A Survey on Potentially Beneficial and Hazardous Bioactive Compounds in Cocoa Powder Samples Sourced from the European Market.

Cocoa (Theobroma cacao, L.) represents an important market that gained relevance and became an esteemed commodity thanks to cocoa powder, chocolate, and other related products. This work analyzed 59 cocoa powder samples from the European market. Three distinct subgroups were identified: organic or conventional, alkalized or not alkalized, and raw or roasted processing. The impact of the technological process on their pH, color, and compositional traits, as well as their content of biogenic amines and salsolinol, was evaluated. The phenolic fraction was also investigated through both common and emerging methods. The results depict that the influence of the agronomical practices (organic/conventional) did not significantly (p < 0.05) affect the composition of the cocoa powders; similarly, the roasting process was not a determinant of the compounds traced. On the other hand, the alkalinization process greatly impacted color and pH, no matter the cocoa's provenience or obtention or other processes, also resulting in reducing the phenolic fraction of the treated samples. Principal component analysis confirmed that the alkali process acts on pH, color, and phenolic composition but not on the content of other bioactive molecules (biogenic amines and salsolinol). All the samples were safe, while the alkalized powders saw a great reduction in beneficial biocompounds. A novel strategy could be to emphasize on the label whether cocoa powder is non-alkalized to meet the demand for more beneficial products.

Salsolinol improves angiotensin II­induced myocardial fibrosis in vitro via inhibition of LSD1 through regulation of the STAT3/Notch­1 signaling pathway.

The clinical incidence of congestive heart failure (CHF) is very high and it poses a significant threat to the health of patients. The traditional Chinese medicine monomer salsolinol is widely used to treat similar symptoms of CHF. However, there have been no reports on the effect of salsolinol for the management of CHF and its effects on myocardial fibrosis. In the present study, salsolinol was used to treat angiotensin II (AngII)-induced human cardiac fibroblasts (HCFs) and cell proliferation and migration were assessed using a CCK-8, EdU staining assay and wound healing assay. Subsequently, immunofluorescence, western blotting and other techniques were used to detect indicators associated with cell fibrosis and relevant kits were used to detect markers of cellular inflammation and reactive oxygen species (ROS) production. Molecular docking analysis was used to predict the relationship between salsolinol and lysine-specific histone demethylase 1A (LSD1). Subsequently, the expression of LSD1 in the serum of CHF patients was detected by reverse transcription-quantitative PCR. Finally, LSD1 was overexpressed in cells to explore the regulatory mechanism of salsolinol in AngII-induced HFCs. Salsolinol reduced the proliferation and migration. Salsolinol reduced the expression of fibrosis marker proteins α-smooth muscle actin, Collagen I and Collagen III in a concentration-dependent manner, thereby reducing cell fibrosis. In addition, salsolinol reduced the levels of TNF-α and IL-6 in the cell supernatant and ROS production following AngII induction. Salsolinol inhibited LSD1 expression and regulated the STAT3/Notch-1 signaling pathway. Upregulation of LSD1 reversed the effects of salsolinol on AngII-induced HCFs. Salsolinol inhibited LSD1 via regulation of the STAT3/Notch-1 signaling pathway to improve Ang II-induced myocardial fibrosis in vitro.

The rostromedial tegmental nucleus RMTg is not a critical site for ethanol-induced motor activation in rats.

RATIONALE: Opioid drugs indirectly activate dopamine (DA) neurons in the ventral tegmental area (VTA) through a disinhibition mechanism mediated by mu opioid receptors (MORs) present both on the GABA projection neurons located in the medial tegmental nucleus/tail of the VTA (RMTg/tVTA) and on the VTA GABA interneurons. It is well demonstrated that ethanol, like opioid drugs, provokes VTA DA neuron disinhibition by interacting (through its secondary metabolite, salsolinol) with MORs present in VTA GABA interneurons, but it is not known whether ethanol could disinhibit VTA DA neurons through the MORs present in the RMTg/tVTA. OBJECTIVES: The objective of the present study was to determine whether ethanol, directly microinjected into the tVTA/RMTg, is also able to induce VTA DA neurons disinhibition. METHODS: Disinhibition of VTA DA neurons was indirectly assessed through the analysis of the motor activity of rats. Cannulae were placed into the tVTA/RMTg to perform microinjections of DAMGO (0.13 nmol), ethanol (150 or 300 nmol) or acetaldehyde (250 nmol) in animals pre-treated with either aCSF or the irreversible antagonist of MORs, beta-funaltrexamine (beta-FNA; 2.5 nmol). After injections, spontaneous activity was monitored for 30 min. RESULTS: Neither ethanol nor acetaldehyde directly administered into the RMTg/tVTA were able to increase the locomotor activity of rats at doses that, in previous studies performed in the posterior VTA, were effective in increasing motor activities. However, microinjections of 0.13 nmol of DAMGO into the tVTA/RMTg significantly increased the locomotor activity of rats. These activating effects were reduced by local pre-treatment of rats with beta-FNA (2.5 nmol). CONCLUSIONS: The tVTA/RMTg does not appear to be a key brain region for the disinhibiting action of ethanol on VTA DA neurons. The absence of dopamine in the tVTA/RMTg extracellular medium, the lack of local ethanol metabolism or both could explain the present results.

Role of Metabolism on Alcohol Preference, Addiction, and Treatment.

Studies presented in this chapter show that: (1) in the brain, ethanol is metabolized by catalase to acetaldehyde, which condenses with dopamine forming salsolinol; (2) acetaldehyde-derived salsolinol increases the release of dopamine mediating, via opioid receptors, the reinforcing effects of ethanol during the acquisition of ethanol consumption, while (3) brain acetaldehyde does not influence the maintenance of chronic ethanol intake, it is suggested that a learned cue-induced hyperglutamatergic system takes precedence over the dopaminergic system. However, (4) following a prolonged ethanol deprivation, the generation of acetaldehyde in the brain again plays a role, contributing to the increase in ethanol intake observed during ethanol re-access, called the alcohol deprivation effect (ADE), a model of relapse behavior; (5) naltrexone inhibits the high ethanol intake seen in the ADE condition, suggesting that acetaldehyde-derived salsolinol via opioid receptors also contributes to the relapse-like drinking behavior. The reader is referred to glutamate-mediated mechanisms that trigger the cue-associated alcohol-seeking and that also contribute to triggering relapse.

Salsolinol Protects SH-SY5Y Cells Against MPP+ Damage and Increases Enteric S100-Immunoreactivity in Wistar Rats.

A dopamine derivative, 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, known as salsolinol (SAL), has increasingly gained attention since its first detection in the urine of Parkinson's disease patients treated with levodopa, and has been proposed as a possible neurotoxic contributor to the disease. Yet, so far, the neurobiological role of SAL remains unclear. Thus, the main aims of our study were to compare the neurotoxic potential of SAL with MPP+ (1-methyl-4-phenylpyridinium ion) in vitro, and to examine intestinal and metabolic alterations following intraperitoneal SAL administration in vivo. In vitro, SH-SY5Y neuroblastoma cell line was monitored following MPP+ and SAL treatment. In vivo, Wistar rats were subjected to SAL administration by either osmotic intraperitoneal mini-pumps or a single intraperitoneal injection, and after two weeks, biochemical and morphological parameters were assessed. SH-SY5Y cells treated with MPP+ (1000 µM) and SAL (50 µM) showed increase in cell viability and fluorescence intensity in comparison with the cells treated with MPP+ alone. In vivo, we predominantly observed decreased collagen content in the submucosal layer, decreased neuronal density with comparable ganglionic area in the jejunal myenteric plexus, and increased glial S100 expression in both enteric plexuses, yet with no obvious signs of inflammation. Besides, glucose and triglycerides levels were lower after single SAL-treatment (200 mg/kg), and low- to high-density lipoprotein (LDL/HDL) ratio and aspartate to alanine aminotransferases (AST/ALT) ratio levels were higher after continuous SAL-treatment (200 mg/kg in total over 2 weeks). Low doses of SAL were non-toxic and exhibited pronounced neuroprotective properties against MPP+ in SH-SY5Y cell line, which supports the use of SAL as a reference compound for in vitro studies. In vivo results give insight into our understanding of gastrointestinal remodeling following intraperitoneal SAL administration, and might represent morphological correlates of a microglial-related enteric neurodegeneration and dopaminergic dysregulation.

Salsolinol Induces Parkinson's Disease Through Activating NLRP3-Dependent Pyroptosis and the Neuroprotective Effect of Acteoside.

Endogenous neurotoxin 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroiso-quinoline (Salsolinol, SAL) is a dopamine metabolite that is toxic to dopaminergic neurons in vitro and in vivo, and is involved in the pathogenesis of Parkinson's disease (PD). However, the molecular mechanism by which SAL induces neurotoxicity in PD remains challenging for future investigations. This study found that SAL induced neurotoxicity in SH-SY5Y cells and mice. RNA sequencing (RNAseq) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were used to detect differentially expressed genes in SAL-treated SH-SY5Y cells. We found that NLR family pyrin domain-containing 3 (NLRP3)-dependent pyroptosis was enriched by SAL, which was validated by in vitro and in vivo SAL models. Further, NLRP3 inflammasome-related genes (ASC, NLRP3, active caspase 1, IL-1ß, and IL-18) were increased at the mRNA and protein level. Acteoside mitigates SAL-induced neurotoxicity by inhibiting NLRP3 inflammasome-related pyroptosis in in vitro and in vivo PD models. In summary, the present study suggests for the first time that NLRP3-dependent pyroptosis plays a role in the pathogenesis of SAL-induced PD, and acteoside mitigates SAL-induced pyroptosis-dependent neurotoxicity in in vitro and in vivo PD models. The present results demonstrated a new mechanism whereby SAL mediates neurotoxicity by activating NLRP3-dependent pyroptosis, further highlighting SAL-induced pyroptosis-dependent neurotoxicity as a potential therapeutic target in PD.

Sal synthase induced cytotoxicity of PC12 cells through production of the dopamine metabolites salsolinol and N-methyl-salsolinol.

As a catechol isoquinoline, salsolinol (Sal) is widely distributed in mammalian brains, and is increased in the cerebrospinal fluid (CSF) and urine of Parkinsonian patients. Sal can be metabolized to N-methyl-salsolinol (NM-Sal), an MPP+-like neurotoxin, and impairs the function of dopaminergic neurons, causing the clinical symptoms of Parkinson's disease (PD). Sal synthase, which catalyzes the production of Sal from dopamine and acetaldehyde, may be the important enzyme in the metabolism of catechol isoquinolines (CTIQs). Previously, our work demonstrated the existence of Sal synthase in rat brain and identified its amino acid sequence. However, the biological function of Sal synthase has not been thoroughly explored, especially its role in dopaminergic neuronal degeneration. In this study, we tried to clarify the catalytic role of Sal synthase in the formation of CTIQs which are endogenous neurotoxins in the mammalian brain. Furthermore, the cytotoxicity of Sal synthase was also observed in dopaminergic PC12 cells. The results demonstrated that Sal synthase overexpression can increase the level of Sal and NM-Sal, and ultimately cause mitochondria damage and apoptosis.

Suppressive effects of processed aconite root on dexamethasone-induced muscle ring finger protein-1 expression and its active ingredients.

Processed aconite root (PA), the tuberous root of Aconitum carmichaelii prepared by autoclaving, is a crude drug used in Japanese traditional Kampo medicine and traditional Chinese medicine for the symptoms of kidney deficiency, that is related to the muscle atrophy in modern medicine. The objective of the present study is to evaluate the effectiveness of PA on muscle atrophy and to find its active ingredients using dexamethasone-induced muscle ring finger protein-1 (MuRF1) mRNA expression in murine myoblast C2C12 cells. Dexamethasone-induced MuRF1 expression was significantly suppressed by methanol-soluble part of boiling water extract of PA in a concentration-dependent manner with its IC50 value of 1.5 mg/ml. By the activity-guided fractionations of PA extract using the partition between organic solvents and its aqueous solution, the activity of PA did not transfer into the fraction containing aconitine-type diterpenoid alkaloids but into BuOH layer. Then, we found higenamine and salsolinol as the active ingredients in PA. Higenamine and salsolinol significantly suppressed dexamethasone-induced MuRF1 expression, and their IC50 values were 0.49 and 50 µM, respectively. The contents of higenamine and salsolinol in the decoctions of commercially available fourteen PA products are 0.12 and 14 µg/ml as the average values, and varied with the coefficient of variation (CV) values of 97 and 63%, respectively. Higenamine also significantly suppressed dexamethasone-induced mRNA expressions of muscle atrophy F-box protein (MAFbx)/atrogin1, casitas B-lineage lymphoma-b (Cbl-b), troponin, branched-chain amino acid aminotransferase 2 (BCAT2), and Bcl-2 binding and pro-apoptotic protein3 (Bnip3). Although the quality control of PA is regulated by the contents of diterpene alkaloids, salsolinol and higenamine can be used as the marker compounds to certificate the pharmacological activities of PA.

Behavioral consequences of the downstream products of ethanol metabolism involved in alcohol use disorder.

Research concerning Alcohol Use Disorder (AUD) has previously focused primarily on either the behavioral or chemical consequences experienced following ethanol intake, but these areas of research have rarely been considered in tandem. Compared with other drugs of abuse, ethanol has been shown to have a unique metabolic pathway once it enters the body, which leads to the formation of downstream metabolites which can go on to form biologically active products. These metabolites can mediate a variety of behavioral responses that are commonly observed with AUD, such as ethanol intake, reinforcement, and vulnerability to relapse. The following review considers the preclinical and chemical research implicating these downstream products in AUD and proposes a chemobehavioral model of AUD.

Ethanol-Dependent Synthesis of Salsolinol in the Posterior Ventral Tegmental Area as Key Mechanism of Ethanol's Action on Mesolimbic Dopamine.

Abnormal consumption of ethanol, the ingredient responsible for alcoholic drinks' addictive liability, causes millions of deaths yearly. Ethanol's addictive potential is triggered through activation, by a still unknown mechanism, of the mesolimbic dopamine (DA) system, part of a key motivation circuit, DA neurons in the posterior ventral tegmental area (pVTA) projecting to the ipsilateral nucleus accumbens shell (AcbSh). The present in vivo brain microdialysis study, in dually-implanted rats with one probe in the pVTA and another in the ipsilateral or contralateral AcbSh, demonstrates this mechanism. As a consequence of the oral administration of a pharmacologically relevant dose of ethanol, we simultaneously detect a) in the pVTA, a substance, 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (salsolinol), untraceable under control conditions, product of condensation between DA and ethanol's first by-product, acetaldehyde; and b) in the AcbSh, a significant increase of DA release. Moreover, such newly generated salsolinol in the pVTA is responsible for increasing AcbSh DA release via µ opioid receptor (µOR) stimulation. In fact, inhibition of salsolinol's generation in the pVTA or blockade of pVTA µORs prevents ethanol-increased ipsilateral, but not contralateral, AcbSh DA release. This evidence discloses the long-sought key mechanism of ethanol's addictive potential and suggests the grounds for developing preventive and therapeutic strategies against abnormal consumption.

Novel pentacyclic triterpenes exhibiting strong neuroprotective activity in SH-SY5Y cells in salsolinol- and glutamate-induced neurodegeneration models.

Novel triterpene derivatives were prepared and evaluated in salsolinol (SAL)- and glutamate (Glu)-induced models of neurodegeneration in neuron-like SH-SY5Y cells. Among the tested compounds, betulin triazole 4 bearing a tetraacetyl-ß-d-glucose substituent showed a highly potent neuroprotective effect. Further studies revealed that removal of tetraacetyl-ß-d-glucose part (free triazole derivative 10) resulted in strong neuroprotection in the SAL model at 1 µM, but this derivative suffered from cytotoxicity at higher concentrations. Both compounds modulated oxidative stress and caspase-3,7 activity, but 10 showed a superior effect comparable to the Ac-DEVD-CHO inhibitor. Interestingly, while both 4 and 10 outperformed the positive controls in blocking mitochondrial permeability transition pore opening, only 4 demonstrated potent restoration of the mitochondrial membrane potential (MMP) in the model. Derivatives 4 and 10 also showed neuroprotection in the Glu model, with 10 exhibiting the strongest oxidative stress reducing effect among the tested compounds, while the neuroprotective activity of 4 was probably due recovery of the MMP.

Cytokinin Plant Hormones Have Neuroprotective Activity in In Vitro Models of Parkinson's Disease.

Cytokinins are adenine-based phytohormones that regulate key processes in plants, such as cell division and differentiation, root and shoot growth, apical dominance, branching, and seed germination. In preliminary studies, they have also shown protective activities against human neurodegenerative diseases. To extend knowledge of the protection (protective activity) they offer, we investigated activities of natural cytokinins against salsolinol (SAL)-induced toxicity (a Parkinson's disease model) and glutamate (Glu)-induced death of neuron-like dopaminergic SH-SY5Y cells. We found that kinetin-3-glucoside, cis-zeatin riboside, and N6-isopentenyladenosine were active in the SAL-induced PD model. In addition, trans-, cis-zeatin, and kinetin along with the iron chelator deferoxamine (DFO) and the necroptosis inhibitor necrostatin 1 (NEC-1) significantly reduced cell death rates in the Glu-induced model. Lactate dehydrogenase assays revealed that the cytokinins provided lower neuroprotective activity than DFO and NEC-1. Moreover, they reduced apoptotic caspase-3/7 activities less strongly than DFO. However, the cytokinins had very similar effects to DFO and NEC-1 on superoxide radical production. Overall, they showed protective activity in the SAL-induced model of parkinsonian neuronal cell death and Glu-induced model of oxidative damage mainly by reduction of oxidative stress.

In Vitro Protective Action of Monomeric and Fibrillar α-Synuclein on Neuronal Cells Exposed to the Dopaminergic Toxins Salsolinol and DOPAL.

The aggregation of α-synuclein (aSyn) is believed to be mechanistically linked to the degeneration of dopamine (DA)-producing neurons in Parkinson's disease (PD). In this respect, one crucial question that yet remains unsolved is whether aSyn aggregation is associated with either a gain- or loss-of-function of the protein in neuronal cells. Herein, we investigated the effect of monomeric versus fibrillar aSyn on mesencephalic dopaminergic neurons in primary cultures challenged with the neurotoxic catechols: salsolinol (SALSO; 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline) and 3,4-dihydroxyphenylacetaldehyde (DOPAL). aSyn monomer protected cells against either SALSO- or DOPAL-induced toxicity via inhibition of caspase-3-mediated apoptosis. While fibrillar aSyn failed in attenuating SALSO neurotoxicity, it increased the viability of DOPAL-treated cells, which was apparently not associated with the inhibition of caspase-3 cleavage. The fact that DOPAL-derived aSyn adducts exhibit lower toxicity compared with DOPAL itself raises the question of whether the generation of these adducts could be part of or a collateral effect of aSyn-mediated protection in neurons exposed to DOPAL. Overall, our work provides important evidence on the impact of the fibrillation of aSyn on its protective role in neuronal cells exposed to the toxic catechols SALSO and DOPAL.

The Mechanism of Action of Salsolinol in Brain: Implications in Parkinson's Disease.

1-Methyl-1,2,3,4-tetrahydroisoquinoline-6,7-diol, commonly known as salsolinol, is a compound derived from dopamine. It was first discovered in 1973 and has gained attention for its role in Parkinson's disease. Salsolinol and its derivatives were claimed to play a role in the pathogenesis of Parkinson's disease as a neurotoxin that induces apoptosis of dopaminergic neurons due to its structural similarity to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its ability to induce Parkinsonism. In this article, we discussed the biosynthesis, distribution and blood-brain barrier permeability of salsolinol. The roles of salsolinol in a healthy brain, particularly the interactions with enzymes, hormone and catecholamine, were reviewed. Finally, we discussed the involvement of salsolinol and its derivatives in the pathogenesis of Parkinson's disease.

Synergistic Neuroprotective Effect of Endogenously-Produced Hydroxytyrosol and Synaptic Vesicle Proteins on Pheochromocytoma Cell Line Against Salsolinol.

Oxidative stress triggers a lethal cascade, leading to Parkinson's disease by causing degeneration of dopaminergic neurons. In this study, eight antioxidants were screened for their neuroprotective effect on PC12 cells (pheochromocytoma cell line) under oxidative stress induced by salsolinol (OSibS). Hydroxytyrosol was found to be the strongest neuroprotective agent; it improved viability of PC12 cells by up to 81.69% under OSibS. Afterward, two synaptic vesicle proteins, synapsin-1 and septin-5, were screened for their neuroprotective role; the overexpression of synapsin-1 and the downregulation of septin-5 separately improved the viability of PC12 cells by up to 71.17% and 67.00%, respectively, compared to PC12 cells only treated with salsolinol (PoTwS) under OSibS. Subsequently, the PC12+syn++sep- cell line was constructed and pretreated with 100 µM hydroxytyrosol, which improved its cell viability by up to 99.03% and led to 14.71- and 6.37-fold reductions in the levels of MDA and H2O2, respectively, and 6.8-, 12.97-, 10.57-, and 7.57-fold increases in the activity of catalase, glutathione reductase, superoxide dismutase, and glutathione peroxidase, respectively, compared to PoTwS under OSibS. Finally, alcohol dehydrogenase-6 from Saccharomyces cerevisiae was expressed in PC12+syn++sep- cells to convert 3,4-dihydroxyphenylacetaldehyde (an endogenous neurotoxin) into hydroxytyrosol. The PC12+syn++sep-+ADH6+ cell line also led to 22.38- and 12.33-fold decreases in the production of MDA and H2O2, respectively, and 7.15-, 13.93-, 12.08-, and 8.11-fold improvements in the activity of catalase, glutathione reductase, superoxide dismutase, and glutathione peroxidase, respectively, compared to PoTwS under OSibS. Herein, we report the endogenous production of a powerful antioxidant, hydroxytyrosol, from 3,4-dihydroxyphenylacetaldehyde, and evaluate its synergistic neuroprotective effect, along with synapsin-1 and septin-5, on PC12 cells under OSibS.

Salsolinol-neurotoxic or Neuroprotective?

Salsolinol (6,7-dihydroxy-1-methyl-1,2,3,4-tetrahydroisoquinoline), widely available in many edibles, is considered to alter the function of dopaminergic neurons in the central nervous system and thus, multiple hypotheses on its either physiological and/or pathophysiological role have emerged. The aim of our work was to revisit its potentially neurotoxic and/or neuroprotective role through a series of both in vitro and in vivo experiments. Salsolinol in the concentration range 10-250 µM did not show any significant release of lactate dehydrogenase from necrotic SH-SY5Y cells and was able in the concentration of 50 and 100 µM to rescue SH-SY5Y cells from death induced by H2O2. Its neuroprotective effect against neurotoxin 6-hydroxydopamine was also determined. Salsolinol was found to decrease significantly the reactive oxygen species level in SH-SY5Y cells treated by 500 µM H2O2 and the caspase activity induced by 300 µM of H2O2 or 100 µM of 6-hydroxydopamine. Serum levels of TNFα and CRP of salsolinol-treated rats were not significantly different from control animals. Both TNFα and CRP served as indirect markers of neurotoxicity and/or neuroprotection. Although the neurotoxic properties of salsolinol have numerously been emphasized, its neuroprotective properties should not be neglected and need greater consideration.

Transcriptome analysis of Aconitum carmichaelii and exploration of the salsolinol biosynthetic pathway.

Aconitum carmichaelii has been used in traditional Chinese medicine for treating various diseases for several thousand years. Based on the biosynthetic pathway of some alkaloids such as C19-diterpenoid alkaloids and obvious differences in alkaloid content between leaves of two A. carmichaelii varieties has been reported, we performed leaves transcriptome analysis of two A. carmichaelii varieties. Besides we characterized the biosynthetic pathway of salsolinol. A total of 56 million raw reads (8.28 G) and 55 million clean reads (8.24 G) were obtained from two varieties (Z175 and R184) leaves transcriptome, respectively, and 176,793 unigenes were annotated. 281 and 843 unigenes are involved in the salsolinol biosynthetic pathway and the formation of C19-diterpenoid alkaloids respectively. And including 34 and 24 unigenes are the differentially expressed genes (DEGs) in the biosynthesis pathway for C19-diterpenoid alkaloids and salsolinol between Z175 and R184 respectively, which were target genes to explore differences in C19-diterpenoid alkaloid and salsolinol biosynthesis in Z175 and R184. Thus genes involved in alkaloid biosynthesis and accumulation differ between varieties leaves. The mechanisms underlying the differences and their relevance require further exploration. The results expand our knowledge of alkaloids biosynthesis in A. carmichaelii leaves, and provide a theoretical basis for analysis differences in alkaloids biosynthesis patterns in different varieties.

Salsolinol Attenuates Doxorubicin-Induced Chronic Heart Failure in Rats and Improves Mitochondrial Function in H9c2 Cardiomyocytes.

Backgrounds: Salsolinol (SAL), a plant-based isoquinoline alkaloid, was initially isolated from Aconiti Lateralis Radix Praeparata (ALRP) and identified as the active cardiotonic component of ALRP. This study was aimed to explore the therapeutic effect and mechanism by which SAL attenuates doxorubicin (DOX)-induced chronic heart failure (CHF) in rats and improves mitochondrial function in H9c2 cardiomyocytes. Methods: Rats were intraperitoneally injected with DOX to establish CHF model. Therapeutic effects of SAL on hemodynamic parameters, serum indices, and the histopathology of the heart were analyzed in vivo. Moreover, H9c2 cardiomyocytes were pretreated with SAL for 2 h before DOX treatment in all procedures in vitro. Cell viability, cardiomyocyte morphology, proliferation, and mitochondrial function were detected by a high-content screening (HCS) assay. In addition, a Seahorse Extracellular Flux (XFp) analyzer was used to evaluate the cell energy respiratory and energy metabolism function. To further investigate the potential mechanism of SAL, relative mRNA and protein expression of key enzymes in the tricarboxylic acid cycle in vivo and mitochondrial calcium uniporter (MCU) signaling pathway-related molecules in vitro were detected. Results: The present data demonstrated the pharmacological effect of SAL on DOX-induced CHF, which was through ameliorating heart function, downregulating serum levels of myocardial injury markers, alleviating histological injury to the heart, increasing the relative mRNA expression levels of key enzymes downstream of the tricarboxylic acid cycle in vivo, and thus enhancing myocardial energy metabolism. In addition, SAL had effects on increasing cell viability, ameliorating DOX-induced mitochondrial dysfunction, and increasing mitochondrial oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) in H9c2 cardiomyocyte. Moreover, we found that SAL might have an effect on improving mitochondrial respiratory function and energy metabolism via inhibiting excessive activation of MCU pathway in H9c2 cells. However, the protective effect could be ameliorated by ruthenium red (an MCU inhibitor) and abrogated by spermine (an MCU activator) in vitro. Conclusion: The therapeutic effects of SAL on CHF are possibly related to ameliorating cardiomyocyte function resulting in promotion of mitochondrial respiratory and energy metabolism. Furthermore, the potential mechanism might be related to downregulating MCU pathway. These findings may provide a potential therapy for CHF.

Stimulatory effect of dopamine derivative, salsolinol, on pulsatile luteinizing hormone secretion in seasonally anestrous sheep: Focus on dopamine, kisspeptin and gonadotropin-releasing hormone.

In the present study, there was testing of the hypothesis that a centrally administered dopamine (DA) derivative, salsolinol, could affect pulsatile luteinizing hormone (LH) secretion in seasonally anestrous sheep by affecting the neuronal components of the estradiol (E2) negative feedback. In two experiments performed during early spring (increasing day length - March/April), salsolinol or Ringer-Locke solution (control) were administered into the third brain ventricle (IIIv): 1) in several injections for three consecutive days; and 2) in several hour-long infusions. In addition to determining the LH concentration (in both experiments), the abundances of gonadotropin-releasing hormone (GnRH) and kisspeptin mRNA were examined in the hypothalamus and LHß subunit mRNA in the pituitary (Experiment 1). In Experiment 2, concentrations of DA and 3,4-dihydroxyphenylacetic acid (DOPAC) were determined in perfusates collected from the infundibular nucleus/median eminence (IN/ME) by the push-pull method. In both experiments, salsolinol increased both LH pulse frequency (P < 0.05) and plasma LH concentration (P < 0.001) compared to controls. The injected salsolinol also increased (P < 0.05) the abundance of GnRH mRNA in the mediobasal hypothalamus and kisspeptin mRNA in the arcuate nucleus. The two doses of infused salsolinol decreased DA to undetectable concentrations and DOPAC concentration by 60% in perfusates collected from the IN/ME. In conclusion, exogenous salsolinol functioning centrally stimulates pulsatile LH secretion in sheep during seasonal anestrus. It is suggested that salsolinol may have this effect by reducing the activity of the hypothalamic neuroendocrine dopaminergic system, which results in an increase in both kisspeptin and GnRH neurons activity.

Discovery and Validation of Banana Intake Biomarkers Using Untargeted Metabolomics in Human Intervention and Cross-sectional Studies.

BACKGROUND: Banana is one of the most widely consumed fruits in the world. However, information regarding its health effects is scarce. Biomarkers of banana intake would allow a more accurate assessment of its consumption in nutrition studies. OBJECTIVES: Using an untargeted metabolomics approach, we aimed to identify the banana-derived metabolites present in urine after consumption, including new candidate biomarkers of banana intake. METHODS: A randomized controlled study with a crossover design was performed on 12 healthy subjects (6 men, 6 women, mean ± SD age: 30.0 ± 4.9 y; mean ± SD BMI: 22.5 ± 2.3 kg/m2). Subjects underwent 2 dietary interventions: 1) 250 mL control drink (Fresubin 2 kcal fiber, neutral flavor; Fresenius Kabi), and 2) 240 g banana + 150 mL control drink. Twenty-four-hour urine samples were collected and analyzed with ultra-performance liquid chromatography coupled to a quadrupole time-of-flight MS and 2-dimensional GC-MS. The discovered biomarkers were confirmed in a cross-sectional study [KarMeN (Karlsruhe Metabolomics and Nutrition study)] in which 78 subjects (mean BMI: 22.8; mean age: 47 y) were selected reflecting high intake (126-378 g/d), low intake (47.3-94.5 g/d), and nonconsumption of banana. The confirmed biomarkers were examined singly or in combinations, for established criteria of validation for biomarkers of food intake. RESULTS: We identified 33 potentially bioactive banana metabolites, of which 5 metabolites, methoxyeugenol glucuronide (MEUG-GLUC), dopamine sulfate (DOP-S), salsolinol sulfate, xanthurenic acid, and 6-hydroxy-1-methyl-1,2,3,4-tetrahydro-ß-carboline sulfate, were confirmed as candidate intake biomarkers. We demonstrated that the combination of MEUG-GLUC and DOP-S performed best in predicting banana intake in high (AUCtest = 0.92) and low (AUCtest = 0.87) consumers. The new biomarkers met key criteria establishing their current applicability in nutrition and health research for assessing the occurrence of banana intake. CONCLUSIONS: Our metabolomics study in healthy men and women revealed new putative bioactive metabolites of banana and a combined biomarker of intake. These findings will help to better decipher the health effects of banana in future focused studies. This study was registered at clinicaltrials.gov as NCT03581955 and with the Ethical Committee for the Protection of Human Subjects Sud-Est 6 as CPP AU 1251, IDRCB 2016-A0013-48; the KarMeN study was registered with the German Clinical Trials Register (DRKS00004890). Details about the study can be obtained from https://www.drks.de.