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Cannabinoid receptor agonists are mitochondrial inhibitors: A unified hypothesis of how cannabinoids modulate mitochondrial function and induce cell death

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Abstract

Time-lapse microscopy of human lung cancer (H460) cells showed that the endogenous cannabinoid anandamide (AEA), the phyto-cannabinoid Delta-9-tetrahydrocannabinol (THC) and a synthetic cannabinoid HU 210 all caused morphological changes characteristic of apoptosis. Janus green assays of H460 cell viability showed that AEA and THC caused significant increases in OD 595 nm at lower concentrations (10-50 microM) and significant decreases at 100 microM, whilst HU 210 caused significant decreases at all concentrations. In rat heart mitochondria, all three ligands caused significant decreases in oxygen consumption and mitochondrial membrane potential. THC and HU 210 caused significant increases in mitochondrial hydrogen peroxide production, whereas AEA was without significant effect. All three ligands induced biphasic changes in either mitochondrial complex I activity and/or mitochondrial complex II-III activity. These data demonstrate that AEA, THC, and HU 210 are all able to cause changes in integrated mitochondrial function, directly, in the absence of cannabinoid receptors.

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... Data suggests that metformin possesses direct anti-cancer activity mediated by effective inhibition of mitochondrial complex I [7]. In addition, a number of other agents have been identified that act at the mitochondria to cause cancer cell death in vitro, including tricyclic antidepressants such as chlorimipramine [8]; vanilloids such as capsaicin [9]; and cannabinoids such as Δ9-tetrahydrocannabinol (THC), the psychoactive compound in Cannabis sativa [10]. ...
... The increase in mitochondrial complex IV activity that occurred between concentrations of 0.01 μM and 0.4 μM may have been due to changes in mitochondrial membrane fluidity; alterations in the fluidity of biological membranes are known to alter the maximum rate of an enzymecatalysed reactions (V max ) and the substrate concentration at which these proceed at half-maximal velocity (K m ) [28]. Furthermore, multi-phasic, drug concentration-dependent changes in mitochondrial complex activity similar to those measured here have previously been reported in relation to the effects of cannabinoid receptor agonists anandamide and Δ-9-tetrahydrocannabinol on the activity of mitochondrial complex I and complex II-III [10]. The decreased complex IV activity induced by combretastatin A4 at concentrations of 12.5 μM and above was likely due to a direct inhibitory effect following stabilisation of the increased fluidity of the membrane that occurred at lower concentrations [10]. ...
... Furthermore, multi-phasic, drug concentration-dependent changes in mitochondrial complex activity similar to those measured here have previously been reported in relation to the effects of cannabinoid receptor agonists anandamide and Δ-9-tetrahydrocannabinol on the activity of mitochondrial complex I and complex II-III [10]. The decreased complex IV activity induced by combretastatin A4 at concentrations of 12.5 μM and above was likely due to a direct inhibitory effect following stabilisation of the increased fluidity of the membrane that occurred at lower concentrations [10]. ...
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... Fisher's exact test). Specific subsets of genes involved in the glutamate receptor pathway and mitochondrial function were altered in response to acute or chronic THC exposure (Supplementary Table S4 8,24,25 . These results provide data to support the use of hiPSC-derived neurons as a model for investigating THC responses in an in vitro human neuronal system. ...
... Others have similarly reported that treatment of hiPSC-derived dopaminergic neurons with THC reproduces effects observed in other mouse and human models 27 . Mitochondrial pathway dysfunction has been linked to THC exposure 24,25,[28][29][30] and schizophrenia 26,31 , a convergence that is captured in our system (Figs. 1b-d; 2c). ...
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There is a strong association between cannabis use and schizophrenia but the underlying cellular links are poorly understood. Neurons derived from human-induced pluripotent stem cells (hiPSCs) offer a platform for investigating both baseline and dynamic changes in human neural cells. Here, we exposed neurons derived from hiPSCs to Δ9-tetrahydrocannabinol (THC), and identified diagnosis-specific differences not detectable in vehicle-controls. RNA transcriptomic analyses revealed that THC administration, either by acute or chronic exposure, dampened the neuronal transcriptional response following potassium chloride (KCl)-induced neuronal depolarization. THC-treated neurons displayed significant synaptic, mitochondrial, and glutamate signaling alterations that may underlie their failure to activate appropriately; this blunted response resembles effects previously observed in schizophrenia hiPSC- derived neurons. Furthermore, we show a significant alteration in THC-related genes associated with autism and intellectual disability, suggesting shared molecular pathways perturbed in neuropsychiatric disorders that are exacerbated by THC.
... In different models of animal epilepsy, such as maximal electroshock model of grand-mal seizure, the pilocarpine model of acquired epilepsy in rats, and the mouse pentylenetetrazole (PTZ) model of myoclonic seizures, a remarkable anticonvulsant effect was detected through CB1 receptor activation [7][8][9][10]. In addition to mentioned information, interestingly, past studies have shown that CBR agonist could inhibit the mitochondrial function [11]. Also, recent reports revealed that CB1 is functionally present at mitochondrial membranes (mtCB1) in the brain [12]. ...
... Past reports have demonstrated that CBR agonist could inhibit the mitochondrial function by decreasing the ATP level, complex I/II/III activity, and oxygen consumption [11]. It has been reported that the CBRs and K channels like K ATP and K ir 6.2 channel are in interaction with each other [22,23]. ...
Article
Available online xxxx Cannabinoid receptor (CBR) agonist could act as a protective agent against seizure susceptibility in animal models of epilepsy. Studies have shown that potassium channels could play a key role in ameliorating neuronal excitability. In this study, we attempted to evaluate how CBRs and Adenosine Tri-Phosphate (ATP)-sensitive potassium channels collaborate to affect seizure susceptibility by changing the clonic seizure threshold (CST). We used male Naval Medical Research Institute (NMRI) mice and treated them with the following drugs: cromakalim (a potassium channel opener, 10 μg/kg), glibenclamide (a potassium channel blocker, 0.03 and 1 mg/kg), 0.5 mg/kg of AM-251 (a selective CB1 antagonist), AM-630 (a selective CB2 antagonist), and 0.5, 3, and 10 mg/kg of WIN 55,212-2 (a nonselective agonist of CBRs); and CST was appraised after each type of administration. Also, we evaluated the ATP level of the hippocampus in each treatment to clarify the interaction between the cannabinoid system and potassium channel. Our results showed that administration of WIN 55,212-2 at 10 mg/kg significantly increased CST (P b 0.001). This change could be reversed by using AM-251(P b 0.001) but not AM-630. Also, either cromakalim (10 μg/kg) or glibenclamide (0.03 and 1 mg/kg) could not significantly affect the CST. In addition, glibenclamide (1 mg/kg) could reverse the anticonvulsant effect of WIN 55,212-2 (10 mg/kg) on CST (P b 0.001). However, the anticonvulsant effect was observed when cromakalim (10 μg/kg) was added to WIN 55,212-2 at its subeffective dose (3 mg/kg) in comparison to single-treated animals. Interestingly , we observed that CB1 agonist could significantly decrease ATP level. In conclusion, CB1 agonist accomplishes at least a part of its anticonvulsant actions through ATP-sensitive potassium channels, probably by decreasing the mitochondrial ATP level to open the potassium channel to induce its anticonvulsant effect.
... For instance, quercetin can modulate complex-1 of the ETC at low concentrations (<10 μM), reducing mitochondrial hydrogen peroxide production and inhibiting apoptosis (Lagoa, Graziani, Lopez-Sanchez, Garcia-Martinez, & Gutierrez-Merino, 2011). Similarly, berberine can also inhibit complex 1 (Turner et al., 2008), as can CBD, THC and resveratrol (Athanasiou et al., 2007;Fisar et al., 2014;Zini, Morin, Bertelli, Bertelli, & Tillement, 1999). Resveratrol, EGCG, curcumin and quercetin have also been shown to have effects on F0F1-ATPase/ATP synthase (Zheng & Ramirez, 2000). ...
... of the mode of action of many plant compounds, at least in animals, seems to involve upregulation of mitochondrial function as part of an anti-oxidant mechanism(Stevenson, 2012), as well as displaying ROS-scavenging independent actions (Sandoval-Acuna, Ferreira, & Speisky, 2014). Indeed, several well-known medicinal plant compounds can directly modulate mitochondrial function, including epigallocatechin gallate (EGCG), resveratrol(Battaglia, Salvi, & Toninello, 2005;Demos, Woolwine, Wilson, & McMillan, 1975;Oliveira, Nabavi, Daglia, Rastrelli, & Nabavi, 2016;Pereira et al., 2007;Ravanel, Tissut, & Douce, 1982;Usta et al., 2009;van Ginkel et al., 2007;Xie, Bezard, & Zhao, 2005), tetrahydrocannabinol (THC)(Athanasiou et al., 2007;Bartova & Birmingham, 1976;Fisar, Singh, & Hroudova, 2014;Mahoney & Harris, 1972) and CBD(Fisar et al., 2014;Mato, Victoria Sanchez-Gomez, & Matute, 2010;Olivas-Aguirre et al., 2019;Rimmerman et al., 2013;Ryan, Drysdale, Lafourcade, Pertwee, & Platt, 2009), as well as salicylic acid(de Souza et al., 2011;Gordon et al., 2002;Norman et al., 2004;Nulton-Persson, Szweda, & Sadek, 2004;Vlot, Dempsey, & Klessig, 2009) and curcumin ...
Article
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Medicine has utilised plant‐based treatments for millennia, but precisely how they work is unclear. One approach is to use a thermodynamic viewpoint that life arose by dissipating geothermal and/or solar potential. Hence, the ability to dissipate energy to maintain homeostasis is a fundamental principle in all life, which can be viewed as an accretion system where layers of complexity have built upon core abiotic molecules. Many of these compounds are chromophoric and are now involved in multiple pathways. Plants have further evolved a plethora of chromophoric compounds that can not only act as sunscreens and redox modifiers, but also have now become integrated into a generalised stress adaptive system. This could be an extension of the dissipative process. In animals, many of these compounds are hormetic, modulating mitochondria and calcium signalling. They can also display anti‐pathogen effects. They could therefore modulate bioenergetics across all life due to the conserved electron transport chain and proton gradient. In this review paper, we focus on well‐described medicinal compounds, such as salicylic acid and cannabidiol and suggest, at least in animals, their activity reflects their evolved function in plants in relation to stress adaptation, which itself evolved to maintain dissipative homeostasis.
... Additionally, maternal undernutrition in rats induces impaired placental mitochondrial function and results in fetal and placental growth restriction 33 . Recently, associations between mitochondrial dysfunction and THC exposure have been suggested 34 . THC treatment in human lung cancer cells (H460) was shown to reduce mitochondrial complex I and complex II-III activities, reduce mitochondrial membrane potential, and induce oxidative stress and apoptosis 34 . ...
... Recently, associations between mitochondrial dysfunction and THC exposure have been suggested 34 . THC treatment in human lung cancer cells (H460) was shown to reduce mitochondrial complex I and complex II-III activities, reduce mitochondrial membrane potential, and induce oxidative stress and apoptosis 34 . ...
Article
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Prenatal cannabis use is a significant problem and poses important health risks for the developing fetus. The molecular mechanisms underlying these changes are not fully elucidated but are thought to be attributed to delta-9-tetrahydrocannabinol (THC), the main bioactive constituent of cannabis. It has been reported that THC may target the mitochondria in several tissue types, including placental tissue and trophoblast cell lines, and alter their function. In the present study, in response to 48-h THC treatment of the human extravillous trophoblast cell line HTR8/SVneo, we demonstrate that cell proliferation and invasion are significantly reduced. We further demonstrate THC-treatment elevated levels of cellular reactive oxygen species and markers of lipid damage. This was accompanied by evidence of increased mitochondrial fission. We also observed increased expression of cellular stress markers, HSP70 and HSP60, following exposure to THC. These effects were coincident with reduced mitochondrial respiratory function and a decrease in mitochondrial membrane potential. Taken together, our results suggest that THC can induce mitochondrial dysfunction and reduce trophoblast invasion; outcomes that have been previously linked to poor placentation. We also demonstrate that these changes in HTR8/SVneo biology may be variably mediated by cannabinoid receptors CB1 and CB2.
... CB1 receptor is present at membrane of neuronal mitochondria, and get activated by exogenous cannabinoid or endocannabinoid which decreases cyclic AMP concentration, protein kinase A activity, enzymatic activity and respiration in neuronal mitochondria (Bénard et al., 2012). Thus, activation of mitochondrial CB1 receptor reduces oxidative stress, ROS production and oxidative phosphorylation (Athanasiou et al., 2007). Cannabinoids plays protective role against mitochondrial damage during oxidative stress and also decrease Ca2+ induced cytochrome c release from mitochondria, further preventing neuronal cell from apoptosis (Velez-Pardo et al., 2010;Catanzaro et al., 2009). ...
... Moreover, CB1 receptor are present at membrane of neuronal mitochondria, and get activated by exogenous cannabinoid or endocannabinoid which decreases cyclic AMP concentration, protein kinase A activity, enzymatic activity and respiration in neuronal mitochondria (Bénard et al., 2012). Thus, activation of mitochondrial CB1 receptor reduces oxidative stress, ROS production and oxidative phosphorylation (Athanasiou et al., 2007). Cannabinoids plays protective role against mitochondrial damage during oxidative stress and also decrease Ca2+ induced cytochrome c release from mitochondria, further preventing neuronal cell from apoptosis (Velez-Pardo et al., 2010;Catanzaro et al., 2009). ...
Article
Alzheimer's disease is a neurodegenerative disease characterized by progressive decline of cognitive function in combination with neuronal death. Current approved treatment target single dysregulated pathway instead of multiple mechanism, resulting in lack of efficacy in slowing down disease progression. The proclivity of endocannabinoid system to exert neuroprotective action and mitigate symptoms of neurodegeneration condition has received substantial interest. Growing evidence suggest the endocannabinoids (eCB) system, viz. anadamide (AEA) and arachidonoyl glycerol (2-AG), as potential therapeutic targets with the ability to modify Alzheimer's pathology by targeting the inflammatory, neurodegenerative and cognitive aspects of the disease. In order to modulate endocannabinoid system, number of agents have been reported amongst which are inhibitors of the monoacylglycerol (MAGL) and fatty acid amide hydrolase (FAAH), the enzymes that hydrolyses 2-AG and AEA respectively. However, little is known regarding the exact mechanistic signalling and their effects on pathophysiology and cognitive decline associated with Alzheimer's disease. Both MAGL and FAAH inhibitors possess fascinating properties that may offer a multi-faceted approach for the treatment of Alzheimer's disease such as potential to protect neurons from deleterious effect of amyloid-β, reducing phosphorylation of tau, reducing amyloid-β induced oxidative stress, stimulating neurotrophin to support brain intrinsic repair mechanism etc. Based on empirical evidence, MAGL and FAAH inhibitors might have potential for therapeutic efficacy against cognitive impairment associated with Alzheimer's disease. The aim of this review is to summarize the experimental studies demonstrating the polyvalent properties of MAGL or FAAH inhibitor compounds for the treatment of Alzheimer's disease, and also effect of these on learning and types of memories, which together encourage to study these compounds over other therapeutics targets. Further research in this direction would enhance the molecular mechanisms and development of applicable interventions for the treatment of Alzheimer's disease, which nevertheless stay as the primary unmet need.
... More recently, Δ9-THC has been reported to be utilized as an adjunct treatment for various neurological diseases including PD [4]. Since in PD, mitochondrial impairment plays a crucial role in cell degeneration, the use of phytocannabinoids is not quite obvious, as studies demonstrated mitochondrial toxicity of phytocannabinoids leading to mitochondrial dysfunction and an increase of oxidative stress [7,8]. But in a PD cell culture model, phytocannabinoids, especially Δ9-THC, showed antioxidant properties against MPP + and rotenone. ...
... The authors reported that exposure of human SH-SY5Y cells to 250 μM H 2 O 2 for 24 h led to 50 % cell cytotoxicity [31]. Athanasiou and colleagues [7] could show that production of H 2 O 2 was significantly triggered by μM range concentrations of THC. The same effects were found by Wolff and her colleagues [8]. ...
Article
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Delta-9-Tetrahydrocannabinol and other phytocannabinoids have been previously demonstrated to possess neuroprotective effects in murine mesencephalic cell culture models of Parkinson’s disease, in which increased levels of superoxide radicals led to the loss of dopaminergic neurons. In these models, delta-9-tetrahydrocannabinol did not scavenge these radicals but displayed antioxidative capacity by increasing glutathione levels. Based on these findings, in the present study, we investigated whether the neuroprotective effect of delta-9-tetrahydrocannabinol can also be detected in FeSO4- and H2O2-stressed cells. Mesencephalic cultures were concomitantly treated with FeSO4 (350 μM) or H2O2 (150 μM) and delta-9-tetrahydrocannabinol (0.01, 0.1, 1, 10 μM) on the 12th days in vitro for 48 h. On the 14th DIV, dopaminergic neurons were stained immunocytochemically by tyrosine hydroxylase, and fluorescently using crystal violet, Hoechst 33342, and JC-1. FeSO4 and H2O2 significantly reduced the number of dopaminergic neurons by 33 and 36%, respectively, and adversely affected the morphology of surviving neurons. Moreover, FeSO4, but not H2O2, significantly decreased the fluorescence intensity of crystal violet and Hoechst 33342, and reduced the red/green ratio of JC-1. Co-treatment with delta-9-tetrahydrocannabinol at the concentrations 0.01 and 0.1 μM significantly rescued dopaminergic neurons in FeSO4 and H2O2-treated cultures by 16 and 30%, respectively. delta-9-Tetrahydrocannabinol treatment also led to a higher fluorescence intensity of crystal violet and Hoechst 33342, and increased the red/green fluorescence ratio of JC-1 when concomitantly administered with FeSO4 but not H2O2. To conclude, delta-9-tetrahydrocannabinol rescues dopaminergic neurons against FeSO4- and H2O2-induced neurotoxicity. Using fluorescence dyes, this effect seems to be mediated partially by restoring mitochondrial integrity and decreasing cell death, particularly in FeSO4-treated cultures.
... The major anticancerogenic effects shown in vitro and in vivo include reduction of proliferation, induction of apoptosis and autophagy, inhibition of invasion and angiogenesis, enhancement of tumor immune surveillance, and improved chemosensitivity to anticancer drugs [149,150]. Regarding the induction of apoptosis, cannabinoids have been reported to induce mitochondrial damage and increase ROS production [151][152][153]. However, as in the case of NSAIDs, less is known about the direct effects of cannabinoids on mitochondrial respiration and bioenergetics in cancer cells. ...
... As a consequence of reduced respiration, Δ9-THC decreased cellular ATP levels by 64% compared to control cells (Table 4). A possible mechanism by which Δ9-THC interferes with mitochondrial respiration could be its potential to interfere with mitochondrial respiratory chain complexes, as shown in mitochondria isolated from rat heart and liver [151,152]. However, caution is needed with respect to Δ9-THC. ...
Article
Although we have entered the era of personalized medicine and tailored therapies, drugs that target a large variety of cancers regardless of individual patient differences would be a major advance nonetheless. This review article summarizes current concepts and therapeutic opportunities in the area of targeting aerobic mitochondrial energy metabolism in cancer. Old drugs previously used for diseases other than cancer, such as antibiotics and antidiabetics, have the potential to inhibit the growth of various tumor entities. Many drugs are reported to influence mitochondrial metabolism. However, here we consider only those drugs which predominantly inhibit oxidative phosphorylation.
... The uptake and accumulation of JG-B by actively respiring mitochondria with a negative membrane potential in adherent cells has been employed previously to analyze the cell numbers by some research groups [5,[24][25][26][27], although the assay is time-consuming and requires error prone steps of ethanol-fixation, washing and de-staining preceding spectroscopic analysis of JG-B at 595 nm [5,26]. Moreover, the analysis of cellular uptake of JG-B as reported previously [5,26] could lead to erroneous results because JG-B can nonspecifically bind to nuclei and dead cells as well as non-mitochondrial cellular compartments [28]. ...
... JG-B is well known to biomedical science researchers as it has been extensively used for the staining of mitochondria and assessing its purity, integrity and metabolic activity using microscopybased methods [34][35][36][37][38][39][40][41][42][43]. Of note, JG-B has also been employed in the past to assess the cell numbers by some research groups [5,[24][25][26][27]. However, all these studies have evaluated the cellular uptake of JG-B utilizing the property of JG-B to accumulate in actively respiring mitochondria with a negative membrane potential, and, as such, the assay requires ethanol-fixation, washing and de-staining steps preceding spectroscopic analysis of JG-B at 595 nm [5,26]. ...
Article
Purpose: Evaluation of cell viability and toxicity in adherent culture systems is of critical relevance for a wide range of disciplines of biomedical sciences research, including cancer research, toxicology, pharmacology, cell biology, neurology and nanomedicine. Several well-established cytotoxicity assays are widely used by researchers, including the most-preferred MTT assay. Nevertheless, there are problems associated with them, for example; in terms of the time-factor and solubilization of the formzan crystals before its spectroscopic quantification. In this study, we propose a simple, fast and cost-effective colorimetric assay that is free of these issues. Methods: Our assay was based upon reductive splitting of blue-green colored supravital safranin derivative dye Janus green B (JG-B) to pink colored diethyl safranin by oxidoreductases of the electron transport chain (ETC) of actively respiring mitochondria. Because this conversion can be easily and reliably followed spectroscopically, measure of diethyl safranin formed from extraneously added JG-B provides a proficient indicator of cellular health and viability. Results: Using MCF-7, a breast cancer cell line, we provide a proof of concept for the suggested assay and compare it with the MTT assay. Conclusion: Unlike the MTT assay, our JG-B assay does not require a solubilization/extraction step, and hence follows a much simpler and time-efficient protocol suitable for high-throughput analysis of cell viability in anchorage-dependent cell culture models. Additionally, the JG-B cell viability assay reported here can be suitably applied either independently or in complementation with other assays for the analysis of cellular viability and toxicity in both analytic and therapeutic aspects of research.
... In bulls, both AEA and met-AEA (a non-hydrolyzing analogue of AEA), when used at micromolarconcentrations, induced a persistent decrease in sperm motility (Gervasi et al., 2009) and inhibited heparin-induced sperm capacitation (Schuel et al., 1999). The AEA, at micromolar concentrations, inhibits mitochondrial membrane potential and oxygen consumption in spermatozoa, thus interfering with mitochondrial electron transport (Sarafian et al., 2003;Athanasiou et al., 2007). Such an inhibitory role was, however, not observed when used at nanomolar concentrations. ...
... Animal Reproduction Science 189 (2018) 77-83 on post-thaw spermatozoa was studied. Cannabinoids inhibit oxygen consumption and ATP production by preventing the mitochondrial respiratory chain (Athanasiou et al., 2007) in spermatozoa, which preserves energy and ensures gradual acquisition of sperm fertilizing capacity during ascent through the decreasing AEA concentrations in the female reproductive tract (Rossato et al., 2005). During cryopreservation, if the spermatozoa are active and motile for a long period, the accumulating metabolites induce damage to spermatozoa. ...
Article
Anandamide (AEA), an endocannabinoid, has been shown to reduce capacitation and acrosomal exocytosis in human spermatozoa. Because buffalo spermatozoa are highly susceptible to cryopreservation induced damage, AEA was assessed as to whether it could protect spermatozoa from cryo-damage. Six ejaculates from six Murrah buffalo bulls (total 36 ejaculates) were utilized for the study. Each ejaculate was divided into four aliquots; spermatozoa in Aliquot 1 were extended in Tris-Citrate-Egg Yolk and frozen as per the standard protocol. Spermatozoa in Aliquots 2, 3 and 4 were incubated with AEA at 1 nM, 1 μM and 10 μM, respectively in Tris-Citrate extender for 15 min at 37 °C before cryopreservation. Cryopreserved spermatozoa were thawed at 37 °C for 30 s before assessment of sperm motility, membrane integrity, capacitation, acrosome reaction, mitochondrial membrane potential (MMP) and lipid peroxidation status. The proportion of motile and membrane intact spermatozoa were greater (P < 0.05) with use of 1 μM AEA incorporated group compared with other groups. The proportion of un-capacitated and acrosome intact spermatozoa was greater (P < 0.05) with use of 1 or 10 μM of AEA compared with the other groups. When compared to the control group, use of 1 μM AEA resulted in a greater proportion of spermatozoa with high MMP (P < 0.05). There was no significant difference in the lipid peroxidation status of spermatozoa among any of the four groups. It was inferred that the protective role of AEA during cryopreservation of buffalo spermatozoa was dose dependent and incubation of spermatozoa with AEA at 1 μM concentration prior to cryopreservation reduced cryo-capacitation and improved post-thaw sperm quality in buffalo.
... antagonists not used [193] rats cardiac mitochondria THC, HU-210, AEA THC and HU-210 ↓oxygen consumption and ↓mitochondrial membrane potential ↑mitochondrial hydrogen peroxide production ↓mitochondrial oxygen consumption; antagonists not used [194] Wistar rats cardiac mitochondria THC up to 500 µM ↔ROS production, no mitochondrial swelling ↔membrane potential, no oxidative stress, no lipid peroxidation THC is not directly toxic in isolated cardiac mitochondria, and may even be helpful in reducing mitochondrial toxicity [195] SD rats neonatal ventricular myocytes CB13 ...
... However, the results obtained so far are contradictory. Thus, on the one hand, THC or AEA and HU-210 not only at high (100-120 or 1-20 µM) but even at low (0.1 or 0.2 µM) concentrations led to a decrease in oxygen consumption in bovine [193], rat [194] and mouse [191,192] cardiac tissue or mitochondria. The latter was connected with a lower mitochondrial membrane potential and an enhanced mitochondrial hydrogen peroxide production ( Table 4). ...
Article
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The use of cannabis preparations has steadily increased. Although cannabis was traditionally assumed to only have mild vegetative side effects, it has become evident in recent years that severe cardiovascular complications can occur. Cannabis use has recently even been added to the risk factors for myocardial infarction. This review is dedicated to pathogenetic factors contributing to cannabis-related myocardial infarction. Tachycardia is highly important in this respect, and we provide evidence that activation of CB1 receptors in brain regions important for cardiovascular regulation and of presynaptic CB1 receptors on sympathetic and/or parasympathetic nerve fibers are involved. The prototypical factors for myocardial infarction, i.e., thrombus formation and coronary constriction, have also been considered, but there is little evidence that they play a decisive role. On the other hand, an increase in the formation of carboxyhemoglobin, impaired mitochondrial respiration, cardiotoxic reactions and tachyarrhythmias associated with the increased sympathetic tone are factors possibly intensifying myocardial infarction. A particularly important factor is that cannabis use is frequently accompanied by tobacco smoking. In conclusion, additional research is warranted to decipher the mechanisms involved, since cannabis use is being legalized increasingly and Δ9-tetrahydrocannabinol and its synthetic analogue nabilone are indicated for the treatment of various disease states.
... TRPV1 activation potentially results in increase in mitochondrial activity as well as higher oxygen consumption by sperm cells and thereby promotes acquisition of sperm hypermotility and induction of capacitation, while cannabinoids lower the mitochondrial activity [1,24]. TRPV1 activation inhibits premature acrosome reaction and promotes egg-sperm fusion just before fertilization. ...
Article
In view of the limited information available on functional significance of TRPV1 in regulating sperm functions, present study was undertaken on bull spermatozoa. Sixty four ejaculates were collected from four Hariana bulls and were used for molecular and functional characterisation of TRPV1. Immunoblotting using TRPV1 specific antibody revealed the presence of a single band of 104 kDa corresponding to TRPV1 in Hariana bull spermatozoa. Indirect immuno fluorescence revealed positive immune-reactivity to TRPV1 at acrosomal, pre-acrosomal, post acrosomal and flagellar regions of spermatozoa. Based on the results of pilot study dose-response analysis, doses of anandamide (AEA; 0.3 μM) and capsazepine (Cp; 10 μM) were selected for further studies. Three groups of semen samples (control 100 μL diluted semen having 1 × 10 ⁶ spermatozoa; anandamide (3 μL AEA+97 μL of diluted semen containing 1 × 10 ⁶ spermatozoa and Cp (1 μL Cp+99 μL of diluted semen containing 1 × 10 ⁶ spermatozoa) were used to study the functional involvement of TRPV1 in bull spermatozoa. Blocking of TRPV1 with Cp resulted in significant (P < 0.05) reduction in progressive sperm motility (PSM) as compared to control. With activation of TRPV1 using AEA also, PSM was significantly (P < 0.05) decreased till 1h and thereafter the PSM was sustained to the level as observed in control. However, both during blocking and activation of TRPV1, per cent spermatozoa showing hyperactive motility were significantly (P < 0.05) increased (20–30%) compared to the control. Treatment with both Cp and AEA revealed significant (P < 0.05) increase in B-pattern of spermatozoa in chlortetracycline hydrochloride (CTC) staining indicating induction of capacitation. Inhibition of soluble adenyl cyclase (sAC) with 99 nM KH7and protein kinase A (PKA) with 3 μM P9115 significantly (P < 0.05) decreased PSM both in the presence of Cp and AEA. Blocking as well as activation of TRPV1 showed significant (P < 0.05) reduction in sperm livability, intact membrane, intact acrosome, high mitochondrial transmembrane potential; hence indicating the involvement of TRPV1 in regulation of sperm functions in bulls. From the study-it was concluded that TRPV1 channels are found in bull spermatozoa and mediate number of sperm functions like motility, hypermotility, capacitation and acrosome reaction. Further studies are required to find out the possible relationship between TRPV1 channels and other channels in regulating spermatozoa function and possible mechanisms associated with TRPV1 activation as well as its role in sperm function regulation.
... The receptors represent therapeutic opportunities in the treatment of pain, inflammation, and chemotherapy-induced nausea or vomiting since they cause inhibitory effects in these pathological processes [76]. Aside from that, modulation of the ECS by natural and synthetic ligands may also result in the induction of apoptosis, inhibition of cancer cell invasion, and neoangiogenesis [77][78][79][80][81]. ...
Article
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The endocannabinoid system (ECS) is a multifunctional homeostatic system involved in many physiological and pathological conditions. The ligands of the ECS are the endo­cannabinoids, whose actions are mimicked by exogenous cannabinoids, such as phytocannabinoids and synthetic cannabinoids. Responses to the ligands of the ECS are mediated by numerous receptors like the classical cannabinoid receptors (CB<sub>1</sub> and CB<sub>2</sub>) as well as ECS-related receptors, e.g., G protein-coupled receptors 18 and 55 (GPR18 and GPR55), transient receptor potential ion channels, and nuclear peroxisome proliferator-activated receptors. The ECS regulates almost all levels of female reproduction, starting with oocyte production through to parturition. Dysregulation of the ECS is associated with the development of gynecological disorders from fertility disorders to cancer. Cannabinoids that act at the ECS as specific agonists or antagonists may potentially influence dysregulation and, therefore, represent new therapeutic options for the therapy of gynecological disorders.
... For instance, Beńard et al. (2012) used anti-CB1R antibodies, disclosed the protein localization of CB1R nearly 30 percent of neuronal mitochondria, which when triggered by exogenous/endogenous cannabinoids lowers the respiratory chain complex-I activity and oxygen consumption, probably via cyclic adenosine monophosphate (cAMP) and protein kinase-A (PKA) signaling. These results are supported by Athanasiou et al. (2007) findings, which reveal that all of the partial CB1R agonists including AEA, D-9-THC, and HU-210 markedly reduced oxygen consumption ( Figure 3) and mitochondrial membrane potential. However, care must be taken to interpret these findings due to using commercial anti-CB1R antibodies (Morozov et al., 2013). ...
Article
Alzheimer’s disease (AD) is an irreversible chronic neurodegenerative disorder that occurs when neurons in the brain degenerate and die. Pain frequently arises in older patients with neurodegenerative diseases including AD. However, the presence of pain in older people is usually overlooked with cognitive dysfunctions. Most of the times dementia patients experience moderate to severe pain but the development of severe cognitive dysfunctions tremendously affects their capability to express the presence of pain. Currently, there are no effective treatments against AD that emphasize the necessity for increasing research to develop novel drugs for treating or preventing the disease process. Furthermore, the prospective therapeutic use of cannabinoids in AD has been studied for the past few years. In this regard, targeting the endocannabinoid system has considered as a probable therapeutic strategy to control several associated pathological pathways, such as mitochondrial dysfunction, excitotoxicity, oxidative stress, and neuroinflammation for the management of AD. In this review, we focus on recent studies about the role of cannabinoids for the treatment of pain and related neuropathological changes in AD.
... One possible mechanism for this elevation is that cannabis activates free radicals as measured from the current results as MDA levels in the serum. Also, (Athanasiou et al., 2007) confirmed that THC causes a marked increase in H2O2 production in the mitochondria. ...
Research
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Background and Objective: Addiction with Cannabis represents one of the major spreading phenomena worldwide. The objective of this study is to evaluate the oxidative stress enzyme markers and lipid peroxidation marker in cannabis administered female rats via intraperitoneal injection. MATERIALS AND METHODS: Thirty female rats were randomly allocated into three groups. First group regarded as control, while the animals of the second and third groups were daily injected intraperitoneally with (5 and 10 mg Cannabis/kg b.w.) for seven successive days. Enzymatic activities of alkaline phosphatase (ALP), Xanthine Oxidase (XO), Glucose-6-phosphate dehydrogenase (G6PDH) and lactate dehydrogenase (LDH) in serum were analyzed, and concentration of Malondialdehyde (MDA) was assayed. RESULTS: The activity of ALP, G6PDH and LDH significantly affected, whereas XO not affected. The lipid peroxidation marker; MDA significantly elevated in high dose treatment. CONCLUSION: This finding suggests that intraperitoneal injection of cannabis causes alterations in oxidation process.
... THC stimulated sphingomyelin hydrolysis in glioma cells. These effects were explained by Athanasiou et al. [33] in their study, where they found that the THC causes morphological changes characteristic to apoptosis. In rat mitochondria, it causes significant decreases in oxygen consumption and mitochondrial membrane potential. ...
Article
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Introduction: Cannabis is extensively abused in Egypt; approximately 800 million dollars are paid annually for management. Cannabis neurophysiologic disturbances represent an annoying health issue, thus, efficient safer treatments are needed. The neurosteroid pregnenolone has been recently observed as a potent inhibitor of cannabis on its receptors. Objectives: To capture cannabis leaves induced brain pathological changes, to assess the myelin basic protein as prognostic biomarker and the potential therapeutic use of pregnenolone against these changes. Materials and Methods: This study was carried out on 160 adult albino rats divided equally into: Group I (negative control), group II (pregnenolone group) gavaged pregnenolone orally, group III (cannabis leaves extract) rats gavaged orally once daily 1/10 of the LD50 of cannabis leaves extract (172.9 mg/kg) and group IV (cannabis leaves extract+pregnenolone) gavaged orally same as before. This study was carried for 12 weeks. Results: There was a significant increase in values of serum myelin basic protein in group (III) compared to group (I); upon supplementation of pregnenolone to intoxicated rats, significant improvement of its values was noticed. Microscopic and immunohistochemistry brain tissues examination of cannabis leaves extract group revealed shrunken cell with dense nuclei and vacuolated cytoplasm & negative immunoreactivity of myelin basic protein antibodies in comparison to control group. Pregnenolone administration to intoxicated rats induced cells improvement and increased the immunoreactivity. Conclusion: cannabis leaves induced brain toxicity in rats with administration of prgnenolone protects brain tissues. Key words: Abuse; Brain; Cannabis leaves; Pregnenolone; Toxicity
... THC stimulated sphingomyelin hydrolysis in glioma cells. These effects were explained by Athanasiou et al. [33] in their study, where they found that the THC causes morphological changes characteristic to apoptosis. In rat mitochondria, it causes significant decreases in oxygen consumption and mitochondrial membrane potential. ...
... Although further studies (particularly in skeletal muscle) are warranted to elucidate the underlying biological mechanisms, these results indicate that endocannabinoids and sphingomyelins act in concert to regulate EE and are in line with previous reports of a cellular link between these lipid moieties (20)(21)(22)(23). As seen for sphingolipids, previous studies reported a negative effect of the ECS on mitochondrial respiration (20,45). Reduction of mitochondrial respiratory activity and disruption of mitochondrial integrity attributed to sphingolipid action in skeletal muscle was proposed to involve inhibition of the key positive mitochondrial regulator of mitochondrial respiratory activity protein kinase B (20,46,47). ...
Article
Context Skeletal muscle endocannabinoids and sphingolipids (particularly, sphingomyelins) are inversely associated with sleeping energy expenditure (SLEEP) in humans. The endocannabinoid system may increase sphingolipid synthesis via cannabinoid receptor-1. Objective To investigate in human skeletal muscle whether endocannabinoids are responsible for the effect of sphingomyelins on SLEEP. Design Muscle endocannabinoid (anandamide, AEA; 2-arachidonoylglycerol, 2-AG), endocannabinoid congeners (oleoylethanolamide, OEA; palmitoylethanolamide, PEA) and sphingomyelin content were measured using liquid chromatography/mass spectrometry. SLEEP was assessed in a whole-room indirect calorimeter. Mediation analyses tested whether the inverse associations between sphingomyelins and SLEEP depended on endocannabinoids and endocannabinoid-related OEA and PEA. Setting Inpatient study. Participants Fifty-three overweight Native Americans. Main Outcome Measure SLEEP. Results AEA (r = 0.45, P = 0.001), 2-AG (r = 0.47, P = 0.0004), OEA (r = 0.27, P = 0.05) and PEA (r = 0.53, P < 0.0001) concentrations were associated with the total sphingomyelin content. AEA, OEA, and PEA correlated with specific sphingomyelins (SM18:1/23:0, SM18:1/23:1, and SM18:1/26:1) previously reported to be determinants of SLEEP in Native Americans (all r > 0.31, all P < 0.03). Up to 1/2 of the negative effect of these specific sphingomyelins on SLEEP was accounted by AEA (all P < 0.04), rendering the direct effect by sphingomyelin per se on SLEEP negligible (P > 0.05). Conclusions In skeletal muscle, AEA is responsible for the sphingomyelin effect on SLEEP, indicating that endocannabinoids and sphingomyelins may jointly reduce human whole-body energy metabolism.
... Numerous studies have shown that mitochondrial function is altered in the early stages of AD, with a reduced en-ergy production as well as increased demand and consequent excessive oxidative stress being observed [2]. It has been suggested that the ECBS may prevent ROS production and lipid peroxidation [55] and reduce NO levels by inhibiting the expression of iNOS, an enzyme that is responsible for NO synthesis [51,56,57]. ...
Article
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Background: The pathogenesis of Alzheimer's disease (AD) is somewhat complex and has yet to be fully understood. As the effectiveness of the therapy currently available for AD has proved to be limited, the need for new drugs has become increasingly urgent. The modulation of the endogenous cannabinoid system (ECBS) is one of the potential therapeutic approaches that is attracting a growing amount of interest. The ECBS consists of endogenous compounds and receptors. The receptors CB1 and CB2 have already been well characterized: CB1 receptors, which are abundant in the brain, particularly in the hippocampus, basal ganglia and cerebellum, regulate memory function and cognition. It has been suggested that the activation of CB1 receptors reduces intracellular Ca concentrations, inhibits glutamate release and enhances neurotrophin expression and neurogenesis. CB2 receptors are expressed, though to a lesser extent, in the central nervous system, particularly in microglia and in immune system cells involved in the release of cytokines. CB2 receptors have been shown to be upregulated in neuritic plaque-associated migroglia in the hippocampus and entorhinal cortex of patients, which suggests that these receptors play a role in the inflammatory pathology of AD. The role of the ECBS in AD is supported by cellular and animal models. By contrast, few clinical studies designed to investigate therapies aimed at reducing behaviour disturbances, especially night-time agitation, eating behaviour and aggressiveness, have yielded positive results. In this review, we will describe how the manipulation of the ECBS offers a potential approach to the treatment of AD.
... CB 1 R agonists have also been reported to decrease temperature (25,49), which could potentially be harmful and exacerbate immune dysfunction in sepsis (50,51). Conversely, D9-THC has been reported to decrease oxygen consumption (52,53), suggesting that it might be protective in conditions in which there is low oxygen delivery such as septic shock and ischemiareperfusion injury. We also observed that D9-THC reduces LPSinduced tachycardia in the first hours following injection. ...
Article
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Cannabis sativa and its principal components, Δ9-tetrahydrocannabinol (Δ9-THC) and cannabidiol, are increasingly being used to treat a variety of medical problems, including inflammatory conditions. Although studies suggest that the endocannabinoid system has immunomodulatory properties, there remains a paucity of information on the effects of cannabinoids on immunity and on outcomes of infection and injury. We investigated the effects and mechanism(s) of action of cannabinoid receptor agonists, including Δ9-THC, on inflammation and organ injury in endotoxemic mice. Administration of Δ9-THC caused a dramatic early upregulation of plasma IL-10 levels, reduced plasma IL-6 and CCL-2 levels, led to better clinical status, and attenuated organ injury in endotoxemic mice. The anti-inflammatory effects of Δ9-THC in endotoxemic mice were reversed by a cannabinoid receptor type 1 (CB1R) inverse agonist (SR141716), and by clodronate-induced myeloid-cell depletion, but not by genetic invalidation or blockade of other putative Δ9-THC receptors, including cannabinoid receptor type 2, TRPV1, GPR18, GPR55, and GPR119. Although Δ9-THC administration reduced the activation of several spleen immune cell subsets, the anti-inflammatory effects of Δ9-THC were preserved in splenectomized endotoxemic mice. Finally, using IL-10-GFP reporter mice, we showed that blood monocytic myeloid-derived suppressive cells mediate the Δ9-THC-induced early rise in circulating IL-10. These results indicate that Δ9-THC potently induces IL-10, while reducing proinflammatory cytokines, chemokines, and related organ injury in endotoxemic mice via the activation of CB1R. These data have implications for acute and chronic conditions that are driven by dysregulated inflammation, such as sepsis, and raise the possibility that CB1R-signaling may constitute a novel target for inflammatory disorders.
... In isolated rat liver mitochondria, AEA inhibits oxidative phosphorylation by blocking F 0 /F 1 ATP synthase activity [65]. In addition, endocannabinoid, phytocannabinoid and synthetic CB receptor agonists such as AEA, D9-THC and HU 210 reduce mito-chondrial oxygen consumption in a dose-dependent manner in rat heart mitochondria [66]. This study further demonstrated that CB 1 agonists induce biphasic changes in complex I and/or complex II/III activities. ...
Article
Dysfunction of the endocannabinoid system (ECS) has been identified in metabolic disease. Cannabinoid receptor 1 (CB1) is abundantly expressed in the brain but also expressed in the periphery. Cannabinoid receptor 2 (CB2) is more abundant in the periphery, including the immune cells. In obesity, global antagonism of overexpressed CB1 reduces bodyweight but leads to centrally mediated adverse psychological outcomes. Emerging research in isolated cultured cells or tissues has demonstrated that targeting the endocannabinoid system in the periphery alleviates the pathologies associated with metabolic disease. Further, peripheral specific cannabinoid ligands can reverse aspects of the metabolic phenotype. This Keynote review will focus on current research on the functionality of peripheral modulation of the ECS for the treatment of obesity.
... Through this interaction, AEA mediates cellular death in rat and human neurons [46,47], lymphoma cells (47) and cytotrophoblasts (48) . Ligands of cannabinoid receptors generally induce apoptosis inhibiting mitochondrial activity because they increase mitochondrial hydrogen peroxide production and decrease the consumption of oxygen and the mitochondrial membrane potential (49) . Another important effect of cannabinoid receptor activation is the inhibition of cancer cells invasion. ...
Article
Objectives: The endocannabinoid system (ECS) is made up of an array of endogenous bioactive lipids, their receptors and enzymes for their synthesis and degradation. The main endogenous ligands are unsaturated fatty acid derivatives such as anandamide (AEA) and 2-arachidonoylglycerol (2-AG), but many others are still under study. Endocannabinoids are involved in both physiological and pathological conditions and could play an important role in the regulation of processes which lead to cancer. Methods: With focus on gynaecological cancers, main papers and review articles, up to September 2018, on the role of the ECS, were acquired by PubMed searches using the search terms: ‘cannabinoid’, ‘endocannabinoid’, ‘gynaecology’, ‘cancer’, and ‘malignancy’. Results: The present review showed the involvement of the endocannabinoid system in numerous physiological and pathological conditions of the female genital tract up to the development of gynaecological malignancy as cervical, endometrial and ovarian cancer. Conclusion: The endocannabinoid system has an important role in antitumor actions involving different signalling receptor and receptor-independent pathways. It represents an exciting challenge to researchers for its potential use in diagnosis and treatment of all gynaecological malignancies.
... While it has been demonstrated that THC impairs mitochondrial function in neuronal systems (Athanasiou et al., 2007;Wolff et al., 2015), the consequences of THC on trophoblast mitochondrial dynamics, respiration, and associated stress responses have not been fully explored. Given that mitochondrial dysfunction, oxidative stress, and poor trophoblast outcomes (Walker et al., 2020), and THC exposure during pregnancy leading to poor placentation and restricted intrauterine growth (Natale et al., 2020) has been demonstrated in rats, we hypothesize that THC may directly increase oxidative stress and reduce ATP generation, and alter trophoblast gene expression resulting in attenuated syncytialization. ...
Article
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The psychoactive component in cannabis, delta-9-tetrahydrocannabinol, can restrict fetal growth and development. Delta-9-tetrahydrocannabinol has been shown to negatively impact cellular proliferation and target organelles like the mitochondria resulting in reduced cellular respiration. In the placenta, mitochondrial dysfunction leading to oxidative stress prevents proper placental development and function. A key element of placental development is the proliferation and fusion of cytotrophoblasts to form the syncytium that comprises the materno-fetal interface. The impact of delta-9-tetrahydrocannabinol on this process is not well understood. To elucidate the nature of the mitochondrial dysfunction and its consequences on trophoblast fusion, we treated undifferentiated and differentiated BeWo human trophoblast cells, with 20 µM delta-9-tetrahydrocannabinol for 48 hr. At this concentration, delta-9-tetrahydrocannabinol on BeWo cells reduced the expression of markers involved in syncytialization and mitochondrial dynamics, but had no effect on cell viability. Delta-9-tetrahydrocannabinol significantly attenuated the process of syncytialization and induced oxidative stress responses in BeWo cells. Importantly, delta-9-tetrahydrocannabinol also caused a reduction in the secretion of human chorionic gonadotropin and the production of human placental lactogen and insulin growth factor 2, three hormones known to be important in facilitating fetal growth. Furthermore, we also demonstrate that delta-9-tetrahydrocannabinol attenuated mitochondrial respiration, depleted adenosine triphosphate, and reduced mitochondrial membrane potential. These changes were also associated with an increase in cellular reactive oxygen species, and the expression of stress responsive chaperones, HSP60 and HSP70. These findings have important implications for understanding the role of delta-9-tetrahydrocannabinol-induced mitochondrial injury and the role this might play in compromising human pregnancies.
... Siegmund reported that the CB 2-AG stimulation increases mitochondrial ROS productions and mitochondrial depolarization in hepatic stellate cells. Different other CB receptor agonists (anandamide, phytocannabinoid D-9-tetrahydrocannabinol, and synthetic CB HU 210) have also been reported to decrease mitochondrial membrane potential and increase mitochondrial ROS production (Athanasiou et al., 2007). Interestingly, in a recent report, ROS has been shown to be involved in MK maturation . ...
Article
Endocannabinoids are well‐known regulators of neurotransmission by activating the cannabinoid (CB) receptors. Endocannabinoids are being used extensively for the treatment of various neurological disorders such as Alzheimer's and Parkinson's diseases. Although endocannabinoids are well studied in cell survival, proliferation, and differentiation in various neurological disorders and several cancers, the functional role in the regulation of blood cell development is less examined. In the present study, virodhamine, which is an agonist of CB receptor‐2, was used to examine its effect on megakaryocytic development from a megakaryoblastic cell. We observed that virodhamine increases cell adherence, cell size, and cytoplasmic protrusions. Interestingly, we have also observed large nucleus and increased expression of megakaryocytic marker (CD61), which are the typical hallmarks of megakaryocytic differentiation. Furthermore, the increased expression of CB2 receptor was noticed in virodhamine‐induced megakaryocytic cells. The effect of virodhamine on megakaryocytic differentiation could be mediated through CB2 receptor. Therefore, we have studied virodhamine induced molecular regulation of megakaryocytic differentiation; mitogen‐activated protein kinase (MAPK) activity, mitochondrial function, and reactive oxygen species (ROS) production were majorly affected. The altered mitochondrial functions and ROS production is the crucial event associated with megakaryocytic differentiation and maturation. In the present study, we report that virodhamine induces megakaryocytic differentiation by triggering MAPK signaling and ROS production either through MAPK effects on ROS‐generating enzymes or by the target vanilloid receptor 1‐mediated regulation of mitochondrial function. Virodhamine, an endocannabinoid, induces megakaryocyte differentiation by regulating mitogen‐activated protein kinase activity and function of mitochondria.
... This corresponds with the available studies that express that the activation of the mitochondrial CB1 receptor reduces the oxidative stress, ROS production, and oxidative phosphorylation. 64,65 The eCB also decreases Ca 2+ -induced cytochrome c release from the mitochondria and thus prevents the apoptosis of neuronal cells. 66 Drugs such as JWH-133, a CB-2 agonist, also reduce the derivative of lipid peroxidation and increase SOD levels in transgenic mice. ...
... CB1 is also localized on skeletal muscle mitochondria where it directly controls cellular respiration and ATP production, 25 and its activation is known to impair mitochondrial function and biogenesis. [25][26][27][28][29] This process is in part mediated through the production of ceramides, 29 lipid derivatives associated with the development of lipotoxicity and insulin resistance commonly seen in sarcopenia. 5 Lastly, CB1 overactivation is also associated with skeletal muscle development and function. ...
Article
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Background: Activation of the endocannabinoid system (ECS) is associated with the development of obesity and insulin resistance, and with perturbed skeletal muscle development. Age-related sarcopenia is a progressive and generalized skeletal muscle disorder involving an accelerated loss of muscle mass and function, with changes in skeletal muscle protein homeostasis due to lipid accumulation and anabolic resistance. Hence, both obesity and sarcopenia share a common set of pathophysiological alterations leading to skeletal muscle impairment. The aim of this study was to characterize how sarcopenia impacts the ECS and if these modifications were related to the loss of muscle mass and function associated with aging in rats. Methods: Six-month-old and 24-month-old male rats were used to measure the contractile properties of the plantarflexors (isometric torque-frequency relationship & concentric power-velocity relationship) and to evaluate locomotor activity, motor coordination, and voluntary gait by open field, rotarod, and catwalk tests, respectively. Levels of endocannabinoids (AEA & 2-AG) and endocannabinoid-like molecules (OEA & PEA) were measured by LCF-MS/MS in plasma, skeletal muscle, and adipose tissue, while the expression of genes coding for the ECS were investigated by quantitative reverse transcription PCR (RT-qPCR). Results: Sarcopenia in old rats was exemplified by a 49% decrease in hindlimb muscle mass (P < 0.01), which was associated with severe impairment of isometric torque, power, voluntary locomotor activity, motor coordination, and gait quality. Sarcopenia was associated with (1) increased 2-AG (+32%, P = 0.07) and reduced PEA and OEA levels in the plasma (-25% and -40%, respectively, P < 0.01); (2) an increased content of AEA, PEA, and OEA in subcutaneous adipose tissue (P < 0.01); and (3) a four-fold increase of 2-AG content in the soleus (P < 0.01) and a reduced OEA content in EDL (-80%, P < 0.01). These alterations were associated with profound modifications in the expression of the ECS genes in the adipose tissue and skeletal muscle. Conclusions: Taken together, these findings demonstrate that circulating and peripheral tissue endocannabinoid tone are altered in sarcopenia. They also demonstrate that OEA plasma levels are associated with skeletal muscle function and loss of locomotor activity in rats, suggesting OEA could be used as a circulating biomarker for sarcopenia.
... ATP provides energy essential for flagellar movements of spermatozoa, and a decrease in the energy supply due to mitochondrial injuries can impair sperm motility [67]. The highly lipophilic feature of cannabinoids grants them access to the intracellular compartment, with mitochondria being reported as targets for cannabinoids [69,70]. Rimmerman et al. [71] showed that CBD directly modulates the outer mitochondrial membrane channel in BV-2 microglial cells, as an agonist of the voltage-dependent anion channel 1 (VDAC1), related to the regulation of cell energy. ...
Article
Cannabidiol (CBD) is a natural cannabinoid present in the Cannabis sativa plant, widely prescribed as an anticonvulsant drug, especially for pediatric use. However, its effects on male reproduction are still little investigated. Therefore, the present study assessed the effects of CBD on the spermatogenesis and sperm quality. For this, twenty-one-day-old Swiss mice received CBD for 34 consecutive days by gavage at doses of either 15 or 30 mg/kg. Chronic exposure to CBD decreased the frequency of stages VII-VIII and XII of spermatogenesis and an increase in the frequency of stage IX were noted. Furthermore, the seminiferous epithelium height reduced at stage IX and increased at stage XII in both CBD-treated groups. There was a significant rise of sperm DNA damage, while no genotoxic effects were observed in leukocytes. The activities of superoxide dismutase and catalase decreased, while malondialdehyde levels increased in the sperm of mice treated with a higher dose of CBD. Mice exposed to 30 mg/kg of CBD showed a reduction in the mobile spermatozoa percentage and in curvilinear velocity, while straight line and average path velocity decreased in both treated groups. The number of acrosome-intact spermatozoa declined in the CBD 30 group, and the number of abnormal acrosomes raised in both CBD groups. On the other hand, the weight of reproductive organs, sperm count, and hormone levels were not affected by CBD treatment. These findings show that deregulation of the endocannabinoid system by CBD can reduce sperm quality. The mechanisms responsible may be associated with disorders during spermatogenesis, especially during the final stages of nuclear remodeling and assembly of acrosome. However, changes in mitochondrial function, as well as the reduction on the antioxidant enzyme activities during epididymal transit, at least partly, may also be involved.
... Previously, it has been shown that cannabinoids can stimulate mitochondrial ROS generation in different cells, leading to the impairment of mitochondrial function and cytochrome C release [37][38][39]. In rat decidual cells, CB1 activation led to ROS generation related to increased ceramide synthesis [39]. ...
Article
Full-text available
Endometriosis is characterized by the formation and development of endometrial tissues outside the uterus, based on an imbalance between proliferation and cell death, leading to the uncontrolled growth of ectopic foci. The potential target for the regulation of these processes is the endocannabinoid system, which was found to be involved in the migration, proliferation, and survival of tumor cells. In this paper, we investigated the effect of endocannabinoid-like compounds from the N-acyl dopamine (NADA) family on the viability of stromal cells from ectopic and eutopic endometrium of patients with ovarian endometriosis. N-arachidonoyldopamine, N-docosahexaenoyldopamine, and N-oleoyldopamine have been shown to have a five-times-more-selective cytotoxic effect on endometrioid stromal cells. To study the mechanisms of the toxic effect, inhibitory analysis, measurements of caspase-3/9 activity, reactive oxygen species, and the mitochondrial membrane potential were performed. It was found that NADA induced apoptosis via an intrinsic pathway through the CB1 receptor and downstream serine palmitoyltransferase, NO synthase activation, increased ROS production, and mitochondrial dysfunction. The higher selectivity of NADA for endometriotic stromal cells and the current lack of effective drug treatment can be considered positive factors for further research of these compounds as possible therapeutic agents against endometriosis.
... Histopathological changes can be explained by oxidative stress in tissues causing cell apoptosis. Athanasiou et al. (2007), studied the effect of THC, endocannabinoids and synthetic cannabinoids on mitochondrial function. The study was carried out on in vitro cells. ...
... Direct effects could also be at play during fetal development given that cannabinoids can cross the placenta and ∆9-THC has been demonstrated to inhibit cardiomyocyte growth in isolated rat cardiomyocytes [149]. Moreover, CB1 antagonists prevent Doxorubicin-(a chemotherapy medication)-induced apoptosis in embryonic heart cells [150], while CB1 agonists (AEA, ∆9-THC, and HU-210) have been demonstrated to decrease mitochondria respiration and mitochondrial membrane potential in rat heart mitochondria [151]. Overall, these data suggest that maternal exposure to ∆9-THC leads to early onset of cardiac dysfunction associated with postnatal catch-up growth. ...
Article
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Up to 20% of pregnant women ages 18–24 consume cannabis during pregnancy. Moreover, clinical studies indicate that cannabis consumption during pregnancy leads to fetal growth restriction (FGR), which is associated with an increased risk of obesity, type II diabetes (T2D), and cardiovascular disease in the offspring. This is of great concern considering that the concentration of Δ9- tetrahydrocannabinol (Δ9-THC), a major psychoactive component of cannabis, has doubled over the last decade and can readily cross the placenta and enter fetal circulation, with the potential to negatively impact fetal development via the endocannabinoid (eCB) system. Cannabis exposure in utero could also lead to FGR via placental insufficiency. In this review, we aim to examine current pre-clinical and clinical findings on the direct effects of exposure to cannabis and its constituents on fetal development as well as indirect effects, namely placental insufficiency, on postnatal metabolic diseases.
... Recent studies showed that THC is able to induce mitochondrial toxicity and oxidative stress in brain mitochondria (Wolff et al. 2015;Beiersdorf et al. 2020). Athanasiou et al., demonstrated that THC is able to cause changes in integrated mitochondrial function, directly, in the absence of cannabinoid receptors in H460 cells (a human non-small cell lung cancer line) (Athanasiou et al. 2007). In addition, Valvassori et al., both acute and chronic injection of CBD increased the activity of the mitochondrial complexes (I, II, II-III, and IV) and creatine kinase (CK) in the rat brain (Valvassori et al. 2013). ...
Article
Mitochondria have the main roles in myocardial tissue homeostasis, through providing ATP for the vital enzymes in intermediate metabolism, contractile apparatus and maintaining ion homeostasis. Mitochondria-related cardiotoxicity results from the exposure with illicit drugs have previously reported. These illicit drugs interference with processes of normal mitochondrial homeostasis and lead to mitochondrial dysfunction and mitochondrial-related oxidative stress. Cannabis consumption has been shown to cause ventricular tachycardia, to increase the risk of myocardial infarction (MI) and potentially sudden death. Here, we investigated this hypothesis that delta-9-tetrahydrocannabinol (Delta-9-THC) as a main cannabinoid found in cannabis could directly cause mitochondrial dysfunction. Cardiac mitochondria were isolated with mechanical lysis and differential centrifugation form rat heart. The isolated cardiac mitochondria were treated with different concentrations of THC (1, 5, 10, 50, 100 and 500 µM) for 1 hour at 37 °C. Then succinate dehydrogenase (SDH) activity, mitochondrial swelling, reactive oxygen species (ROS) formation, mitochondrial membrane potential (MMP) collapse and lipid peroxidation were measured in the treated and non-treated isolated cardiac mitochondria. Our observation showed that THC did not cause a deleterious alteration in mitochondrial functions, ROS production, MMP collapse, mitochondrial swelling, oxidative stress and lipid peroxidation in used concentrations (5-100 µM), even in several tests, toxicity showed a decreasing trend. Altogether, the results of the current study showed that THC is not directly toxic in isolated cardiac mitochondria, and even may be helpful in reducing mitochondrial toxicity.
... Additionally, ∆ 9 -THC has been shown to affect mitochondrial function in several tissues, including the placenta [148,149,222,223]. Human trophoblast cells exposed to ∆ 9 -THC have diminished mitochondrial respiration and ATP-coupling due to decreased abundance of mitochondrial chain complex proteins [148], as well as increased mitochondrial fission and decreased mitochondrial membrane potential [149]. ...
Article
Full-text available
Cannabis use during pregnancy has continued to rise, particularly in developed countries, as a result of the trend towards legalization and lack of consistent, evidence-based knowledge on the matter. While there is conflicting data regarding whether cannabis use during pregnancy leads to adverse outcomes such as stillbirth, preterm birth, low birthweight, or increased admission to neonatal intensive care units, investigations into long-term effects on the offspring’s health are limited. Historically, studies have focused on the neurobehavioral effects of prenatal cannabis exposure on the offspring. The effects of cannabis on other physiological aspects of the developing fetus have received less attention. Importantly, our knowledge about cannabinoid signaling in the placenta is also limited. The endocannabinoid system (ECS) is present at early stages of development and represents a potential target for exogenous cannabinoids in utero. The ECS is expressed in a broad range of tissues and influences a spectrum of cellular functions. The aim of this review is to explore the current evidence surrounding the effects of prenatal exposure to cannabinoids and the role of the ECS in the placenta and the developing fetus.
... Histopathological changes can be explained by oxidative stress in tissues causing cell apoptosis. Athanasiou et al. (2007), studied the effect of THC, endocannabinoids and synthetic cannabinoids on mitochondrial function. The study was carried out on in vitro cells. ...
Article
Full-text available
Abstract Background: Synthetic cannabinoid (SCs) substances are intended for drug addiction while they cannot be easily detected on a regular drug screen. The danger of these substances is not only being undetected, but also their health effects are not well studied and cannot be predicted. This is one of the recent major health problems that threaten populations around the world. Aim of the study: This study is an experimental study to detect the toxic effect of acute exposure to a synthetic cannabinoid substance “AB-CHMINACA’ clinically and histopathologically in different organs in adult male albino rats. Material and methods: AB-CHMINACA was tested for dissolution in different solvents to choose the best vehicle. Doses were selected according to "Guidance on dose level selection for regulatory general toxicology studies for pharmaceuticals". Animals were injected intraperitoneal and after 24 hours, animals were sacrificed and the lung, heart, and liver were examined for histopathological changes. Results: AB-CHMINACA dissolves best in organic solvents like ethanol and DMSO. The most suitable vehicle for intraperitoneal injection of animals was ethanol-saline. After injection, animals showed CNS manifestations; depression or excitation followed by depression according to the dose. Histopathological examination of the lung, heart, and liver tissues showed generalized congestion, hemorrhage, inflammatory cell infiltration and degeneration, which increased by increasing the dose. Conclusion: AB-CHMINACA has toxic histopathological effects on the lung, heart, and liver on single-dose exposure even with minimal clinical manifestations. These effects are dose-related. Key words AB-CHMINACA, Synthetic Cannabinoids, Experimental Study, Lung, Heart, Liver
... At the cellular level, involvement of the ECS has been demonstrated in metabolic processes relevant to fetal development. Indeed, mitochondrial and endoplasmic reticulum (ER) stress contribute to gestational complications, such as FGR (60), and 9-THC has been shown to decreases oxygen consumption and membrane potential of rat heart mitochondria, an effect that appears to be independent of cannabinoid receptor activation (61). Similarly, in the brain, 9-THC impedes mitochondrial respiratory rate, both through CB 1 R and nonreceptor-mediated mechanisms (62). ...
Article
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Clinical reports of cannabis use prevalence during pregnancy vary widely from 3% to upwards of 35% in North America; this disparity likely owing to underestimates from self-reporting in many cases. The rise in cannabis use is mirrored by increasing global legalization and the overall perceptions of safety, even during pregnancy. These trends are further compounded by a lack of evidence-based policy and guidelines for prenatal cannabis use, which has led to inconsistent messaging by healthcare providers and medically licensed cannabis dispensaries regarding prenatal cannabis use for treatment of symptoms, such as nausea. Additionally, the use of cannabis to self-medicate depression and anxiety during pregnancy is a growing medical concern. This review aims to summarize recent findings of clinical and preclinical data on neonatal outcomes, as well as long-term physiological and neurodevelopmental outcomes of prenatal cannabis exposure. Although many of the outcomes under investigation have produced mixed results, we consider these data in light of the unique challenges facing cannabis research. In particular, the limited longitudinal clinical studies available have not previously accounted for the exponential increase in (-)-Δ9– tetrahydrocannabinol (Δ9–THC; the psychoactive compound in cannabis) concentrations found in cannabis over the past two decades. Polydrug use and the long-term effects of individual cannabis constituents [Δ9–THC vs. cannabidiol (CBD)] are also understudied, along with sex-dependent outcomes. Despite these limitations, prenatal cannabis exposure has been linked to low birth weight, and emerging evidence suggests that prenatal exposure to Δ9–THC, which crosses the placenta and impacts placental development, may have wide-ranging physiological and neurodevelopmental consequences. The long-term effects of these changes require more rigorous investigation, though early reports suggest Δ9–THC increases the risk of cognitive impairment and neuropsychiatric disease, including psychosis, depression, anxiety, and sleep disorders. In light of the current trends in the perception and use of cannabis during pregnancy, we emphasize the social and medical imperative for more rigorous investigation of the long-term effects of prenatal cannabis exposure.
... Since neurons are permanent cells, they are most susceptible to hypoxic injury as that occurs with cannabis. Cannabis causes significant decrease in oxygen consumption and significant increase in mitochondrial hydrogen peroxide production and so changes in integrated mitochondrial function and inhibitions of mitochondrial respiratory chain (Athanasiou et al., 2007). ...
... Activation of CB1 receptors on the neuronal membrane decreases firing frequency and protects against depletion of energy sources, whereas activation of mitochondrial CB1 receptors decreases mitochondria activity, thus enabling a coupling between firing activity and energy need of the neurons [59]. Moreover, CB1r agonists decrease oxygen consumption, ROS production [60], and oxidative phosphorylation [61], and, under cellular stress, cannabinoids protect mitochondria [62], which together could be essential for the survival of noradrenergic neurons. Cellular stress resistance is largely dependent on cell metabolism and also on mitochondrial function. ...
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Our laboratory and others have previously shown that cannabinoid receptor type-1 (CB1r) activity is neuroprotective and a modulator of brain ageing; a genetic disruption of CB1r signaling accelerates brain ageing, whereas the pharmacological stimulation of CB1r activity had the opposite effect. In this study, we have investigated if the lack of CB1r affects noradrenergic neurons in the locus coeruleus (LC), which are vulnerable to age-related changes; their numbers are reduced in patients with neurodegenerative diseases and probably also in healthy aged individuals. Thus, we compared LC neuronal numbers between cannabinoid 1 receptor knockout (Cnr1-/-) mice and their wild-type littermates. Our results reveal that old Cnr1-/- mice have less noradrenergic neurons compared to their age-matched wild-type controls. This result was also confirmed by the analysis of the density of noradrenergic terminals which proved that Cnr1-/- mice had less compared to the wild-type controls. Additionally, we assessed pro-inflammatory glial activity in the LC. Although the density of microglia in Cnr1-/- mice was enhanced, they did not show enhanced inflammatory profile. We hypothesize that CB1r activity is necessary for the protection of noradrenergic neurons, but its anti-inflammatory effect probably only plays a minor role in it.
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Mitochondria are key organelles providing energy supply and many other vital functions to cells. Shortly after the discovery of plant-derived cannabinoid compounds, some studies indicated their impact onto mitochondrial functions. The later identification of cannabinoid receptors as classical seven-transmembrane G protein-coupled receptors suggested that these mitochondrial effects might be due to unspecific membrane-altering properties of cannabinoids. However, the recent discovery that brain mitochondria contain significant amounts of functional type-1 cannabinoid receptors (CB1) shed new light on cannabinoid physiology and pharmacology. In this chapter, we will summarize historical and recent evidence of the cannabinoid impact on mitochondrial functions in peripheral and central organs of the body.
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CB1 receptors are functionally present within brain mitochondria (mtCB1), although they are usually considered specifically targeted to plasma membrane. Acute activation of mtCB1 alters mitochondrial ATP generation, synaptic transmission, and memory performance. However, the detailed mechanism linking disrupted mitochondrial metabolism and synaptic transmission is still uncharacterized. CB1 receptors are among the most abundant G protein-coupled receptors in the brain and impact on several processes, including fear coping, anxiety, stress, learning, and memory. Mitochondria perform several key physiological processes for neuronal homeostasis, including production of ATP and reactive oxygen species, calcium buffering, metabolism of neurotransmitters, and apoptosis. It is therefore possible that acute activation of mtCB1 impacts on these different mitochondrial functions to modulate synaptic transmission. In reviewing and integrating across the literature in this area, we describe the possible mechanisms involved in the regulation of brain physiology by mtCB1 receptors.
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Cannabinoid receptors have been shown to interact with other receptors, including Tumor Necrosis Factor Receptor Superfamily (TNFRS) members, to induce cancer cell death. When cannabinoids and death-inducing ligands (including TRAIL) are administered together, they have been shown to synergize and demonstrate enhanced antitumor activity in vitro. Certain cannabinoid ligands have been shown to sensitize cancer cells and synergistically interact with members of the TNFRS, thus suggesting that the combination of cannabinoids with death receptor (DR) ligands induces additive or synergistic tumor cell death. This review summarizes recent findings on the interaction of the cannabinoid and DR systems and suggests possible clinical co-application of cannabinoids and DR ligands in the treatment of various malignancies. This article is protected by copyright. All rights reserved.
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Cannabis sativa has long been used for medicinal purposes. To improve safety and efficacy, compounds from C. sativa were purified or synthesized and named under an umbrella group as cannabinoids. Currently, several cannabinoids may be prescribed in Canada for a variety of indications such as nausea and pain. More recently, an increasing number of reports suggest other salutary effects associated with endogenous cannabinoid signaling including cardioprotection. The therapeutic potential of cannabinoids is therefore extended; however, evidence is limited and mechanisms remain unclear. In addition, the use of cannabinoids clinically has been hindered due to pronounced psychoactive side effects. This review provides an overview on the endocannabinoid system, including known physiological roles, and conditions in which cannabinoid receptor signaling has been implicated.
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Cannabis is the most widely used illegal drug during pregnancy, however, the effects of gestational exposure to Cannabis smoke (CS) on the central nervous system development remain uncharacterised. This study investigates the effects of maternal CS inhalation on brain function in the offspring. Pregnant mice were exposed daily to 5 min of CS during gestational days (GD) 5.5–17.5. On GD 18.5 half of the dams were euthanized for foetus removal. The offspring from the remaining dams were euthanized on postnatal days (PND) 20 and 60 for evaluation. Brain volume, cortex cell number, SOX2, histone-H3, parvalbumin, NeuN, and BDNF immunoreactivity were assessed in all groups. In addition, levels of NeuN, CB1 receptor, and BDNF expression were assessed and cortical primary neurons from rats were treated with Cannabis smoke extract (CSE) for assessment of cell viability. We found that male foetuses from the CS exposed group had decreased brain volume, whereas mice at PND 60 from the exposed group presented with increased brain volume. Olfactory bulb and diencephalon volume were found lower in foetuses exposed to CS. Mice at PND 60 from the exposed group had a smaller volume in the thalamus and hypothalamus while the cerebellum presented with a greater volume. Also, there was an increase in cortical BDNF immunoreactivity in CS exposed mice at PND 60. Protein expression analysis showed an increase in pro-BDNF in foetus brains exposed to CS. Mice at PND 60 presented an increase in mature BDNF in the prefrontal cortex (PFC) in the exposed group and a higher CB1 receptor expression in the PFC. Moreover, hippocampal NeuN expression was higher in adult animals from the exposed group. Lastly, treatment of cortical primary neurons with doses of CSE resulted in decreased cell viability. These findings highlight the potential negative neurodevelopmental outcomes induced by gestational CS exposure.
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Toll-like receptors (TLRs) are sensors of pathogen-associated molecules that trigger inflammatory signalling in innate immune cells including macrophages. All TLRs, with the exception of TLR3, promote intracellular signalling via recruitment of the myeloid differentiation factor 88 (MyD88) adaptor, while TLR3 signals via Toll-Interleukin-1 Receptor (TIR)-domain-containing adaptor-inducing interferon (IFN)-β (TRIF) adaptor to induce MyD88-independent signalling. Furthermore, TLR4 can activate both MyD88-dependent and –independent signalling (via TRIF). The study aim was to decipher the impact of the highly purified plant-derived (phyto) cannabinoids Δ⁹-tetrahydrocannabinol (THC) and cannabidiol (CBD), when delivered in isolation and in combination (1:1), on MyD88-dependent and -independent signalling in macrophages. We employed the use of the viral dsRNA mimetic poly(I:C) and endotoxin lipopolysaccharide (LPS), to induce viral TLR3 and bacterial TLR4 signalling in human Tamm-Horsfall protein-1 (THP-1)-derived macrophages, respectively. TLR3/TLR4 stimulation promoted the activation of interferon (IFN) regulatory factor 3 (IRF3) and TLR4 promoted the activation of nuclear factor (NF)-κB signalling, with downstream production of the type I IFN-β, the chemokines CXCL10 and CXCL8, and cytokine TNF-α. THC and CBD (both at 10 μM) attenuated TLR3/4-induced IRF3 activation and induction of CXCL10/IFN-β, while both phytocannabinoids failed to impact TLR4-induced IκB-α degradation and TNF-α/CXCL8 expression. The role of CB1, CB2 and PPARγ receptors in mediating the effect of THC and CBD on MyD88-independent signalling was investigated. TLRs are attractive therapeutic targets given their role in inflammation and initiation of adaptive immunity, and data herein indicate that both CBD and THC preferentially modulate TLR3 and TLR4 signalling via MyD88-independent mechanisms in macrophages. This offers mechanistic insight into the role of phytocannabinoids in modulating cellular inflammation.
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The endocannabinoid system plays an important role in the pathophysiology of various neurological disorders, such as anxiety, depression, neurodegenerative diseases, and schizophrenia; however, little information is available on the coupling of the endocannabinoid system with the monoaminergic systems in the brain. In the present study, we tested four endocannabinoids and two anandamide analogs for inhibition of recombinant human MAO-A and -B (monoamine oxidase). Virodhamine inhibited both MAO-A and -B (IC50 values of 38.70 and 0.71 μM, respectively) with ∼55-fold greater inhibition of MAO-B. Two other endocannabinoids (noladin ether and anandamide) also showed good inhibition of MAO-B with IC50 values of 18.18 and 39.98 μM, respectively. Virodhamine was further evaluated for kinetic characteristics and mechanism of inhibition of human MAO-B. Virodhamine inhibited MAO-B (Ki value of 0.258 ± 0.037 μM) through a mixed mechanism/irreversible binding and showed a time-dependent irreversible mechanism. Treatment of Neuroscreen-1 (NS-1) cells with virodhamine produced significant inhibition of MAO activity. This observation confirms potential uptake of virodhamine by neuronal cells. A molecular modeling study of virodhamine with MAO-B and its cofactor flavin adenine dinucleotide (FAD) predicted virodhamine's terminal -NH2 group to be positioned near the N5 position of FAD, but for docking to MAO-A, virodhamine's terminal -NH2 group was far away (∼6.52 Å) from the N5 position of FAD, and encountered bad contacts with nearby water molecules. This difference could explain virodhamine's higher potency and preference for MAO-B. The binding free energies for the computationally-predicted poses also showed that virodhamine was selective for MAO-B. These findings suggest potential therapeutic applications of virodhamine for the treatment of neurological disorders.
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Aims To evaluate the specific role of the endocannabinoid/CB1R system in modulating mitochondrial dynamics in the metabolically active renal proximal tubular cells (RPTCs). Materials and methods We utilized mitochondrialy‐targeted GFP in live cells (wild‐type and null for the CB1R), and electron microscopy in kidney sections of RPTC‐CB1R‐/‐ mice and their littermate controls. In both in vitro and in vivo conditions, we assessed the ability of CB1R agonism or fatty acid flux to modulate mitochondrial architecture and function. Results Direct stimulation of CB1R resulted in mitochondrial fragmentation in RPTCs. This process was mediated, at least in part, by modulating the phosphorylation levels of the canonical fission protein dynamin‐related protein 1 on both S637 and S616 residues. CB1R‐induced mitochondrial fission was associated with mitochondrial dysfunction, as documented by reduced oxygen consumption and ATP production, increased reactive oxygen species and cellular lactate levels, as well as a decline in mitochondrial biogenesis. Likewise, we documented that exposure of RPTCs to a fatty acid flux induced CB1R‐depended mitochondrial fission, lipotoxicity, and cellular dysfunction. Conclusions CB1R plays a key role in inducing mitochondrial fragmentation in RPTCs, leading to a decline in organelle’s function, and contributing to renal tubular injury associated with lipotoxicity and other metabolic diseases. This article is protected by copyright. All rights reserved.
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While studies have demonstrated that the main psychoactive component of cannabis, Δ9-tetrahydrocannabinol (Δ9-THC)alone induces placental insufficiency and fetal growth restriction, the underlying mechanisms remain elusive. Given that both (i)endoplasmic reticulum (ER)stress in pregnancy and (ii)gestational exposure to Δ9-THC leads to placental deficiency, we hypothesized that Δ9-THC may directly induce placental ER stress, influencing trophoblast gene expression and mitochondrial function. BeWo human trophoblast cells treated with Δ9-THC (3–30 μM)led to a dose-dependent increase in all ER stress markers and CHOP; these effects could be blocked with CB1R/CB2R antagonists. Moreover, expression of ER stress-sensitive genes ERRγ, VEGFA, and FLT-1 were increased by Δ9-THC, and abrogated with the ER stress inhibitor TUDCA. Δ9-THC also diminished mitochondrial respiration and ATP-coupling due to decreased abundance of mitochondrial chain complex proteins. Collectively, these findings indicate that Δ9-THC can directly augment ER stress resulting in aberrant placental gene expression and impaired mitochondrial function.
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Aim: The aim of this review article is to summarize current knowledge about the role of cannabinoids and cannabinoid receptors in tumor disease modulation and to evaluate comprehensively the use of cannabinoids in cancer patients. Method: According to the PRISMA protocol, we have included data from a total of 105 articles. Results: Cannabinoids affect cancer progression by three mechanisms. The most important mechanism is the stimulation of autophagy and affecting the signaling pathways leading to apoptosis. The most important mechanism of this process is the accumulation of ceramide. Cannabinoids also stimulate apoptosis by mechanisms independent of autophagy. Other mechanisms by which cannabinoids affect tumor growth are inhibition of tumor angiogenesis, invasiveness, metastasis, and the modulation of the anti-tumor immune response. Conclusion: In addition to the symptomatic therapy of cancer patients, the antitumor effects of cannabinoids (whether in monotherapy or in combination with other cancer therapies) have promising potential in the treatment of cancer patients. More clinical trials are needed to demonstrate the antitumor effect of cannabinoids (Tab. 1, Fig. 1, Ref. 167).
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Diabetic complications, chiefly seen in long-term situations, are persistently deleterious to a large extent, requiring multi-factorial risk reduction strategies beyond glycemic control. Diabetic cardiomyopathy is one of the most common deleterious diabetic complications, being the leading cause of mortality among diabetic patients. The mechanisms of diabetic cardiomyopathy are multi-factorial, involving increased oxidative stress, accumulation of advanced glycation end products (AGEs), activation of various pro-inflammatory and cell death signaling pathways, and changes in the composition of extracellular matrix with enhanced cardiac fibrosis. The novel lipid signaling system, the endocannabinoid system, has been implicated in the pathogenesis of diabetes and its complications through its two main receptors: Cannabinoid receptor type 1 and cannabinoid receptor type 2, alongside other components. However, the role of the endocannabinoid system in diabetic cardiomyopathy has not been fully investigated. This review aims to elucidate the possible mechanisms through which cannabinoids and the endocannabinoid system could interact with the pathogenesis and the development of diabetic cardiomyopathy. These mechanisms include oxidative/ nitrative stress, inflammation, accumulation of AGEs, cardiac remodeling, and autophagy. A better understanding of the role of cannabinoids and the endocannabinoid system in diabetic cardiomyopathy may provide novel strategies to manipulate such a serious diabetic complication.
Chapter
Cannabis psychoactive and medicinal properties have been known for thousands of years, but it is only recently that we started to understand the cellular mechanisms triggered by phytocannabinoids, such as Δ⁹-tetrahydrocannabinol and cannabidiol. The discovery of the endocannabinoid system, which includes cannabinoid receptors and cannabinoids produced endogenously in animal cells, allowed a better understanding of the impact of these phytocannabinoids on human health. Early findings suggested that these molecules disrupt mitochondrial functions via direct binding to the organelle membranes and subsequent alterations of their structure and functions. The multiple tools generated after the discovery of the cannabinoid receptors in the early 1990s allowed to show that specific cannabinoids can also modulate mitochondrial activity via activation of cannabinoid receptors localized at the plasma membrane or directly within mitochondrial membranes. Considering that mitochondria are involved in the onset and progression of various diseases, it appears crucial to better understand the mechanisms linking cannabinoids and mitochondria. The aim of this chapter is to provide an overview of the multiple mechanisms triggered by cannabinoids that affects mitochondria.
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Exploring molecular mechanisms of aging and determinants of lifespan will help reduce age-related morbidity, thus facilitating healthy brain aging. Recently, it has been demonstrated that nutritional polyphenols, the main constituents of the Mediterranean diet, maintain redox balance and neuroprotection through the activation of hormetic vitagene pathway. Mitochondria play pivotal roles in the mechanisms of cellular aging and lifespan extension, although further studies are required concerning optimal bioenergetic mechanisms promoting aerobic energy production and the underlying detrimental effects of reactive oxygen species (ROS) by-production with the interplayed nutrition and caloric intake modulatory effects. Consistently, ROS acting as sensors of intracellular nutrients and energy state regulate functional mitochondrial state. Interestingly, increasing evidence reports a functional crosstalk between ROS production by mitochondria and longevity pathways modulating lifespan across species thus ensuring healthy aging. Nrf2-dependent pathways of cellular stress response with their target antioxidant vitagenes are emerging as powerful systems capable to preserve redox homeostasis under environmental and metabolic stresses. Vitagenes encode redox longevity genes induced by oxidative damage including heat shock family (Hsp) Hsp32, Hsp70, glutathione, thioredoxin and sirtuin protein systems. During aging process, a gradual decline of the heat shock response occurs and this may prevent repair of protein damage. Therefore, there is a growing interest by scientific community in developing of novel preventive andpharmacological agents capable of inducing stress responses at the minimum dose within the broad frame of hormesis as therapeutic strategy in patients suffering from chronic degenerative diseases. The specialattention of this paper is focusedonpotential neuroprotective mechanisms of nutrition, in particular dietary polyphenols involved in the activation of vitagenes resulting in improved intracellular antioxidant defense systems against ROS damage leading to degeneration and death with considerable impact on brain health and longevity processes.
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–: The endocannabinoid system (ECS) is a key cellular signalling system that has been implicated in the regulation of diverse cellular functions. Importantly, growing evidence suggests that the biological actions of the ECS may, in part, be mediated through its ability to regulate the production and/or release of nitric oxide, a ubiquitous bioactive molecule, which functions as a versatile signalling intermediate. Herein, we review and discuss evidence pertaining to ECS-mediated regulation of nitric oxide production, as well as the involvement of reactive nitrogen species in regulating ECS-induced signal transduction by highlighting emerging work supporting nitrergic modulation of ECS function. Importantly, the studies outlined reveal that interactions between the ECS and nitrergic signalling systems can be both stimulatory and inhibitory in nature, depending on cellular context. Moreover, such crosstalk may act to maintain proper cell function, whereas abnormalities in either system can undermine cellular homoeostasis and contribute to various pathologies associated with their dysregulation. Consequently, future studies targeting these signalling systems may provide new insights into the potential role of the ECS nitric oxide signalling axis in disease development and/or lead to the identification of novel therapeutic targets for the treatment of nitrosative stress-related neurological, cardiovascular, and metabolic disorders.
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Delta9-Tetrahydrocannabinol (delta9-THC), the active ingredient of marihuana was found to be a highly effective inhibitor in vitro of the NADH-oxidase activity of rat brain and heart mitochondria. The degree of inhibition of the enzyme system obtained from rat brain tissue varied with the region from which it was derived as follows, in the presence of 10(-5) M delta9-THC: hypothalamus plus thalamus plus midbrain, 73 +/- 4%; cerebellum, 66 +/- 4%; medulla oblongata plus pons, 63 +/- 6%; cerebral cortex, 50 +/- 8%. The same concentration inhibited rat heart NADH-oxidase activity 69 +/- 9%. Inhibition of NADH-oxidase activity by a corresponding concentration of deoxycorticosterone was significantly less in all tissue preparations tested, ranging from 11% to 26%. The inhibition of delta9-THC appeared to be competitive and near the amytal-sensitive site of the electron transport system. Suggestive evidence was also obtained for a second site of action, above the cytochrome c site.
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The role of endocannabinoid signaling in the response of the brain to injury is tantalizing but not clear. In this study, transient middle cerebral artery occlusion (MCAo) was used to produce ischemia/reperfusion injury. Brain content of N-arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol were determined during MCAo. Whole brain AEA content was significantly increased after 30, 60 and 120 min MCAo compared with sham-operated brain. The increase in AEA was localized to the ischemic hemisphere after 30 min MCAo, but at 60 and 120 min, was also increased in the contralateral hemisphere. 2-Arachidonoylglycerol content was unaffected by MCAo. In a second set of studies, injury was assessed 24 h after 2 h MCAo. Rats administered a single dose (3 mg/kg) of the cannabinoid receptor type 1 (CB1) receptor antagonist SR141716 prior to MCAo exhibited a 50% reduction in infarct volume and a 40% improvement in neurological function compared with vehicle control. A second CB1 receptor antagonist, LY320135 (6 mg/kg), also significantly improved neurological function. The CB1 receptor agonist, WIN 55212-2 (0.1-1 mg/kg) did not affect either infarct volume or neurological score.
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Mitochondria have been suggested to be a potential intracellular target for cancer chemotherapy. In this report, we demonstrate the ability of the tricyclic antidepressant chlorimipramine to kill human glioma cells in vitro by a molecular mechanism resulting in an increase in caspase 3 activity following inhibition of glioma oxygen consumption. Studies with isolated rat mitochondria showed that chlorimipramine specifically inhibited mitochondrial complex III activity, which causes decreased mitochondrial membrane potential as well as mitochondrial swelling and vacuolation. The use of chlorimipramine in human as an effective, non-toxic cancer therapeutic having a strong selectivity between cancer cells and normal cells on the basis of their mitochondrial function is discussed.
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Habitual marijuana smoking is associated with inflammation and atypia of airway epithelium accompanied by symptoms of chronic bronchitis. We hypothesized that Delta(9)-tetrahydrocannabinol (THC), the primary psychoactive component of marijuana, might contribute to these findings by impairing cellular energetics and mitochondrial function. To test this hypothesis, we examined particulate smoke extracts from marijuana cigarettes, tobacco cigarettes, and placebo marijuana (0% THC) cigarettes for their effects on the mitochondrial function of A549 cells in vitro. Only extracts prepared from marijuana cigarettes altered mitochondrial staining by the potentiometric probe JC-1. With the use of a cross-flow, nose-only inhalation system, rats were then exposed for 20 min to whole marijuana smoke and examined for its effects on airway epithelial cells. Inhalation of marijuana smoke produced lung tissue concentrations of THC that were 8-10 times higher than those measured in blood (75 +/- 38 ng/g wet wt tissue vs. 9.2 +/- 2.0 ng/ml), suggesting high local exposure. Intratracheal infusion of JC-1 immediately following marijuana smoke exposure revealed a diffuse decrease in lung cell JC-1 red fluorescence compared with tissue from unexposed or placebo smoke-exposed rats. Exposure to marijuana smoke in vivo also decreased JC-1 red fluorescence (54% decrease, P < 0.01) and ATP levels (75% decrease, P < 0.01) in single-cell preparations of tracheal epithelial cells. These results suggest that inhalation of marijuana smoke has deleterious effects on airway epithelial cell energetics that may contribute to the adverse pulmonary consequences of marijuana smoking.
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The impaired mitochondrial function hypothesis in schizophrenia is based on evidence of altered brain metabolism, morphology, biochemistry and gene expression. Mitochondria have their own genome, which is needed to synthesize some of the subunits of the respiratory chain enzymes. Mitochondrial DNA (mtDNA) is maternally inherited and we observed an excess of maternal transmission of schizophrenia in a set of parent-offspring affected pairs. We therefore hypothesized that mutations in the mtDNA may contribute to the complex genetic basis of schizophrenia. The entire mtDNA of six schizophrenic patients with an apparent maternal transmission of the disease was sequenced and compared to the reference sequence. We have identified 50 variants and among these six have not been previously reported. Three of them were missense variants: MTCO2 7750C>A, MTATP6 8857G>A and MTND4 12096T>A. These were maternally inherited because they were also present in the mtDNA of their respective schizophrenic mothers and none of them were found in 95 control individuals. The MTND4 12096T>A (Leu446His) is a heteroplasmic variant present in five of the six mother-offspring patient pairs that triggers a non-conservative substitution in the ND4 subunit of complex I. Sequence alignment of 110 ND4 peptides from all eukaryotic kingdoms shows that only hydrophobic amino acids are found in this position. Moreover, leucine was conserved or substituted by an isoleucine in all mammalian species. This indicates that the presence of histidine could affect complex I activity in patients with schizophrenia.
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Mammalian tissues express at least two types of cannabinoid receptor, CB1 and CB2, both G protein coupled. CB1 receptors are expressed predominantly at nerve terminals where they mediate inhibition of transmitter release. CB2 receptors are found mainly on immune cells, one of their roles being to modulate cytokine release. Endogenous ligands for these receptors (endocannabinoids) also exist. These are all eicosanoids; prominent examples include arachidonoylethanolamide (anandamide) and 2-arachidonoyl glycerol. These discoveries have led to the development of CB1- and CB2-selective agonists and antagonists and of bioassays for characterizing such ligands. Cannabinoid receptor antagonists include the CB1-selective SR141716A, AM251, AM281 and LY320135, and the CB2-selective SR144528 and AM630. These all behave as inverse agonists, one indication that CB1 and CB2 receptors can exist in a constitutively active state. Neutral cannabinoid receptor antagonists that seem to lack inverse agonist properties have recently also been developed. As well as acting on CB1 and CB2 receptors, there is convincing evidence that anandamide can activate transient receptor potential vanilloid type 1 (TRPV1) receptors. Certain cannabinoids also appear to have non-CB1, non-CB2, non-TRPV1 targets, for example CB2-like receptors that can mediate antinociception and "abnormal-cannabidiol" receptors that mediate vasorelaxation and promote microglial cell migration. There is evidence too for TRPV1-like receptors on glutamatergic neurons, for alpha2-adrenoceptor-like (imidazoline) receptors at sympathetic nerve terminals, for novel G protein-coupled receptors for R-(+)-WIN55212 and anandamide in the brain and spinal cord, for novel receptors for delta9-tetrahydrocannabinol and cannabinol on perivascular sensory nerves and for novel anandamide receptors in the gastro-intestinal tract. The presence of allosteric sites for cannabinoids on various ion channels and non-cannabinoid receptors has also been proposed. In addition, more information is beginning to emerge about the pharmacological actions of the non-psychoactive plant cannabinoid, cannabidiol. These recent advances in cannabinoid pharmacology are all discussed in this review.
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It is now well established that the psychoactive effects of Cannabis sativa are primarily mediated through neuronal CB1 receptors, while its therapeutic immune properties are primarily mediated through CB2 receptors. Two endocannabinoids, arachidonoylethanolamide and 2-arachidonoylglycerol, have been identified, their action on CB1 and CB2 thoroughly characterized, and their production and inactivation elucidated. However, many significant exceptions to these rules exist. Here we review the evidence suggesting that cannabinoids can modulate synaptic transmission, the cardiovascular system, and the immune system through receptors distinct from CB1 and CB2, and that an additional "independent" endocannabinoid signaling system that involves palmitoylethanolamide may exist.
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Early and late effects of alloxan diabetes and insulin treatment on mitochondrial membrane structure and function were evaluated by studying the kinetic properties of mitochondrial membrane marker enzyme FoF1-ATPase and its modulation by membrane lipid/phospholipid composition and membrane fluidity. Under all experimental conditions the enzyme displayed three kinetically distinguishable components. In 1 wk-old diabetic animals the enzyme activity was unchanged; however, K(m) and V(max) of component I increased and K(m) of component II decreased. Insulin treatment resulted in lowering of K(m) and V(max) of components II and Ill. One-mon diabetic state resulted in decreased enzyme activity, whereas insulin treatment caused hyperstimulation. K(m) of components I and II decreased together with decreased V(max) of all the components. Insulin treatment restored the K(m) and V(max) values. In late-stage diabetes the catalytic efficiency of components I and II increased; insulin treatment had drastic adverse effect. Binding pattern of ATP was unchanged under all experimental conditions. Diabetic state resulted in progressive decrease in energy of activation in the low temperature range (E(L)). Insulin treatment lowered the energy of activation in the high temperature range (E(H)) without correcting the E(L) values. The phase transition temperatures increased in diabetic state and were not corrected by insulin treatment. Long-term diabetes lowered the total phospholipid content and elevated the cholesterol content; insulin treatment had partial restorative effect. The membrane fluidity decreased in general in diabetic condition and was not corrected by insulin treatment at late stage. Regression analysis studies suggest that specific phospholipid classes and/or their ratios may play a role in modulation of the enzyme activity.
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Cannabinoids have been shown to possess anti-inflammatory and neuroprotective properties, which were proposed to occur mainly via activation of the G-protein-coupled receptor CB(1) (cannabinoid receptor 1). Recently, certain cannabinoids have been reported to be ligands for members of the nuclear receptor transcription factor superfamily known as PPARs (peroxisome-proliferator-activated receptors). This review summarizes the evidence for cannabinoid activation of PPARs and identifies a new intracellular target for cannabinoids as therapeutic agents for neuroprotective treatment.
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Mitochondria play a pivotal role in the regulation of apoptotic cell death as well as in several cellular metabolic processes, including energy supply.1, 2 The latter is achieved by oxidative phosphorylation of ADP to ATP using the electrochemical proton gradient generated by the stepwise transport of electrons from oxidizable substrates to molecular oxygen mediated by the mitochondrial respiratory chain. One component of the respiratory chain is cytochrome c, which transfers electrons from Complex III to Complex IV. In apoptosis signaling, however, this vital function of cytochrome c is gradually lost.3 Once the outer mitochondrial membrane (OMM) has been permeabilized by proapoptotic members of the Bcl-2 family of proteins, cytochrome c is released from the mitochondrial intermembrane space into the cytosol. Here, it triggers apoptosome formation and the activation of the caspase cascade, which leads to the cleavage of a host of cellular proteins and dismantling of the cell (Figure 1). Within the mitochondria, cytochrome c is bound to the outer surface of the mitochondrial inner membrane (IMM) by its association with cardiolipin, an anionic phospholipid present predominantly in the mitochondria. We and others have previously suggested that the interaction of cytochrome c with cardiolipin critically determines the amount of the hemoprotein that can be released during apoptosis signaling.4, 5 Furthermore, there is emerging evidence that proapoptotic Bcl-2 family proteins might require cardiolipin for permeabilization of the mitochondria during apoptosis.6 However, the precise role of cardiolipin in the release of cytochrome c from mitochondria during apoptosis is still unclear and is the subject of this commentary.
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Mammalian tissues express at least two types of cannabinoid receptor, CB1 and CB2, both G protein coupled. CB1 receptors are expressed predominantly at nerve terminals where they mediate inhibition of transmitter release. CB2 receptors
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The major psychoactive component of marijuana, Δ9-tetrahydrocannabinol (THC), strongly affected rat liver mitochondria in vitro. At concentrations of 15–60 nmoles/mg of mitochondrial protein, THC uncoupled state IV respiration and decreased respiratory control and ADP/O ratios. Energy-linked changes in fluorescence of 8-anilino-1-naphthalene sulfonate were prevented or reversed by THC. THC also produced large amplitude swelling of mitochondria and release of matrix enzymes. These effects were greatly potentiated by Mg2+. Likewise, flocculation of mixed phospholipid micelles by Mg2+ was potentiated greatly by low concentrations of THC. Studies with micelles prepared from purified phospholipids suggest that THC may specifically destabilize the cardiolipin in mixed micelles.
The present study was undertaken in an attempt to establish whether or not some of the many biochemical effects of cannabinoids could be explained by inhibition of energy generation. Of particular interest was the relationship of inhibition of DNA synthesis in L1210 murine leukemia cells by certain cannabinoids to possible effects on respiration. In studies using these cells, delta9-tetrahydrocannabinol (delta9-THC) and delta8-tetrahydrocannabinol (delta8-THC) were inactive when tested both in vitro and in vivo. Since these cannabinoids have previously been shown to be inhibitors of DNA synthesis in L1210 cells, there is no necessary relationship between inhibition of the two processes.
The effect of cannabinoid derivatives on thymidine-3H uptake in L1210 murine leukemia was determined. In experiments at 200 mg/kg 3 hrs after treatment, the order of activity was delta9-tetrahydrocannabinol less than cannabinol less than cannabidiol less than abnormal cannabidiol less than 11-hydroxy-delta9-tetrahydrocannabinol less than delta8-tetrahydrocannabinol. The inhibitory effect of delta8-tetrahydrocannabinol was 99%. When animals were dosed on consecutive days with delta9-tetrahydrocannabinol and killed on the third day, thymidine-3H incorporation was increased while delta8-tetrahydrocannabinol retained its inhibitory activity under the same conditions. Delta-9-tetrahydrocannabinol and delta8-tetrahydrocannabinol inhibited RNA and protein synthesis in a fashion analagous to the inhibition of DNA synthesis.
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affects the configurational integrity of rat liver mitochondria to a degree dependent upon its concentration. When concentration is increased, the mitochondria pass through three different morphological stages, which are visualized by electron microscopy. These stages are correlated with changes in oxygen uptake, ATPase activity and turbidity. Starting with control mitochondria, designated as Stage A, addition of 15 μg per mg protein results in maximum stimulation of oxygen uptake (Stage B). In the presence of 50 μg per mg protein this stimulation vanishes, the inner mitochondrial structure is disrupted and maximum values of swelling and ATPase activity are attained (Stage C). At higher concentrations, up to more than 100 μg per mg protein, both stimulation of ATPase activity and swelling diminish. These effects are proposed to be associated with bonding of the hydrophobic to some protein-phospholipid receptors in the mitochondrial membrane.
Article
Four persons developed marked parkinsonism after using an illicit drug intravenously. Analysis of the substance injected by two of these patients revealed primarily 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP) with trace amounts of 1-methyl-4-phenyl-4-propionoxy-piperidine (MPPP). On the basis of the striking parkinsonian features observed in our patients, and additional pathological data from one previously reported case, it is proposed that this chemical selectively damages cells in the substantia nigra.
Article
The postnatal development of the complexes of the electron transport chain in isolated rat brain mitochondria were investigated. Nonsynaptosomal brain mitochondria were isolated from rats aged 1-60 days, and the activities of mitochondrial complexes I, II-III, IV, V and citrate synthase were measured. There was a significant increase in the activity of complex I from postnatal day 1 to day 21, and in the activities of complex II-III, complex IV and citrate synthase from postnatal day 1 to day 60. In contrast, the activity of complex V increased significantly between postnatal day 1 and day 10 where it attained adult levels. These data are consistent with the increasing demand for mitochondrial ATP production as the brain develops and as aerobic glycolysis becomes the major pathway for energy production.
Article
The effects of 1-methyl-4-phenylpyridinium (MPP+) on the oxygen consumption, ATP production, H2O2 production, and mitochondrial NADH-CoQ1 reductase (complex I) activity of isolated rat brain mitochondria were investigated. Using glutamate and malate as substrates, concentrations of 10-100 microM MPP+ had no effect on state 4 (-ADP) respiration but decreased state 3 (+ADP) respiration and ATP production. Incubating mitochondria with ADP for 30 min after loading with varying concentrations of MPP+ produced a concentration-dependent decrease in H2O2 production. Incubation of mitochondria with ADP for 60 min after loading with 100 microM MPP+ caused no loss of complex I activity after washing of MPP+ from the mitochondrial membranes. These data are consistent with MPP+ initially binding specifically to complex I and inhibiting both the flow of reducing equivalents and the production of H2O2 by the mitochondrial respiratory chain, without irreversibly damaging complex I. However, mitochondria incubated with H2O2 in the presence of Cu2+ ions showed decreased complex I activity. This study provides additional evidence that cellular damage initiated by MPP+ is due primarily to energy depletion caused by specific binding to complex I, any increased damage due to free radical production by mitochondria being a secondary effect.
Article
The postnatal development of the complexes of the electron transport chain in mitochondria isolated from rat brain synaptosomes was investigated. Synaptosomal brain mitochondria were isolated from rats aged 10-60 days, and the activities of mitochondrial complex I, complex II-III, complex IV and complex V were measured. There was a significant increase in the activity of II-III from day 10 to day 15 and complex IV from day 10 to day 21, thereafter the activities of complexes I-III and IV did not change significantly. The activity of complex I did not change significantly during the period 10-60 days post partum. In synaptic mitochondria, complex V activity was higher than in non-synaptic mitochondria, whereas the activity of complex I was lower than in non-synaptic mitochondria. These data show that the complexes of the respiratory chain within synaptic mitochondria have activities different from those of non-synaptic mitochondria and may have major implications for the relative susceptibility of mitochondria in different brain cell types to neurotoxins such as MPP+, hypoxic/ischaemic damage and oxidative stress.
Article
A colorimetric assay using the basic azo dye Janus green has been developed to assess cell numbers in anchorage-dependent cell cultures, with special regard to the enumeration of osteoblastic cells. Therefore, cells are fixed in ethanol and stained with a 0.2% solution of Janus green for 3 min, followed by a destaining step of 1 min in tap water. The addition of diluted hydrochloric acid easily and immediately leads to dye elution from stained cell layers into the acidic supernatant which consequently is transferred into 96-well plates and read on a microplate reader at 595 nm. Working under standardized conditions, Janus green uptake in several cell lines is shown to be linearly correlated with cell numbers over a broad range of cell densities, in MC3T3-E1 cells from about 3% up to more than 300% of confluency. Absolute sensitivity of the assay allows detection of less than 1000 cells/cm(2). In comparison to many other colorimetric assays, the Janus green technique is simple to perform, fast, precise, stable, cheap, and well suited for processing large quantities of samples. Moreover, it is applicable to any culture formate and size, from irregular formed carriers up to 96-multiwell plates.
Article
It is becoming increasingly evident that the mitochondrial genome may play a key role in neurodegenerative diseases. Mitochondrial dysfunction is characteristic of several neurodegenerative disorders, and evidence for mitochondria being a site of damage in neurodegenerative disorders is partially based on decreases in respiratory chain complex activities in Parkinson's disease, Alzheimer's disease, and Huntington's disease. Such defects in respiratory complex activities, possibly associated with oxidant/antioxidant balance perturbation, are thought to underlie defects in energy metabolism and induce cellular degeneration. Efficient functioning of maintenance and repair process seems to be crucial for both survival and physical quality of life. This is accomplished by a complex network of the so-called longevity assurance processes, which are composed of genes termed vitagenes. A promising approach for the identification of critical gerontogenic processes is represented by the hormesis-like positive effect of stress. In the present review, we discuss the role of energy thresholds in brain mitochondria and their implications in neurodegeneration. We then review the evidence for the role of oxidative stress in modulating the effects of mitochondrial DNA mutations on brain age-related disorders and also discuss new approaches for investigating the mechanisms of lifetime survival and longevity.
Article
The central nervous system is one of the main target organs in cyanide toxicity. In this study, primary cultures of chick embryonic neurons were used to characterize sodium cyanide (NaCN)-induced cell death and to investigate the mechanism of NaCN-mediated preconditioning. After treatment of the cells with 1mM NaCN for 1h followed by a NaCN-free incubation period of 23 h, we observed features of apoptosis such as a reduction in nuclear size, chromatin condensation and nuclear fragmentation as evaluated by nuclear staining with Hoechst 33258 and electron microscopy. In addition, NaCN-induced neurotoxicity was reduced by the protein synthesis inhibitor cycloheximide (CHX) suggesting an active type of cell death. Most of the neurons with condensed chromatin and a shrunken nuclei also showed membrane damage at a late stage. Mitochondrial membrane potential as well as the protein levels of Bcl-2 and Bcl-x(L) decreased 15-60 min and 1-3 h after the exposure to NaCN (1mM, 1h), respectively. Preconditioning caused by incubating chick neurons with 100 microM NaCN for 30 min followed by a NaCN-free interval of 24h significantly protected the neurons against subsequent NaCN (1mM, 1h)-induced damage. Preconditioning prevented NaCN-induced decrease in the mitochondrial membrane potential as well as in the protein levels of Bcl-2 and Bcl-x(L) suggesting that preconditioning-induced neuroprotection is mediated by preserving mitochondrial function.
Article
This study presents a new animal model, the Large White Pig, which was tested for studying cannabinoids metabolism. The first step has focused on determination of plasma kinetics after injection of Delta(9)-tetrahydrocannabinol (THC) at different dosages. Seven pigs received THC by intravenous injections (50, 100 or 200 microg/kg). Plasma samples were collected during 48 h. Determination of cannabinoids concentrations were performed by gas chromatography/mass spectrometry. Results showed that plasma kinetics were comparable to those reported in humans. Terminal half-life of elimination was 10.6 h and a volume of distribution of 32 l/kg was calculated. In a second step, this model was used to determine the kinetic profile of cannabinoids distribution in tissues. Eight Large White male pigs received an injection of THC (200 microg/kg). Two pigs were sacrificed 30 min after injection, two others after 2, 6 and 24 h. Different tissues were sampled: liver, kidney, heart, lung, spleen, muscle, fat, bile, blood, vitreous humor and several brain areas. The fastest THC elimination was noted in liver tissue, where it was completely eliminated in 6 h. THC concentrations decreased in brain tissue slower than in blood. The slowest THC elimination was observed for fat tissue, where the molecule was still present at significant concentrations 24 h later. After 30 min, THC concentration in different brain areas was highest in the cerebellum and lowest in the medulla oblongata. THC elimination kinetics noted in kidney, heart, spleen, muscle and lung were comparable with those observed in blood. 11-Hydroxy-THC was only found at high levels in liver. THC-COOH was less than 5 ng/g in most tissues, except in bile, where it increased for 24 h following THC injection. This study confirms, even after a unique administration, the prolonged retention of THC in brain and particularly in fat, which could be at the origin of different phenomena observed for heavy users such as prolonged detection of THC-COOH in urine or cannabis-related flashbacks. Moreover, these results support the interest for this animal model, which could be used in further studies of distribution of cannabinoids in tissues.
Article
Time-lapse photomicroscopy of human H460 lung cancer cells demonstrated of the transient receptor potential V1 (TRPV1) channel agonists, (E)-capsaicin and resiniferatoxin, and the TRPV1 antagonists, capsazepine, and SB366791, were able to bring about morphological changes characteristic of apoptosis and/or necrosis. Immunoblot analysis identified immunoreactivity for the transient receptor potential V1 (TRPV1) channel in rat brain samples, but not in rat heart mitochondria or in H460 cells. In isolated rat heart mitochondria, all four ligands caused concentration-dependent decreases in oxygen consumption and mitochondrial membrane potential. (E)-Capsaicin and capsazepine evoked concentration-dependent increases and decreases, respectively, in mitochondrial hydrogen peroxide production, whilst resiniferatoxin and SB366791 were without significant effect. These data support the hypothesis that (E)-capsaicin, resiniferatoxin, capsazepine, and SB366791 are all mitochondrial inhibitors, able to activate apoptosis and/or necrosis via non-receptor mediated mechanisms, and also support the use of TRPV1 ligands as anti-cancer agents.
  • C I Ragan
  • Nadh-Ubiquinone Oxidoreductase
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Chronic Parkinsonism in humans due to a product of meperidine-analog synthesis
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  • J W Tetrud
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Chronic Parkinsonism in humans due to a product of meperidine-analog synthesis
  • Langston