Increasing Temperature Hot-plate Test Research Articles

Chronic, combined treatment with desipramine and mianserin: Enhanced 5-HT 1A receptor function and altered 5-HT 1A/5-HT 2 receptor interaction in rats

A greater percentage of depressed patients respond to combined treatment with a tricyclic antidepressant and the tetracyclic antidepressant mianserin than to treatment with these drugs given alone. The functional sensitivity of the 5-hydroxytryptamine (5-HT) 1A receptor, and the functional interaction between the 5-HT 1A and the 5-HT 2 receptors were investigated after treatment with despiramine and mianserin either alone or combined for 21–28 days. Pretreatment with desipramine and mianserin in combination induced the most intense 5-HT syndrome and the greatest fall in colonic temperature after injection of the 5-HT 1A agonist 8-hydroxy-2-(di- n-propylamino)-tetralin (8-OH-DPAT). The rats pretreated with desipramine alone had the largest elevation of the response temperature in the increasing temperature hot-plate test after injection of 8-OH-DPAT. After the combined pretreatment with desipramine and mianserin, no enhanced functional response in these tests was found when the 5-HT 1A and the 5-HT 2 receptors were stimulated simultaneously using 8-OH-DPAT and the 5-HT 2 agonist, (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropine HCl (DOI), contrasting the findings for desipramine or mianserin treatments given alone, where an increased functional response was found for the colonic temperature and the response temperature in the increasing temperature hot-plate test. In vitro receptor binding using [ 3H]-8-OH-DPAT as ligand revealed an increase in K d and K max in the spinal cord after chronic treatment with the combination of desipramine and mianserin. In the hippocampus and the frontal cortex the changes were small. Thus, the combination of desipramine and mianserin increased the functional response to 5-HT 1A receptor stimulation, and decreased the response to simultaneous stimulation of the 5-HT 1A and 5-HT 2 receptors, when compared to treatments with either one of the antidepressants alone, or controls. These rather large functional changes were not clearly reflected in the receptor binding study, indicating that changes in the postreceptor signal transduction may be of importance.

The role of 5-HT1A and 5-HT1B receptors in spinal nociceptive transmission and in the modulation of NMDA induced behaviour.

The effects on nociception of intrathecal (i.th.) administration of selective 5-HT1A and 5-HT1B agonists were studied in rats. Nociception was evaluated using the tail-flick test with adjustments for tail-skin temperature, the increasing temperature hot-plate test and the scoring of biting and scratching behaviour after i.th. N-methyl-D-aspartate (NMDA). Activation of the spinal 5-HT1A receptor induced an antinociceptive effect in the increasing temperature hot-plate test and produced a dose dependent decrease in NMDA-receptor mediated behaviour. No significant change in nociception measured by either of the nociceptive tests was found after administration of the 5-HT1B agonist. These results support the hypothesis that spinal 5-HT1A receptor activation has an antinociceptive effect, and indicate a possible interaction between the serotonergic and glutaminergic transmitter systems.

Acute and Long Term Effects of 1‐Methyl‐4‐phenyl‐1, 2, 3, 6‐tetrahydropyridine (MPTP) in Tests of Nociception in Mice

Acute and long term changes in nociception after administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) 80 mg/kg (four injections of 20 mg/kg given at two hr intervals) were investigated in mice. MPTP caused shivering, lacrimation, salivation, teeth chattering and fur erection a few minutes after drug injection, but all these behavioural changes were normalized within 30 min., when the first behavioural testing was performed. No significant alteration in general behaviour, sensorimotor performance or body temperature could be detected at the time of nociceptive testing. The acute effects of MPTP on nociception were a reduced response latency in the tail flick test and a prolonged response latency compared to controls in the constant temperature hot plate test. No significant effects of MPTP were found in the increasing temperature hot plate test. The long term effects were a reduced response latency both in the tail flick test and the constant temperature hot plate test, indicating that the MPTP induced lesions of dopaminergic pathways result in hyperalgesia. In the increasing temperature hot plate test and the formalin test, no significant long term changes were demonstrated. Seven days after injection, the dopamine content was reduced to 62% of control values in striatum, to 51% in the rest of the forebrain, and to 41% in the spinal cord. Noradrenaline levels were only slightly and transiently reduced. Serotonin levels were not affected 7 days after injection, but 14 days after injection, a great increase was found in the forebrain and in the spinal cord. The results suggest that dopaminergic systems tonically inhibit nociception.(ABSTRACT TRUNCATED AT 250 WORDS)

Desipramine, administered chronically, influences 5-hydroxytryptamine 1A-receptors, as measured by behavioural tests and receptor binding in rats

The 5-hydroxytryptamine 1A (5-HT 1A) receptor subtype seems to be of importance in the pathogenesis of depression and in the mode of action of antidepressants. In this study, behavioural experiments were performed in rats after oral administration of desipramine for 18–20 days, followed by an acute injection of the selective 5-HT 1A receptor agonist, 8-hydroxy-2-(di- n-propylamino)tetralin (8-OH-DPAT), either systemically or intrathecally. Chronic administration of desipramine prolonged the behavioural 5-HT syndrome in the animals injected systemically with 8-OH-DPAT. Treatment with desipramine was also found to potentiate and prolong the antinociceptive effect of an acute injection, systemically or intrathecally, of 8-OH-DPAT in the increasing temperature hot plate test. After systemic administration of 8-OH-DPAT, the colonic temperature was lowered similarly in the desipramine-treated group and in controls, whereas an intrathecal injection of 8-OH-DPAT resulted in a fall in the colonie temperature in the desipramine-treated group only. In vitro receptor binding studies, using [ 3H]8-OH-DPAT as the ligand, showed a statistically significant reduction of K d and B max in the frontal cortex and of K d in the spinal cord, after treatment with desipramine. No changes of K d and B max were found in the hippocampus after this treatment. Thus, desipramine, administered chronically, resulted in a functional up-regulation of the 5-HT 1A-receptors, both spinally and supraspinally, whereas in the in vitro receptor binding, a slight down-regulation or no change was found. It seems therefore that the results of in vitro receptor binding studies do not necessarily reflect the functional state of the neuronal system. The functional up-regulation of the system may be due to changes in the signal transduction processes, beyond the receptors.

The increasing-temperature hot-plate test: An improved test of nociception in mice and rats

The increasing-temperature hot-plate test has several advantages compared to the conventional hot-plate test, but available equipment has been impractical and restricted with regard to stimulus control. We now describe an apparatus consisting of an aluminum plate that is heated and cooled by Peltier elements in contact with its lower surface. Several plates can be used simultaneously, individually controlled by electronic proportional feedback circuits. The set temperature of the feedback circuit is controlled by a computer program run on an IBM XT-compatible PC, so that a linear increase in temperature is achieved. Experiments were performed using rats and mice, with hindpaw licking as an end-point criterion. Experiments with various heating rates showed that 3.0° C/min is the lowest rate that can be applied without signs of stress in the animals. On the basis of the recorded data, nociceptive temperature thresholds were calculated to be approximately 44.5°C for both rats and mice. Inspection of the paws after analgesic treatment and exposure to different end-point temperatures suggested that a cutoff temperature of 50°C should be employed to minimize tissue damage. Testing at ambient temperatures of 18° and 28°C yielded similar results for rats, whereas mice responded at significantly higher plate temperatures in the colder environment. Dose-related antinociceptive effects were demonstrated for morphine and paracetamol in both species. The results confirm that the increasingtemperature hot-plate test is a valuable test of nociception, which is also suitable for demonstrating the antinociceptive effects of nonopioid analgesics. The test may also be used to estimate the nociceptive temperature threshold.

Chronic administration of desipramine and zimelidine changes the behavioural response in the formalin test in rats

In studies of the effect on nociception of chronic administration of antidepressants, the stress of the injections may influence the results. In this experiment, desipramine or zimelidine were administered in the drinking water of rats, in a concentration yielding a dose of approximately 8 mg/kg/24 hr. Desipramine, given both for a short time (24 hr) and chronically (14 days), induced antinociception in the increasing temperature hot-plate test; zimelidine did not significantly influence the results of this test. In the tail-flick test, neither short-term nor chronic administration of these antidepressants had any effect on nociception, when correction was made for the changes in the temperature of the tail skin. In the formalin test, nine behavioural categories were scored for 1 hr and the data were treated statistically, using a multivariate analysis. Chronic administration of desipramine increased nociceptive behaviour during the first 10 min of the test. Desipramine and, to a lesser extent, zimelidine, changed the response in the late phase (10–60 min), showing less focussed pain-related behaviour (jerks and shaking, licking and biting of the injected paw) and more non-focussed pain-related behaviour (activity states with elevation or protection of the injected paw). It was concluded that desipramine is antinociceptive in the increasing temperature hot-plate test. Desipramine and zimelidine, administered chronically, modify the late phase of the formalin test towards less focussed pain-related behaviour, suggesting an antinociceptive effect. Multivariate analysis of the data of the formalin test seemed to be of value for the interpretation of the data.

Antinociceptive effect of intrathecally-administered desipramine and zimelidine in rats

The effect of intrathecally (i.th.) administered desipramine and zimelidine, in doses of 5, 10 and 50 μg, were investigated in the tail-flick test with simultaneous measurement of the temperature of the tail skin and in the increasing temperature hot-plate test. A constant negative correlation between the temperature of the tail skin and tail-flick latency, as described previously, was found. For all doses tested, desipramine induced longer tail-flick latencies, 10 min after injection than vehicle and the temperature of the tail skin tended to increase less in this group than in controls. After adjustment of the tail-flick latencies for the changes in the temperature of the tail skin, an antinociceptive effect of desipramine was still found. For zimelidine, only the largest dose (50 μg) was found to be antinociceptive, after adjustment for the tail skin temperature. In the increasing temperature hot-plate test, no antinociceptive effect of these antidepressants was found. For desipramine and zimelidine, the effect in the tail-flick test, 10 min after injection, indicates that the antinociceptive effect of these drugs may have, at least partly, a spinal site of action. In the increasing temperature hot-plate test, the response is integrated supraspinally. This may partly explain the lack of effect in this test when desipramine and zimelidine were administered intrathecally.

Diazepam Attenuates Morphine Antinociception Test‐Dependently in Mice

The influence of diazepam on the antinociceptive effect of morphine was studied using four different nociceptive tests in mice. In the tail flick test, diazepam induced a dose-dependent reduction of the morphine effect, with an almost total reversed morphine effect following diazepam 2 mg/kg. The effect could not be explained by altered tail skin temperature or pharmacokinetic changes. Diazepam 1 mg/kg and higher induced sedation and significantly impaired the performance in a rotarod test, a dose of 0.5 mg/kg diazepam was therefore used in the other nociceptive tests. This dose of diazepam significantly attenuated the antinociceptive effect of morphine in the constant temperature hot plate test and the tail flick test. In the increasing temperature hot plate test and in the formalin test, no effect of diazepam on the nociceptive effect of morphine was observed. The results indicate that diazepam antagonizes the effect of morphine dependent upon the test employed. No antagonism could be observed in tests with a long-lasting stimulus, and a response integration probably at a rather high level in the CNS. In the two tests showing antagonism, the stimulus is more short-lasting, and at least for the tail flick test, the integration takes place at a lower level in the CNS.

Desipramine in small doses induces antinociception in the increasing temperature hot-plate test, but not in the tail-flick test

The data reported for the antinociceptive effect of tricyclic antidepressants are conflicting. In this investigation, the effect of acute intraperitoneal (i.p.) administration of desipramine (2 and 5 mg/kg) was studied in rats, using the tail-flick test with simultaneous measurement of tail-skin temperature, and the increasing temperature hot-plate test. A constant negative correlation between tail-flick latency and tail-skin temperature, as described earlier, was also found in this study. Different ambient temperatures influenced the results of the tail-flick test. At an ambient temperature of 24–25°C, desipramine gave rise to an apparent antinociception in the tail-flick test, which was found to be caused by a relative fall in tail-skin temperature. At 21–22°C, no change in tail-flick latency was found after the administration of desipramine. In the increasing temperature hot-plate test, however, a dose-dependent antinociceptive effect of desipramine was observed at both ambient temperatures. The strong influence of moderate differences in ambient temperature on the results of the tail-flick test may explain some of the conflicting results reported in the literature. Whenever this test is used, the temperature of the tail should be recorded and taken into account in the evaluation of the data.

Mechanisms of orhenadrine-induced antinociception in mice: a role for serotonergic pathways

The possible involvement of central serotonergic pathways in the mechanism of action of orphenadrine citrate was investigated in male albino mice. Orphenadrine (20 mg/kg) did not alter the concentration of 5-hydroxytryptamine (5-HT) or its metabolite 5-hydroxyindole acetic acid in the frontal cortex or spinal cord, nor did it, in moderate concentrations, inhibit the uptake of [ 14C]5-HT, [ 3H]noradrenaline ([ 3H]NA) or [ 3H]dopamine ([ 3H]DA) into crude synaptosomal preparations from the cortex. The antinociceptive effect of orphenadrine was studied in the formalin test and in the increasing temperature hot plate test. No sensorimotor impairment was observed for doses of 30 mg/kg or lower. A general depletion of serotonin by means of p-chlorophenylalanine significantly reduced the effect of orphenadrine in both tests, while lesion of the ascending serotonergic systems by means of p-chloroamphetamine did not affect the analgesia. It is concluded that the antinociceptive effect of orphenadrine may be mediated in part via the raphe-spinal serotonergic systems.

The relative contribution of ascending and descending serotonergic pathways in p-Chloroamphetamine-induced antinociception

Systemic administration in rats of p-chloroamphetamine (PCA; 2×10 mg/kg) reduced the in vitro uptake of 14C-5-hydroxytrypamine ( 14C-5-HT) in cortical synaptosomes by 76% and in spinal cord synaptosomes by 35%. Intrathecal injection of 5,6-dihydroxytryptamine (20 μg/rat) selectively lesioned the descending serotonergic pathways (83% reduction in uptake of 14C-5-HT in spinal synaptosomes, no significant change in uptake in cortical synaptosomes). Administration of PCA or 5,6-DHT did not significantly alter the uptake of 3H-noradrenalin into cortical or spinal synaptosomes. The response thresholds of the rats in the increasing temperature hot plate test (1 to 7 days after administration) were unaffected by either type of lesion. Interference with the antinociceptive effect of PCA (2.5 mg/kg) was evaluated 7 days after administration of the neurotoxins. PCA pretreatment strongly reduced the peak of the PCA-induced antinociception while 5,6-DHT reduced its duration. Thus, both ascending and descending serotonergic pathways contribute to PCA-induced antinociception.

Modification of the antinociceptive effect of morphine by acute and chronic administration of clomipramine in mice

The antinociception effect of clomipramine and the combination of clomipramine and morphine was evaluated using the constant temperature hot plate test, the increasing temperature hot plate test and the formalin test in mice. Both a single dose of clomipramine (10 mg/kg) given intraperitoneally (i.p.) and chronic clomipramine treatment (10 mg/kg/day, i.p., for 7 days) induced antinociception. When morphine (5 mg/kg), given subcutaneously (s.c.), and clomipramine were combined, an increased antinociceptive effect compared to morphine alone was observed in all tests. In the increased temperature hot plate test and in the formalin test increased antinociception was also observed when morphine was combined with chronic clomipramine treatment. In the constant temperature hot plate test no additional effect compared to morphine alone was observed when morphine and chronic clomipramine treatment were combined. These results indicate that clomipramine has an antinociceptive effect in itself, and that it may enhance the antinociceptive effects of morphine. The differences between the tests indicate that different nociceptive mechanisms are involved, and that the combination of clomipramine and morphine may be beneficial for some types of nociceptive stimuli, but not for others.

Mechanisms by which the putative serotonin receptor antagonist metitepin alters nociception in mice.

The putative serotonin (5-HT) receptor antagonist metitepin (0.5 mg/kg, intraperitoneally) produced hypoalgesia in the increasing temperature hot-plate test and hyperalgesia in the tail-flick test in mice. The effects of metitepin were not altered after depletion of 5-HT by the neurotoxin 5,7-dihydroxytryptamine (5,7-DHT, 80 micrograms free base, intracerebroventricularly) or the serotonin synthesis inhibitor p-chlorophenylalanine (PCPA, 400 mg/kg for 10 consecutive days). After chronic administration (2 or 5 mg/kg for 18 consecutive days) tolerance to the effect of metitepin (0.5 mg/kg) and cross-tolerance to the antinociceptive effect of the 5-HT agonist 5-methoxy-N,N-dimethyltryptamine (5-MeODMT, 3 mg/kg) was found in the hot-plate test but not in the tail-flick test. It is suggested that metitepin may block descending 5-HT transmission while more complex mechanisms of action are involved at supraspinal level. One possibility is that metitepin exhibits partial agonist properties or, alternatively, that the drug may block 5-HT subsystems which tonically enhance nociception.

5-HT depletion with 5,7-DHT, PCA and PCPA in mice: differential effects on the sensitivity to 5-MeODMT, 8-OH-DPAT and 5-HTP as measured by two nociceptive tests

Depletion of 5-hydroxytryptamine (5-HT) in mice was produced by intracerebroventricular injection of 5,7-dihydroxytryptamine (5,7-DHT, 80 μg) or by systemic injections of p-chloroamphetamine (PCA, 3 × 40 or 4 × 40 mg/kg), p-chlorophenylalanine (PCPA, 5 × 400 or 14 × 400 mg/kg) or combined PCA (3 × 40 mg/kg) + PCA (11 × 400 mg/kg). Neither of the pretreatments altered nociception in the increasing temperature hot-plate test, whereas hyperalgesia was demonstrated in 5,7-DHT lesioned animals in the tail-flick test, 5,7-DHT-pretreatment enhanced the antinociceptive effect of the 5-HT agonists 5-methoxy-N,N-dimethyltryptamine (5-MeODMT), 8-hydroxy-2-(di- n-propylamino)tetralin (8-OH-DPAT) and 5-hydroxytryptophan (5-HTP). This effect was observed after 2.5 and 8 days in the tail-flick test and after 5 and 8 days in the hot-plate test. However, pretreatment with PCPA or PCA failed to alter the antinociception elicited by the 5-HT agonists, although a tendency towards enhancement of antinociception was found after combined treatment with PCA and PCPA. It is suggested that the injection of 5,7-DHT induces denervation supersensitivity of post-synaptic 5-HT receptors. The lack of such supersensitivity after PCPA-pretreatment which induces similar 5-HT depletion to 5,7-DHT, may suggest that other factors than the absence of 5-HT may contribute to the development of denervation supersensitivity. Alternatively, the three 5-HT depleting agents may produce a qualitatively different reduction of 5-HT.

Test-dependent changes in nociception after administration of the putative serotonin antagonist metitepin in mice

Intraperitoneal administration of the putative serotonin receptor antagonist metitepin (0.06–1.0 mg/kg) in mice induced dose-dependent antinociception in the increasing temperature hot-plate test and the formalin test, but elicited hyperalgesia in the tail-flick test. Reduced motor activity was observed after injection of the largest dose of metitepin, but did not influence the behavioural responses in the tests. Selective lesions of ascending serotonergic pathways induced by administration of the neurotoxin p-chloroamphetamine 5 and 6 days before testing (40 mg/kg each day) did not directly affect the responsiveness in any of the tests but enhanced the metitepin-induced antinociception in the hot-plate and formalin tests. The hyperalgesia in the tail-flick test was not affected by the lesions. The results suggest that metitepin may alter nociception in mice by exhibiting both agonist and antagonist properties on central serotonergic receptors.

Involvement of central serotogenic pathways in nefopam-induced antiniciception

The possible involvement of central serotonergic pathways in the mechanism of action of nefopam was investigated in male albino mice. Nefopam (15 mg/kg i.p.) did not alter the concentration of serotonin or its metabolite 5-hydroxyindole acetic acid in frontal cortex or spinal cord. Lesions of the ascending serotonergic pathways were made by systematic administration of p-chloroamphetamine (PCA). Serotonin depletion in all serotonergic systems was obtained by means of p-chlorophenylalanine (PCPA). Two different nociceptive assays were used, the formalin test and the increasing temperature hot plate test. PCPA pretreatment significantly reduced the effect of nefopam (15 mg/kg) in the formalin test. In contrast, nefopam-induced analgesia was not affected by PCA pretreatment, either in the formalin test or in the increasing temperature hot plate test. In conclusion, the data suggest that descending serotonergic pathways are involved in nefopam-induced antinociception.

A modified hot-plate test sensitive to mild analgesics.

The present study compares a modified hot-plate test in which the temperature is slowly increased from non-noxious levels with a standard constant temperature hot-plate test. In both tests, hindpaw lick was found to be a more reliable criterion response than forepaw lick, and was employed throughout the experiments. In the constant temperature hot-plate test, 1-min exposure to the test apparatus the day before testing significantly reduced the response latencies of both rats and mice. No effect of pre-exposure was found in the increasing temperature test. In both tests and in both species, sessions of repeated testing were conducted with only insignificant alterations in responsiveness between trials. In both rats and mice, dose-related increases in response temperature were obtained in the increasing temperature hot-plate test after administration of morphine, paracetamol and acetylsalicylic acid, whereas only morphine had consistent effects in the constant temperature test. Thus, the increasing temperature hot-plate is a useful analgesimetric test in both rats and mice, superior to the conventional hot-plate test with regard to consistency of results and sensitivity to non-narcotic analgesics.

Orphenadrine citrate increases and prolongs the antinociceptive effects of paracetamol in mice.

Orphenadrine, a muscle relaxant with antinociceptive effects, was shown to increase and prolong the antinociceptive effects of paracetamol in mice. Both in the increasing temperature hot plate test and in the formalin test, a combination of the two drugs showed a significantly improved effect compared to either of the drugs alone. The time course of the effects was tested in the increasing temperature hot plate test. The group treated with the drug combination showed a prolonged effect compared to both single drug treated groups, the effect lasting longer than 120 min. for the combination and about 80 min. for the single drugs. Orphenadrine and paracetamol increased antinociception even when orphenadrine was injected 90 min. after paracetamol, which by that time did not exert antinociceptive effects by itself. Thus the combination of orphenadrine and paracetamol enhances the antinociceptive effect of either drug in mice.

Antinociceptive effects of orphenadrine citrate in mice

The antinociceptive effect of orphenadrine citrate, a muscle relaxant, was investigated in mice. Four different pain tests were selected to involve different noxious stimuli. Clear antinociceptive effects were found in the formalin test. The increasing temperature hot-plate test showed a biphasic dose-response relationship with slight hyperalgesia after low doses and hypoalgesia after higher doses. No significant effects of orphenadrine (0–25 mg/kg) were found in the tail flick and constant temperature hot-plate tests. The data suggest that orphenadrine may reduce, enhance or leave unaffected different types of nociceptive transmission. Orphenadrine may also possess analgesic properties in conditions not involving muscle spasm.