Downregulation Of Tyrosine Hydroxylase Research Articles

Adrenal G protein-coupled Receptor Kinase (GRK)-2 Downregulation And Regulator of G protein Signaling (RGS)-4 Upregulation Mediate The Sympatholytic Effect Of Dapagliflozin

Dapagliflozin is a sodium/­glucose co-­transporter (SGLT)-­2 inhibitor with beneficial cardiovascular effects independent of its anti­diabetic actions. Among these is sympatholysis, i.e., norepinephrine (NE) lowering, beneficial in chronic human heart failure (HF). Adrenal G protein­-coupled receptor kinase (GRK)-­2 upregulation is a major driver of circulating catecholamine (CA) elevation and sympathetic nervous system (SNS) hyperactivity in HF due to dysregulation of adrenal sympatho-­inhibitory α2­-adrenergic receptors (ARs). On the other hand, we recently reported that Regulator of G protein Signaling (RGS)-­4 inhibits free fatty acid receptor (FFAR)-­3-stimulated NE release from sympathetic neurons, while it is also known to inhibit adrenal CA secretion induced by acetylcholine. Thus, we hypothesized herein that dapagliflozin may lower SNS hyperactivity by enhancing adrenal α2AR and RGS4 functions. We used the rat pheochromocytoma PC12 cell line expressing human α2A­AR, as well as freshly isolated adrenal glands from rats treated with dapagliflozin in vivo. Dapagliflozin treatment for 7 consecutive days (20 mg/kg/d in drinking water) led to a significant reduction in blood circulating NE levels (217±67 pg/ml, n=6), compared to vehicle-­treated rats (363±77 pg/ml, n=6, p<.05), suggesting reduced SNS activity. This was accompanied by reduced GRK2 and tyrosine hydroxylase (TH) mRNA and protein levels in dapagliflozin-­treated vs. vehicle­-treated adrenal glands. In contrast, RGS4 mRNA and protein levels were increased by dapagliflozin ­treatment. Adrenal α2AR density was higher in dapagliflozin- ­vs. vehicle-­treated rats (51.3±7.3 vs. 26.1±8.1 fmol/mg of protein, respectively; n=12 glands from 6 animals per group, p<.05). These results (i.e., GRK2 & TH downregulation with RGS4 upregulation) were completely recapitulated in PC12 cells in culture, treated with 5 μM dapagliflozin (or vehicle) for 24 hours. Importantly, α2AR signaling towards inhibition of CA secretion was markedly enhanced, while the cholinergic agonist carbachol-induced CA secretion was reduced post-­dapagliflozin treatment of PC12 cells. GRK2 overexpression and RGS4 siRNA-mediated knockdown partially reversed dapagliflozin‘s effect on CA secretion, confirming the involvement of GRK2 and RGS4 in dapagliflozin‘s sympatholytic effects in PC12 cells. In conclusion, dapagliflozin exerts adrenal sympatholysis via: a) downregulation of TH, which reduces CA biosynthesis, and GRK2, which reduces α2AR desensitization (i.e., enhances α2AR sympatho-inhibitory signaling); and b) upregulation of RGS4, which directly opposes cholinergic-stimulated CA secretion. 1) NIH/NHLBI #HL155718-01. 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.

Parvalbumin interneuron deficiency in the prefrontal and motor cortices of spontaneously hypertensive rats: an attention-deficit hyperactivity disorder animal model insight.

Attention deficit hyperactivity disorder (ADHD) is characterized by impairments in developmental-behavioral inhibition, resulting in impulsivity and hyperactivity. Recent research has underscored cortical inhibition deficiencies in ADHD via the gamma-aminobutyric acid (GABA)ergic system, which is crucial for maintaining excitatory-inhibitory balance in the brain. This study explored postnatal changes in parvalbumin (PV) immunoreactivity, indicating GABAergic interneuron types, in the prefrontal (PFC) and motor (MC) cortices of spontaneously hypertensive rats (SHRs), an ADHD animal model. Examining PV- positive (PV+) cells associated with dopamine D2 receptors (D2) and the impact of dopamine on GABA synthesis, we also investigated changes in the immunoreactivity of D2 and tyrosine hydroxylase (TH). Brain sections from 4- to 10-week-old SHRs and Wistar Kyoto rats (WKYs) were immunohistochemically analyzed, comparing PV+, D2+ cells, and TH+ fiber densities across age-matched SHRs and WKYs in specific PFC/MC regions. The results revealed significantly reduced PV+ cell density in SHRs: prelimbic (~20% less), anterior cingulate (~15% less), primary (~15% less), and secondary motor (~17% less) cortices. PV+ deficits coincided with the upregulation of D2 in prepubertal SHRs and the downregulation of TH predominantly in pubertal/postpubertal SHRs. Reduced PV+ cells in various PFC regions could contribute to inattention/behavioral alterations in ADHD, while MC deficits could manifest as motor hyperactivity. D2 upregulation and TH deficits may impact GABA synthesis, exacerbating behavioral deficits in ADHD. These findings not only shed new light on ADHD pathophysiology but also pave the way for future research endeavors.

Effects of latroeggtoxin-VI on dopamine and α-synuclein in PC12 cells and the implications for Parkinson's disease.

Parkinson's disease (PD) is characterized by death of dopaminergic neurons leading to dopamine deficiency, excessive α-synuclein facilitating Lewy body formation, etc. Latroeggtoxin-VI (LETX-VI), a proteinaceous neurotoxin discovered from the eggs of spider L. tredecimguttatus, was previously found to promote the synthesis and release of PC12 cells, showing a great potential as a drug candidate for PD. However, the relevant mechanisms have not been understood completely. The present study explored the mechanism underlying the effects of LETX-VI on dopamine and α-synuclein of PC12 cells and the implications for PD. After PC12 cells were treated with LETX-VI, the level of dopamine was significantly increased in a dose-dependent way within a certain range of concentrations. Further mechanism analysis showed that LETX-VI upregulated the expression of tyrosine hydroxylase (TH) and L-dopa decarboxylase to enhance the biosynthesis of dopamine, and downregulated that of monoamine oxidase B to reduce the degradation of dopamine. At the same time, LETX-VI promoted the transport and release of dopamine through modulating the abundance and/or posttranslational modification of vesicular monoamine transporter 2 (VMAT2) and dopamine transporter (DAT). While the level of dopamine was increased by LETX-VI treatment, α-synuclein content was reduced by the spider toxin. α-Synuclein overexpression significantly decreased the dopamine level and LETX-VI efficiently alleviated the inhibitory action of excessive α-synuclein on dopamine. In the MPTP-induced mouse model of PD, application of LETX-VI ameliorated parkinsonian behaviors of the mice, and reduced the magnitude of MPTP-induced α-synuclein upregulation and TH downregulation. In addition, LETX-VI displayed neuroprotective effects by inhibiting MPTP-induced decrease in the numbers of TH-positive and Nissl-stained neurons in mouse brain tissues. All the results demonstrate that LETX-VI promotes the synthesis and release of dopamine in PC12 cells via multiple mechanisms including preventing abnormal α-synuclein accumulation, showing implications in the prevention and treatment of PD.

Dapagliflozin downregulates G protein-coupled receptor Kinase-2 and upregulates regulator of g protein signaling-4 in adrenals to exert sympatholysis in heart failure

Abstract Dapagliflozin is a sodium-glucose co-transporter (SGLT)-2 inhibitor with beneficial cardiovascular effects independent of its anti-diabetic actions. Among these is sympatholysis, i.e., norepinephrine (NE) lowering which is beneficial in chronic human heart failure (HF). Adrenal G protein-coupled receptor kinase (GRK)-2 upregulation is a major driver of circulating catecholamine (CA) elevation and sympathetic nervous system (SNS) hyperactivity in HF due to dysregulation of adrenal sympatho-inhibitory α2-adrenergic receptors (ARs) [1]. On the other hand, we recently reported that signaling by neuronal free fatty acid receptor (FFAR)-3, which stimulates NE release from sympathetic neurons, is blocked by Regulator of G protein Signaling (RGS)-4 [2,3]. We hypothesized herein that dapagliflozin may lower SNS hyperactivity in the adrenal gland by improving α2AR, and antagonizing FFAR3 signaling in adrenal chromaffin cells. We used the rat pheochromocytoma PC12 cell line expressing human α2A-AR, as well as freshly isolated adrenal glands from rats treated with dapagliflozin in vivo. Dapagliflozin treatment for 7 consecutive days (20 mg/kg/d in drinking water) led to a significant reduction in blood circulating NE levels (217+67 pg/ml, n=6), compared to control, vehicle-treated rats (363+77 pg/ml, n=6, p<.05), suggesting reduced SNS activity. This was accompanied by reduced GRK2 and tyrosine hydroxylase (TH) mRNA and protein levels in dapagliflozin-treated rat adrenals vs. vehicle-treated animal-derived glands. In contrast, RGS4 mRNA levels were increased in dapagliflozin-treated adrenals. Adrenal α2AR density was higher in dapagliflozin- vs. vehicle-treated rats (51.3+7.3 vs. 26.1+8.1 fmol/mg of protein, respectively; n=12 glands from 6 animals per group, p<.05). These results (i.e., GRK2 & TH downregulation with RGS4 upregulation) were completely recapitulated in PC12 cells in culture, treated with 5 microM dapagliflozin (or vehicle) for 24 hours. Importantly, α2AR-dependent G protein-mediated signaling towards inhibition of CA secretion was markedly enhanced, whereas FFAR3-dependent phospholipase C/calcium signaling was reduced, at 24 hrs post-dapagliflozin treatment of PC12 cells. GRK2 overexpression partially reversed dapagliflozin`s effect on cholinergic-induced CA secretion. In conclusion, dapagliflozin exerts a sympatholytic action in the adrenal medulla via a) downregulation of TH, which reduces CA biosynthesis, and GRK2, which reduces α2AR desensitization; and b) upregulation of RGS4 to block FFAR3-stimulated CA secretion (Figure).Figure

FTO-targeted siRNA delivery by MSC-derived exosomes synergistically alleviates dopaminergic neuronal death in Parkinson's disease via m6A-dependent regulation of ATM mRNA

BackgroundParkinson's disease (PD), characterized by the progressive loss of dopaminergic neurons in the substantia nigra and striatum of brain, seriously threatens human health, and is still lack of effective treatment. Dysregulation of N6-methyladenosine (m6A) modification has been implicated in PD pathogenesis. However, how m6A modification regulates dopaminergic neuronal death in PD remains elusive. Mesenchymal stem cell-derived exosomes (MSC-Exo) have been shown to be effective for treating central nervous disorders. We thus propose that the m6A demethylase FTO-targeted siRNAs (si-FTO) may be encapsulated in MSC-Exo (Exo-siFTO) as a synergistic therapy against dopaminergic neuronal death in PD.MethodsIn this study, the effect of m6A demethylase FTO on dopaminergic neuronal death was evaluated both in vivo and in vitro using a MPTP-treated mice model and a MPP + -induced MN9D cellular model, respectively. The mechanism through which FTO influences dopaminergic neuronal death in PD was investigated with qRT-PCR, western blot, immumohistochemical staining, immunofluorescent staining and flow cytometry. The therapeutic roles of MSC-Exo containing si-FTO were examined in PD models in vivo and in vitro.ResultsThe total m6A level was significantly decreased and FTO expression was increased in PD models in vivo and in vitro. FTO was found to promote the expression of cellular death-related factor ataxia telangiectasia mutated (ATM) via m6A-dependent stabilization of ATM mRNA in dopaminergic neurons. Knockdown of FTO by si-FTO concomitantly suppressed upregulation of α-Synuclein (α-Syn) and downregulation of tyrosine hydroxylase (TH), and alleviated neuronal death in PD models. Moreover, MSC-Exo were utilized to successfully deliver si-FTO to the striatum of animal brain, resulting in the significant suppression of α-Syn expression and dopaminergic neuronal death, and recovery of TH expression in the brain of PD mice.ConclusionsMSC-Exo delivery of si-FTO synergistically alleviates dopaminergic neuronal death in PD via m6A-dependent regulation of ATM mRNA.

Neuroprotective effect of quercetin nanoparticles: A possible prophylactic effect in cerebellar neurodegenerative disorders

Memory deficit, anxiety, coordination deficit and depression are common neurological disorders attributed to aluminum (Al) buildup in the nervous system. Quercetin nanoparticles (QNPs) are a newly developed effective neuroprotectant. We aimed to investigate the potential protective and therapeutic effects of QNPs in Al induced toxicity in rat cerebellum. A rat model of Al-induced cerebellar damage was created by AlCl3 (100 mg/kg) administration orally for 42 days. QNPs (30 mg/kg) was administered for 42-days as a prophylactic (along with AlCl3 administration) or therapeutic for 42-days (following AlCl3 induced cerebellar damage). Cerebellar tissues were assessed for structural and molecular changes. The results showed that Al induced profound cerebellar structural and molecular changes, including neuronal damage, astrogliosis and tyrosine hydroxylase downregulation. Prophylactic QNPs significantly reduced Al induced cerebellar neuronal degeneration. QNPs is a promising neuroprotectant that can be used in elderly and vulnerable subjects to protect against neurological deterioration. It could be a promising new line for therapeutic intervention in neurodegenerative diseases.

The role of tyrosine hydroxylase within dapoxetine-assisted therapy against premature ejaculation.

There are several investigations that have revealed that cerebral dopamine (DA) plays a pivotal role in the occurrence of premature ejaculation (PE). Although tyrosine hydroxylase (TH) is an essential enzyme for the synthesis of DA, only few investigations have described the role of TH in regulation mechanisms for ejaculation till now. To investigate whether there is a correlation between TH expression level in the brain and different ejaculation behavior in rats. Then explore whether the TH expression in the brain will change after acute dapoxetine treatment in rats with Rapid ejaculation. Rats (male, S-D rats, 6-8 weeks) were separated into three groups based on their ejaculation frequency: Rapid, Normal, and Sluggish. Expression level of DA in the brain was determined by enzyme-linked immune sorbent assay (ELISA) kit, TH expression level in the brain was determined by immunohistochemistry and Western Blot (WB) techniques. Among the three groups, DA and TH expression level were the highest in the Rapid ejaculation group, while the lowest was the Sluggish ejaculation group. The results also showed that TH level was positively associated with ejaculation frequency (r = 0.8038, P < 0.001) and negatively associated with ejaculation latency (r=-0.6199, P = 0.018). Furthermore, acute dapoxetine therapy in rats with Rapid ejaculation downregulated TH level in the brain. Changes in ejaculation behavior were significantly linked with TH level. Upregulated TH in selected brain regions related with ejaculation could cause rapid ejaculation. The effect of dapoxetine in prolonging ejaculation could be related to TH downregulation within the brain.

Abstract 11977: Dapagliflozin Downregulates Adrenal G Protein-Coupled Receptor-Kinase-2 to Exert Sympatholysis in Heart Failure

Introduction: Dapagliflozin is a sodium-glucose co-transporter (SGLT)-2 inhibitor exerting several cardiorenal protective effects independently of blood glucose regulation. Adrenal G protein-coupled receptor kinase (GRK)-2 upregulation is a major driver of circulating catecholamine (CA) elevation and sympathetic nervous system (SNS) hyperactivity, especially in heart failure (HF). GRK2 severely dysregulates adrenal sympatho-inhibitory α 2 -adrenergic receptors (ARs), leading to unchecked, chronically elevated CA secretion. Hypothesis: SGLT2 inhibition with dapagliflozin lowers SNS hyperactivity in part by opposing adrenal GRK2 actions. Methods: We used the rat pheochromocytoma PC12 cell line expressing human α 2A AR, as well as freshly isolated adrenal glands from rats treated with dapagliflozin in vivo. Results: Dapagliflozin treatment for 7 consecutive days (20 mg/kg/d in drinking water) led to a significant reduction in blood circulating NE levels (217+67 pg/ml, n=6), compared to control, vehicle-treated rats (363+77 pg/ml, n=6, p&lt;.05), suggesting reduced SNS activity. This was accompanied by reduced GRK2 and tyrosine hydroxylase (TH) mRNA and protein levels in dapagliflozin-treated rat adrenals vs. vehicle-treated animal-derived glands, indicating reduced adrenal CA synthesis and secretion. Finally, adrenal α 2 AR density was higher in dapagliflozin- vs. vehicle-treated rats(51.3+7.3 vs. 26.1+8.1 fmol/mg of protein, respectively; n=12 glands from 6 animals per group, p&lt;.05). GRK2 and TH downregulation were also observed in PC12 cells treated with 5 μM dapagliflozin for 24 hours. Importantly, α 2 AR-dependent inhibition of CA secretion was markedly enhanced 24 hrs post-dapagliflozin application. This was due to reduced GRK2-dependent receptor desensitization induced by dapagliflozin, since this effect was abolished in PC12 cells overexpressing GRK2. Conclusions: Dapagliflozin exerts a sympatholytic action in the adrenal medulla via downregulation of both TH, thus reducing CA biosynthesis, and GRK2, thus reducing α 2 AR desensitization to lower CA secretion. This adrenal-mediated sympatholysis may be part of the increasingly apparent cardiovascular benefits of SGLT2 inhibitors in human HF irrespective of diabetic status.

Association of Hepatitis C Virus Replication with the Catecholamine Biosynthetic Pathway.

A bidirectional negative relationship between Hepatitis C virus (HCV) replication and gene expression of the catecholamine biosynthetic enzyme L-Dopa decarboxylase (DDC) was previously shown in the liver and attributed at least to an association of DDC with phosphatidylinositol 3-kinase (PI3K). Here, we report that the biosynthesis and uptake of catecholamines restrict HCV replication in hepatocytes, while HCV has developed ways to reduce catecholamine production. By employing gene silencing, chemical inhibition or induction of the catecholamine biosynthetic and metabolic enzymes and transporters, and by applying the substrates or the products of the respective enzymes, we unravel the role of the different steps of the pathway in viral infection. We also provide evidence that the effect of catecholamines on HCV is strongly related with oxidative stress that is generated by their autoxidation in the cytosol, while antioxidants or treatments that lower cytosolic catecholamine levels positively affect the virus. To counteract the effect of catecholamines, HCV, apart from the already reported effects on DDC, causes the down-regulation of tyrosine hydroxylase that encodes the rate-limiting enzyme of catecholamine biosynthesis and suppresses dopamine beta-hydroxylase mRNA and protein amounts, while increasing the catecholamine degradation enzyme monoamine oxidase. Moreover, the NS4B viral protein is implicated in the effect of HCV on the ratio of the ~50 kDa DDC monomer and a ~120 kDa DDC complex, while the NS5A protein has a negative effect on total DDC protein levels.

Dapagliflozin Exerts Adrenal Sympatholysis via G protein-Coupled Receptor Kinase-2 &amp; Tyrosine Hydroxylase Downregulation

Abstract The sodium-glucose co-transporter (SGLT)-2 inhibitor dapagliflozin was recently reported to reduce renal tyrosine hydroxylase (TH) and norepinephrine levels, lowering blood pressure and preventing endothelial dysfunction, in a murine model of neurogenic hypertension. This suggested that dapagliflozin may combat sympathetic nervous system (SNS) hyperactivity, which is known to accompany and aggravate chronic heart failure (CHF). Adrenal G protein-coupled receptor kinase (GRK)-2 upregulation is a major driver of circulating catecholamine (CA) elevation and SNS hyperactivity, especially in CHF. GRK2 severely dysregulates adrenal sympatho-inhibitory α 2-adrenergic receptors (ARs), leading to unchecked, chronically elevated CA secretion. Therefore, we hypothesized herein that SGLT2 inhibition with dapagliflozin may lower SNS hyperactivity in the adrenal gland by antagonizing GRK2 actions on α 2ARs in adrenal chromaffin cells. We used the rat pheochromocytoma PC12 cell line expressing human α 2AAR, as well as freshly isolated adrenal glands from adult male Sprague-Dawley rats treated with dapagliflozin in vivo. We measured circulating norepinephrine (NE) in vivo via RIA, GRK2 &amp; TH expression levels via real-time PCR and immunoblotting, adrenal α 2AR density (Bmax) via saturation radioligand binding with [methyl-3H]-rauwolscine, and G protein activation via the GTPγS assay. Dapagliflozin treatment for 7 consecutive days (20 mg/kg/d in drinking water) led to a significant reduction in blood circulating NE levels (217+67 pg/ml, n=6), compared to control, vehicle-treated rats (363+77 pg/ml, n=6, p&amp;lt;.05), suggesting reduced SNS activity. This was accompanied by reduced GRK2 and TH mRNA and protein levels in dapagliflozin-treated rat adrenals vs. vehicle-treated animal-derived glands, indicating reduced adrenal CA synthesis and secretion. Finally, adrenal α 2AR density was higher in dapagliflozin- vs. vehicle-treated rats (51.3+7.3 vs. 26.1+8.1 fmol/mg of protein, respectively; n=12 glands from 6 animals per group, p&amp;lt;.05). These results (i.e. GRK2 and TH downregulation) were completely recapitulated in PC12 α 2AAR-expressing cells in culture, treated with 5 μM dapagliflozin (or vehicle) for 24 hours. Importantly, α 2AR-dependent G protein-mediated signaling towards inhibition of CA secretion was markedly enhanced at 24 hrs post-dapagliflozin application in PC12 cells. This was the result of reduced GRK2-dependent receptor desensitization, since dapagliflozin lacked this effect in cells co-transfected with a GRK2-encoding adenovirus to acutely overexpress GRK2. In conclusion, dapagliflozin exerts a sympatholytic action in the adrenal medulla via downregulation of both TH, which reduces CA biosynthesis, and GRK2, which reduces α 2AR desensitization in favor of enhanced inhibition of CA secretion.

Neuronal degeneration and oxidative stress in the SNc of 6-OHDA intoxicated rats; improving role of silymarin long-term treatment.

Progressive loss in dopaminergic neurons (DA) of substantia nigra pars compacta (SNc) leads to Parkinson's disease with a hypothesis of oxidative stress generation. The present study was conducted to determine the long-term efficacy of silymarin (SM) post-treatment on 6-OHDA-induced oxidative stress in the SNc of male rats. Male Wistar rats were received 6-OHDA (8μg/rat) into SNc. After 3weeks, as recovery period, the animals were treated with i.p. injection of SM at different doses of 100, 200, or 300mg/kg for 15days. At the end of the treatment, motor function, neuronal cell count, antioxidant enzymes, and lipid peroxidation and tyrosine hydroxylase (TH) activities were evaluated in the ventral midbrain tissue. The 6-OHDA significantly decreased (p ≤ 0.05) motor function, antioxidant enzyme activity, GSH level, and GSH/GSSG ratio and caused an augmentation in GSSG and lipid peroxidation level. The 6-OHDA also reduced the population of neurons and TH expression. The SM repaired the 6-OHDA-induced motor impairment, antioxidant enzyme suppression, and TH down-regulation. All three doses of SM could restore the MDA level to the normal range in the 6-OHDA-lesioned rats and could reversed the effect of 6-OHDA on GSH, GSSG level, and GSH/GSSG ratio. The SM treatment significantly and dose-dependently increased (p ≤ 0.001) the total number of surviving neurons in the SNc. Silymarin chronic treatment restored the brain's antioxidant capacity and salvaged neurons from oxidative stress-induced neurodegeneration. The SM could also improve motor function in parkinsonian animals by increasing TH expression. These results recommend that application of SM over initial clinical stages may depict a hopeful approach versus PD. However, more research is needed to confirm this issue.

Neuroprotective effect of quercetin nanoparticles: A possible prophylactic and therapeutic role in alzheimer’s disease

BackgroundAlzheimer’s disease (AD) is the most common cause of dementia in elderly. Quercetin is a well-known flavonoid with low bioavailability. Recently, quercetin nanoparticles (QNPs) has been shown to have a better bioavailability. AimsThis study aimed to investigate the protective and therapeutic effects of QNPs in Aluminum chloride (AlCl3) induced animal model of AD. Materials and MethodsAD was induced in rats by oral administration of AlCl3 (100 mg/kg/day) for 42 days. QNPs (30 mg/kg) was given along with AlCl3 in the prophylactic group and following AD induction in the treated group. Hippocampi were harvested for assessments of the structural and ultrastructural changes using histological and histochemical approaches. Results and DiscussionAD hippocampi showed a prominent structural and ultrastructural disorders both neuronal and extraneuronal. Including neuronal degeneration, formation of APs and NFTs, downregulation of tyrosine hydroxylase (TH), astrogliosis and inhibition of the proliferative activity (all P ≤ 0.05). Electron microscopy showed signs of neuronal degeneration with microglia and astrocyte activation and disruption of myelination and Blood Brain Barrier (BBB). Interestingly, QNPs administration remarkably reduced the neuronal degenerative changes, APs and NFTs formation (all P ≤ 0.05). Furthermore, it showed signs of regeneration (all P ≤ 0.05) and upregulation of TH. The effect was profound in the prophylactic group. Thus, QNPs reduced the damaging effect of AlCl3 on hippocampal neurons at the molecular, cellular and subcellular levels. ConclusionFor the best of our knowledge this is the first study to show a prophylactic and therapeutic effect for QNPs in AD model. This might open the gate for further research and provide a new line for therapeutic intervention in AD.

Downregulation of tyrosine hydroxylase phenotype after AAV injection above substantia nigra: Caution in experimental models of Parkinson's disease.

Adeno-associated virus (AAV) vector-mediated delivery of human α-synuclein (α-syn) gene in rat substantia nigra (SN) results in increased expression of α-syn protein in the SN and striatum which can progressively degenerate dopaminergic neurons. Therefore, this model is thought to recapitulate the neurodegeneration in Parkinson's disease. Here, using AAV to deliver α-syn above the SN in male and female rats resulted in clear expression of human α-syn in the SN and striatum. The protein was associated with moderate behavioral deficits and some loss of tyrosine hydroxylase (TH) in the nigrostriatal areas. However, the immunohistochemistry results were highly variable and showed little to no correlation with behavior and the amount of α-syn present. Expression of green fluorescent protein (GFP) was used as a control to monitor gene delivery and expression efficacy. AAV-GFP resulted in a similar or greater TH loss compared to AAV-α-syn and therefore an additional vector that does not express a protein was tested. Vectors with double-floxed inverse open reading frame (DIO ORF) encoding fluorescent proteins that generate RNA that is not translated also resulted in TH downregulation in the SN but showed no significant behavioral deficits. These results demonstrate that although expression of wild-type human α-syn can cause neurodegeneration, the variability and lack of correlation with outcome measures are drawbacks with the model. Furthermore, design and control selection should be considered carefully because of conflicting conclusions due to AAV downregulation of TH, and we recommend caution with having highly regulated TH as the only marker for the dopamine system.

Tyrosine hydroxylase down-regulation after loss of Abelson helper integration site 1 (AHI1) promotes depression via the circadian clock pathway in mice

Abelson helper integration site 1 (AHI1) is associated with several neuropsychiatric and brain developmental disorders, such as schizophrenia, depression, autism, and Joubert syndrome. Ahi1 deficiency in mice leads to behaviors typical of depression. However, the mechanisms by which AHI1 regulates behavior remain to be elucidated. Here, we found that down-regulation of expression of the rate-limiting enzyme in dopamine biosynthesis, tyrosine hydroxylase (TH), in the midbrains of Ahi1-knockout (KO) mice is responsible for Ahi1-deficiency-mediated depressive symptoms. We also found that Rev-Erbα, a TH transcriptional repressor and circadian regulator, is up-regulated in the Ahi1-KO mouse midbrains and Ahi1-knockdown Neuro-2a cells. Moreover, brain and muscle Arnt-like protein 1 (BMAL1), the Rev-Erbα transcriptional regulator, is also increased in the Ahi1-KO mouse midbrains and Ahi1-knockdown cells. Our results further revealed that AHI1 decreases BMAL1/Rev-Erbα expression by interacting with and repressing retinoic acid receptor-related orphan receptor α, a nuclear receptor and transcriptional regulator of circadian genes. Of note, Bmal1 deficiency reversed the reduction in TH expression induced by Ahi1 deficiency. Moreover, microinfusion of the Rev-Erbα inhibitor SR8278 into the ventral midbrain of Ahi1-KO mice significantly increased TH expression in the ventral tegmental area and improved their depressive symptoms. These findings provide a mechanistic explanation for a link between AHI1-related behaviors and the circadian clock pathway, indicating an involvement of circadian regulatory proteins in AHI1-regulated mood and behavior.

Reduced TH expression and α-synuclein accumulation contribute towards nigrostriatal dysfunction in experimental hepatic encephalopathy.

The present work examines α-synuclein expression in the nigrostriatal system of a rat chronic hepatic encephalopathy model induced by portacaval anastomosis (PCA). There is evidence that dopaminergic dysfunction in disease conditions is strongly associated with such expression. Possible relationships among dopaminergic neurons, astroglial cells and α-synuclein expression were sought. Brain tissue samples from rats at 1 and 6 months post-PCA, and controls, were analysed immunohistochemically using antibodies against tyrosine hydroxylase (TH), α-synuclein, glial fibrillary acidic protein (GFAP) and ubiquitin (Ub). In the control rats, TH immunoreactivity was detected in the neuronal cell bodies and processes in the substantia nigra pars compacta (SNc). A dense TH-positive network of neurons was also seen in the striatum. In the PCA-exposed rats, however, a reduction in TH-positive neurons was seen at both 1 and 6 months in the SNc, as well as a reduction in TH-positive fibres in the striatum. This was coincident with the appearance of α-synuclein-immunoreactive neurons in the SNc; some of the TH-positive neurons also showed α-synuclein immunoreactivity. In addition, α-synuclein accumulation was seen in the SNc and striatum at both 1 and 6 months post-PCA, whereas α-synuclein was only mildly expressed in the nigrostriatal pathway of the controls. Astrogliosis was also seen following PCA, as revealed by increased GFAP expression from 1 month to 6 months post-PCA in both the SN and striatum. The astroglial activation level in the SN paralleled the reduced neuronal expression of TH throughout PCA exposure. α-synuclein accumulation following PCA may induce dopaminergic dysfunction via the downregulation of TH, as well as astroglial activation.

Impact of low-dose chronic exposure to Bisphenol A (BPA) on adult male zebrafish adaption to the environmental complexity: Disturbing the color preference patterns and reliving the anxiety behavior

The extensive usage of xenobiotic endocrine disrupting chemicals (XEDCs), such as Bisphenol A (BPA), has created obvious threat to aquatic ecosystems worldwide. Although a comprehensive understanding of the adverse effect of BPA on behaviors and physiology have been proven, the potential impact of low-dose BPA on altering the basic ability of aquatic organism in adapting to the surrounded complex environment still remains elusive. In this research, we report that treatment of adult male zebrafish with chronic (7 weeks) low-dose (0.22 nM–2.2 nM) BPA, altered the ability in adapting the complex environment by disturbing the natural color preference patterns. In addition, chronic 50 ng/L (0.22 nM) BPA exposure alleviated the anxiety behavior of male zebrafish confronted with the novel environment by enhancing the preference towards light in the light/dark preference test. This phenotype was associated with less expression of serotonin (5-TH) in the hypothalamus and the down-regulation of tyrosine hydroxylase (TH) in brain tissues. As such, our results show that low-dose BPA remnant in surface waters altered zebrafish behavior that are known to have ecological and evolutionary consequences. HighlightsHere we reported that the impact of chronic low-dose BPA exposure on the basic capability of zebrafish to adapt to the environmental complexity. Specifically, BPA at low concentration, under the environmental safety level and 3000-fold lower than the accepted human daily exposure, interfered with the ability to discriminate color and alleviate anxiety induced by the novel environment, which finally altered the capability of male zebrafish to adapt to the environmental complexity. These findings revealed the ecological effect of low-dose BPA and regular BPA concentration standard are not necessarily safe. The result also provided the consideration of retuning the hazard concentration level of BPA.

Parkinsonian features in aging GFAP.HMOX1 transgenic mice overexpressing human HO-1 in the astroglial compartment

Epigenetic influences mediating brain iron deposition, oxidative mitochondrial injury, and macroautophagy in Parkinson disease and related conditions remain enigmatic. Here, we show that selective overexpression of the stress protein, heme oxygenase-1 (HO-1) in astrocytes of GFAP.HMOX1 transgenic mice between 8.5 and 19 months of age results in nigrostriatal hypodopaminergia associated with locomotor incoordination and stereotypy; downregulation of tyrosine hydroxylase, DAT, LMX1B, Nurr1, Pitx3 and DJ-1 mRNA and/or protein; overproduction of α-synuclein and ubiquitin; oxidative stress; basal ganglia siderosis; mitochondrial damage/mitophagy; and augmented GABAergic systems (increased GABA, GAD67 and reelin). The neurophenotype of these GFAP.HMOX18.5-19m mice is highly consistent with parkinsonism and differs dramatically from the schizophrenia-like features previously documented in younger GFAP.HMOX10-12m mice. Common stressors may elicit either early-onset developmental (schizophrenia) or later-life degenerative (PD) brain disorders depending on whether the glial HO-1 response is engaged prior to or following the maturation of dopaminergic circuitry. Curtailment of glial HO-1 transduction at strategic points of the life course may confer neuroprotection in human degenerative and developmental central nervous system disorders.

From the Cover: Prenatal Nicotinic Exposure Attenuates Respiratory Chemoreflexes Associated With Downregulation of Tyrosine Hydroxylase and Neurokinin 1 Receptor in Rat Pup Carotid Body.

Maternal cigarette smoke is the major risk of sudden infant death syndrome (SIDS). A depressed ventilatory response to hypoxia (HVR) and hypercapnia (HCVR) is thought to be responsible for the pathogenesis of SIDS and the carotid body is critically involved in these responses. We have recently reported that prenatal nicotinic exposure (PNE) over the full gestation induces depressed HVR in rat pups. Here, we asked whether PNE (1) depressed not only HVR but also HCVR that were dependent on the carotid body, (2) affected some important receptors and neurochemicals expressed in the carotid body, such as tyrosine hydroxylase (TH), neurokinin-1 receptor (NK1R), and α7 nicotinic acetylcholine receptor (α7nAChR), and (3) blunted the ventilatory responses to activation of these receptors. To this end, HVR and HCVR in Ctrl and PNE pups were measured with plethysmography before and after carotid body ablation (Series I), mRNA expression and/or immunoreactivity (IR) of TH, NK1R, and α7nAChR in the carotid body were examined by RT-PCR and immunohistochemistry (Series II), and the ventilatory responses were tested before and after intracarotid injection of substance P (NK1R agonist) and AR-R17779 (α7nAChR agonist) (Series III). Our results showed that PNE (1) significantly depressed both HVR and HCVR and these depressions were abolished by carotid body ablation, (2) reduced the relative population of glomus cells, mRNA NK1R, and α7nAChR and IR of NK1R and TH in the carotid body, and (3) decreased ventilatory responses to intracarotid injection of substance P or AR-R17779. These results suggest that PNE acting via the carotid body could strikingly blunt HVR and HCVR, likely through downregulating TH and NK1R.

A regulatable AAV vector mediating GDNF biological effects at clinically-approved sub-antimicrobial doxycycline doses

Preclinical and clinical data stress the importance of pharmacologically-controlling glial cell line-derived neurotrophic factor (GDNF) intracerebral administration to treat PD. The main challenge is finding a combination of a genetic switch and a drug which, when administered at a clinically-approved dose, reaches the brain in sufficient amounts to induce a therapeutic effect. We describe a highly-sensitive doxycycline-inducible adeno-associated virus (AAV) vector. This vector allowed for the first time a longitudinal analysis of inducible transgene expression in the brain using bioluminescence imaging. To evaluate the dose range of GDNF biological activity, the inducible AAV vector (8.0 × 109 viral genomes) was injected in the rat striatum at four delivery sites and increasing doxycycline doses administered orally. ERK/Akt signaling activation as well as tyrosine hydroxylase downregulation, a consequence of long-term GDNF treatment, were induced at plasmatic doxycycline concentrations of 140 and 320 ng/ml respectively, which are known not to increase antibiotic-resistant microorganisms in patients. In these conditions, GDNF covered the majority of the striatum. No behavioral abnormalities or weight loss were observed. Motor asymmetry resulting from unilateral GDNF treatment only appeared with a 2.5-fold higher vector and a 13-fold higher inducer doses. Our data suggest that using the herein-described inducible AAV vector, biological effects of GDNF can be obtained in response to sub-antimicrobial doxycycline doses.

Transcription factors FOXA1 and FOXA2 maintain dopaminergic neuronal properties and control feeding behavior in adult mice

Midbrain dopaminergic (mDA) neurons are implicated in cognitive functions, neuropsychiatric disorders, and pathological conditions; hence understanding genes regulating their homeostasis has medical relevance. Transcription factors FOXA1 and FOXA2 (FOXA1/2) are key determinants of mDA neuronal identity during development, but their roles in adult mDA neurons are unknown. We used a conditional knockout strategy to specifically ablate FOXA1/2 in mDA neurons of adult mice. We show that deletion of Foxa1/2 results in down-regulation of tyrosine hydroxylase, the rate-limiting enzyme of dopamine (DA) biosynthesis, specifically in dopaminergic neurons of the substantia nigra pars compacta (SNc). In addition, DA synthesis and striatal DA transmission were reduced after Foxa1/2 deletion. Furthermore, the burst-firing activity characteristic of SNc mDA neurons was drastically reduced in the absence of FOXA1/2. These molecular and functional alterations lead to a severe feeding deficit in adult Foxa1/2 mutant mice, independently of motor control, which could be rescued by L-DOPA treatment. FOXA1/2 therefore control the maintenance of molecular and physiological properties of SNc mDA neurons and impact on feeding behavior in adult mice.