Interaction between Ukrain and Naltrexone in the writhing syndrome test in mice

The interaction between Ukrain and Naltrexone, a nonselective opioid receptor antagonist, was studied in the 'writhing syndrome' test in mice. Naltrexone (1 mg/kg s.c.) was administered i.p. 30 min before Ukrain (2.375, 4.75, 9.5 and 19 mg/kg). In addition, the antinociceptive properties of Ukrain and Naltrexone in 10-day treatment in mice were studied. The results show that the antinociceptive effect of both single dose and prolonged administration of Ukrain is completely antagonized by Naltrexone.     

Ellagic acid enhances the antinociceptive action of carbamazepine in the acetic acid writhing test with mice

Context: Ellagic acid (EA) produced antinociceptive and anti-inflammatory effects through the central and peripheral sites of action.

Objective: The objective of the current study was to examine the functional interaction between ellagic acid and carbamazepine (CBZ) on pain.

Materials and methods: Fourteen groups of mice (8–10 each) were used in this study. Pain was induced by intraperitoneal acetic acid in mice (writhing test) and the functional interaction was analyzed using the isobolographic method. EA at doses 0.3, 1, 3, and 10?mg/kg and carbamazepine at doses 3, 10, 20, and 30?mg/kg, alone and also in combination (1/2, 1/4, and 1/8 of the drug’s ED₅₀) were intraperitoneally administered 30?min before acetic acid (0.6% v/v). Then, the abdominal writhes were counted during a 25-min period.

Results: EA (0.3–10?mg/kg, i.p.) and CBZ (3–30?mg/kg, i.p.) inhibited the writhing response evoked by acetic acid. Fifty percent effective dose (ED₅₀) values against this tonic pain were 1.02?mg/kg and 6.40?mg/kg for EA and CBZ, respectively. The antinociception induced by EA showed higher potency than that of carbamazepine. Co-administration of increasing fractional increments of ED₅₀ values of EA and CBZ produced additive interaction against writhing responses, as revealed by isobolographic analysis.

Discussion and conclusion: These results suggest that a combination of carbamazepine and ellagic acid may be a new strategy for the management of neuropathic pain such as what occurs in trigeminal neuralgia, since the use of carbamazepine is often limited by its adverse effects and by reduction of its analgesic effect through microsomal enzyme induction.     

Possible role of nitric oxide in the antinociceptive action of intraventricular bradykinin in mice

The i.c.v. administration of bradykinin (4, 8 and 16 μg) induced antinociception in mice which was resistant to naloxone; furthermore, the induction of tolerance to morphine by a single s.c. injection (100 mg/kg, 24 h before test doses of the peptide) did not affect antinociception. Since bradykinin is known to increase nitric oxide (NO) in peripheral tissues, we studied the possibility that its antinociceptive action may be related to NO effects in the central nervous system. Bradykinin effects were antagonized by previous treatment with N^G-nitro- -arginine or concomitant i.c.v. administration of bradykinin and methylene blue. The immediate precursor of NO, -arginine, which by itself produces analgesia, also reduced bradykinin effects; moreover, tolerance to -arginine significantly decreased the response to the peptide. These results suggest that NO is involved in antinociception induced by i.c.v. administration of bradykinin.     

Antidepressants enhance the antinociceptive effects of carbamazepine in the acetic acid-induced writhing test in mice

Some antidepressants, as well as antiepileptics, are effective for treating pain of varying etiology. The present study was designed to characterize the antinociceptive effects of imipramine, a tricyclic antidepressant, fluvoxamine, a selective serotonin reuptake inhibitor, milnacipran, a serotonin noradrenaline reuptake inhibitor, and carbamazepine, an antiepileptic drug, using the acetic acid-induced writhing test in mice. Imipramine (1.25–10 mg/kg, i.p.), fluvoxamine (5–40 mg/kg, i.p.) and milnacipran (2.5–20 mg/kg, i.p.) all dose-dependently and significantly reduced the number of writhes induced by the injection of acetic acid (0.8% (v/v)), although the maximal effect of milnacipran was weaker than those of imipramine and fluvoxamine. Similarly, carbamazepine (5–20 mg/kg, i.p.) also showed a dose-dependent and significant antinociceptive effect. In combination studies, the co-administration of a sub-effective dose of carbamazepine (5 mg/kg, i.p.) with imipramine (1.25 and 2.5 mg/kg, i.p.), fluvoxamine (10 mg/kg, i.p.) or milnacipran (1.25 and 2.5 mg/kg, i.p.) significantly reduced the number of writhes. Additionally, the hole-board test revealed that the medications with significant antinociceptive effects barely produced changes in motor activity that could possibly affect writhing behavior. Thus, the present study demonstrated that the antinociceptive effect of carbamazepine is enhanced by combination with imipramine, fluvoxamine and milnacipran. Therefore, the combined therapy using antidepressants and carbamazepine may be useful clinically for the control of pain.     

Modification of antinociceptive action of morphine by dopamine agonist and antagonist in mice

A marked potentiation of the antinociceptive action of morphine was produced by haloperidol, but apomorphine had no effect on the antinociceptive action of morphine in acute experiments. Following chronic treatment with haloperidol, the antinociceptive action of morphine was significantly suppressed by apomorphine, and apomorphine shifted the dose-response curve of morphine to the right and increased the ED50 value of morphine by 2.3-fold. These results suggest that the suppressive action of apomorphine on the antinociceptive effect of morphine in chronic haloperidol-treated mice may be due to an increased sensitivity of post-synaptic dopaminergic receptors to apomorphine.     

The development of morphine antinociceptive tolerance in nitric oxide synthase-deficient mice

Elevations in nitric oxide (NO) have been implicated in the development of morphine antinociceptive tolerance. This study was conducted to establish the role of specific isoforms of NO synthase (NOS) in morphine tolerance development using genetically modified mice. METHODS: Three groups of mice (endothelial NOS [eNOS]-deficient, neuronal NOS [nNOS]-deficient, and NOS-competent) were used in this experiment. On Day 1, the analgesic response (radiant heat tail-flick) to a challenge dose of morphine (4 mg/kg) was determined over 3 hr. Tolerance was induced on Days 1-5 by administering morphine subcutaneously (10 mg/kg) or L-arginine, a NO precursor, intraperitoneally (200 mg/kg), twice daily. Analgesic response to the challenge dose was determined again on Day 6. RESULTS: Following sustained morphine administration, nNOS-deficient mice exhibited less tolerance development when compared to the control group, although measurable tolerance still occurred. Mice deficient in eNOS evidenced a degree of tolerance similar to that of control. Prolonged L-arginine administration produced significant functional tolerance to morphine in NOS-competent and eNOS-deficient mice. The loss of morphine responsivity after L-arginine administration was similar to that after morphine pretreatment. L-Arginine did not affect the antinociceptive response to morphine in mice deficient in nNOS, suggesting that the small degree of morphine-induced tolerance in this group occurs through an alternate pathway. CONCLUSIONS: These data demonstrate the pivotal role of the neuronal isoform of NOS in development of morphine antinociceptive tolerance. Furthermore, tolerance development appears to be predominantly a NO-mediated process, but likely is augmented by a secondary (non-NO) pathway.     

The role of neuronal nitric oxide synthase on hypobaric hypoxiainduced antinociception in writhing test

It has been reported that hypobaric hypoxia exposure by high altitude is responsible for neuropsychological impairment. In the present study, we examined an effect of hypobaric hypoxia on the writhing test. The ICR mice were exposed in hypobaric chamber with several altitudes (5000, 10,000 or 20,000 ft) for 1 or 2 h, and then immediately injected intraperitoneally (i.p.) with 1% acetic acid for writhing test. Our results show that both 10,000 ft and 20,000 ft exposure induce antinociceptive effect in writhing test, but 5,000 ft does not. In addition, this antinociceptive effect was abolished by L-NAME (nitric oxide synthase inhibitor) pre-treated intraperitoneally, but not naloxone (non-specific opioid receptor antagonist). Furthermore, we examined that neuronal NOS immunoreactivities in the hypothalamus (paraventricular nucleus and arcuate nucleus) were increased by hypobaric hypoxic exposure (10,000ft). These results suggest that hypobaric hypoxic-induced antinociception may be associated with neuronal NOS IR in the hypothalamus.     

Role of nitric oxide in diabetes-induced attenuation of antinociceptive effect of morphine in mice

The study was designed to investigate the role of nitric oxide (NO) in the diabetes-induced decrease of the antinociceptive effect of morphine. The nociceptive threshold in diabetic and non-diabetic mice was measured in the tail-flick test. Streptozotocin (200 mg/kg i.p.) was administered to induce experimental diabetes in the mice. Four weeks after the administration of streptozotocin, the tail-flick test was performed and urinary nitrite concentration was estimated using Greiss reagent. Experimental diabetes markedly decreased the antinociceptive effect of morphine (10 microg in 5 microl/mice i.c.v.) and significantly increased the urinary nitrite concentration. Administration of aminoguanidine (12 mg/mice) markedly improved the antinociceptive effect of morphine and attenuated the increase in urinary nitrite concentration in diabetic mice. It may be tentatively concluded that an increase in NO formation may be responsible for the observed decrease in antinociceptive effect of morphine in diabetic mice.     

Effect of nitric oxide synthase inhibition on antinociceptive action of different doses of acetaminophen

As demonstrated in previous studies, both cyclooxygenases (COXs) and nitric oxide synthases (NOS) localized peripherally and/or centrally participate in the antinociceptive action of acetaminophen (ACETA). We showed that opioidergic system(s) was involved in the mechanism of ACETA action, as well. In previous and recent studies, the changes in nociceptive threshold were estimated using a mechanical and chemical stimulus. In this study, the influence of nonspecific inhibitor of NOS [N(G)-nitro-L-arginine (L-NOArg)] on antinociceptive action of ACETA administered icv and it was studied in rats. ACETA increased threshold for electrical stimuli, however, its analgesic activity was not dose-dependent. Independently of the route of administration, the existence of a ceiling dose of ACETA was observed above which the activity of ACETA was self-limited. Pretreatment with L-NOArg (ip) markedly increased the action of higher doses of ACETA. It suggests that the attenuation of analgesic action of higher doses of ACETA may be due to increased activity of NOS.     

A test for analgesics: incoordination in writhing mice

Acetic acid administered to mice resulted in post-writhing incoordination and a subsequent fall in the rota-rod test. This fall from rota-rod induced by 3% acetic acid was dose-dependently protected by the non-narcotic analgesics. While suprofen was found to be the most potent (ED50 = 15.29 mg/kg), aspirin was found to be the least potent (ED50 = 81.54 mg/kg). The model appears to be more sensitive than the conventional methods for testing non-narcotic analgesics.     

Tipepidine enhances the antinociceptive-like action of carbamazepine in the acetic acid writhing test

Several antidepressants have been used to treat severe pain in clinics. Recently, we reported that the centrally acting non-narcotic antitussive (cough suppressant drug), tipepidine produces an antidepressant-like effect in the forced swimming test, although the mechanism of action appears to be quite different from that of known antidepressants. In the present study, we investigated whether a combination of tipepidine and carbamazepine acts synergistically to induce an antinociceptive effect in the acetic acid-induced writhing test in mice. Prior to studying the combination of tipepidine and carbamazepine, the analgesic action of tipepidine alone was also examined in mice. Tipepidine at 5-40mg/kg i.p. significantly reduced the number of writhes induced by acetic acid in mice. Carbamazepine at 20mg/kg i.p. also significantly reduced the writhing reaction. Furthermore, co-administration of carbamazepine (5 and 10mg/kg, i.p.) and tipepidine (2.5mg/kg i.p.) significantly decreased the number of writhes induced by acetic acid. This finding suggests that a combination of carbamazepine and tipepidine may be a new strategy for the treatment of neuropathic pain such as what occurs in trigeminal neuralgia, because the use of carbamazepine is often limited by its adverse effects and by reduction of its analgesic efficacy by microsomal enzyme induction.     

Further analysis of the antinociceptive action caused by p-methoxyl-diphenyl diselenide in mice

The objective of this study was to extend our previous findings by investigating in greater detail the mechanisms that might be involved in the antinociceptive action of p-methoxyl-diphenyl diselenide, (MeOPhSe)₂, in mice. The pretreatment with nitric oxide precursor, l-arginine (600 mg/kg, intraperitoneal, i.p.), reversed antinociception caused by (MeOPhSe)₂ (10 mg/kg, p.o.) or N^G-nitro-l-arginine (l-NOARG, 75 mg/kg, i.p.) in the glutamate test. Ondansetron (0.5 mg/kg, i.p., a 5-HT₃ receptor antagonist) and SCH23390 (0.05 mg/kg, i.p.., a D₁ receptor antagonist) blocked the antinociceptive effect caused by (MeOPhSe)₂. Conversely, pindolol (1 mg/kg, i.p., a 5-HT_1A/_1B receptor/β adrenoceptor antagonist), WAY 100635 (0.7 mg/kg, i.p., a selective 5-HT_1A receptor antagonist), ketanserin (0.3 mg/kg, i.p., a selective 5-HT_2A receptor antagonist), prazosin (0.15 mg/kg, i.p., an α₁-adrenoreceptor antagonist), yohimbine (1.0 mg/kg, i.p., an α₂-adrenoreceptor antagonist), sulpiride (5 mg/kg, i.p., a D₂ receptor antagonist), naloxone (1 mg/kg, i.p., a non-selective opioid receptor antagonist) and caffeine (3 mg/kg, i.p., a non-selective adenosine receptor antagonist) did not change the antinociceptive effect of (MeOPhSe)₂. (MeOPhSe)₂ significantly inhibited nociception induced by intraplantar (i.pl.) injection of bradykinin (10 nmol/paw) and Des-Arg⁹-bradykinin (10 nmol/paw, a B₁ receptor agonist). (MeOPhSe)₂ significantly inhibited phorbol myristate acetate (PMA, 0.03 μg/paw, a protein kinase C (PKC) activator)-induced licking response. These results indicate that (MeOPhSe)₂ produced antinociception in mice through mechanisms that involve an interaction with nitrergic system, 5-HT₃ and D₁ receptors. The antinociceptive effect is related to (MeOPhSe)₂ ability to interact with kinin B₁ and B₂ receptors and PKC pathway mediated mechanisms.     

Evidence for the participation of the nitric oxide-cyclic GMP pathway in the antinociceptive action of meloxicam in the formalin test

The involvement of the nitric oxide-cyclic GMP pathway in the antinociceptive action of the cyclooxygenase-2 preferential inhibitor meloxicam was assessed in the rat formalin test. Rats received local pretreatment with saline or meloxicam and then 50 microl of dilute formalin (1%). Local administration of meloxicam produced a dose-dependent antinociception in the second phase of the formalin test. The antinociception produced by meloxicam was due to a local action as its administration in the contralateral paw was ineffective. Local pretreatment of the paws with saline or N(G)-D-nitro-arginine methyl ester (D-NAME) did not affect the antinociception produced by meloxicam. However, N(G)-L-nitro-arginine methyl ester (L-NAME, a NO synthesis inhibitor) or 1H-(1,2,4)-oxadiazolo(4,2-a)quinoxalin-1-one (ODQ, a soluble guanylyl cyclase inhibitor) blocked in a dose-dependent way the effect of meloxicam. It is concluded that the peripheral antinociceptive effect of meloxicam involves a local NO-cyclic GMP pathway.     

Modulation of adjuvant arthritis by endogenous nitric oxide.

1. The role of endogenous nitric oxide (NO) in adjuvant arthritis in Lewis rats has been studied by use of L-arginine, the amino acid from which NO is synthesized, and NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthase. Prolonged modulation (35 days) of the L-arginine: NO pathway in rats was achieved by dissolving test compounds in the drinking water (L-arginine: 3, 10 and 30 mg ml-1; L-NAME: 0.1, 1 and 10 mg ml-1). 2. Arthritis was exacerbated by L-arginine and suppressed by L-NAME in a dose-related fashion. Combined treatment with L-NAME (1 mg ml-1) and L-arginine (30 mg ml-1) did not modify the arthritis. 3. Reduced weight gain, which is a feature of adjuvant arthritis, was modified by these compounds so that L-arginine reduced weight gain whereas L-NAME increased weight gain compared with that in control animals. 4. D-Arginine (30 mg ml-1), NG-nitro-D-arginine methyl ester (D-NAME: 1 mg ml-1) and L-lysine (30 mg ml-1), an amino acid not involved in the generation of NO, were without effect on either arthritis or body weight gain. 5. Antigen-stimulated proliferation of T-lymphocytes as well as generation of nitrite (NO2-) and release of acid phosphatase from macrophages were all enhanced in L-arginine-treated arthritic rats and reduced in L-NAME-treated animals. 6. These results suggest that endogenous NO modulates adjuvant arthritis, possibly by interfering with the activation of T-lymphocytes and/or macrophages.     

Modulation of acute inflammation by endogenous nitric oxide.

The role of endogenous nitric oxide (NO) in acute inflammation was investigated using two inhibitors of NO synthase (NG-nitro-L-arginine methyl ester(L-NAME) and NG-monomethyl-L-arginine (L-NMMA)) as well as L- or D-arginine. The effect of test compounds was studied on the carrageenin-induced increase in vascular permeability in rat skin and in dextran- and carrageenin-induced paw oedema. Both L-NAME and L-NMMA dose dependently inhibited the increase in vascular permeability and oedema formation. L- but not D-arginine increased these inflammatory responses and reversed the inhibitory effects of L-NAME and L-NMMA. In dexamethasone-treated rats L-arginine enhanced the dextran-induced oedema and the early phase of carrageenin-induced oedema but did not modify the inhibition by dexamethasone of the late phase of carrageenin-induced oedema. These results suggest that endogenous NO is released at the site of acute inflammation and modulates oedema formation. Depending on the time course or on the type of inflammation, NO may be predominantly generated by the constitutive or by the inducible NO synthase.     

Antinociceptive activity of sildenafil and adrenergic agents in the writhing test in mice

The authors investigated the antinociceptive activity of sildenafil and adrenergic agents co-administered in the writhing test in mice. The intensity of nociception was quantified by the number of writhes occurring between 0 and 30 min after stimulus injection. Nontreated groups (NT) received acid intraperitoneally (ip) followed by sterile saline (ip). Animals received (ip) sildenafil (2.5 or 5 mg/kg), propranolol (0.5 or 2 mg/kg), atenolol (0.05 or 2 mg/kg), prazosin (0.05 or 0.25 mg/kg) or clonidine (0.01 or 0.1 mg/kg) 30 min before acid injection. It was observed that only the largest doses of every drug inhibited the number of writhes in mice. In another series of experiments, animals were pretreated with the lower ineffective doses of propranolol, atenolol, prazosin or clonidine. After 30 min, mice also received the lower ineffective dose of sildenafil followed by acid injection. The combination of ineffective doses of propranolol, atenolol, prazosin or clonidine with sildenafil significantly inhibited the nociceptive response induced by acetic acid injection. Data obtained from these experiments showed that ineffective doses of sildenafil associated with ineffective doses of adrenergic agents provided analgesic effects in the writhing test.     

Inhibition of nitric oxide synthase by isothioureas: Cardiovascular and antinociceptive effects

A range of substituted isothiourea compounds including S-isopropylisothiourea (IPTU), S-methylisothiourea (SMT), S-ethylisothiourea (ETU), N-pentylisothiourea (PTU), S-(2 aminoethyl)isothiourea (AETU), and S-acetamidoisothiourea (AATU) inhibit mouse spinal cord/cerebellar neuronal nitric oxide synthase (nNOS) and bovine aortic endothelial cell eNOS in vitro. IP administration of isothioureas increased mean arterial blood pressure of the urethane-anaesthetised mouse (rank order of effect: IPTU > ETU > SMT > AETU). PTU and AATU were without vasopressor activity. IPTU (50 mg/kg, IP) inhibited late phase formalin-induced hindpaw licking behaviour in the mouse while SMT (50 mg/kg, IP) was without effect. Neither compound influenced the formalin-induced increase in hindpaw weight reflecting a lack of significant peripheral antioedema effect in this model. IPTU (50 mg/kg, IP) but not SMT (50 mg/kg, IP) inhibited mouse spinal cord and cerebellar NOS activity measured ex vivo in animals killed 45 min after injection. The present study confirms the potent NOS inhibitory effect of selected substituted isothioureas in vitro. Little or no isoform selectivity (i.e., nNOS vs. eNOS) was apparent. The potent vasopressor effect of isothioureas indicates that these compounds may be of limited use as tools to study the role of nitric oxide in pain perception.     

Modulation of morphine antinociception in the mouse by endogenous nitric oxide.

1. L-Arginine (100-1000 mg kg-1) administered orally (p.o.) or intraperitoneally (i.p.), but not intracerebroventricularly (i.c.v., 0.08 mg per mouse), reduced the antinociceptive effect of morphine (0.5-10 mg kg-1 s.c.) assessed in mice using three different tests: hot plate, tail-flick and acetic acid-induced writhing. D-Arginine (up to 1000 mg kg-1 p.o. or i.p.) was ineffective. 2. NG-Monomethyl-L-arginine (L-NMMA, 5-50 mg kg-1 i.p.) and NG-nitro-L-arginine methyl ester (L-NAME, 5- 30 mg kg-1 i.p.), but not NG-nitro-D-arginine methyl ester (D-NAME, 30 mg kg-1 i.p.), reversed in all assays the effect of L-arginine on morphine-induced antinociception. 3. Morphine (10 mg kg-1 s.c.), L-arginine (1000 mg kg-1 p.o.) or L-NAME (30 mg kg-1 i.p.), either alone or in combination, did not produce changes in locomotor activity or sensorimotor performance of animals. 4. These results suggest that the L-arginine-nitric oxide pathway plays a modulating role in the morphine-sensitive nociceptive processes.     

The synergistic interaction between rilmenidine and paracetamol in the writhing test in mice

The aim of the study was to ascertain antinociceptive effects of rilmenidine, a second-generation imidazoline-alpha-2-adrenoreceptor agonist, and to see whether rilmenidine was able to increase the analgesic effects of paracetamol in the writhing test in mice. An acetic acid (0.7%) solution was injected into the peritoneal cavity and the number of writhes was counted. The influence on locomotor performance was tested using the rotarod test. Rilmenidine, paracetamol, and rilmenidine–paracetamol fixed-ratio combinations produced dose-dependent antinociceptive effects. ED₅₀ values were estimated for the individual drugs and an isobologram was constructed. The derived theoretical additive ED₅₀ value for the rilmenidine–paracetamol combination was 109.23 ± 35.05 mg/kg. This value was significantly greater than the observed ED₅₀ value which was 56.35 ± 20.86 mg/kg, indicating a synergistic interaction. Rilmenidine did not impair motor coordination, as measured by the rotarod test, at antinociceptive and higher doses.