Plasma Cannabinoid Pharmacokinetics After Controlled Smoking and Ad libitum Cannabis Smoking in Chronic Frequent Users.

More Americans are dependent on cannabis than any other illicit drug. The main analytes for cannabis testing include the primary psychoactive constituent, Δ(9)-tetrahydrocannabinol (THC), equipotent 11-hydroxy-THC (11-OH-THC) and inactive 11-nor-9-carboxy-THC (THCCOOH). Eleven adult chronic frequent cannabis smokers resided on a closed research unit with unlimited access to 5.9% THC cannabis cigarettes from 12:00 to 23:00 during two ad libitum smoking phases, followed by a 5-day abstinence period in seven participants. A single cigarette was smoked under controlled topography on the last day of the smoking and abstinence phases. Plasma cannabinoids were quantified by two-dimensional gas chromatography-mass spectrometry. Median plasma maximum concentrations (Cmax) were 28.3 (THC), 3.9 (11-OH-THC) and 47.0 µg/L (THCCOOH) 0.5 h after controlled single cannabis smoking. Median Cmax 0.2-0.5 h after ad libitum smoking was higher for all analytes: 83.5 (THC), 14.2 (11-OH-THC) and 155 µg/L (THCCOOH). All 11 participants' plasma samples were THC and THCCOOH-positive, 58.3% had THC ≥5 µg/L and 79.2% were 11-OH-THC-positive 8.1-14 h after last cannabis smoking. Cannabinoid detection rates in seven participants 106-112 h (4-5 days) after last smoking were 92.9 (THC), 35.7 (11-OH-THC) and 100% (THCCOOH), with limits of quantification of 0.5 µg/L for THC and THCCOOH, and 1.0 µg/L for 11-OH-THC. These data greatly expand prior research findings on cannabinoid excretion profiles in chronic frequent cannabis smokers during ad libitum smoking. Smoking multiple cannabis cigarettes led to higher Cmax and AUC compared with smoking a single cigarette. The chronic frequent cannabis smokers exhibited extended detection windows for plasma cannabinoids, reflecting a large cannabinoid body burden.

Hydroxycarbamide modulates components involved in the regulation of adenosine levels in blood cells from sickle-cell anemia patients.

Recent studies have demonstrated the role of adenosine (ADO) in sickle-cell anemia (SCA). ADO is produced by CD39 and CD73 and converted to inosine by adenosine deaminase (ADA). We evaluated the effects of hydroxycarbamide (HU) treatment on the modulation of adenosine levels in SCA patients. The expressions of CD39, CD73, and CD26 were evaluated by flow cytometry on blood cells in 15 HU-treated and 17 untreated patients and 10 healthy individuals. RNA was extracted from monocytes, and ADA gene expression was quantified by real-time PCR. ADA activity was also evaluated. We found that ADA transcripts were two times higher in monocytes of HU-treated patients, compared with untreated (P = 0.039). Monocytes of HU-treated patients expressed CD26, while monocytes of controls and untreated patients did not (P = 0.023). In treated patients, a lower percentage of T lymphocytes expressed CD39 compared with untreated (P = 0.003), and the percentage of T regulatory (Treg) cells was reduced in the treated group compared with untreated (P = 0.017) and controls (P = 0.0009). Besides, HU-treated patients displayed increased ADA activity, compared with untreated. Our results indicate a novel mechanism of action of HU mediated by the reduction of adenosine levels and its effects on pathophysiological processes in SCA.

Rimonabant effects on anxiety induced by simulated public speaking in healthy humans: a preliminary report.

OBJECTIVE: We investigated the hypothesis that rimonabant, a cannabinoid antagonist/inverse agonist, would increase anxiety in healthy subjects during a simulation of the public speaking test. METHODS: Participants were randomly allocated to receive oral placebo or 90 mg rimonabant in a double-blind design. Subjective effects were measured by Visual Analogue Mood Scale. Physiological parameters, namely arterial blood pressure and heart rate, also were monitored. RESULTS: Twelve participants received oral placebo and 12 received 90 mg rimonabant. Rimonabant increased self-reported anxiety levels during the anticipatory speech and performance phase compared with placebo. Interestingly, rimonabant did not modulate anxiety prestress and was not associated with sedation, cognitive impairment, discomfort, or blood pressure changes. CONCLUSIONS: Cannabinoid-1 antagonism magnifies the responses to an anxiogenic stimulus without interfering with the prestress phase. These data suggest that the endocannabinoid system may work on-demand to counteract the consequences of anxiogenic stimuli in healthy humans.

Impact of enzymatic and alkaline hydrolysis on CBD concentration in urine.

A sensitive and specific analytical method for cannabidiol (CBD) in urine was needed to define urinary CBD pharmacokinetics after controlled CBD administration, and to confirm compliance with CBD medications including Sativex-a cannabis plant extract containing 1:1 ∆(9)-tetrahydrocannabinol (THC) and CBD. Non-psychoactive CBD has a wide range of therapeutic applications and may also influence psychotropic smoked cannabis effects. Few methods exist for the quantification of CBD excretion in urine, and no data are available for phase II metabolism of CBD to CBD-glucuronide or CBD-sulfate. We optimized the hydrolysis of CBD-glucuronide and/or -sulfate, and developed and validated a GC-MS method for urinary CBD quantification. Solid-phase extraction isolated and concentrated analytes prior to GC-MS. Method validation included overnight hydrolysis (16 h) at 37 °C with 2,500 units ß-glucuronidase from Red Abalone. Calibration curves were fit by linear least squares regression with 1/x (2) weighting with linear ranges (r(2) > 0.990) of 2.5-100 ng/mL for non-hydrolyzed CBD and 2.5-500 ng/mL for enzyme-hydrolyzed CBD. Bias was 88.7-105.3 %, imprecision 1.4-6.4 % CV and extraction efficiency 82.5-92.7 % (no hydrolysis) and 34.3-47.0 % (enzyme hydrolysis). Enzyme-hydrolyzed urine specimens exhibited more than a 250-fold CBD concentration increase compared to alkaline and non-hydrolyzed specimens. This method can be applied for urinary CBD quantification and further pharmacokinetics characterization following controlled CBD administration.

Impact of prolonged cannabinoid excretion in chronic daily cannabis smokers' blood on per se drugged driving laws.

BACKGROUND: Cannabis is the illicit drug most frequently reported with impaired driving and motor vehicle accidents. Some "per se" laws make it illegal to drive with any amount of drug in the body, while others establish blood, saliva, or urine concentrations above which it is illegal to drive. The persistence of Δ(9)-tetrahydrocannabinol (THC) in chronic daily cannabis smokers' blood is unknown. METHODS: Thirty male chronic daily cannabis smokers resided on a secure research unit for up to 33 days, with daily blood collection. Samples were processed in an ice bath during sample preparation to minimize cannabinoid adsorption onto precipitant material. We quantified THC by 2-dimensional GC-MS. RESULTS: Of the 30 participants, 27 were THC-positive on admission, with a median (range) concentration of 1.4 µg/L (0.3-6.3). THC decreased gradually; only 1 of 11 participants was negative at 26 days, 2 of 5 remained THC-positive (0.3 µg/L) for 30 days, and 5.0% of participants had THC ≥ 1.0 µg/L for 12 days. Median 11-hydroxy-THC concentrations were 1.1 µg/L on admission, with no results ≥ 1.0 µg/L 24 h later. 11-Nor-9-carboxy-THC (THCCOOH) detection rates were 96.7% on admission, decreasing slowly to 95.7% and 85.7% on days 8 and 22, respectively; 4 of 5 participants remained THCCOOH positive (0.6-2.7 µg/L) after 30 days, and 1 remained positive on discharge at 33 days. CONCLUSIONS: Cannabinoids can be detected in blood of chronic daily cannabis smokers during a month of sustained abstinence. This is consistent with the time course of persisting neurocognitive impairment reported in recent studies.

Anxiolytic-like effect of oxytocin in the simulated public speaking test.

Oxytocin (OT) is known to be involved in anxiety, as well as cardiovascular and hormonal regulation. The objective of this study was to assess the acute effect of intranasally administered OT on subjective states, as well as cardiovascular and endocrine parameters, in healthy volunteers (n = 14) performing a simulated public speaking test. OT or placebo was administered intranasally 50 min before the test. Assessments were made across time during the experimental session: (1) baseline (-30 min); (2) pre-test (-15 min); (3) anticipation of the speech (50 min); (4) during the speech (1:03 h), post-test time 1 (1:26 h), and post-test time 2 (1:46 h). Subjective states were evaluated by self-assessment scales. Cortisol serum and plasma adrenocorticotropic hormone (ACTH) were measured. Additionally, heart rate, blood pressure, skin conductance, and the number of spontaneous fluctuations in skin conductance were measured. Compared with placebo, OT reduced the Visual Analogue Mood Scale (VAMS) anxiety index during the pre-test phase only, while increasing sedation at the pre-test, anticipation, and speech phases. OT also lowered the skin conductance level at the pre-test, anticipation, speech, and post-test 2 phases. Other parameters evaluated were not significantly affected by OT. The present results show that OT reduces anticipatory anxiety, but does not affect public speaking fear, suggesting that this hormone has anxiolytic properties.

Mesenchymal stromal cells up-regulate CD39 and increase adenosine production to suppress activated T-lymphocytes.

Mesenchymal stromal cells (MSCs) suppress T cell responses through mechanisms not completely understood. Adenosine is a strong immunosuppressant that acts mainly through its receptor A(2a) (ADORA2A). Extracellular adenosine levels are a net result of its production (mediated by CD39 and CD73), and of its conversion into inosine by Adenosine Deaminase (ADA). Here we investigated the involvement of ADO in the immunomodulation promoted by MSCs. Human T lymphocytes were activated and cultured with or without MSCs. Compared to lymphocytes cultured without MSCs, co-cultured lymphocytes were suppressed and expressed higher levels of ADORA2A and lower levels of ADA. In co-cultures, the percentage of MSCs expressing CD39, and of T lymphocytes expressing CD73, increased significantly and adenosine levels were higher. Incubation of MSCs with media conditioned by activated T lymphocytes induced the production of adenosine to levels similar to those observed in co-cultures, indicating that adenosine production was mainly derived from MSCs. Finally, blocking ADORA2A signaling raised lymphocyte proliferation significantly. Our results suggest that some of the immunomodulatory properties of MSCs may, in part, be mediated through the modulation of components related to adenosine signaling. These findings may open new avenues for the development of new treatments for GVHD and other inflammatory diseases.

[Interchangeability among therapeutics equivalents of lamotrigine in the treatment of refractory epilepsy patients: risks and benefits]. / Intercambiabilidad entre equivalentes terapéuticos de lamotrigina en pacientes con epilepsia refractaria: riesgos y beneficios.

INTRODUCTION: Epilepsy is a condition characterized by signs and symptoms of neurological disorder. Lamotrigine has been widely used, mainly due to their greater tolerability and lower rate of drug interactions with other antiepileptic drugs however the newest antiepileptic drugs have high cost to patient. In Brazil there are three different sort of pharmaceutical equivalents (reference, generic and similar), and the Brazilian health care authorities offers to users the possibility to receive them free of charge. Moreover these pharmaceutical equivalents can change during the treatment of epilepsy because this authorities buy the cheapest by public tender two or three times a year. AIM: To evaluate the clinical and laboratory findings related to the most frequently used therapeutic equivalents of lamotrigine (reference drugs and similar products). PATIENTS AND METHODS: Two similar formulations (A and B) and one reference (C) were tested in nine epileptic refractory patients. The study was divided into three periods of 42 days, one for each formulation, and medical data about the frequency of seizures, the occurrence of side effects and measurement of plasma concentrations of lamotrigine were collected. RESULTS: The average number of seizures/week and plasma concentration of lamotrigine for formulations A, B and C were not statistically significant differences. Three patients during the use of the formulation C presented mild and transitory side effects. CONCLUSION: Similar or reference drugs showed satisfactory results, however the interchangeability among the formulations raise the difficulty for the management of seizures in refractory epilepsy.

Quantification of nicotine in commercial brand cigarettes: How much is inhaled by the smoker?

The main objective of this experiment is to determine the amount of nicotine in commercial brand cigarettes by means of a nonaqueous acid-base titration. A simple glass device simulating a smoker is proposed, which allows the determination of the volatilized, filter retained, and inhaled portions. Students will readily see that the amount of nicotine/cigarette stated on the label (∼0.5-1.0 mg) refers indeed to the inhaled portion only, rather than to the total amount/cigarette (usually more than 10 mg). Even so, values for inhaled nicotine may be significantly higher than those reported for several brands. Students will also be able to make a critical evaluation of the true content of nicotine in the inhaled portion and confront it with the reported value for a given brand. In addition, the theoretical approach, supported by HPLC data, provides an excellent experience on nonaqueous acid-base volumetric analysis.

Socioeconomic differences in preferences and willingness-to-pay for insulin delivery systems in type 1 and type 2 diabetes.

BACKGROUND: An evaluation of patients' preferences is necessary to understand the demand for different insulin delivery systems. The aim of this study was to investigate the association between socioeconomic status (SES) and patients' preferences and willingness to pay (WTP) for various attributes of insulin administration for diabetes management. METHODS: We conducted a discrete choice experiment (DCE) to determine patients; preferences and their WTP for hypothetical insulin treatments. Both self-reported annual household income and education completed were used to explore differences in treatment preferences and WTP for different attributes of treatment across different levels of SES. RESULTS: The DCE questionnaire was successfully completed by 274 patients. Overall, glucose control was the most valued attribute by all socioeconomic groups, while route of insulin delivery was not as important. Patients with higher incomes were willing to pay significantly more for better glucose control and to avoid adverse events compared to lower income groups. In addition, they were willing to pay more for an oral short-acting insulin ($Can 71.65 [95% confidence interval, $40.68, $102.62]) compared to the low income group ($Can 9.85 [95% confidence interval, 14.86, 34.56; P < 0.01]). Conversely, there were no differences in preferences when the sample was stratified by level of education. CONCLUSIONS: This study revealed that preferences and WTP for insulin therapy are influenced by income but not by level of education. Specifically, the higher the income, the greater desire for an oral insulin delivery system, whereas an inhaled route becomes less important for patients.

A valuation of patients' willingness-to-pay for insulin delivery in diabetes.

OBJECTIVES: The aim of this study was to determine the insulin-delivery system and the attributes of insulin therapy that best meet patients' preferences, and to estimate patients' willingness-to-pay (WTP) for them. METHODS: This was a cross-sectional discrete choice experiment (DCE) study involving 378 Canadian patients with type 1 or type 2 diabetes. Patients were asked to choose between two hypothetical insulin treatment options made up of different combinations of the attribute levels. Regression coefficients derived using conditional logit models were used to calculate patients' WTP. Stratification of the sample was performed to evaluate WTP by predefined subgroups. RESULTS: A total of 274 patients successfully completed the survey. Overall, patients were willing to pay the most for better blood glucose control followed by weight gain. Surprisingly, route of insulin administration was the least important attribute overall. Segmented models indicated that insulin naïve diabetics were willing to pay significantly more for both oral and inhaled short-acting insulin compared with insulin users. Surprisingly, type 1 diabetics were willing to pay $C11.53 for subcutaneous short-acting insulin, while type 2 diabetics were willing to pay $C47.23 to avoid subcutaneous short-acting insulin (p < .05). These findings support the hypothesis of a psychological barrier to initiating insulin therapy, but once that this barrier has been overcome, they accommodate and accept injectable therapy as a treatment option. CONCLUSIONS: By understanding and addressing patients' preferences for insulin therapy, diabetes educators can use this information to find an optimal treatment approach for each individual patient, which may ultimately lead to improved control, through improved compliance, and better diabetes outcomes.

Chronic ethanol intake inhibits in vitro osteogenesis induced by osteoblasts differentiated from stem cells.

The study investigated whether chronic ethanol (ETH) intake and subsequent ETH exposure of cell cultures affects osteoblast differentiation by evaluating key parameters of in vitro osteogenesis. Rats were treated with 5-20% (0.85-3.43 mm) ETH, increasing by 5% per week for a period of 4 weeks (habituation), after which the 20% level was maintained for 15 days (chronic intake). Bone-marrow stem cells from control (CONT) or ETH-treated rats were cultured in osteogenic medium which was either supplemented (ETH) or not supplemented (CONT) with 1.3 mm ethanol. Thus, four groups relating to rat treatment/culture supplementation were evaluated: (1) CONT/CONT, (2) ETH/CONT, (3) CONT/ETH and (4) ETH/ETH. Cell morphology, proliferation and viability, total protein content, alkaline phosphatase (ALP) activity and bone-like nodule formation were evaluated. Chronic ethanol intake significantly reduced both food and liquid consumption and body weight gain. No difference was seen in cell morphology among treatments. Cell number was affected at 7 and 10 days as follows: CONT/CONT = CONT/ETH < ETH/CONT = ETH/ETH. Doubling time between 3 and 10 days was greater in groups of CONT animals: ETH/ETH = ETH/CONT < CONT/ETH = CONT/CONT. Cell viability and ALP activity were not affected by either animal treatment or culture exposure to ethanol. At day 21, the total protein content was affected as follows: ETH/ETH = CONT/ETH < ETH/CONT = CONT/CONT. Bone-like nodule formation was affected as follows: ETH/ETH < CONT/ETH < ETH/CONT < CONT/CONT. These results show that chronic ethanol intake, followed by the exposure of osteoblasts to ethanol, inhibited the differentiation of osteoblasts, as indicated by an increased proliferation rate and reduced bone-like nodule formation.

Leaching and half-life of the herbicide tebuthiuron on a recharge area of Guarany aquifer in sugarcane fields in Brazil.

This study was undertaken to evaluate the degradation and mobility of the herbicide tebuthiuron (N-[5-(1,1-dimethylethyl)-1,3,4-thiadiazol-2-yl]-N,N'-dimethylurea) in soil under field conditions, and its potential for leaching and groundwater contamination. A watershed, Espraiado, located over a recharge area in Brazil, was chosen for soil and water studies. At Espraiado, water samples were collected from seven wells at intervals of three months from March 2004 to June 2006 and analyzed for tebuthiuron. Other samples were taken from city wells located outside of the recharge area. To assess the potential movement to the aquifer, tebuthiuron was also applied to trial plots at the recommended label rate of 1.0 kg/ha a.i. in May of 2004, with and without sugarcane coverage, on sandy soil. Soil samples were collected during the years of 2004 and 2005, at depths intervals of 20 cm from soil surface down to 120 cm and analyzed for tebuthiuron at zero, 3, 30, 60, 90, 120, 150, 180, 240, and 300 days after application. There was no clear effect of sugarcane coverage on the tebuthiuron degradation in soils, but it moved faster into the soil where there was no cover. After 180 days there were no measurable residues in the soil, and tebuthiuron was not found below 40 cm depth in any time. Tebuthiuron had a half-life of 20 days under those conditions. No tebuthiuron residue was found in ground water samples at any sampling time.

Tolerability and effectiveness of fluoxetine, metformin and sibutramine in reducing anthropometric and metabolic parameters in obese patients / Tolerabilidade e eficácia da fluoxetina, metformina e sibutramina na redução de parâmetros antropométricos e metabólicos em pacientes obesos

The aim of this study is to assess the effects of sibutramine (S) 15 mg/day, fluoxetine (F) 60 mg/day, and metformin (M) 1,700 mg/day, as an adjunct therapy to a 1,500 kcal/day diet, in reducing anthropometric and metabolic parameters. S (n= 8), F (n= 9), and M (n= 8) were compared to placebo (n= 10) in 35 obese patients in a 90-day trial. Side effects were also studied during the treatment. The data demonstrated that F therapy resulted in a greater average reduction in BMI (11.0 percent), weight (10.0 percent), abdominal circumference (11.0 percent) and percentfatty-tissue (12.8). An elevation in HDL-cholesterol (25.8 percent) and a reduction in average triglyceride levels (28.3 percent) were also shown. S presented a 7.91 percent reduction in the abdominal circumference and a 9.65 reduction in percentfatty-tissue was also found. M group presented reductions in BMI (4.03 percent), waist circumference (6.92 percent), HOMA (23.5 percent) and blood pressure (6.08 percent in systolic and 2.08 percent in diastolic). In general, the three drugs can be considered well tolerated. We concluded that F and S demonstrated a greater mean reduction in anthropometric and metabolic parameters when compared to M, however all of them are useful for that purpose, when the subjectsÆ characteristics are considered.

O objetivo deste estudo foi avaliar o efeito da sibutramina (S) 15 mg/dia, fluoxetina (F) 60 mg/dia, e metformina (M) 1.700 mg/dia, associadas a uma dieta de 1.500 kcal/dia, na redução de parâmetros antropométricos e metabólicos. S (n= 8), F (n= 9) e M (n= 8) foram comparadas ao placebo (n= 10) em 35 pacientes obesos durante 90 dias de tratamento. As reações adversas também foram avaliadas durante o tratamento. O grupo F demonstrou uma redução no IMC (11,0 por cento), peso (10,0 por cento), circunferência abdominal (11,0 por cento) e por cento de tecido adiposo (12,8). Também foram observados um aumento nos níveis de HDL-colesterol (25,8 por cento) e uma redução nos níveis de triglicérides (28,3 por cento), no grupo F. O grupo S apresentou uma redução de 7,91 por cento na circunferência abdominal e de 9,65 na por cento de tecido adiposo. Já o grupo M apresentou reduções no IMC (4,03 por cento), circunferência abdominal (6,92 por cento), HOMA (23,5 por cento) e pressão arterial (6,08 por cento na sistólica, 2,08 por cento na diastólica). Os três fármacos analisados foram bem tolerados durante o tratamento. Concluímos que a F e a S demonstraram maior eficácia na redução dos parâmetros antropométricos e metabólicos dos pacientes obesos quando comparadas à M, entretanto todas podem ser prescritas para essa finalidade, desde que sejam consideradas as características individuais dos pacientes.

Tolerability and effectiveness of fluoxetine, metformin and sibutramine in reducing anthropometric and metabolic parameters in obese patients.

The aim of this study is to assess the effects of sibutramine (S) 15 mg/day, fluoxetine (F) 60 mg/day, and metformin (M) 1,700 mg/day, as an adjunct therapy to a 1,500 kcal/day diet, in reducing anthropometric and metabolic parameters. S (n= 8), F (n= 9), and M (n= 8) were compared to placebo (n= 10) in 35 obese patients in a 90-day trial. Side effects were also studied during the treatment. The data demonstrated that F therapy resulted in a greater average reduction in BMI (11.0%), weight (10.0%), abdominal circumference (11.0%) and %fatty-tissue (12.8). An elevation in HDL-cholesterol (25.8%) and a reduction in average triglyceride levels (28.3%) were also shown. S presented a 7.91% reduction in the abdominal circumference and a 9.65 reduction in %fatty-tissue was also found. M group presented reductions in BMI (4.03%), waist circumference (6.92%), HOMA (23.5%) and blood pressure (6.08% in systolic and 2.08% in diastolic). In general, the three drugs can be considered well tolerated. We concluded that F and S demonstrated a greater mean reduction in anthropometric and metabolic parameters when compared to M, however all of them are useful for that purpose, when the subjects characteristics are considered.

Determination of paroxetine in geriatric depression by high-performance liquid chromatography.

An HPLC method was developed for the determination of paroxetine in depressed geriatric patients. This method includes only one-step extraction of paroxetine from 1 ml of plasma with hexane:isoamyl alcohol (99:1, v/v). Separation was obtained using an RP select B column (250 mm x 4 mm, 5 microm) under isocratic conditions with UV detection (lambda=205 nm). The mobile phase was 0.05 mol/l sodium phosphate buffer, pH 5.0, and acetonitrile (50:50, v/v). The intra- and inter-assay accuracy and precision, determined as relative error and relative standard deviation, respectively, were less than 10%. The lower limit of quantification, with relative error and relative standard deviation of less than 10%, was 5 ng/ml. Linearity was assessed in the 5-1250 ng/ml range and recovery was 80%. Neither endogenous compounds nor other drugs were found to interfere. The assay showed high specificity, even when patients were found to be on multiple medications. In conclusion, the present method, when compared to others reported in the literature, has advantages such as simplicity, selectivity and sensitivity as a routine procedure and can be applied to the therapeutic monitoring of depressed geriatric patients.