Guanosine-Mediated Anxiolytic-Like Effect: Interplay with Adenosine A1 and A2A Receptors.

Acute or chronic administration of guanosine (GUO) induces anxiolytic-like effects, for which the adenosine (ADO) system involvement has been postulated yet without a direct experimental evidence. Thus, we aimed to investigate whether adenosine receptors (ARs) are involved in the GUO-mediated anxiolytic-like effect, evaluated by three anxiety-related paradigms in rats. First, we confirmed that acute treatment with GUO exerts an anxiolytic-like effect. Subsequently, we investigated the effects of pretreatment with ADO or A1R (CPA, CCPA) or A2AR (CGS21680) agonists 10 min prior to GUO on a GUO-induced anxiolytic-like effect. All the combined treatments blocked the GUO anxiolytic-like effect, whereas when administered alone, each compound was ineffective as compared to the control group. Interestingly, the pretreatment with nonselective antagonist caffeine or selective A1R (DPCPX) or A2AR (ZM241385) antagonists did not modify the GUO-induced anxiolytic-like effect. Finally, binding assay performed in hippocampal membranes showed that [3H]GUO binding became saturable at 100-300 nM, suggesting the existence of a putative GUO binding site. In competition experiments, ADO showed a potency order similar to GUO in displacing [3H]GUO binding, whereas AR selective agonists, CPA and CGS21680, partially displaced [3H]GUO binding, but the sum of the two effects was able to displace [3H]GUO binding to the same extent of ADO alone. Overall, our results strengthen previous data supporting GUO-mediated anxiolytic-like effects, add new evidence that these effects are blocked by A1R and A2AR agonists and pave, although they do not elucidate the mechanism of GUO and ADO receptor interaction, for a better characterization of GUO binding sites in ARs.

Guanosine may increase absence epileptic activity by means of A2A adenosine receptors in Wistar Albino Glaxo Rijswijk rats.

The non-adenosine nucleoside guanosine (Guo) was demonstrated to decrease quinolinic acid(QA)-induced seizures, spontaneously emerged absence epileptic seizures and lipopolysaccharide(LPS)-evoked induction of absence epileptic seizures suggesting its antiepileptic potential. It was also described previously that intraperitoneal (i.p.) injection of 20 and 50mg/kg Guo decreased the number of spike-wave discharges (SWDs) in a well investigated model of human absence epilepsy, the Wistar Albino Glaxo Rijswijk (WAG/Rij) rats during 4th (20mg/kg Guo) and 3rd as well as 4th (50mg/kg Guo) measuring hours. Guanosine can potentially decrease SWD number by means of its putative receptors but absence epileptic activity changing effects of Guo by means of increased extracellular adenosine (Ado) cannot be excluded. An increase in the dose of i.p. injected Guo is limited by its low solubility in saline, therefore, we addressed in the present study whether higher doses of Guo, diluted in sodium hydroxide (NaOH) solution, have more potent antiepileptic effect in WAG/Rij rats. We confirmed that i.p. 50mg/kg Guo decreased but, surprisingly, i.p. 100mg/kg Guo enhanced the number of SWDs in WAG/Rij rats. Combined i.p. injection of a non-selective Ado receptor antagonist theophylline (5mg/kg) or a selective Ado A2A receptor (A2AR) antagonist SCH 58261 (7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine) (1mg/kg) and a cyclooxygenase 1 and 2/COX-1 and COX-2 inhibitor indomethacin (10mg/kg) with 100mg/kg Guo decreased the SWD number compared to i.p. 100mg/kg Guo alone. The results suggest that i.p. 100mg/kg Guo can increase SWD number by means of the adenosinergic system.

Intervention of Toll-like receptor-mediated human innate immunity and inflammation by synthetic compounds and naturally occurring products.

Human innate immunity plays a pivotal role in host defense against various microbial challenges. Mediated by a family of Toll-like-receptors (TLR) and associated intracellular downstream signaling molecules, human innate immunity can specifically recognize diverse microbial products and many other non-microbial environmental cues. Beyond its role of providing first line of defense, activation of innate immunity signaling can lead to expression of diverse pro- and anti- inflammatory mediators, which are critical for regulating various cell and tissue metabolism. Alteration in innate immunity signaling may therefore lead to infection and inflammatory diseases such as atherosclerosis, diabetes, and cancer. TLR receptors as well as intracellular signaling proteins can serve as therapeutic targets for treating various inflammatory diseases. Several synthetic ligands of TLR receptors such as lipid A analogs, poly(I:C), loxoribine, oligodeoxynucleotides have been shown to be effective in regulating innate immune response. This review discusses the potential, challenge, and recent progress of developing synthetic as well as naturally occurring TLR ligands in regulating innate immunity and treating inflammatory diseases.

Oral administration of guanosine impairs inhibitory avoidance performance in rats and mice.

Extracellular guanine-based purines, mainly the nucleoside guanosine, have recently been shown to exert neuroprotective effects, which seem to be related to antagonism of the glutamatergic system. In this study, we investigated the effects of acute oral administration of guanosine on inhibitory avoidance task in rats and mice. We also studied its effects on locomotor activity, anxiety-related behaviors and mechanisms of action involving the purinergic system. Guanosine (2.0 and 7.5mg/kg, per os), administered 75min pretraining, dose-dependently impaired retention of the inhibitory avoidance task in rats and mice, an effect not prevented by the adenosine receptor antagonist caffeine. Guanosine presented no effects on locomotor activity and anxiety-related behaviors. This amnesic effect of guanosine may be compatible with inhibition of glutamatergic system and seems to be not mediated by adenosine.

Cyclic GMP protein kinase activity is reduced in thyroxine-induced hypertrophic cardiac myocytes.

1. We tested the hypothesis that the cGMP-dependent protein kinase has major negative functional effects in cardiac myocytes and that the importance of this pathway is reduced in thyroxine (T4; 0.5 mg/kg per day for 16 days) hypertrophic myocytes. 2. Using isolated ventricular myocytes from control (n = 7) and T4-treated (n = 9) rabbit hypertrophic hearts, myocyte shortening was studied with a video edge detector. Oxygen consumption was measured using O2 electrodes. Protein phosphorylation was measured autoradiographically. 3. Data were collected following treatment with: (i) 8-(4-chlorophenylthio)guanosine-3',5'-monophosphate (PCPT; 10-7 or 10-5 mol/L); (ii) 8-bromo-cAMP (10-5 mol/L) followed by PCPT; (iii) beta-phenyl-1,N2-etheno-8-bromoguanosine-3',5'-monophosphorothioate, SP-isomer (SP; 10-7 or 10-5 mol/L); or (iv) 8-bromo-cAMP (10-5 mol/L) followed by SP. 4. There were no significant differences between groups in baseline percentage shortening (Pcs; 4.9 +/- 0.2 vs 5.6 +/- 0.4% for control and T4 groups, respectively) and maximal rate of shortening (Rs; 64.8 +/- 5.9 vs 79.9 +/- 7.1 micro m/ s for control and T4 groups, respectively). Both SP and PCPT decreased Pcs (-43 vs-21% for control and T4 groups, respectively) and Rs (-36 vs-22% for control and T4 groups, respectively), but the effect was significantly reduced in T4 myocytes. 8-Bromo-cAMP similarly increased Pcs (28 vs 23% for control and T4 groups, respectively) and Rs (20 vs 19% for control and T4 groups, respectively). After 8-bromo-cAMP, SP and PCPT decreased Pcs (-34%) and Rs (-29%) less in the control group. However, the effects of these drugs were not altered in T4 myocytes (Pcs -24%; Rs -22%). Both PCPT and cAMP phosphorylated the same five protein bands. In T4 myocytes, these five bands were enhanced less. 5. We conclude that, in control ventricular myocytes, the cGMP-dependent protein kinase exerted major negative functional effects but, in T4-induced hypertrophic myocytes, the importance of this pathway was reduced and the interaction between cAMP and the cGMP protein kinase was diminished.

Enhancement by O6-benzyl-N2-acetylguanosine of N'-[2-chloroethyl]-N-[2-(methylsulphonyl)ethyl]-N'-nitrosourea therapeutic index on nude mice bearing resistant human melanoma.

The exposure of cells to O6-benzyl-N2-acetylguanosine (BNAG) and several guanine derivatives is known to reduce the activity of O6-alkylguanine-DNA alkyltransferase (MGMT) and to enhance the sensitivity of Mer+ (methyl enzyme repair positive) tumour cells to chloroethylnitrosoureas (CENUs) in vitro and in vivo. High water solubility and the pharmacokinetic properties of BNAG make it a candidate for simultaneous administration with CENUs by the i.v. route in human clinical use. In vivo we have shown previously that BNAG significantly increases the efficiency of N'-[2-chloroethyl]-N-[2-(methylsulphonyl)ethyl]-N'-nitrosourea (cystemustine) against M4Beu melanoma cells (Mer+) through its cytostatic activity by the i.p. route, but also increases its toxicity. To investigate the toxicity of BNAG and cystemustine when administered simultaneously in mice, we compared the maximum tolerated dose and LD50 doses of cystemustine alone or in combination with 40 mg kg(-1) BNAG by the i.p. route. The toxicity of cystemustine was enhanced by a factor of almost 1.44 when combined with BNAG. To compare the therapeutic index of cystemustine alone and the cystemustine/BNAG combination, pharmacological tests were carried out in nude mice bearing Mer+ M4Beu human melanoma cells. Isotoxic doses were calculated using the 1.44 ratio. The treatments were administered three times by the i.v. route on days 1, 5 and 9 after s.c. inoculation of tumour cells. Although the toxicities of the treatments were equal, BNAG strongly enhanced tumour growth inhibition. These results demonstrate the increase of the therapeutic index of cystemustine by BNAG and justify the use of BNAG to enhance nitrosourea efficiency in vivo by i.v. co-injection.

Clinical studies of beta-thioguanine deoxyriboside alone and in combination with arabinosyl cytosine.

Beta-thioguanine deoxyriboside (betaTGdR) is a purine nucleoside derivative which was studied alone or in combination with arabinosyl cytosine (Ara-C) in patients with solid tumors and acute leukemia. No significant responses were observed in 22 patients with solid tumors. The response rate with betaTGdR alone in acute leukemia was 26% and in combination with Ara-C was 24%. Responses were generally of short duration. Toxicity included myelosuppression, nausea, stomatitis, hyperpigmentation, photosensitivity, and liver function abnormalities.