relative expression level of the pPDYN mRNA and PDYN peptide in PBLs were significantly up-regulated in SOD, MMT, and abstinent groups compared to control subjects. No significant difference was found in the plasma DYN concentration between study groups. The expression level of the KOR mRNA in PBLs was significantly decreased in all three study groups compared to the control subjects. CONCLUSION: the expression changes in the DYN/KOR system after chronic exposure to opioids, including methadone, seems to be stable and does not return to normal levels even after 12 months abstinence. These long-time and permanent changes in PBLs may serve as a biomarker and footprint of SOD development in the periphery.

Tramadol induces changes in Δ-FosB, µ-opioid receptor, and p-CREB level in the nucleus accumbens and prefrontal cortex of male Wistar rat.

BACKGROUND: Besides the analgesic effect of tramadol, prolonged exposure to tramadol can induce adaptive changes thereby leading to dependence and tolerance. Tramadol induces its effect via µ-opioid receptor (MOR). However, tramadol has other targets such as serotonin and epinephrine transporters. OBJECTIVE: CREB and ΔFosB are transcriptional factors, which are involved in the behavioral abnormalities underlying drug abuse. In this study, the effects of acute and chronic tramadol treatments on MOR, ΔFosB, and CREB levels were studied. METHODS: For this purpose, 36 male Wistar rats were used. The animals were divided into two main groups. A total of 18 animals received tramadol (0, 5, and 10 mg/kg) acutely and 18 animals received the same doses for the following 14 days. One hour after the last injection, the NAC and PFC were dissected and kept at -80°C in liquid nitrogen. Using western blotting technique, the levels of MOR, ΔFosB, and p-CREB were evaluated. RESULTS: In the NAC, acute tramadol exposure increases the levels of MOR and p-CREB. Moreover, chronic tramadol administration in this region results in elevated levels of MOR, ΔFosB and p-CREB compared with saline-treated rats. The levels of MOR and p-CREB in the PFC increased in both acute and chronic tramadol exposure. Also, ΔFosB levels increased only following chronic tramadol administration. The results revealed that adaptive changes occurred during drug exposure. CONCLUSION: We concluded that both CREB and ΔFosB played a role in tramadol dependence. Additionally, increased MOR levels during tramadol treatments might be due to receptor desensitization.

Morphine exposure before conception affects anxiety-like behavior and CRF level (in the CSF and plasma) in the adult male offspring.

It has been proven that exposure to some drugs even before gestation had transgenerational effects. To investigate the changes which induced by parental morphine exposure before gestation; mainly the anxiety-like behavior, Corticotropin Releasing Factor (CRF) level in the CSF and plasma, CRF Receptor 1 (CRFR1), and the level of protein kinase C (PKC-α) were evaluated in the male offspring. Male and female Wistar rats were exposed to morphine for 21 following days. Ten days after last drug exposure, animals were prepared for mating in 4 distinct groups as follow: drug-naïve female and male (used as control), drug-naïve female and morphine-abstinent male, drug-naïve male and morphine-abstinent female, and morphine abstinent male and female. Offspring were subjected to assess anxiety-like behavior (using elevated plus maze test). CSF and plasma were gathered, and the CRF level was evaluated by ELISA. Using real-time PCR, the CRFR1 level in the brain was evaluated. Results showed that anxiety-like behavior increased in the offspring of morphine-abstinent parent(s) compared with the control group. CRF level in the plasma and CSF also increased in the litter of morphine-abstinent parent(s). CRFR1 mRNA level was upregulated in the brain of offspring with one and/or two morphine-abstinent parent(s). Furthermore, the level of PKC-α was decreased in the brain of offspring which had one and/or two morphine-abstinent parent(s). Taken together, our findings indicated that morphine exposure even before gestation induced transgenerational effects via dysregulation of HPA axis which results in anxiety in the adult male offspring.

Correction: Sensitivity of self-reported opioid use in case-control studies: Healthy individuals versus hospitalized patients.

[This corrects the article DOI: 10.1371/journal.pone.0183017.].

Sensitivity of self-reported opioid use in case-control studies: Healthy individuals versus hospitalized patients.

BACKGROUND: Several case-control studies have shown associations between the risk of different cancers and self-reported opium use. Inquiring into relatively sensitive issues, such as the history of drug use, is usually prone to information bias. However, in order to justify the findings of these types of studies, we have to quantify the level of such a negative bias. In current study, we aimed to evaluate sensitivity of self-reported opioid use and suggest suitable types of control groups for case-control studies on opioid use and the risk of cancer. METHODS: In order to compare the validity of the self-reported opioid use, we cross-validated the response of two groups of subjects 1) 178 hospitalized patients and 2) 186 healthy individuals with the results of their tests using urine rapid drug screen (URDS) and thin layer chromatography (TLC). The questioners were asked by trained interviewers to maximize the validity of responses; healthy individuals were selected from the companions of patients in hospitals. RESULTS: Self-reported regular opioid use was 36.5% in hospitalized patients 19.3% in healthy individuals (p-value> 0.001).The reported frequencies of opioid use in the past 72 hours were 21.4% and 11.8% in hospitalized patients and healthy individuals respectively. Comparing their responses with the results of urine tests showed a sensitivity of 77% and 69% among hospitalized patients and healthy individuals for self-reports (p-value = 0.4). Having corrected based on the mentioned sensitivities; the frequency of opioid regular use was 47% and 28% in hospitalized patients and healthy individuals, respectively. Regular opioid use among hospitalized patients was significantly higher than in healthy individuals (p-value> 0.001). CONCLUSION: Our findings showed that the level of opioid use under-reporting in hospitalized patients and healthy individuals was considerable but comparable. In addition, the frequency of regular opioid use among hospitalized patients was significantly higher than that in the general population. Altogether, it seems that, without corrections for these differences and biases, the results of many studies including case-control studies on opioid use might distort findings substantially.