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The present study examined the anti-hypernociceptive effects of agmatine (AGM) in acute and chronic models of behavioural pain in mice. Agmatine (30 mg/kg, i.p. 30 min early), produced time-dependent inhibition of mechanical hypernociception induced by Complete Freund's Adjuvant (CFA) injected in the mice paw (inhibition of 52+/-7%) after 4 h. Given chronically (twice a day) during 10 days, AGM significantly reversed the mechanical hypernociception caused by CFA (inhibition of 43+/-6% to 67+/-5%). Moreover, AGM also significantly reduced the mechanical hypernociception caused by partial sciatic nerve ligation (PSNL) during 6 h, with inhibition of 81+/-8%. In thermal hypernociception (cold stimuli) caused by PSNL the antinociceptive effect of AGM was prolonged by 4 h with inhibition of 97+/-3% observed 1 h after the treatment. Nevertheless, AGM failed to inhibit the paw oedema caused by CFA and the myeloperoxidase enzyme activity. Of note, AGM (10-100 mg/kg, i.p., 30 min before) also elicited a pronounced inhibition of the biting response induced by TNF-alpha and IL-1beta in mice, with mean ID(50) values of 61.3 mg/kg (47.7-78.6 mg/kg) and 30.4 mg/kg (18.6-49.8 mg/kg) and inhibitions of 75+/-5% and 66+/-6%, respectively. Together, present and previous findings show that AGM given systemically is effective in inhibiting mechanical and thermal hypernociception present in chronic inflammatory processes caused by CFA and also the neuropathic pain caused by PSNL.

The role of the cellular prion protein (PrP(c)) in neuronal functioning includes neuronal excitability, cellular adhesion, neurite outgrowth and maintenance. Here we investigated the putative involvement of the PrP(c) function on the nociceptive response using PrP(c) null (Prnp(0/0)) and wild-type (Prnp(+/+)) mice submitted to thermal and chemical models of nociception. PrP(c) null mice were more resistant than wild-type mice to thermal nociception of the tail-flick test. However, no significant difference was found on the hot plate test. In the acetic acid-induced visceral nociception, PrP(c) null mice showed an enhanced response when compared to wild-type mice. However, there was no difference between Prnp(0/0) and wild-type mice on glutamate- and formalin-induced licking behaviour and Freund's Complete Adjuvant (FCA)-induced mechanical allodynia. PrP(c) null mice developed significantly lower paw edema than wild-type mice. In addition, the visceral conditioning stimuli produced by a previous injection of acetic acid (20 days before testing) significantly reduced early and late phases of formalin-induced nociception in wild-type mice. In contrast, the same pre-treatment did not alter the formalin response in PrP(c) null mice. These results indicate a role of PrP(c) in the nociceptive transmission, including the thermal tail-flick test and visceral inflammatory nociception (acetic acid-induced abdominal constriction). Our findings show that PrP(c) is involved with a response mediated by inflammation (paw edema) and by visceral conditioning stimuli.

This study was designed to evaluate the role of spinal glutamatergic receptors in the antinociception elicited by agmatine in mice. Intraperitoneal (i.p.) administration of agmatine (1.0-100.0 mg/kg) dose dependently inhibited the nociceptive response induced by intrathecal (i.t.) injection of glutamate, N-methy-D-aspartate (NMDA) and (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD), with mean ID(50) values of 16.7, 6.8 and 27.0 mg/kg, respectively. However, agmatine completely failed to affect the nociception induced by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) or kainic acid (kainate). Agmatine injected by i.t. route (10-100 microg/site) also produced dose-related inhibition of NMDA- and trans-ACPD-induced biting response with mean ID(50) values of 29.6 and 36.0 mug/site, respectively. The nitric oxide synthase inhibitor N(omega)-nitro-L-arginine (l-NOARG) (75.0 mg/kg, i.p.) also consistently inhibited glutamate-, NMDA- and trans-ACPD-induced nociception (41 +/- 13, 100 and 83 +/- 6%, of inhibition, respectively) but had no effect on the same response caused by AMPA and kainate agonists. The selective NMDA receptor antagonist (5S,10R)-(+)-5-Methyl-10,11-dihydro-5H-dibenzo[a,d] (MK-801) at a low dose (0.05 mg/kg, i.p.) inhibited the nociceptive response caused by both glutamate and NMDA agonists (inhibitions of 35 +/- 1 and 72 +/- 2%, respectively). At a high dose, MK-801 (0.5 mg/kg, i.p.) significantly inhibited the biting response induced by i.t. administration of all the glutamatergic agonists tested: glutamate, AMPA, NMDA, kainate and trans-ACPD, with inhibitions of 49 +/- 8, 84 +/- 16, 84 +/- 3, 76 +/- 8 and 97 +/- 2%, respectively. Together, these results provide experimental evidence indicating that agmatine given systemically and spinally produce marked antinociception at spinal sites in mice. Furthermore, an interaction with glutamate receptors, namely NMDA and trans-ACPD, metabotropic and NMDA-ionotropic origin, by a mechanism similar to that of nitric oxide (NO) inhibitors, seems to account for the agmatine antinociceptive action.

This study investigated the involvement of 5-HT(1) and 5-HT(2) receptors in the antidepressant-like effect of agmatine in the mouse forced swimming test (FST). Pretreatment with p-chlorophenylalanine methyl ester (PCPA; 100 mg/kg, intraperitoneally (i.p.), an inhibitor of serotonin synthesis, for 4 consecutive days), methysergide (5 mg/kg, i.p., a serotonin (5-HT) antagonist), pindolol (32 mg/kg, i.p., a 5-HT(1A/1B) receptor/beta-adrenoceptor antagonist), N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridynyl)cyclohexanecarboxamide (WAY 100635; 0.3 mg/kg, subcutaneously (s.c.), a selective 5-HT(1A) receptor antagonist), 1-(2-methoxyphenyl)-4[-(2-phthalimido)butyl]piperazine) (NAN-190; 0.5 mg/kg, i.p., a 5-HT(1A) receptor antagonist), 1-(2-(1-pyrrolyl)-phenoxy)-3-isopropylamino-2-propanol (isamoltane; 2.5 mg/kg, i.p., a 5-HT(1B) receptor antagonist), cyproheptadine (3 mg/kg, i.p., a 5-HT(2) antagonist) or ketanserin (5 mg/kg, i.p., a 5-HT(2A/2C) receptor antagonist), but not with propranolol (2 mg/kg, i.p., a beta-adrenoceptor antagonist), prevented the effect of agmatine (10 mg/kg, i.p.) in the FST. A subeffective dose of agmatine (0.001 mg/kg, i.p.) produced a synergistic antidepressant-like effect with pindolol (32 mg/kg), NAN-190 (0.5 mg/kg, i.p.), WAY 100635 (0.03 mg/kg, s.c.), (+)-8-hydroxy-2-(di-n-propylamino)tetralin HBr (8-OH-DPAT; 0.01 mg/kg, i.p., a 5-HT(1A) receptor agonist), R(-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl (DOI; 1 mg/kg, i.p., a preferential 5-HT(2A) receptor agonist), or fluoxetine (10 mg/kg, i.p., a selective serotonin reuptake inhibitor, SSRI) but not with isamoltane (2.5 mg/kg, i.p.), ritanserin (4 mg/kg, i.p., a 5-HT(2A/2C) receptor antagonist) or ketanserin (5 mg/kg, i.p.). Taken together, the results firstly demonstrate that agmatine antidepressant-like effects in the FST seem to be mediated, at least in part, by an interaction with 5-HT(1A/1B) and 5-HT(2) receptors.

Recently, our laboratory has proposed the avulsion of rat brachial plexus as a new and reliable model for the study of neuropathic pain. In this model, the neuropathy can be detected even at distant sites from the injury, both in ipsilateral and contralateral hindpaws. The purpose of this study was to pharmacologically characterise this behavioural model of persistent peripheral neuropathic pain by assessing the effects of several analgesic drugs currently used in clinical practice. For this purpose, the effects of these drugs on the mechanical and cold allodynia were analysed 20-40 days after rat brachial plexus avulsion. Injection of saline, administered by the same route as the other drugs, did not significantly affect the nociceptive threshold either in sham-operated or in neuropathic rats. However, administration of the opioid analgesic morphine (5 mg/kg, s.c.), the alpha2 adrenoceptor agonist clonidine (300 microg/kg, i.p.), the NMDA receptor antagonist ketamine (25 mg/kg, i.p.) or the anticonvulsant drug gabapentin (70 mg/kg, p.o.) consistently reduced both mechanical and cold allodynia following avulsion of rat brachial plexus. The administration of the selective COX-2 inhibitor celecoxib (10 mg/kg, p.o.) blocked mechanical allodynia, but not cold allodynia, whereas the sodium channel blocker lidocaine (40 mg/kg, i.p.) attenuated only cold allodynia. The non-steroidal anti-inflammatory drug diclofenac (100 mg/kg, i.p.), the steroidal anti-inflammatory dexamethasone (1.5 mg/kg, i.p.) and the antidepressant imipramine (10 mg/kg, i.p.) all failed to significantly attenuate both mechanical and cold allodynia in the rats following avulsion of brachial plexus. These findings suggest that avulsion-associated mechanical and cold allodynia, two classic signs of persistent neuropathic pain, were consistently prevented by several analgesics currently available in clinical practice, namely morphine, clonidine, ketamine and gabapentin, and to a lesser extent by celecoxib and lidocaine. Therefore, this new proposed model of persistent nociception seems to be suitable for the study of the underlying mechanisms involved in neuropathic pain and for the identification of potential clinically relevant drugs to treat this aspect of peripheral neuropathy.

In the present study, we sought to characterise a behavioural model of persistent peripheral neuropathic pain produced by avulsion of the right brachial plexus in rats. In addition, we compared the effects of avulsion with those of ligation or crush injury of the brachial plexus. Avulsion and, to a lesser extent, ligation and crushing of brachial plexus caused a long-lasting (up to 90 days) and highly reproducible mechanical hyperalgesia, in both ipsilateral and contralateral hindpaws. However, the same injury did not produce thermal hyperalgesia. The avulsion and, to a lesser extent, ligation and crushing of the brachial plexus elicited a significant and long-lasting (up to 90 days) ipsilateral and contralateral cold and mechanical allodynia. Furthermore, the brachial plexus injury caused a significant decrease in functional activity of the forepaws as assessed in the grasping strength test, but did not alter the locomotor activity of the rats in the open field test in comparison with control or sham groups. Taken together these results show that avulsion of the brachial plexus in rat produces persistent mechanical and cold allodynia and mechanical hyperalgesia, and might represent a valuable method for understanding the mechanisms underlying the aetiology of neuropathic pain.

This study sought to establish whether sensory neuropeptides and the capsaicin-sensitive fibres are involved in the nociception and oedema formation caused by intraplantar (i.pl.) injection of glutamate into the mouse paw. The i.pl. co-injection of the selective neurokinin (NK) NK(2) (SR 48968, 0.05-0.5 nmol/paw), and to a lesser extent the selective NK(1) (FK 888, 0.25-1.0 nmol/paw) receptor antagonists, resulted in a significant inhibition of glutamate-induced nociception. The percentages of inhibition were 82 and 37%, respectively. In contrast, the selective NK(3) receptor antagonist (SR 142801, 0.25-1.0 nmol/paw) failed to significantly affect glutamate-induced nociception. SR 48968, but not FK 888 or SR 142801, significantly inhibited (36%) glutamate-induced paw oedema formation. The i.pl. injection of kinin B(1) receptor antagonist des-Arg(9)-[Leu(8)]-BK (0.2-0.8 nmol/paw), but not the B(2) receptor antagonist HOE 140 (1.0-4.0 nmol/paw), together with glutamate, also inhibited glutamate-induced nociception (53%) in a graded manner, without affecting glutamate-induced paw oedema. The i.pl. co-injection of the calcitonin gene-related peptide (CGRP) receptor antagonist CGRP(8-37) (1 nmol/paw) failed to significantly inhibit glutamate-induced nociception or oedema. Finally, neonatal-capsaicin (50 mg/kg, s.c.) treatment inhibited glutamate-induced nociception by 69% and to a lesser extent glutamate-mediated oedema formation (30%). Collectively, the current results indicate that the nociception caused by i.pl. injection of glutamate in mice is clearly mediated by capsaicin-sensitive fibres and by release of neurokinins from sensory neurones that activate NK(2) receptors and to a lesser extent NK(1) receptors. Furthermore, kinins acting at B(1) (but not at B(2)) receptors also largely account for glutamate-mediated nociceptive behaviour response. In contrast, glutamate-induced paw oedema seems to be primarily mediated via activation of NK(2) receptors and stimulation of capsaicin-sensitive C-fibres. CGRP receptors do not seem to be involved in either of the glutamate responses.

The pregnane compound MV8612 isolated from the rhizome of the plant Mandevilla velutina administered by intraperitoneal (i.p.), intrathecal (i.t.) or by intracerebroventricular (i.c.v.) routes caused graded and complete inhibition of the thermal hyperalgesia caused by i.t. injection of bradykinin (BK) in mice with mean ID(50) values of 7.8 micromol/kg, 33.6 and 4.6 nmol/site, respectively. Compound MV8612 (i.p.) also inhibited both the neurogenic and inflammatory pain responses to formalin with mean ID(50) values of 5.6 and 10.6 micromol/kg, respectively. Given i.t., MV8612 produced significant inhibition of both phases of the formalin-induced licking (inhibition of 34+/-5 and 36+/-4%, respectively). Given by i.c.v. route MV8612 inhibited both phases of formalin-induced pain (32+/-6 and 63+/-5%) with mean ID(50) of 8.4 nmol/site against the late phase. MV8612, given by i.p., i.c.v. or i.t. routes, also inhibited capsaicin-induced pain (51+/-4, 25+/-8 and 39+/-6%, respectively). The i.t. injection of potassium (K(+)) channel blockers, apamin and charybdotoxin given 15 min before, markedly prevented the antinociception of MV8612 against both phases of formalin-induced nociception. In contrast, tetraethylammonium (TEA) or glibenclamide had no effect. The i.c.v. treatment with pertussis toxin resulted in a significant inhibition of both MV8612- and morphine-induced antinociception against both phases of formalin-induced pain. Taken together these results confirm and also extend our previous data by demonstrating that the greater part of the antinociception caused by MV8612 seems to be associated with its ability to interfere with BK action. Finally, both the low and high conductance calcium (Ca(2+))-activated K(+) channels and the activation of G(i/o) pertussis sensitive G-proteins take part in the mechanism by which compound MV8612 produces antinociception.

This study characterizes the receptor subtypes and investigates some of the mechanisms by which glutamate, injected intraplantarly (i.pl.) into the mouse paw, produces nociception and paw oedema. I.pl. injection of glutamate induced a rapid-onset, dose-related pain response associated with oedema formation, with mean ED(50) values of 2.6 (1.6-4.3) and 0.5 (0.4-0.7) micromol/kg, respectively. Pretreatment with Chicago sky blue 6B (100 microg/kg), an inhibitor of glutamate uptake, caused a significant (about sixfold) reduction of the mean ED(50) value for glutamate-induced nociception, but not paw oedema. NMDA receptor antagonist MK 801, given by systemic (i.p.), intracerebroventricular (i.c.v.), i.pl. or intrathecal (i.t.) routes, produced graded inhibition of glutamate-induced nociception. Non-NMDA receptor antagonists NBQX or GAMS, metabotropic antagonist E4CPG, and also the antagonist that acts at the NMDA receptor-associated glycine binding site felbamate, significantly inhibited the nociception induced by glutamate. L(omega)-N-nitro-arginine (given i.p., i.t., i.pl. or i.c.v.) prevented the nociception and paw oedema caused by glutamate, an effect that was reversed by L-arginine but not by D-arginine. S-nitroso-N-acetyl-D,L-penicillamine (SNAP), given i.pl., greatly potentiated glutamate-induced nociception and oedema formation. Finally, the i.pl. injection of glutamate was accompanied by a graded increase in the nitrite levels of the hindpaw exudate. It is concluded that the nociception caused by i.pl. injection of glutamate probably involves the activation of NMDA and non-NMDA receptors by a mechanism which largely depends on the activation of L-arginine-nitric oxide pathway. Glutamate-induced paw oedema seems to be primarily mediated by non-NMDA ionotropic glutamate receptors and release of nitric oxide.

Slices of frog cerebrum, optic tectum, medulla and spinal cord rapidly accumulate [3H]GABA and [3H]glycine from the surrounding medium so that after 10 min tissue:medium ratios as high as 113 for GABA (optic tectum) and 18.5 for glycine (medulla) may be achieved. Kinetic analysis revealed two distinct saturable uptake systems for each amino acid in the 4 CNS areas. The high affinity systems (apparent Km: 9-22 muM for GABA; 5-35 muM for glycine) required sodium ions in the medium and were relatively substrate specific. Significant release of [3H]GABA and [3H]glycine, but not of L-[3H]leucine, was evoked by exposure to medium containing potassium ions in a concentration of 40 mM. The process of release was calcium-dependent. The importance of these results with regard to the roles of GABA and glycine as neurotransmitters in both spinal and supraspinal levels of the amphibian neuraxis is discussed.