[Effects of periodontitis on bone and tryptophan metabolism of gut microbiota in estrogen-deficient mice].

Objective: To study the effects of periodontitis on bone and tryptophan metabolism of gut microbiota in the context of estrogen deficiency. Methods: Thirty-two female C57BL6/J mice were randomly divided into four groups based on table of random numbers (n=8 in each group): Sham group, in which mice were given sham surgery; Sham_Lig group, in which mice were given sham surgery and were induced to periodontitis by ligating the bilateral maxillary second molars with 5-0 silk threads at the fourth week; Ovx group, in which mice were given bilateral ovariectomy; Ovx_Lig group, in which mice were given bilateral ovariectomy and were induced to periodontitis at the fourth week. After 8 weeks of ligation, the mice of 4 groups were euthanized for collecting the samples of femur, tibia, mandible and skull. Those samples were scanned by micro-CT to measure the bone mineral density (BMD), bone volume versus total volume ratio (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th) and trabecular spacing (Tb.Sp). The cecum contents of 4 groups of mice were collected for gut microbiota 16S rRNA gene sequencing. The tryptophan and its metabolites in intestinal tracts were detected by liquid chromatography-mass spectrometry. Pearson correlation analysis was performed to analyze the correlation between the abundance of gut microbiota and the content of tryptophan and its metabolites. Results: Femur BMD [(82.23±3.97) mg/cm3], BV/TV [(9.25±1.37)%] and Tb.Th [(70.95±5.70) µm] in Ovx_Lig group were significantly lower than Ovx group [(96.30±3.76) mg/cm3 (P=0.004); (14.45±1.55)% (P=0.022) and (87.58±8.02) µm (P<0.001), respectively]. The ß-diversity analysis of gut microbiota based on Bray-Curtis distance showed that samples of Ovx_Lig group and Ovx group were obviously grouped. Linear discriminant analysis effect size (LEfSe) showed that Alistipes was the representative genus in Ovx_Lig group. The relative abundance of Alistipes in Ovx_Lig group [(0.42±0.14)%] were significantly higher than that in Ovx group [(0.17±0.05)%] (t=4.45, P<0.001). Tryptophan metabolism analysis showed that the content of kynurenic acid [(531.12±158.60) ng/g] in Ovx_Lig group were significantly higher than that in Ovx group [(400.42±57.96) ng/g] (t=2.19, P=0.046). And the content of indole-3-carbaldehyde [(383.37±144.06) ng/g] in Ovx_Lig group were significantly lower than Ovx group [(701.72±141.93) ng/g] (t=4.45, P<0.001). Correlation analysis showed that relative abundance of Alistipes was positively correlated with kynurenic acid (r=0.32, P=0.088), while negatively correlated with indole-3-carbaldehyde (r=-0.32, P=0.088). Conclusions: Periodontitis can induce bone destruction of femur in estrogen-deficient mice, the mechanism of which may be related to Alistipes in gut and the tryptophan metabolites kynurenic acid and indole-3-carbaldehyde.

Integrative Pharmacology in the Treatment of Substance Use Disorders.

The detrimental physical, mental, and socioeconomic effects of substance use disorders (SUDs) have been apparent to the medical community for decades. However, it has become increasingly urgent in recent years to develop novel pharmacotherapies to treat SUDs. Currently, practitioners typically rely on monotherapy. Monotherapy has been shown to be superior to no treatment at all for most substance classes. However, many randomized controlled trials (RCTs) have revealed that monotherapy leads to poorer outcomes when compared with combination treatment in all specialties of medicine. The results of RCTs suggest that monotherapy frequently fails since multiple dysregulated pathways, enzymes, neurotransmitters, and receptors are involved in the pathophysiology of SUDs. As such, research is urgently needed to determine how various neurobiological mechanisms can be targeted by novel combination treatments to create increasingly specific yet exceedingly comprehensive approaches to SUD treatment. This article aims to review the neurobiology that integrates many pathophysiologic mechanisms and discuss integrative pharmacology developments that may ultimately improve clinical outcomes for patients with SUDs. Many neurobiological mechanisms are known to be involved in SUDs including dopaminergic, nicotinic, N-methyl-D-aspartate (NMDA), and kynurenic acid (KYNA) mechanisms. Emerging evidence indicates that KYNA, a tryptophan metabolite, modulates all these major pathophysiologic mechanisms. Therefore, achieving KYNA homeostasis by harmonizing integrative pathophysiology and pharmacology could prove to be a better therapeutic approach for SUDs. We propose KYNA-NMDA-α7nAChRcentric pathophysiology, the "conductor of the orchestra," as a novel approach to treat many SUDs concurrently. KYNA-NMDA-α7nAChR pathophysiology may be the "command center" of neuropsychiatry. To date, extant RCTs have shown equivocal findings across comparison conditions, possibly because investigators targeted single pathophysiologic mechanisms, hit wrong targets in underlying pathophysiologic mechanisms, and tested inadequate monotherapy treatment. We provide examples of potential combination treatments that simultaneously target multiple pathophysiologic mechanisms in addition to KYNA. Kynurenine pathway metabolism demonstrates the greatest potential as a target for neuropsychiatric diseases. The investigational medications with the most evidence include memantine, galantamine, and N-acetylcysteine. Future RCTs are warranted with novel combination treatments for SUDs. Multicenter RCTs with integrative pharmacology offer a promising, potentially fruitful avenue to develop novel therapeutics for the treatment of SUDs.

Inactivation of the paraventricular nucleus attenuates the cardiogenic sympathetic afferent reflex in the spontaneously hypertensive rat.

BACKGROUND: Myocardial ischemia causes the release of bradykinin, which stimulates cardiac afferents, causing sympathetic excitation and chest pain. Glutamatergic activation of the paraventricular hypothalamic nucleus (PVN) in the spontaneously hypertensive rat (SHR) drives elevated basal sympathetic activity. Thus, we tested the hypothesis that inactivation of the PVN attenuates the elevated reflex response to epicardial bradykinin in the SHR and that ionotropic PVN glutamate receptors mediate the elevated reflex. METHODS: We recorded the arterial pressure and renal sympathetic nerve activity (RSNA) response to epicardial bradykinin application in anesthetized SHR and Wistar Kyoto (WKY) rats before and after PVN microinjection of GABA A agonist muscimol or ionotropic glutamate receptor antagonist kynurenic acid. RESULTS: Muscimol significantly decreased the arterial pressure response to bradykinin from 180.4 ±â€Š5.8 to 119.5 ±â€Š6.9 mmHg in the SHR and from 111.8 ±â€Š7.0 to 84.2 ±â€Š8.3 mmHg in the WKY and the RSNA response from 186.2 ±â€Š7.1 to 142.7 ±â€Š7.3% of baseline in the SHR and from 201.0 ±â€Š11.5 to 160.2 ±â€Š9.3% of baseline in the WKY. Kynurenic acid significantly decreased the arterial pressure response in the SHR from 164.5 ±â€Š5.0 to 126.2 ±â€Š7.7 mmHg and the RSNA response from 189.9 ±â€Š13.7to 168.5 ±â€Š12.7% of baseline but had no effect in the WKY. CONCLUSION: These results suggest that tonic PVN activity is critical for the full manifestation of the CSAR in both the WKY and SHR. Glutamatergic PVN activity contributes to the augmented CSAR observed in the SHR.

The steroid hormone ADIOL promotes learning by reducing neural kynurenic acid levels.

Reductions in brain kynurenic acid levels, a neuroinhibitory metabolite, improve cognitive function in diverse organisms. Thus, modulation of kynurenic acid levels is thought to have therapeutic potential in a range of brain disorders. Here we report that the steroid 5-androstene 3ß, 17ß-diol (ADIOL) reduces kynurenic acid levels and promotes associative learning in Caenorhabditis elegans We identify the molecular mechanisms through which ADIOL links peripheral metabolic pathways to neural mechanisms of learning capacity. Moreover, we show that in aged animals, which normally experience rapid cognitive decline, ADIOL improves learning capacity. The molecular mechanisms that underlie the biosynthesis of ADIOL as well as those through which it promotes kynurenic acid reduction are conserved in mammals. Thus, rather than a minor intermediate in the production of sex steroids, ADIOL is an endogenous hormone that potently regulates learning capacity by causing reductions in neural kynurenic acid levels.

Effect of probenecid on blood levels and renal elimination of furosemide and endogenous compounds in rats: Discovery of putative organic anion transporter biomarkers.

Transporter-mediated drug-drug interactions (DDIs) are assessed using probe drugs and in vitro and in vivo models during drug development. The utility of endogenous metabolites as transporter biomarkers is emerging for prediction of DDIs during early phases of clinical trials. Endogenous metabolites such as pyridoxic acid and kynurenic acid have shown potential to predict DDIs mediated by organic anion transporters (OAT1 and OAT3). However, these metabolites have not been assessed in rats as potential transporter biomarkers. We carried out a rat pharmacokinetic DDI study using probenecid and furosemide as OAT inhibitor and substrate, respectively. Probenecid administration led to a 3.8-fold increase in the blood concentrations and a 3-fold decrease in renal clearance of furosemide. High inter-individual and intra-day variability in pyridoxic acid and kynurenic acid, and no or moderate effect of probenecid administration on these metabolites suggest their limited utility for prediction of Oat-mediated DDI in rats. Therefore, rat blood and urine samples were further analysed using untargeted metabolomics. Twenty-one m/z features (out of >8000 detected features) were identified as putative biomarkers of rat Oat1 and Oat3 using a robust biomarker qualification approach. These m/z features belong to metabolic pathways such as fatty acid analogues, peptides, prostaglandin analogues, bile acid derivatives, flavonoids, phytoconstituents, and steroids, and can be used as a panel to decrease variability caused by processes other than Oats. When validated, these putative biomarkers will be useful in predicting DDIs caused by Oats in rats.

Kynurenic acid inhibits macrophage pyroptosis by suppressing ROS production via activation of the NRF2 pathway.

Macrophage pyroptosis and related inflammatory responses play an important role in periodontitis. Kynurenic acid (KA) is hypothesized to have anti­inflammatory potential, but whether KA can inhibit macrophage pyroptosis and the underlying mechanisms remain unclear. Lipopolysaccharide (LPS) was used to induce pyroptosis in THP­1­derived macrophages. KA or ML385 was used to pretreat macrophages, after which, cell viability, NOD­like receptor protein 3 (NLRP3) inflammasome­related protein expression, oxidative stress levels and nuclear factor erythroid 2­related factor 2 (NRF2) expression were measured. The results showed that KA improved the LPS­induced decrease in macrophage viability and lactate dehydrogenase release. KA prevented THP­1 macrophage pyroptosis induced by LPS by reducing the expression of NLRP3, Gasdermin­D, and Caspase1, and decreased the expression of inflammatory factors. KA suppressed NLRP3 inflammasome activation by inhibiting ROS overproduction and increasing Heme Oxygenase 1 and glutathione levels. Moreover, KA promoted NRF2 translocation from the cytoplasm to the nucleus. In addition, the anti­pyroptotic and antioxidant effects of KA were reversed by ML385 inhibition of NRF2. In the present study, it was found that KA significantly suppressed macrophage pyroptosis induced by LPS. It was further demonstrated that the anti­pyroptotic effects of KA were mediated by activation of the NRF2 pathway.

Kynurenic acid, a key L-tryptophan-derived metabolite, protects the heart from an ischemic damage.

BACKGROUND: Renal injury induces major changes in plasma and cardiac metabolites. Using a small- animal in vivo model, we sought to identify a key metabolite whose levels are significantly modified following an acute kidney injury (AKI) and to analyze whether this agent could offer cardiac protection once an ischemic event has occurred. METHODS AND RESULTS: Metabolomics profiling of cardiac lysates and plasma samples derived from rats that underwent AKI 1 or 7 days earlier by 5/6 nephrectomy versus sham-operated controls was performed. We detected 26 differential metabolites in both heart and plasma samples at the two selected time points, relative to sham. Out of which, kynurenic acid (kynurenate, KYNA) seemed most relevant. Interestingly, KYNA given at 10 mM concentration significantly rescued the viability of H9C2 cardiac myoblast cells grown under anoxic conditions and largely increased their mitochondrial content and activity as determined by flow cytometry and cell staining with MitoTracker dyes. Moreover, KYNA diluted in the drinking water of animals induced with an acute myocardial infarction, highly enhanced their cardiac recovery according to echocardiography and histopathology. CONCLUSION: KYNA may represent a key metabolite absorbed by the heart following AKI as part of a compensatory mechanism aiming at preserving the cardiac function. KYNA preserves the in vitro myocyte viability following exposure to anoxia in a mechanism that is mediated, at least in part, by protection of the cardiac mitochondria. A short-term administration of KYNA may be highly beneficial in the treatment of the acute phase of kidney disease in order to attenuate progression to reno-cardiac syndrom and to reduce the ischemic myocardial damage following an ischemic event.

The kynurenic acid analog SZR104 induces cytomorphological changes associated with the anti-inflammatory phenotype in cultured microglia.

We previously showed the anti-inflammatory effects of kynurenic acid (KYNA) and its brain-penetrable analog N-(2-(dimethylamino)ethyl)-3-(morpholinomethyl)-4-hydroxyquinoline-2-carboxamide (SZR104) both in vivo and in vitro. Here, we identified the cytomorphological effects of KYNA and SZR104 in secondary microglial cultures established from newborn rat forebrains. We quantitatively analyzed selected morphological aspects of microglia in control (unchallenged), lipopolysaccharide (LPS)-treated (challenged), KYNA- or SZR104-treated, and LPS + KYNA or LPS + SZR104-treated cultures. Multicolor immunofluorescence labeling followed by morphometric analysis (area, perimeter, transformation index, lacunarity, density, span ratio, maximum span across the convex hull, hull circularity, hull area, hull perimeter, max/min radii, mean radius, diameter of bounding circle, fractal dimension, roughness, circularity) on binary (digital) silhouettes of the microglia revealed their morphological plasticity under experimental conditions. SZR104 and, to a lesser degree, KYNA inhibited proinflammatory phenotypic changes. For example, SZR104 treatment resulted in hypertrophied microglia characterized by a swollen cell body, enlarged perimeter, increased transformation index/decreased circularity, increased convex hull values (area, perimeter, mean radius, maximum span, diameter of the bounding circle and hull circularity), altered box-counting parameters (such as fractal dimension), and increased roughness/decreased density. Taken together, analysis of cytomorphological features could contribute to the characterization of the anti-inflammatory activity of SZR104 on cultured microglia.

Kynurenic acid promotes osteogenesis via the Wnt/ß-catenin signaling.

The role of kynurenic acid (KynA) in neurological and mental diseases has been widely studied. Emerging studies disclosed that KynA has a protective effect on tissues including heart, kidney, and retina. However, the role of KynA in osteoporosis has not been reported so far. To elucidate the role of KynA in age-related osteoporosis, both control and osteoporosis mice were administrated KynA for three consecutive months, and micro-computed tomography (µCT) analysis was then performed. In addition, primary bone marrow mesenchymal stem cells (BMSCs) were isolated for osteogenic differentiation induction and treated with KynA in vitro. Our data suggested that KynA administration rescued age-related bone loss in vivo, and KynA treatment promotes BMSC osteogenic differentiation in vitro. Moreover, KynA activated the Wnt/ß-catenin signaling during BMSC osteogenic differentiation. Wnt inhibitor MSAB inhibited KynA-induced osteogenic differentiation. Further data demonstrated that KynA exerted its effect on BMSC osteogenic differentiation and Wnt/ß-catenin signaling activation via G protein-coupled receptor 35 (GPR35). In conclusion, the protective effect of KynA on age-related osteoporosis was disclosed. Additionally, the promoting effect of KynA on osteoblastic differentiation via Wnt/ß-catenin signaling was verified and the effect dependent on GPR35. These data suggest that KynA administration potentially contributes to the treatment of age-related osteoporosis.

Recent advances in GPR35 pharmacology; 5-HIAA serotonin metabolite becomes a ligand.

GPR35, an orphan receptor, has been waiting for its ligand since its cloning in 1998. Many endogenous and exogenous molecules have been suggested to act as agonists of GPR35 including kynurenic acid, zaprinast, lysophosphatidic acid, and CXCL17. However, complex and controversial responses to ligands among species have become a huge hurdle in the development of therapeutics in addition to the orphan state. Recently, a serotonin metabolite, 5-hydroxyindoleacetic acid (5-HIAA), is reported to be a high potency ligand for GPR35 by investigating the increased expression of GPR35 in neutrophils. In addition, a transgenic knock-in mouse line is developed, in which GPR35 was replaced with a human ortholog, making it possible not only to overcome the different selectivity of agonists among species but also to conduct therapeutic experiments on human GPR35 in mouse models. In the present article, I review the recent advances and prospective therapeutic directions in GPR35 research. Especially, I'd like to draw attention of readers to the finding of 5-HIAA as a ligand of GPR35 and lead to apply the 5-HIAA and human GPR35 knock-in mice to their research fields in a variety of pathophysiological conditions.

Kynurenic acid blunts A1 astrocyte activation against neurodegeneration in HIV-associated neurocognitive disorders.

Despite extensive astrocyte activation in patients suffering from HIV-associated neurocognitive disorders (HAND), little is known about the contribution of astrocytes to HAND neuropathology. Here, we report that the robust activation of neurotoxic astrocytes (A1 astrocytes) in the CNS promotes neuron damage and cognitive deficits in HIV-1 gp120 transgenic mice. Notably, knockout of α7 nicotinic acetylcholine receptors (α7nAChR) blunted A1 astrocyte responses, ultimately facilitating neuronal and cognitive improvement in the gp120tg mice. Furthermore, we provide evidence that Kynurenic acid (KYNA), a tryptophan metabolite with α7nAChR inhibitory properties, attenuates gp120-induced A1 astrocyte formation through the blockade of α7nAChR/JAK2/STAT3 signaling activation. Meanwhile, compared with gp120tg mice, mice fed with tryptophan showed dramatic improvement in cognitive performance, which was related to the inhibition of A1 astrocyte responses. These initial and determinant findings mark a turning point in our understanding of the role of α7nAChR in gp120-mediated A1 astrocyte activation, opening up new opportunities to control neurotoxic astrocyte generation through KYNA and tryptophan administration.

Modulating the Kynurenine pathway or sequestering toxic 3-hydroxykynurenine protects the retina from light-induced damage in Drosophila.

Tissue health is regulated by a myriad of exogenous or endogenous factors. Here we investigated the role of the conserved Kynurenine pathway (KP) in maintaining retinal homeostasis in the context of light stress in Drosophila melanogaster. cinnabar, cardinal and scarlet are fly genes that encode different steps in the KP. Along with white, these genes are known regulators of brown pigment (ommochrome) biosynthesis. Using white as a sensitized genetic background, we show that mutations in cinnabar, cardinal and scarlet differentially modulate light-induced retinal damage. Mass Spectrometric measurements of KP metabolites in flies with different genetic combinations support the notion that increased levels of 3-hydroxykynurenine (3OH-K) and Xanthurenic acid (XA) enhance retinal damage, whereas Kynurenic Acid (KYNA) and Kynurenine (K) are neuro-protective. This conclusion was corroborated by showing that feeding 3OH-K results in enhanced retinal damage, whereas feeding KYNA protects the retina in sensitized genetic backgrounds. Interestingly, the harmful effects of free 3OH-K are diminished by its sub-cellular compartmentalization. Sequestering of 3OH-K enables the quenching of its toxicity through conversion to brown pigment or conjugation to proteins. This work enabled us to decouple the role of these KP genes in ommochrome formation from their role in retinal homeostasis. Additionally, it puts forward new hypotheses on the importance of the balance of KP metabolites and their compartmentalization in disease alleviation.

Impact of the glutamatergic neurotransmission within the A5 region on the cardiorespiratory response evoked from the midbrain dlPAG.

Stimulation of the dorsolateral periaqueductal grey matter (dlPAG) in rats evokes an active defensive behaviour together with a cardiorespiratory response characterised by tachypnoea, tachycardia and hypertension. The dlPAG neurons involved in these responses are excitatory, presumably glutamatergic, due to the presence of vesicular glutamate transporter VGLUT2 within their axon terminals. Previously, our group described a functional interaction between dlPAG and the pontine A5 region. Accordingly, in the present work, in order to characterize the role of glutamate within this interaction, experiments were carried out in spontaneously breathing anaesthetized rats (sodium pentobarbitone 60 mg/kg i.p., suplemented with 20 mg/kg i.p.). The cardiorespiratory response evoked by electrical stimulation of the dlPAG (1 ms pulses, 20-50 µA, given at 100 Hz, during 5 s) was analysed before and after the microinjection, within the A5 region, of either kynurenic acid (non-specific glutamate receptor antagonist; 5-10 nmol), DAP-5 (NMDA antagonist; 1 pmol), CNQX (non-NMDA antagonist; 1 pmol) or MCPG (metabotropic antagonist; 0,1 nmol). Kynurenic acid decreased the intensity of both the tachypnoea (p < 0,001) and tachycardia (p < 0,001) induced by dl-PAG stimulation. Blockade of no-NMDA receptors reduced the increase of respiratory frequency, heart rate and pressor response to dl-PAG stimulation (p < 0,01, p < 0,001, p < 0,05 respectively). Blockade of either NMDA or metabotropic receptors reduced the dlPAG-evoked tachycardia and pressor response (p < 0,01; p < 0,05 respectively). These results suggest a neuromodulatory role for A5 region via glutamate neurotransmission of the dlPAG-evoked cardiorespiratory response, confirming the role of the ventrolateral pons in the neuronal circuits involved in respiratory and heart rate control.

Differential Levels of Tryptophan-Kynurenine Pathway Metabolites in the Hippocampus, Anterior Temporal Lobe, and Neocortex in an Animal Model of Temporal Lobe Epilepsy.

Glutamate-receptor-mediated hyperexcitability contributes to seizure generation in temporal lobe epilepsy (TLE). Tryptophan-kynurenine pathway (TKP) metabolites regulate glutamate receptor activity under physiological conditions. This study was designed to investigate alterations in the levels of TKP metabolites and the differential regulation of glutamatergic activity by TKP metabolites in the hippocampus, anterior temporal lobe (ATL), and neocortex samples of a lithium-pilocarpine rat model of TLE. We observed that levels of tryptophan were reduced in the hippocampus and ATL samples but unaltered in the neocortex samples. The levels of kynurenic acid were reduced in the hippocampus samples and unaltered in the ATL and neocortex samples of the TLE rats. The levels of kynurenine were unaltered in all three regions of the TLE rats. The magnitude of reduction in these metabolites in all regions was unaltered in the TLE rats. The frequency and amplitude of spontaneous excitatory postsynaptic currents were enhanced in hippocampus ATL samples but not in the neocortex samples of the TLE rats. The exogenous application of kynurenic acid inhibited glutamatergic activity in the slice preparations of all these regions in both the control and the TLE rats. However, the magnitude of reduction in the frequency of kynurenic acid was higher in the hippocampus (18.44 ± 2.6% in control vs. 30.02 ± 1.5 in TLE rats) and ATL (16.31 ± 0.91% in control vs. 29.82 ± 3.08% in TLE rats) samples of the TLE rats. These findings suggest the differential regulation of glutamatergic activity by TKP metabolites in the hippocampus, ATL, and neocortex of TLE rats.

Kynurenic acid ameliorates NLRP3 inflammasome activation by blocking calcium mobilization via GPR35.

The inflammasome has been linked to diverse inflammatory and metabolic diseases, and tight control of inflammasome activation is necessary to avoid excessive inflammation. Kynurenic acid (KA) is a tryptophan metabolite in the kynurenine pathway. However, the roles and mechanisms of the regulation of inflammasome activation by KA have not yet been fully elucidated. Here, we found that KA suppressed caspase-1 activation and IL-1ß production in macrophages by specifically inhibiting canonical and noncanonical activation of the NLRP3 inflammasome. Mechanistically, KA reduced calcium mobilization through G-protein receptor 35 (GPR35), resulting in reduced mitochondrial damage and decreased mtROS production, thus blocking NLRP3 inflammasome assembly and activation. Importantly, KA prevented lipopolysaccharide-induced systemic inflammation, monosodium urate-induced peritoneal inflammation, and high-fat diet-induced metabolic disorder. Thus, KA ameliorated inflammation and metabolic disorders by blocking calcium mobilization-mediated NLRP3 inflammasome activation via GPR35. Our data reveal a novel mechanism for KA in the modulation of inflammasome activation and suggest that GPR35 might be a promising target for improving NLRP3 inflammasome-associated diseases by regulating calcium mobilization.

Binge-like mephedrone treatment induces memory impairment concomitant with brain kynurenic acid reduction in mice.

While mephedrone (4-methylmethcathinone), a synthetic cathinone derivative, is widely abused by adolescents and young adults, the knowledge about its long-term effects on memory processes is limited. Kynurenic acid (KYNA) is a neuroactive metabolite of the kynurenine pathway of tryptophan degradation. KYNA is considered an important endogenous modulator influencing physiological and pathological processes, including learning and memory processes. The aim of this study was to determine whether (A) binge-like mephedrone administration (10.0 and 30.0 mg/kg, intraperitoneally, in 4 doses separated by 2 h) induces memory impairments, assessed 2, 8 and 15 days after mephedrone cessation in the passive avoidance test in mice, and whether (B) KYNA is involved in these memory processes. To clarify the role of KYNA in the mephedrone effects, its production in the murine brain in vivo, and in cortical slices in vitro, as well as the activities of kynurenine aminotransferases (KATs) I and II were assessed. Furthermore, cell line experiments were conducted to investigate the effects of mephedrone on normal human brain cells. Our results showed memory impairments 8 and 15 days after binge-like mephedrone administration. At the same time, reduction in the KYNA level in the murine brain was noted. In vitro studies showed no effect of mephedrone on the production of KYNA in cortical slices or on the activity of the KAT I and II enzymes. Finally, exposure of normal cells to mephedrone in vitro resulted in a modest reduction of cell viability and proliferation.

2-month ketogenic diet preferentially alters skeletal muscle and augments cognitive function in middle aged female mice.

The effect of a ketogenic diet (KD) on middle aged female mice is poorly understood as most of this work have been conducted in young female mice or diseased models. We have previously shown that an isocaloric KD started at middle age in male mice results in enhanced mitochondrial mass and function after 2 months on diet and improved cognitive behavior after being on diet for 14 months when compared with their control diet (CD) fed counterparts. Here, we aimed to investigate the effect of an isocaloric 2-month KD or CD on healthy 14-month-old female mice. At 16 months of age cognitive behavior tests were performed and then serum, skeletal muscle, cortex, and hippocampal tissues were collected for biochemical analysis. Two months on a KD resulted in enhanced cognitive behavior associated with anxiety, memory, and willingness to explore. The improved neurocognitive function was associated with increased PGC1α protein in the gastrocnemius (GTN) muscle and nuclear fraction. The KD resulted in a tissue specific increase in mitochondrial mass and kynurenine aminotransferase (KAT) levels in the GTN and soleus muscles, with a corresponding decrease in kynurenine and increase in kynurenic acid levels in serum. With KAT proteins being responsible for converting kynurenine into kynurenic acid, which is unable to cross the blood brain barrier and be turned into quinolinic acid-a potent neurotoxin, this study provides a potential mechanism of crosstalk between muscle and brain in mice on a KD that may contribute to improved cognitive function in middle-aged female mice.

Mitochondrial remodeling and ischemic protection by G protein-coupled receptor 35 agonists.

Kynurenic acid (KynA) is tissue protective in cardiac, cerebral, renal, and retinal ischemia models, but the mechanism is unknown. KynA can bind to multiple receptors, including the aryl hydrocarbon receptor, the a7 nicotinic acetylcholine receptor (a7nAChR), multiple ionotropic glutamate receptors, and the orphan G protein-coupled receptor GPR35. Here, we show that GPR35 activation was necessary and sufficient for ischemic protection by KynA. When bound by KynA, GPR35 activated Gi- and G12/13-coupled signaling and trafficked to the outer mitochondria membrane, where it bound, apparantly indirectly, to ATP synthase inhibitory factor subunit 1 (ATPIF1). Activated GPR35, in an ATPIF1-dependent and pertussis toxin-sensitive manner, induced ATP synthase dimerization, which prevented ATP loss upon ischemia. These findings provide a rationale for the development of specific GPR35 agonists for the treatment of ischemic diseases.

Kynurenine metabolism is altered in mdx mice: a potential muscle to brain connection.

NEW FINDINGS: What is the central question in this study? Promoting muscle health with regular aerobic exercise can improve mental health through a kynurenine metabolic pathway: do conditions of muscle disease such as muscular dystrophy negatively influence this pathway? What is the main finding and its importance? The DBA/2J mdx model of Duchenne muscular dystrophy exhibits altered kynurenine metabolism with less kynurenic acid and peroxisome proliferator-activated receptor-γ coactivator 1-α and higher levels of tumour necrosis factor α mRNA - results associated with anxiety-like behaviour. ABSTRACT: Regular exercise can direct muscle kynurenine (KYN) metabolism toward the neuroprotective branch of the kynurenine pathway thereby limiting the accumulation of neurotoxic metabolites in the brain and contributing to mental resilience. However, the effect of muscle disease on KYN metabolism has not yet been investigated. Previous work has highlighted anxiety-like behaviours in approximately 25% of patients with Duchenne muscular dystrophy (DMD), possibly due to altered KYN metabolism. Here, we characterized KYN metabolism in mdx mouse models of DMD. Young (8-10 week old) DBA/2J (D2) mdx mice, but not age-matched C57BL/10 (C57) mdx mice, had lower levels of circulating kynurenic acid (KYNA) and lower KYNA:KYN ratio compared with their respective wild-type (WT) controls. While both C57 and D2 mdx mice displayed signs of anxiety-like behaviour, spending more time in the corners of the arena during a novel object recognition test, this effect was more prominent in D2 mdx mice. Correlational analysis detected a significant negative association between KYNA:KYN levels and time spent in corners in D2 mice, but not C57 mice. In extensor digitorum longus muscles from D2 mdx mice, but not C57 mdx mice, we found lowered protein levels of peroxisome proliferator-activated receptor-γ coactivator 1-α and kynurenine amino transferase-1 enzyme when compared with WT. Furthermore, D2 mdx quadriceps muscles had the highest level of tumour necrosis factor α expression, which is suggestive of enhanced inflammation. Thus, our pilot work shows that KYN metabolism is altered in D2 mdx mice, with a potential contribution from altered muscle health.

Peripheral N-methyl-D-aspartate receptor activation contributes to monosodium glutamate-induced headache but not nausea behaviours in rats.

Monosodium glutamate induces behaviors thought to reflect headache and nausea in rats. We explored the effects of the N-methyl-D-aspartate receptor antagonist (2R)-amino-5-phosphonovaleric acid, the inotropic glutamate receptor antagonist kynurenic acid, and the CGRP receptor antagonist olcegepant, on monosodium glutamate-induced increases in nocifensive, headache-like and nausea behaviours. Effects of these antagonists on motor function were examined with a rotarod. The effect of the dopamine receptor antagonist metoclopramide and the serotonin 3 receptor antagonist ondansetron on nausea behaviour was also assessed. (2R)-amino-5-phosphonovaleric acid, and to a lesser extent, kynurenic acid and olcegepant, reduced nocifensive and headache-like behaviours evoked by monosodium glutamate. No alteration in motor function by (2R)-amino-5-phosphonovaleric acid, kynurenic acid or olcegepant was observed. No sex-related differences in the effectiveness of these agents were identified. Nausea behaviour was significantly more pronounced in male than in female rats. Olcegepant, ondansetron and metoclopramide ameliorated this nausea behaviour in male rats. Ondansetron and metoclopramide also reduced headache-like behaviour in male rats. These findings suggest that peripheral N-methyl-D-aspartate receptor activation underlies monosodium glutamate-induced headache-like behaviour but does not mediate the nausea behaviour in rats.