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Abstract: Ropinirole, an agonist of the post-synaptic dopamine D2-receptor, exerts neuroprotective activity. The mechanism is still under discussion. Assuming that this neuroprotection might be associated with inhibition of the apoptotic cascade underlying cell death, we examined a possible effect of ropinirole on the permeability transition pore (mtPTP) in the mitochondrial inner membrane. Using isolated rat liver mitochondria, the effect of ropinirole was studied on Ca2+-triggered large amplitude swelling, membrane depolarization and cytochrome c release. In addition, the effect of ropinirole on oxidation of added, membrane-impermeable NADH was investigated. The results revealed doubtlessly, that ropinirole can inhibit permeability transition. In patch-clamp experiments on mitoplasts, we show directly that ropinirole interacts with the mtPTP. Thus, ropinirole reversibly inhibits the opening of mtPTP with an IC50 of 3.4µM and a Hill coefficient of 1.3. In both systems (i.e. energized mitochondria and mitoplasts) the inhibitory effect on permeability transition was attenuated by increasing concentrations of inorganic phosphate. In addition, we showed with antimycin A-treated mitochondria that ropinirole failed to suppress respiratory chain-linked reactive oxygen species release. In conclusion, our data suggest that the neuroprotective activity of ropinirole is due to the blockade of the Ca2+-triggered permeability transition. [Copyright &y& Elsevier]

Abstract: We previously demonstrated that the progesterone (PROG) metabolite allopregnanolone (AP) is more potent than PROG in the treatment of traumatic brain injury (TBI) and stroke, but the mechanisms for this differential effect are little understood. The mitochondrial permeability transition pore (mtPTP) appears to be a key player in the intrinsic pathway of apoptosis-induced loss of neurons. Its activation is accompanied by the release of cytochrome c (cyt c) from the intermembrane gap and subsequent cell death. We investigated whether mtPTP is implicated in the mechanisms of PROG and AP neuroprotection following traumatic and ischemic brain injury. To assess the neurosteroids'' direct effects on mtPTP activity at the single-channel level, recordings from the inner mitochondrial membrane were obtained by a patch-clamp approach in rat liver mitoplasts. AP but not PROG strongly inhibited mtPTP currents. Interaction of AP with the PTP was further supported by a swelling assay demonstrating that AP inhibited Ca2+-triggered swelling in functionally intact rat liver and brain mitochondria. If AP inhibits the mtPTP, it should prevent the mitochondrial cyt c release seen in stroke and TBI. To test this idea, we subjected one group of rats to cortical contusion injury (CCI) and another to transient middle cerebral artery occlusion (MCAO). AP-treated animals showed substantially decreased cyt c release and AP was more potent than PROG in inhibiting mitochondrial cyt c release at 24 h post-CCI and -MCAO. Our results demonstrate that AP inhibits the mtPTP current. This may help to explain its more potent anti-apoptotic and neuroprotective effects compared to PROG. [Copyright &y& Elsevier]

[Display omitted] Traumatic brain injury (TBI) triggers a bevy of changes including mitochondrial dysfunction, apoptosis, oxidative stress, neurobehavioural impairment, and neuroinflammation, among others. Dantrolene (DNT), a muscle relaxant which inhibits intracellular Ca2+ signaling from the ER, has been repurposed as a potential neuroprotective agent in various neurological diseases. However, there have been limited studies on whether it can mitigate TBI-induced deficits and restore impaired mitochondrial dynamics. This study sought to evaluate whether Dantrolene can potentially provide neuroprotection in an in vivo model of TBI. Male wistar rats subjected to TBI were treated with DNT (10 mg/kg) 1 h and 12 h post surgery. Animals were assessed 24 h post-TBI to evaluate neurobehavioural deficits and cerebral edema. We evaluated the protein expressions of apoptotic, autophagic, and neuroinflammatory markers by immunoblotting, as well as Mitochondrial Membrane Potential (MMP) and Reactive Oxygen Species (ROS) via Flow Cytometry to ascertain the effects of DNT on TBI. We further analysed immunofluorescence staining with Glial Fibrillary Acidic Protein (GFAP) and immunohistochemistry with NF-κβ to investigate neuroinflammation. H&E staining was also performed post-TBI. Our findings revealed DNT administration inhibits mitochondria-mediated apoptotis and reduces heightened oxidative stress. DNT treatment was also found to reverse neurobehavioural impairments and offer neuroprotection by preserving neuronal architechture. We also demonstrated that DNT inhibits neuronal autophagy and alleviates neuroinflammation following TBI by modulating the NF-κβ/Akt signaling pathway. Thus, our results suggest a novel application of DNT in ameliorating the multitude of deficits induced by TBI, thereby conferring neuroprotection. [ABSTRACT FROM AUTHOR]

Ropinirole (ROP) is a dopamine agonist that can cross the blood-brain barrier (BBB), which is crucial for drugs targeting neurological conditions like Alzheimer's disease (AD). The rationale for the current research is to investigate the potential of ROP as an inhibitor of Microtubule affinity regulating kinase 4 (MARK4)-NFκβ in neurodegenerative diseases, specifically AD. The interaction between ROP and MARK4-NFκβ holds significant promise in the realm of drug discovery and therapeutic interventions for diseases like AD. Molecular docking and biophysical characterization demonstrate how ROP effectively hinders MARK4 activity, offering detailed insights into their molecular interactions. The present research also investigates the biological aspect of MARK4 shows promise in treating AD, with neuroinflammation playing a crucial role in the disease's progression. Aβ 42 and ROP were co-administered directly into the cells for the establishment of the AD model. We confirmed that ROP can inhibit the path of MARK4 activity, as evidenced by biophysical characterization, and can enhance the cell viability, lowers the expression of MARK4, decrease the rate of oxidative stress, and attenuate the expression of NFκβ, leading to reduced neuronal apoptosis in an in vitro -induced Aβ model. Overall, this research provides valuable mechanistic insights into the neuroprotective potential of ROP and its ability to target the MARK4-NFκβ pathway. The amyloid hypothesis (AH) suggests that Alzheimer's disease (AD) begins with the breakdown of amyloid precursor protein (APP) by β-secretase (BACE1), forming APP carboxyl-terminal fragments (CTFs). Subsequent processing by γ-secretase produces Aβ, leading to the formation of senile plaques in the extracellular space involving proteoglycans and apolipoproteins. Extracellular Aβ oligomers can trigger caspases via cell surface receptors, while intracellularly, Aβ may activate caspases through endoplasmic reticulum or mitochondrial stress. Immune response dysregulation can harm cells at a molecular level, releasing inflammatory markers and reactive oxygen species, culminating in cell death and plaque development in AD. The NFκβ and MARK4 pathway is highlighted as a key contributor to AD pathogenesis. [Display omitted] • Biophysical/kinetic confirming that ROP has inhibitory potential in the path of MARK4 activity. • The neuroprotective efficacy of ROP against the Aβ 42 model by enhancing cell viability • Lowering the expression of MARK4 can reduce oxidative stress and neuronal apoptosis by attenuating the NF-κB pathway. • The findings suggest that MARK4 inhibition can be a potential therapeutic approach for managing MARK4-associated diseases [ABSTRACT FROM AUTHOR]

• Aβ (1-42) significantly increases AChE activity, induces of ER stress and apoptosis. • Aβ (1-42) treatment causes increase in oxidative stress and neuronal degeneration. • Aβ (1-42) treatment also results in activation of microglial cells. • Salubrinal attenuates Aβ (1-42) -induced ER and oxidative stress and apoptosis. • Salubrinal attenuates Aβ (1-42) -induced neurodegeneration and microglial activation. Amyloid-β (Aβ) accumulation in the brain is a pathological hallmark of Alzheimer's disease (AD). Endoplasmic reticulum (ER) stress has been implicated in aetiology of neurodegenerative disorders. We studied the involvement of ER stress in Aβ-induced neuronal degeneration in rat brain to correlate it with cellular and molecular modifications in Aβ-induced Alzheimer's like neuropathological process. Aβ (1-42) (5 μg) was administered by bilateral intracerebroventricular (icv) injection in the brain of adult male Wistar rats. Acetylcholinesterase (AChE) activity and histological alterations were observed in different brain regions. ER stress-associated proteins- glucose regulated protein-78 (GRP78), eukaryotic translation initiation factor-2α (eIF2α) and growth arrest and DNA damage-inducible protein-153 (GADD153), neuronal marker- microtubule associated protein-2 (MAP-2) and microglial protein- ionized calcium binding adaptor molecule-1 (Iba-1) were measured by western blot. Reduced glutathione (GSH), nitrite level and levels of caspase-12 and caspase-3 were also measured. ER stress inhibitor, salubrinal (1 mg/kg, intraperitoneally, ip) was used to assess the specific role of ER stress. Aβ (1-42) -induced increase in AChE activity, GRP78 and GADD protein levels, dephosphorylation of eIF2-α and caspase-12 and caspase-3 levels and decrease in GSH and MAP-2 levels were attenuated by salubrinal. Increase in Iba-1 protein and nitrite levels after Aβ (1-42) administration were partially attenuated by salubrinal. Aβ (1-42) -induced histological alterations were correlated with findings of ER stress. Results of present study implicate ER stress as a potential molecular mechanism in Aβ-induced Alzheimer's like neuropathology which could serve as surrogate biomarker for study of AD progression and efficacy of therapeutic interventions for AD management. [ABSTRACT FROM AUTHOR]

Oxidative stress-related inflammation and apoptosis are important pathogenic consequences, which result in acute pulmonary toxicity. Bleomycin (BLM) is used to treat various forms of cancers. However, its prolonged administration is associated with major toxicity to respiratory system. We studied the effect of walnut ( Juglans regia) extract in a rat model of BLM-induced pulmonary toxicopathy. We also studied parameters of inflammation, apoptosis and oxidative stress in various groups of animals. Prophylactic treatment of total methanolic extract of walnut at the dose of 150 mg/kg b.w. was given per os to Wistar rats for 14 days prior to BLM exposure. A single intratracheal injection of BLM (10 U/kg b.w.) was administered on the eleventh day of the treatment. There was a marked increase in the hydroxyproline level, lipid peroxidation, nitric oxide production, and in the activities of xanthine oxidase and myeloperoxidase in the lung tissue in BLM-treated animals when compared to control animals. BLM also decreased the activities of antioxidant enzymes such as glutathione reductase and catalase and increased the lung inflammation and apoptosis by upregulating the NF-κB signaling pathway and caspase-3 expression. Treatment with walnut extract attenuated these changes in a significant manner. Walnut extract significantly modulated the lung injury as measured by markers of cellular injury such as lactate dehydrogenase and alkaline phosphatase, total cell count, total protein and reduced glutathione in bronchoalveolar lavage fluid. Histological findings supported the protective effects of walnut extract against BLM-induced lung injury. Walnut which has been shown to have numerous medicinally valuable constituents including ellagic acid showed efficacy in preventing the various toxicopathological effects of BLM in rat lungs. Overall, walnut extract decreases BLM-induced oxidative stress and lung inflammation by modulating the alveolar macrophage inflammatory response in rats and thus protecting them from the pathological effect of BLM. [ABSTRACT FROM AUTHOR]

Ischemic reperfusion (I/R) injury causes a wide array of functional and structure alternations of mitochondria, associated with oxidative stress and increased the severity of injury. Despite the previous evidence for N -acetyl-L-cysteine (NAC) provide neuroprotection after I/R injury, it is unknown to evaluate the effect of NAC on altered mitochondrial autophagy forms an essential axis to impaired mitochondrial quality control in cerebral I/R injury. Male wistar rats subjected to I/R injury were used as transient Middle Cerebral Artery Occlusion (tMCAO) model. After I/R injury, the degree of cerebral tissue injury was detected by infarct volume, H&E staining and behavioral assessment. We also performed mitochondrial reactive oxygen species and mitochondrial membrane potential by flow cytometry and mitochondrial respiratory complexes to evaluate the mitochondrial dysfunction. Finally, we performed the western blotting analysis to measure the apoptotic and autophagic marker. We found that NAC administration significantly ameliorates brain injury, improves neurobehavioral outcome, decreases neuroinflammation and mitochondrial mediated oxidative stress. We evaluated the neuroprotective effect of NAC against neuronal apoptosis by assessing its ability to sustained mitochondrial integrity and function. Further studies revealed that beneficial effects of NAC is through targeting the mitochondrial autophagy via regulating the GSK-3β/Drp1mediated mitochondrial fission and inhibiting the expression of beclin-1 and conversion of LC3, as well as activating the p-Akt pro-survival pathway. Our results suggest that NAC exerts neuroprotective effects to inhibit the altered mitochondrial changes and cell death in I/R injury via regulation of p-GSK-3β mediated Drp-1 translocation to the mitochondria. [ABSTRACT FROM AUTHOR]

Ischemic stroke (IS) is a rapidly increasing global burden and is associated with severe neurological decline and mortality. There is urgent requirement of the efforts, aimed to identify therapeutic strategies that are effective in clinic to promote significant recovery from IS. Studies have shown that mitochondria mediated neuroprotection can be a competent target against ischemic damage. Therefore, we examined whether mitochondrial impairment is regulated by Piperine (PIP), an alkaloid of Piper Longum , which has neuroprotective activity against ischemic brain injury. In this study, transient middle cerebral artery occlusion (tMCAO) surgery was performed on male Wistar rats for 90 min followed by 22.5 h of reperfusion for mimicking the IS condition. This study consisted of three groups: sham, tMCAO and tMCAO + PIP (10 mg/kg b.wt., p.o/day for 15 days), and studied for behavioral tests, infarct volume, brain pathological changes, mitochondrial dysfunction, inflammation alongwith cell survival status. PIP pre-treatment showed reduction in neurological alterations and infarct volume. In addition, PIP pre-treatment suppressed the mitochondrial dysfunction and might have anti-apoptotic potential by preventing Cytochrome c (Cyt c) release from mitochondria to cytoplasm and caspase 3 activation. It also regulates pro-apoptotic, Bax and anti-apoptotic, Bcl-2 proteins accompanied by glial fibrillary acidic protein (GFAP) positive cells in cortex region. Quantitative Reverse transcription-polymerase chain reaction (qRT-PCR) results also showed that PIP reduced the expression of pro-inflammatory protein, interleukin-1 β (IL-1β) and enhanced cell survival by restoring the activity of brain derived neurotrophic factor (BDNF) and its transcription protein, cAMP response element binding protein (CREB). Taken together, PIP reduced the mitochondrial dysfunction, neurological impairment, and enhanced neuronal survival. In conclusion, our findings reinforce PIP as an effective neuroprotective agent and provide important evidence about its role as a potential target to serve as a promising therapy for treatment of IS. Schematic representation of mitochondria mediated mechanisms of neuroprotection against ischemic stroke provided by Piperine. Piperine can inhibit mitochondrial dysfunction by suppressing inflammation and mitochondrial ROS mediated alterations in ETC and reduced mitochondrial membrane permeability (Δѱm) thereby preserving mitochondrial Cyt c level through blockage of mitochondrial permeability transition pore (mPTP). Finally, it might increase anti-apoptotic (Bcl-2) protein and decrease pro-apoptotic (Bax) protein, and alleviated neuronal cell death. Piperine also involved in cell survival processes and ultimately rescued neuronal morphological alterations and neurobehavioral abnormalities. IL-1β (Interleukin-1β), GFAP (Glial Fibrillary Acidic Protein), LPO (Lipid Peroxidation), GSH (Glutathione), Cyt c (Cytochrome c), Electron transport chain (ETC), Reactive oxygen species (ROS). [Display omitted] • Ischemic animals pre-treated with piperine showed reduced neurological alterations and infarct volume. • Piperine improved mitochondrial dysfunction. • Piperine showed anti-apoptotic potential by preventing cytochrome c release from mitochondria to cytoplasm. • PIP reduced neurological impairment and enhanced cell survival. [ABSTRACT FROM AUTHOR]