Effects of repeated as compared to single aggressive confrontation on nociception and defense behavior in C57BL/6 and DBA/2 mice

Behavioral reactions (submissive postures, escape, immobility, activity, locomotion) in C57BL/6 and DBA/2 test mice were recorded during single (50 bites) or three repeated (3 X 50 bites, separated by 24 hr) aggressive confrontations, as well as during a nonaggressive confrontation 24 hr after the last aggressive confrontation with opponents of the opposite strain. Nociception (hot plate response latency) was measured 1 min after aggressive or nonaggressive confrontations. During repeated aggressive confrontation, DBA mice reacted with a stable pattern of escape and analgesia, whereas C57 mice failed to develop an analgesic response and changed their behavioral defense strategy during repeated aggressive confrontations (decrease of escape, increase of defensive upright). The conditioned display of submission and of escape behavior during nonaggressive confrontation did not change as a function of earlier repeated aggressive confrontations in DBA mice, while C57 mice showed a significant increase of defensive upright postures and immobility. Conditioned analgesia was not observed after nonaggressive confrontations. The results point toward a dissociation between attack-elicited behavior and antinociception and suggest that encounter-induced analgesia may influence the processing of aversive experience.     

Systemic administration of naloxone produces analgesia in BALB/c mice in the formalin pain test

Systemic administration of naloxone usually produces either hyperalgesia or no change in nociception depending on the animal species used and/or the pain test employed. This study, however, demonstrates that naloxone produces a dose-dependent analgesia in the formalin pain test using an inbred strain of albino mouse. Female BALB/c, C57BL/6 and CD1 mice were injected subcutaneously with naloxone HCl in saline (0.1 10.0 mg/kg) or saline alone, and tested for analgesia using the formalin test. Naloxone produced a statistically significant dose-dependent analgesia in the BALB/c mice, with an ED50 of 0.24 mg/kg and almost total analgesia at doses of 1 mg/kg or greater. No changes in pain behaviour were observed in the C57BL/6 or CD1 strains of mice. We believe this to be the first report of analgesia following administration of doses of naloxone normally used for opioid antagonism. To determine if this effect was specific to the formalin test, the 3 strains of mice were injected subcutaneously with naloxone HCl and tested in the tail-flick test. Naloxone had no analgesic action in this test in any of the strains.     

Involvement of endogenous opioids with forced swimming-induced immobility in mice

The present study investigated the involvement of endogenous opioid mechanisms with the immobility response induced in mice by forced swimming. Pretreatment with the narcotic antagonist naloxone (0.625--40.0 mg/kg) caused a dose-dependent decrease in the duration of immobility in mice subjected to a 10 min swim test. This effect was more pronounced in C57BL/6J mice than in BALB/C mice. A low dose of morphine (0.15 mg/kg) potentiated immobility whereas higher doses (0.625/10.0 mg/kg) had no demonstrable effect on immobility in these strains. The results suggest that release of endogenous opioids may be a physiological event promoting natural cataleptic-like behaviors in mice.     

Social conflict activates status-dependent endogenous analgesic or hyperalgesic mechanisms in male mice: effects of naloxone on nociception and behaviour

The concept of environmentally-induced activation of endogenous analgesia mechanisms rests, almost exclusively, upon studies which have involved the use of rather intense artificial stimuli. The current study was therefore designed to assess the validity of this concept under the more naturalistic conditions of social conflict between isolated resident mice and group-housed intruders. Agonistic experience was found to result in a potent, naloxone-reversible (10 mg/kg) analgesia in intruder mice while, in residents, it produced a moderate hyperalgesic reaction which was very sensitive to naloxone antagonism (0.1 mg/kg). Detailed videotape analyses revealed that only the behaviour of residents was significantly altered by naloxone treatment, with a highly selective inhibition of attack observed at 10 mg/kg. These data suggest that (1) social conflict in mice is a potent, and biologically-relevant, stimulus in the activation of endogenous naloxone-sensitive pain control mechanisms, (2) social status is an important determinant of nociceptive response to such experience and (3) inescapability from attack may be a critical factor in the development of encounter-induced analgesia.     

Naloxone partially antagonizes post-encounter analgesia and enhances defensive responding in male rats exposed to attack from lactating conspecifics

Recent evidence indicates that inter-male conflict in mice, but not rats, results in opiate-like analgesia in defeated animals. To test the hypothesis that differences in attack intensity/controllability may underlie this apparent species difference, male rats were exposed to non-ritualized attack from lactating conspecifics. In response to such encounters, male intruders exhibited significant elevations in tail-flick latency. Pretreatment with low (0.1–1.0 mg/kg), but not high (10 mg/kg), doses of naloxone partially antagonized the development of post-encounter analgesia and enhanced several components of male defensiveness during encounters. These results suggest that exposure to attack from lactating conspecifics activates both opioid and nonopioid analgesia mechanisms in male intruders. Correlational analyses revealed, however, that intruder analgesia was not related to the number of attacks received or the display of submissive postures. Data are discussed with reference to the possible importance of psychological factors in the activation of endogenous pain inhibitory mechanisms in rats.     

Acute non-opioid analgesia in defeated male mice

Exposure to repeated attack induces a long-lasting analgesia in male mice. Although this reaction has been linked to the special biological significance of defeat, earlier research has confounded defeat and exposure to further attack. In the present studies, DBA/2 intruder mice were individually placed into the home cages of aggressive conspecifics and removed immediately upon display of the species-characteristic upright submissive posture. Under these test conditions, intruders did indeed show a profound analgesia. However, in marked contrast to the antinociceptive effects of repeated attack, this reaction was of short duration (less than 10 min), was not blocked by naloxone (1-10 mg/kg, IP) and did not show cross-tolerance either to or from morphine (5 mg/kg, IP). These findings are discussed in relation to multiple endogenous pain inhibitory systems and their possible adaptive significance in murine social behaviour.     

Quaternary naltrexone reverses morphine-induced behaviors

This study explored the relative role of the peripheral and central nervous systems (CNS) in the production of morphine-induced behavioral changes. Toward this end we used a quaternary derivative of an opiate antagonist (naltrexone methobromide, NM) that presumably does not cross the blood-brain barrier. Naltrexone methobromide (20, 40 and 80 mg/kg, IP) was used to challenge the stereotypic locomotion, analgesia and elevated "Straub" tail response observed in C57BL/6J mice after a 30-mg/kg (IP) injection of morphine. The quaternary derivative of naltrexone reversed the locomotor hyperactivity, "Straub" tail and analgesia normally observed in the opiate-treated C57BL/6J mouse. The data reported here, if taken at face value, suggest an important role for peripheral opiate receptors in morphine-induced behavioral changes. However, these conclusions are contingent on further research to more fully evaluate NM's capacity to cross the blood-brain barrier of the C57BL/6J mouse.     

Naloxone fails to produce conditioned place aversion in mu-opioid receptor knock-out mice.

There is growing evidence that tonic activity of the opioid system may be important in the modulation of affective state. Naloxone produces a conditioned place aversion in rodents, an effect that is centrally mediated. Previous pharmacological data using antagonists with preferential actions at mu-, delta-, and kappa-opioid receptors indicate the importance of the mu-opioid receptor in mediating this effect. We sought to test the mu-opioid receptor selectivity of naloxone aversion using mu-opioid receptor knock-out mice. mu-Opioid receptor knock-out and wild-type mice were tested for naloxone (10 mg/kg, s.c.) aversion using a place conditioning paradigm. As a positive control for associative learning, knock-out mice were tested for conditioned place aversion to a kappa agonist, U50,488H (2 mg/kg, s.c.). Naloxone produced a significant place aversion in wild-type mice, but failed to have any effect in mu-opioid receptor knock-out mice. On the other hand, both knock-out and wild-type mice treated with U50,488H spent significantly less time in the drug-paired chamber compared to their respective vehicle controls. We conclude that the mu-opioid receptor is crucial for the acquisition of naloxone-induced conditioned place aversion. Furthermore, in a separate experiment using C57BL/6 mice, the delta-selective antagonist naltrindole (10 or 30 mg/kg, s.c.) failed to produce conditioned place aversion.Taken together, these data further support the notion that naloxone produces aversion by antagonizing tonic opioid activity at the mu-opioid receptor.     

Effects of pre-training morphine on spatial memory acquisition and retrieval in mice

The opioid system plays an important role in memory processess. Morphine mimics endogenous opioids by acting on opioid receptor in brain to regulate memory. However, the effects of morphine on spatial memory acquisition are controversial. Also, little evidence has suggested that morphine could affect the retrieval of spatial memory. In the current study, effects of pre-training morphine and naloxone on the acquisition vs. retrieval of spatial reference vs. working memory were examined using discrete water maze tasks in C57BL/6 mice. Pre-training morphine administration (7.5 and 15 mg/kg, i.p.) impaired the acquisition of both spatial reference memory and working memory. Motivation to escape from the water maze was not affected by morphine. Pre-test morphine also inhibited the retrieval of spatial working memory but not reference memory. The effects of morphine on the acquisition and retrieval of spatial working memory were eliminated by naloxone pretreatment (1 mg/kg). These results indicate that morphine could differentially modulate a variety of aspects of spatial memory and these effects are mediated by the mu-opioid receptor.     

The effect of estrogens and antiestrogens in a rat model for hot flush

The present studies evaluated the effect of estrogens and the selective estrogen receptor modulator (SERM) tamoxifen and raloxifene in a rat model for hot flush. In this model, ovariectomized rats were treated for 8 or 9 days either sc or po. Rats were dependent to morphine by implanting a morphine pellet (75 mg each) sc on days 3 and 5 of treatment. On the last day of treatment, a thermistor, connected to a data acquisition system, was placed on the tail of each animal and morphine addiction was withdrawn by naloxone injection (1.0 mg/kg, sc). Temperature measurements were taken for 1 h under ketamine (80 mg/kg, im) anesthesia. In general, vehicle treated rats showed a 5–6°C elevation of their tail skin temperature with the peak occurring about 15 min after naloxone injection. 17-Ethinyl estradiol (EE) was evaluated both sc and po using a broad range of doses. The IC₅₀ for inhibition of tail skin temperature rise was approximately 0.1 mg/kg, sc and 0.2 mg/kg, po. 17β-Estradiol and 17-estradiol were also active in this model whereas non-estrogenic steroids were inactive. Raloxifene and tamoxifen were tested for estrogen agonist and antagonist activity administered sc and po. Raloxifene did not demonstrate reproducible estrogen agonist activity at doses up to 10 mg/kg, whereas it demonstrated significant antagonistic activity at the 10 mg/kg dose regardless of the route of administration. Tamoxifen exhibited significant estrogen agonist activity at all doses tested (0.1–10.0 mg/kg) and was a significant antagonist of EE at the 1.0 mg/kg dose. Our results demonstrate the potential utility of this model to evaluate and discriminate among classes of compounds with varying degrees of estrogen agonist and antagonist activity.     

Morphine influences on classical aversive conditioned heart rate in rats

The present series of three experiments was concerned with the effects of morphine and the morphine antagonist naloxone on the development of a classical aversive heart rate (HR) conditioned response (CR) to a tone conditioned stimulus (CS) paired with an electric shock unconditioned stimulus (US). In the first study, separate groups of rats received preconditioning sc injections of either 0.25 mg/kg, 5 mg/kg, or 10 mg/kg of morphine. Three other groups were given 0.1 mg/kg, 5 mg/kg, or 10 mg/kg of naloxone alone. All of the morphine groups showed attenuation HR responses to the CS on preconditioning CS-alone trials. During conditioning, the 10-mg/kg morphine group showed a markedly decremented bradycardia CR and tachycardia unconditioned response (UR), whereas the 5-mg/kg morphine group showed a normal CR in combination with a decremented UR. Naloxone had no measurable effects on HR. In the second study, naloxone (1 mg/kg) given after conditioning failed to reverse the CR and UR losses produced by 10 mg/kg of morphine given prior to conditioning. Administration of 10 mg/kg of morphine produced only a minor reduction in a HR CR established in a drug-free state, but the tachycardia UR was severely reduced. The results of the third study showed that 1 mg/kg of naloxone was effective in reversing analgesia induced by 10 mg/kg of morphine, as indexed by the tail-flick test. Taken together, the results suggest that the 10-mg/kg dose of morphine interfered with the learning of a HR CR, perhaps principally by reducing the aversive or emotional consequences of the shock US. Direct cardiovascular effects of morphine seemed to interfere with the performance of the tachycardia UR, but not with the performance of the bradycardia CR.     

Unique behavioural phenotypes of recombinant-inbred CXBK mice: partial deficiency of sensitivity to mu- and kappa-agonists.

Recombinant-inbred CXBK mice have been used for various studies as putative mu-opioid-receptor deficient mice. However, CXBK mice have never been compared with gene-targeting mice lacking the mu-opioid receptor (muKO) and the K-opioid receptor (kappaKO). Here we report that CXBK mice show distinct behavioural phenotype in opioid-induced analgesia and sedation. Intraperitoneal (i.p.) administration of morphine (3 and 10 mg kg(-1)) induced significantly lower levels of analgesia in CXBK mice than in the control C57BL/6 mice, while higher doses of morphine (30 and 100 mg kg(-1)) induced marked analgesia in CXBK mice. CXBK mice also showed lower analgesia and sedation levels than did C57 mice after i.p. administration of U-50488 (10 and 30 mg kg(-1)). The partial deficiency of sensitivity to morphine and U-50488 of CXBK mice is in sharp contrast to the complete lack of sensitivity to morphine and U-50488 in muKO and kappaKO mice, respectively. Furthermore, CXBK mice showed a lower threshold for nociceptive stimuli when they were not given an opioid, suggesting that CXBK mice could have alterations in the genes related to the nociceptive threshold. These unique behavioural phenotypes of CXBK mice suggest unique genetic alterations in CXBK mice.     

Synergistic interaction between mazindol, an anorectic drug, and swim-stress on analgesic responses in the formalin test in mice

The present study examined the interaction between mazindol (MZ), an anorectic drug extensively used in Brazil and opioid/non-opioid endogenous analgesic systems activated by swim-stress. Further, the role of opioid, dopamine and N-methyl-D-aspartate (NMDA) receptors in mediating the analgesic effect was evaluated. The stress-induced analgesia of a 3-min swimming at 32 degrees C (opioid/non-opioid) and 20 degrees C (non-opioid) were assessed using the formalin test. Male Swiss mice were intraperitoneally injected with naloxone (1.0 mg/kg), sulpiride (3.0 mg/kg), MK-801 (0.075 mg/kg) or saline/vehicle 15 min prior, and with MZ (0.5 mg/kg) or saline/vehicle 5 min prior to swimming. The dose of MZ (0.5 mg/kg) did not cause analgesic effect, however, the association of MZ and swim-stress at both temperatures displayed synergistic interaction on analgesia that was blocked by sulpiride and MK-801 but not by naloxone. The present results suggest that MZ and swim-stress acted synergistically on analgesic responses, involving mainly the non-opioid component and possibly mediated by dopamine D2 receptors and NMDA receptors.     

A genetic study of male social aggression in wild and laboratory mice

For a genetic analysis of aggression, wild males were crossed to two inbred strains, BALB/c and C57BL, and to wild females. The male progeny were tested against BALB/c and subsequently against C57BL standard opponents in a neutral arena during two successive 5-min encounters. Attack latency, number of bites, number of attacks, and total attack time were highly correlated within each 5-min period but the correlations between periods were relatively low. In the first 5 min, mice with a BALB/c mother were far more aggressive toward the C57BL opponent, and vice versa. This maternal effect complicated the genetic analysis, but there was considerable additive genetic variation for all the main measures, the narrow heritabilities ranged from 0.56 to 0.75, and there was little directional dominance for high aggression. The genetic architecture suggested that stabilizing selection is the main selective force behind the evolution of variation in male social aggression.     

Opioid modulation of reflex versus operant responses following stress in the rat

In pre-clinical models intended to evaluate nociceptive processing, acute stress suppresses reflex responses to thermal stimulation, an effect previously described as stress-induced "analgesia." Suggestions that endogenous opioids mediate this effect are based on demonstrations that stress-induced hyporeflexia is enhanced by high dose morphine (>5 mg/kg) and is reversed by naloxone. However, reflexes and pain sensations can be modulated differentially. Therefore, in the present study direct comparisons were made of opioid agonist and antagonist actions, independently and in combination with acute restraint stress in Long Evans rats, on reflex lick-guard (L/G) and operant escape responses to nociceptive thermal stimulation (44.5 degrees C). A high dose of morphine (>8 mg/kg) was required to reduce reflex responding, but a moderate dose of morphine (1 mg/kg) significantly reduced escape responding. The same moderate dose (and also 5 mg/kg) of morphine significantly enhanced reflex responding. Naloxone (3 mg/kg) significantly enhanced escape responding but did not affect L/G responding. Restraint stress significantly suppressed L/G reflexes (hyporeflexia) but enhanced escape responses (hyperalgesia). Stress-induced hyperalgesia was significantly reduced by morphine and enhanced by naloxone. In contrast, stress-induced hyporeflexia was blocked by both naloxone and 1 mg/kg of morphine. Thus, stress-induced hyperalgesia was opposed by endogenous opioid release and by administration of morphine. Stress-induced hyporeflexia was dependent upon endogenous opioid release but was counteracted by a moderate dose of morphine. These data demonstrate a differential modulation of reflex and operant outcome measures by stress and by separate or combined opioid antagonism or administration of morphine.     

Loss of Aggression, After Transfer onto a C57BL/6J Background, in Mice Carrying a Targeted Disruption of the Neuronal Nitric Oxide Synthase Gene

Phenotypic differences among mice with disrupted genes and those with wild-type alleles have not provided the necessary evidence for desired gene/phenotype correlations. These differences could be due to "passenger genes" from the donor 129 strains that are used to produce stem cells. Three variations of attack behavior were measured, using mice carrying a disruption of the neural nitric oxide synthase gene. In the first population, the disrupted gene had been maintained on a mixed background including C57BL/6J and 129 alleles. We have developed a second population in which the disrupted gene was transferred onto a C57BL/6J background during five backcross generations. On the mixed C57BL/6J-129 background, mice homozygous for disrupted Nos1 alleles attacked more frequently, had shorter attack latencies, and presented a greater number of attacks than mice carrying nondisrupted alleles. On the C57BL/6J background, no significant difference persisted between the carriers of the disrupted gene and their noncarrier siblings. The noncarriers on the mixed C57BL/6J-129 background, and the carriers or noncarriers on the C57BL/6J background, did not differ from C57BL/6J. The frequency of attacking males was identical in the homozygous carriers of the disrupted gene, in the mixed C57BL/6J-129 background, and in the 129/SvPas, which approximates the 129/SvJae strain from which the stem cells were derived to produce the disrupted Nos1 gene. These results suggest that Nos1 disruption was not implicated in attack behavior. A possible passenger-gene effect from the 129 donor strain is discussed.     

The calls of murine predators activate endogenous analgesia mechanisms in laboratory mice.

In view of the suggested role of endogenous analgesia mechanisms as an antipredator defense mechanism, the effects on nociception of exposure to the calls of various murine predatory and nonpredatory species were assessed. Data revealed that the calls of the Tawny Owl, Barn Owl and Common Gull all induced significant analgesia following exposure to 2 min of birdsong. Time course analysis revealed the analgesia induced by the Tawny Owl call to have a duration in excess of 40 min while the Barn Owl and Gull call-induced analgesias were much shorter lasting (approximately 10 min or less). Five mg/kg naloxone was found to attenuate the analgesia induced by the Tawny and Barn Owls but not the Common Gull. Together, these data suggest that brief exposure to the calls of night-hunting, aerial predators activate endogenous opioid-mediated analgesia mechanisms in mice.     

Dose-related induction of hepatic preneoplastic lesions by diethylnitrosamine in C57BL/6 mice

The C57BL/6 mouse strain (or derivation of this strain) is used as a background for many transgenic mouse models. This strain has a relatively low susceptibility to chemically induced hepatocarcinogenesis compared with other commonly used experimental mouse strains. In the present study, the authors treated C57BL/6 mice with 25, 50, and 75 mg/kg of diethylnitrosamine (DEN) for 4 or 8 weeks by intraperitoneal injection to investigate the dose-response pattern of preneoplastic and neoplastic lesion formation in the liver. DEN induced preneoplastic lesions and cytokeratin 8/18-positive foci in a dose-dependent manner. In the 75 mg/kg for 8 weeks treatment group, hepatocellular adenoma, cholangioma and hemangioma, and cytokeratin 19-positive foci were also induced, but a significant decrease in body weight was observed. The suitable DEN treatment range for this strain was concluded to be from 75 mg/kg for 4 weeks (total amount = 300 mg/kg) to 50 mg/kg for 8 weeks (total amount = 400 mg/kg). These results should prove useful for future studies investigating hepatocarcinogenesis in both the background C57BL/6 strain and other transgenic mouse models derived from it.     

Dual role of interleukin-4 (IL-4) in pulmonary paracoccidioidomycosis: endogenous IL-4 can induce protection or exacerbation of disease depending on the host genetic pattern

Resistance to paracoccidioidomycosis, the most important endemic mycosis in Latin America, is thought to be primarily mediated by cellular immunity and the production of gamma interferon. To assess the role of interleukin-4 (IL-4), a Th2 cytokine, pulmonary paracoccidioidomycosis in IL-4-depleted susceptible (B10.A) and intermediate (C57BL/6) mice was studied. Two different protocols were used to neutralize endogenous IL-4 in B10.A mice: 1 mg of anti-IL-4 monoclonal antibody (MAb)/week and 8 mg 1 day before intratracheal infection with 10(6) Paracoccidioides brasiliensis yeast cells. Unexpectedly, both protocols enhanced pulmonary infection but did not alter the levels of pulmonary cytokines and specific antibodies. Since in a previous work it was verified that C57BL/6 mice genetically deficient in IL-4 were more resistant to P. brasiliensis infection, we also investigated the effect of IL-4 depletion in this mouse strain. Treatment with the MAb at 1 mg/week led to less severe pulmonary disease associated with impaired synthesis of Th2 cytokines in the lungs and liver of control C57BL/6 mice. Conversely, in IL-4-depleted C57BL/6 mice, increased levels of tumor necrosis factor alpha and IL-12 were found in the lungs and liver, respectively. In addition, higher levels of immunoglobulin G2a (IgG2a) and lower levels of IgG1 antibodies were produced by IL-4-depleted mice than by control mice. Lung pathologic findings were equivalent in IL-4-depleted and untreated B10.A mice. In IL-4-depleted C57BL/6 mice, however, smaller and well-organized granulomas replaced the more extensive lesions that developed in untreated mice. These results clearly showed that IL-4 can have a protective or a disease-promoting effect in pulmonary paracoccidioidomycosis depending on the genetic background of the host.     

美法仑治愈荷瘤小鼠的过程与TNFα的关系

目的探讨单一剂量的美法仑治愈荷瘤野生型C57BL/6小鼠的过程与肿瘤坏死因子α(TNFα)的关系。方法以3种遗传背景相同、肿瘤坏死因子受体1(TNFR1)基因型不同的TNFR1 / 、TNFR1 /-和TNFR1-/-C57BL/6小鼠为实验动物,皮下接种数量相同的小鼠淋巴瘤EL4细胞。接种瘤细胞后12d,给基因型不同的各组荷瘤小鼠腹腔内单次注射7.5mg/kg的美法仑。以荷瘤野生型C57BL/6小鼠(TNFR1 / )为对照,观察美法仑对荷瘤TNFR1 /-C57BL/6小鼠和荷瘤TNFR1-/-C57BL/6小鼠的治疗效应。结果在美法仑(7.5mg/kg)治疗后的1周内,基因型不同的各组荷瘤小鼠肿瘤消退的速度基本相同。在随后的2月内,荷瘤TNFR1 / 和TNFR1 /-C57BL/6小鼠的肿瘤结节逐渐消退、肿瘤治愈;而多数荷瘤TNFR1-/-C57BL/6小鼠的肿瘤结节缩小后又再次出现并逐渐长大、肿瘤复发。结论TNFα与美法仑治愈肿瘤的过程密切相关,其中美法仑的抗肿瘤作用与荷瘤小鼠TNFR1的表达无关,但在美法仑治疗后,机体预防或避免肿瘤复发方面需要TNFR1在机体细胞的表达,而不是在肿瘤细胞的表达。