6-OHDA Injection Research Articles

AdipoRon improves mitochondrial homeostasis and protects dopaminergic neurons through activation of the AMPK signaling pathway in the 6-OHDA-lesioned rats

The progressive decline of dopaminergic neurons in Parkinson's disease (PD) has been linked to an imbalance in energy and the failure of mitochondrial function. AMP-activated protein kinase (AMPK), the major intracellular energy sensor, regulates energy balance, and damage to nigral dopaminergic neurons induced by 6-hydroxydopamine (6-OHDA) is exacerbated in the absence of AMPK activity. This study aimed to examine the potential therapeutic advantages of AdipoRon, an AMPK activator, on motor function and mitochondrial homeostasis in a 6-OHDA-induced PD model. Male Wistar rats were subjected to unilateral injection of 6-OHDA (10 μg) into the left medial forebrain bundle at two points, and after 7 days, they were treated with intranasal AdipoRon (0.1, 1, and 10 μg) or Levodopa (10 mg/kg, p. o.) for 21 successive days. Following the last treatment day, motor behavior was evaluated through the Murprogo's test, bar test, beam walking test, and apomorphine-induced rotation test. After euthanasia, the left substantia nigra (SN) was separated for evaluation of ATP, mitochondrial membrane potential (MMP), and protein expressions of AMPK, p-AMPK, and mitochondrial dynamics markers (Mfn-2 and Drp-1). Moreover, the number of tyrosine hydroxylase-positive (TH+) cells was quantified in the left substantia nigra. Intranasal AdipoRon effectively reversed muscle rigidity, akinesia, bradykinesia, and rotation caused by 6-OHDA. Moreover, AdipoRon increased the phospho-AMPK/AMPK ratio, mitigated mitochondrial dysfunction, and improved mitochondrial dynamics in the SN. Furthermore, AdipoRon increased the number of TH+ cells in the SN of PD animals. These findings suggest that AdipoRon could protect dopaminergic neurons by activating the AMPK pathway and improving mitochondrial dysfunction.

Intranasal AdipoRon Mitigated Anxiety and Depression-Like Behaviors in 6-OHDA-Induced Parkinson 's Disease Rat Model: Going Beyond Motor Symptoms.

Depression and anxiety are prevalent neuropsychiatric conditions among patients with Parkinson's disease (PD), which may manifest prior to motor symptoms. As levodopa, a prominent treatment for PD motor symptoms, provides few benefits for mood-related abnormalities, tackling non-motor symptoms is particularly important. AdipoRon (Ad), an adiponectin agonist, has demonstrated neuroprotective effects by suppressing neuroinflammatory responses and activating the AMPK/Sirt-1 signaling pathway. This study looked at the potential advantages and underlying mechanisms of intranasal Ad in a rat model of PD induced by 6-hydroxydopamine (6-OHDA). We found that Ad at doses of 1 and 10µg for 21 days exhibited anxiolytic- and antidepressant effects in the open field (OF) test, elevated plus maze (EPM), sucrose splash test, and forced swimming test in a PD model caused by a unilateral 6-OHDA injection into the medial forebrain bundle (MFB). The Ad also lowered the levels of corticosterone in the blood, decreased inflammasome components (NLRP3, caspase 1, and IL-1β), and increased Sirt-1 protein levels in the prefrontal cortex (PFC) of PD rats. We conclude that Ad ameliorates anxious and depressive-like behaviors in the PD rat model through stimulating the AMPK/Sirt-1 signaling and blocking the NLRP3 inflammasome pathways in the PFC.

Impact of 6-OHDA injection and microtrauma in the rat substantia nigra on local brain amyloid beta protein concentrations in the affected area.

Amyloid beta peptides (Aβ) are key indicators of Alzheimer's disease and are also linked to cognitive decline in Parkinson's disease (PD) and other neurodegenerative disorders. This study explored the accumulation of Aβ in a standard 6-Hydroxydopamine (6-OHDA) model of PD. We unilaterally injected 6-OHDA into the substantia nigra of Wistar rats to induce dopaminergic cell degeneration and death, a characteristic of PD. The goal was to detect Aβ protein in tissues and blood vessels showing inflammation or degeneration from the 6-OHDA injection. Our results showed that 6-OHDA injection produced a statistically significant rise in Aβ concentration at the injection site 60 minutes after injection, which was slightly reduced 24 hours post-injection but still significantly higher than in controls. We also tried Gp120 injection in the same zone but it only produced effects comparable to control needle trauma. The presence of Aβ in tissues and blood vessel walls after injection was confirmed through ELISA tests and was supported by immunohistochemical staining of injection areas. We found that the increased Aβ concentration was visible in and around blood vessels and inside blood vessel walls, and also, to a lesser extent in some cells, most probably neurons, in the area. This research highlights the connection between dopaminergic cell poisoning and the accumulation of Aβ, offering insights into the progression of PD to cognitive disorders and dementia.

Effects of dopaminergic neuron degeneration on osteocyte apoptosis and osteogenic markers in 6-OHDA male rat model of Parkinson's disease

Parkinson's disease (PD) and osteoporosis are prevalent chronic conditions that impact a significant proportion of the aging population. Observational and longitudinal studies consistently demonstrate that individuals with PD face an elevated risk of osteoporosis and reduced bone mineral density compared to control groups. However, there is currently no experimental evidence demonstrating the impact of dopaminergic neuron degeneration on bone metabolism. In the present study, we used a male rat model of PD induced by unilateral injection of 6-hydroxydopamine (6-OHDA) in the left medial forebrain bundle (MFB) to evaluate the effect of dopaminergic neuron lesion on certain parameters of bone metabolism. To confirm the dopaminergic neuron lesion, cylinder and Rotarod tests were applied to rats injected with 6-OHDA or vehicle. Osteocyte density and viability were determined through histology and TUNEL assay. Western Blot and immunohistochemistry analysis were performed to investigate whether dopaminergic degeneration influences the expression of some apoptotic markers (Caspase 3 and Cytochrome C) and some osteogenic markers (ALP, OCN, and RUNX2). Our findings show that the dopaminergic lesion resulting from the injection of 6-OHDA was successfully confirmed through behavioral tests. Furthermore, the degeneration of dopaminergic neurons induced by 6-OHDA leads to apoptosis of osteocytes associated with a significant reduction in the tissue expression of the studied osteogenic markers. Thus, our study provides evidence that 6-OHDA-induced degeneration of dopaminergic neurons leads to osteocyte apoptosis, which may contribute to the development of some signs of osteoporosis.

Insulin-Like Growth Factor Binding Protein 2 Drives Neurodegeneration in Parkinson's Disease: Insights From InVivo and InVitro Studies.

Insulin-like growth factor binding protein 2 (IGFBP2) is implicated in various neurodegenerative diseases. However, its role in Parkinson's disease (PD) is unclear. PD rat model was established by 6-OHDA injection. After 3 weeks, mRNA-seq was conducted. Rats received rIGFBP2 via intra-MFB injection 6 h prior to 6-OHDA infusion, and the effect of IGFBP2 in PD rats was investigated by western blotting, IHC, specific kits, JC-1 staining, and TUNEL analysis. Invitro, PC12 cells were treated with 6-OHDA, and CCK-8, specific kits, Hoechst 33258 staining, Western blotting, and JC-1 staining were performed to assess the IGFBP2's role. mRNA-seq revealed DEGs in PD, with attention to downregulated IGFBP2. rIGFBP2 treatment aggravated neurobehavioral deficits, decreased TH expression, Ψm, ATP level and SOD, GSH-Px activities but increased α-synuclein, ROS, MDA, mitochondrial cytochrome c contents, cell apoptosis in 6-OHDA-lesioned rats, which might be mediated through inactivating IGF-1R/AKT pathway. In 6-OHDA-treated PC12 cells, rIGFBP2 aggravated cell injury, demonstrated by decreased cell viability and increased apoptosis, oxidative stress, and mitochondrial dysfunction. Co-treatment with rIGFBP2 and rIGF-1 partially reversed the effect of rIGFBP2 on cell damage. IGFBP2 exacerbates neurodegeneration in PD through increasing oxidative stress, mitochondrial dysfunction, and apoptosis via inhibiting IGF-1R/AKT pathway.

Study of Nootropic Effect of Improved Nano Formulation Version of Levodopa in a Rat Model of Parkinson’s Disease Induced by 6-Hydroxydopamine.

Background: More studies on Parkinson’s disease (PD) symptoms have demonstrated that the condition is multifaceted. PD is associated with cognitive problems & motor impairments, which lower quality of life. 6-hydroxydopamine is a common rodent Parkinsonism neurotoxin. 6-OHDA may damage neurons by increasing oxidative stress from hydroxyl radical production during autoxidation. 6-OHDA and oxidative stress have been shown to impair memory in rats. Thus, 6-OHDA mimics rat Parkinsonian and cognitive decline. Objective: To construct a stable and easily administered nano-formulation with enhanced bioavailability by adding Mucuna pruriens seed phytoactive Levodopa into Nanocochleates. Materials and Methods: Free L-Dopa, nanoliposomes, and levodopa nanocochleates were tested in 6-OHDA-treated rats with PD on the 14th day following the 6-OHDA injection, open-field, morris-water-maze, and Y-maze tests assessed learning and memory. Results: Oral LDNC may reduce 6-OHDA-induced catecholamine neurotoxicity, according to this study. In this 6-OHDAinduced PD model, the nano-formulation appeared to restore injured striatal cells via antioxidant and DA-enhancing pathways.

Tet1-mediated activation of the Ampk signaling by Trpv1 DNA hydroxymethylation exerts neuroprotective effects in a rat model of Parkinson's disease.

Epigenetic regulation plays a role in Parkinson's disease (PD), and ten-eleven translocation methylcytosine dioxygenase 1 (TET1) catalyzes the first step in DNA demethylation by converting 5-methylcytosine to 5-hydroxymethylcytosine. We investigated whether TET1 binds to the promoter of the transient receptor potential cation channel subfamily V member 1 (TRPV1) and regulates its expression, thereby controlling oxidative stress in PD. TRPV1 was identified as an oxidative stress-associated gene in the GSE20186 dataset including substantia nigra from 14 patients with PD and 14 healthy controls and the Genecards database. Lentiviral vectors were used to manipulate Trpv1 expression in rats, followed by 6-hydroxydopamine hydrochloride (6-OHDA) injection for modeling. Behavioral tests, immunofluorescence, Nissl staining, western blot assays, DHE fluorescent probe, biochemical analysis, and ELISA were conducted to assess oxidative stress and neurotoxicity. Trpv1 expression was significantly reduced in the brain tissues of 6-OHDA-treated Parkinsonian rats. Trpv1 alleviated behavioral dysfunction, oxidative stress, and dopamine neuron loss in rats. TET1 mediated TRPV1 hydroxymethylation to promote its expression, and Trpv1 inhibition reversed the mitigating effect of Tet1 on oxidative stress and behavioral dysfunction in PD. TRPV1 activated the AMPK signaling by promoting AMPK phosphorylation to alleviate neurotoxicity and oxidative stress in SH-SY5Y cells. Tet1-mediated Trpv1 hydroxymethylation modification promotes the Ampk signaling activation, thereby eliciting neuroprotection in 6-OHDA-treated Parkinsonian rats. These findings provide experimental evidence that targeting the TET1/TRPV1 axis may be neuroprotective for PD by acting on the AMPK signaling.

The effect of growth hormone on motor findings and dendrite morphology in an experimental Parkinson's disease model.

Approaches for the induction of neurogenesis and neuronal recovery through several modalities are gaining popularity in Parkinson's disease (PD). Growth hormone (GH) seems to have a role in the reversal of neural function following brain injury as well as in normal brain development and function; therefore, the use of GH may represent a feasible strategy in the management of PD. This experimental study aimed to evaluate the effect of growth hormone on motor function and dendrite morphology in rats with 6-hydroxydopamine (6-OHDA)-induced PD model. Thirty-six Sprague Dawley rats were included and randomly allocated into one of the six study groups: two controls and four treatment groups that received daily subcutaneous growth hormone injections for 21days, 1, 2, and 3months. PD model was induced through unilateral 6-OHDA injection to the nigrostriatal pathway. The following assessments were made: apomorphine rotation test, stepping test, and tissue examinations for tyrosine hydroxylase and dendrite morphology. The apomorphine rotation test and the stepping test confirmed the presence of PD. These tests as well as dendritic spine density/number and length assessments showed improvement in PD findings over time with GH administration. Findings of this study suggest that GH administration may improve dendrite morphology and motor function in the PD model, which may translate into symptom relief and quality of life improvement in patients with PD. Such potential benefits should be tested in robust clinical studies.

Modelling of the early stage of retinal neurodegeneration via a topical injection of dopaminergic neurotoxin

Purpose: to develop a model of the early stage of retinal neurodegeneration via an intravitreal injection of the dopaminergic neurotoxin. Material and methods. The experiment was carried out on 20 Chinchilla rabbits. The experimental groups received intravitreal injection of 0.1 ml of 6-hydroxydopamine (6-OHDA) in NaCl 0.9 % with 0.5 % ascorbate, containing 0.25 or 0.5 mg 6-OHDA. The control group received injections of 0.1 ml of NaCl-ascorbate solution without neurotoxin. On the 7th and the 14th days after the injection, intraocular pressure (IOP), pupillary light reaction and ocular blood flow were estimated and the fundus was examined. On the 7th day after an injection of 0.25 mg 6-OHDA, we used ELISA to measure dopamine concentration in retinal homogenates, while on the 14th day after an injection of 0.5 mg 6-OHDA, the concentrations of dopamine, norepinephrine, TNF-a and endothelin-1 were measured by the same method. Results. On the 7th day after a 0,25 mg injection of neurotoxine, we revealed a decrease of dopamine in the retina (0.043 ± 0.130 pg/mg protein compared with 0.10 ± 0,03 pg/mg protein in the controls, р < 0.01). On the 14th day, dopamine and norepinephrine levels showed a fourfold increase (р < 0.05). An injection of 0.5 mg of 6-OHDA caused a threefold increase of dopamine (р < 0.05) and a fivefold increase of norepinephrine (р < 0.01) concentration in comparison with the controls. On the 14th day, endotheline-1 level was 65 % higher than in the controls after a 0.25 mg 6-OHDA injection (р < 0.01) and 45 % higher after a 0,5 mg injection (р<0.05). At the same time, TNF-α levels increased by 43 % (р < 0.05) and 20 % (р < 0.05) respectively. We also revealed a dose-dependent change of IOP, a disturbance of pupillary light reaction and decreased ocular blood flow after an injection of neurotoxin. Conclusion. A single intravitreal injection of dopaminergic neurotoxin 6-OHDA in the amount of 0.25 or 0.5 mg per eye leads to a neuroinflammation and vascular disorders, which are the main pathogenetic pathways of neurodegeneration. To model its early phase, the 0.25 mg dose is preferable as it causes less dramatic functional disorders. The latter model can be useful for the investigation of retinal neurodegeneration pathogenesis, the search for early diagnostic and prognostic markers thereof and the estimation of therapy effectiveness.

Nasal application of kisspeptin-54 mitigates motor deficits by reducing nigrostriatal dopamine loss in hemiparkinsonian rats

Parkinson’s Disease is a progressive neurodegenerative disorder characterized by motor symptoms resulting from the loss of nigrostriatal dopaminergic neurons. Kisspeptins (KPs) are a family of neuropeptides that are encoded by the Kiss-1 gene, which exert their physiological effects through interaction with the GPR54 receptor. In the current investigation, we investigated the prospective protective effects of central KP-54 treatments on nigrostriatal dopaminergic neurons and consequent motor performance correlates in 6-hydroxydopamine (6-OHDA)-lesioned rats. Male adult Sprague Dawley rats underwent stereotaxic injection of 6-OHDA into the right medial forebrain bundle to induce hemiparkinsonism. Following surgery, rats received chronic central treatments of nasal or intracerebroventricular KP-54 (logarithmically increasing doses) for seven consecutive days. Motor performance was evaluated seven days post-surgery utilizing the open field test and catalepsy test. The levels of dopamine in the striatum were determined with mass spectrometry. Immunohistochemical analysis was conducted to assess the immunoreactivities of tyrosine hydroxylase (TH) and the GPR54 in the substantia nigra. The dose-response curve revealed a median effective dose value of ≈3 nmol/kg for both central injections. Due to its non-invasive and effective nature, nasal administration was utilized in the second phase of our study. Chronic administration of KP-54 (3nmol/kg, nasally) significantly protected 6-OHDA-induced motor deficits. Nasal KP-54 attenuated the loss of nigrostriatal dopaminergic neurons induced by 6-OHDA. Additionally, significant correlations were observed between motor performance and nigrostriatal dopamine levels. Immunohistochemical analysis demonstrated the localization of the GPR54 within TH-positive nigral cells. These findings suggest the potential efficacy of central KP-54 on motor impairments in hemiparkinsonism.

Inspiratory Neural Network Complexity Quantified by Approximate Entropy (ApEn) Analysis in SN 6-OHDA-induced Rat Model of Parkinson’s Disease

Parkinson’s disease (PD) is a multifaceted chronic and progressive neurodegenerative disease that includes a broad range of motor and non-motor symptoms (NMS). Breathing abnormalities are a common NMS in PD, and despite respiratory dysfunction being a major problem in PD as well as respiratory complications being the most common cause of death in PD patients, this NMS receives little attention. Over the last decade, a number of preclinical experimental rat models of PD generated by nigrostriatal site-targeted administration of the neurotoxin 6-hydroxydopamine (6-OHDA) have been used to evaluate various aspects of ventilatory control in PD. While all of these models have been shown to exhibit basal and chemical control of breathing abnormalities that are consistent with those in clinical reports, no studies have assessed the complex intrinsic dynamics that are associated with the central inspiratory neural network in PD. Since preclinical rat models of PD have also been shown exhibit neuroanatomical changes in brainstem regions involved in ventilatory control, it is likely that changes in inspiratory neural network dynamics, which can be assessed using nonlinear dynamical analysis methods, are also present in PD. To begin to assess this possibility, we quantified the complexity of 20 consecutive inspiratory (diaphragm EMG) bursts recorded during basal breathing in urethane-anesthetized spontaneously breathing adult female rats at 4-weeks after unilateral injection of 6-OHDA (n=18) or vehicle (VEH; n=14) directly into the substantia nigra (SN). For these analyses, we calculated approximate entropy (ApEn), which provides a statistical measure of regularity (orderliness) of a signal in the time domain, with values for the embedding dimension m and the tolerance r set to m=3 and r=0.44 SD. ApEn values were significantly higher in 6-OHDA-injected rats compared to VEH-injected rats (0.6169±0.0069 vs 0.5894±0.0087; P=0.017) albeit TI and Tpeak/TI were similar in both groups (TI: 305.8±9.5 vs 300.0±12.7 ms, P=0.713; Tpeak/TI: 51.0±1.3 vs 51.3±1.1%, P=0.904). Other significant differences in temporal features included higher breathing frequencies (93.4±2.4 vs 82.0±3.2 bpm; P=0.007), reduced TE (in ms: 334.7±12.7 vs 445.5±27.4; P=0.001), and decreased inspiratory duty cycle (in %: 47.2±1.1 vs 40.8±2.0; P=0.006) in 6-OHDA (compared to VEH)-injected rats. These observations suggest that respiratory dysfunction in PD is also accompanied by reorganization of the central inspiratory neural network that is associated with greater irregularity or less system order. Additional experiments are needed to identify the precise neural mechanisms underlying this higher system complexity. NIH NS101737; Thomas Hartman Center for Parkinson's Disease Research at Stony Brook University. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 1124: Sympathetic Denervation Models In Murine Femoral Arteries: Acute To Chronic Effects Of 6-hydroxydopamine

Introduction: The autonomic nervous system (ANS) drives blood flow regulation in peripheral arteries. This study aims to develop novel models of sympathetic denervation in peripheral arteries and to characterize subsequent vascular remodeling. We hypothesize that sympathetic denervation of murine femoral arteries can be achieved through local administration of neurotoxin 6-hydroxydopamine (6-OHDA). Methods: BALB/c mice underwent local administration of 6-OHDA to the left femoral artery, while the contralateral artery received vehicle solution. Two approaches were tested: (1) Acute: a one-time topical application following surgical exposure of femoral arteries (N=8, male, Fig. A ), and (2) Chronic: weekly subcutaneous injections near the femoral arteries (N=11, male and female, Fig. B ). Hindlimb perfusion was assessed with laser Doppler imaging, and arterial structure and remodeling were evaluated with histology and immunohistochemistry at one, two, or four weeks. Results: One-time 6-OHDA application (Acute) led to temporary femoral artery denervation (reduction in tyrosine hydroxylase) for one week ( Fig. C – D ), with inflammation observed in both 6-OHDA and vehicle-treated arteries. No vascular wall remodeling was observed after temporary denervation. Weekly subcutaneous 6-OHDA injections (Chronic) resulted in persistent denervation throughout the four-week period in male mice, while female mice displayed variable outcomes ( Fig. E – F ). Sustained sympathetic denervation in the chronic model led to reduced hindlimb blood perfusion at low core temperatures ( Fig. G – H ), though no significant differences were detected in vascular smooth muscle cell phenotype or extracellular matrix composition. Conclusions: Local administration of 6-OHDA is an effective means of peripheral artery sympathetic denervation with controllable duration. This innovative approach of targeting the ANS unveils new prospects for advancing vascular therapies.

Impact of Serotonergic 5HT1A and 5HT2A Receptor Activation on the Respiratory Response to Hypercapnia in a Rat Model of Parkinson's Disease.

In Parkinson's disease (PD), along with typical motor dysfunction, abnormal breathing is present; the cause of which is not well understood. The study aimed to analyze the effects of stimulation of the serotonergic system with 5-HT1A and 5-HT2A agonists in a model of PD induced by injection of 6-hydroxydopamine (6-OHDA). To model PD, bilateral injection of 6-OHDA into both striata was performed in male Wistar rats. Respiratory disturbances in response to 7% hypercapnia (CO2 in O2) in the plethysmographic chamber before and after stimulation of the serotonergic system and the incidence of apnea were studied in awake rats 5 weeks after 6-OHDA or vehicle injection. Administration of 6-OHDA reduced the concentration of serotonin (5-HT), dopamine (DA) and norepinephrine (NA) in the striatum and the level of 5-HT in the brainstem of treated rats, which have been associated with decreased basal ventilation, impaired respiratory response to 7% CO2 and increased incidence of apnea compared to Sham-operated rats. Intraperitoneal (i.p.) injection of the 5-HT1AR agonist 8-OH-DPAT and 5-HT2AR agonist NBOH-2C-CN increased breathing during normocapnia and hypercapnia in both groups of rats. However, it restored reactivity to hypercapnia in 6-OHDA group to the level present in Sham rats. Another 5-HT2AR agonist TCB-2 was only effective in increasing normocapnic ventilation in 6-OHDA rats. Both the serotonergic agonists 8-OH-DPAT and NBOH-2C-CN had stronger stimulatory effects on respiration in PD rats, compensating for deficits in basal ventilation and hypercapnic respiration. We conclude that serotonergic stimulation may have a positive effect on respiratory impairments that occur in PD.

Experimental rationale for the use of thioctic acid and indomethacin in the therapy of the early stage of retinal neurodegeneration

Purpose: to assess the possibility of using thioctic acid and indomethacin tin eyedrops for the therapy of early stages of retinal neurodegeneration.Material and methods. The study was performed on 20 Chinchilla rabbits, including 16 rabbits in whom the neurodegenerative process in the retina was produced by intravitreal installation of 6-hydroxydopamine (6-OHDA), 0.25 mg per eye, and 4 rabbits (8 eyes) served as control. Thioctic acid 0.5% and Indocollir 0.1% were instilled separately and in combination 5 times daily for 7 days after the injection of 6-OHDA. Pupillary light reaction was estimated and eye fundus condition analyzed. Alpha2-macroglobuline ( 2-MG) and SOD activity, dopamine and protein concentrations were measured in retinal homogenates.Results. Instillations of thioctic acid reduced the retinal damage area almost by half as compared to the untreated group (p d 0.005). Indomethacin was less effective, but the combination of the two drugs brought about a maximal reduction of the damage area (p d 0.05 against the untreated group). Both preparations normalized the disturbed pupillary light reaction. The injection of 6-OHDA caused a decrease of dopamine concentration in the retina (0.050 ± 0.009 pg/mg as against 0.095 ± 0.031 pg/mg in healthy controls, р d 0,05). After instillations of indomethacin and thioctic acid, the concentration tended to increase, and it increased significantly after treatment with their combination (0.141 ± 0.037 pg/mg, p d 0.01 as against untreated animals). The activity of 2-MG in retina decreased after a 6-OHDA injection (2.93 ± 0.42 pg/mg as against 5.5 ± 1.2 pg/mg in healthy controls, р < 0 005) just as they increased the decreased SOD (13.30 ± 5.68 U/mg in healthy controls, 5.41 ± 1.50 U/mg in untreated animals, р < 0.05). Both preparations applied separately or in combination caused a significant increase of 2-MG activity (thioctic acid — 4.19 ± 0.91 pg/mg, indomethacin — 6.95 ± 1.09 pg/mg, combination — 7.25 ± 0.63 pg/mg, р < 0.05 as against the untreated animals) and SOD activity (thioctic acid — 8.25 ± 1.27 U/mg, indomethacin — 29.84 ± 6.63 U/mg, combination — 23.99 ± 3.78 U/mg, р < 0.01 as against the untreated animals).Conclusion. The combination of indomethacin and thioctic acid in eyedrops is the most effective for the prevention of eye damage caused by 6-OHDA, which makes this method promising for the treatment of eye diseases accompanied by neurodegeneration.

Possible role of lncRNAs in amelioration of Parkinson’s disease symptoms by transplantation of dopaminergic cells

Long non-coding RNAs (lncRNAs) are biomarkers for diagnosis and treatment of Parkinson’s disease (PD). Since dopaminergic cell transplantation is a clinical method to treat PD, this study investigated the effects of dopaminergic cell therapy on the expression of some lncRNAs and genes related to PD. In this study, Twenty-eight rats were randomly assigned to four experimental groups. The control group (Sal group) received saline injections. The Par group was a PD rat model with 6-hydroxydopamine (6-OHDA) injection in right striatum (ST). PD animals were transplanted by undifferentiated P19 stem cells (Par-E group), and P19-derived dopaminergic cells (Par-N group). Cell transplant effects were evaluated using behavioral tests (cylinder, open field, and rotarod tests), and histological methods (H&E and Nissl staining, and immunohistochemistry). Moreover, the expression of lncRNAs MALAT1, MEG3, and SNHG1, alongside specific neuronal (synaptophysin) and dopaminergic (tyrosine hydroxylase) markers was evaluated by qRT-PCR. Behavioral and histopathological examinations revealed that cell transplantation partially compensated dopaminergic cell degeneration in ST and substantia nigra (SN) of PD rats. The expression of MALAT1, SNHG1, and MEG3 was decreased in the ST of the Par group, while MEG3 and SNHG1 gene expression was increased in PBMC relative to the Sal group. In PBMC of the Par-N group, all three lncRNAs showed a reduction in their expression. Conversely, MALAT1 and SNHG1 expression was increased in ST tissue, while MEG3 gene expression was decreased compared to the Sal group. In conclusion, dopaminergic cell transplantation could change the lncRNAs expression. Furthermore, it partially improves symptoms in PD rats.

Apnea behavior in early- and late-stage mouse models of Parkinson's disease: Cineradiographic analysis of spontaneous breathing, acute stress, and swallowing

This study aimed to evaluate the timing and frequency of spontaneous apneas during breathing and swallowing by using cineradiography on mouse models of early/initial or late/advanced Parkinson’s disease (PD). C57BL/6 J mice received either 6-OHDA or vehicle injections into their right striatum, followed by respiratory movement recordings during spontaneous breathing and swallowing, and a stress challenge, two weeks later. Experimental group animals showed a significantly lower respiratory rate (158.66 ± 32.88 breaths/minute in late PD, 173.16 ± 25.19 in early PD versus 185.27 ± 25.36 in controls; p<0.001) and a significantly higher frequency of apneas (median 1 apnea/minute in both groups versus 0 in controls; p<0.001). Other changes included reduced food intake and the absence of swallow apneas in experimental mice. 6-OHDA-induced nigrostriatal degeneration in mice disrupted respiratory control, swallowing, stress responsiveness, and feeding behaviors, potentially hindering airway protection and elevating the risk of aspiration.

Aerobic exercise modulates the striatal Erk/MAPK signaling pathway and improves LID in a mouse model of Parkinson's disease

ObjectiveTo investigate the role of the striatal extracellular signal-regulated kinase (Erk1/2) and its phosphorylation (p-Erk1/2) in aerobic training to alleviate the development of the L-DOPA induced dyskinesia (LID) in PD mice. MethodsForty-eight male C57BL/6 N mice were randomly divided into the 6-OHDA surgery group (6-OHDA, n=42) and the sham surgery group (Sham, n=6). A two-point injection of 6-OHDA into the right striatum was used to establish a lateralized injury PD model. PD mice were randomly divided into a PD control group (PD, n=13) and a PD exercise group (PDE, n=16), this is followed by 4 weeks of L-DOPA treatment, and PDE mice received concurrent running table training (18 m/min, 40 min/day, 5 times/week). AIM scores were performed weekly, and mice were assessed for motor function after 4 weeks using the rotarod, open field, and gait tests. Immunohistochemistry was used to test nigrostriatal TH expression, Western blot was used to determine Erk1/2 and p-Erk1/2 protein expression, and immunofluorescence double-labeling technique was used to detect Erk1/2 and p-Erk1/2 co-expression with prodynorphin (PDYN). Results(1) All AIM scores of PD and PDE mice increased significantly (P < 0.05) with the prolongation of L-DOPA treatment. Compared with PD, all AIM scores were significantly lower in PDE mice (P < 0.05). (2) After 4 weeks, the motor function of PD mice was significantly reduced compared with Sham (P < 0.05 or P < 0.01); compared with PD, the motor function of PDE mice was significantly increased (P < 0.05). (3) Compared with Sham, the expression of Erk1/2 protein, the number of positive cells of Erk1/2 and p-Erk1/2 and the number of positive cells co-expressed with PDYN were significantly increased in PD mice (P < 0.05); compared with PD, Erk1/2 protein expression was significantly decreased in PDE mice (P < 0.05), and the number of Erk1/2 and p-Erk1/2 positive cells was significantly reduced (P < 0.05). Conclusion4 weeks of aerobic exercise can effectively alleviate the development of L-DOPA-induced dyskinesia and improve motor function in PD mice. The related mechanism may be related to the inhibition of striatal Erk/MAPK signaling pathway overactivation by aerobic exercise, but this change did not occur selectively in D1-MSNs.

Characterization of graded 6-Hydroxydopamine unilateral lesion in medial forebrain bundle of mice

Parkinson’s disease (PD) is the second most common age-related neurodegenerative disease, with a progressive loss of dopaminergic cells and fibers. The purpose of this study was to use different doses of 6-hydroxydopamine (6-OHDA) injection into the medial forebrain bundle (MFB) of mice to mimic the different stages of the disease and to characterize in detail their motor and non-motor behavior, as well as neuropathological features in the nigrostriatal pathway. MFB were injected with 0.5 μg, 1 μg, 2 μg of 6-OHDA using a brain stereotaxic technique. 6-OHDA induced mitochondrial damage dose-dependently, as well as substantia nigra pars compacta (SNpc) tyrosine hydroxylase-positive (TH+) cell loss and striatal TH fiber loss. Activation of astrocytes and microglia in the SNpc and striatum were consistently observed at 7 weeks, suggesting a long-term glial response in the nigrostriatal system. Even with a partial or complete denervation of the nigrostriatal pathway, 6-OHDA did not cause anxiety, although depression-like behavior appeared. Certain gait disturbances were observed in 0.5 μg 6-OHDA lesioned mice, and more extensive in 1 μg group. Despite the loss of more neurons from 2 μg 6-OHDA, there was no further impairment in behaviors compared to 1 μg 6-OHDA. Our data have implications that 1 μg 6-OHDA was necessary and sufficient to induce motor and non-motor symptoms in mice, thus a valuable mouse tool to explore disease progression and new treatment in PD.

Calcitriol restrains microglial M1 polarization and alleviates dopaminergic degeneration in hemiparkinsonian mice by boosting regulatory T-cell expansion.

Vitamin D deficiency is a risk factor for Parkinson's disease (PD) and vitamin D supplementation robustly alleviates neurodegeneration in PD models. However, the mechanisms underlying this effect require further clarification. Current evidence suggests that harnessing regulatory T cells (Treg) may mitigate neuronal degeneration. In this study, we investigated the therapeutic effects of vitamin D receptor activation by calcitriol on PD, specifically focusing on its role in Treg. Hemiparkinsonian mice model was established through the injection of 6-OHDA into the striatum. Mice were pretreated with calcitriol before 6-OHDA injection. The motor performance, dopaminergic neuronal survival, contents of dopamine, and dopamine metabolites were evaluated. The pro-inflammatory cytokines levels, T-cell infiltration, mRNA expression of indicated microglial M1/M2 phenotypic markers, and microglial marker in the midbrain were detected. Populations of Treg in the splenic tissues were assessed using a flow cytometry assay. PC61 monoclonal antibody was applied to deplete Treg in vivo. We show that calcitriol supplementation notably improved motor performance and reduced dopaminergic degeneration in the 6-OHDA-induced PD model. Mechanistically, calcitriol promoted anti-inflammatory/neuroprotective Treg and inhibited pro-inflammatory/neurodestructive effector T-cell generation in this model. This process significantly inhibited T-cell infiltration in the midbrain, restrained microglial activation, microglial M1 polarization, and decreased pro-inflammatory cytokines release. This more favorable inflammatory microenvironment rescued dopaminergic degeneration. To further verify that the anti-inflammatory effects of calcitriol are associated with Treg expansion, we applied an antibody-mediated Treg depletion assay. As predicted, the anti-inflammatory effects of calcitriol in the PD model were diminished following Treg depletion. These findings suggest that calcitriol's anti-inflammatory and neuroprotective effects in PD are associated with its potential to boost Treg expansion.

TREM2 gene induces differentiation of induced pluripotent stem cells into dopaminergic neurons and promotes neuronal repair via TGF-β activation in 6-OHDA-lesioned mouse model of Parkinson's disease.

Induced pluripotent stem cells (iPSCs) hold a promising potential for rescuing dopaminergic neurons in therapy for Parkinson's disease (PD). This study clarifies a TREM2-dependent mechanism explaining the function of iPSC differentiation in neuronal repair of PD. PD-related differentially expressed genes were screened by bioinformatics analyses and their expression was verified using RT-qPCR in nigral tissues of 6-OHDA-lesioned mice. Following ectopic expression and depletion experiments in iPSCs, cell differentiation into dopaminergic neurons as well as the expression of dopaminergic neuronal markers TH and DAT was measured. Stereotaxic injection of 6-OHDA was used to develop a mouse model of PD, which was injected with iPSC suspension overexpressing TREM2 to verify the effect of TREM2 on neuronal repair. TREM2 was poorly expressed in the nigral tissues of 6-OHDA-lesioned mice. In the presence of TREM2 overexpression, the iPSCs showed increased expression of dopaminergic neuronal markers TH and DAT, which facilitated the differentiation of iPSCs into dopaminergic neurons. Mechanistic investigations indicated that TREM2 activated the TGF-β pathway and induced iPSC differentiation into dopaminergic neurons. Invivo data showed that iPSCs overexpressing TREM2 enhanced neuronal repair in 6-OHDA-lesioned mice. This work identifies a mechanistic insight for TREM2-mediated TGF-β activation in the regulation of neuronal repair in PD and suggests novel strategies for neurodegenerative disorders.