Cannabidiol-induced panicolytic-like effects and fear-induced antinociception impairment: the role of the CB1 receptor in the ventromedial hypothalamus.

RATIONALE: The behavioural effects elicited by chemical constituents of Cannabis sativa, such as cannabidiol (CBD), on the ventromedial hypothalamus (VMH) are not well understood. There is evidence that VMH neurons play a relevant role in the modulation of unconditioned fear-related defensive behavioural reactions displayed by laboratory animals. OBJECTIVES: This study was designed to explore the specific pattern of distribution of the CB1 receptors in the VMH and to investigate the role played by this cannabinoid receptor in the effect of CBD on the control of defensive behaviours and unconditioned fear-induced antinociception. METHODS: A panic attack-like state was triggered in Wistar rats by intra-VMH microinjections of N-methyl-D-aspartate (NMDA). One of three different doses of CBD was microinjected into the VMH prior to local administration of NMDA. In addition, the most effective dose of CBD was used after pre-treatment with the CB1 receptor selective antagonist AM251, followed by NMDA microinjections in the VMH. RESULTS: The morphological procedures demonstrated distribution of labelled CB1 receptors on neuronal perikarya situated in dorsomedial, central and ventrolateral divisions of the VMH. The neuropharmacological approaches showed that both panic attack-like behaviours and unconditioned fear-induced antinociception decreased after intra-hypothalamic microinjections of CBD at the highest dose (100 nmol). These effects, however, were blocked by the administration of the CB1 receptor antagonist AM251 (100 pmol) in the VMH. CONCLUSION: These findings suggest that CBD causes panicolytic-like effects and reduces unconditioned fear-induced antinociception when administered in the VMH, and these effects are mediated by the CB1 receptor-endocannabinoid signalling mechanism in VMH.

Role of cholinergic, opioidergic and GABAergic neurotransmission of the dorsal hippocampus in the modulation of nociception in guinea pigs.

AIMS: Several physiological, pharmacological and behavioral lines of evidence suggest that the hippocampal formation is involved in nociception. The hippocampus is also believed to play an important role in the affective and motivational components of pain perception. Thus, our aim was to investigate the participation of cholinergic, opioidergic and GABAergic systems of the dorsal hippocampus (DH) in the modulation of nociception in guinea pigs. MAIN METHODS: The test used consisted of the application of a peripheral noxious stimulus (electric shock) that provokes the emission of a vocalization response by the animal. KEY FINDINGS: Our results showed that, in guinea pigs, microinjection of carbachol, morphine and bicuculline into the DH promoted antinociception, while muscimol promoted pronociception. These results were verified by a decrease and an increase, respectively, in the vocalization index in the vocalization test. This antinociceptive effect of carbachol (2.7 nmol) was blocked by previous administration of atropine (0.7 nmol) or naloxone (1.3 nmol) into the same site. In addition, the decrease in the vocalization index induced by the microinjection of morphine (2.2 nmol) into the DH was prevented by pretreatment with naloxone (1.3 nmol) or muscimol (0.5 nmol). At doses of 1.0 nmol, muscimol microinjection caused pronociception, while bicuculline promoted antinociception. SIGNIFICANCE: These results indicate the involvement of the cholinergic, opioidergic and GABAergic systems of the DH in the modulation of antinociception in guinea pigs. In addition, the present study suggests that cholinergic transmission may activate the release of endorphins/enkephalin from interneurons of the DH, which would inhibit GABAergic neurons, resulting in antinociception.

Modulation of tonic immobility in guinea pig PAG by homocysteic acid, a glutamate agonist.

Tonic immobility (TI) is an innate defensive behavior elicited by physical restriction and postural inversion, and is characterized by a profound and temporary state of motor inhibition. The participation of the periaqueductal gray matter (PAG) in TI modulation has previously been described. In addition, the excitatory amino acids (EAA) are important mediators involved in the adjustment of several defensive responses produced by PAG. In the present study, we investigated the effect of microinjection of the EAA agonist dl-homocysteic acid (DLH) and the N-methyl-d-aspartate (NMDA) receptor antagonist (MK-801) into the ventrolateral and dorsal PAG over the duration of TI in guinea pigs. Microinjection of 15 nmol/0.2 microl of DLH into the ventrolateral PAG (vlPAG) and 30 nmol/0.2 microl of DLH into the dorsal PAG (dPAG) promoted an increase and decrease in TI duration, respectively. These responses were blocked by prior microinjection of the NMDA receptor antagonist, MK-801 (3.6 nmol/0.2 microl) at the same site. Microinjection of MK-801 alone into the vlPAG and dPAG did not alter the duration of TI episodes. These results suggest that NMDA receptors are involved in the modulation of TI in both the vlPAG and dPAG. In addition, PAG excitatory amino acids modulate the TI response via columnar organization of the PAG. In this manner, the vlPAG facilitates TI modulation whereas dPAG has an inhibitory role in TI.

Role of opioidergic and GABAergic neurotransmission of the nucleus raphe magnus in the modulation of tonic immobility in guinea pigs.

Tonic immobility (TI) is an inborn defensive behavior characterized by a temporary state of profound and reversible motor inhibition elicited by some forms of physical restraint. Previous results from our laboratory have demonstrated that nucleus raphe magnus (NRM) is also a structure involved in the modulation of TI behavior, as chemical stimulation through carbachol decreases the duration of TI in guinea pigs. In view of the fact that GABAergic and opioidergic circuits participate in the regulation of neuronal activity in the NRM and since these neurotransmitters are also involved in the modulation of TI, the objective of the present study was to evaluate the role of these circuits of the NRM in the modulation of the behavioral TI response. Microinjection of morphine (4.4 nmol/0.2 microl) or bicuculline (0.4 nmol/0.2 microl) into the NRM increased the duration of TI episodes while muscimol (0.5 nmol/0.2 microl) decreased it. The effect of morphine injection into the NRM was blocked by previous microinjection of naloxone (2.7 nmol/0.2 microl). Muscimol at 0.25 nmol did not produce any change in TI duration; however, it blocked the increased response induced by morphine. Our results indicate a facilitatory role of opioidergic neurotransmission in the modulation of the TI response within the NRM, whereas GABAergic activity plays an inhibitory role. In addition, in the present study the modulation of TI in the NRM possibly occurred via an interaction between opioidergic and GABAergic systems, where the opioidergic effect might be due to inhibition of tonically active GABAergic interneurons.

Cholinergic/opioid interaction in anterior cingulate cortex reduces the nociceptive response of vocalization in guinea pigs.

The anterior cingulate cortex (ACC) is crucial in the modulation of the sensory, affective and cognitive aspects of nociceptive processing. Also, it participates in the planning and execution of behavioral responses evoked by nociceptive stimuli via descending projections to the brainstem. In laboratory animals nociceptive experimental tests evaluate behavioral responses that preferentially express the sensory-discriminative or affective-motivational component of pain. The objective of this study was to investigate the participation of opioid and cholinergic neurotransmission in the ACC on different nociceptive responses in guinea pigs. We used nociceptive tests of formalin and vocalization evoked by peripheral noxious stimuli (electric shock) to evaluate the behavioral expression of the sensory-discriminative and affective motivational components, respectively. We verified that the microinjection of morphine (4.4nmol) in the ACC of guinea pigs promotes antinociception in the two experimental tests investigated. This effect is blocked by prior microinjection of naloxone (2.7nmol). On the other hand, the microinjection of carbachol (2.7nmol) in the ACC induces antinociception only in the vocalization test. This effect was prevented by prior microinjection of atropine (0.7nmol) and naloxone (2.7nmol). In fact, the blockade of µ-opioids receptors with naloxone in ACC prevented the antinociceptive effect of carbachol in the vocalization test. Accordingly, we suggest that the antinociception promoted by carbachol was mediated by the activation of muscarinic receptors on local ACC opioid interneurons. The release of endogenous opioids seems to inhibited the expression of the behavioral response of vocalization. Therefore, we verified that the antinociceptive effect of morphine microinjection in ACC is broader and more robust than that promoted by carbachol.

Cannabinoid system of dorsomedial telencephalon modulates behavioral responses to noxious stimulation in the fish Leporinus macrocephalus.

Fish dorsomedial telencephalon has been considered a pallial region homologous to mammals amygdala, being considered a possible substrate for nociception modulation in this animal group. The present study aimed to evaluate the participation of the cannabinoid system of Dm telencephalon on nociception modulation in the fish Leporinus macrocephalus. We demonstrated that cannabidiol microinjection in Dm telecephalon inhibits the behavioral nociceptive response to the subcutaneous injection of 3% formaldehyde, and this antinociception is blocked by previous treatment with AM251 microinjection. Furthermore, AM251 microinjection in Dm prior to restraint stress also blockades the stress-induced antinociception. These results reinforce the hypothesis that this pallial telencephalic structure has a pivotal role in nociception modulation in fish.

Serotoninergic activation of the basolateral amygdala and modulation of tonic immobility in guinea pig.

Tonic immobility (TI), also known as death feigning or animal hypnosis, is a reversible state of motor inhibition that is not only triggered by postural inversion and/or movement restraining maneuvers but also by repetitive stimulation and pressure on body parts. Evidence has demonstrated that the basolateral nucleus of the amygdala (BLA) is particularly associated with defensive behavior that involves the emotional states of fear and anxiety. In addition, some reports have demonstrated that serotonin (5-HT) released in the amygdala is increased during states of stress and anxiety, principally in the BLA. In the present study, we investigated the effects of serotonergic activation of the BLA on the duration of TI. The results showed that the microinjection of 5-HT (3.0 microg) into the BLA decreased the duration of TI. Similarly, the administration of a 5-HT1A agonist (0.1 microg of 8-hydroxy-dipropylaminotretalin) or 5-HT2 agonist (0.1 microg of alpha-methyl-5-HT) into the BLA reduced the TI duration. The effect of 5-HT2 agonist was reversed by pretreatment with a dose that had no effect per se (0.01 microg) of ketanserin (5-HT2 receptor antagonists) into the BLA. Moreover, the activation of 5-HT1A and 5-HT2 receptors in the BLA did not alter the spontaneous motor activity in the open field test. The results of the present study indicate that the serotonergic system of the BLA possibly produces a reduction in fear and/or anxiety that reduces the TI duration in guinea pigs, but this is not due to increased spontaneous motor activity induced by serotonergic activation, which might affect TI duration non-specifically.

Cholinergic modulation of tonic immobility and nociception in the NRM of guinea pig.

Tonic immobility (TI) is an inborn defensive behavior characterized by a temporary state of profound and reversible motor inhibition elicited by some forms of physical restraint. It is known that endogenous antinociceptive systems are activated during the emission of defensive behaviors including TI. The nucleus raphe magnus (NRM) is related to the modulation of nociceptive and behavioral responses. In the present study, we investigated the role of the cholinergic system of the NRM in the modulation of TI and nociception in guinea pigs. Microinjection of the cholinergic agonist carbachol (0.5 microg/0.2 microl) into the NRM promoted a reduction in the duration of TI episodes and nociception, the latter measured by the vocalization test in guinea pigs. The effect of microinjection of carbachol on TI reduction and antinociception was blocked by the previous microinjection of the cholinergic antagonist atropine (0.5 microg/0.2 microl and 1 microg/0.2 microl, respectively), demonstrating the participation of muscarinic receptors in the modulation of these responses. Microinjection of atropine per se did not interfere with the duration of TI episodes. In summary, the present results demonstrate that cholinergic stimulation of the NRM promoted analgesia and a reduction in the duration of TI in guinea pigs. These data indicate that the NRM possibly contributes to the modulation of defensive and nociceptive behavioral responses, probably by modulating the activity of neurons in the ventral and dorsal horn of the spinal cord, respectively.

GABAA-benzodiazepine receptors in the dorsomedial (Dm) telencephalon modulate restraint-induced antinociception in the fish Leporinus macrocephalus.

The possibility that fish experience pain has been denied based on the absence of the neural substrates to support this "experience". In this context, the identification of brain regions involved in nociception modulation could provide important insights regarding the processing of nociceptive information in fish. Our study evaluated the participation of the GABAA-benzodiazepine receptor in the dorsomedial (Dm) telencephalon in restraint-induced antinociception in the fish Leporinus macrocephalus through the microinjection of the anxiolytic drug midazolam. The microinjection of midazolam in the Dm did not alter the nocifensive response; however, this drug did block the inhibition of the nocifensive response to formaldehyde promoted by restraint stress. The fish that received midazolam (40nmol) microinjection prior to restraint (3 or 5min), followed by subcutaneous injection with formaldehyde presented a higher distance traveled than the fish that received saline microinjection. This effect might reflect the specific action of midazolam on benzodiazepine receptors in the Dm telencephalon, as pre-treatment with flumazenil, a benzodiazepine receptor antagonist, inhibited the effects of this drug. In the present study, we present the first evidence demonstrating a role for the dorsomedial telencephalic region in the modulation of stress-induced antinociception in fish, revealing new perspectives in the understanding of nociceptive information processing in this group.

Acute administration of a cannabinoid CB1 receptor antagonist impairs stress-induced antinociception in fish.

This study evaluated the influence of the pre-treatment with AM251 (a cannabinoid type I receptor (CB1) selective antagonist) on the stress-induced antinociception promoted by restraint in the fish Leporinus macrocephalus. The application of 3 and 5 min of restraint stress promoted an inhibition of the behavioural response to the subcutaneous injection of 3% formaldehyde (increase in locomotor activity), suggesting the activation of an antinociceptive system. The acute intraperitoneal administration of AM251 (3 mg·kg(-1)) impaired this antinociceptive response induced by 3 and 5 min of restraint stress. The fish treated with AM251 before the application of restraint stress presented an increase in locomotor activity after the subcutaneous injection of formaldehyde, similar to fish not exposed to restraint, suggesting that the stress-induced antinociception promoted by restraint in fish is probably mediated by cannabinoid CB1 receptors. The results presented in this paper suggest the participation of the endocannabinoid system in nociception modulation in fish, supporting the hypothesis that an endogenous antinociceptive system activated by restraint stress is present in fish and that the modulation of antinociception by the CB1 receptor is evolutionary well-conserved across vertebrates.

Serotonin 5-HT2 and 5-HT1A receptors in the periaqueductal gray matter differentially modulate tonic immobility in guinea pig.

Tonic immobility (TI) is an inborn defensive behavior characterized by a temporary state of profound and reversible motor inhibition elicited by some forms of physical restraint. We have previously reported that cholinergic stimulation of the dorsal periaqueductal gray matter (PAG) decreases the duration of TI episodes, while stimulation of the ventrolateral region increases it. The ventrolateral PAG modulates this behavior via a similar neural circuit proposed to be involved in the antinociceptive system. Some studies have indicated that alterations in the levels of cerebral 5-hydroxytryptamine (5-HT) mediate or modulate the analgesic effect of PAG stimulation. Thus, in this study we investigated the possibility that the serotoninergic system is involved in the modulation of TI by this neural substrate. Our results showed that the effect of serotonin into the ventrolateral and dorsal PAG seems to be biphasic and dose dependent. The microinjection of low doses (0.1 microg) of 5-HT into the PAG increased the duration of TI, while high doses (1, 3 and 6 microg) decreased this behavior. Our results also showed that microinjection of a 5-HT(1A) agonist (0.003, 0.01 and 0.1 microg of 8-hydroxy-dipropylaminotretalin (8-OH-DPAT)) into the PAG increased the duration of TI episodes. However, the microinjection of 5-HT(2) agonist (0.01 and 0.1 microg of alpha-methyl-5-HT) into the PAG decreased the duration of TI and this effect could be reversed by pretreatment with an ineffective dose (0.01 microg) of ketanserin. In contrast, ketanserin (0.03 and 0.16 microg) increased this behavior in a dose-dependent manner. These results suggest that the PAG 5-HT(1A) and 5-HT(2) receptors have different roles in the modulation of TI in guinea pigs, since the 5-HT(1A) and 5-HT(2) agonists, respectively, increased and decreased the duration of TI.

Central nucleus of the amygdala and the control of tonic immobility in guinea pigs.

Tonic immobility (TI) is a temporary state of profound motor inhibition induced by situations that supposedly generate intense fear, with the objective to protect the animal from attacks by predators. A previous study by our group demonstrated that cholinergic stimulation of the central, basolateral, and lateral posterior nuclei of the amygdala decreases the duration of TI in guinea pigs. In the current study, we attempted to investigate the effects of cholinergic, opioidergic, and gamma-aminobutyric acid (GABA)ergic stimulation of the central amygdala (CEA) on TI modulation. We observed that both cholinergic (carbachol, 2.7 nmol/0.2 microl) and opioidergic (morphine, 2.2 and 4.4 nmol/0.2 microl) stimulation of the CEA decreased the duration of TI and that these effects could be reversed by pretreatment with naloxone (1.3 and 2.7 nmol/0.2 microl). Our results also showed that microinjection of the GABAergic agonist muscimol (0.26 nmol/0.2 microl) reduced the duration of TI episodes, whereas microinjection of the GABAergic antagonist bicuculline (BIC, 1 nmol/0.2 microl) increased it. Thus, the present experiments demonstrated that cholinergic, opioidergic, and GABAergic systems of the CEA have an inhibitory action on the duration of TI in guinea pigs. Furthermore, the current study suggests an interaction of cholinergic and opioidergic mechanisms. In addition, the GABAergic circuit of the CEA has a tonic inhibitory influence on the duration of TI and is mediated by GABA(A) receptors.

The cholinergic stimulation of the central amygdala modifying the tonic immobility response and antinociception in guinea pigs depends on the ventrolateral periaqueductal gray.

Tonic immobility (TI), also known as death feigning or animal hypnosis, is a reversible state of motor inhibition that is triggered by postural inversion and/or movement restraining maneuvers but also by repetitive stimulation and pressure on body parts. Our previous studies demonstrated that cholinergic stimulation of the central amygdala (CEA) decreases the duration of TI in guinea pigs. Some reports have demonstrated that electrical or chemical stimulation of the CEA promotes antinociception. Evidence suggests that the CEA performs part of its functions by means of a connection with the ventrolateral periaqueductal gray (vlPAG). In the current study, we investigated the participation of a possible functional and anatomical CEA-vlPAG connection in guinea pigs in the regulation of the TI response and antinociception. Our results showed that the functional CEA-vlPAG connection is essential for the participation of the CEA in the modulation of TI and of antinociception. The reversible exclusion of the vlPAG by means of microinjection of 2% lidocaine blocked the inhibitory effect on TI duration and the antinociceptive effect, as determined by a decrease of the vocalization index (VI) obtained with the administration of carbachol (2.7 nmol/0.2 microl) into the CEA. On the other hand, the exclusion of the CEA by lidocaine did not block the antinociception or the increase in TI induced by microinjection of CCh into the vlPAG. Finally, microinjection of the retrograde neurotracer Fast Blue into the CEA or into the vlPAG demonstrated the existence of a reciprocal anatomical connection between the CEA and vlPAG.

Nociceptive vocalization response in guinea pigs modulated by opioidergic, GABAergic and serotonergic neurotransmission in the dorsal raphe nucleus.

The dorsal raphe nucleus (DRN) is involved in the control of several physiological functions, including nociceptive modulation. This nucleus is one of the main sources of serotonin to the CNS and neuromodulators such as opioids and GABA may be are important for its release. This study evaluated the influence of serotonergic, GABAergic and opioidergic stimulation, as well as their interactions in the DRN, on vocalization nociceptive response during a peripheral noxious stimulus application in guinea pigs. Morphine (1.1 nmol), bicuculline (0.50 nmol) and alpha-methyl-5-HT (1.6 nmol) microinjection on the DRN produces antinociception. The antinociception produced by morphine (1.1 nmol) and alpha-methyl-5-HT (1.6 nmol) into the DRN was blocked by prior microinjection of naloxone (0.7 nmol). The alpha-methyl-5-HT effect blocked by naloxone may indicate the existence of 5-HT2A receptors on enkephalinergic interneurons within the dorsal raphe. Pretreatment with muscimol (0.26 nmol) also prevented the antinociceptive effect caused by morphine (1.1 nmol) when administered alone at the same site, indicating an interaction between GABAergic and opioidergic interneurons. The antinociception produced by bicuculline (0.5 nmol) in the DRN was blocked by prior administration of 8-OH-DPAT (0.5 nmol), a 5-HT1A agonist. This may indicate that the 5-HT autoreceptor activation by 8-OH-DPAT at DRN effector neurons can oppose the bicuculline disinhibition effect applied to the same effectors. Thus, we suggest that 5-HT2 receptor activation in the DRN promotes endorphin/enkephalin release that may disinhibit efferent serotonergic neurons of this present structure by inhibiting GABAergic interneurons, resulting in antinociception.

Glutamate/GABA balance in ACC modulates the nociceptive responses of vocalization: an expression of affective-motivational component of pain in guinea pigs.

Evidence corroborates the role of the anterior cingulate cortex (ACC) in the modulation of cognitive and emotional functions. Its involvement in the motivational-affective component of pain has been widely investigated using different methods to elucidate the specific role of different neurotransmitter systems. We used the peripheral noxious stimulus-induced vocalization algesimetric test to verify glutamatergic and GABAergic neurotransmission in the guinea pig ACC. Microinjection of homocysteic acid (DLH; 30 nmol) in the left guinea pig ACC increased the amplitude of vocalizations (pronociception) compared to controls injected with saline. Moreover, microinjection of MK-801 (3.6 nmol), an NMDA receptor antagonist, did not alter the amplitude of vocalizations, but its microinjection prior to DLH prevented the increase in vocalizations induced by this drug. Regarding the GABAergic system, blockade of GABAA receptors with bicuculline (1 nmol) increased the amplitude of vocalizations, while three different doses of the GABAA agonist muscimol (0.5, 1 and 2 nmol) did not influence nociceptive vocalization responses. Finally, a combination of MK-801 (3.6 nmol) and muscimol (1 nmol) reduced the amplitude of vocalizations (antinociception), suggesting that a combination of glutamate and GABA in the ACC modulates the expression of affective-motivational pain response. We suggest that activation of NMDA receptors or blockade of GABAergic neurotransmission promotes pronociception and that the antinociceptive effect of muscimol depends on the blockade of NMDA receptors.

Rostral ventromedial medulla connections in Cavia porcellus and their relation with tonic immobility defensive behavior: a biotinylated dextran amine neurotracing study.

The rostroventromedial medulla (RVM), together with the periaqueductal gray matter (PAG), constitutes the descendent antinociceptive system. Additionally, these structures mutually regulate defensive behaviors, including tonic immobility (TI) in guinea pigs. The current study was undertaken to evaluate the connections of the RVM with the PAG and the spinal cord in guinea pigs in order to provide an anatomical basis for the role played by RVM in the modulation of TI. To address this goal, five guinea pigs were treated with non-fluorescent biotinylated dextran amine (BDA) neurotracer by injection into the RVM. After four days of survival, the encephalon and spinal cord were removed from each rodent, and BDA labeling was visualized with a standard avidin-biotinylated horseradish peroxidase method through reaction with nickel-intensified peroxidase 3,3'-diaminobenzidine dihydrochloride. The microinjection of BDA into the RVM stained fibers in the ventral horn, dorsal horn and intermediate gray matter of the spinal cord. BDA-labeled fibers, terminal buttons suggesting synaptic contacts, and perikarya were found in the dorsomedial, dorsolateral, lateral and ventrolateral PAG, and neuronal somata were identified in the cuneiform nucleus. Together, the current data demonstrate neuroanatomical evidence that supports the role of the RVM in the modulation of TI defensive behavior.

Stress-induced antinociception in fish reversed by naloxone.

Pain perception in non-mammalian vertebrates such as fish is a controversial issue. We demonstrate that, in the fish Leporinus macrocephalus, an imposed restraint can modulate the behavioral response to a noxious stimulus, specifically the subcutaneous injection of 3% formaldehyde. In the first experiment, formaldehyde was applied immediately after 3 or 5 min of the restraint. Inhibition of the increase in locomotor activity in response to formaldehyde was observed, which suggests a possible restraint-induced antinociception. In the second experiment, the noxious stimulus was applied 0, 5, 10 and 15 min after the restraint, and both 3 and 5 min of restraint promoted short-term antinociception of approximately 5 min. In experiments 3 and 4, an intraperitoneal injection of naloxone (30 mg.kg(-1)) was administered 30 min prior to the restraint. The 3- minute restraint-induced antinociception was blocked by pretreatment with naloxone, but the corresponding 5-minute response was not. One possible explanation for this result is that an opioid and a non-preferential µ-opioid and/or non-opioid mechanism participate in this response modulation. Furthermore, we observed that both the 3- and 5- minutes restraint were severely stressful events for the organism, promoting marked increases in serum cortisol levels. These data indicate that the response to a noxious stimulus can be modulated by an environmental stressor in fish, as is the case in mammals. To our knowledge, this study is the first evidence for the existence of an endogenous antinociceptive system that is activated by an acute standardized stress in fish. Additionally, it characterizes the antinociceptive response induced by stress in terms of its time course and the opioid mediation, providing information for understanding the evolution of nociception modulation.

Opioidergic, GABAergic and serotonergic neurotransmission in the dorsal raphe nucleus modulates tonic immobility in guinea pigs.

Tonic immobility (TI) is an innate defensive behavior that can be elicited by physical restriction and postural inversion and is characterized by a profound and temporary state of akinesis. Our previous studies demonstrated that the stimulation of serotonin receptors in the dorsal raphe nucleus (DRN) appears to be biphasic during TI responses in guinea pigs (Cavia porcellus). Serotonin released by the DRN modulates behavioral responses and its release can occur through the action of different neurotransmitter systems, including the opioidergic and GABAergic systems. This study examines the role of opioidergic, GABAergic and serotonergic signaling in the DRN in TI defensive behavioral responses in guinea pigs. Microinjection of morphine (1.1 nmol) or bicuculline (0.5 nmol) into the DRN increased the duration of TI. The effect of morphine (1.1 nmol) was antagonized by pretreatment with naloxone (0.7 nmol), suggesting that the activation of µ opioid receptors in the DRN facilitates the TI response. By contrast, microinjection of muscimol (0.5 nmol) into the DRN decreased the duration of TI. However, a dose of muscimol (0.26 nmol) that alone did not affect TI, was sufficient to inhibit the effect of morphine (1.1 nmol) on TI, indicating that GABAergic and enkephalinergic neurons interact in the DRN. Microinjection of alpha-methyl-5-HT (1.6 nmol), a 5-HT(2) agonist, into the DRN also increased TI. This effect was inhibited by the prior administration of naloxone (0.7 nmol). Microinjection of 8-OH-DPAT (1.3 nmol) also blocked the increase of TI promoted by morphine (1.1 nmol). Our results indicate that the opioidergic, GABAergic and serotonergic systems in the DRN are important for modulation of defensive behavioral responses of TI. Therefore, we suggest that opioid inhibition of GABAergic neurons results in disinhibition of serotonergic neurons and this is the mechanism by which opioids could enhance TI. Conversely, a decrease in TI could occur through the activation of GABAergic interneurons.

Rostral ventromedial medulla modulates nociception and tonic immobility behavior through connections with the A7 catecholaminergic region.

Cholinergic stimulation of the rostral ventromedial medulla (RVM) produces antinociception and reduces the duration of tonic immobility (TI) behavior in guinea pigs. Previous studies indicated that cholinergic antinociception in the RVM is mediated through connections with the A7 catecholaminergic cell group (A7). In the current study, we tested the role of the A7 in both the antinociception and reduction of TI duration mediated by cholinergic stimulation of the RVM. In addition, we used biotinylated dextran amines (BDA) to evaluate the connections between the RVM and A7. The microinjection of the cholinergic agonist carbachol into the RVM produced antinociception and reduced TI behavior duration. These effects were blocked by prior administration of lidocaine to the A7. However, the microinjection of lidocaine into the A7 prior to saline injection into the RVM had no effect on either the nociceptive or TI responses. The microinjection of the neurotracer BDA into the RVM positively stained fibers and synaptic boutons in the A7, indicating that there are direct projections from the RVM to the A7. Taken together, our results indicate that the antinociception and reduction of TI behavior duration after cholinergic stimulation of the RVM depends on connections with the A7.

The functional role of ascending nociceptive control in defensive behavior.

Ascending nociceptive control is a novel spino-striato-rostral ventral medulla pain modulation pathway that mediates heterosegmental pain-induced analgesia, i.e., noxious stimulus-induced antinociception. In this study, we used the dorsal immobility response in rats as a model of the defensive responses. We demonstrated that the activation of ascending nociceptive control by peripheral noxious stimulation and spinal AMPA and mGluR1 receptor blockade significantly potentiated the duration of the dorsal immobility response in rats via an opioid-dependent mechanism in the nucleus accumbens. These results demonstrated the functional role of ascending nociceptive control in the modulation of defensive responses and spinal glutamatergic receptors in the dorsal immobility response. The immobility response is an antipredator behavior that reflects the underlying state of fear, and ascending nociceptive control may modulate fear.