Tyrosine Hydroxylase Expression Research Articles

Abstract 4125419: Very-Low-Density Lipoprotein Induces Neuronal Growth with Increased Tyrosine Hydroxylase Expression through Cardiomyocyte-Secreted Exosomes in Metabolic Syndrome

Introduction: Metabolic syndrome (MS) is an important predisposing factor for atrial fibrillation, and is highly related to the autonomic nervous system. Triglyceride-rich very-low-density lipoprotein (VLDL) has specific VLDL receptors (VLDLR), which are abundantly expressed in cardiomyocytes. Hypothesis: VLDL affects neuronal growth through cardiomyocyte-secreted exosomes in metabolic syndrome. Goals/Aims: To determine whether exosomes from VLDL-incubated cardiomyocytes can alter neuronal proliferation. Methods/ Approach: HL-1 cardiomyocytes were incubated with VLDL isolated from human subjects with MS, and exosomes were extracted at the end of the incubation period. VLDLR knockdown HL-1 cells were used to determine the role of VLDLR in exosome secretion. The molecular contents of exosomes were determined by mass spectrometry, which identified neurotropic proteins only detected in the exosomes of MS VLDL-incubated cardiomyocytes. We selected transmembrane protein 14B (TMEM14B) to investigate its effect on neuronal proliferation in P19 cells. Results: Regardless of the knockdown or overexpression of VLDLR in HL-1 cardiomyocytes, exosomes isolated from after VLDL incubation were shown to cause 2.062-fold and 2.084-fold expression of tyrosine hydroxylase (TH) in P19 cells, relative to P19 cells induced by non-VLDL-treated HL-1 cardiomyocyte-secreted exosomes. A similar trend could be seen with HL-1 cardiomyocytes without VLDLR editing, as VLDL-incubated HL-1 cardiomyocytes produced exosomes that could cause 1.926-fold increase in P19 TH expression, relative to treatment by exosomes from HL-1 cardiomyocytes without VLDL-incubation. The P19 cells incubated with TMEM14B demonstrated increased neurite length, with an average of 2.342-fold neurite length when compared with that of control cells. Conclusions: VLDL in MS induced HL-1 cardiomyocytes to secrete exosomes containing neurotrophic molecules and increased dendritic growth of neuronal cells. This effect was VLDLR-independent and was partially caused by TMEM14B in the exosomes.

Inhibition of sympathetic tone via hypothalamic descending pathway propagates glucocorticoid-induced endothelial impairment and osteonecrosis of the femoral head

Osteonecrosis of the femoral head (ONFH) is a common complication of glucocorticoid (GC) therapy. Recent advances demonstrate that sympathetic nerves regulate bone homeostasis, and GCs lower the sympathetic tone. Here, we show that the dramatically decreased sympathetic tone is closely associated with the pathogenesis of GC-induced ONFH. GCs activate the glucocorticoid receptor (GR) but hinder the activation of the mineralocorticoid receptor (MR) on neurons in the hypothalamic paraventricular nucleus (PVN). This disrupts the balance of corticosteroid receptors (GR/MR) and subsequently reduces the sympathetic outflow in the PVN. Vascular endothelial cells rapidly react to inhibition of sympathetic tone by provoking endothelial apoptosis in adult male mice treated with methylprednisolone (MPS) daily for 3 days, and we find substantially reduced H-type vessels in the femoral heads of MPS-treated ONFH mice. Importantly, treatment with a GR inhibitor (RU486) in the PVN promotes the activation of MR and rebalances the ratio of GR and MR, thus effectively boosting sympathetic outflow, as shown by an increase in tyrosine hydroxylase expression in both the PVN and the sympathetic postganglionic neurons and an increase in norepinephrine levels in both the serum and bone marrow of the femoral head of MPS-treated mice. Rebalancing the corticosteroid receptors mitigates GC-induced endothelial impairment and ONFH and promotes angiogenesis coupled with osteogenesis in the femoral head, while these effects are abolished by chemical sympathectomy with 6-OHDA or adrenergic receptor-β2 (Adrb2) knockout. Furthermore, activating Adrb2 signaling in vivo is sufficient to rescue the GC-induced ONFH phenotype. Mechanistically, norepinephrine increases the expression of the key glycolytic gene 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) via Adrb2-cyclic AMP response element-binding protein (CREB) signaling. Endothelial-specific overexpression of PFKFB3 attenuates endothelial impairment and prevents severe osteonecrosis in MPS-treated Adrb2 knockout mice. Thus, GC inhibits sympathetic tone via the hypothalamic descending pathway, which, in turn, acts as a mediator of GC-induced ONFH.

Diagnostic Potential of NEAT1, hsa-let-7a-5p, and miR-506-3p in Early-stage Parkinson's Disease.

Parkinson's disease (PD) is a multifaceted disease that is influenced by both genetic and environmental parameters. Non-coding RNAs have been shown to be ideal biomarkers for several diseases, including PD. This study was conducted to evaluate the expression levels of NEAT1, hsa-let-7a-5p, and miR-506-3p in individuals with PD to assess their efficacy for early-stage PD diagnosis. Twenty-four patients with PD and 29 healthy individuals participated in this study. The IntaRNA tool was used to predict potential base pairings between NEAT1 and let-7a-5p, and NEAT1 and miR-506-3p. RT-qPCR was employed to measure the relative expression of tyrosine hydroxylase (TH), as well as nuclear enriched abundant transcript 1 (NEAT1), hsa-let-7a-5p, and miR-506-3p levels in both groups. The area under the receiver operating characteristic curve (AUC) was calculated to evaluate the diagnostic value. The PD group exhibited significantly elevated NEAT1 expression levels compared to the healthy control group. While the PD group exhibited an insignificant decreased TH expression level relative to the healthy group. Furthermore, the levels of hsa-let-7a-5p and miR-506-3p expression were seen to be decreased in patients with PD in comparison to the control group. Integration of NEAT1, hsa-let-7a-5p, and miR-506-3p levels significantly enhanced diagnostic capabilities and increased the AUC to 0.9501 (95% confidence interval: 0.8978-1.000, p < .0001). The elevated NEAT1 expression and the decreased expression of hsalet- 7a-5p and miR-506-3p in PD patients indicate that these factors might contribute to the disease's pathogenesis. Combining the ROC curves of NEAT1 and hsa-let-7a-5p with miR-506-3p showed improved sensitivity and specificity, facilitating more accurate PD diagnosis. More importantly, they may contribute as promising non-invasive biomarkers for PD diagnosis.

Mechanical acupuncture at HT7 attenuates alcohol self-administration in rats by modulating neuroinflammation and altering mPFC-habenula-VTA circuit activity.

Alcohol use disorder is a chronic disorder with significant limitations in pharmacological treatments, necessitating the exploration of non-pharmacological interventions. We used a model of alcohol self-administration (10% v/v) to analyze behavioral, neurochemical, and signaling mechanisms. Our findings demonstrate that stimulation of the HT7 acupuncture point significantly decreased the frequency of active lever presses in rats self-administering alcohol (p < 0.05). Alcohol self-administration increased microglial activity and sigma 1 receptor expression in the habenula (Hb), while HT7 stimulation mitigated these effects, decreasing microglial activity and sigma 1 receptor levels (p < 0.05). Additionally, alcohol self-administration reduced brain-derived neurotrophic factor (BDNF) expression in the medial prefrontal cortex (mPFC) and increased tyrosine hydroxylase (TH) levels in the ventral tegmental area (VTA) (p < 0.05). HT7 stimulation reversed these alterations by increasing BDNF expression in the mPFC and decreasing TH levels in the VTA (p < 0.05). Further investigation revealed that BDNF microinjection into the mPFC inhibited sigma 1 receptor activity in the Hb, while microglial inhibition in the Hb decreased TH expression in the VTA (p < 0.05). The administration of the microglial inhibitor MINO to the Hb also reduced alcohol self-administration (p < 0.05). These results suggest that HT7 stimulation regulates the mPFC-Hb-VTA circuit, leading to decreased alcohol-seeking behavior. Our study demonstrates that HT7 acupuncture can modulate the mPFC-Hb-VTA circuit, providing a potential non-pharmacological treatment for alcohol-seeking behavior by influencing microglial activity, sigma 1 receptor expression, and TH levels. These findings contribute to a deeper understanding of the neural mechanisms underlying acupuncture's therapeutic effects on alcohol use disorder.

Unveiling the Therapeutic Potential of Small Molecule of SVAK-12: A Comprehensive In Silico, In Vitro, and In Vivo Studies on its Neuroprotective Effects and Molecular Interactions in Parkinson's Disease.

Parkinson's disease (PD) is a neurodegenerative disorder associated with a progressive loss of dopaminergic cells and as of now, there is no established definitive treatment available for this condition. In this study, the focus was on investigating the impact of SVAK-12, a small molecule that can cross the blood-brain barrier and remain stable without structural changes. The effect of SVAK-12 was investigated in vitro on neurotoxicity, in vivo model of Parkinson's Diseases and in silico. Through in vitro and in vivo experiments, as well as molecular docking simulations, it was found that SVAK-12 (375 ng.ml) led to increased cell viability, reduced cellular damage, and decreased production of NO and ROS. Additionally, it boosted levels of important neurotrophic factors like BDNF (130.49%) and GDNF (116.38%), potentially aiding in alleviating motor disability and depression. The study also highlighted SVAK-12's potential as a therapeutic candidate for neurological disorders due to its ability to increase tyrosine hydroxylase expression and dopamine levels (4.84 times). While it did not significantly improve motor symptoms in vivo, it did enhance motor asymmetry in the forelimbs and gene expression related to brain regions. Besides, it induced significant BMP-2 gene expression in substantial nigra regions without significant changes in GDNF and Nurr1 gene expression in the striatum expression. The docking of SVAK-12, Levodopa, Amantadine, Biperiden, Selegiline, and Rasagiline to the binding site of GFRα1, sortilin, and TrkB showed that SVAK-12 had greater MolDock score than Selegiline and Amantadine for GFRα1 and greater than amantadine for Sortilin and TrKB. Overall, the study suggests that SVAK-12's neuro-biocompatibility, ability to reduce free radicals, and enhanced neurotrophic factors make it a promising candidate as a neuroprotective drug.

RETRACTION: The oxysterol 27-hydroxycholesterol regulates α-synuclein and tyrosine hydroxylase expression levels in human neuroblastoma cells through modulation of liver X receptors and estrogen receptors-relevance to Parkinson's disease.

Retraction: G. Marwarha, T. Rhen, T. Schommer, and O. Ghribi, "The Oxysterol 27-Hydroxycholesterol Regulates α-Synuclein and Tyrosine Hydroxylase Expression Levels in Human Neuroblastoma Cells Through Modulation of Liver X Receptors and Estrogen Receptors-Relevance to Parkinson's Disease," Journal of Neurochemistry 119, no. 5 (2011): 1119-1136, https://doi.org/10.1111/j.1471-4159.2011.07497.x. The above article, published online on 23 September 2011 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor-in-Chief, Andrew Lawrence; the International Society for Neurochemistry; and John Wiley & Sons Ltd. The retraction has been agreed upon following an investigation by the authors' institution, the University of North Dakota, which determined that Figure2E was falsified by the corresponding author Othman Ghribi. Source data were not available for the article. The other authors were unaware of Ghribi's actions and not in any way involved. All authors were notified of the retraction decision, but did not respond.

Differential roles of NR4A2 (NURR1) paralogs in the brain and behavior of zebrafish.

Dopaminergic (DAnergic) dysfunction and imbalanced dopamine (DA) levels are known contributors to the pathogenesis of numerous psychiatric and neurodegenerative disorders. Of the many identified risk factors for DA-associated disorders, nuclear receptor subfamily 4 group A2 (NR4A2; or nuclear receptor related-1 protein (NURR1)), a transcription factor involved in DAnergic differentiation, has been associated with Parkinson's disease and attention deficit hyperactive disorder (ADHD). In zebrafish, transient loss of nr4a2 was previously shown to decrease tyrosine hydroxylase (TH) expression and impair locomotion. To further characterize the roles of the two zebrafish nr4a2 paralogs, nr4a2a, and nr4a2b, we produced targeted loss-of-function mutants and examined DAnergic neuron regeneration, oxidative respiration, and behavioral traits. The loss of nr4a2a function more closely recapitulated Parkinsonian phenotypes and affected neurotrophic factor gene expression. Conversely, nr4a2b mutants displayed behavioral symptoms reminiscent of mice deficient in Nr4a2 with increased neurotrophic output. In contrast, nr4a2b mutants also displayed increased metabolic input from non-mitochondrial sources indicative of high cytosolic reactive oxygen species and perturbed mitochondrial function. The nr4a2a mutants also showed increased maximal respiration, which may suggest a compensatory mechanism to meet the metabolic requirements of DAnergic neuron health. Overall, the zebrafish mutants generated in this study helped uncover molecular mechanisms involved in DA-related disease pathologies, and in the regeneration of DAnergic neurons.

Bisphenol F-induced precocious genesis of aggressive neurobehavioral response is associated with heightened monoamine oxidase activity and neurodegeneration in zebrafish brain

The production and use of plastics and plastics products has increased dramatically in recent decades. Moreover, their unprotected disposal into ambient life sustaining environment poses a significant health risk. Bisphenol F (BPF) an alternative to bisphenol A (BPA) has been extensively employed for making of plastics. Recent reports have documented the neurotoxic potential of BPF through induction of altered neurochemical profile, microglia-astrocyte-mediated neuroinflammation, oxidative stress, transformed neurobehavioral response, cognitive dysfunction, etc. In the present study, our approach was to understand the underlying mechanism of BPF-persuaded genesis of aggressive neurobehavioral response in zebrafish. The basic findings advocated a temporal transformation in native explorative behaviour and progressive induction of aggressive behavioural response in zebrafish following exposure to BPF. Our neurobehavioral findings supported the argument of oxidative stress-mediated neuromorphological transformation in the periventricular grey zone (PGZ) of the zebrafish brain. In line with earlier reports, our findings also showed that heightened monoamine oxidase (MAO) activity and downregulation in tyrosine hydroxylase expression in the zebrafish brain is associated with the precocious genesis of aggressive neurobehavioral response in zebrafish brain. Our findings also shed light on BPF-instigated apoptotic neuronal death as revealed by augmented chromatin condensation and cleaved caspase-3 expression. Further observation showed that the downregulation of NeuN (a marker of post-mitotic mature neuron) expression provided substantial neurotoxicity, leading to neurodegeneration in the PGZ region of the zebrafish brain. These basic findings grossly advocate that BPF acts as a potent neurotoxicant in transmuting native neurobehavioral response through the induction of oxidative stress, heightened MAO activity and neuromorphological transformation in the zebrafish brain.

Carvedilol through ß1-Adrenoceptor blockade ameliorates glomerulonephritis via inhibition of oxidative stress, apoptosis, autophagy, ferroptosis, endoplasmic reticulum stress and inflammation

Glomerulonephritis (GN) is one of the main causes of end stage renal disease and requires an effective treatment for inhibiting GN. Renal nerves through efferent (RENA) and afferent (RANA) innervation to glomeruli regulate the glomerular function. We delineated the role of RENA and RANA on anti-Thy1.1-induced GN. Female Wistar rats were divided into Control, Thy1.1 plus anti-Thy1.1, bilaterally renal nerve denervation (DNX) plus anti-Thy1.1, and topical capsaicin to bilateral renal nerves for selective ablation of RANA (DNAX) plus anti-Thy1.1. We examined RANA and RENA response to anti-Thy1.1 and compared the effect of DNX or DNAX on urinary oxidative stress, renal gp91, tyrosine hydroxylase (TH), calcitonin gene-related peptide (CGRP), apoptosis, autophagy, ferroptosis, antioxidant enzymes, endoplasmic reticulum (ER) stress and inflammation by western blot. Anti-Thy1.1 significantly enhanced RENA, but did not affect RANA. DNX significantly decreased TH and CGRP expression, whereas DNAX only reduced CGRP expression. Anti-Thy1.1 significantly increased glomerulosclerosis injury, urinary protein, electron paramagnetic resonance signals of alpha-(4-pyridyl-N-oxide)-N-tert-butylnitrone adducts, 8-isoprostane and nitrotyrosine levels, NADPH oxidase gp91phox (gp91), macrophage/monocyte (ED-1), GRP-78, Beclin-1/LC3-II, Bax/caspase-3/poly(ADP-ribose) polymerase expression, inflammatory cytokines levels and decreased renal Copper/Zinc superoxide dismutase, Cystine/glutamate transporter (xCT) and Glutathione peroxidase 4 (GPX4) expression vs. Control. The enhanced oxidative parameters or reduced antioxidant defense by anti-Thy1.1 were significantly attenuated by DNX but not DNAX. Additionally, oral ß1-adrenoceptor antagonist-Carvedilol at an early stage reduced anti-Thy1.1 increased proteinuria level and oxidative parameters. Our data suggest that DNX and ß1-adrenoceptor antagonist-Carvedilol efficiently attenuate oxidative stress, inflammation, ER stress, autophagy, ferroptosis and apoptosis in GN

Bee Venom Reduces Early Inflammation and Oxidative Stress Associated with Lipopolysaccharide-induced Alpha-synuclein in the Substantia Nigra-striatum Axis.

Neuroinflammation and oxidative stress are important features in the pathogenesis and development of synucleinopathies, the glial activation and upregulation of pro-inflammatory and oxidative mediators induce alpha-synuclein (α-syn) accumulation. Recent studies have shown that bee venom (BV) has beneficial effects on symptoms of these neurodegenerative diseases. BV is known to exert anti-inflammatory and anti-oxidative effects. Here, we investigated the effects of BV over the different inflammatory and oxidative markers, and in the expression of α-syn and tyrosine hydroxylase (TH) in a lipopolysaccharide (LPS)-induced rat model of synucleinopathies. We examined whether BV (1.5 mg/kg by acupoint injection ST36 six times every 48 h) could change the α-syn and TH expression measured by western blotting, also, observed the activation of microglia and astrocytes by immunofluorescence, quantified the proinflammatory cytokines levels of tumoral necrosis factor-α (TNF-α) and Interleukin-1β (IL-1β) by enzyme-linked immunosorbent assay (ELISA), and estimated the lipid peroxidation and the activity of superoxide dismutase (SOD) and catalase (CAT) by colorimetric kits in LPS-treated rats (2.5 µg by a single dose intranigral injection) in substantia nigra (SN) and striatum (STR) brain areas. In the LPS-injected rat brain, BV treatment reduced α-syn levels and increased the TH levels. In addition, we observed lower microglia and astrocyte activation in SN and STR. Furthermore, BV decreases IL-1β and lipid peroxidation and increases the CAT activity in the STR. These results indicate that BV can restore the α-syn and TH levels possibly by the inhibition of LPS-induced neuroinflammation and oxidation, also, these results suggest that BV could be a promising treatment option for synucleinopathies.

Neuroprotective effects of intranasal extracellular vesicles from human platelet concentrates supernatants in traumatic brain injury and Parkinson’s disease models

BackgroundThe burgeoning field of regenerative medicine has significantly advanced with recent findings on biotherapies using human platelet lysates (HPLs), derived from clinical-grade platelet concentrates (PCs), for treating brain disorders. These developments have opened new translational research avenues to explore the neuroprotective effects of platelet-extracellular vesicles (PEVs). Their potential in managing neurodegenerative conditions like traumatic brain injury (TBI) and Parkinson’s disease (PD) warrants further exploration. We aimed here to characterize the composition of a PEV preparation isolated from platelet concentrate (PC) supernatant, and determine its neuroprotective potential and neurorestorative effects in cellular and animal models of TBI and PD.MethodsWe isolated PEVs from the supernatant of clinical-grade PC collected from healthy blood donors utilizing high-speed centrifugation. PEVs were characterized by biophysical, biochemical, microscopic, and LC–MS/MS proteomics methods to unveil biological functions. Their functionality was assessed in vitro using SH-SY5Y neuronal cells, LUHMES dopaminergic neurons, and BV-2 microglial cells, and in vivo by intranasal administration in a controlled cortical impact (CCI)-TBI model using 8-weeks-old male C57/BL6 mice, and in a PD model induced by MPTP in 5-month-old male C57/BL6 mice.ResultsPEVs varied in size from 50 to 350 nm, predominantly around 200 nm, with concentrations ranging between 1010 and 1011/mL. They expressed specific platelet membrane markers, exhibited a lipid bilayer by cryo-electron microscopy and, importantly, showed low expression of pro-coagulant phosphatidylserine. LC–MS/MS indicated a rich composition of trophic factors, including neurotrophins, anti-inflammatory agents, neurotransmitters, and antioxidants, unveiling their multifaceted biological functions. PEVs aided in the restoration of neuronal functions in SH-SY5Y cells and demonstrated remarkable neuroprotective capabilities against erastin-induced ferroptosis in dopaminergic neurons. In microglial cells, they promoted anti-inflammatory responses, particularly under inflammatory conditions. In vivo, intranasally delivered PEVs showed strong anti-inflammatory effects in a TBI mouse model and conserved tyrosine hydroxylase expression of dopaminergic neurons of the substantia nigra in a PD model, leading to improved motor function.ConclusionsThe potential of PEV-based therapies in neuroprotection opens new therapeutic avenues for neurodegenerative disorders. The study advocates for clinical trials to establish the efficacy of PEV-based biotherapies in neuroregenerative medicine.Graphical

Tyrosine hydroxylase expression and neuronal loss in the male and female adolescent ventral tegmental area

Adolescence is a critical period of development characterized by numerous behavioral and neuroanatomical changes. While studies of adolescent neurodevelopment typically compare adolescent age groups with young adults, there are fewer studies that assess developmental trajectories within the adolescent period. In the adolescent prefrontal cortex, some maturational changes take place linearly/chronologically, while others are associated specifically with pubertal onset. The adolescent ventral tegmental area (VTA), a primary source of forebrain dopamine, is relatively understudied during this period. In the present study, dopamine neuron number, total neuron number and tyrosine hydroxylase expression are assessed in the male and female rat VTA at three timepoints: postnatal day(P) 30 (pre-pubertal), P40 (post-pubertal for females, pre-pubertal for males) and P60 (post-pubertal). There was a non-significant trend for a reduction in total VTA neuron number between P30 and P60, but there was a significant reduction in dopamine neuron number across age. The expression of tyrosine hydroxylase did not change with age. However, in a second cohort of subjects, brain tissue was collected pre-pubertal, from recently post-pubertal males and females, and young adults. In this cohort, there was a sex-specific and transient decrease in tyrosine hydroxylase expression in recently post-pubertal males. These results suggest a selective pruning of VTA dopamine cells between early adolescence and young adulthood, while pubertal onset may coincide with a rapid maturation of these neurons. These findings may have implications for psychiatric disorders associated with dopamine dysfunction that tend to manifest during adolescence.

Activating α7nAChR suppresses systemic inflammation by mitigating neuroinflammation of the medullary visceral zone in sepsis in a rat model.

Our previous studies have shown that activating α7nAChRs suppresses systemic inflammation and immunity through the cholinergic anti-inflammatory pathway (CAP) in early sepsis. Now that the medullary visceral zone (MVZ) is the center of CAP and responsible for regulating systemic inflammation, what changes will occur in MVZ's pathology and function in sepsis, especially when interfering with α7nAChRs? Does activation of MVZ's α7nAChRs contribute to the inhibition of systemic inflammation? To clarify these issues, we explored the systemic inflammation and immunity state by detecting serum levels of TNF-α, IL-6, HMGB1, sCD14, and CD4+CD25+Treg and TH17 lymphocytes percentage, meanwhile, we analyzed the apoptosis of cholinergic and catecholaminergic neurons and the expressions of tyrosine hydroxylase (TH) and choline acetyltransferase (CHAT) in MVZ in sepsis and the interfering effects on α7nAChRs. In this study, we found that in sepsis, serum TNF-α, IL-6, HMGB1, sCD14, CD4+CD25+Treg, and TH17 lymphocytes significantly increased and the ratio of Treg/TH17 significantly decreased, cholinergic and catecholaminergic neurons underwent apoptosis with low expressions of TH and CHAT in MVZ; activation of α7nAChRs not only significantly decreased the levels of septic serum TNF-α, IL-6, HMGB1, sCD14, and TH17 lymphocytes (P ＜ 0.05), but also significantly reduced cholinergic and catecholaminergic neurons' apoptosis, and promoted expressions of TH/CHAT. Our study reveals that sepsis undermines MVZ through neuroinflammation which contributes to the uncontrolled systemic inflammation. Activating central α7nAChRs is not only helpful to restore MVZ's structure and function but also beneficial to subside the inflammatory storm in sepsis. Even if MVZ is damaged in sepsis, cholinergic neurons in MVZ still regulate the systemic inflammation stably.

Basal release and relaxation responses to 6-nitrodopamine in swine carotid, coronary, femoral, and renal arteries

Mammalian and reptilian vascular tissues present basal release of 6-nitrodopamine, which is reduced when the tissues are pre-incubated with the NO synthase inhibitor L-NG-Nitro arginine methyl ester (L-NAME), or when the endothelium is mechanically removed. 6-Nitrodopamine induces vasorelaxation in pre-contracted vascular rings by antagonizing the dopaminergic D2-like receptor. Here it was investigated whether male swine vessels (including carotid, left descendent coronary, renal, and femoral arteries) release 6-nitrodopamine, dopamine, noradrenaline, and adrenaline, as measured by liquid chromatography coupled to tandem mass spectrometry. The in vitro vasorelaxant action of 6-nitrodopamine was evaluated in carotid, coronary, renal, and femoral arteries precontracted by U-46619 (3 nM), and compared to that induced by the dopamine D2-receptor antagonist L-741,626. Expression of tyrosine hydroxylase and the neuromaker calretinin was investigated by immunohistochemistry. All vascular tissues presented basal release of endothelium-derived catecholamines. The relaxation induced by 6-nitrodopamine was not affected by preincubation of the tissues with either L-NAME (100 μM, 30-min preincubation) or the heme-site inhibitor of soluble guanylyl cyclase ODQ (100 μM, 30-min preincubation). Electrical field stimulation (EFS)-induced contractions were significantly potentiated by previous incubation with L-NAME, but unaffected by ODQ preincubation. The contractions induced by EFS were reduced by preincubation with either 6-nitrodopamine or L-741,626. Immunohistochemistry in all arteries revealed the presence of tyrosine hydroxylase in the endothelium, whereas immunoreactivity for calretinin was negative. Swine vessels present basal release of endothelium-derived catecholamines and expression of tyrosine hydroxylase in the endothelium. The vasodilation induced by 6-nitrodopamine is due to blockade of dopaminergic D2-like receptors.

Oxytocin Protects Nigrostriatal Dopamine Signal via Activating GABAergic Circuit in the MPTP-Induced Parkinson's Disease Model.

The most pronounced neuropathological feature of Parkinson's disease (PD) is the loss of dopamine (DA) neurons in the substantia nigra compacta (SNc), which depletes striatal DA. Hypothalamic oxytocin is found to be reduced in PD patients and closely interacts with the DA system, but the role of oxytocin in PD remains unclear. Here, the disturbances of endogenous oxytocin level and the substantia nigra (SN) oxytocin receptor expression in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse model is observed, correlated with the striatal tyrosine hydroxylase (TH) expression reduction. Killing/silencing hypothalamic oxytocin neurons aggravates the vulnerability of nigrostriatal DA signal to MPTP, whereas elevating oxytocin level by intranasal delivery or microinjecting into the SN promotes the resistance. In addition, knocking out SN oxytocin receptors induces the time-dependent reductions of SNc DA neurons, striatal TH expression, and striatal DA level by increasing neuronal excitotoxicity. These results further uncover that oxytocin dampens the excitatory synaptic inputs onto DA neurons via activating oxytocin receptor-expressed SN GABA neurons, which target GABA(B) receptors expressed in SNc DA neuron-projecting glutamatergic axons, to reduce excitotoxicity. Thus, besides the well-known prosocial effect, oxytocin acts as a key endogenous factor in protecting the nigrostriatal DA system.

Abstract Mo073: Elucidating the Role of Cardiac Radiotherapy on the Sympathetic Nervous System as a Treatment for Ventricular Tachycardia

Ventricular tachycardia (VT) is a dangerous arrhythmia that occurs in structurally and electrically heterogenous regions of the diseased heart, leading to sudden cardiac arrest and death if left untreated. One of the main drivers underlying the arrhythmogenic substrate is abnormal remodeling of the autonomic nervous system after cardiac injury, resulting in a heterogenous distribution of sympathetic innervation and subsequent ventricular arrhythmia. In patients with VT refractory to conventional therapy, noninvasive cardiac stereotactic body radiotherapy (SBRT) has emerged as a promising new treatment. Here, we investigated whether cardiac radiotherapy homogenizes the arrhythmogenic substrate through induction of cardiac sympathetic nerve innervation and activity in a closed-chest ischemia reperfusion murine model of ischemic heart failure. Using single nucleus-RNA sequencing, we identified global transcriptomic changes in the left ventricle 6 weeks after 25 Gray (Gy) cardiac X-ray radiation. Notably, these changes included increased expression of neurotrophins such as brain derived neurotrophic factor, a growth factor essential for neuronal survival and development, in irradiated hearts compared to sham controls. Consequently, Western blot and immunofluorescence showed increased expression of the sympathetic neuronal markers tyrosine hydroxylase and neuropeptide Y at the scar border zone of irradiated mice. Similar immunofluorescence results were observed at the irradiated border zone from the explanted heart of a patient previously targeted with 25 Gy compared to a nontargeted remote region within the same heart. These findings suggest a novel role for radiation-induced remodeling of the sympathetic nervous system in the anti-arrhythmic response to SBRT.

Rotenone and Its Derivative, Rotenoisin A, Induce Neurodegeneration Differentially in SH-SY5Y Cells.

Rotenone (ROT), the most significant rotenoid, which has shown anticancer activity, has also been reported to be toxic to normal cells, inducing Parkinson's disease (PD)-like neuronal loss with aggregation of α-synuclein (α-syn). To reduce the adverse effects of ROT, its derivative, rotenoisin A (ROA), is obtained by directly irradiating a ROT solution in methanol using γ-rays, which has been reported for potential anticancer properties. However, its PD-inducing effects have not yet been researched or reported. This study sought to compare the activities of ROA and ROT on the aggregation of α-syn, apoptosis, and autophagy in SH-SY5Y cells. ROA decreased cell survival less when compared with ROT on SH-SY5Y cells at 48 h in a dose-dependent manner. ROT (0.5 and 1 μM) and ROA (4 and 5 μM) decreased the expression of tyrosine hydroxylase. Western blot analysis of the Triton X-100 insoluble fraction revealed that both ROT and ROA significantly increased the levels of oligomeric, dimeric, and monomeric phosphorylated Serine129 α-syn and total monomeric α-syn. Moreover, both compounds decreased the proportion of neuronal nuclei, the neurofilament-heavy chain, and β3-tubulin. The phosphorylation of ERK and SAPK were reduced, whereas ROA did not act on Akt. Additionally, the increased Bax/Bcl-2 ratio further activated the downstream caspases cascade. ROT promoted the LC3BII/I ratio and p62 levels; however, different ROA doses resulted in different effects on autophagy while inducing PD-like impairments in SH-SY5Y cells.

Effect of chlorpyrifos on cypermethrin-induced dopaminergic neurotoxicity in rats.

The study aimed to investigate the combined effects of chlorpyrifos and cypermethrin combined on dopaminergic neurotoxicity, motor behaviours and level of selected inflammatory proteins in rats compared to either alone for delineating an interaction between these two pesticides. The rotarod and grip strength tests were employed to assess neurobehavioural changes. The striatal dopamine content and expression of tyrosine hydroxylase (TH), α-synuclein, cyclooxygenase-2 (COX-2), and tumour necrosis factor-α (TNF-α) proteins in the nigrostriatal tissue were measured. Chlorpyrifos impaired the neurobehavioural indexes, reduced the striatal dopamine level, augmented the level of α-synuclein, COX-2, and TNF-α and attenuated the expression of TH similar to but a little less than cypermethrin. Half the dose of both pesticides together produced additional neurotoxicity compared with the usual (highest employed) dose of either alone. The results showed that chlorpyrifos induced moderately less dopaminergic neurotoxicity than cypermethrin. In the combination, they produced a little higher toxicity than either pesticide alone.

Moxibustion improves motor function by down-regulating SNX5 and inhibiting ferroptosis in neurons of corpus striatum in mice with Parkinson's disease.

To explore the effect of moxibustion on the expression of sorting nexin 5 (SNX5), glutathione peroxidase (GPX4) and ferritin heavy chain (FTH1) in the corpus striatum in mice with Parkinson's disease (PD), so as to explore its mechanisms underlying improvement of PD by ameliorating ferroptosis in the substantia nigra striatum. C57BL/6J mice were randomly divided into normal, sham operation, model, and moxibustion groups, with 10 mice in each group. The PD model was established by unilateral injection of 6-hydroxydopamine (3.5 μL) into the right medial forebrain bundle (AP=-1.2 mm, ML=-1.3 mm, DV=-4.75 mm). The mice in the moxibustion group received moxibustion at "Baihui"(GV20) and "Sishencong"(EX-HN1) for 20 min each time, once a day, 6 times a week for 4 weeks. After the intervention, mice received apomorphine rotation behavior detection and pole climbing test. The expression of tyrosine hydroxylase (TH) in the substantia nigra was detected by immunofluorescence, the contents of Fe2+, malondialdehyde (MDA), the ratio of glutathione/oxidized glutathione (GSH/GSSG) in the corpus striatum were detected by using photocolorimetric method, and the expression levels of SNX5 (endocytosomal protein), GPX4 (one of the key targets for inhibiting ferroptosis) and FTH1 proteins and mRNAs in the corpus striatum were detected by Western blot and qPCR, respectively. Behavior tests showed that the pole climbing time and number of body rotation were significantly increased in the model group relevant to the sham operation group (P<0.01), and strikingly decreased in the moxibustion group relevant to the model group (P<0.01). The immunofluorescence intensity of TH in the substantia nigra, the ratio of GSH/GSSG, and the expression levels of GPX4 and FTH1 mRNAs and proteins in the corpus striatum were markedly decreased (P<0.01, P<0.05), while the contents of Fe2+ and MDA and the expression levels of SNX5 mRNA and protein in the corpus striatum significantly increased in the model group relevant to the sham operation group (P<0.01, P<0.05). Compared with the model group, the decreased immunofluorescence intensity of TH, GSH/GSSH, and the expression levels of GPX4 and FTH1 mRNAs and proteins, and the increased contents of Fe2+ and MDA and the expression levels of SNX5 mRNA and protein were reversed in the moxibustion group relevant to the model group (P<0.01, P<0.05). Moxibustion may improve motor dysfunction in PD mice, which may be related to its effects in down-regulating the expression of SNX5, promoting the synthesis of GSH, decreasing the contents of Fe2+ and MDA, up-regulating the ratio of GSH/GSSG and the expression of GPX4 and FTH1 mRNAs and proteins in the corpus striatum, and inhibiting the occurrence of ferroptosis.

Basal release of 6-cyanodopamine from rat isolated vas deferens and its role on the tissue contractility.

6-Cyanodopamine is a novel catecholamine released from rabbit isolated heart. However, it is not known whether this catecholamine presents any biological activity. Here, it was evaluated whether 6-cyanodopamine (6-CYD) is released from rat vas deferens and its effect on this tissue contractility. Basal release of 6-CYD, 6-nitrodopamine (6-ND), 6-bromodopamine, 6-nitrodopa, and 6-nitroadrenaline from vas deferens were quantified by LC-MS/MS. Electric-field stimulation (EFS) and concentration-response curves to noradrenaline, adrenaline, and dopamine of the rat isolated epididymal vas deferens (RIEVD) were performed in the absence and presence of 6-CYD and /or 6-ND. Expression of tyrosine hydroxylase was assessed by immunohistochemistry. The rat isolated vas deferens released significant amounts of both 6-CYD and 6-ND. The voltage-gated sodium channel blocker tetrodotoxin had no effect on the release of 6-CYD, but it virtually abolished 6-ND release. 6-CYD alone exhibited a negligible RIEVD contractile activity; however, at 10 nM, 6-CYD significantly potentiated the noradrenaline- and EFS-induced RIEVD contractions, whereas at 10 and 100 nM, it also significantly potentiated the adrenaline- and dopamine-induced contractions. The potentiation of noradrenaline- and adrenaline-induced contractions by 6-CYD was unaffected by tetrodotoxin. Co-incubation of 6-CYD (100 pM) with 6-ND (10 pM) caused a significant leftward shift and increased the maximal contractile responses to noradrenaline, even in the presence of tetrodotoxin. Immunohistochemistry revealed the presence of tyrosine hydroxylase in both epithelial cell cytoplasm of the mucosae and nerve fibers of RIEVD. The identification of epithelium-derived 6-CYD and its remarkable synergism with catecholamines indicate that epithelial cells may regulate vas deferens smooth muscle contractility.