Frances V. Abbott

The formalin test: scoring properties of the first and second phases of the pain response in rats

&NA; The formalin test is increasingly used as a model of injury‐produced pain but there is no generally accepted method of pain rating. To examine the properties of various pain rating methods we established dose‐response relations for formalin injected in the plantar surface of one hind paw, and the analgesic effects of morphine and amphetamine using the most frequently reported behavioural measures of pain (favouring, lifting, licking and flinching/shaking of the injured paw) and combinations of these. Licking, elevation and favouring of the injected paw showed a biphasic response at all formalin doses. Flinching varied in form across the time course of formalin, and the biphasic nature of the behaviour was not as apparent. In untreated rats all these behaviours were infrequent. Flinching and favouring were increased after injection of local anaesthetic into the paw but remained negligible relative to the effect of formalin. Grooming other than that directed paw was elevated in a dose‐dependent manner by formalin. Intercorrelations between the behaviours were different for the initial response and the second phase. Correlational analysis indicated that no single behavioural measure was a strong predictor of formalin, morphine and amphetamine dose. A simple sum of time spent licking plus elevating the paw, or the weighted pain score of Dubuisson and Dennis (1977), were superior to any single measure (r ranging from 0.75 to 0.86). Addition of flinching and favouring to the combined pain score using multiple regression did not increase variance explained. Depending on the measure used, a sedative dose of pentobarbital produced apparent analgesia, hyperalgesia or no effect. The interphase depression of pain, as well as the analgesic effects of morphine and amphetamine, were all associated with increased motor activation. Power analysis indicated that using a moderate dose of formalin and a combined pain score gave the greatest power to detect differences in pain. It was also found that pain scores increase with ambient temperature and that rat strains may differ in formalin pain sensitivity.

Vestibular tactile and pain thresholds in women with vulvar vestibulitis syndrome.

&NA; Vulvar vestibulitis syndrome (VVS) is a common cause of dyspareunia in pre‐menopausal women. Little is known about sensory function in the vulvar vestibule, despite Kinseys assertion that it is important for sexual sensation. We examined punctate tactile and pain thresholds to modified von Frey filaments in the genital region of women with VVS and age‐ and contraceptive‐matched pain‐free controls. Women with VVS had lower tactile and pain thresholds around the vulvar vestibule and on the labium minus than controls, and these results were reliable over time. Women with VVS also had lower tactile, punctate pain, and pressure‐pain tolerance over the deltoid muscle on the upper arm, suggesting that generalized systemic hypersensitivity may contribute to VVS in some women. In testing tactile thresholds, 20% of trials were blank, and there was no group difference in the false positive rate, indicating that response bias cannot account for the lower thresholds. Women with VVS reported significantly more catastrophizing thoughts related to intercourse pain, but there was no difference between groups in catastrophizing for unrelated pains. Pain intensity ratings for stimuli above the pain threshold increased in a parallel fashion with log stimulus intensity in both groups, but the ratings of distress were substantially greater in the VVS group than in controls at equivalent levels of pain intensity. The data imply that VVS may reflect a specific pathological process in the vestibular region, superimposed on systemic hypersensitivity to tactile and pain stimuli.

Morphine-6-glucuronide: Analgesic effects and receptor binding profile in rats

The antinociceptive effects of morphine-6-glucuronide (M6G) were examined in two animal models of pain, the tail immersion test (reflex withdrawal to noxious heat) and the formalin test (behavioral response to minor tissue injury). In the tail immersion test, M6G produced an increase in withdrawal latency that rose rapidly between 0.01 and 0.025 ug ICV or 1 and 2 mg/kg SC. A further increase occurred at doses greater than 0.2 ug ICV or 4 mg/kg SC and was associated with marked catalepsy and cyanosis. Naloxone, 0.1 mg/kg SC, shifted the lower component of the dose-effect relation by a factor of 24. In the formalin test, 0.01 ug M6G ICV produced hyperalgesia, while between 0.05 and 0.2 ug ICV, antinociception increased rapidly without toxicity. The dose effect relations for hyperalgesia and antinociception were shifted to the right by factors of 20- and 3-fold, respectively. By comparison, ICV morphine was 60 (formalin test) to 145-200 (tail immersion test) times less potent than M6G. At sub-nanomolar concentrations, M6G enhanced the binding of [3H]-etorphine, [3H]-dihydromorphine and [3H]-naloxone to rat brain membrane receptors by 20-40%. At higher concentrations, M6G displaced each ligand from binding sites, with Ki values of about 30 nM, as compared to morphine Ki values of about 3 nM. The data indicate that the in vivo and in vitro effects of M6G are complex and that M6G may play an important role in analgesia in experimental animals, and by implication, in man.

Morphine analgesia in the tail-flick and Formalin pain tests is mediated by different neural systems

Abstract Morphine analgesia was examined in rats with the Formalin test, which produces moderate pain that lasts about 2 h, and with the tail-flick test which measures brief, threshold-level pain. Lesions of the nucleus raphe magnus and the caudal periaqueductal gray attenuate the effects of morphine in the tail-flick test but do not influence analgesia in the Formalin test. The results indicate that different neural mechanisms underlie the analgesic effects of morphine in different types of pain.

Brainstem lesions dissociate neural mechanisms of morphine analgesia in different kinds of pain.

The effects of brainstem lesions on morphine analgesia were examined using the formalin test which produces moderate pain that lasts about 2 h, and the tail-flick test which measures brief threshold-level pain. Lesions of the nucleus raphe magnus attenuated and small lesions of the central tegmental nucleus potentiated the effects of morphine in the tail-flick test. Lesions of the median raphe nucleus potentiated the effects of morphine in the formalin test. Large lesions of the pontine reticular formation had no effect in either pain test. These results indicate that the neural mechanisms underlying morphine analgesia are different in different kinds of pain.

Reduction of post- operative morbidity following patient- controlled morphine

The present study examined the impact of two methods of pain management on recovery in 38 women undergoing hysterectomy. One group received IV morphine in the recovery room and IM morphine on the ward on a PRN basis (PRN group). In the other group, a loading dose of morphine 8 mg IV was given when the patient first complained of pain and patientcontrolled IV morphine (PCA) was initiated and continued for 48 h (PCA group). Both groups received similar amounts of morphine overall, differently distributed over time. The PCA patients received 8 mg · h− 1 in the recovery room (approximately 2.5 hrs) and less thereafter. The PRN patients received approximately 2 mg · h− 1 for the entire 48- hr period. Pain control was better throughout convalescence and less variable across time with PCA management. Minute ventilation also recovered faster and by day four was 25 per cent above the preoperative baseline in the PCA group. In addition, oral temperature became normal one day earlier, ambulation recovered more rapidly and patients were discharged from hospital earlier. The data suggest that early treatment with relatively high, selftitrated morphine doses may alter the course of the metabolic response to surgery.RésuméNous avons mesuré l’impact de deux méthodes d’analgésie sur la récupération posthystérectomie de 38 patientes. A celles du groupe PRN, on injectait de la morphine IV à la salle de réveil puis de la morphine IM au besoin. Aux autres (groupe PCA), on donnait 8 mg de morphine IV dès l’avènement des douleurs et on les laissaient ensuite elles-mêmes contrôler l’injection de la morphine IV pendant 48 hre. Les patientes des deux groupes utilisèrent des quantités semblables de morphine quoique vec des profits temporels différents soil 8 mg · h− 1 en salle de réveil (∼2,5 hre) et moins par la suite pour celles du groupe PCA contre approximativement 2 mg · h− 1 pendant 48 hre pour celles du groupe PRN. L’analgésie était plus stable et de meilleure qualité pendant la convalescence chez le groupe PCA. La ventilation/minute remontait aussi plus vite dans ce groupe, dépassant même les valeurs pré- opératoires de 25 pour cent au quatrième jour post- opératoire. De plus la température orale s’y normalisait un jour plus tôt, la mobilisation y était plus rapide et le congé y survenait plus précocement. Il semble done que l’auto- injection précoce de doses relativement importantes de morphine puisse modifier la réponse métabolique au stress chirurgical.

Pain on a surgical ward: a survey of the duration and intensity of pain and the effectiveness of medication

&NA; The effectiveness of analgesic medication for post‐surgical pain was surveyed in a surgical ward of a large general hospital. Since earlier studies have shown that pain generally decreases rapidly and is negligible by the fourth day after surgery, the patients in the survey were assigned to 2 groups: (1) those given analgesics during the first 4 days after surgery, and (2) those given analgesics for pain after the fourth day. The results show that the patients with pain that persists beyond day 4 comprise a substantial proportion of the patients in a surgery ward (31%), are older, tend to use more words to describe their pain, and are helped less by their prescribed analgesic medications. This group is prescribed lower doses of analgesics and receives them more frequently; however, this prescription strategy appears to be ineffective since 26% of these patients report increased pain after medication compared to only 2% in the group that received analgesics during the first 4 days.

The behavioral response to formalin in preweanling rats

&NA; The behavioural response of rat pups, 1–20 days of age, to subcutaneous injection of formalin in a rear paw is described. Formalin‐injected pups were compared to handled controls and to pups that received an injection of normal saline. Ongoing behaviour was recorded every 2 min for 60 min after injection. Injection of normal saline produced little disorganization of behaviour, although day‐1, ‐3 and ‐6 pups did frequently flex the limb on the injected side early in the session. Injection of 10 &mgr;l of 1% formalin depressed active and quiet sleep in pups 10 days old and younger. Much less disruption of sleep was observed in day‐15 pups, and in day‐20 pups it was necessary to increase the concentration of formalin to 2.5% to produce a consistent behavioural response. The specific responses of pups to formalin injection were flexion of the limb, shaking the limb, and licking the injected paw. Pups of all ages displayed all of these responses, but in pups younger than 10 days, only limb flexion was consistent. Shaking became a consistent response in day‐10 pups and licking in day‐15 and ‐20 pups. Non‐specific behaviours (squirming, vigorous rear kicks with both hind limbs and convulsive whole body jerks) were markedly increased by formalin in younger pups with a developmental pattern: squirming and kicking in day‐1 pups, kicking and jerking in day‐3 to ‐15 pups. Non‐specific behaviours decreased and specific behaviours increased with age. In addition, the overall intensity and duration of the response decreased with age. The biphasic time course of the response of adult rats to formalin injection did not appear until 15 days of age.

The formalin test: a dose–response analysis at three developmental stages

&NA; We investigated the behavioral response of rat pups to intraplantar injection of varying formalin concentrations using a time‐sampling method. At 3 days of age, the response was monophasic and persisted for the whole hour, even at low formalin concentrations. Flexion, shaking and licking the injected limb and hind‐limb kicking correlated strongly with log formalin concentration (r=0.82); behavioral state was altered only at the highest concentration. The response on day 15 was also monophasic, but it waned in 30 min, even at the highest formalin concentration tested. Flexion, shaking and licking of the injected limb were strong pain measures (r=0.83). The response at 25 days was biphasic, and the adult measures, paw lifting and licking, produced a good formalin concentration–effect relationship (r=0.80). The log concentration–effect relationships for formalin at the three developmental stages and for adult rats were parallel, but between 3 days and 15 days of age, the relationship shifted to the right by 2.5‐fold, and by a further 4‐fold between 15 and 25 days, when the sensitivity to formalin‐induced pain was similar to that in adults. The data describe efficient, quantitative measures of formalin‐induced pain for developing rats, show that the pain response is log‐linearly related to formalin concentration throughout development, and demonstrate that the sensitivity to formalin‐induced pain is about 10‐fold higher in neonatal rats than in weanlings. the data imply that there are major qualitative changes in pain processing as the nervous system develops.

Peripheral opioid modulation of pain and inflammation in the formalin test

The effects of local treatment with opioid receptor agonists on the early (0-10 min) and late (20-40 min) behavioural response and extravasation induced by intraplantar injection of 1% formalin in rats were examined. The mu-opioid receptor agonist [D-Ala2,N-Me-Phe4,Gly5-ol]enkephalin (DAMGO) depressed pain behaviour in the late phase, and extravasation in both phases. The kappa-opioid receptor agonist trans-(+/-)-3,4-dichloro-N-methyl-N-(2-[1-pyrrolidinyl] benzeneacetamide methanesulfonate (U50,488H) suppressed the behavioural response in both phases, but extravasation was enhanced in the early phase and not altered in the late phase. The delta-opioid receptor agonist [D-Pen2,5]enkephalin (DPDPE) enhanced the behavioural response in the late phase, but inhibited extravasation in the both early and late phases. Systemic injection of the agonists had no effects, and pretreatment with s.c. naloxone methiodide reversed the effects of locally administered agonists. These data (1) support the notion that different pathophysiological mechanisms underlie the two phases of the formalin test, and (2) indicate that depending on the receptor specificity, opioid receptor agonists have both pro- and antinociceptive effects, as well as pro- and antiinflammatory activity.