Acute and Chronic Exposure to Linagliptin, a Selective Inhibitor of Dipeptidyl Peptidase-4 (DPP-4), Has an Effect on Dopamine, Serotonin and Noradrenaline Level in the Striatum and Hippocampus of Rats.

Linagliptin is a selective dipeptidyl peptidase-4 (DPP-4) inhibitor that indirectly elevates the glucagon-like peptide-1 (GLP-1) level. The aim of the present study was to check whether linagliptin has an influence on neurotransmission in rat brain. Rats were acutely and chronically exposed to linagliptin (10 and 20 mg/kg, intraperitoneally (i.p.)). Twenty-four hours later, the striatum and hippocampus were selected for further studies. In neurochemical experiments, using high-performance liquid chromatography with electrochemical detection (HPLC-ED), the concentrations of three major neurotransmitters-dopamine, serotonin and noradrenaline-and their metabolites were measured. The analysis of mRNA expression of dopamine (D1 and D2), serotonin (5-HT-1 and 5-HT-2) and noradrenaline (α1 and α2a) receptors was also investigated using real-time quantitative reverse transcription polymerase chain reaction (RQ-PCR) in the same brain areas. Linagliptin has the ability to influence the dopaminergic system. In the striatum, the elevation of dopamine and its metabolites was observed after repeated administration of that linagliptin, and in the hippocampus, a reduction in dopamine metabolism was demonstrated. Acute linagliptin exposure increases the serotonin level in both areas, while after chronic linagliptin administration a tendency for the mRNA expression of serotoninergic receptors (5-HT1A and 5-HT2A) to increase was observed. A single instance of exposure to linagliptin significantly modified the noradrenaline level in the striatum and intensified noradrenaline turnover in the hippocampus. The recognition of the interactions in the brain between DPP-4 inhibitors and neurotransmitters and/or receptors is a crucial step for finding novel discoveries in the pharmacology of DPP-4 inhibitors and raises hope for further applications of DPP-4 inhibitors in clinical practices.

Chronic and Cycling Hypoxia: Drivers of Cancer Chronic Inflammation through HIF-1 and NF-κB Activation: A Review of the Molecular Mechanisms.

Chronic (continuous, non-interrupted) hypoxia and cycling (intermittent, transient) hypoxia are two types of hypoxia occurring in malignant tumors. They are both associated with the activation of hypoxia-inducible factor-1 (HIF-1) and nuclear factor κB (NF-κB), which induce changes in gene expression. This paper discusses in detail the mechanisms of activation of these two transcription factors in chronic and cycling hypoxia and the crosstalk between both signaling pathways. In particular, it focuses on the importance of reactive oxygen species (ROS), reactive nitrogen species (RNS) together with nitric oxide synthase, acetylation of HIF-1, and the action of MAPK cascades. The paper also discusses the importance of hypoxia in the formation of chronic low-grade inflammation in cancerous tumors. Finally, we discuss the effects of cycling hypoxia on the tumor microenvironment, in particular on the expression of VEGF-A, CCL2/MCP-1, CXCL1/GRO-α, CXCL8/IL-8, and COX-2 together with PGE2. These factors induce angiogenesis and recruit various cells into the tumor niche, including neutrophils and monocytes which, in the tumor, are transformed into tumor-associated neutrophils (TAN) and tumor-associated macrophages (TAM) that participate in tumorigenesis.

The role of linagliptin, a selective dipeptidyl peptidase-4 inhibitor, in the morphine rewarding effects in rats.

Linagliptin is a selective dipeptidyl peptidase-4 (DPP-4) inhibitor which suppresses the rapid degradation of endogenous glucagon-like peptide-1 (GLP-1). In clinical practice, it is used as an antidiabetic drug, but recent studies have confirmed its role in the activity of the central nervous system (CNS). The reported study focused on the role of linagliptin (10 and 20 mg/kg, ip) in the morphine rewarding effect, analyzing how the agent had influenced the conditioned place preference (CPP) in rats via the expression, acquisition, extinction and reinstatement of the morphine rewarding effect. The obtained results clearly demonstrated linagliptin to inhibit the expression and acquisition, to accelerate the extinction and, eventually, to reduce the reinstatement of morphine-induced CPP. The undertaken experiments significantly extended our knowledge on the mechanisms behind the morphine rewarding effect.

Phosphodiesterase inhibitors say NO to Alzheimer's disease.

Phosphodiesterases (PDEs) consisted of 11 subtypes (PDE1 to PDE11) and over 40 isoforms that regulate levels of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP), the second messengers in cell functions. PDE inhibitors (PDEIs) have been attractive therapeutic targets due to their involvement in diverse medical conditions, e.g. cardiovascular diseases, autoimmune diseases, Alzheimer's disease (AD), etc. Among them; AD with a complex pathology is a progressive neurodegenerative disorder which affect mostly senile people in the world and only symptomatic treatment particularly using cholinesterase inhibitors in clinic is available at the moment for AD. Consequently, novel treatment strategies towards AD are still searched extensively. Since PDEs are broadly expressed in the brain, PDEIs are considered to modulate neurodegenerative conditions through regulating cAMP and cGMP in the brain. In this sense, several synthetic or natural molecules inhibiting various PDE subtypes such as rolipram and roflumilast (PDE4 inhibitors), vinpocetine (PDE1 inhibitor), cilostazol and milrinone (PDE3 inhibitors), sildenafil and tadalafil (PDE5 inhibitors), etc have been reported showing encouraging results for the treatment of AD. In this review, PDE superfamily will be scrutinized from the view point of structural features, isoforms, functions and pharmacology particularly attributed to PDEs as target for AD therapy.

‬The expression of purinergic P2X4 and P2X7 receptors in selected mesolimbic structures during morphine withdrawal in rats.

Morphine is one of the most potent analgesics used in medicine and it's long-term use is associated with the risk of the state of dependence. The cessation of chronic morphine administration leads to withdrawal signs which are associated with neurotransmitter dysregulations within mesolimbic system. Adenosine 5'-triphosphate (ATP) and purinergic system play an important role in the activity of central nervous system (CNS). Purinergic receptors are widely distributed in neurons and glial cells throughout the CNS taking part in integration of functional activity between neurons, glial and vascular cells. In the present study the mRNA and protein expression of purinergic P2X4 and P2X7 receptors in selected mesolimbic structures (striatum, hippocampus and prefrontal cortex) during morphine withdrawal in rats was investigated by RT-PCR and Western Blot analysis. Two experimental models of morphine withdrawal were studied: single and repeated morphine withdrawal. We demonstrated that expression of P2X4 and P2X7 receptors was altered during morphine withdrawal period in rats. These alterations were varied in particular mesolimbic areas depending on the scheme of morphine administration. Our results extend the current knowledge on morphine withdrawal and for the first time high-light interactions between purinergic system and morphine withdrawal. It seems, the purinergic system may be a new, valuable tool in searching for a new strategy of management of opioid dependence.

SB-334867 (an Orexin-1 Receptor Antagonist) Effects on Morphine-Induced Sensitization in Mice-a View on Receptor Mechanisms.

The present study focused upon the role of SB-334867, an orexin-1 receptor antagonist, in the acquisition of morphine-induced sensitization to locomotor activity in mice. Behavioral sensitization is an enhanced systemic reaction to the same dose of an addictive substance, which assumingly increases both the desire for the drug and the risk of relapse to addiction. Morphine-induced sensitization in mice was achieved by sporadic doses (five injections every 3 days) of morphine (10 mg/kg, i.p.), while a challenge dose of morphine (10 mg/kg) was injected 7 days later. In order to assess the impact of orexin system blockade on the acquisition of sensitization, SB-334867 was administered before each morphine injection, except the morphine challenge dose. The locomotor activity test was performed on each day of morphine administration. Brain structures (striatum, hippocampus, and prefrontal cortex) were collected after behavioral tests for molecular experiments in which mRNA expression of orexin, dopamine, and adenosine receptors was explored by the qRT-PCR technique. Additionally, the mRNA expression of markers, such as GFAP and Iba-1, was also analyzed by the same technique. SB-334867 inhibited the acquisition of morphine-induced sensitization to locomotor activity of mice. Significant alterations were observed in mRNA expression of orexin, dopamine, and adenosine receptors and in the expression of GFAP and Iba-1, showing a broad range of interactions in the mesolimbic system among orexin, dopamine, adenosine, and glial cells during behavioral sensitization. Summing up, the orexin system may be an effective measure to inhibit morphine-induced behavioral sensitization.

Neuroprotective effects of honokiol: from chemistry to medicine.

The incidence of neurological disorders is growing in developed countries together with increased lifespan. Nowadays, there are still no effective treatments for neurodegenerative pathologies, which make necessary to search for new therapeutic agents. Natural products, most of them used in traditional medicine, are considered promising alternatives for the treatment of neurodegenerative diseases. Honokiol is a natural bioactive phenylpropanoid compound, belonging to the class of neolignan, found in notable amounts in the bark of Magnolia tree, and has been reported to exert diverse pharmacological properties including neuroprotective activities. Honokiol can permeate the blood brain barrier and the blood-cerebrospinal fluid to increase its bioavailability in neurological tissues. Diverse studies have provided evidence on the neuroprotective effect of honokiol in the central nervous system, due to its potent antioxidant activity, and amelioration of the excitotoxicity mainly related to the blockade of glutamate receptors and reduction in neuroinflammation. In addition, recent studies suggest that honokiol can attenuate neurotoxicity exerted by abnormally aggregated Aß in Alzheimer's disease. The present work summarizes what is currently known concerning the neuroprotective effects of honokiol and its potential molecular mechanisms of action, which make it considered as a promising agent in the treatment and management of neurodegenerative diseases. © 2017 BioFactors, 43(6):760-769, 2017.

The adenosinergic system is involved in sensitization to morphine withdrawal signs in rats-neurochemical and molecular basis in dopaminergic system.

RATIONALE: Experimental data informs that not only do the dose and time duration of dependent drugs affect the severity of withdrawal episodes. Previous withdrawal experiences may intensify this process, which is referred as sensitization to withdrawal signs. Adenosine and dopamine (DA) receptors may be involved in this sensitization. OBJECTIVES: Rats were continuously and sporadically treated with increasing doses of morphine for 8 days. In rats, sporadically treated with morphine, morphine administration was modified by adding three morphine-free periods. Adenosine agonists were given during each of the morphine-free periods (six injections in total). On the 9th day, morphine was injected. One hour later, naloxone was administered to induce morphine withdrawal signs. Then, the animals were placed into cylinders and the number of jumpings was recorded. Next, the rats were decapitated and brain and brain structures (striatum, hippocampus, and prefrontal cortex) were dissected for neurochemical, molecular, and immunohistochemical experiments within DAergic pathways. RESULTS: We demonstrated that previous experiences of opioid withdrawal intensified subsequent withdrawal signs. Adenosine ligands attenuated the sensitization to withdrawal signs. In a neurochemical study, the release of DA and its metabolites was impaired in all structures. Significant alterations were also observed in mRNA and protein expression of DA receptors. CONCLUSIONS: Results demonstrate that intermittent treatment with morphine induces alterations in the DAergic system which may be responsible for sensitization to morphine withdrawal signs. Although adenosine ligands attenuate this type of sensitization, they are not able to fully restore the physiological brain status.

Effects of perinatal exposure to lead (Pb) on purine receptor expression in the brain and gliosis in rats tolerant to morphine analgesia.

The aim of the present study was to investigate the molecular effects of perinatal exposure to lead (Pb) on protein and mRNA expression of purine receptors: P2X4, P2X7, adenosine receptor A1; and astrocytes (GFAP mRNA expression) and on microglia activation (Iba1 mRNA expression) in several structures of the mesolimbic system (striatum, hippocampus, prefrontal cortex) in rats expressing tolerance to the antinociceptive effect of morphine. Rat mothers were orally treated with 0.1% lead acetate from conception, through gestation, and postnatally, as well as to offspring up to day (PND) 28; subsequently molecular studies were conducted on adult (PND 60) male rats. Morphine tolerance developed more strongly in rats perinatally exposed to Pb. The analysis revealed a significant up-regulation of protein and mRNA P2X4 receptor expression in the striatum and prefrontal cortex but not in the hippocampus; P2X7 protein and mRNA receptor expression in the striatum and hippocampus, but not in the prefrontal cortex; A1 protein receptor expression in all investigated structures and A1 mRNA expression in the striatum and hippocampus; Iba1 mRNA expression in the striatum and hippocampus; and GFAP mRNA expression in the striatum and prefrontal cortex. Immunohistochemical analysis has also revealed significant alterations. Strong expressions of P2X4, P2X7, A1 receptors, astrocytes and microglia activation were observed in the hippocampus in Pb and/or morphine treated rats. The higher expression of purine receptors and glial cell activation are important markers of neuroinflammatory processes. Therefore, we conclude that Pb-induced neuroinflammation may be responsible for the intensification of morphine tolerance in the Pb-treated rats. Additionally, the dysregulation of A1 adenosine receptors, mainly in the hippocampus, may also be involved in the intensification of morphine tolerance in Pb-treated rats. Our study demonstrates the significant participation of environmental factors in addictive process; additionally, it shows the necessity of modification of addictive disorder with neuroprotective agents.

Effects of the adenosinergic system on the expression and acquisition of sensitization to conditioned place preference in morphine-conditioned rats.

In the presented study, we attempt to investigate if the sensitization to conditioned place preference (CPP) induced by low doses of morphine was developed in rats which have been previously conditioned with morphine. The experiments were performed in the CPP test. Firstly, it has been demonstrated that administration of ineffective dose of morphine on the 9th day induces the increase in time spent of rats at a morphine-paired compartment, confirming that sensitization to CPP has been developed in these animals. Secondly, it has been shown that stimulation of A1 receptor significantly inhibits the expression of morphine-induced of sensitization, and blockade of these receptors produces the opposite effect. Finally, it has been indicated that both stimulation and blockade of A1 and/or A2A receptors inhibit the acquisition of sensitization to CPP. The obtained results have strongly supported the significance of adenosinergic system in both expression and acquisition of studied sensitization. These results seem to be important for the identification of connections in the central nervous system which can help finding new strategies to attenuate rewarding action of morphine.

Attenuating effect of adenosine receptor agonists on the development of behavioral sensitization induced by sporadic treatment with morphine.

The aim of the study is to investigate the effect of adenosine receptor agonists on the development of morphine-induced sensitization to the locomotor activity of mice. Selective A1 (N6-cyclopentyladenosine - CPA) and A2A (2-p-(2-carboxyethyl) phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride - CGS 21680) adenosine receptor agonists or non-selective A1/A2A (5'-N-ethylcarboxamidoadenosine - NECA) adenosine agonists as representatives of adenosinergic drugs have been used in the experiment. Behavioral sensitization has been obtained by sporadic treatment with morphine (10.0mg/kg, i.p.). We have shown that adenosine receptor agonists co-administered with morphine significantly attenuates increase in the locomotor activity of mice evoked by challenge dose of morphine. These effects have been observed after stimulation of the selective A1 or A2A and non-selective A1/A2A adenosine receptors, namely both receptors were involved in morphine-induced sensitization. Thus, we have demonstrated that adenosine agonists are able to inhibit behavioral sensitization induced by sporadic applications of morphine.

Adenosinergic system is involved in development of diazepam tolerance in mice.

In the present study the effect of adenosinergic system on the development of diazepam tolerance to motor disturbances in mice was investigated. Diazepam tolerance was obtained by administration of diazepam at a dose of 5.0 mg/kg, s.c. for ten consecutive days. On the 1st and the 10th day of the experiment motor impairments were measured in two behavioural tests: rota-rod and chimney test. We showed that acute diazepam injection produced significant motor impairments in mice and that effect was decreased by repeated diazepam treatment, confirming the development of tolerance to the motor impairing effect of diazepam. We demonstrated that adenosine A(1) and/or A(2A) receptor agonists: CPA (0.025 and 0.05 mg/kg, i.p.), CGS 21680 (0.1 and 0.2 mg/kg, i.p.), NECA (0.005 and 0.01 mg/kg, i.p.) pretreatment with diazepam were able to attenuate the development of diazepam tolerance and adenosine receptor antagonists: DPCPX (1.0 and 3.0 mg/kg, i.p.), DMPX (3.0 and 6.0 mg/kg, i.p.) and caffeine (10.0 and 20.0mg/kg, i.p.) induced the opposite effect. The most apparent effects were obtained by non-selective agonist (NECA) and antagonist (caffeine) of adenosine receptors. We conclude that adenosinergic system plays an important role in mechanisms underlying the development of benzodiazepine tolerance.

Involvement of adenosine receptor agonists on the development of hypersensitivity to acute dose of morphine during morphine withdrawal period.

In the present study, the involvement of the selective adenosine A1 (CPA) and A2A (CGS 21680) and non-selective adenosine A1/A2A (NECA) receptor agonists on the development of hypersensitivity to acute morphine injection given during opiate withdrawal was investigated. Intraperitioneal (ip) injections of morphine at increasing doses (10, 20, 30, 40, 50 mg/kg) for 6 consecutive days produced a state of dependence. On the 6th day, in the morning, animals were injected with the last dose of morphine (50 mg/kg, ip). Each day, 20 min before each injection of morphine, adenosine receptor agonists were also administered. Seven days after cessation of the morphine treatment, on the 13th day of the experiment, all animals were challenged with a dose of morphine (10 mg/kg, ip). A clear increase in locomotor activity was observed, indicating that hypersensitivity had developed. Our study has demonstrated the presence of an attenuating effect of adenosinergic drugs, such as CGS 21680 and NECA, but not CPA, on the development of hypersensitivity. The results indicate that stimulation of the adenosine A2A receptor plays some role in modulating the neuroadaptive changes appearing during chronic opioid treatment and that adenosine A2A receptor agonists may serve as useful drugs in relapse protection. Our investigations focused on adenosine A2A agonists as possible vehicles for pharmacotherapy for morphine addiction.

Adenosine receptor agonists attenuate the development of diazepam withdrawal-induced sensitization in mice.

In the present study, the effects of adenosine agonists on the development of sensitization to withdrawal signs precipitated after sporadic treatment with diazepam, in mice, were investigated. To obtain the sensitization, the animals were divided into groups: continuously and sporadically treated with diazepam (15.0 mg/kg, s.c.). The adenosine receptor agonists (CPA, CGS 21,680 and NECA) were administered in sporadically diazepam treated mice during two diazepam-free periods. Concomitant administration of pentetrazole (55.0 mg/kg, s.c.) with flumazenil (5.0 mg/kg, i.p.) after the last injection of diazepam or vehicle, induced the withdrawal signs, such as clonic seizures, tonic convulsion and death episodes. The major finding of our experiments is attenuation of withdrawal signs in sensitized mice, inducing by all adenosine agonists. Only higher dose of CPA produced significantly decreased the number of withdrawal incidents, while both used doses of CGS 21,680 and NECA produced more clear effects. These results support the hypothesis that adenosinergic system is involved in the mechanisms of sensitization to the benzodiazepine withdrawal signs, and adenosine A(2A) receptors play more important role in that process.

Adenosine receptor antagonists intensify the benzodiazepine withdrawal signs in mice.

The aim of the present experiment was to assess the involvement of adenosine receptor antagonists in benzodiazepine (BDZ) withdrawal signs, observed as the seizure susceptibility in mice. The discontinuation of chronic treatment with temazepam or diazepam decreased seizure threshold (one of BDZ withdrawal signs). The concomitant application of subconvulsive dose of pentetrazole (55.0 mg/kg) with low dose of flumazenil (5.0 mg/kg) - a BDZ receptor antagonist, immediately induced BDZ withdrawal signs in these animals. The non-selective adenosine receptor antagonist (caffeine), and the selective adenosine A1 receptor antagonist (DPCPX), injected 15 min before the application of pentetrazole and flumazenil, were able to intensify BDZ withdrawal signs in mice. The most apparent effects were observed after administration of DPCPX, indicating that the adenosine A1 receptor may play a more important role in these effects. The obtained data demonstrate that the adenosinergic system is involved in BDZ withdrawal signs in mice, and adenosine A1 receptor plays an important role in this process.

Influence of adenosine receptor agonists on benzodiazepine withdrawal signs in mice.

The involvement of adenosine receptor agonists in benzodiazepine withdrawal signs was evaluated as the seizure susceptibility of mice. The concomitant administration of subthreshold dose of pentetrazole (55.0 or 60.0 mg/kg, s.c.) with flumazenil (10.0 mg/kg, i.p.) in mice chronically treated with temazepam or diazepam induced the appearance of withdrawal signs: clonic seizures, tonic convulsions and death episodes. The administration of the selective A1 (CPA-N6-cyclopentyladenosine), A2A (CGS 21680-2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride) and the non-selective A1/A2A (NECA-5'-N-ethylcarboxamidoadenosine) adenosine receptor agonists (i.p.) evoked the significant attenuation of benzodiazepine withdrawal signs, and these effects were more expressed in temazepam- than in diazepam-dependent mice. CPA has shown the most apparent and dose-dependent attenuating effect. The results confirm that adenosine A1 and A2A receptors are involved in benzodiazepine withdrawal signs, and adenosine A1 receptor plays a predominant role in this phenomenon.