ACS Chemical Neuroscience最新文献

Neuromelanin (NM) is an iron-rich, insoluble brown or black pigment that exhibits protective properties. However, its accumulation over time may render it a source of free radicals. In Parkinson's disease, dopaminergic neurons with the highest NM levels and increased iron content are preferentially vulnerable to degeneration. Considering NM's iron binding capacity and the critical role of iron in ferroptosis, we aimed to investigate the interplay between neuromelanin and ferroptosis in dopaminergic cells. We prepared two NM pigments: iron-free NM (ifNM) and iron-containing NM (Fe³⁺NM) and, exposed to cells. After verifying NM accumulation, cell viability was assessed in the absence or presence of antioxidants (NAC (1 mM), Trolox (100 μM)) and specific inhibitors of cell death types. Ferroptosis-related parameters, including lipid peroxidation byproducts (4-HNE), lipid ROS, glutathione, intracellular iron, GPX4, and ACSL4, and cellular iron metabolism-related proteins (TfR1, ferroportin, ferritin, IREB2) were evaluated following ifNM and Fe³⁺NM treatments, with or without Ferrostatin-1, Liproxstatin-1 and deferoxamine. Both NMs induced cell death via distinct mechanisms. Ferroptotic cell death by ifNM and Fe³⁺NM was reversed by ferrostatin-1 and NAC (p < 0.05). Significant alterations in lipid peroxidation, GPX4 levels, and iron metabolism were observed independent of NM's iron composition (p < 0.05). Ferritin levels increased following ifNM treatment, reflecting an adaptive response to iron overload, while Fe³⁺NM treatment led to ferritin depletion, possibly via ferritinophagy. Our findings reveal a distinct role of iron-rich and iron-free neuromelanin in modulating ferroptotic pathways, highlighting the potential of targeting neuromelanin-iron interactions as a therapeutic strategy to mitigate neuronal ferroptosis in Parkinson's disease.

Nuclear factor erythroid 2 related factor 2 (Nrf2) is closely associated with neurodegenerative diseases, and the Nrf2-mediated activation of antioxidant response elements (AREs) brings about validated strategies for treating neurodegenerative diseases. Here, we discovered that troglitazone, a clinical medication for diabetes mellitus, could serve as a Nrf2 activator to rescue neuronal damages both in vitro and in vivo. The mechanism of troglitazone action involves binding with kelch-like ECH-associated protein 1 (Keap1) and the activation of Nrf2. This process leads to the migration of Nrf2 to the cell nucleus and transactivates the AREs. Troglitazone exhibits significant alleviation of oxidative stress in PC12 cells induced by hydrogen peroxide or 6-hydroxydopamine (6-OHDA). In vivo studies indicate that troglitazone could rescue the motor activity and neurodevelopmental deficiency in zebrafish induced by 6-OHDA. Additionally, mass spectrometry imaging demonstrates that troglitazone could cross the zebrafish blood-brain barrier, supporting the application of troglitazone in treating neurodegenerative diseases. Overall, this work reveals that the novel Nrf2 activator troglitazone has potential therapeutic value for neurodegeneration and provides a foundation for its repurposing.

Traditional analgesic opioid compounds, which act through μ opioid receptors (MORs), engender a high risk for misuse and dependence. κ opioid receptor (KOR) activation, a potential target for pain treatment, produces antinociception without euphoric side effects but results in dysphoria and aversion. Triazole 1.1 is a KOR agonist biased toward G-protein coupled signaling, potentially promoting antinociception without dysphoria. We tested whether triazole 1.1 could provide antinociception and its effects in combination with morphine. We employed a lactic acid abdominal pain model, which induced acute pain behaviors, decreased basal dopamine levels in the nucleus accumbens (NAc), and increased KOR function. We administered several interventions including triazole 1.1 (30 mg/kg) and morphine (12 or 24 mg/kg), individually and in combination. Triazole 1.1 alone reduced the pain behavioral response and changes to KOR function but did not prevent the reduction in basal dopamine levels. Morphine not only dose-dependently prevented behavioral pain responses but also elevated NAc dopamine and did not prevent the pain-induced increase in KOR function. However, combining low-dose morphine with triazole 1.1 prevents behavioral pain responses, changes to NAc dopamine levels, and changes to KOR function. Therefore, we present triazole 1.1 as a dose-sparing pain treatment to be used in combination with a lower dose of morphine, thus reducing the potential for opioid misuse.

There currently are no medications proven to be effective for the treatment of stimulant-use disorder (SUD). Sigma-receptor (σR) antagonists block many effects of stimulant drugs but not the reinforcing effects assessed with self-administration in rats. However, a recent study suggests that σR antagonism combined with a dopamine (DA) transporter (DAT) blockade selectively attenuates stimulant self-administration. A compound with potential for dual DAT/σR inhibition, CM699, was synthesized and had the necessary ex vivo affinities of 311 and 14.1 nM at DAT and σ₁Rs, respectively. CM699 inhibited DA uptake ex vivo. Antagonist effects at σ₁Rs by CM699 were confirmed with a recently reported pharmacological assay: CM699 increased, whereas the σ₁R agonist, (+)-pentazocine, decreased σ₁R multimers detected in nondenaturing protein gels, and CM699 blocked the effects of (+)-pentazocine. CM699 after intravenous administration (5.0 mg/kg) in rats had an elimination half-life of 4.4 h. In rats, CM699 after intraperitoneal administration blunted the stimulatory effects of cocaine on DA levels in the nucleus accumbens and insurmountably blocked cocaine self-administration, indicating efficacy as a cocaine antagonist in vivo. When given alone, CM699 was not self-administered nor had significant effects on nucleus accumbens DA, suggesting minimal, if any, abuse potential. Further, in a biochemical assay designed to probe the conformation of DAT, (+)-pentazocine potentiated cocaine-induced cysteine accessibility of DAT transmembrane domain 6a, suggesting a shift in the conformational equilibrium of DAT toward outward-facing, whereas CM699 blocked this effect. The results provide preclinical proof of concept for dual DAT/σR inhibition as a novel DAT-conformational approach for the development of medications to treat SUD.

Cathepsin B has been shown to contribute to deficits in traumatic brain injury (TBI), an important risk factor for Alzheimer’s disease (AD). Cathepsin B is elevated in TBI and AD patients, as well as in animal models of these conditions. Knockout of the cathepsin B gene results in amelioration of TBI-induced motor dysfunction and improvement of AD memory deficit in mice. The mechanism of cathepsin B pathogenesis in these brain disorders has been hypothesized to involve its translocation to the cytosol from its normal lysosomal location. This study, therefore, evaluated brain cytosolic cathepsin B activity in the controlled cortical impact (CCI) mouse model of TBI. CCI-TBI resulted in motor deficits demonstrated by the rotarod assay, brain tissue lesions, and disorganization of the hippocampus. Significantly, CCI-TBI increased cytosolic cathepsin B activity in the brain cortex in the ipsilateral brain hemisphere that received the CCI-TBI injury, with a concomitant decrease in the lysosomal fraction. Cathepsin B activity was monitored using the substrate Z-Nle-Lys-Arg-AMC which specifically detects cathepsin B activity but not other cysteine proteases. The normal lysosomal distribution of cathepsin B was observed by its discrete localization in brain cortical cells. CCI-TBI resulted in a more diffuse cellular distribution of cathepsin B consistent with translocation to the cytosol. Further studies utilized the novel neutral pH-selective inhibitor, Z-Arg-Lys-AOMK, that specifically inhibits cathepsin B at neutral pH 7.2 of the cytosol but not at acidic pH 4.6 of lysosomes. Daily administration of Z-Arg-Lys-AOMK (ip), beginning 1 day before CCI-TBI, resulted in the reduction of the increased cytosolic cathepsin B activity induced by CCI-TBI. The inhibitor also reduced cathepsin B activities in homogenates of the brain cortex and hippocampus which were increased by CCI-TBI. Furthermore, the Z-Arg-Lys-AOMK inhibitor resulted in the reduction of motor function deficit resulting from CCI-TBI. These findings demonstrate the activation of cytosolic cathepsin B activity in CCI-TBI mouse brain injury.

Recently, we disclosed VU0467319, an M₁ positive allosteric modulator (PAM) clinical candidate that had successfully completed a phase I single ascending dose clinical trial. Pharmacokinetic assessment revealed that, in humans upon increasing dose, a circulating, inactive metabolite constituted a major portion of the total drug-related area under the curve (AUC). One approach the team employed to reduce inactive metabolite formation in the back-up program was the kinetic isotope effect, replacing the metabolically labile C-H bonds with shorter, more stable C-D bonds. The C-D dipole afforded VU6045422, a more potent M₁ PAM (human EC₅₀ = 192 nM, 80% ACh Max) than its proteocongener VU0467319 (human EC₅₀ = 492 nM, 71% ACh Max), and retained the desired profile of minimal M₁ agonism. Overall, the profile of VU6045422 supported advancement, as did greater in vitro metabolic stability in both microsomes and hepatocytes than did VU0467319. In both rat and dog in vivo, low doses proved to mirror the in vitro profile; however, at higher doses in 14-day exploratory toxicology studies, the amount of the same undesired metabolite derived from VU6045422 was equivalent to that produced from VU0467319. This unexpected IVIVC result, coupled with less than dose-proportional increases in exposure and no improvement in solubility, led to discontinuation of VU0467319/VU6045422 development.

Parkinson’s disease (PD) is one of the most common progressive neurodegenerative pathologies that leads to dopaminergic deficiency and motor manifestations. Alpha-synuclein aggregation is a characteristic hallmark of PD pathogenesis. These aggregates facilitate the formation of Lewy bodies and degeneration. The epidemiological evidence demonstrates a definitive association of diabetes with PD risk. Considering this, many antidiabetic agents such as GLP-1 agonists and DPP-4 inhibitors are being explored as alternative PD therapeutics. This study evaluated the neuroprotective effect of the DPP-4 inhibitor sitagliptin mediated by the PI3K/AKT and Nrf2 pathways in PD models. In silico studies were conducted to determine the binding affinity, stability, and ADMET properties of DPP-4 inhibitors with target proteins. Sitagliptin (15 mg/kg p.o.) was administered in rotenone (30 mg/kg p.o. for 28 days)-induced and MPTP/P (25 mg/kg i.p. MPTP and 100 mg/kg probenecid i.p. twice a week for 5 weeks)-induced PD mouse (C57/BL6) models. Neurobehavioral assessments were carried out throughout the study. Biochemical (GSH, MDA), molecular estimations (AKT, Nrf2, PI3K, GSK-3β, GLP1, CREB, BDNF, NF-κB, alpha-synuclein), histopathological studies, and immunohistochemistry were carried out at the end of the study. The in silico studies demonstrate better binding, stability, and ADMET profile of sitagliptin with both target proteins. Sitagliptin restored cognitive and motor deficits in both rotenone- and MPTP/P-induced mouse models. There was upregulation of PI3K, AKT, Nrf2, CREB, and BDNF levels and downregulation of GSK-3β, NF-κB, and alpha-synuclein levels in both models after treatment with sitagliptin. However, GLP1 levels were not significantly restored, indicating a GLP1-independent mechanism. It also restored histopathological alterations and TH+ neuronal loss induced by rotenone and MPTP/P. These findings demonstrate that sitagliptin exhibits neuroprotective action mediated by upregulation of the PI3K/AKT and Nrf2 pathways in rotenone and MPTP/P mouse models of PD.

Prolonged exposure to corticosteroids (CORTs) triggers depression and anxiety symptoms either endogenously or exogenously via stimulating endoplasmic reticulum stress (ERS). The study assessed the therapeutic implications of hydrogen sulfide (H₂S) versus sertraline (SERT) in alleviating anxiety and depression induced by CORTs through the modulation of ERS and its inflammatory, oxidative, and apoptotic consequences. Rats were subdivided into four groups: control, CORT (20 mg/kg), NaHS (100 μmol/kg), and SERT (10 mg/kg) for 21 days. Behavioral and histological examinations of the cerebral cortex were performed. The levels of CHOP, GADD34, EIF2AK3, GRP78, caspase 3, and miR-146a were analyzed using qRT-PCR. The levels of CORTs, serotonin, BDNF, TNF-α, BCL2, NRF2, and ATF4 were measured using ELISA, whereas those of IL-1β and BAX were measured using immunohistochemical techniques. Total and phosphorylated PERK were assessed via western blotting, whereas GSH and MDA were assessed via a colorimetric assay. In the present study, CORTs upregulated the gene expression of CHOP, GADD34, EIF2AK3, GRP78, and Caspase 3, whereas it downregulated that of miR-146a. The levels of serotonin, BDNF, BCL2, GSH, and NRF2 were decreased, whereas those of ATF4, TNF-α, IL-1β, BAX, and MDA were elevated. On the contrary, NaHS and SERT reversed all the above-mentioned changes. H₂S shows promise in counteracting anxiety and depression symptoms induced by CORTs by targeting ERS cascades, mitigating inflammation, oxidative insults, and apoptosis in the cerebral cortex. H₂S elicits neuroprotective effects by targeting the miR-146a-3p/GRP78/CHOP/PERK/ATF4/GADD34 signaling pathway and regulating apoptotic markers BAX/BCL2 and inflammatory markers TNF-α and/IL-1β. Compared with SERT, H₂S exhibited superior anxiolytic and antidepressive effects.

The protein DJ-1 appears to play a protective role in the development of Parkinson's disease (PD). Here, we show that endogenous neurotoxins of the 1,2,3,4-tetrahydroisoquinoline family (TIQs), formed upon reaction of various aldehydes such as methylglyoxal (MGO) with the neurotransmitter dopamine, act as irreversible inhibitors of the esterase activity of human DJ-1, with IC50 values between 15 and 57 μM. The presence of a catechol function appears to be essential for these inhibitory effects, which may be at the origin of the oxidation of cysteine 106, a crucial residue in the DJ-1 active site, thereby leading to DJ-1 inhibition. We also show that these endogenous neurotoxins inhibit the protective effects of DJ-1 against glycated guanosine diphosphate (GDP) formation and against alpha-synuclein (aSyn) aggregation induced by MGO. In total, the observed inhibition of DJ-1 by these endogenous neurotoxins may contribute to their damaging effects on the nervous system and, should be taken into account in therapeutic strategies for PD and related disorders.