Synergism between COX-3 inhibitors in two animal models of pain.

OBJECTIVE AND DESIGN: The antinociception induced by the intraperitoneal coadministration in mice of combinations of metamizol and paracetamol was evaluated in the tail flick test and orofacial formalin test. METHODS: The antinociception of each drugs alone and the interaction of the combinations was evaluated by isobolographic analysis in the tail-flick and in the formalin orofacial assay of mice. RESULTS: Mice pretreated with the drugs demonstrated that the antinociception of metamizol and paracetamol is dose-dependent. The potency range on the antinocifensive responses for metamizol or paracetamol was as follows: orofacial (Phase II) > orofacial (Phase I) > tail flick. In addition, the coadministration of metamizol with paracetamol induced a strong synergistic antinociception in the algesiometer assays. Both drugs showed effectiveness in inflammatory pain. CONCLUSION: These actions can be related to the differential selectivity of the drugs for inhibition of COX isoforms and also to the several additional antinociception mechanisms and pathways initiated by the analgesic drugs on pain transmission. Since the efficacy of the combination of metamizol with paracetamol has been demonstrated in the present study, this association could have a potential beneficial effect on the pharmacological treatment of clinical pain.

Lack of effect of naltrindole on the spinal synergism of morphine and non-steroidal anti-inflammatory drugs (NSAIDS).

To enhance analgesia, combination of analgesics drugs of proven efficacy is a strategy which is accompanied by a reduction of adverse effects. The present study was undertaken to characterize the antinociceptive interaction of morphine with different non-steroidal anti-inflammatory drugs (NSAIDs) using isobolographic analysis and the writhing test of mice. One of the possible mechanisms of action of spinally administered morphine with non-steroidal antiinflammatory drugs was investigated using the DOR antagonist naltrindole. The study demonstrated a synergistic antinociception of spinal administered combinations of morphine with the following NSAIDs agents: diclofenac, ketoprofen, meloxicam, metamizol, naproxen, nimesulide, parecoxib and piroxicam. The supraadditive effect seems to be independent of the selectivity of each NSAIDs to inhibit COX-1 or COX-2. The findings of the present work suggest that the combinations of opioids and non-steroidal anti-inflammatory drugs have a direct action on spinal processing of the nociceptive information, which may achieved by additional mechanisms independent of prostaglandin synthesis inhibition and/or activation of opioid receptors. The lack of effect of naltrindole to modify the analgesic activity of the combination of opioids and NSAIDs indicates that others pain regulatory systems are involved in this central action. Therefore, these combinations could be a viable alternative to clinical pain management, especially trough multimodal analgesia.

The effect of opioid antagonists on synergism between dexketoprofen and tramadol.

The antinociceptive activity of dexketoprofen was studied in mice using the formalin assay for orofacial pain. The interaction between dexketoprofen and co-administered tramadol was studied using isobolographic analysis. The intraperitoneal administration of dexketoprofen or tramadol, showed dose-dependent antinociceptive activity in both phases of the assay. When administered together, the interaction was mildly synergistic during the first phase, and antagonistic in the second phase. Selective opioid receptor antagonists where used in order to measure the analgesic activity of tramadol in other regions of the CNS. The co-administration of dexketoprofen and tramadol, with previous administration of naltrexone, showed high synergistic activity during the first phase, and less but still synergistic during the second. When using naltrindole, the interaction was mildly more synergistic than the mixture dexketoprofen+tramadol during both phases. Using norbinaltorphimine, the interaction was synergistic in both phases, more marked in the second. These results suggest that the opioid activity of tramadol has an inhibiting effect in antinociceptive activity of the interaction between dexketoprofen and tramadol during the inflammatory (late) stages of pain.

Isobolographic analysis of the antinociceptive interactions of clonidine with nonsteroidal anti-inflammatory drugs.

The present study was undertaken to characterize the interactions between nonsteroidal anti-inflammatory drugs and the alpha(2)-adrenoceptor agonist clonidine in an acute nociceptive test. The writhing test was selected as a model of acute visceral pain. Isobolograms were constructed to assess the interactions of clonidine and each nonsteroidal anti-inflammatory drugs, when coadministered intraperitoneally and intrathecally (i.t.). The simultaneous intraperitoneal administration of fixed ratios of ED(50) fractions of all nonsteroidal anti-inflammatory drugs (naproxen, piroxicam, paracetamol, dipyrone or metamizol and nimesulide) combined with clonidine resulted in synergistic interactions. The same combinations administered intrathecally were additive. The synergistic interactions between systemic nonsteroidal anti-inflammatory drugs and clonidine may involve supraspinal mechanisms.

Synergy between the antinociceptive effects of morphine and NSAIDs.

The intraperitoneal administration of morphine, diclofenac, ketoprofen, meloxicam, metamizol, paracetamol and piroxicam induced dose-dependent antinociception in mice tested with the acetic acid writhing test. The isobolographic analysis of the simultaneous intraperitoneal administration of fractions of the ED50's of morphine with each nonsteroidal anti-inflammatory drug (NSAID) demonstrated the existence of a supra-additive interaction (synergy). The selective antagonist of micro -opioid receptors naltrexone partially reversed the supra-additive interactions to additive interactions; however, the combinations of morphine/metamizol and morphine/paracetamol were completely antagonized, resulting in subadditive interactions. The selective antagonist of delta-opioid receptors naltrindole failed to significantly attenuate the combinations of morphine with ketoprofen, meloxicam and piroxicam, but decreased the activity of the combinations of morphine with diclofenac, metamizol and paracetamol, transforming the interactions from supra-additive to additive. Nor-binaltorphimine was used to evaluate the involvement of kappa-opioid receptors. Nor-binaltorphimine did not modify the supra-additive interaction of morphine and NSAIDs and the additive interaction of the co-administration of morphine and metamizol. The synergy between morphine and NSAIDs could be related to different pathways of pain transmission, probably related to the different intracellular signal transduction mechanisms of action of opioid and non-opioid agents.

Effect of the inhibition of serotonin biosynthesis on the antinociception induced by nonsteroidal anti-inflammatory drugs.

The antinociceptive activity of nonsteroidal anti-inflammatory drugs (NSAIDs) has been explained mainly on the basis of their inhibition of the enzyme cyclooxygenase (COX); however, this inhibition is not enough to completely explain the analgesic efficacy of these drugs. The modulation exerted by serotonergic systems on antinociception is well known. The purpose of the present work was to further explore the role of serotonin in the antinociceptive activity of NSAIDs using the writhing test and the tail-flick test of the mice after the inhibition of serotonin biosynthesis with intraperitoneal p-chlorophenylalanine (p-CPA). Pretreatment with p-CPA produced a significant decrease in the antinociceptive activity of NSAIDs administered either by the intraperitoneal or intrathecal routes, in both algesiometric tests. These results suggest a complementary mechanism of antinociception for NSAIDs, independent of their ability to inhibit the activity of COX, involving the activation of descending serotonergic pathways. By the pharmacological nature of the study, one limitation was the absence of biochemical measurement of the synthesis of 5-HT, since the reduction of the brain 5-HT synthesis by pretreatment with p-CPA will be expressed as a diminished antinociceptive activity of NSAIDs, which would be a new argument to consider NSAIDs acting as central analgesic agents.

Atropine reverses the antinociception of nonsteroidal anti-inflammatory drugs in the tail-flick test of mice.

The nonsteroidal anti-inflammatory drugs (NSAIDs) clonixin, diclofenac, piroxicam, ketoprofen, meloxicam, and paracetamol induced antinociception after intraperitoneal or intrathecal administration in mice submitted to an acute thermal algesiometric test without inflammation (tail-flick). Antinociception was evaluated by the increase in reaction time difference (Delta latency), between readings obtained before and after the administration of drugs. The antinociception induced by doses of NSAIDs producing between 20% and 30% of the maximum possible effect (MPE) 30 min after intraperitoneal and 15 min after intrathecal injections was compared with the antinociception obtained after pretreatment with 1 mg/kg atropine ip, 30 min before. Systemic atropine (1 mg/kg) significantly antagonized NSAID-induced antinociception in all cases, both after intraperitoneal and intrathecal administration. Cholinergic depletion by intracerebroventricular hemicholinium-3 (HC-3, 5 microg) 5 h before prevented the antinociceptive effect of all NSAIDs. These observations suggest that intrinsic muscarinic cholinergic facilitatory pathways represent an important modulating system in pain perception in this animal model of acute thermal pain. The results of the present work support the increasingly accepted notion that NSAIDs are effective analgesics even when inflammation is not present, acting by mechanisms that involve actions on spinal and supraspinal nociceptive transmission. It is suggested that, similar to morphine and clonidine, the active mechanism of NSAIDs may involve the release of acetylcholine (ACh) in the spinal cord.

Adrenergic mechanisms in antinociceptive effects of non steroidal anti-inflammatory drugs in acute thermal nociception in mice.

OBJECTIVE: The interactions of alpha-adrenoceptors with the antinociceptive effects of non-steroidal antiinflammatory drugs (NSAIDs) were assessed in acute thermal nociception in mice. MATERIALS AND METHODS: The analgesic effect was analyzed by the tail-flick test. RESULTS: The pretreatment with yohimbine (1 mg/kg i.p.), 30 min prior to the intraperitoneal injection of ketoprofen (50 mg/kg), diclofenac (30 mg/kg) and piroxicam (50 mg/kg) antagonized the antinociception induced by these NSAIDs, significantly reducing the tail-flick latency. Yohimbine did not affect paracetamol (125 mg/kg) induced antinociception. Prazosin (1 mg/kg i.p.) antagonized only the effect of paracetamol, without affecting the latency of the other drugs. When NSAIDs were administered i.t. (ketoprofen 2 m/kg; diclofenac 0.9 mg/kg; piroxicam 1.5 mg/kg; paracetamol 3.75 mg/kg), the same results were obtained after i.p. pretreatment with yohimbine and prazosin. The pretreatment of phenoxybenzamine (1 mg/kg i.p.) antagonized all antinociceptive effects. CONCLUSIONS: NSAIDs induced antinociception in an acute thermal pain model without inflammation. The mechanism of antinociception induced by ketoprofen, diclofenac and piroxicam involves an activation of alpha2-adrenoceptors at spinal and supraspinal levels, while paracetamol-induced antinociception is probably due mainly to central activation of the descending noradrenergic inhibitory system by alpha1-adrenoceptors.

Comparing two programs of cognitive training in Alzheimer's disease: a pilot study.

OBJECTIVES: To evaluate the efficacy of two different procedures of individual cognitive training in mild to moderate Alzheimer's Disease (AD). MATERIAL AND METHODS: Twenty-two AD patients entered the study. We compared stimulation of procedural memory (group 1) with training of partially spared cognitive functions (group 2). Assessment included: neuropsychological tests, scales, and the Functional Living Skills Assessment (FLSA), a standardized battery built to directly evaluate patients' performance in everyday life. RESULTS: We observed a significant improvement for both groups after training in FLSA total score (P=0.005) and subscales. For group 1, we also found a slightly improved performance in two tests: Attentional Matrices (P=0.041), and Verbal Fluency for Letters (P=0.059). After 3 months, patients' results showed a tendency to regress to the pre-training level. CONCLUSION: Both AD groups showed a substantial improvement after training in a direct performance measure of everyday functioning. However, results at neuropsychological tests suggest that training activities of daily living (supported by procedural memory) may be more effective than stimulating "residual" cognitive functions.

Carbachol interactions with nonsteroidal anti-inflammatory drugs.

The inhibition of cyclooxygenase enzymes by nonsteroidal anti-inflammatory drugs (NSAIDs) does not completely explain the antinociceptive efficacy of these agents. It is known that cholinergic agonists are antinociceptive, and this study evaluates the interactions between carbachol and some NSAIDs. Antinociceptive activity was evaluated in mice by the acetic acid writhing test. Dose-response curves were constructed for NSAIDs and carbachol, administered either intraperitoneally (i.p.) or intrathecally (i.t.). The interactions of carbachol with NSAIDs were evaluated by isobolographic analysis after the simultaneous administration of fixed proportions of carbachol with each NSAID. All of the drugs were more potent after spinal than after systemic administration. The combinations of NSAIDs and carbachol administered i.p. were supra-additive; however, the i.t. combinations were only additive. Isobolographic analysis of the coadministration of NSAIDs and carbachol and the fact that atropine antagonized the synergistic effect suggest that carbachol may strongly modulate the antinociceptive activity of NSAIDs; thus, central cholinergic modulation would be an additional mechanism for the antinociceptive action of NSAIDs, unrelated to prostaglandin biosynthesis inhibition.

Interaction between the antinociceptive effect of ketoprofen and adrenergic modulatory systems.

The interaction between the antinociceptive activity of ketoprofen and adrenergic agents was evaluated in the writhing test of mice. Dose-response curves were obtained for systemic and intrathecal antinociceptive effects of ketoprofen, phenylephrine, clonidine, desipramine, and prazosin; and ED50 were calculated. The interactions were evaluated by isobolographic analysis of the systemic or intrathecal co-administration of fixed-ratio combinations of ketoprofen with each adrenergic agent. The intraperitoneal combinations of ketoprofen with phenylephrine, clonidine, and prazosin showed supra-additivity, indicating that activation of alpha1 and alpha2 adrenoceptors play a role in nociceptive transmission at supraspinal levels. The same combinations given intrathecal were only additive. Desipramine intraperitoneal was also supra-additive: however, when ketoprofen was administered intrathecally with desipramine, only an additive interaction was obtained. The supra-additive interactions suggest that complementary mechanisms of antinociception have been activated, related with interference with the multiplicity of receptors and systems involved in the transmission of the nociceptive information. Racemic ketoprofen has an antinociceptive activity which is probably not only due to COX inhibition but also involves noradrenergic systems at spinal and supraspinal levels.

An isobolographic analysis of the adrenergic modulation of diclofenac antinociception.

UNLABELLED: We evaluated the noradrenergic modulation of the antinociceptive activity of diclofenac in mice using the acetic acid writhing test. Dose-response curves were obtained for the antinociceptive effect of diclofenac, phenylephrine, clonidine, desipramine, prazosin, and yohimbine administered both systemically and intrathecally, and ED(50)s were calculated. Noradrenergic modulation was evaluated by performing an isobolographic analysis of the systemic or intrathecal coadministration of fixed-ratio combinations of diclofenac with each adrenergic drug. The systemic, but not the intrathecal, combinations of diclofenac with phenylephrine or clonidine showed supraadditivity, suggesting that the activation of alpha(1) and alpha(2) adrenoceptors interfered with the nociceptive transmission at spinal and supraspinal levels. Supraadditive effects were not demonstrated for the intrathecal injection of diclofenac combined with phenylephrine, clonidine and a selective norepinephrine uptake inhibitor (desipramine) or adrenergic antagonists. We conclude that interaction between adrenoceptors and diclofenac can modulate antinociception by activating common or different mechanisms. Diclofenac has an antinociceptive activity that, in addition to cyclooxygenase inhibition, can be modulated by additive and supraadditive interactions with adrenergic drugs. IMPLICATIONS: Diclofenac analgesia in mice can be modulated by interaction with adrenergic drugs. The systemic but not the intrathecal administration of phenylephrine and clonidine produced supraadditive interactions. For desipramine, prazosin, and yohimbine, supraadditive interactions were not statistically demonstrated. The coadministration of drugs inducing supraadditive effects could be clinically relevant for the treatment of chronic pain because of reduction of doses and side effects.

Effects of ketoconazole on oestrus cycle and convulsant action of pentylenetetrazol in mice.

The frequency of oestrus cycles in female mice was significantly reduced by the implantation of a pellet of subcutaneous ketoconazole (50 mg every 6 days). The effect was more pronounced after 22 days than after 13 days and it was probably related with the progressive reduction in steroid synthesis induced by the inhibition of key steroidogenic P450 cytochromes by the drug. In addition, the influence of ketoconazole on the incidence of seizures after the administration of intraperitoneal pentylenetetrazol was evaluated in female mice. Pentylenetetrazol produced a higher percentage of seizures during dioestrus than during oestrus. Pretreatment with ketoconazole significantly increased the percentage of animals with induced seizures in oestrus but not in dioestrus as compared to controls, probably through reduced progesterone levels. The reduced seizure threshold confirm the modulatory role exerted by progesterone on central nervous system excitability, and may be relevant in epileptic patients undergoing antifungal therapy.

NSAID antinociception measured in a chemical and a thermal assay in mice.

The antinociceptive activity of several nonsteroidal anti-inflammatory drugs (NSAIDs) that were administered either intraperitoneally or intrathecally was assessed in mice by two algesiometric tests. The first was the writhing test, which assessed the abdominal constrictions that were induced by intraperitoneal acetic acid (a chemical test), and the second was the tail flick test, which measured pain responses to heat stimuli. The corresponding effective doses and their relative potencies were compared because these tests use different nociceptive stimuli with different transmission pathways. The intraperitoneal and intrathecal dose-response curves for the antinociception induced by every NSAID that was tested were parallel in the writhing test. In the tail flick test, however, only the intraperitoneal and intrathecal dose-response curves for ketoprofen, piroxicam, naproxen, nimesulide, paracetamol and diclofenac were parallel. The results obtained in this study confirm that NSAIDs possess different abilities to induce inhibition of cyclooxygenase, and they can be indirectly assessed by their different antinociceptive activities, depending on the algesiometric assays that are used. The antinociception of most NSAIDs might involve central mechanisms. The findings demonstrate the increasing importance of the spinal cord in processing and modulating nociceptive input, because intrathecal administration of NSAIDs is always more effective (in terms of potency) than systemic administration; thus, the antinociceptive efficacy of NSAIDs strongly depends on the algesiometric assay that is used and on the type of the nociceptive stimulus to which the test subject is exposed.

Progesterone and testosterone modulate the convulsant actions of pentylenetetrazol and strychnine in mice.

The influence of progesterone and testosterone on the incidence of seizures after administration of intraperitoneal pentylenetetrazol and subcutaneous strychnine was evaluated in mice. Pentylenetetrazol and strychnine were administered in doses that induced seizures in 40-50% of control mice in dioestrus (48 and 0.9 mg/kg, respectively). The percentage of seizures induced by pentylenetetrazol and strychnine was significantly lower in female mice in prooestrus/oestrus, when progesterone levels are high, than in dioestrus, when progesterone levels are low. Pretreatment of pentylenetetrazol-challenged mice with progesterone (250 microg/kg) increased the incidence of seizures in prooestrus/oestrus, without affecting seizures in dioestrus. The same pretreatment in strychnine-challenged mice also increased the incidence of seizures in prooestrus-dioestrus, but significantly reduced the incidence of seizures in dioestrus. In addition, progesterone pretreatment significantly increased the percentage of deaths induced by strychnine in prooestrus-oestrus, reducing deaths in dioestrus. Orchidectomized male mice had a significantly higher incidence of seizures after administration of pentylenetetrazol and strychnine than control mice. Administration of 11 daily doses of 250 microg/kg of testosterone to castrated mice significantly reduced the incidence of seizures induced by pentylenetetrazol. These results confirm the modulatory influence of reproductive steroids on the excitability of the central nervous system and the possible clinical importance of progesterone and testosterone in the management of partial epilepsy.

Modulación opioide de la actividad antinociceptiva de L-arginina en ratones / Opioid modulation of the antinociceptive activity of L-arginine in mice

Objetivos: El óxido nítrico (NO) es un neuromodulador intercelular sintetizado por la NO sintetasa (NOS). L-arginina (L-ARG) es convertida a ornitina y NO por la NOS y está envuelta en la vía L-ARG-NO de la transmisión y/o modulación nociceptiva, tanto en el SNC como en la periferia. En el presente trabajo se determinó la actividad antinociceptiva de L-ARG y se evaluó la neuromodulación producida por antagonistas opioides selectivos y no selectivos.Material y métodos: En ratones de la cepa CF-1 se determinó la curva dosisrespuesta de la actividad antinociceptiva de L-ARG administrada por vía intraperitoneal (i.p.), usando el test algesiométrico de las contracciones abdominales inducidas por ácido acético. Las curvas dosis-respuesta se repitieron después del p retratamiento con los siguientes antagonistas de receptores opioides: naloxona y naltrexona (no selectivos), naltrindole (antagonista ) y norbinaltorfimina (antagonista ). La actividad antinociceptiva de una dosis única de L-ARG administrada tanto por vía intratecal (i.t.) como por vía intracere broventricular (i.c.v.) se determinó mediante el mismo test.Resultados : L-ARG por vía i.p. exhibió una actividad antinociceptiva dosis-dependiente, con una DE5 0 de 4,65 mg*k g- 1. La inhibición de NOS redujo significativamente el efecto antinociceptivo de L-ARG. La administración i.t. e i.c.v. de una dosis aproximadamente 45 veces menor (0,16 y 0,15 m g . k g- 1) produjo un efecto antinociceptivo comparable. El p retratamiento con todos los antagonistas de receptores opioides desplazó significativamente la curva dosis-re spuesta de la actividad antinociceptiva de L-ARG hacia la d e recha, indicando un antagonismo farmacológico .Conclusiones : L-ARG administrada por vía sistémica, a través de la formación de NO y activación de GMP cíclico, induce antinocicepción probablemente por efecto directo sobre nociceptores periféricos, activando mecanismos antinociceptivos a nivel espinal y supraespinal. La relación de dosis sistémicas, intratecales e intracere bro-ventriculares equipotentes sugiere que la mayor parte de su acción se ejerce en el SNC. La síntesis y la liberación de NO facilita en alguna forma la activación de receptores opioides tanto a nivel espinal como supraespinal y confirma el efecto modulador del NO en la nocicepción (AU)

Isobolographic analysis of the interaction between fenoldopam and levodopa on arterial blood pressure of the rat.

Fenoldopam (FD) and levodopa (LD) injected intravenously in rats in a noncumulative schedule induced dose-dependent reductions in mean arterial blood pressure. The doses that induced a 50% reduction in the initial control mean arterial pressure (referred as ED50) were calculated by linear regression analysis of the corresponding parallel dose-response curves and were 0.88 and 0.068 mg/kg, respectively. The interaction between the effects of FD and LD on pressure reduction was evaluated by simultaneous administration of different fixed ratios of FD and LD (16:1) and obtaining a dose-response curve. An isobolographic analysis was then performed, which showed that the experimental point for the effect of the simultaneous administration of FD and LD was significantly different from the theoretically calculated additive point, denoting supradditivity. It was concluded that the effect of the combination of FD and LD on mean blood pressure reduction was synergic and was probably due to an activation of D1 vascular receptors by both drugs, in conjunction with an activation of beta2 adrenoceptors by LD and a blocking action of FD on postsynaptic alpha1-adrenoceptors.

Antinociception, tolerance, and physical dependence comparison between morphine and tramadol.

The mechanism of action of tramadol includes the activation of opioid receptors, and the potential ability of the drug to induce tolerance and physical dependence has been evaluated in different animal species and humans. This work was designed to study the involvement of opioid receptors in the antinociceptive activity and the potential ability to develop tolerance, crosstolerance, and/or physical dependence of tramadol. The writhes induced by acetic acid administration was used as algesiometric test. After chronic administration of tramadol, tolerance was evaluated by measuring the antinociceptive activity, and physical dependence was measured by naloxone administration. Morphine was used as drug of comparison. The i.p. administration of tramadol produced a dose-dependent antinociception with an ED50 value of 7.82 +/- 1.16 mg/kg, which was unchanged after chronic administration of either tramadol (39.1 or 100 mg/kg) or morphine (1.05 or 100 mg/kg). By contrast, the ED50 for morphine (0.21 +/- 0.08 mg/kg) was significantly reduced only by chronic pretreatment with both doses of morphine (tolerance). Physical dependence was developed only in mice pretreated with morphine, as evidenced by the presence of jumps, wet-dog shakes, tachypnea, piloerection, seizures, diarrhea, and urination after the administration of naloxone (1 mg/kg). These findings suggest that the antinociceptive activity of tramadol in mice is due to activation of opioid and nonopioid mechanisms, and as opposed to morphine, is not likely to induce tolerance and physical dependence.