Administration Of Δ9-THC Research Articles

Chronic administration of Δ9-tetrahydrocannabinol induces intestinal anti-inflammatory microRNA expression during acute simian immunodeficiency virus infection of rhesus macaques.

Recreational and medical use of cannabis among human immunodeficiency virus (HIV)-infected individuals has increased in recent years. In simian immunodeficiency virus (SIV)-infected macaques, chronic administration of Δ9-tetrahydrocannabinol (Δ9-THC) inhibited viral replication and intestinal inflammation and slowed disease progression. Persistent gastrointestinal disease/inflammation has been proposed to facilitate microbial translocation and systemic immune activation and promote disease progression. Cannabinoids including Δ9-THC attenuated intestinal inflammation in mouse colitis models and SIV-infected rhesus macaques. To determine if the anti-inflammatory effects of Δ9-THC involved differential microRNA (miRNA) modulation, we profiled miRNA expression at 14, 30, and 60 days postinfection (days p.i.) in the intestine of uninfected macaques receiving Δ9-THC (n=3) and SIV-infected macaques administered either vehicle (VEH/SIV; n=4) or THC (THC/SIV; n=4). Chronic Δ9-THC administration to uninfected macaques significantly and positively modulated intestinal miRNA expression by increasing the total number of differentially expressed miRNAs from 14 to 60 days p.i. At 60 days p.i., ∼28% of miRNAs showed decreased expression in the VEH/SIV group compared to none showing decrease in the THC/SIV group. Furthermore, compared to the VEH/SIV group, THC selectively upregulated the expression of miR-10a, miR-24, miR-99b, miR-145, miR-149, and miR-187, previously been shown to target proinflammatory molecules. NOX4, a potent reactive oxygen species generator, was confirmed as a direct miR-99b target. A significant increase in NOX4+ crypt epithelial cells was detected in VEH/SIV macaques compared to the THC/SIV group. We speculate that miR-99b-mediated NOX4 downregulation may protect the intestinal epithelium from oxidative stress-induced damage. These results support a role for differential miRNA induction in THC-mediated suppression of intestinal inflammation. Whether similar miRNA modulation occurs in other tissues requires further investigation. Gastrointestinal (GI) tract disease/inflammation is a hallmark of HIV/SIV infection. Previously, we showed that chronic treatment of SIV-infected macaques with Δ9-tetrahydrocannabinol (Δ9-THC) increased survival and decreased viral replication and infection-induced gastrointestinal inflammation. Here, we show that chronic THC administration to SIV-infected macaques induced an anti-inflammatory microRNA expression profile in the intestine at 60 days p.i. These included several miRNAs bioinformatically predicted to directly target CXCL12, a chemokine known to regulate lymphocyte and macrophage trafficking into the intestine. Specifically, miR-99b was significantly upregulated in THC-treated SIV-infected macaques and confirmed to directly target NADPH oxidase 4 (NOX4), a reactive oxygen species generator known to damage intestinal epithelial cells. Elevated miR-99b expression was associated with a significantly decreased number of NOX4+ epithelial cells in the intestines of THC-treated SIV-infected macaques. Overall, our results show that selective upregulation of anti-inflammatory miRNA expression contributes to THC-mediated suppression of gastrointestinal inflammation and maintenance of intestinal homeostasis.

A vapourized Δ9-tetrahydrocannabinol (Δ9-THC) delivery system part I: Development and validation of a pulmonary cannabinoid route of exposure for experimental pharmacology studies in rodents

IntroductionMost studies evaluating the effects of Δ9-tetrahydrocannabinol (Δ9-THC) in animal models administer it via a parenteral route (e.g., intraperitoneal (IP) or intravenous injection (IV)), however, the common route of administration for human users is pulmonary (e.g., smoking or vapourizing marijuana). A vapourized Δ9-THC delivery system for rodents was developed and used to compare the effects of pulmonary and parenteral Δ9-THC administration on blood cannabinoid levels and behaviour. MethodsSprague–Dawley rats were exposed to pulmonary Δ9-THC (1, 5, and 10mg of inhaled vapour) delivered via a Volcano® vapourizing device (Storz and Bickel, Germany) or to parenteral Δ9-THC (0.25, 0.5, 1.0, and 1.5mg/kg injected IP). Quantification of Δ9-THC and its psychoactive metabolite, 11-hydroxy-Δ9-THC (11-OH-Δ9-THC), in blood was determined by liquid chromatography/mass spectrometry (LC/MS). In order to verify the potential for the vapourization procedure to produce a robust conditioned place preference (CPP) or conditioned place avoidance CPA, classical conditioning procedures were systematically varied by altering the exposure time (10 or 20min) and number of exposed rats (1 or 2) while maintaining the same vapourization dose (10mg). ResultsBlood collected at 20min intervals showed similar dose-dependent and time-dependent changes in Δ9-THC and 11-OH-Δ9-THC for both pulmonary and parenteral administration of Δ9-THC. However, vapourized Δ9-THC induced CPP under certain conditions whereas IP-administered Δ9-THC induced CPA. DiscussionThese results support and extend the limited evidence (e.g., in humans, Naef et al., 2004; in rodents, Niyuhire et al., 2007) that Δ9-THC produces qualitatively different effects on behaviour depending upon the route of administration.

A vapourized Δ9-tetrahydrocannabinol (Δ9-THC) delivery system part II: Comparison of behavioural effects of pulmonary versus parenteral cannabinoid exposure in rodents

IntroductionStudies of the rewarding and addictive properties of cannabinoids using rodents as animal models of human behaviour often fail to replicate findings from human studies. Animal studies typically employ parenteral routes of administration, whereas humans typically smoke cannabis, thus discrepancies may be related to different pharmacokinetics of parenteral and pulmonary routes of administration. Accordingly, a novel delivery system of vapourized Δ9-tetrahydrocannabinol (Δ9-THC) was developed and assessed for its pharmacokinetic, pharmacodynamic, and behavioural effects in rodents. A commercially available vapourizer was used to assess the effects of pulmonary (vapourized) administration of Δ9-THC and directly compared to parenteral (intraperitoneal, IP) administration of Δ9-THC. Methods:Sprague–Dawley rats were exposed to pure Δ9-THC vapour (1, 2, 5, 10, and 20mg/pad), using a Volcano® vapourizing device (Storz and Bickel, Germany) or IP-administered Δ9-THC (0.1, 0.3, 0.5, 1.0mg/kg), and drug effects on locomotor activity, food and water consumption, and cross-sensitization to morphine (5mg/kg) were measured. Results:Vapourized Δ9-THC significantly increased feeding during the first hour following exposure, whereas IP-administered Δ9-THC failed to produce a reliable increase in feeding at all doses tested. Acute administration of 10mg of vapourized Δ9-THC induced a short-lasting stimulation in locomotor activity compared to control in the first of four hours of testing over 7days of repeated exposure; this chronic exposure to 10mg of vapourized Δ9-THC did not induce behavioural sensitization to morphine. Discussion:These results suggest vapourized Δ9-THC administration produces behavioural effects qualitatively different from those induced by IP administration in rodents. Furthermore, vapourized Δ9-THC delivery in rodents may produce behavioural effects more comparable to those observed in humans. We conclude that some of the conflicting findings in animal and human cannabinoid studies may be related to pharmacokinetic differences associated with route of administration.

Chronic administration during early adulthood does not alter the hormonally-dependent disruptive effects of delta-9-tetrahydrocannabinol (Δ9-THC) on complex behavior in female rats

This study examined whether chronic Δ9-THC during early adulthood would produce the same hormonally-dependent deficits in learning that are produced by chronic Δ9-THC during adolescence. To do this, either sham-operated (intact) or ovariectomized (OVX) female rats received daily saline or 5.6mg/kg of Δ9-THC i.p. for 40days during early adulthood. Following chronic administration, and a drug-free period to train both a learning and performance task, acute dose-effect curves for Δ9-THC (0.56–10mg/kg) were established in each of the four groups (intact/saline, intact/THC, OVX/saline and OVX/THC). The dependent measures of responding under the learning and performance tasks were the overall response rate and the percentage of errors. Although the history of OVX and chronic Δ9-THC in early adulthood did not significantly affect non-drug or baseline behavior under the tasks, acute administration of Δ9-THC produced both rate-decreasing and error-increasing effects on learning and performance behavior, and these effects were dependent on their hormone condition. More specifically, both intact groups were more sensitive to the rate-decreasing and error-increasing effects of Δ9-THC than the OVX groups irrespective of chronic Δ9-THC administration, as there was no significant main effect of chronic treatment and no significant interaction between chronic treatment (saline or Δ9-THC) and the dose of Δ9-THC administered as an adult. Post mortem examination of 10 brain regions also indicated there were significant differences in agonist-stimulated GTPγS binding across brain regions, but no significant effects of chronic treatment and no significant interaction between the chronic treatment and cannabinoid signaling. Thus, acute Δ9-THC produced hormonally-dependent effects on learning and performance behavior, but a period of chronic administration during early adulthood did not alter these effects significantly, which is contrary to what we and others have shown for chronic administration during adolescence.

ΔFosB induction correlates inversely with CB1 receptor desensitization in a brain region-dependent manner following repeated Δ9-THC administration

Repeated Δ9-tetrahydrocannabinol (THC) administration produces desensitization and downregulation of cannabinoid type 1 receptors (CB1Rs) in the brain, but the magnitude of these adaptations varies among regions. CB1Rs in the striatum and its output regions exhibit the least magnitude and slowest development of desensitization and downregulation. The molecular mechanisms that confer these region-dependent differences are not known. The stable transcription factor, ΔFosB, is induced in the striatum following repeated THC administration and could regulate CB1Rs. To directly compare the regional profile of ΔFosB induction and CB1R desensitization and downregulation, mice were treated with THC (10 mg/kg) or vehicle for 13.5 days. CP55,940-stimulated [35S]GTPγS autoradiography and immunohistochemistry were performed to measure CB1R desensitization and downregulation, respectively, and ΔFosB expression was measured by immunoblot. Significant CB1R desensitization and downregulation occurred in the prefrontal cortex, lateral amygdala and hippocampus; desensitization was found in the basomedial amygdala and no changes were seen in remaining regions. ΔFosB was induced in the prefrontal cortex, caudate-putamen, nucleus accumbens and lateral amygdala. An inverse regional relationship between ΔFosB expression and CB1R desensitization was found, such that regions with the greatest ΔFosB induction did not exhibit CB1R desensitization and areas without ΔFosB induction had the greatest desensitization, with remaining regions exhibiting intermediate levels of both. Dual immunohistochemistry in the striatum showed both CB1R co-localization with ΔFosB in cells and CB1R puncta surrounding ΔFosB-positive cells. THC-induced expression of ΔFosB was absent in the striatum of CB1R knockout mice. These data suggest that transcriptional targets of ΔFosB might inhibit CB1R desensitization and/or that ΔFosB induction could be limited by CB1R desensitization.

Delta-9-tetrahydrocannabinol-induced dopamine release as a function of psychosis risk: 18F-fallypride positron emission tomography study.

Cannabis use is associated with psychosis, particularly in those with expression of, or vulnerability for, psychotic illness. The biological underpinnings of these differential associations, however, remain largely unknown. We used Positron Emission Tomography and 18F-fallypride to test the hypothesis that genetic risk for psychosis is expressed by differential induction of dopamine release by Δ9-THC (delta-9-tetrahydrocannabinol, the main psychoactive ingredient of cannabis). In a single dynamic PET scanning session, striatal dopamine release after pulmonary administration of Δ9-THC was measured in 9 healthy cannabis users (average risk psychotic disorder), 8 patients with psychotic disorder (high risk psychotic disorder) and 7 un-related first-degree relatives (intermediate risk psychotic disorder). PET data were analyzed applying the linear extension of the simplified reference region model (LSRRM), which accounts for time-dependent changes in 18F-fallypride displacement. Voxel-based statistical maps, representing specific D2/3 binding changes, were computed to localize areas with increased ligand displacement after Δ9-THC administration, reflecting dopamine release. While Δ9-THC was not associated with dopamine release in the control group, significant ligand displacement induced by Δ9-THC in striatal subregions, indicative of dopamine release, was detected in both patients and relatives. This was most pronounced in caudate nucleus. This is the first study to demonstrate differential sensitivity to Δ9-THC in terms of increased endogenous dopamine release in individuals at risk for psychosis.

Induction of Psychosis byΔ9-Tetrahydrocannabinol Reflects Modulation of Prefrontal and Striatal Function During Attentional Salience Processing

The aberrant processing of salience is thought to be a fundamental factor underlying psychosis. Cannabis can induce acute psychotic symptoms, and its chronic use may increase the risk of schizophrenia. We investigated whether its psychotic effects are mediated through an influence on attentional salience processing. To examine the effects of Δ9-tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD) on regional brain function during salience processing. Volunteers were studied using event-related functional magnetic resonance imaging on 3 occasions after administration of Δ9-THC, CBD, or placebo while performing a visual oddball detection paradigm that involved allocation of attention to infrequent (oddball) stimuli within a string of frequent (standard) stimuli. University center. Fifteen healthy men with minimal previous cannabis use. Symptom ratings, task performance, and regional brain activation. During the processing of oddball stimuli, relative to placebo, Δ9-THC attenuated activation in the right caudate but augmented it in the right prefrontal cortex. Δ9-Tetrahydrocannabinol also reduced the response latency to standard relative to oddball stimuli. The effect of Δ9-THC in the right caudate was negatively correlated with the severity of the psychotic symptoms it induced and its effect on response latency. The effects of CBD on task-related activation were in the opposite direction of those of Δ9-THC; relative to placebo, CBD augmented left caudate and hippocampal activation but attenuated right prefrontal activation. Δ9-Tetrahydrocannabinol and CBD differentially modulate prefrontal, striatal, and hippocampal function during attentional salience processing. These effects may contribute to the effects of cannabis on psychotic symptoms and on the risk of psychotic disorders.

Δ9‐THC increases endogenous AHA1 expression in rat cerebellum and may modulate CB1 receptor function during chronic use

To characterize the long-term effects of adolescent marijuana abuse, we performed a proteomic analysis of cerebellar extracts from adult female rats with and without ovariectomy that were treated with Δ9-THC for 40 days during adolescence. Six proteins were found to significantly differ among the four treatment groups, with Δ9-THC and ovariectomy (OVX) decreasing the mitochondrial proteins, pyruvate carboxylase and NADH dehydrogenase, whereas the levels of putative cytosolic molecular chaperones NM23B, translationally controlled tumor protein, DJ-1 and activator of heat-shock 90kDa protein ATPase homolog 1 (AHA1) were increased. We further analyzed the effects of AHA1, a HSP90 co-chaperone, on CB1R and CB2R trafficking and signaling in transfected HEK293T and Neuro-2A cells. In HEK293T cells, AHA1 over-expression enhanced plasma membrane levels of CB1R and increased CB1R-mediated effects on cAMP levels and on MAPK phosphorylation. AHA1 over-expression also enhanced cell surface levels of endogenous CB1R and the effects of Δ9-THC on the cAMP levels in Neuro-2A cells. In contrast, over-expression of AHA1 did not affect the subcellular localization and signaling of CB2R. Our data indicate that chronic Δ9-THC administration in adolescence altered the endogenous levels of specialized proteins in the cerebellum, such as AHA1, and that this protein can change CB1R cell surface levels and signaling.

Role of the central autonomic nervous system in the hypotension and bradycardia induced by (-)-delta 9-trans-tetrahydrocannabinol.

Abstract (-)-Δ9-trans-Tetrahydrocannabinol (Δ9-THC), when given intravenously (2 mg kg−1) to cats, produced marked decreases in blood pressure and heart rate which developed gradually and were of prolonged duration. Cervical spinal transection (C1-C2) abolished these effects whereas surgical removal of neurogenic tone to the myocardium selectively eliminated the bradycardia. Bilateral vagotomy alone did not modify the action of Δ9-THC upon heart rate or blood pressure. Recordings of spontaneous sympathetic outflow in the inferior cardiac nerve indicated a rapid reduction in neural discharge rate after Δ9-THC administration. These observations support the hypothesis that Δ9-THC produces a cardiodecellerator and hypotensive effect by acting at some level within the sympathetic nervous system. Experiments conducted to investigate transmission in the superior cervical and stellate ganglia demonstrated that Δ9-THC did not alter ganglionic function. Also, responses to intravenous isoprenaline and noradrenaline were unchanged which suggested that Δ9-THC did not interact with α- or β- adrenoceptors. The possible action of Δ9-THC on central sympathetic structures was investigated by perfusion of Δ9-THC into the lateral cerebral ventricle. Δ9-THC so administered produced a significant reduction in heart rate without a substantial lowering of blood pressure. Tritiated or 14C-Δ9-THC perfused into the lateral ventricle demonstrated that the amount of radioactive compound passing into the peripheral circulation was insignificant and could not account for the decrease in heart rate. The current data are in agreement with the proposal that Δ9-THC produces cardiovascular alterations by an action on the central nervous system which results in a decrease in sympathetic tone.

Clozapine and SCH 23390 prevent the spatial working memory disruption induced by Δ9-THC administration into the medial prefrontal cortex

Marijuana (Cannabis sativa) is one of the most widely used illicit drugs in the world. Its use is associated with impairments in cognitive function. We previously reported that Δ9-tetrahydrocannabinol (Δ9-THC), the primary psychoactive component of marijuana, impaired spatial working memory in the radial maze task when injected intracortically (IC) into the medial prefrontal cortex (mPFC) of rats. Here, we used this paradigm to evaluate the involvement of prefrontal dopamine receptors in working memory disruption induced by Δ9-THC. Intracortical pre-treatment of animals with either the D1- or D2-like dopamine receptor antagonists SCH 23390 or clozapine, respectively, significantly reduced the number of errors rats made in the radial maze following treatment with Δ9-THC also administered intracortically. These results were obtained in the absence of locomotor impairment, as evidenced by the time spent in each arm a rat visited. Our findings suggest that prefrontal dopamine receptors are involved in Δ9-THC-induced disruption of spatial working memory. This interaction between the cannabinoid system and dopamine release in the PFC contributes to new directions in research and to treatments for cognitive dysfunctions associated with drug abuse and dependence.

Separate and combined effects of the cannabinoid agonists nabilone and Δ9-THC in humans discriminating Δ9-THC

BackgroundAgonist replacement treatment is a promising strategy to manage cannabis-use disorders. The aim of this study was to assess the combined effects of the synthetic cannabinoid agonist nabilone and Δ9-tetrahydrocannabinol (Δ9-THC) using drug-discrimination procedures, which are sensitive to drug interactions. Testing the concurrent administration of nabilone and Δ9-THC was also conducted to provide initial safety and tolerability data, which is important because cannabis users will likely lapse during treatment. MethodsSix cannabis users learned to discriminate 30mg oral Δ9-THC from placebo and then received nabilone (0, 1 and 3mg) and Δ9-THC (0, 5, 15 and 30mg), alone and in combination. Subjects completed the multiple-choice procedure to assess drug reinforcement, and self-report, task performance and physiological measures were collected. ResultsΔ9-THC and nabilone alone shared discriminative-stimulus effects with the training dose of Δ9-THC, increased crossover point on the multiple-choice procedure, produced overlapping subject ratings and decreased skin temperature. Nabilone alone also elevated heart rate. In combination, nabilone shifted the discriminative-stimulus effects of Δ9-THC leftward/upward and enhanced Δ9-THC effects on the other outcome measures. ConclusionsThese results replicate a previous study demonstrating that nabilone shares agonist effects with the active constituent of cannabis in cannabis users, and contribute further by indicating that nabilone would likely be safe and well tolerated when combined with cannabis. These data support the conduct of future studies to determine if nabilone treatment would produce cross-tolerance to the abuse-related effects of cannabis and reduce cannabis use.

Long-term behavioral and pharmacodynamic effects of delta-9-tetrahydrocannabinol in female rats depend on ovarian hormone status

Abuse of Δ⁹-THC by females during adolescence may produce long-term deficits in complex behavioral processes such as learning, and these deficits may be affected by the presence of ovarian hormones. To assess this possibility, 40 injections of saline or 5.6 mg/kg of Δ⁹-THC were administered i.p. daily during adolescence to gonadally intact or ovariectomized (OVX) female rats, yielding four treatment groups (intact/saline, intact/THC, OVX/saline, and OVX/ THC). Δ⁹-THC (0.56-10 mg/kg) was then re-administered to each of the four groups during adulthood to examine their sensitivity to its disruptive effects. The behavioral task required adult subjects to both learn (acquisition component) different response sequences and repeat a known response sequence (performance component) daily. During baseline (no injection) and control (saline injection) sessions, OVX subjects had significantly higher response rates and lower percentages of error in both behavioral components than the intact groups irrespective of saline or Δ⁹-THC administration during adolescence; the intact group that received Δ⁹-THC had the lowest response rates in each component. Upon re-administration of Δ⁹-THC, the groups that received adolescent ovariectomy alone, adolescent Δ⁹-THC administration alone, or both treatments were found to be less sensitive to the rate-decreasing effects, and more sensitive to the error-increasing effects of Δ⁹-THC than the control group (i.e. intact subjects that received saline during adolescence). Neurochemical analyses of the brains from each adolescent-treated group indicated that there were also persistent effects on cannabinoid type-1 (CB-1) receptor levels in the hippocampus and striatum that depended on the brain region and the presence of ovarian hormones. In addition, autoradiographic analyses of the brains from adolescent-treated, but behaviorally naïve, subjects indicated that ovariectomy and Δ⁹-THC administration produced effects on receptor coupling in some of the same brain regions. In summary, chronic administration of Δ⁹-THC during adolescence in female rats produced long-term effects on operant learning and performance tasks and on the cannabinoid system that were mediated by the presence of ovarian hormones, and that altered their sensitivity to Δ⁹-THC as adults.

Oral Δ9-Tetrahydrocannabinol Improved Refractory Gilles de la Tourette Syndrome in an Adolescent by Increasing Intracortical Inhibition

To describe the clinical course of the Delta 9-tetrahydrocannabinol (Delta 9-THC) treatment of a boy with Gilles de la Tourette Syndrome (TS) and comorbid attention-deficit/hyperactivity disorder (ADHD) in relation to Delta 9-THC plasma levels and intracortical inhibition measured by transcranial magnetic stimulation. The clinical course and pharmacological and neurophysiological measures are reported in a 15-year-old boy with treatment refractory TS plus ADHD leading to severe physical and psychosocial impairment. Administration of Delta 9-THC improved tics considerably without adverse effects, allowing parallel stimulant treatment of comorbid ADHD. Along with the Delta 9-THC treatment, intracortical inhibition was increased, reflected in the enhanced short-interval intracortical inhibition and the prolongation of the cortical silent period. Our observation suggests that Delta 9-THC might be a successful alternative in patients with severe TS refractory to classic treatment. Particularly in the case of stimulant-induced exacerbation of tics, Delta 9-THC might enable successful treatment of comorbid ADHD. The enhancement of intracortical inhibition might be mediated by modulating release of several neurotransmitters including dopamine and gamma-aminobutyric acid. Further studies are needed to substantiate our findings.

Two opposite effects of Δ9-tetrahydrocannabinol on subthalamic nucleus neuron activity: Involvement of GABAergic and glutamatergic neurotransmission

Activation of CB1 cannabinoid receptors in the basal ganglia interferes with movement regulation. The aim of this study was to characterize the effect of Delta(9)-tetrahydrocannabinol (Delta(9)-THC) on neurons in the subthalamic nucleus (STN) and to elucidate the mechanisms involved in this effect using single-unit extracellular recordings in anesthetized rats. Administration of Delta(9)-THC (0.25-2 mg/kg, i.v.) stimulated (by 107% +/- 32%) neurons mainly recorded in the ventromedial portion of the caudal STN, whereas it inhibited (by 65% +/- 4%) neurons recorded in the dorsolateral portion of the rostral STN. The CB1 receptor antagonist rimonabant (1 mg/kg, i.v.) completely reverted these effects. The excitatory effect of Delta(9)-THC on STN neurons was not observed after antagonism of GABA(A) receptors by bicuculline administration (10 ng, icv.) or after chemical lesion of the globus pallidus with ibotenic acid. The inhibitory effect was abolished when excitatory amino acid receptors were blocked by kynurenic acid (0.5 mumol, icv.). These results indicate that CB1 receptor activation modulates STN neuron activity by indirect mechanisms involving glutamatergic and GABAergic neurotransmission.

Chronic cannabinoid administration increases CD8+ lymphocyte apoptosis during early Simian Immunodeficiency Virus infection in rhesus macaques.

Δ9‐tetrahydrocannabinol (Δ9‐THC), the primary psychoactive component of marijuana, is used to ameliorate Acquired Immune Deficiency Syndrome associated wasting. Cannabinoid receptors are expressed in cells of the immune system, raising the possibility that chronic Δ9‐THC use may impact on immune function, potentially affecting progression of human immunodeficiency virus infection. We examined the effects of chronic Δ9‐THC administration (0.32mg/kg i.m., 2 X daily, starting 30 d pre‐infection) on the early changes in lymphocyte counts, subpopulations, apoptosis, and proliferation in young adult male rhesus macaques at day 30 post simian immunodeficiency virus (SIVmac251; 100 TCID50/ml, i.v.) infection. SIV+ animals had lower (40%) lymphocyte CD4+/CD8+ ratio, 3‐7‐fold higher CD8+Ki67 expression (proliferation marker), without significant changes in total lymphocyte count, or CD4+ and CD8+caspase 3 expression (apoptosis marker) than time‐matched SIV‐ controls. THC‐treated SIV+ animals showed similarly decreased CD4+/CD8+, increased CD8+Ki67 expression, but higher expression of caspase 3 in CD8+ lymphocytes than time‐matched THC‐treated SIV‐ controls. We speculate that though modest the Δ9‐THC‐induced increase in CD8+ apoptosis may accentuate the CD4+/CD8+ imbalance during later stages of SIV infection. Supported NIDA‐020419‐01A1

Δ9-Tetrahydrocannabinol inhibits cell cycle progression by downregulation of E2F1 in human glioblastoma multiforme cells

Background. The active components of Cannabis sativa L., Cannabinoids, traditionally used in the field of cancer for alleviation of pain, nausea, wasting and improvement of well-being have received renewed interest in recent years due to their diverse pharmacologic activities such as cell growth inhibition, anti-inflammatory activity and induction of tumor regression. Here we used several experimental approaches, which identified delta-9-tetrahydrocannabinol (Δ9-THC) as an essential mediator of cannabinoid antitumoral action. Methods and results. Administration of Δ9-THC to glioblastoma multiforme (GBM) cell lines results in a significant decrease in cell viability. Cell cycle analysis showed G0/1 arrest and did not reveal occurrence of apoptosis in the absence of any sub-G1 populations. Western blot analyses revealed a THC altered cellular content of proteins that regulate cell progression through the cell cycle. The cell content of E2F1 and Cyclin A, two proteins that promote cell cycle progression, were suppressed in both U251-MG and U87-MG human glioblastoma cell lines, whereas the level of p16INK4A, a cell cycle inhibitor was upregulated. Transcription of thymidylate synthase (TS) mRNA, which is promoted by E2F1, also declined as evident by QRT-PCR. The decrease in E2F1 levels resulted from proteasome mediated degradation and was prevented by proteasome inhibitors. Conclusions. Δ9-THC is shown to significantly affect viability of GBM cells via a mechanism that appears to elicit G1 arrest due to downregulation of E2F1 and Cyclin A. Hence, it is suggested that Δ9-THC and other cannabinoids be implemented in future clinical evaluation as a therapeutic modality for brain tumors.

Effects of Δ9-tetrahydrocannabinol, ( R)-methanandamide, SR 141716, and d-amphetamine before and during daily Δ9-tetrahydrocannabinol dosing

We examined the effects of Δ9-tetrahydrocannabinol (Δ9-THC), ( R)-(+)-arachidonyl-1′-hydroxy-2′-propylamide (( R)-methanandamide, AM 356), SR 141716, and d-amphetamine on fixed-ratio (FR) responding maintained by food in rats before and during daily dosing with Δ9-THC. Rats responded under a FR 10 schedule of food reinforcement. Cumulative dose–response curves for the various drugs were determined before and during daily Δ9-THC administration. All four drugs dose-dependently decreased responding both before and during daily dosing with Δ9-THC (18 mg/kg/day). The dose–response curves for both Δ9-THC and ( R)-methanandamide were shifted to the right with daily dosing with Δ9-THC, indicating tolerance to the effects of Δ9-THC and cross-tolerance to the effects of ( R)-methanandamide. The doses of d-amphetamine examined produced similar effects both before and during daily dosing with Δ9-THC. The effects of SR 141716 were not consistently altered by daily Δ9-THC administration. These results indicate that tolerance develops to the effects of Δ9-THC, when Δ9-THC is administered repeatedly. These results also indicate that cross-tolerance to ( R)-methanandamide develops with repeated Δ9-THC administration.

Increased mortality, hypoactivity, and hypoalgesia in cannabinoid CB1 receptor knockout mice.

Delta9-Tetrahydrocannabinol (Delta9-THC), the major psychoactive ingredient in preparations of Cannabis sativa (marijuana, hashish), elicits central nervous system (CNS) responses, including cognitive alterations and euphoria. These responses account for the abuse potential of cannabis, while other effects such as analgesia suggest potential medicinal applications. To study the role of the major known target of cannabinoids in the CNS, the CB1 cannabinoid receptor, we have produced a mouse strain with a disrupted CB1 gene. CB1 knockout mice appeared healthy and fertile, but they had a significantly increased mortality rate. They also displayed reduced locomotor activity, increased ring catalepsy, and hypoalgesia in hotplate and formalin tests. Delta9-THC-induced ring-catalepsy, hypomobility, and hypothermia were completely absent in CB1 mutant mice. In contrast, we still found Delta9-THC-induced analgesia in the tail-flick test and other behavioral (licking of the abdomen) and physiological (diarrhea) responses after Delta9-THC administration. Thus, most, but not all, CNS effects of Delta9-THC are mediated by the CB1 receptor.

Mesolimbic dopaminergic decline after cannabinoid withdrawal.

The mesolimbic dopamine system has recently been implicated in the long-term aversive consequences of withdrawal from major drugs of abuse. In the present study we sought to determine whether mesolimbic dopamine neurons are involved in the neurobiologic mechanisms underlying withdrawal from chronic cannabinoid exposure. Rats were treated chronically with the major psychoactive ingredient of hashish and marijuana, Delta9-tetrahydrocannabinol (Delta9-THC). Administration of the cannabinoid antagonist SR 141716A precipitated an intense behavioral withdrawal syndrome, whereas abrupt Delta9-THC suspension failed to produce overt signs of abstinence. In contrast, both groups showed a reduction in dopamine cells activity as indicated by extracellular single unit recordings from antidromically identified meso-accumbens dopamine neurons. The administration of Delta9-THC to spontaneously withdrawn rats restored neuronal activity. Conversely, SR 141716A produced a further decrease of spontaneous activity in cannabinoid-treated although it was ineffective in control rats. These data indicate that withdrawal from chronic cannabinoid administration is associated with reduced dopaminergic transmission in the limbic system, similar to that observed with other addictive drugs; these changes in neuronal plasticity may play a role in drug craving and relapse into drug addiction.

Effects of Chronic Treatment with Δ9-Tetrahydrocannabinol on Cannabinoid-Stimulated [35S]GTPγS Autoradiography in Rat Brain

Chronic Delta9-tetrahydrocannabinol (Delta9-THC) administration produces tolerance to cannabinoid effects, but alterations in signal transduction that mediate these changes are not yet known. The present study uses in vitro autoradiography of agonist-stimulated [35S]GTPgammaS binding to localize cannabinoid receptor-activated G-proteins after chronic Delta9-THC treatment. Cannabinoid (WIN 55212-2)-stimulated [35S]GTPgammaS binding was performed in brain sections from rats treated chronically with 10 mg/kg Delta9-THC for 21 d. Control animals received saline or an acute injection of Delta9-THC. Acute Delta9-THC treatment had no effect on basal or WIN 55212-2-stimulated [35S]GTPgammaS binding. After chronic Delta9-THC treatment, net WIN 55212-2-stimulated [35S]GTPgammaS binding was reduced significantly (up to 70%) in most brain regions, including the hippocampus, caudate-putamen, perirhinal and entorhinal cortex, globus pallidus, substantia nigra, and cerebellum. In contrast, chronic Delta9-THC treatment had no effect on GABAB-stimulated [35S]GTPgammaS binding. In membranes and brain sections, Delta9-THC was a partial agonist, stimulating [35S]GTPgammaS by only 20% of the level stimulated by WIN 55212-2 and inhibiting WIN 55212-2-stimulated [35S]GTPgammaS at high concentrations. Because the EC50 of WIN 55212-2-stimulated [35S]GTPgammaS binding and the KD of cannabinoid receptor binding were unchanged by chronic Delta9-THC treatment, the partial agonist actions of Delta9-THC did not produce the decrease in cannabinoid-stimulated [35S]GTPgammaS binding. These results suggest that profound desensitization of cannabinoid-activated signal transduction mechanisms occurs after chronic Delta9-THC treatment.