Neuroprotective effect of guanosine against glutamate-induced cell death in rat hippocampal slices is mediated by the phosphatidylinositol-3 kinase/Akt/ glycogen synthase kinase 3ß pathway activation and inducible nitric oxide synthase inhibition.

Excitotoxicity and cell death induced by glutamate are involved in many neurodegenerative disorders. We have previously demonstrated that excitotoxicity induced by millimolar concentrations of glutamate in hippocampal slices involves apoptotic features and glutamate-induced glutamate release. Guanosine, an endogenous guanine nucleoside, prevents excitotoxicity by its ability to modulate glutamate transport. In this study, we have evaluated the neuroprotective effect of guanosine against glutamate-induced toxicity in hippocampal slices and the mechanism involved in such an effect. We have found that guanosine (100 µM) was neuroprotective against 1 mM glutamate-induced cell death through the inhibition of glutamate release induced by glutamate. Guanosine also induced the phosphorylation and, thus, activation of protein kinase B (PKB/Akt), a downstream target of phosphatidylinositol-3 kinase (PI3K), as well as phosphorylation of glycogen synthase kinase 3ß, which has been reported to be inactivated by Akt after phosphorylation at Ser9. Glutamate treated hippocampal slices showed increased inducible nitric oxide synthase (iNOS) expression that was prevented by guanosine. Slices preincubated with SNAP (an NO donor), inhibited the protective effect of guanosine. LY294002 (30 µM), a PI3K inhibitor, attenuated guanosine-induced neuroprotection, guanosine prevention of glutamate release, and guanosine-induced GSK3ß(Ser9) phosphorylation but not guanosine reduction of glutamate-induced iNOS expression. Taken together, the results of this study show that guanosine protects hippocampal slices by a mechanism that involves the PI3K/Akt/GSK3ß(Ser9) pathway and prevention of glutamate-induced glutamate release. Furthermore, guanosine also reduces glutamate-induced iNOS by a PI3K/Akt-independent mechanism.

Involvement of glutathione, ERK1/2 phosphorylation and BDNF expression in the antidepressant-like effect of zinc in rats.

We investigated the antidepressant-like effect of zinc chloride (zinc) administered acutely during 7 days (i.p. route), or chronically during 30 days (oral route) in the forced swimming test (FST) in rats. It was also investigated whether the antidepressant-like effect of zinc is associated with changes in the glutathione antioxidant system in the Wistar rat brain. Animals receiving a single zinc dose (5, 15 and 30 mg/kg, i.p.) 24 h prior to analysis showed no changes in the FST, but glutathione reductase and glutathione S-transferase activity were reduced in the hippocampus and cerebral cortex. This treatment did not, however, affect the glutathione status (GSH and GSSG) in both brain structures. The 7-day zinc treatment (1, 5 and 15 mg/kg, i.p.) caused a mild though significant antidepressant-like effect in the FST at the highest dosing, without affecting the glutathione antioxidant system. Finally, a consistent antidepressant-like effect was achieved in the FST after chronic (30 days) zinc treatment (300 mg/L, p.o.). This was accompanied by a significant increase in total glutathione levels in the hippocampus and cerebral cortex. The good response to oral treatment in the FST led us to investigate other variables, such as ERK phosphorylation and BDNF expression. Similar to therapeutic antidepressants, zinc in chronic oral treatment produced an increase in ERK phosphorylation and BDNF expression in the cerebral cortex. It is our hypothesis that up-regulation of neuroprotective effectors (GSH, ERK and BDNF) may be related to the antidepressant properties of zinc, but this will require additional work to be confirmed.

Effects of potassium channel inhibitors in the forced swimming test: possible involvement of L-arginine-nitric oxide-soluble guanylate cyclase pathway.

The effects of inhibitors of different subtypes of potassium (K+) channels were investigated in the mouse forced swimming test (FST). The treatment of animals with tetraethylammonium (TEA, a non-specific inhibitor of potassium channels, 0.25-2.5 ng/site, intracerebroventricular, i.c.v.), glibenclamide (an ATP-sensitive potassium channels (K(ATP) inhibitor, 0.05-5 ng/site, i.c.v.), apamine (a small conductance calcium-activated potassium channels inhibitor (SKCa), 0.1-1 ng/site, i.c.v.), charybdotoxin (a large- (big, BK) and intermediate- (IK) conductance calcium-activated potassium channels inhibitor, 2.5-25 ng/site, i.c.v.) produced an anti-depressant-like effect in the FST. At the highest effective doses, none of the drugs affected the locomotor activity in an open-field. Besides that, the pre-treatment of animals with l-arginine (a nitric oxide (NO) precursor, 750 mg/kg, intraperitoneal, i.p.) or sildenafil (a specific phosphodiesterase type 5 (PDE5) inhibitor, 5 mg/kg, i.p.) prevented the anti-depressant-like effect of all K+ channel inhibitors. The present results demonstrate that the decrease in the immobility time in the FST elicited by the inhibition of several subtypes of K+ channels is also dependent on the inhibition of NO-cGMP synthesis.

Evidence for the involvement of the opioid system in the agmatine antidepressant-like effect in the forced swimming test.

This study investigated the involvement of the opioid system in the antidepressant-like effect of agmatine in the mouse forced swimming test (FST). The antidepressant-like effects of agmatine (10 mg/kg, i.p.), as well as those of fluoxetine (32 mg/kg, i.p, a selective serotonin reuptake inhibitor, SSRI) or morphine (5 mg/kg, s.c., a nonselective opioid receptor agonist) in the FST was completely blocked by pretreatment of mice with naloxone (1 mg/kg, i.p., a nonselective opioid receptor antagonist). Pretreatment of mice with naltrindole (3 mg/kg, i.p., a selective delta-opioid receptor antagonist), clocinnamox (1 mg/kg, i.p., an irreversible mu-opioid receptor antagonist), but not with 2-(3,4-dichlorophenyl)-N-methyl-N-[(1S)-1-(3-isothiocyanatophenyl)-2-(1-pyrrolidinyl)ethyl]acetamide (DIPPA; 1 mg/kg, i.p., a selective kappa-opioid receptor antagonist) completely blocked the anti-immobility effect of agmatine (10 mg/kg, i.p.) in the FST. These results firstly demonstrate that the antidepressant-like effects of agmatine in the FST seem to be mediated, at least in part, by an interaction with the opioid system, that involves an activation of delta- and mu-opioid receptors.

Involvement of 5-HT1A receptors in the antidepressant-like effect of adenosine in the mouse forced swimming test.

This study investigated the involvement of 5-HT1 and 5-HT2 receptors in the antidepressant-like effect of adenosine in the mouse forced swimming test (FST). The pre-treatment of mice with PCPA (100mg/kg, i.p., an inhibitor of serotonin synthesis, for four consecutive days), NAN-190 (0.5mg/kg, i.p., a 5-HT1A receptor antagonist), pindolol (32 mg/kg, i.p., a 5-HT1A/1B receptor/beta-adrenoceptor antagonist) or WAY100635 (0.1 and 0.3mg/kg, s.c., a selective 5-HT1A receptor antagonist), but not with ketanserin (5mg/kg, i.p., a 5-HT2A/2C receptor antagonist), prevented the antidepressant-like effect of adenosine (10mg/kg, i.p.) in the FST. Moreover, the pre-treatment of animals with WAY100635 (0.1mg/kg, s.c.) blocked the decrease in immobility time in the FST elicited by adenosine (5 or 10mg/kg, i.p.), but produced a synergistic effect with a sub-effective dose of adenosine (1mg/kg, i.p.) and did not cause any alteration at the highest dose of adenosine administered (50mg/kg, i.p.). Adenosine (1mg/kg, i.p.) produced a synergistic antidepressant-like effect with pindolol (32 mg/kg), NAN-190 (0.5mg/kg, i.p.), WAY100635 (0.03 mg/kg, s.c.), 8-OH-DPAT (1mg/kg, i.p., a 5-HT1A receptor agonist), but not with DOI (1mg/kg, i.p., a preferential 5-HT2A receptor agonist) or ketanserin. The pre-treatment of mice with DPCPX (2mg/kg, i.p., a selective adenosine A1 receptor antagonist) or ZM241385 (1mg/kg, i.p., a selective adenosine A2A receptor antagonist) did not prevent the effect of fluoxetine (32 mg/kg, i.p., a preferential serotonin reuptake inhibitor) in the FST. Besides that, adenosine (1mg/kg, i.p.) did not produce a synergistic antidepressant-like effect with fluoxetine (10mg/kg, i.p.). Taken together, the results indicate that the antidepressant-like effect of adenosine in the FST appears to be mediated, at least in part, by an interaction with 5-HT1A receptors.