Involvement of the Serotonergic System in Analgesia Induced by the Influence of Low-Intensity Microwaves on an Antipain Acupuncture Point

In experiments on mice, we studied changes in the level of analgesia induced by irradiation of the antipain acupuncture point (AP) E36 by low-intensity microwaves under conditions of modification of the serotonin level in the brain; this level was modified by injection of 300 mg/kg DL-p-chlorophenylalanine (pCPA). The duration of the nociceptive behavioral reaction (licking the limb) caused by injection of the formalin solution into the foot dorsal surface increased 24, 48, and 72 h after pCPA injection by 99.9, 84.4, and 114.4%, as compared with those in animals subjected to microwave irradiation of the AP E36 with no preliminary pCPA injection. It is concluded that the brain serotonergic system is actively involved in the analgesia effects induced by irradiation of the AP by low-intensity microwaves.     

Microwave-induced and pharmacological suppression of somatic pain under conditions of the formalin test in mice

We examined pain-related behavioral reactions and non-pain behavioral manifestations in mice under conditions of the formalin test. Levels of analgesia induced by i.p. injections of analgin, microwave irradiation of an antinociceptive acupuncture point (AP), E-36, or combined application of the above factors were measured. The duration of the pain behavioral reaction (licking of the injured limb) decreased due to irradiation of the AP with microwaves and to injection of 8.3 mg/kg analgin by 24.3% and 53.8%, on average, respectively. Combination of injection of analgin in a smaller dose (4.2 mg/kg) and microwave irradiation of the AP suppressed manifestations of the pain behavioral reaction by 43.4%. Thus, combination of pharmacologically induced analgesia with the action of microwaves on the antinociceptive AP allows one to significantly decrease the doses of analgesic preparations necessary to provide a full-level analgesic effect; in such a way, side effects of the respective drugs can be weakened. Neirofiziologiya/Neurophysiology, Vol. 38, No. 1, pp. 46–51, January–February, 2006.     

Analgesia induced by combined action of Tramadol and microwave irradiation in somatic pain in mice

In experiments on albino male mice weighing 24 to 32 g, we studied the suppression of the pain reaction induced by injection of formalin into the dorsal surface of the foot, using a combination of injection of tramadol hydrochloride (Tramal) in a dose of 0.42 mg/kg and irradiation of the acupuncture point (AP) E-36 with low-intensity microwaves (frequency 30 to 300 GHz and power rate density 3·10⁻⁹ W/cm²). The combined use of tramadol injection and the action of microwaves on the AP led to a decrease in the duration of the pain reaction (licking of the pain nidus), on average, by 58.2%, as compared with that in the control animal group. The analgesic effect was more strong in the case of such a combination than after isolated injection of tramadol in doses of 0.83 and 0.42 mg/kg (by 14.4 and by 10.0%, respectively) and isolated microwave irradiation of the AP TA E-36 (by 48.7%). The data obtained demonstrate the possibility of effective analgesia using a combination of microwave irradiation of the AP and the pharmacological influence with a significant decrease in the dose of the analgesic. Neirofiziologiya/Neurophysiology, Vol. 38, No. 4, pp. 314–319, July–August, 2006.     

Involvement of the serotonergic cerebral system in microwave-induced analgesia in mice of different genotypes

Under conditions of the formalin test, we studied changes in the level of analgesia induced by the action of low-intensity microwaves on the antinociceptive acupuncture point (AP) E36 in mice of strains CBA/CaLac (CBA) and C57BL/6j (C57) and in albino mongrel mice. Measurements were performed under control conditions and with experimentally induced decrease in the serotonin level in the brain (by injections of DL-parachlorophenylalanine, p-CPA). In the latter cases, the duration of the pain behavioral reaction increased despite irradiation of the AP E36. In mongrel, CBA, and C57 mice, the intensity of pain manifestations was 114.4, 29.0, and 21.1% greater, respectively, than in mice of these groups with no injections of p-CPA. These facts show that the serotonergic brain system is profoundly involved in the formation of analgesia after irradiation of the AP by low-intensity microwaves, and this involvement significantly depends on the genotype of the animals. Neirofiziologiya/Neurophysiology, Vol. 38, Nos. 5/6, pp. 495–497, September–December, 2006.     

Combination of pharmacological analgesics and microwave irradiation of an acupuncture point for suppression of visceral pain in mice: Role of the opioid and serotonergic cerebral systems

We studied suppression of pain-related reactions induced in mice by i.p. injection of 0.08 ml of a 2% solution of acetic acid using pharmacological analgesics (analgin and tramadol) combined with low-intensity microwave irradiation of an acupuncture point (AP) E-36 (frequency 30 to 300 GHz and power rate density 3·10⁻⁹ W/cm²). The respective effects were also observed under conditions of suppression of the functions of opioid and serotonergic cerebral systems using injections of, respectively, naloxone and DL-p-chlorophenylalanine (p-CPA). We found that antinociceptive effects provided by analgesics used in a 50% mean single dose in the combination with microwave irradiation of the AP were significantly more intense than those induced by isolated injection of analgesics used in both 50% and full mean single doses and isolated microwave irradiation of the AP E-36. After injections of naloxone, analgesic effects caused by the combined action of analgin and microwave irradiation of the AP were considerably smaller. At the same time, after injection of p-CPA, analgesic effects, provided by the combination of injection of pharmacological agents and microwave irradiation of the AP, weakened in the case of use of both analgesics. This was manifested in a significant increase in the total duration of pain-related behavioral reactions. Therefore, the studied analgesic effects observed in the examined animal groups are realized due to the involvement of the opioid and serotonergic cerebral systems. Neirofiziologiya/Neurophysiology, Vol. 39, No. 6, pp. 468–477, November–December, 2007.     

Corticosterone does not change open elevated plus maze-induced antinociception in mice

It has been demonstrated that the exposure of rodents to the standard elevated plus-maze (sEPM: 2 open and 2 enclosed arms) elicits defensive behavioral reactions and antinociception and also activates the hypothalamo-pituitary-adrenal (HPA) axis. We have recently reported that EPM-induced antinociception is particularly observed when rats and mice are exposed to a totally open EPM (oEPM: 4 open arms). Given that the oEPM seems to be a more aversive situation than the sEPM, we hypothesized that oEPM exposure would induce higher plasma levels of corticosterone than sEPM exposure in mice. In this study, we investigated the influence of exposure to eEPM (enclosed EPM: 4 enclosed arms), sEPM or oEPM on plasma corticosterone levels in mice, with or without prior nociceptive stimulation (2.5% formalin injection into the right hind paw). We also tested whether the nociceptive response in the formalin test and oEPM-induced antinociception are altered by adrenalectomy. Results showed that oEPM-exposed mice spent less time licking the injected paw than sEPM- and eEPM-exposed animals. All three types of EPM exposure increased plasma corticosterone when compared to the basal group, but sEPM- and oEPM-exposed mice showed higher corticosterone levels than eEPM-exposed mice. Prior nociceptive stimulation (formalin injection) did not enhance the plasma corticosterone response induced by the three types of EPM exposure. Indeed, formalin injection appeared to provoke a ceiling effect on plasma corticosterone concentration. Furthermore, neither the nociceptive response in the formalin test nor oEPM-induced antinociception was changed by adrenalectomy. Present results suggest that oEPM antinociception does not depend on corticosterone release in mice.     

Inhibition of <Emphasis Type="SmallCaps">d</Emphasis>-Amino-Acid Oxidase Activity Induces Pain Relief in Mice

(1). We investigated the effects of inhibiting d-amino-acid oxidase (DAO) activity on nociceptive responses through the use of mutant ddY/DAO⁻ mice, which lack DAO activity, and through the application of a selective inhibitor of DAO, sodium benzoate, in the tail flick test, hot-plate test, formalin test, and acetic acid-induced writhing test. (2). Compared with normal ddY/DAO⁺ mice, ddY/DAO⁻ mice showed significantly prolonged tail withdrawal latency in the tail flick test and licking/jumping latency in the hot-plate test, as well as significantly reduced duration of licking/biting in the late phase of the formalin test and the number of abdominal writhing in the acetic acid-induced writhing test. (3). In addition, we investigated the effects of sodium benzoate in Kunming mice having normal DAO activity. (4). Intravenous administration of sodium benzoate (400 mg/kg) significantly inhibited pain responses of the late phase of the formalin test and abdominal writhing responses in the acetic acid-induced writhing test, with no effects on the early phase flinch responses in the formalin test, nociceptive responses in the tail flick test, or hot-plate test. (5). These results suggest that DAO acts as a pro-nociceptive factor in pain, particularly chronic pain, transmission and modulation, and may be a target for pain treatment.     

Long-term ovariectomy changes formalin-induced licking in female rats: the role of estrogens

Gonadal hormones have been shown to exert modulatory effects on nociception and analgesia. To investigate the role of gonadal hormones in the response by female rats to both phasic and persistent nociceptive stimulation, we evaluated the effects of long-term ovariectomy (OVX, 6 months) on the thermal pain threshold and on formalin-induced responses. The thermal pain threshold was evaluated with the plantar test apparatus, while persistent pain was induced by a subcutaneous injection of dilute formalin (50 microliter, 10%) in the dorsal hind paw. The formalin test was carried out in an open field apparatus where the animal's spontaneous behavior and formalin-induced responses (licking duration, flinching frequency and flexing duration of the injected paw) were recorded for 60 min. Estradiol and corticosterone plasma levels were determined in blood collected from the anesthetized animals at the end of the test. In OVX females, the duration of formalin-induced licking was longer than in Intact females during both the first and the second phase; flinching and flexing did not differ from Intact. The thermal pain threshold was only slightly affected by OVX. Estradiol and corticosterone were lower in OVX females than Intact ones. These data indicate that long-term depletion of gonadal hormones in female rats modulates the pain-induced behavioral responses related to supraspinal neural circuits (licking of the injected paw) rather than more spinally mediated responses such as formalin-induced flinching and withdrawal latency in the plantar test.     

Effects of ethanol on deep pain evoked by formalin injected in TMJ of rat

It has been reported that ethanol can alter nociceptive sensitivity from superficial tissues, such as skin and subcutaneous region. However, the influence of ethanol on deep pain conditions is not understood. The aim of this study was to demonstrate the acute, chronic and ethanol withdrawal effects on nociceptive behavioral responses induced by the injection of formalin into the temporomandibular joint (TMJ) region of rats. In experiment 1, rats were injected with ethanol (2,5 g/Kg, i.p.) or an equal volume of saline 15 min before the administration of formalin (1.5%) into the TMJ. Rats pretreated with ethanol showed a decrease in nociceptive behavioral responses. In experiment 2, rats were given an ethanol solution (6.5%) or tap water to drink for 4 and 10 days. On day 4, the animals (ethanol group) showed amounts of analgesia when submitted to the TMJ formalin test. Tolerance to the antinociceptive effects was observed on day 10. Behavioral hyperalgesia was verified 12 hr after withdrawal in another group that drank ethanol for 10 days. These results show that ethanol can affect the nociceptive responses related to deep pain evoked by the TMJ formalin test.     

Behavioral responses to phasic nociceptive stimulation occurring on the background of a tonic pain focus in ontogeny of the rabbit

The effect of a focus of tonic pain (a subcutaneous injection of formalin into the dorsal field of the shin) on the thresholds of a defense reaction, an attempt to jump out of the chamber in response to a nociceptive electrocutaneous stimulation of the hindpaw1, was studied in the 20– 25-day-old and adult rabbits. The tonic pain produces a biphasic reduction of the defense reaction threshold. At the first phase, hyperalgesia is more pronounced than in the second one, but its duration is shorter. Changes in pain sensitivity in the rabbits proceed in the same direction in both age groups and coincide in time with increase of specific behavioral responses to the formalin injection (licking and shaking the paw). In the 20–25-day-old rabbits the reduction of the threshold of the defense reaction and duration of hyperalgesia phases are more pronounced than in adult animals. Deceased.     

Effects of nociceptive stimuli on the pulmonary circulation in the ovine fetus

The fetus is able to exhibit a stress response to painful events, and stress hormones have been shown to modulate pulmonary vascular tone. At birth, the increased level of stress hormones plays a significant role in the adaptation to postnatal life. We therefore hypothesized that pain may alter pulmonary circulation in the perinatal period. The hemodynamic response to subcutaneous injection of formalin, which is used in experimental studies as nociceptive stimulus, was evaluated in chronically prepared, fetal lambs. Fetal lambs were operated on at 128 days gestation. Catheters were placed into the ascending aorta, superior vena cava, and main pulmonary artery. An ultrasonic flow transducer was placed around the left pulmonary artery. Three subcutaneous catheters were placed in the lambs' limb. The hemodynamic responses to subcutaneous injection of formalin, to formalin after fetal analgesia by sufentanil, and to sufentanil alone were recorded. Cortisol and catecholamine concentrations were also measured. Pulmonary vascular resistances (PVR) increased by 42% (P < 0.0001) after formalin injection. Cortisol increased by 54% (P = 0.05). During sufentanil infusion, PVR did not change significantly after formalin. Cortisol increased by 56% (P < 0.05). PVR did not change during sufentanil infusion. Norepinephrine levels did not change during any of the protocols. Our results indicate that nociceptive stimuli may increase the pulmonary vascular tone. This response is not mediated by an increase in circulating catecholamine levels. Analgesia prevents this effect. We speculate that this pulmonary vascular response to nociceptive stimulation may explain some hypoxemic events observed in newborn infants during painful intensive care procedures.     

Induction of nociceptive responses by intrathecal injection of interleukin-1 in mice.

Intrathecal (i.t.) injection (between lumbar vertebrae 5 and 6) into mice of a markedly low dose of IL-1alpha (3x10(-4) fmol or 5.4 fg in 5 microl per mouse) induced behaviors involving scratching, biting, and licking of non-stimulated hindpaws. The IL-1-induced behaviors appeared within 10 min of the injection of IL-1alpha, peaked at 20-40 min, and had disappeared 60 min after the injection. The IL-1-induced behaviors were similar to the nociceptive responses induced in mice by i.t. injection of substance P (SP) or subcutaneous (s.c.) injection of formalin into the footpad. The IL-1-induced behaviors were suppressed by intraperitoneal morphine, indicating that they are nociceptive responses. The nociceptive responses induced by 3x10(-4) (5.4 fg) of IL-1alpha were almost completely suppressed by co-injection of 0.3 fmol (7.2 pg) of an IL-1 receptor antagonist (IL-1ra). An antiserum against substance P, but not an antiserum against somatostatin, suppressed the IL-1-induced nociceptive responses. The nociceptive responses induced by s.c. injection of 2% formalin into the footpad were also inhibited by i.t. injection of 30 pmol (720 ng) of IL-1ra. These results suggest that IL-1 may play a role in hyperalgesia in mice by acting as a factor augmenting pain transmission in the spinal cord at least in part by either directly or indirectly releasing substance P.     

Antinociceptive effect of spinally injected L-NAME on the acute nociceptive response induced by low concentrations of formalin

The formalin test has been proposed as an animal model of pain produced by tissue injury. Although biphasic nociceptive responses to formalin injection have been well documented, low concentrations (0.125 and 0.5%) of formalin injected into the mouse hindpaw produced only the phasic (acute) paw-licking response, lasting the first 5 min after the formalin injection. To explore the involvement of nitric oxide (NO) in the spinal cord and peripheral system during the acute phase of the formalin test, we examined the effect of intrathecal (i.t.) or intraplantar (i.pl.) injection of L-N(G)-nitro arginine methyl ester (L-NAME), a NO synthase inhibitor in mice. Pretreatment with L-NAME (160 nmol), injected i.t., resulted in a significant inhibition of the paw-licking response induced by 0.125 and 0.5% of formalin. L-Arginine (600 mg/kg, i.p.) but not D-arginine (600 mg/kg, i.p.) reversed the antinociceptive effect of L-NAME on the acute nociceptive response induced by low concentrations of formalin. The i.pl. injection of L-NAME (160 nmol) produced a significant decrease of the late (tonic) phase response evoked by 2.0% formalin without affecting the early (acute) phase response. Similar results have been reported in the case of i.t. injected L-NAME as assayed by the 2.0% formalin test. L-NAME (160 nmol), injected into the plantar paw, gave no significant effect on the acute nociceptive response induced by a low concentration of formalin (0.125%). These results suggest that NO in the spinal cord may be involved in not only the late phase response of the formalin (2.0%)-induced paw-licking, but also at least the acute phase response induced by low concentrations (0.125 and 0.5%) of formalin, while peripheral NO has little effect on the early (acute) phase nociceptive response evoked by formalin (0.125--2.0%) injection.     

Altered nociceptive behavior and emotional contagion of pain in mouse models of autism

Individuals with autism spectrum disorder (ASD) have altered sensory processing but may ineffectively communicate their experiences. Here, we used a battery of nociceptive behavioral tests to assess sensory alterations in two commonly used mouse models of ASD, BTBR T⁺Itpr3^tf/J (BTBR), and fragile-X mental retardation-1 knockout (Fmr1-KO) mice. We also asked whether emotional contagion, a primitive form of empathy, was altered in BTBR and Fmr1 KO mice when experiencing pain with a social partner. BTBR mice demonstrated mixed nociceptive responses with hyporesponsivity to mechanical/thermal stimuli and intraplantar injections of formalin and capsaicin while displaying hypersensitivity on the acetic acid test. Fmr1-KO mice were hyposensitive to mechanical stimuli and intraplantar injections of capsaicin and formalin. BTBR and Fmr1-KO mice developed significantly less mechanical allodynia following intraplantar injections of complete Freund's adjuvant, while BTBR mice developed slightly more thermal hyperalgesia. Finally, as measured by the formalin and acetic acid writhing tests, BTBR and Fmr1-KO mice did not show emotional contagion of pain. In sum, our findings indicate that depending on the sensation, pain responses may be mixed, which reflects findings in ASD individuals.     

Peripheral effect of a kappa opioid receptor antagonist on nociception evoked by formalin injected in TMJ of pregnant rats

The effect of sex hormones on orofacial pain modulation is poorly understood. Therefore, this study aimed to investigate the effect of hormonal changes as a result of pregnancy, as well as that of the kappa (kappa) opioid receptor antagonist on female rats' sensitivity to the temporomandibular joint (TMJ) formalin test. Initially, female rats at estrus and pregnant females on day 19 of pregnancy received a 50 microl formalin (1.5%) injection in the right TMJ. The pregnant females showed a reduction in nociceptive responses to the TMJ formalin test when compared with those at estrus. Then, the selective kappa-opioid receptor antagonist nor-Binaltorphimine (nor-BNI), was co-administered with the formalin. Next, additional groups received the kappa (200 microg) receptor antagonist or 0.9% NaCl 24 hours prior to the periarticular injection of formalin. Co-administration of nor-BNI with formalin into the TMJ region had no significant effect. The pre-injection of selective kappa-opioid receptor antagonist, nor-BNI, significantly enhanced the nociceptive behavioral responses in pregnant females. When applied in the contralateral TMJ, nor-BNI did not affect the magnitude of the nociceptive response induced by formalin. It can be concluded that: 1) The increase of the sex hormone levels, as result of pregnancy, induces a reduction of nociceptive behavioral responses to the TMJ formalin test; 2) the peripheral kappa opioid receptor activation, by endogenous opioid agonists release, is involved in the antinociception to TMJ formalin test, induced by pregnancy.     

Effects of Stimulation of the Periaqueductal Gray and <Emphasis Type="Italic">Locus Coeruleus</Emphasis> on Postsynaptic Reactions of Cat Somatosensory Cortex Neurons Activated by Nociceptors

In cats, we studied the influences of stimulation of the periaqueductal gray (PAG) and locus coeruleus (LC) on postsynaptic processes evoked in neurons of the somatosensory cortex by stimulation of nociceptive (intensive stimulation of the tooth pulp) and non-nociceptive (moderate stimulations of the infraorbital nerve and ventroposteromedial nucleus of the thalamus) afferent inputs. Twelve cells activated exclusively by nociceptors and 16 cells activated by both nociceptive and non-nociceptive influences (hereafter, nociceptive and convergent neurons, respectively) were recorded intracellularly. In neurons of both groups, responses to nociceptive stimulation (of sufficient intensity) looked like an EPSP-spike-IPSP (the latter, of significant duration, up to 200 msec) complex. Electrical stimulation of the PAG (which could itself evoke activation of the cortical neurons under study) resulted in long-term suppression of synaptic responses evoked by excitation of nociceptors (inhibition reached its maximum at a test interval of 600 to 800 msec). We observed a certain parallelism between conditioning influences of PAG activation and effects of systemic injections of morphine. Isolated stimulation of LC by a short high-frequency train of stimuli evoked primary excitatory responses (complex EPSPs) in a part of the examined cortical neurons, while in other cells high-amplitude and long-lasting IPSP (up to 120 msec) were observed. Independently of the type of the primary response to PAG stimulation, the latter resulted in long-term (several seconds) suppression of the responses evoked in cortical cells by stimulation of the nociceptive inputs. The mechanisms of modulatory influences coming from opioidergic and noradrenergic brain systems to somatosensory cortex neurons activated due to excitation of high-threshold (nociceptive) afferent inputs are discussed.Neirofiziologiya/Neurophysiology, Vol. 37, No. 1, pp. 61–73, January–February, 2005.     

Antinociceptive effect induced by intraperitoneal administration of vitamin K2 (menatetrenone) in ICR mice

The antinociceptive effect of vitamin K2 (menatetrenone) in mice was examined using tail-flick and formalin test. Menatetrenone at doses of 10, 50 and 100 mg/kg, i.p. produced a dose-dependent and significant inhibition of the tail-flick response in mice. Menatetrenone (50 and 100 mg/kg, i.p.) had no significant effect on the duration of the first phase of the formalin-induced flinching. However, menatetrenone (100 mg/kg, i.p.) significantly inhibited the second phase of the formalin-induced flinching. I.p. administration of menatetrenone (100 mg/kg) significantly reduced the duration of nociceptive responses induced by i.t. injection of bradykinin, but not of substance P, prostaglandin E2 or N-methyl-D-aspartate (NMDA). These present data suggest that i.p. pretreatment with menatetrenone produced dose-dependent antinociceptive effect in mice. This effect may be, at least in part, mediated by the inhibition of bradykinin dependent nociceptive transmission in the spinal cord.     

Interaction of Tonic and Phasic Pain in Rabbit Ontogenesis

In the experiments on the 20–25-day-old and adult rabbits, effects of tonic pain focus (a subcutaneous injection of formalin into leg dorsal surface) on behavioral and electrophysiological characteristics of acute pain were studied. The effect of the 40–60-min-long tonic pain was seen as a decrease of defensive reaction threshold and an increase of inhibitory effect of brain rewarding zones on evoked potential recorded in thalamus parafascicular complex in response to a nociceptive electrocutaneous stimulation in narcotized rabbits. The changes observed were biphasic and coincided in time with an enhancement of the earlier described [26] specific behavioral responses to formalin injection. It is established that the effect of tonic pain is more expressed by its intensity and duration in the 20–25-day-old than in adult rabbits.