Effects of Δ8- and Δ9-Tetrahydrocannabinol on experimentally induced seizures

Effects of Δ⁸- and Δ⁹-tetrahydrocannabinol (Δ⁸- and Δ⁹-THC) on three experimentally induced seizure models, i.e., audiogenic seizure (AS) test, maximal electroshock seizure (MES) test and pentylenetetrazol (PTZ)-induced seizure test were determined in the audiogenic rat. Both tetrahydrocannabinols possess a dose-related anticonvulsant effect against AS, MES and PTZ-induced maximal seizure. Although anticonvulsant potencies do not significantly differ, Δ⁸THC is three times more neurotoxic than Δ⁹THC. In addition, both THC's are without effect on minimal seizure and lethality induced by PTZ. Furthermore, the low protective indexes (TD50/ED50) determined in this study suggest that Δ⁸ and Δ⁹ THC may have poor therapeutic potentials as antiepileptic drugs.     

Interactions between cannabidiol and delta9-THC during abstinence in morphine-dependent rats.

Male rats, each implanted with a pellet containing 75 mg morphine, were administered naloxone 72 hours later to precipitate abstinence. Two hours before naloxone, animals were pretreated acutely with either 10 mg/kg cannabidiol (CBD) or the vehicle. One hour later, an injection of the vehicle or a low dose of Δ⁹-THC that we have shown to exhibit slight efficacy in attenuating morphine abstinence signs was administered to each of the groups previously receiving the vehicle or CBD. Interactions between CBD and Δ⁹-THC were assessed during abstinence, precipitated one hour after the last series of injections. CBD had little effect on abstinence scores, but significantly increased the abstinence attenuating properties of Δ⁹-THC, Rotational behavior (turning), induced by Δ⁹-THC during abstinence, was also potentiated by CBD. These data extend previous reports of potentiation of pharmacological effects of THC by CBD to abstinence-attenuating properties and other effects of THC in morphine-dependent rats.     

Blocking of the effect of delta1 tetrahydrocannabinol (delta1THC) on spontaneous motor activity in the rat by immunization with a protein conjugate of delta1THC

Rats actively immunized with porcine gamma globulin- hemisuccinate-Δ¹-tetrahydrocannabinol (PγG-HS-Δ¹THC) showed higher spontaneous motor activity after intraperitoneal administration of Δ¹THC at a dose of 10 mg. per kg. than did rats immunized with a control antigen, porcine gamma globulin-hemisuccinate-phenol (PγG-HS-Phenol). The capacity to neutralize the effect of Δ¹THC was found to depend on the degree of immunization; thus, the difference in mean spontaneous motor activity after injection of Δ¹THC was significant in rats which had received five injections of the immunogen over a period of 86 days, and not in those which had received only two injections over a period of 34 days.In view of the observations that Δ¹-tetrahydrocannabinol induces a decrease in spontaneous motor activity in rats, the observed neutralization of the effect of δ¹THC in animals receiving multiple injections of protein conjugates of Δ¹THC may be due to the binding of the drug by anti-THC antibodies (which are expected to be produced on active immunization with these conjugates), thus preventing Δ¹THC from reaching drug-receptor sites.     

A Structural Insight into the Molecular Recognition of a (−)-Δ-Tetrahydrocannabinol and the Development of a Sensitive, One-Step, Homogeneous Immunocomplex-Based Assay for Its Detection

(−)-Δ⁹-Tetrahydrocannabinol (THC) is the main psychoactive compound found in cannabis. In this study, an anti-THC Fab fragment, designed T3, was isolated from a display library cloned from the spleen cells of a mouse immunized with a THC-bovine serum albumin conjugate, and the crystal structures of the T3 Fab in its free form and in complex with THC were determined at 1.9 Å and 2.0 Å resolution, respectively. The THC binding site of the T3 Fab is a narrow cavity: the n-pentyl group of THC protrudes deep into the interface area between the variable domains and the C₁₀ monoterpene moiety of the hapten is partially exposed to solvent. The metabolites of THC, with modifications in the C₁₀ monoterpene moiety, 11-nor-9-carboxy-Δ⁹-tetrahydrocannabinol and 11-hydroxy-Δ⁹-tetrahydrocannabinol, are bound by the T3 Fab with a higher affinity than THC. The crystal structures suggest that Ser52H and Arg53H of the T3 Fab are able to make hydrogen bonds with the metabolites, which leads to an increased binding against these metabolites. By developing a T3 Fab-Δ⁹-THC immunocomplex binding antibody from a naïve antibody phage display library, the specificity of the Δ⁹-THC binding is highly increased, which allows a one-step, homogeneous, fluorescence resonance energy transfer-based sensitive immunoassay, with a detection limit of 20 ng/ml from saliva samples.     

Differential effects of cannabinoid exposure and stress on plasma prolactin, growth hormone and corticosterone levels in male mice

Plasma prolactin (PRL) levels were reduced in stressed and non-stressed male mice after a single dose of Δ⁹-tetrahydrocannabinol (THC), the main psychoactive constituent of marihuana, while growth hormone (GH) levers were reduced only in non-stressed animals. Chronic treatment with THC did not affect PRL or GH levels under either condition. Neither acute nor repeated exposure to THC affected plasma corticosterone levels.In contrast to the affects of THC, acute exposure to cannabinol (CBN), a non-psychoactive ingredient in marihuana, increased plasma GH levels in non-stressed mice, while repeated CBN treatments reduced GH levels in stressed animals. Moreover, chronic CBN exposure resulted in decreased peripheral levels of corticosterone in both stressed and non-stressed mice, and reduced plasma PRL levels in stressed mice.Psychoactive and non-psychoactive components of marihuana can exert different effects on endocrine function and on responsivity to stress in male mice.     

Further evidence for a role of 2-phenylethylamine in the mode of action of delta 9-tetrahydrocannabinol

In rabbits, Δ⁹-tetrahydrocannabinol (Δ⁹-THC) increased the recovery of labeled 2-phenylethylamine (PEA) from brain following its intraventricular administration. Δ⁹-THC also enhanced the excitatory effect of iontophoretic PEA on cortical unit potentials. Although Δ⁹-THC induced sedation in mice, the subsequent injection of reserpine induced transient excitement. Low doses of PEA, which do not significantly alter the behavior of mice, induced marked excitement in mice pretreated with Δ⁹-THC. In mice treated with pargyline, Δ⁹-THC induced excitement (instead of sedation); this excitement was increased by PEA and reduced by phenylethanolamine. These results suggest that Δ⁹-THC inhibits the disposition of PEA. Since endogenous PEA may be one of the adrenergic ergotropic modulators, it may play a role in the euphoriant effect of marihuana.     

Effects of Δ9-tetrahydrocannabinol on the disposition of d-amphetamine in the rat

Δ⁹-Tetrahydrocannabinol (THC), 10 or 50 mg/kg, administered intragastrically one hour before intraperitoneal injection of ¹⁴C-d-amphetamine, 4 mg/kg, did not modify the disappearance from the blood or the tissue distribution of amphetamine in fasted rats. Furthermore, THC did not influence the urinary excretion of unchanged amphetamine or its major metabolite, p-hydroxyamphetamine, in these animals. However, when the interval between drug treatments was increased to two hours, THC, 10 mg/kg, minimally reduced the rate of disappearance of ¹⁴C-amphetamine from the blood of fasted rats. This effect was much more pronounced in rats which had food available throughout the experiment. THC also inhibited the urinary excretion of total radioactivity as well as ¹⁴C-amphetamine metabolites in fed but not in fasted animals during the first 4 hours following ¹⁴C-amphetamine injection. In addition, fasted rats excreted significantly more total radioactivity and unchanged ¹⁴C-amphetamine than fed rats during the 0 – 4 hour urine collection interval. The pH of urine collected during this and all other periods was significantly more acid for faster than fed rats. It is concluded that THC can inhibit amphetamine metabolism in rats depending upon the time interval between the administration of the two drugs and the dietary state of the animals.     

Stimulation of ALK by the growth factor midkine renders glioma cells resistant to autophagy-mediated cell death

Δ⁹-tetrahydrocannabinol (THC), the main active component of marijuana, promotes cancer cell death via autophagy stimulation. We find that activation of the tyrosine kinase receptor ALK by its ligand midkine interferes with the signaling mechanism by which THC promotes autophagy-mediated glioma cell death.     

Oxidative stress and cannabinoid receptor expression in type‐2 diabetic rat pancreas following treatment with Δ9‐THC

The objectives of study were (a) to determine alteration of feeding, glucose level and oxidative stress and (b) to investigate expression and localization of cannabinoid receptors in type‐2 diabetic rat pancreas treated with Δ⁹‐tetrahydrocannabinol (Δ⁹‐THC). Rats were randomly divided into four groups: control, Δ⁹‐THC, diabetes and diabetes + Δ⁹‐THC groups. Diabetic rats were treated with a single dose of nicotinamide (85 mg/kg) 15 min before injection of streptozotocin (65 mg/kg). Δ⁹‐THC was administered intraperitoneally at 3 mg/kg/day for 7 days. Body weights and blood glucose level of rats in all groups were measured on days 0, 7, 14 and 21. On day 15 after the Δ⁹‐THC injections, pancreatic tissues were removed. Blood glucose levels and body weights of diabetic rats treated with Δ⁹‐THC did not show statistically significant changes when compared with the diabetic animals on days 7, 14 and 21. Treatment with Δ⁹‐THC significantly increased pancreas glutathione levels, enzyme activities of superoxide dismutase and catalase in diabetes compared with non‐treatment diabetes group. The cannabinoid 1 receptor was found in islets, whereas the cannabinoid 2 receptor was found in pancreatic ducts. Their localization in cells was both nuclear and cytoplasmic. We can suggest that Δ⁹‐THC may be an important agent for the treatment of oxidative damages induced by diabetes. However, it must be supported with anti‐hyperglycaemic agents. Furthermore, the present study for the first time emphasizes that Δ⁹‐THC may improve pancreatic cells via cannabinoid receptors in diabetes. The aim of present study was to elucidate the effects of Δ⁹‐THC, a natural cannabinoid receptor agonist, on the expression and localization of cannabinoid receptors, and oxidative stress statue in type‐2 diabetic rat pancreas. Results demonstrate that the cannabinoid receptors are presented in both Langerhans islets and duct regions. The curative effects of Δ⁹‐THC can be occurred via activation of cannabinoid receptors in diabetic rat pancreas. Moreover, it may provide a protective effect against oxidative damage induced by diabetes. Thus, it is suggested that Δ⁹‐THC can be a candidate for therapeutic alternatives of diabetes symptoms. Copyright © 2014 John Wiley & Sons, Ltd.     

Structural divergence among cannabinoids influences membrane dynamics: A H Solid-State NMR analysis

Cannabinoids are compounds that can modulate neuronal functions and immune responses via their activity at the CB₁ receptor. We used ²H NMR order parameters and relaxation rate determination to delineate the behavior of magnetically aligned phospholipid bilayers in the presence of several structurally distinct cannabinoid ligands. THC (Δ⁹-Tetrahydrocannabinol) and WIN-55,212-2 were found to lower the phase transition temperature of the DMPC and to destabilize their acyl chains leading to a lower average S_CD (≈ 0.13), while methanandamide and CP-55,940 exhibited unusual properties within the lipid bilayer resulting in a greater average S_CD (≈ 0.14) at the top of the phospholipid upper chain. The CB₁ antagonist AM281 had average S_CD values that were higher than the pure DMPC lipids, indicating a stabilization of the lipid bilayer. R_1Z versus |S_CD|² plots indicated that the membrane fluidity is increased in the presence of THC and WIN-55,212-2. The interaction of CP-55,940 with a variety of zwitterionic and charged membranes was also assessed. The unusual effect of CP-55,940 was present only in bicelles composed of DMPC. These studies strongly suggest that cannabinoid action on the membrane depends upon membrane composition as well as the structure of the cannabinoid ligands.     

A qualitative and quantitative HPTLC densitometry method for the analysis of cannabinoids in Cannabis sativa L.

Introduction – Cannabis and cannabinoid based medicines are currently under serious investigation for legitimate development as medicinal agents, necessitating new low‐cost, high‐throughput analytical methods for quality control. Objective – The goal of this study was to develop and validate, according to ICH guidelines, a simple rapid HPTLC method for the quantification of Δ⁹‐tetrahydrocannabinol (Δ⁹‐THC) and qualitative analysis of other main neutral cannabinoids found in cannabis. Methodology – The method was developed and validated with the use of pure cannabinoid reference standards and two medicinal cannabis cultivars. Accuracy was determined by comparing results obtained from the HTPLC method with those obtained from a validated HPLC method. Results – Δ⁹‐THC gives linear calibration curves in the range of 50–500 ng at 206 nm with a linear regression of y = 11.858x + 125.99 and r² = 0.9968. Conclusion – Results have shown that the HPTLC method is reproducible and accurate for the quantification of Δ⁹‐THC in cannabis. The method is also useful for the qualitative screening of the main neutral cannabinoids found in cannabis cultivars. Copyright © 2009 John Wiley & Sons, Ltd.     

TRB3 links ER stress to autophagy in cannabinoid antitumoral action

Δ⁹-tetrahydrocannabinol (THC), the main active component of marijuana, is being investigated as a potential anti-tumoral agent. We find that THC stimulates an endoplasmic reticulum (ER) stress-related signaling pathway, which activates autophagy via inhibition of the Akt/mTORC1 axis. We also show that autophagy is upstream of apoptosis in cannabinoid-induced cancer cell death and that activation of this pathway is necessary for the anti-tumoral action of cannabinoids in vivo.     

Influence of anticonvulsant cannabinoids on posttetanic potentiation at isolated bullfrog ganglia

The effects of anticonvulsant cannabinoids on posttetanic potentiation (PTP) at bullfrog paravertebral ganglia $\text{in vitro}$ were investigated electrophysiologically. Two Δ⁹-tetrahydrocannabinol metabolites, 11-hydroxy-Δ⁹-tetrahydrocannabinol and 8α, 11-dihydroxy-Δ⁹-tetrahydrocannabinol, as well as cannabidiol, markedly depressed PTP. In contrast, Δ⁹-tetrahydrocannabinol had no such effect. Thus, the findings of the present study clearly demonstrate that pharmacological properties of these hydroxylated metabolites of Δ⁹-tetrahydrocannabinol are not identical to those of their parent compound.     

The effect of hypothermic doses of 1-Δ9-tetrahydrocannabinol on biogenic amine metabolism in selected parts of the rat brain

Hypothalamic and brainstem biogenic amine metabolism was investigated in rats following the administration of hypothermic doses of 1-Δ⁹-tetrahydrocannabinol (THC). The dose-dependent fall in body temperature induced by THC was both rapid in onset and prolonged in duration. The disruption in thermoregulation, however, was unaccompanied by any observed alteration in the concentration of turnover rate of 5-hydroxytryptamine (5-HT) in the brain tissues studied. Norepinephrine (NE) was also unchanged, with the exception of a reduction in the amount of brainstem NE 30 min after the administration of 50 mg/kg THC. These observations indicate that the hypothermic effect of THC is not mediated by changes in brain 5-HT or NE metabolism.     

Behavioral comparisons of the stereoisomers of tetrahydrocannabinols

The potencies of (?)-$\text{trans}$-Δ⁹-THC, (+)-$\text{trans}$-Δ⁹-THC, (+)-$\text{cis}$-Δ⁹-THC, (?)-$\text{trans}$-Δ⁸-THC and (+)-$\text{trans}$-Δ⁸-THC were compared in several different species. (?)-$\text{trans}$-Δ⁹-THC was 100 times more potent than (+)-$\text{trans}$-Δ⁹-THC in depressing schedule-controlled responding in monkeys. The (+)-$\text{trans}$ isomers were less effective than their corresponding (?)-$\text{trans}$ isomers in the dog static-ataxia test, but potency ratios could not be determined due to a lack of dose-responsiveness of the (+)-$\text{trans}$ isomers. However, it appeared that their potency differed by at least ten fold. The potency of (+)-$\text{cis}$-Δ⁹-THC in the dog static-ataxia test was comparable to that of (+)-$\text{trans}$-Δ⁹-THC. The hypothermia in mice produced by the (?) isomers of $\text{trans}$-Δ⁹-THC and $\text{trans}$-Δ⁸-THC were 9.1 and 30.4 times greater than that produced by their respective (+)-isomers. Also, the potency ratio of the (+)- and (?)-$\text{trans}$-Δ⁹-THC was 5.6 as measured by depression of spontaneous activity in mice. The magnitude of the potency ratios of the THC stereo-isomers is dependent upon the species and the pharmacological test used.     

Potentials evoked in the intermediolateral column by hypothalamic stimulation - suppression by delta 9-tetrahydrocannabinol

Δ⁹-Tetrahydrocannabinol (THC), the primary psychoactive component of marihuana produces pronounced effects on the cardiovascular system including bradycardia and hypotension. A decrease in sympathetic activity may contribute to these actions. In chloralose urethane anesthetized cats, THC (2 mg/kg, i.v.) produced significant bradycardia, hypotension and attenuation of threshold pressor responses induced by hypothalamic stimulation. Evoked potentials recorded in the intermediolateral cell column (ILC) by stimulation of these hypothalamic pressor sites were significantly altered after THC. Hypotension induced by histamine administration (5 μg/kg, i.v.) altered ILC potentials before and after THC. These results support the hypothesis that THC reduces sympathetic outflow and reversibly resets the level of central cardiovascular homeostasis.     

A cannabinoid analogue of Δ9‐tetrahydrocannabinol disrupts neural development in chick

Marijuana is the most commonly abused illicit drug by pregnant women. Its major psychoactive constituent, Δ⁹‐THC (Δ⁹‐tetrahydrocannabinol), crosses the placenta and accumulates in the f?tus, potentially harming its development. In humans, marijuana use in early pregnancy is associated with miscarriage, a fetal alcohol‐like syndrome, as well as learning disabilities, memory impairment, and ADHD in the offspring. Classical studies in the 1970 s have reached disparate conclusions as to the teratogenic effects of cannabinoids in animal models. Further, there is very little known about the immediate effects of Δ⁹‐THC on early embryogenesis. We have used the chick embryo as a model in order to characterize the effects of a water‐soluble Δ⁹‐THC analogue, O‐2545, on early development. Embryos were exposed to the drug (0.035 to 0.35 mg/ml) at gastrulation and assessed for morphological defects at stages equivalent to 9–14 somites. We report that O‐2545 impairs the formation of brain, heart, somite, and spinal cord primordia. Shorter incubation times following exposure to the drug show that O‐2545 interferes with the initial steps of head process and neural plate formation. Our results indicate that the administration of the cannabinoid O‐2545 during early embryogenesis results in embryotoxic effects and serves to illuminate the risks of marijuana exposure during the second week of pregnancy, a time point at which most women are unaware of their pregnancies. Birth Defects Res (Part B) 83:477–488, 2008. © 2008 Wiley‐Liss, Inc.     

Genetic differences in Δ9-tetrahydrocannabinol-induced facilitation of brain stimulation reward as measured by a rate-frequency curve-shift electrical brain stimulation paradigm in three different rat strains

Lewis, Fischer 344, and Sprague-Dawley rats were implanted with electrodes in the medial forebrain bundle and trained to lever press for brain stimulation reward using a ratefrequency curve-shift electrical brain stimulation paradigm based on a series of 16 pulse frequencies ranging from 25 to 141 Hz in descending order. Once reward thresholds were stable, rats were given 1.0 mg/kg Δ⁹-tetrahydrocannabinol (Δ⁹-THC), the psychoactive constituent in marijuana and hashish, or vehicle, by intraperitoneal injection. Lewis rats showed the most pronounced Δ⁹-THC-induced enhancement of brain reward functions. Sprague-Dawley rats showed an enhancement of brain reward functions that was approximately half that seen in Lewis rats. Brain reward functions in Fischer 344 rats were unaffected by Δ⁹-THC at the dose tested. These results are consistent with previous work showing Lewis rats to be highly sensitive to the rewarding properties of a variety of drugs of abuse, including opiates, cocaine, and alcohol, while Fischer 344 rats are relatively less sensitive. They extend such previous findings to cannabinoids, and further suggest that genetic variations to other cannabinoid effects may also exist.     

Lipid phase transitions in cytoplasmic and outer membranes of Escherichia coli

The cytoplasmic and outer membranes containing either trans-Δ⁹-octadecenoate, trans-Δ⁹-hexadecenoate or cis-Δ⁹-octadecenoate as predominant unsaturated fatty acid residues in the phospholipids were prepared from a fatty acid auxotroph, Escherichia coli strain K1062. Order-disorder transitions of the phospholipids were revealed in both fractions of the cell envelope by fluorescent probing or wide angle X-ray diffraction. The mid-transition temperatures, $\text{T}\text{t}$, and the range of the transition, $\text{ΔT}$, are similar in the outer and cytoplasmic membrane. Relative to the corresponding extracted lipids, 60–80% of the hydrocarbon chains take part in the transition in the cytoplasmic membrane whereas in the outer membrane only 25–40% of the chains become ordered. The results suggest that in the outer membrane part of the lipids form fluid domains in the form of mono- and/or bilayers.     

Effects of environmental stress or ACTH treatment during pregnancy on maternal and fetal plasma androstenedione in the rat

Prenatal stress applied during the last trimester of pregnancy has been shown to alter fetal development and influence adult sexual behavior. Since androstenedione (Δ⁴) has the potential to participate in differentiation processes, this study was designed to assess the effect of prenatal stress on maternal and fetal Δ⁴ titers. Restraint/illumination/heat (environmental stress) or ACTH injections were used to stress pregnant rat dams beginning on Day 14 of pregnancy. Blood samples and organ weights were obtained from nonpregnant animals, pregnant rats on Days 5, 10, 15, 18, and 20 of pregnancy, and fetuses on Days 18 and 20 of gestation. Maternal and male and female fetal Δ⁴ titers were determined by radioimmunoassay. ACTH and environmental stress significantly reduced fetal body weight and male anogenital distance. Environmental stress also significantly reduced the size of 20-day fetal adrenals and testes. Each treatment caused significant short-term (1 hr after treatment) and long-term (16 hr after treatment) elevation of maternal plasma Δ⁴ on Days 15 and 18 of gestation, but only short-term elevation of Δ⁴ titers on Day 20. ACTH treatment did not cause long-term elevation of fetal Δ⁴ although both ACTH treatment and environmental stress generated a significant short-term increase in fetal Δ⁴ titers. Environmental stress produced long-term elevation of fetal Δ⁴ in 18-day fetuses of both sexes and in 20-day female fetuses. It is concluded that maternal stress and exogenous ACTH significantly elevate maternal and fetal Δ⁴ titers during the prenatal period postulated to be critical in sexual differentiation of the rat brain.