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We developed a focused series of original phenyl-glycinamide derivatives which showed potent activity across in vivo mouse seizure models, namely, maximal electroshock (MES) and 6 Hz (using both 32 and 44 mA current intensities) seizure models. Following intraperitoneal ( i.p .) administration, compound ( R )-32 , which was identified as a lead molecule, demonstrated potent protection against all seizure models with ED 50 values of 73.9 mg/kg (MES test), 18.8 mg/kg (6 Hz, 32 mA test), and 26.5 mg/kg (6 Hz, 44 mA test). Furthermore, ( R )-32 demonstrated efficacy in both the PTZ-induced kindling paradigm and the iv PTZ seizure threshold test. The expression of neurotrophic factors, such as mature brain-derived neurotrophic factor (mBDNF) and nerve growth factor (NGF), in the hippocampus and/or cortex of mice, and the levels of glutamate and GABA were normalized after PTZ-induced kindling by ( R )-32 . Importantly, besides antiseizure activity, ( R ) -32 demonstrated potent antinociceptive efficacy in formalin-induced pain, capsaicin-induced pain, as well as oxaliplatin- and streptozotocin-induced peripheral neuropathy in mice ( i.p .). No influence on muscular strength and body temperature in mice was observed. Pharmacokinetic studies and in vitro ADME-Tox data ( i.e. , high metabolic stability in human liver microsomes, a weak influence on CYPs, no hepatotoxicity, satisfactory passive transport, etc. ) proved favorable drug-like properties of ( R ) -32 . Thermal stability of ( R )- 32 shown in thermogravimetry and differential scanning calorimetry gives the opportunity to develop innovative oral solid dosage forms loaded with this compound. The in vitro binding and functional assays indicated its multimodal mechanism of action. ( R ) -32 , beyond TRPV1 antagonism, inhibited calcium and sodium currents at a concentration of 10 μM. Therefore, the data obtained in the current studies justify a more detailed preclinical development of ( R ) -32 for epilepsy and pain indications.

In the present study, a series of original alaninamide derivatives have been designed applying a combinatorial chemistry approach, synthesized, and characterized in the in vivo and in vitro assays. The obtained molecules showed potent and broad-spectrum activity in basic seizure models, namely, the maximal electroshock (MES) test, the 6 Hz (32 mA) seizure model, and notably, the 6 Hz (44 mA) model of pharmacoresistant seizures. Most potent compounds 26 and 28 displayed the following pharmacological values: ED 50 = 64.3 mg/kg (MES), ED 50 = 15.6 mg/kg (6 Hz, 32 mA), ED 50 = 29.9 mg/kg (6 Hz, 44 mA), and ED 50 = 34.9 mg/kg (MES), ED 50 = 12.1 mg/kg (6 Hz, 32 mA), ED 50 = 29.5 mg/kg (6 Hz, 44 mA), respectively. Additionally, 26 and 28 were effective in the iv PTZ seizure threshold test and had no influence on the grip strength. Moreover, lead compound 28 was tested in the PTZ-induced kindling model, and then, its influence on glutamate and GABA levels in the hippocampus and cortex was evaluated by the high-performance liquid chromatography (HPLC) method. In addition, 28 revealed potent efficacy in formalin-induced tonic pain, capsaicin-induced pain, and oxaliplatin- and streptozotocin-induced peripheral neuropathy. Pharmacokinetic studies and in vitro ADME-Tox data proved favorable drug-like properties of 28 . The patch-clamp recordings in rat cortical neurons showed that 28 at a concentration of 10 μM significantly inhibited fast sodium currents. Therefore, 28 seems to be an interesting candidate for future preclinical development in epilepsy and pain indications.

In this study, we explored the potential antidepressant-like properties of the brain-penetrant Y2 receptor (Y2R) antagonist SF-11 [N-(4-ethoxyphenyl)- 4-(hydroxydiphenylmethyl)- 1-piperidinecarbothioamide] in the astroglial degeneration model of depression with an emphasis on checking the possible mechanisms implicated in this antidepressant-like effect. The model of depression relies on the loss of astrocytes in the medial prefrontal cortex (mPFC) in Sprague-Dawley rats after administering the gliotoxin L-alpha-aminoadipic acid (L-AAA). SF-11 was administered intraperitoneally (i.p.) once (10 mg/kg) or for three consecutive days (10 mg/kg/day), and the effects of L-AAA and SF-11 injected alone or in combination were investigated using the forced swim test (FST), sucrose intake test (SIT), Western blotting, immunohistochemical staining, and microdialysis. SF-11 produced an antidepressant-like effect after single or three-day administration in rats subjected to astrocyte impairment, as demonstrated by the FST and SIT, respectively. Immunoblotting and immunohistochemical analyses showed that SF-11 reversed the L-AAA-induced astrocyte cell death in the mPFC, suggesting it is glioprotective. Microdialysis studies showed that SF-11 decreased extracellular glutamate (Glu) levels compared to basal value when administered alone and compared to the basal value and control group in LAAA-treated rats. The results from immunoblotting analysis indicated the involvement of Y2Rs in the astrocyte ablation model of depression and the antidepressant-like effect of SF-11. In addition, we observed the participation of the caspase-3 apoptotic pathway in the mechanism of gliotoxin action induced by L-AAA. These findings demonstrate that SF-11, a Y2R antagonist, elicited a rapid antidepressant-like response, possibly linked to its ability to inhibit glutamatergic neurotransmission., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Excessive fructose consumption may lead to metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MAFLD) and hypertension. α1-adrenoceptors antagonists are antihypertensive agents that exert mild beneficial effects on the metabolic profile in hypertensive patients. However, they are no longer used as a first-line therapy for hypertension based on Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial (ALLHAT) outcomes. Later studies have shown that quinazoline-based α1-adrenolytics (prazosin, doxazosin) induce apoptosis; however, this effect was independent of α1-adrenoceptor blockade and was associated with the presence of quinazoline moiety. Recent studies showed that α1-adrenoceptors antagonists may reduce mortality in COVID-19 patients due to anti-inflammatory properties. MH-76 (1-[3-(2,6-dimethylphenoxy)propyl]-4-(2-methoxyphenyl)piperazine hydrochloride)) is a non-quinazoline α1-adrenoceptor antagonist which, in fructose-fed rats, exerted anti-inflammatory, antihypertensive properties and reduced insulin resistance and visceral adiposity. In this study, we aimed to evaluate the effect of fructose consumption and treatment with α1-adrenoceptor antagonists of different classes (MH-76 and prazosin) on liver tissue of fructose-fed rats. Livers were collected from four groups (Control, Fructose, Fructose + MH-76 and Fructose + Prazosin) and subjected to biochemical and histopathological studies. Both α1-adrenolytics reduced macrovesicular steatosis and triglycerides content of liver tissue and improved its antioxidant capacity. Treatment with MH-76, contrary to prazosin, reduced leucocytes infiltration as well as decreased elevated IL-6 and leptin concentrations. Moreover, the MH-76 hepatotoxicity in hepatoma HepG2 cells was less than that of prazosin. The use of α1-adrenolytics with anti-inflammatory properties may be an interesting option for treatment of hypertension with metabolic complications.

The pharmacological activation of the GPR39 receptor has been proposed as a novel strategy for treating seizures; however, this hypothesis has not been verified experimentally. TC-G 1008 is a small molecule agonist increasingly used to study GPR39 receptor function but has not been validated using gene knockout. Our aim was to assess whether TC-G 1008 produces anti-seizure/anti-epileptogenic effects in vivo and whether the effects are mediated by GPR39. To obtain this goal we utilized various animal models of seizures/epileptogenesis and GPR39 knockout mice model. Generally, TC-G 1008 exacerbated behavioral seizures. Furthermore, it increased the mean duration of local field potential recordings in response to pentylenetetrazole (PTZ) in zebrafish larvae. It facilitated the development of epileptogenesis in the PTZ-induced kindling model of epilepsy in mice. We demonstrated that TC-G 1008 aggravated PTZ-epileptogenesis by selectively acting at GPR39. However, a concomitant analysis of the downstream effects on the cyclic-AMP-response element binding protein in the hippocampus of GPR39 knockout mice suggested that the molecule also acts via other targets. Our data argue against GPR39 activation being a viable therapeutic strategy for treating epilepsy and suggest investigating whether TC-G 1008 is a selective agonist of the GPR39 receptor., (© 2023. The Author(s).)

Many studies have shown the high efficacy of histamine H 3 receptor ligands in preventing weight gain. In addition to evaluating the efficacy of future drug candidates, it is very important to assess their safety profile, which is established through numerous tests and preclinical studies. The purpose of the present study was to evaluate the safety of histamine H 3 /sigma-2 receptor ligands by assessing their effects on locomotor activity and motor coordination, as well as on the cardiac function, blood pressure, and plasma activity of certain cellular enzymes. The ligands tested at a dose of 10 mg/kg b.w. did not cause changes in locomotor activity (except for KSK-74) and did not affect motor coordination. Significant reductions in blood pressure were observed after the administration of compounds KSK-63, KSK-73, and KSK-74, which seems logically related to the increased effect of histamine. Although the results of in vitro studies suggest that the tested ligands can block the human ether-a-go-go-related gene (hERG) potassium channels, they did not affect cardiac parameters in vivo. It should be noted that repeated administration of the tested compounds prevented an increase in the activity of alanine aminotransferase (AlaT) and gamma-glutamyl transpeptidases (gGT) observed in the control animals fed a palatable diet. The obtained results show that the ligands selected for this research are not only effective in preventing weight gain but also demonstrate safety in relation to the evaluated parameters, allowing the compounds to proceed to the next stages of research.

Phosphodiesterase (PDE) inhibitors represent a wide class of chemically different compounds that have been extensively studied in recent years. Their anti-inflammatory and anti-fibrotic effects are particularly desirable in the treatment of chronic respiratory diseases, including asthma and chronic obstructive pulmonary disease (COPD). Due to diversified expression of individual PDEs within cells and/or tissues as well as PDE signaling compartmentalization, pan-PDE inhibitors (compounds capable of simultaneously blocking various PDE subtypes) are of particular interest. Recently, a large group of 7,8-disubstituted derivatives of 1,3-dimethyl-7H-purine-2,6-dione (theophylline) was designed and synthesized. These compounds were characterized as potent pan-PDE inhibitors and their prominent anti-inflammatory and anti-fibrotic activity in vitro has been proved. Herein, we investigated a general in vitro safety profile and pharmacokinetic characteristics of two leading compounds from this group: a representative compound with N'-benzylidenebutanehydrazide moiety (38) and a representative derivative containing N-phenylbutanamide fragment (145). Both tested pan-PDE inhibitors revealed no cytotoxic, mutagenic, and genotoxic activity in vitro, showed moderate metabolic stability in mouse and human liver microsomes, as well as fell into the low or medium permeation category. Additionally, 38 and 145 revealed a lack of interaction with adenosine receptors, including A 1 , A 2A , and A 2B . Pharmacokinetic analysis revealed that both tested 7,8-disubstituted derivatives of 1,3-dimethyl-7H-purine-2,6-dione were effectively absorbed from the peritoneal cavity. Simultaneously, they were extensively distributed to mouse lungs and after intraperitoneal (i.p.) administration were detected in bronchoalveolar lavage fluid. These findings provide evidence that investigated compounds represent a new drug candidates with a favorable in vitro safety profile and satisfactory pharmacokinetic properties after a single i.p. administration. As the next step, further pharmacokinetic studies after multiple i.p. and p.o. doses will be conducted to ensure effective 38 and 145 serum and lung concentrations for a longer period of time. In summary, 7,8-disubstituted derivatives of 1,3-dimethyl-7H-purine-2,6-dione represent a promising compounds worth testing in animal models of chronic respiratory diseases, the etiology of which involves various PDE subtypes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The adenosine A 2A and A 2B receptors are promising therapeutic targets in the treatment of obesity and diabetes since the agonists and antagonists of these receptors have the potential to positively affect metabolic disorders. The present study investigated the link between body weight reduction, glucose homeostasis, and anti-inflammatory activity induced by a highly potent and specific adenosine A 2B receptor antagonist, compound PSB-603. Mice were fed a high-fat diet for 14 weeks, and after 12 weeks, they were treated for 14 days intraperitoneally with the test compound. The A 1 /A 2A /A 2B receptor antagonist theophylline was used as a reference. Following two weeks of treatment, different biochemical parameters were determined, including total cholesterol, triglycerides, glucose, TNF-α, and IL-6 blood levels, as well as glucose and insulin tolerance. To avoid false positive results, mouse locomotor and spontaneous activities were assessed. Both theophylline and PSB-603 significantly reduced body weight in obese mice. Both compounds had no effects on glucose levels in the obese state; however, PSB-603, contrary to theophylline, significantly reduced triglycerides and total cholesterol blood levels. Thus, our observations showed that selective A 2B adenosine receptor blockade has a more favourable effect on the lipid profile than nonselective inhibition.

( R )-7 [ ( R )-AS-1 ] showed broad-spectrum antiseizure activity across in vivo mouse seizure models: maximal electroshock (MES), 6 Hz (32/44 mA), acute pentylenetetrazol (PTZ), and PTZ-kindling. A remarkable separation between antiseizure activity and CNS-related adverse effects was also observed. In vitro studies with primary glia cultures and COS-7 cells expressing the glutamate transporter EAAT2 showed enhancement of glutamate uptake, revealing a stereoselective positive allosteric modulator (PAM) effect, further supported by molecular docking simulations. ( R )-7 [ ( R )-AS-1 ] was not active in EAAT1 and EAAT3 assays and did not show significant off-target activity, including interactions with targets reported for marketed antiseizure drugs, indicative of a novel and unprecedented mechanism of action. Both in vivo pharmacokinetic and in vitro absorption, distribution, metabolism, excretion, toxicity (ADME-Tox) profiles confirmed the favorable drug-like potential of the compound. Thus, ( R )-7 [ ( R )-AS-1 ] may be considered as the first-in-class small-molecule PAM of EAAT2 with potential for further preclinical and clinical development in epilepsy and possibly other CNS disorders.

Clinical and preclinical studies show evidence that chronic stress or nutritional deficits in dietary zinc (Zn) intake may be risk factors for developing major depressive disorder (MDD). Furthermore, there may be possible links between low serum Zn levels and development of treatment-resistant depression. In the present work, we combined chronic restraint stress (CRS) and a low-zinc diet (ZnD) in mice and carried out a set of behavioral and biochemical studies. The mice were treated with four different antidepressant compounds, namely, ketamine, Ro 25-6981 (Ro), hyperforin and lanicemine (Hyp + Lan), and imipramine (IMI). We show that CRS or ZnD alone or a combination of CRS and ZnD (CRS + ZnD) induces anhedonia observed in the sucrose preference test (SPT). The behavioral effects of CRS were restored by ketamine or IMI. However, only Hyp + Lan restored the deficits in behavioral phenotype in mice subjected to CRS + ZnD. We also showed that the antidepressant-like effects observed in Hyp + Lan-treated CRS + ZnD mice were associated with changes in the morphology of the dendritic spines (restored physiological level) in the hippocampus (Hp). Finally, we studied the metabolism of ketamine and its brain absorption in CRS and CRS + ZnD mice. Our results suggest that CRS + ZnD does not alter the metabolism of ketamine to (2R,6R;2S,6S)-HNK; however, CRS + ZnD can induce altered bioavailability and distribution of ketamine in the Hp and frontal cortex (FC) in CRS + ZnD animals compared to the control and CRS groups., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Pochwat, Misztak, Masternak, Bączyńska, Bijata, Roszkowska, Bijata, Włodarczyk, Szafarz, Wyska, Muszyńska, Krakowska, Opoka, Nowak and Szewczyk.)

The aim of this study was to determine, in the diet-induced obesity model in rats, the potential of Guanabenz to reduce body weight and ameliorate some metabolic disturbances. Obesity was induced in rats by a high-fat diet. After 10 weeks, rats were treated intraperitoneally with Guanabenz at the two doses: 2 or 5 mg/kg b.w./day, once daily for 25 days. The spontaneous activity of rats was measured for 24 h on the 1st and 24th day of the Guanabenz treatment with a special radio-frequency identification system. Gastric emptying was measured in intragastric phenol red-treated mice by measuring the color of the stomach homogenate 30 min after phenol red administration. Intraperitoneal administration of Guanabenz for 25 days to obese rats resulted in a significant decrease in body weight compared to the baseline values (about 11% at a dose of 5 mg/kg). Both body weight and the amount of adipose tissue in the groups receiving Guanabenz decreased to the levels observed in the control rats fed only standard feed. The anorectic effect occurred in parallel with a reduction in plasma triglyceride levels. We also confirmed the beneficial effect of Guanabenz on plasma glucose level. The present study demonstrates that the administration of Guanabenz strongly inhibits gastric emptying (about 80% at a dose of 5 mg/kg). Guanabenz can successfully and simultaneously attenuate all the disorders and risk factors of metabolic syndrome: hypertension, hyperglycemia, obesity, and dyslipidemia. However, the exact cellular mechanisms of its action require further research., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Many studies involving compounds that enhance histamine release, such as histamine H 3 receptor (H 3 R) antagonists, have shown efficacy in inhibiting weight gain, but none have passed clinical trials. As part of the search for H 3 receptor ligands that have additional properties, the aim of this study is to evaluate the activity in the reduction in weight gain in a rat model of excessive eating, as well as the impact on selected metabolic parameters, and the number and size of adipocytes of two new H 3 R antagonists, KSK-60 and KSK-74, which also exert a significant affinity at the sigma-2 receptor. Compounds KSK-60 and KSK-74 are homologues and the elongation of the distal part of the molecule resulted in an approximate two-fold reduction in affinity at H 3 R, but simultaneously an almost two-fold increase in affinity at the sigma-2 receptor. Animals fed palatable feed and receiving KSK-60 or KSK-74 both at 10 mg/kg b.w. gained significantly less weight than animals in the control obese group. Moreover, KSK-74 significantly compensated for metabolic disturbances that accompany obesity, such as an increase in plasma triglyceride, resistin, and leptin levels; improved glucose tolerance; and protected experimental animals against adipocyte hypertrophy. Furthermore, KSK-74 inhibited the development of inflammation in obesity-exposed adipose tissue. The in vivo pharmacological activity of the tested ligands appears to correlate with the affinity at the sigma-2 receptors; however, the explanation of this phenomenon requires further and extended research.

We report herein a series of water-soluble analogues of previously described anticonvulsants and their detailed in vivo and in vitro characterization. The majority of these compounds demonstrated broad-spectrum anticonvulsant properties in animal seizure models, including the maximal electroshock (MES) test, the pentylenetetrazole-induced seizure model ( sc PTZ), and the psychomotor 6 Hz (32 mA) seizure model in mice. Compound 14 showed the most robust anticonvulsant activity (ED 50 MES = 49.6 mg/kg, ED 50 6 Hz (32 mA) = 31.3 mg/kg, ED 50 sc PTZ = 67.4 mg/kg). Notably, it was also effective in the 6 Hz (44 mA) model of drug-resistant epilepsy (ED 50 = 63.2 mg/kg). Apart from favorable anticonvulsant properties, compound 14 revealed a high efficacy against pain responses in the formalin-induced tonic pain, the capsaicin-induced neurogenic pain, as well as in the oxaliplatin-induced neuropathic pain in mice. Moreover, compound 14 showed distinct anti-inflammatory activity in the model of carrageenan-induced aseptic inflammation. The mechanism of action of compound 14 is likely complex and may result from the inhibition of peripheral and central sodium and calcium currents, as well as the TRPV1 receptor antagonism as observed in the in vitro studies. This lead compound also revealed beneficial in vitro ADME-Tox properties and an in vivo pharmacokinetic profile, making it a potential candidate for future preclinical development. Interestingly, the in vitro studies also showed a favorable induction effect of compound 14 on the viability of neuroblastoma SH-SY5Y cells.

Rationale: Pterostilbene is the 3,5-dimethoxy derivative of resveratrol with numerous beneficial effects including neuroprotective properties. Experimental studies revealed its anticonvulsant action in the acute seizure tests., Objectives: The purpose of the present study was to evaluate the effect of pterostilbene in the pentetrazol (PTZ)-induced kindling model of epilepsy in mice as well as to assess some possible mechanisms of its anticonvulsant action in this model., Methods: Mice were repeatedly treated with pterostilbene (50-200 mg/kg) and its effect on the development of seizure activity in the PTZ kindling was estimated. Influence of pterostilbene on the locomotor activity and anxiety- and depression-like behavior in the PTZ-kindled mice was also assessed. To understand the possible mechanisms of anticonvulsant activity of pterostilbene, γ-aminobutyric acid (GABA) and glutamate concentrations in the prefrontal cortex and hippocampus of the PTZ-kindled mice were measured using LC-MS/MS method. Moreover, mRNA expression of BDNF, TNF-α, IL-1β, IL-6, GABRA1A, and GRIN2B was determined by RT-qPCR technique., Results: We found that pterostilbene at a dose of 200 mg/kg considerably reduced seizure activity but did not influence the locomotor activity and depression- and anxiety-like behavior in the PTZ-kindled mice. In the prefrontal cortex and hippocampus, pterostilbene reversed the kindling-induced decrease of GABA concentration. Neither in the prefrontal cortex nor hippocampus pterostilbene affected mRNA expression of IL-1β, IL-6, GABRA1A, and GRIN2B augmented by PTZ kindling. Pterostilbene at a dose of 100 mg/kg significantly decreased BDNF and TNF-α mRNA expression in the hippocampus of the PTZ-kindled mice., Conclusions: Although further studies are necessary to understand the mechanism of anticonvulsant properties of pterostilbene, our findings suggest that it might be considered a candidate for a new antiseizure drug., (© 2021. The Author(s).)

Noting the worldwide rapid increase in the prevalence of overweight and obesity new effective drugs are now being sought to combat these diseases. Histamine H 3 receptor antagonists may represent an effective therapy as they have been shown to modulate histamine synthesis and release and affect a number of other neurotransmitters (norepinephrine, acetylcholine, γ-aminobutyric acid, serotonin, substance P) thus influencing the food intake. Based on the preliminary studies determining affinity, intrinsic activity, and selected pharmacokinetic parameters, two histamine H 3 receptor ligands were selected. Female rats were fed palatable food for 28 days and simultaneously administered the tested compounds intraperitoneally (i.p.) at a dose of 10 or 1 mg/kg b.w./day. Weight was evaluated daily and calorie intake was evaluated once per week. The plasma levels of cholesterol, triglycerides, leptin, adiponectin, ghrelin, corticosterone, CRP and IL-6 were determined at the end of experiment. The glucose tolerance test was also performed. To exclude false positives, the effect of tested compounds on spontaneous activity was monitored during the treatment, as well as the amount of consumed kaolin clay was studied as a reflection of possible gastrointestinal disturbances comparable to nausea. The histamine H 3 receptor antagonists KSK-59 and KSK-73 administered i.p. at a dose of 10 mg/kg b.w. prevented weight gain in a rat model of excessive eating. They reduced adipose tissue deposits and improved glucose tolerance. Both compounds showed satisfying ability to penetrate through biological membranes determined in in vitro studies. Compound KSK-73 also reduced the caloric intake of the experimental animals what indicates its anorectic effect. These results show the pharmacological properties of histamine H 3 receptor antagonists, (4-pyridyl)piperazine derivatives, as the compounds causing not only slower weight gain but also ameliorating some metabolic disorders in rats having the opportunity to overeat.

Aims: One of the therapeutic approaches in the treatment of obesity is the use of histamine H 3 receptor ligands. Histamine plays a significant role in eating behavior because it causes a loss of appetite and is considered to be a satiety signal released during food intake., Material and Methods: Histamine ligands were selected based on the preliminary studies which included determination of intrinsic activity and selected pharmacokinetic parameters. Female Wistar rats were fed palatable feed for 28 days and simultaneously the tested compounds were administered intraperitoneally at a dose of 10 mg/kg b.w./day. Rats' weight was evaluated daily and calories intake was evaluated once per week. At the end of experiment insulin and glucose tolerance tests was performed. Plasma levels of cholesterol, triglycerides, leptin, insulin, glucose, C-peptide and CRP were also determined. In order to rule out false-positive results the influence of tested compounds on spontaneous activity of rats was monitored., Results: Animals fed palatable feed and treated with KSK-61 or KSK-63 compounds showed the slowest weight gain which was comparable to the one observed in control animals. Both compounds with the highest pharmacological activity have also similar pharmacokinetic properties with quite long half-life and high volume of distribution indicating that they can freely cross most biological barriers. Some compounds, especially KSK-63, compensated for metabolic disorders., Conclusion: The presented study proves that search among the active histamine H 3 receptor ligands for the new therapeutic agents to treat obesity is justified. Compounds KSK-61 and KSK-63 can be considered as the leading structures., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

It is important to search for a promising therapeutic target or small molecules that can control excessive eating since limiting the intake of foods, especially tasty ones, could be effective in the treatment or prevention of obesity. Some studies indicate betahistine as an unique drug having the ability to ameliorate, for example, antipsychotic-induced weight gain. This study aimed to determine whether repeated administration of betahistine (histamine H1R agonist and H3R antagonist) could be beneficial in reducing the intake of tasty foods or the body's response to overeating via mechanisms such as by influencing the levels of hormones involved in the regulation of food intake or the levels of selected metabolic parameters. Studies were performed in the excessive eating model in rats, which perfectly illustrates the harmful high-caloric intake from freely available tasty products rich in sugar and fat. Our results indicated that repeated administration of betahistine to rats caused lower gain of body mass compared to the control rats fed palatable feed. Interestingly, betahistine treatment increased the consumption of cheese, which is a source of histamine. Although betahistine did not prevent the development of metabolic disorders, such as reduced glucose tolerance, in test animals, it significantly increased the level of high-density lipoprotein cholesterol, which could certainly be considered beneficial. Further studies should be conducted to investigate the effect of repeated administration of betahistine on satiety, gastrointestinal disorders, and the preference for histamine-containing foods., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

GPR18 is an orphan GPCR that is activated by the cannabinoid tetrahydrocannabinol (THC). Emerging evidence indicates its involvement in the control of cardiovascular functions, including heart rate, contractility, vascular tone, as well as blood pressure. Therefore, we investigated the effects of selective GPR18 receptor ligands, namely PSB-KD-107 (agonist) and PSB-CB-92 (antagonist), on blood pressure, electrocardiogram (ECG), and vascular dilatation in vitro and in vivo, as well as their anti-oxidative potential in in vitro ferric reducing antioxidant power (FRAP) and 2,2-diphenyl-1-picryl-hydrazyl-hydrate free radical (DPPH) assays. Our results clearly show that PSB-KD-107 dilates blood vessels. This effect is related to its activation of GPR18 as it can be blocked by the GPR18 antagonist PSB-CB-92. Moreover, our finding confirms the presence of GPR18 in blood vessels. The mechanism of the vasorelaxant activity of PSB-KD-107 is mainly related to endothelial nitric oxide generation; however, we cannot exclude additional nitric oxide-independent mechanisms or a direct influence on K + channels. PSB-KD-107 may affect blood pressure and heart function after a single administration; however, this effect was no longer observed after repeated administrations once daily for eight days. PSB-KD-107 does not affect platelet aggregation-an important feature considering the safety of its administration. PSB-KD-107 also shows a significant anti-oxidant effect and further studies of its antioxidant activity in vivo are justified.

Ischemic stroke is the third leading cause of death in the world, which accounts for almost 12% of the total deaths worldwide. Despite decades of research, the available and effective pharmacotherapy is limited. Some evidence underlines the beneficial properties of hydrogen sulfide (H 2 S) donors, such as NaSH, in an animal model of brain ischemia and in in vitro research; however, these data are ambiguous. This study was undertaken to verify the neuroprotective activity of AP39, a slow-releasing mitochondria-targeted H 2 S delivery molecule. We administered AP39 for 7 days prior to ischemia onset, and the potential to induce brain tolerance to ischemia was verified. To do this, we used the rat model of 90-min middle cerebral artery occlusion (MCAO) and used LC-MS/MS, RT-PCR, Luminex TM assays, Western blot and immunofluorescent double-staining to determine the absolute H 2 S levels, inflammatory markers, neurotrophic factor signaling pathways and apoptosis marker in the ipsilateral frontal cortex, hippocampus and in the dorsal striatum 24 h after ischemia onset. AP39 (50 nmol/kg) reduced the infarct volume, neurological deficit and reduced the microglia marker (Iba1) expression. AP39 also exerted prominent anti-inflammatory activity in reducing the release of Il-1β, Il-6 and TNFα in brain areas particularly affected by ischemia. Furthermore, AP39 enhanced the pro-survival pathways of neurotrophic factors BDNF-TrkB and NGF-TrkA and reduced the proapoptotic proNGF-p75 NTR -sortilin pathway activity. These changes corresponded with reduced levels of cleaved caspase 3. Altogether, AP39 treatment induced adaptative changes within the brain and, by that, developed brain tolerance to ischemia.

Background: Quinazoline α 1 -adrenoceptors antagonists have been shown to exert moderately favorable effects on the metabolic profile in hypertensive patients. However, based on AntiHypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial (ALLHAT) results, they are no longer recommended as a first line therapy of hypertension. Recent studies have shown that quinazoline-based α 1 -adrenoceptors antagonists (prazosin, doxazosin) induce the apoptosis and necrosis, which may be responsible for ALLHAT outcomes; however, these effects were proven to be independent of α 1 -adrenoceptor blockade and were associated with the presence of quinazoline moiety. MH-76 (1-[3-(2,6-dimethylphenoxy)propyl]-4-(2-methoxyphenyl)piperazine hydrochloride)) is a non-quinazoline α 1 -adrenoceptor antagonist which, in fructose-fed rats, exerted antihypertensive effect, and, contrary to prazosin, reduced insulin resistance and abdominal adiposity. In this study we aimed to further investigate and compare the effects of MH-76 and prazosin on inflammation in adipose tissue of fructose-fed rats., Methods: Abdominal adipose tissue was collected from four groups of fructose-fed rats (Control, Fructose, Fructose + MH-76 and Fructose + Prazosin) and subjected to biochemical, histopathological and immunohistochemical studies. Moreover, selected tissue distribution studies were performed., Results: Treatment with MH-76 but not with prazosin improved endothelial integrity, reduced adipose tissue inflammation and infiltration by immune cells, resulting in lowering leptin, MCP-1, IL-6, TNF-α and PAI-1 levels. In adipose tissue from Fructose + MH-76 animals, a higher amount of eosinophils accompanied with higher IL-4 concentration was observed. Treatment with MH-76 but not with prazosin markedly reduced phosphorylation of IRS-1 at Ser307., Conclusion: MH-76 may improve insulin signaling in adipose tissue by reducing the pro-inflammatory cytokine production and inhibiting the inflammatory cells recruitment. In contrast, in adipose tissue from animals treated with prazosin, the inflammatory effect was clearly enhanced.

GPR18 has been proposed to play a role in the progression of metabolic disease and obesity. Therefore, the aim of this study was to determine the effects of selective GRP18 ligands (the antagonists PSB-CB5 and PSB-CB27 and the agonist PSB-KK1415) on body mass and the development of metabolic disorders commonly accompanying obesity. Experiments were carried out on female Wistar rats. In order to determine the anorectic activity of the investigated ligands, their effect on food and water intake in a model of excessive eating was assessed. Lipid profile, glucose and insulin levels as well as alanine aminotransferase, aspartate aminotransferase, and γ-glutamyl transpeptidase activity in plasma were also evaluated. Potential side effects were examined in rat models of pica behavior and conditioned taste aversion. Animals treated with different ligands gained significantly less weight than rats from the obese control group. Effects of GPR18 antagonists on food intake and body weight were specific and unrelated to visceral illness, stress or changes in spontaneous activity. However, the GPR18 agonist is likely to affect body weight by inducing gastrointestinal disorders such as nausea. The presented preliminary data support the idea that the search for selective GPR18 antagonists for the treatment of obesity might be promising.

A 2B adenosine receptors are present in a wide spectrum of tissues, especially on cells of the immune system. Since these particular receptors have the lowest, of all adenosine receptor subtypes, affinity for adenosine they are believed to play a special role in immunological processes associated with elevated adenosine levels such as inflammation. The aim of this preliminary study was to determine the potential anti-inflammatory properties of compound PSB-603, a potent and selective adenosine A 2B receptor antagonist, in two different experimental models of local and systemic inflammation. In a model of inflammation induced by local carrageenan administration paw edema was measured using a pletysmometer. Additionally, levels of C-reactive protein (CRP), interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α) and reactive oxygen species (ROS) were determined in the inflamed paw. Using the mouse model of peripheral inflammation induced by intraperitoneal (ip) administration of zymosan A, the influence of the A 2B antagonist on the infiltration of neutrophils into the peritoneum and its effect on the plasma levels of CRP, TNF-α, and IL-6 were investigated. The results showed that PSB-603 administered at a dose of 5 mg/kg b.w. ip significantly reduced inflammation in both tested models. Particularly, it significantly decreased levels of the inflammatory cytokines IL-6, TNF-α and of ROS in the inflamed paw and reduced inflammation of the peritoneum by significantly decreasing the infiltration of leukocytes. Additionally, in the latter model, no statistically significant difference was observed in the CRP level between the control group without inflammation and the group which has been treated with the PSB-603 compound. Thus, the results may indicate the anti-inflammatory activity of adenosine A 2B receptor antagonists in two different models of inflammation., (Copyright © 2020 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Background: Our previous studies showed that xanthone derivatives with N -(2-methoxyphenyl)piperazine fragment have an affinity to the 5-HT1A receptor and show antidepressant-like properties in rodents. In this study, we tested three xanthone derivatives, HBK-1 (R, S) and its enantiomers, in which we increased the distance between the piperazine and xanthone fragments by using a hydroxypropoxy linker. We hypothesized that this would increase the binding to the 5-HT1A receptor and consequently, pharmacological activity., Aims: We aimed to assess the in vitro and in vivo pharmacological activity of the xanthone derivatives., Methods: We evaluated the in vitro affinity for serotonin 5-HT1A and 5-HT2A receptors and serotonin transporter. We also determined the intrinsic activity at the 5-HT1A receptor. We investigated the antidepressant-like properties and safety after acute administration (dose range: 1.25-20 mg/kg) using the forced swim, tail suspension, locomotor activity, rotarod and chimney tests in mice. We also evaluated the basic pharmacokinetic parameters., Results: Our results indicated that the compounds showed a high affinity for the 5-HT1A receptor but very weak antagonistic properties in the Ca 2+ mobilization assay; however, they showed significant agonistic properties in the β-arrestin recruitment assay. In both behavioural tests the studied xanthone derivatives showed antidepressant-like activity. Pre-treatment with p-chlorophenylalanine or WAY-100635 abolished their antidepressant-like activity. None of the compounds caused motor impairments at antidepressant-like doses. The racemate penetrated the blood-brain barrier and had a relatively high bioavailability after intraperitoneal administration., Conclusions: Xanthone derivatives with N -(2-methoxyphenyl)piperazine fragment and hydroxypropoxy linker show increased binding to the 5-HT1A receptor and may represent an attractive putative treatment candidate for depression.

The A2 adenosine receptors play an important role, among others, in the regulation of inflammatory process and glucose homeostasis in diabetes and obesity. Thus, the presented project evaluated of influence of the selective antagonist of A2A adenosine receptor-KD-64 as compared to the known non-selective antagonist-caffeine on these two particular processes. Two different inflammation models were induced namely local and systemic inflammation. Obesity was induced in mice by high-fat diet and the tested compounds (KD-64 and caffeine) were administrated for 21 days. KD-64 showed anti-inflammatory effect in both tested inflammation models and administered at the same dose as ketoprofen exerted stronger effect than this reference compound. Elevated levels of IL-6 and TNF-α observed in obese control mice were significantly lowered by the administration of KD-64 and were similar to the values observed in control non-obese mice. Interestingly, caffeine increased the levels of these parameters. In contrast to caffeine which had no influence on AlaT activity, KD-64 administration significantly lowered AlaT activity in the obese mice. Although, contrary to caffeine, KD-64 did not reduce diet-induced obesity in mice, it improved glucose tolerance. Thus, the activity of the selective adenosine A2A receptor antagonist was quite different from that of the non-selective., Competing Interests: The authors have declared that no competing interests exist.

Introduction: High dose methotrexate (HD-Mtx) is highly effective and significantly improves overall acute lymphoblastic leukemia (ALL) patients survival. The pharmacodynamics of Mtx depends on the polymorphism of genes encoding proteins engaged in the folate metabolism pathway. The aim of the current study is to determine the relationship between variants of folate metabolism-related genes and the frequency of acute toxicities of HD-Mtx. Material and Methods: A group of 133 patients aged 1.5-18.1 years (median: 6.3) was treated in accordance with the ALL-IC-2002 and ALL-IC-2009 protocols. The following polymorphisms were determined: 80 G>A SLC19A1 (solute carrier family 19 member 1; rs1051266) with direct DNA sequencing, as well as 677 C>T MTHFR (methylenetetrahydrofolate reductase; rs1801133) and the tandem repeats of the TS (thymidylate synthase) with PCR technique. HD-Mtx organ toxicities were evaluated based on the laboratory tests results and the National Cancer Institute criteria. Results: In patients with genotypes AA for SLC19A1 and CC or CT for MTHFR Mtx steady state concentrations (C ss ) and AUC inf were distinctly higher. In patients with genotype 3R/3R for TS initial elimination rate constant was significantly higher ( P = 0.003). Patients receiving Mtx at the dose of 5 g/m 2 had lower clearance (4.35 vs. 8.92 L/h/m 2 ) as compared to the ones receiving 2 g/m 2 that indicates non-linear Mtx elimination at the higher dose. Liver impairment was the most frequently observed toxicity. The homozygous genotype was associated with a significantly higher incidence of hepatic toxicity for both the SLC19A1 ( P = 0.037) and TS ( P = 0.002). Logistic regression analysis indicated an increased risk of vomiting for the 2R/3R genotype of the TS gene (OR 3.20, 95% CI 1.33-7.68, P = 0.009) and for vomiting and hepatic toxicity for the 3R/3R genotype (vomiting: OR 3.39, 95% CI 1.12-10.23, P = 0.031; liver toxicity: OR 2.28, 95% CI 1.05-4.95, P = 0.038). None of the acute toxicities differed between the analyzed dosing groups. Conclusions: Determination of polymorphisms of SLC19A1, MTHFR , and TS genes might allow for a better prior selection of patients with higher risk of elevated Mtx levels. Our study is the first one to report the increased risk of hepatotoxicity and vomiting in patients with TS polymorphisms., (Copyright © 2020 Cwiklinska, Czogala, Kwiecinska, Madetko-Talowska, Szafarz, Pawinska, Wieczorek, Klekawka, Rej, Stepien, Halubiec, Lazarczyk, Miklusiak, Bik-Multanowski, Balwierz and Skoczen.)

Cannabidiol and cannabidiol-enriched products have recently attracted much attention as an add-on therapy for epilepsy, especially drug-resistant seizures. It should be, however, remembered that concomitant use of cannabidiol and antiepileptic drugs may pose a risk of interactions between them. For this reason, the aim of our study was to assess the effect of cannabidiol on the activity of selected new antiepileptic drugs in the electrically-induced seizure models in mice. We studied the effect of cannabidiol on the anticonvulsant action of topiramate, oxcarbazepine, lamotrigine, and pregabalin in the maximal electroshock-induced seizure test as well as on the activity of levetiracetam, tiagabine, lacosamide, and gabapentin in the 6 Hz seizure test in mice. We showed that cannabidiol increased the activity of topiramate, oxcarbazepine, pregabalin, tiagabine, and gabapentin. It did not affect the anticonvulsant effect of lamotrigine and lacosamide. Interestingly, cannabidiol attenuated the anticonvulsant activity of levetiracetam. Co-administration of antiepileptic drugs with cannabidiol did not cause adverse effects such as impairment of motor coordination, changes in neuromuscular strength or potentiation of the cannabidiol-induced hypolocomotion. Serum and brain levels of antiepileptic drugs and cannabidiol were determined by using HPLC in order to ascertain any pharmacokinetic contribution to the observed behavioral effects. Only interaction with levetiracetam was purely pharmacodynamic in nature because no changes in serum and brain concentration of either levetiracetam or cannabidiol were observed. Increased anticonvulsant activity of topiramate, oxcarbazepine, pregabalin, tiagabine, and gabapentin could be, at least in part, related to pharmacokinetic interactions with cannabidiol because there were changes in serum and/or brain concentrations of antiepileptic drugs and/or cannabidiol. Pharmacokinetic interactions cannot be also excluded between lacosamide and cannabidiol because cannabidiol increased brain concentration of lacosamide and lacosamide increased brain concentration of cannabidiol. Further pharmacokinetic studies are required to evaluate the type of interactions between cannabidiol and novel antiepileptic drugs., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

A novel series of 4-pyridylpiperazine derivatives with varying alkyl linker length and eastern part substituents proved to be potent histamine H 3 receptor (hH 3 R) ligands in the nanomolar concentration range. While paying attention to their alkyl linker length, derivatives with a six methylene linker tend to be more potent than their five methylene homologues. Moreover, in the case of both phenoxyacetyl- and phenoxypropionyl- derivatives, an eight methylene linkers possess lower activity than their seven methylene homologues. However, in global analysis of collected data on the influence of alkyl linker length, a three methylene homologues appeared to be of highest hH 3 R affinity among all described 4-pyridylpiperazine derivatives from our group up to date. In the case of biphenyl and benzophenone derivatives, compounds with para- substituted second aromatic ring were of higher affinity than their meta analogues. Interestingly, benzophenone derivative 18 showed the highest affinity among all tested compounds (hH 3 R K i  = 3.12 nM). The likely protein-ligand interactions, responsible for their high affinity were demonstrated using molecular modeling techniques. Furthermore, selectivity, intrinsic activity at H 3 R, as well as drug-like properties of selected ligands were evaluated using in vitro methods., (Copyright © 2019. Published by Elsevier Inc.)

Aims: Histamine H 3 receptors ligands act anorectic by blocking the H 3 autoreceptors in the CNS, that results in increased synthesis and disinhibition of histamine release. Histamine further influencing H 1 receptors participates in the leptin-dependent inhibition of food intake. It also affects the peripheral metabolism by increasing white adipose tissue lipolysis. Thus, ligands such as KSK19 with significant antagonistic properties at the H 3 receptor might serve as an useful treatment for obesity., Materials and Methods: To induce obesity, female CD-1 mice were fed a high-fat diet for 14 weeks. The test compound at the doses of 10 or 15 mg/kg, i.p. was administrated for 21 days. Glucose and insulin tolerance tests was performed at the beginning of week 15. At the end of study, amount of intraperitoneal fat pads, AlAT, IL-6 and TNF-α plasma levels were determined., Results: Animals fed with high-fat diet and treated with test compound at the dose of 15 mg/kg showed significantly less weight gain, than mice from the control group. The use of KSK19 for 21 days in obese mice also significantly improved glucose tolerance and insulin resistance. In the tested doses KSK19 did not affect locomotor activity neither in lean nor in obese mice after single i.p. administration, but spontaneous activity increased during three hour after twentieth administrations., Conclusion: KSK19 is a strong, selective histamine H 3 receptor antagonist with a favorable influence on body weight after multiple administration at the dose of 15 mg/kg. Moreover it significantly improves glucose tolerance., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background and Aims: Metabolic syndrome associated with insulin resistance and hypertension is often caused by excessive fructose consumption. Treatment of hypertension in patients with metabolic syndrome is a difficult task as many antihypertensive drugs have adverse effects on the metabolic profile. We investigated if MH-76 and MH-79, non-quinazoline α 1 -adrenoceptor antagonists with an additional ability to stimulate NO/cGMP/K + pathway, ameliorates metabolic syndrome in fructose-fed rats. As reference compound prazosin was used., Methods and Results: Male rats were divided into 5 groups (n = 8) and studied for 18 weeks: group control: standard diet and drinking water; group Fructose: high-fructose diet (20% fructose in drinking water); groups Fructose + MH-76, Fructose + MH-79, Fructose + prazosin: high-fructose diet with subsequent MH-76, MH-79 (5 mg/kg/day ip) or prazosin (0.2 mg/kg/day ip) treatment 12 weeks later. In addition to their antihypertensive effect, the studied compounds reversed endothelial dysfunction, decreased hyperglycemia and hypertriglyceridemia, as well as prevented abdominal adiposity. Moreover, MH-76 reduced insulin resistance and decreased TNF-α concentration and lipid peroxidation in adipose tissue. Prazosin treatment exerted an antihypertensive effect, reduced hyperglycemia but did not improve endothelial dysfunction, insulin resistance, and abdominal adiposity. The lower efficacy of prazosin may be the result of its short half-time and the lack of described pleiotropic effects., Conclusions: α 1 -adrenoceptor blockade, endothelial protection, TNF-α suppressing and antioxidant activity together with favorable pharmacokinetic parameters determines high efficacy of MH-76, leading to the effective improvement of hemodynamic and metabolic disturbances in metabolic syndrome. The use of non-quinazoline, multiple-targeted α 1 -blockers may be an interesting option for treatment of hypertension with metabolic complications., (Copyright © 2019 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.)

Tadalafil, a selective phosphodiesterase type 5 inhibitor, is a long-acting oral agent for the treatment of erectile dysfunction of multiple etiologies. Although generalized tonic-clonic seizures were reported in a healthy man after taking tadalafil, the influence of tadalafil on seizure susceptibility has not been studied so far. Therefore, the aim of the present study was to investigate the effect of tadalafil on seizure threshold as well as on the activity of some first- and second-generation antiepileptic drugs in three acute seizure tests in mice. The obtained results showed that tadalafil, at the highest dose tested (20 mg/kg), significantly decreased the threshold for the first myoclonic twitch in the intravenous pentylenetetrazole (i.v. PTZ) seizure test. It did not affect the threshold for generalized clonic seizure and forelimb tonus in the i.v. PTZ, for tonic hindlimb extension in the maximal electroshock seizure threshold test, and for psychomotor seizure in the 6-Hz-induced seizure threshold test. Tadalafil did not alter the anticonvulsant activity of any of the studied antiepileptic drugs in electrically induced seizure tests. Interestingly, tadalafil potentiated the anticonvulsant activity of clonazepam and decreased the anticonvulsant activity of oxcarbazepine in the i.v. PTZ test. These interactions were pharmacodynamic in nature, as tadalafil did not alter clonazepam and oxcarbazepine concentrations both in serum and brain tissue. Furthermore, neither tadalafil alone nor its combinations with the studied antiepileptic drugs produced any significant impairment of motor coordination (assessed in the chimney test), muscular strength (investigated in the grip-strength test), and long-term memory (assessed in the passive avoidance task). In conclusion, tadalafil may increase the risk of myoclonic seizure and decrease the anticonvulsant efficacy of oxcarbazepine. Further studies are warranted to evaluate the safety of tadalafil usage in patients with epilepsy.

Tianeptine is an atypical antidepressant with a unique mechanism of action and recently it has been also reported that its major metabolite, compound MC5, possesses pharmacological activity similar to that of the parent drug. The current study aims to investigate the pharmacokinetics (PK) of both tianeptine and MC5 after intravenous or intraperitoneal administration of the parent drug as well as the metabolic ratio of MC5 in rats. To achieve these goals an LC-MS/MS method using the small sample volume for the quantitation of tianeptine and its active metabolite MC5 in rat plasma and liver perfusate has been developed and validated. Following an intravenous administration of tianeptine pharmacokinetic parameters were calculated by non-compartmental analysis. The average tianeptine volume of distribution at steady state was 2.03 L/kg and the systemic clearance equaled 1.84 L/h/kg. The mean elimination half-lives of tianeptine and MC5 metabolite were 1.16 and 7.53 h, respectively. The hepatic clearance of tianeptine determined in the isolated rat liver perfusion studies was similar to the perfusate flow rate despite the low metabolic ratio of MC5. Mass spectrometric analysis of rat bile indicated that tianeptine and MC5 metabolite are eliminated with bile as glucuronide and glutamine conjugates. Bioavailability of tianeptine after its intraperitoneal administration was 69%. The PK model with a metabolite compartment developed in this study for both tianeptine and MC5 metabolite after two routes of administration may facilitate tianeptine dosage selection for the prospective pharmacological experiments.

Treatment with 1-methylnicotinamide (MNA), a major metabolite of nicotinamide, exerts antithrombotic, anti-inflammatory, and vasoprotective effects. Yet, pharmacokinetic (PK) profile of MNA has not been fully characterized. In the present work, we analyze the PK profile of the MNA given as a nitrate (MNANO 3 ) in comparison to nitrite (MNANO 2 ) or chloride (MNACl) in rats. The bioavailability of MNA administered as MNANO 3 equaled 22.4% as compared to MNANO 2 or MNACl (9.2% and 9.1%, respectively). Moreover, in single-pass intestinal perfusion experiments, effective permeability of MNA given as MNANO 3 was higher as compared to MNA administered as MNANO 2 or MNACl. In turn, t max was the shortest and C max the highest (0.22 h and 56.65μM) for intragastrically administered MNANO 2 comparing to MNANO 3 (1.92 h, 21.74μM) or MNACl (0.63 h, 16.13μM). Transfer constant between central and peripheral compartments (k cp ) and volume of distribution (V ss ) for MNANO 3 (0.33 h -1 and 1.96 L/kg) were higher as compared to MNANO 2 or MNACl (0.11 h -1 , 0.08 h -1 for k cp and 1.05 L/kg, 0.76 L/kg for V ss , respectively). In conclusion, we characterized PK profile of MNA and demonstrated that nitrate ion augmented bioavailability and favorably modified PK profile of MNA. Furthermore, given vasoprotective properties of MNA as well as nitrate, MNANO 3 represents a bifunctional compound., (Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.)

A sensitive and specific liquid chromatography tandem mass spectrometry method for quantification of 1-methylpyridinium (1-MP) and 1,4-dimethylpyridinium (1,4-DMP) in rat plasma and tissues homogenates was developed. Chromatographic separation was performed on an Aquasil C18 analytical column with an isocratic elution of acetonitrile and water, both with an addition of formic acid (0.1%, v/v). Detection was achieved by triple quadrupole mass spectrometer TSQ Quantum Ultra equipped with a heated electrospray ionization source (HESI). The limit of quantification for both compounds was 0.05 pg/mL in plasma and 0.25 μg/g in studied tissues. The method was applied to pharmacokinetics and bioavailability of both 1-MP and 1,4-DMP with tissue distribution of 1,4-DMP in rats. Pharmacokinetic studies of 1-MP and 1,4-DMP were carried out following their intravenous or intragastric administration to male Wistar rats at the dose of 100 mg/kg. The terminal half-lives of I-MP and 1,4-DMP after their intravenous administration were 55.3 and 70.8 min, respectively. The absolute bioavailability was 51 and 31% for t-MP and 1,4-DMP, respectively.

Exogenous 1-methylnicotinamide (MNA) displays anti-inflammatory activity. The aim of this work was to characterize the profile of release of endogenous MNA during the initiation and progression of murine hepatitis induced by Concanavalin A (ConA). In particular we aimed to clarify the role of interleukin-6 (IL-6) as well as the energy state of hepatocytes in MNA release in early and late phases of ConA-induced hepatitis in mice. Hepatitis was induced by ConA in IL-6(+/+) and IL-6(-/-) mice, and various parameters of liver inflammation and injury, as well as the energy state of hepatocytes, were analysed in relation to MNA release. The decrease in ATP/ADP and NADH/NAD ratios, cytokine release (IL-6, IFN-ɤ), acute phase response (e.g. haptoglobin) and liver injury (alanine aminotransaminase, ALT) were all blunted in ConA-induced hepatitis in IL-6(-/-) mice as compared to IL-6(+/+) mice. The release of MNA in response to Con A was also significantly blunted in IL-6(-/-) mice as compared to IL-6(+/+) mice in the early stage of ConA-induced hepatitis. In turn, nicotinamide N-methyltransferase (NNMT) and aldehyde oxidase (AO) activities were blunted in the liver and MNA plasma concentration was elevated to similar degree in the late stage after Concanavalin A in IL-6(+/+) and IL-6(-/-) mice. In conclusion, we demonstrated that in ConA-induced hepatitis, early, but not late MNA release was IL-6-dependent. Our results suggest that in the initiation and early hepatitis, MNA release is linked to the energy deficit/impaired redox status in hepatocytes, while in a later phase, MNA release is rather linked to the systemic inflammation., (Copyright © 2015 Elsevier B.V. All rights reserved.)

1. The aim of this study was to evaluate pharmacokinetics and tissue distribution of novel histamine H3 receptor antagonist 1-[3-(4-tert-butylphenoxy)propyl]piperidine (compound DL76). 2. Following intravenous administration of DL76 at the dose of 3 mg/kg, pharmacokinetic parameters were calculated using non-compartmental analysis. The systemic serum clearance was 10.08 L/h/kg and the estimated blood clearance was 5.64 L/h/kg. The volume of distribution at steady state was 16.1 L/kg which was greater than total body water, terminal half-life and MRT equalled 1.41 h and 1.6 h, respectively. The two-compartment pharmacokinetic model with enterohepatic circulation was also successfully fitted to the experimental data. 3. After systemic administration, DL76 was rapidly distributed into all organs studied (liver, kidney, brain, and lung). The highest AUC of DL76 was observed in lungs followed by brain, where the exposure to the investigated compound expressed as AUC was almost 30 times higher than in serum. 4. Bioavailability, calculated based on the area-under-the-concentration-time curve extrapolated to infinity after intravenous and intragastric administration of the dose 3 mg/kg, equalled 60.9%.

A sensitive and specific liquid chromatography electrospray ionization-mass spectrometry method for determination of 1,4-dimethylpyridinium (1,4-DMP) in rat plasma has been developed and validated. Chromatography was performed on an Aquasil C(18) analytical column (4.6 × 150 mm, 5 µm, Thermo Scientific, Rockford, IL, USA) with isocratic elution using a mobile phase containing acetonitrile and water with an addition of 0.1% of formic acid. Detection was achieved by an Applied Biosystems MDS Sciex (Concord, Ontario, Canada) API 2000 triple quadrupole mass spectrometer. Electrospray ionization was used for ion production. The limit of detection in the single ion monitoring mode was found to be 10 ng/mL. The limit of quantification was 50 ng/mL. The precision and accuracy for both within-day and between-day determination of 1,4-dimethylpyridinium was 2.4-7.56 and 90.93-111.48%. The results of this analytical method validation allow pharmacokinetic studies to be carried out in rats. The method was used for the pilot study of the pharmacokinetic behavior of 1,4-DMP in rats after intravenous administration., (Copyright © 2012 John Wiley & Sons, Ltd.)

The degree of binding of a drug to plasma proteins has a significant effect on its distribution, elimination, and pharmacological effect since only the unbound fraction is available for distribution into extra-vascular space. The binding of DL76 (1-[3-(4-tert-butyl-phenoxy)propyl]piperidine) to bovine serum albumin (BSA) was studied in viitro by equilibrium dialysis at 37 degrees C and pH 7.4 over the concentration range of 0.32-317.18 microM and at a physiological protein concentration of 602 microM. Drug concentrations were determined by validated LC/MS/MS method. Nonlinear regression analyses of the data pointed to a single class of binding sites (m = 1) with a dissociation constant of DL76 equal 49.20 microM. Scatchard plot concave-down curve might indicate positive cooperativity, which was confirmed by the Hill plot with the slope higher than one.

Plasma protein binding of drugs may have significant effect on its pharmacodynamic, toxicological and pharmacokinetic properties, since only the free drug can pass across biological membrane and get to its specific site of action. Many drugs show a high affinity to albumin which is the most abundant plasma protein. In the present study capillary electrophoresis in the frontal analysis mode (CE/FA), as promising technique for assessment of drug-protein interaction was used. The free drug concentration was measured from height of the frontal peak and calculated based on the external drug standard in absence of protein. With a known concentration of total drug, the percentage of protein bound drug was determined. The binding parameters were also estimated based on the equilibrium dialysis experiment which is considered to be a reference method. This study was designed to examine the interaction of dexamethasone sodium phosphate (DXM) with BSA and HSA under simulated physiological conditions (pH 7.4, 67 mM phosphate buffer, I = 0.17). Using fixed, at physiological level, HSA and BSA concentrations and increasing DXM concentrations, the number of binding sites (n) and binding constant (K(a) ) was calculated from both nonlinear regression fitting and Scatchard Plot. Despite some differences, it can be concluded that the CE/FA is comparable with equilibrium dialysis, but since the first one offers advantages such as low sample consumption, short analysis time, and high separation efficiency, it can be used in high-throughput screening of drug protein binding at the early stage of drug discovery. Interspecies differences in binding of a drug to albumins have been observed and it should be taken into account in interpretation of the results., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Methylnicotinamide (MNA) displays vasoprotective activity, however, the regulation of the activity of nicotinamide-N-methyltransferase (NNMT), is largely unknown. We analyze a possible involvement of IL-6 in the activation of NNMT-MNA pathway during an endurance exercise., Methods: FVB, C57Bl/6J IL6(+/+) and C57Bl/6J IL-6(-/-) mice were subjected to the single bout of endurance exercise consisting of 90 min of swimming. Thereafter, exercise-induced changes in NNMT activity in the liver as well as concomitant changes in the concentration of MNA and its further metabolites in plasma were analyzed., Results: In two strains of mice (FVB and C57Bl/6J IL6(+/+)) 90 min of swimming resulted in approximately 2-3 folds increase in NNMT activity (from 0.14 ± 0.03 to 0.421 ± 0.02 pmol/min/mg, p < 0.05 and from 0.2 ± 0.06 to 0.35 ± 0.07 pmol/min/mg, p < 0.01, respectively) and concomitant increase in the plasma concentration of MNA (from 157 ± 15.06 to 230 ± 16.2 ng/ml, p < 0.01, and from 77.05 ± 14.6 ng/ml to 152.55 ± 58.4 ng/ml; p < 0.01, respectively). However, in C57Bl/6J IL-6(-/-) mice 90 min of swimming did not change liver NNMT activity (from 0.25 ± 0.07 to 0.23 ± 0.06 pmol/min/mg), while MNA concentration in plasma rose approximately two-fold (from 65.3 ± 30.9 ng/ml to 124.8 ± 35.8 ng/ml; p < 0.05)., Conclusions: We demonstrated for the first time that NNMT - MNA pathway is activated by a single bout of endurance exercise. Interestingly, exercise-induced activation of NNMT in the liver involves IL-6, while the rise in MNA concentration in plasma was partially IL-6-independent. Taking into the consideration the pharmacological activity of MNA, IL-6-dependent and IL-6-independent activation of NNMT, may contribute to the exercise capacity. The physiological role of NNMT in the exercise warrant further studies.

A sensitive and specific liquid chromatography electrospray ionisation-tandem mass spectrometry method for determination of new non-imidazole histamine H(3) receptor antagonist 1-[3-(4-tert-butylphenoxy)propyl]piperidine (DL76) in rat serum has been developed and validated. Chromatography was performed on a XBridge™ C18 analytical column (2.1 × 30 mm, 3.5 µm, Waters, Ireland) with gradient elution using a mobile phase containing acetonitrile and water with an addition of 0.1% of formic acid. Detection was achieved by an Applied Biosystems MDS Sciex (Concord, Ontario, Canada) API 2000 triple quadrupole mass spectrometer. Electrospray ionization (ESI) was used for ion production. The limit of detection in the SRM mode was found to be 0.5 ng mL(-1). The limit of quantification was 1 ng mL(-1). The precision and accuracy for both intra- and inter-day determination of DL76 ranged from 1.65 to 15.09% and from 88.74 to 113.43%. The results of this analytical method validation allow to carry out pharmacokinetic studies in rats. The method was used for the pilot study of the pharmacokinetic behavior of DL76 in rats after intravenous administration.

Nicotinamide N-methyltransferase (NNMT), which converts nicotinamide (NA) to 1-methylnicotinamide (MNA), is up-regulated in the cirrhotic liver. Because MNA displays PGI(2)-dependent anti-inflammatory effects, the up-regulation of NNMT may play a regulatory role in liver inflammation. In the present work, we analyzed changes in NNMT activity in the liver and concomitant changes in the concentration of endogenous MNA in plasma in T-cell dependent hepatitis induced by concanavalin A (ConA) in BALB/c mice. Furthermore, we tested whether exogenous MNA possessed a protective effect against ConA-induced hepatitis. Development of liver injury induced by ConA (10 mg/kg, iv) was characterized by measurements of plasma concentration of alanine aminotransaminase (ALT), inflammatory cytokines (INF gamma and TNFalpha) and by histopathological examination. ConA-induced hepatitis was characterized by an early activation of inflammatory cytokines (IFN gamma; from below 0.05 ng/ml to 23.72 +/- 8.80 ng/ml; TNFalpha;from 0.07 +/- 0.01 ng/ml to 0.71 +/- 0.12 ng/ml, 2 h after ConA), an elevation of ALT (from 40.65 +/- 3.2 U/l to 5,092.20 +/- 1,129.05 U/l, 8 h after ConA) and by morphological signs of severe liver inflammation and injury (24 h after ConA). In mice injected with ConA, NNMT activity in the liver was up-regulated approximately 2-fold to 3-fold, 8-24 h after ConA injection. The concentration of MNA and its metabolites (Met-2PY and Met-4PY) in plasma were elevated approximately 2-fold 8 h after ConA injection. Exogenous MNA (100 mg/kg, iv) diminished ConA-induced liver injury, and this effect was reversed by an antagonist of the prostacyclin receptor, RO 3244794 (10 mg/kg, po). In conclusion, the present study demonstrated that hepatic NNMT activity and MNA concentration in plasma significantly increased during the progression of ConA-induced hepatitis in mice. This response may play a hepatoprotective role compatible with the PGI(2)-releasing properties of MNA.

A sensitive and specific liquid chromatography electrospray ionization-tandem mass spectrometry method for the simultaneous quantitation of nicotinic acid (NicA) and its metabolites nicotinamide (NA), 1-methylnicotinamide (MNA), 1-methyl-2-pyridone-5-carboxamide (M2PY) and 1-methyl-4-pyridone-5-carboxamide (M4PY) in rat plasma has been developed and validated. As an internal standard, 6-chloronicotinamide was used. The samples (100 microL) were subjected to deproteinization with acetonitrile (200 microL) and then, after centrifugation, 150 microL of the supernatant was transferred into conical vial and evaporated. Dry residue was reconstituted in 100 microL of the ACN/water (10:90, v/v) mixture. Chromatography was performed on a Waters Spherisorb 5 microm CNRP 4.6 x 150 mm analytical column with gradient elution using a mobile phase containing acetonitrile and water with 0.1% of formic acid. The full separation of all compounds was achieved within 15 min of analysis. Detection was performed by an Applied Biosystems MDS Sciex API 2000 triple quadrupole mass spectrometer set at unit resolution. The mass spectrometer was operated in the selected reactions monitoring mode (SRM), monitoring the transition of the protonated molecular ions m/z 153-110 for M2PY, 153-136 for M4PY, 124-80 for NicA, 123-80 for NA and 137-94 for MNA. The mass spectrometric conditions were optimized for each compound by continuously infusing the standard solution at the rate of 5 microL/min using a Harvard infusion pump. Electrospray ionization (ESI) was used for ion production. The instrument was coupled to an Agilent 1100 LC system. The precision and accuracy for both intra- and inter-day determination of all analytes ranged from 1.3% to 13.3% and from 94.43% to 110.88%. No significant matrix effect (ME) was observed. Stability of compounds was established in a battery of stability studies, i.e. bench-top, autosampler and long-term storage stability as well as freeze/thaw cycles. The method proved to be suitable for various applications. In particular using this method we detected increased concentration of MNA and its metabolites in rat plasma after treatment with exogenous MNA (100 mg/kg), as well as increased concentration of endogenous NA and MNA in rat plasma in the early phase of hypertriglyceridemia development in rats fed high-fructose diet., (2010 Elsevier B.V. All rights reserved.)

A sensitive and specific liquid chromatography electrospray ionization-tandem mass spectrometry method for the enantioselective determination of the novel beta-adrenolytic compound, 1-(1-H-indol-4-yloxy)-3-{[2-(2-methoxyphenoxy)ethylo]amino} propan-2-ol, in rat plasma has been developed and validated. Chromatography was performed on a reversed-phase Chiralcel OD-RH analytical column (150x4.6 mm, 5 microm, Daicel Chemical Industries, Tokyo, Japan) with isocratic elution using a mobile phase containing acetonitrile and water with 0.01% formic acid. Detection was achieved by an Applied Biosystems MDS Sciex (Concord, Ontario, Canada) API 2000 triple quadrupole mass spectrometer. Electrospray ionization (ESI) was used for ion production. The limit of detection in the MRM mode was found to be 1.25 ng/ml. The limit of quantification of both enantiomers was 2.5 ng/ml. The precision and accuracy for both intra- and inter-day determination of 2F109 enantiomers ranged from 2.6 to 12% and from 89.1 to 107.1%. This analytical method allowed us to carry out pharmacokinetic studies in rats. Our findings demonstrate that 2F109 shows stereoselective disposition in rat plasma after i.v. administration. The terminal half-lives of (+)-(R)-2F109 and (-)-(S)-2F109 were 33.5 and 42.6 min, respectively. The AUC0-inf of (+)-(R)-2F109 exceeded that of (-)-(S)-2F109., (Copyright (c) 2007 Wiley-Liss, Inc.)