PD is a progressive neurodegenerative movement disorder caused by a number of factors like oxidative stress, neurotransmitters dysbalance, mitochondrial dysfunction, and neuroinflammation. However, current evidence are contrastingly challenging previous opinions and facts towards an active role of oxidative stress, neurotransmitter dysbalance, and neuroinflammation in the progression of motor disturbances, leading to PD. Therefore, to explore these interactions, pure neuroinflammation is driven (LPS induced) animal model of PD was used in the present study (Farooqui et al. 2017). In this study, we have demonstrated that the administration of L-theanine was able to protect dopaminergic neurons against LPS induced neuroinflammatory cytokine release, neurotransmitters imbalance, and mitochondrial dysfunction in the rat brain through analysis of behavioral parameters, biochemical estimation (GSH, LPO, nitrite, catalase, and SOD), neuroinflammatory markers (IL-1β, TNF-α, and IL-6), and neurotransmitters (serotonin, dopamine, noradrenaline, GABA and glutamate) analysis by using HPLC-ECD.
The outcomes of the study revealed the protective effect of L-theanine against LPS induced PD-like symptoms in experimental rats. In order to overcome this, LPS at a dose of 5ug/5ul in PBS was infused stereotaxically into SNpc of rats. This results in an increase in pro-inflammatory cytokine activation, oxidative stress, and neurotransmitter dysregulation. LPS-induced behavioral and motor coordination deficits in rats are similarly likely to be observed in people with Parkinsonism, as indicated by tremor, bradykinesia, and stiffness. (Stigger et al. 2013). Intranigral unilateral infusion of LPS causes the generation of ROS and OH that caused oxidative damage to membrane lipids, leading to the reduction of antioxidant molecules (GSH, catalase, and SOD) in SNpc (Anusha et al. 2017). GSH is an antioxidant enzyme responsible for buffering free radicals through reducing H2O2 and organic peroxides (Lobo et al. 2010). Moreover, LPS is highly lipophilic, directly inhibiting the mitochondrial complex I and IV and increasing oxidative stress.
Additionally, it has been shown in pre-clinical and clinical research investigations that catalase and SOD enzymes activity decreased in the substantia nigra and putamen part of the brain. These enzyme alterations may be directly linked to substantia nigra neuron loss and show Parkinson's like symptoms. In PD, these enzyme activities were decreased in the substantia nigra, caudate, and putamen (Jenner et al. 1996).
In the present study, unilaterally infused LPS within SNpc resulted in the activation of neuroinflammatory markers. Their activation released numerous pro-inflammatory substances, which have been implicated in dopaminergic neuronal death (Machado at al. 2011). The direct and indirect dopaminergic pathways of basal ganglia are involved in movements, whereas an imbalance between these pathways results in uncontrolled involuntary movements resulting from progressive dopaminergic neuron degeneration and subsequent changes in striatal neurotransmitter signalling (Crittenden et al. 2011).
In the present research, intranigral injection of LPS induced nigrostriatal area regression and substantially reduced levels of neurotransmitters such as dopamine, serotonin, GABA, and increased the level of norepinephrine, glutamate in the striatum. Although excitotoxicity does not directly cause this damage, it is a major role in the oxidation of catecholamines and their processing by monoamine oxidase enzymes (Wajner et al. 2004).
NF-κB is a transcription factor that has a role in the regulation of the inflammatory response. The NF-κB is primarily found in the cytoplasm of resting cells, complexed with the inhibitory IκB family members. When inflammatory stimuli such as LPS is present, the IB protein is phosphorylated by IB kinase and dissociated from NF-κB, activating the NF-κB signalling pathway. When activated, NF-κB translocate into the nucleus and interacts with the promoter regions of downstream genes, thus controlling their expression in cells. LPS is a potent inducer of NF-κB in neuronal cell death through intracellular ROS production and microglial cells activation (An et al. 2020).
L-theanine treatment improved both motor dysfunction and behavioral deficits. The motor changes and behavioral impairments were measured using narrow beam walk, rotarod test, open field, and grip strength. After the restored motor performance, L-theanine (50 and 100 mg/kg; po) treated rats showed a significant and dose-dependent improvement in neuromuscular coordination and grip strength. We also observed that L-theanine treatment restored the levels of GSH through scavenging superoxide anion and peroxyl radicals. Lipid peroxidation within the membrane and its excessive level is measured as an indicator of oxidative stress, which results in cellular damage through peroxidation in phospholipids membranes (Paradies et al. 1999).
However, L-theanine administration reduced the LPO level in LPS infused rats. This reduction in LPO revealed the antioxidant potential of L-theanine through scavenging ROS, involving superoxide, hydroxyl radical, and peroxyl radicals. RNS (Reactive Nitrogen Species) consists of non-reactive molecule nitric oxide (NO), a chemical messenger that participated in the pathogenesis of PD. NO in combination with O2−, produced harmful and reactive peroxynitrite oxidant (ONOO−), responsible for producing lipid peroxidation in the biological membrane (Korkmaz et al. 2006). We also found that administration of L-theanine in LPS treated rats show that neuroinflammatory cytokine markers were brought back to normal. This signified the anti-inflammatory potential of L-theanine towards halting the progression of PD. L-theanine low (50 mg/kg; po) and high dose (100 mg/kg; po) starting from the 7th day of (LPS 5ug/5ul) injection to 21st day, dose-dependently restored the level of dopamine, serotonin, GABA and decreased the level of NE, and glutamate in LPS infused rats. Hence, L-theanine maintained neural circuits and controlled movement impairments. To further define the mechanism by which L-theanine showed its therapeutic benefits in LPS-induced PD, we examined L-theanine's effects on NF-κB signaling pathways. Thus, L-theanine may exert its inhibitory impact on intracellular ROS generation via inhibiting NF-κB activation triggered by LPS. In the previously published paper, green tea extracts L-theanine reduced the activation of NF-κB in various cell types, including neuronal cells exposed to various oxidative or inflammatory stimuli (Kim et al. 2009). Therefore, it is possible that the ability of L-theanine to prevent ROS generation could be related to its inhibitory effect on LPS-induced stratum neuronal death through inhibition of NFκB. In the present study, we found that theanine prevents the LPS induction of NF-κB by anti-inflammatory impact and could be implicated in the protective effect against LPS-induced Parkinsonism-like symptoms in rats.
Based on our findings, L-theanine has therapeutic potential and needs further exploration to enhance scientific understanding of its role in treating and managing PD. Pertinently, it can be hypothesized that L-theanine reported beneficial effects towards improving motor functions and could be attributed to its neuroprotective abilities, which can be further associated with its anti-inflammatory, antioxidant activity, and capacity to restored the level of striatal neurotransmitters. L-theanine reduces neuroinflammation; reduced neurodegeneration, neurotransmitter alteration, and neuronal destruction. Moreover, our findings proved that treatment with L-theanine might be therapeutically beneficial in treating PD-like symptoms because the study results showed dose-dependently improvement in defects.