Pharmacological and toxicological effects of copper and vanadium using in vitro and in vivo models of Parkinson’s Disease

Parkinson’s disease (PD) pathology is characterised by distinct types of cellular defects: notably, oxidative damage and mitochondria dysfunction, leading to selective loss of dopaminergic neurons in the substantia nigra pars compacta. Exposure to heavy metals and some environmental toxicants have been associated for many years with this disease pathogenesis. Raised iron levels have been consistently observed in the nigrostriatal pathway in PD cases. This thesis focused on the effects of an endogenous heavy metal micronutrient (copper - Cu) and an exogenous environmental heavy metal (vanadium - Vd), and explored the interplay with iron (Fe), focusing for the first time on sub-toxic effects of these metals upon neuronal cell oxidative and ER stress, differentiation, calcium signalling, motor activity, oxidative stress and lifespan in an in vitro (Catecholaminergic a-differentiated (CAD) cells) and in vivo (Drosophila melanogaster) model of PD respectively. Undifferentiated CAD cells were more susceptible to vanadium exposure than differentiated cells and this susceptibility was modulated by iron. Both a natural (Aloysia citrodora) and synthetic iron chelator, Deferoxamine (DFO), significantly and efficiently protected against chronic sub-toxic Vd-induced mitochondrial oxidative stress in contrast, iron chelation exacerbated the oxidative stress elicited by Cu. Low dose Cu had no significant effect upon metabolic rate (in both differentiated and differentiating CAD cells) but significantly protected undifferentiated cells, decreased potassium chloride (KCl)-induced depolarisation and positively enhanced the expression of MAP2 in differentiated cells In vivo exposure of Drosophila melanogaster (DM) to sub-toxic doses of Vd had a range of differential biochemical and behavioural effects upon wild-type (WT) and PD Pink-1B9 drosophila fly models. In pink-1 flies, exposure to chronic low dose of vanadium exacerbated the existing motor deficits, reduced survival, increased the production of reactive oxygen species (ROS), as well as T-SH and a reduction in survival. In WT, it caused an enhancement in motor activity (like L-dopa), in parallel with a reduction in brain RONS generation and increased total thiol levels (T-SH), with a resulting lifespan extension. Both Aloysia citrodora L, and DFO significantly protected against the PD-like phenotypes in both models. The results accrued in this thesis favours the case for iron-chelation therapy as a viable option for the symptomatic treatment of PD

Attenuation of Vanadium-Induced Neurotoxicity in Rat Hippocampal Slices (In Vitro) and Mice (In Vivo) by ZA-II-05, a Novel NMDA-Receptor Antagonist

Exposure to heavy metals, such as vanadium, poses an ongoing environmental and health threat, heightening the risk of neurodegenerative disorders. While several compounds have shown promise in mitigating vanadium toxicity, their efficacy is limited. Effective strategies involve targeting specific subunits of the NMDA receptor, a glutamate receptor linked to neurodegenerative conditions. The potential neuroprotective effects of ZA-II-05, an NMDA receptor antagonist, against vanadium-induced neurotoxicity were explored in this study. Organotypic rat hippocampal slices, and live mice, were used as models to comprehensively evaluate the compound’s impact. Targeted in vivo fluorescence analyses of the hippocampal slices using propidium iodide as a marker for cell death was utilized. The in vivo study involved five dams, each with eight pups, which were randomly assigned to five experimental groups (n = 8 pups). After administering treatments intraperitoneally over six months, various brain regions were assessed for neuropathologies using different immunohistochemical markers. High fluorescence intensity was observed in the hippocampal slices treated with vanadium, signifying cell death. Vanadium-exposed mice exhibited demyelination, microgliosis, and neuronal cell loss. Significantly, treatment with ZA-II-05 resulted in reduced cellular death in the rat hippocampal slices and preserved cellular integrity and morphological architecture in different anatomical regions, suggesting its potential in countering vanadium-induced neurotoxicity

The Pathopharmacological Interplay between Vanadium and Iron in Parkinson’s Disease Models

Parkinson’s disease (PD) pathology is characterised by distinct types of cellular defects, notably associated with oxidative damage and mitochondria dysfunction, leading to the selective loss of dopaminergic neurons in the brain’s substantia nigra pars compacta (SNpc). Exposure to some environmental toxicants and heavy metals has been associated with PD pathogenesis. Raised iron levels have also been consistently observed in the nigrostriatal pathway of PD cases. This study explored, for the first time, the effects of an exogenous environmental heavy metal (vanadium) and its interaction with iron, focusing on the subtoxic effects of these metals on PD-like oxidative stress phenotypes in Catecholaminergic a-differentiated (CAD) cells and PTEN-induced kinase 1 (PINK−1)B9Drosophila melanogaster models of PD. We found that undifferentiated CAD cells were more susceptible to vanadium exposure than differentiated cells, and this susceptibility was modulated by iron. In PINK−1 flies, the exposure to chronic low doses of vanadium exacerbated the existing motor deficits, reduced survival, and increased the production of reactive oxygen species (ROS). Both Aloysia citrodora Paláu, a natural iron chelator, and Deferoxamine Mesylate (DFO), a synthetic iron chelator, significantly protected against the PD-like phenotypes in both models. These results favour the case for iron-chelation therapy as a viable option for the symptomatic treatment of PD

Volatile Terpenes and Brain Function: Investigation of the Cognitive and Mood Effects of Mentha × Piperita L. Essential Oil with In Vitro Properties Relevant to Central Nervous System Function

Background: Extracts of several members of the monoterpene-rich Lamiaceae sub-family Nepetoideae, including those from the Salvia (sage), Melissa (Lemon balm) and Rosmarinus (rosemary) genera, evince cognitive and mood effects in humans that are potentially related to their effects on cholinergic and GABAergic neurotransmission. To date, despite promising in vitro properties, the cognitive and mood effects of the closely related Mentha spicata (spearmint) and Mentha piperita (peppermint) remain unexplored. This study therefore assessed the human cognitive/mood effects of the M. spicata/piperita essential oil with the most promising, brain-relevant in vitro properties according to pre-trial in vitro screening. Design: Organic spearmint and peppermint (Mentha spicata/piperita) essential oils were pre-screened for neurotransmitter receptor binding and acetylcholinesterase (AChE) inhibition. In a double-blind, placebo-controlled, balanced cross-over study, 24 participants (mean age 25.2 years) consumed single doses of encapsulated placebo and 50 µL and 100 µL of the most promising essential oil (peppermint with nicotinic/GABAA receptor binding and AChE inhibitory properties, that increased calcium influx in a CAD cell neuronal model). Psychological functioning was assessed with mood scales and a range of standardised, cognitively demanding tasks pre-dose and at 1, 3 and 6 h post-dose. Results: The highest (100 µL) dose of essential oil improved performance on the cognitively demanding Rapid Visual Information Processing task (RVIP) at 1 h and 3 h post-dose and both doses attenuated fatigue and improved performance of the Serial 3 s subtraction task at 3 h post-dose. Conclusion: Peppermint (Mentha piperita) essential oil with high levels of menthol/menthone and characteristic in vitro cholinergic inhibitory, calcium regulatory and GABAA/nicotinic receptor binding properties, beneficially modulated performance on demanding cognitive tasks and attenuated the increase in mental fatigue associated with extended cognitive task performance in healthy adults. Future investigations should consider investigating higher dose