Induction of pain facilitation by sustained opioid exposure: relationship to opioid antinociceptive tolerance

Opioid analgesics are frequently used for the long-term management of chronic pain states, including cancer pain. The prolonged use of opioids is associated with a requirement for increasing doses to manage pain at a consistent level, reflecting the phenomenon of analgesic tolerance. It is now becoming clearer that patients receiving long-term opioid therapy can develop unexpected abnormal pain. Such paradoxical opioid-induced pain, as well as tolerance to the antinociceptive actions of opioids, has been reliably measured in animals during the period of continuous opioid delivery. Several recent studies have demonstrated that such pain may be secondary to neuroplastic changes that result, in part, from an activation of descending pain facilitation mechanisms arising from the rostral ventromedial medulla (RVM). One mechanism which may mediate such pain facilitation is through the increased activity of CCK in the RVM. Secondary consequences from descending facilitation may be produced. For example, opioid-induced upregulation of spinal dynorphin levels seem to depend on intact descending pathways from the RVM reflecting spinal neuroplasticity secondary to changes at supraspinal levels. Increased expression of spinal dynorphin reflects a trophic action of sustained opioid exposure which promotes an increased pain state. Spinal dynorphin may promote pain, in part, by enhancing the evoked release of excitatory transmitters from primary afferents. In this regard, opioids also produce trophic actions by increasing CGRP expression in the dorsal root ganglia. Increased pain elicited by opioids is a critical factor in the behavioral manifestation of opioid tolerance as manipulations which block abnormal pain also block antinociceptive tolerance. Manipulations that have blocked enhanced pain and antinociceptive tolerance include reversible and permanent ablation of descending facilitation from the RVM. Thus, opioids elicit systems-level adaptations resulting in pain due to descending facilitation, upregulation of spinal dynorphin and enhanced release of excitatory transmitters from primary afferents. Adaptive changes produced by sustained opioid exposure including trophic effects to enhance pain transmitters suggest the need for careful evaluation of the consequences of long-term opioid administration to patients.     

Underlying mechanisms of pronociceptive consequences of prolonged morphine exposure

The opioid analgesics, commonly exemplified by morphine, represent the best option for the treatment of severe pain and for the management of chronic pain states, of both malignant and nonmalignant origin. It is well recognized that the prolonged use of opioids is associated with a requirement for ever-increasing doses in order to maintain pain relief at an acceptable and consistent level. This phenomenon is termed analgesic tolerance. While the concept that tolerance can develop as a result of cellular adaptations to the presence of the opioid has been proposed, it is now becoming abundantly clear that tolerance may also be related to a state of hyperalgesia that results from exposure to the opioid itself. Patients who receive long-term opioid therapy sometimes develop unexpected, abnormal pain. Similar paradoxical opioid-induced pain has been confirmed in a number of animal studies, even during the period of continuous opioid delivery. A number of recent studies have demonstrated that such pain may be secondary to neuroplastic changes that occur in the brain and spinal cord. One such change may be the activation of descending pain facilitation mechanisms arising from the rostral ventromedial medulla (RVM) elicited in part by increased activity of cholecystokinin (CCK) in the RVM. A cascade of pronociceptive events may follow, such as opioid-induced upregulation of spinal dynorphin levels that promotes enhanced input from primary afferent nociceptors. This mechanism appears to depend on intact descending pathways from the RVM, since interrupting this pathway abolishes enhanced abnormal pain. Furthermore, extended opioid exposure also can elicit increased calcitonin gene related peptide (CGRP) and substance P expression in the dorsal root ganglia. It is probable that increased pain elicited by opioids is a critical factor in the behavioral manifestation of opioid tolerance because the same manipulations that block abnormal pain also block antinociceptive tolerance. Taken together, such studies show that opioids elicit systems-level adaptations resulting in pain due to descending facilitation, upregulation of spinal dynorphin, and enhanced, evoked release of excitatory transmitters from primary afferents. These adaptive changes in response to sustained exposure to opioids indicate the need for the evaluation of the clinical consequences of long-term opioid administration. Additionally, these findings suggest a need for novel chemistry involving design of agents that may counteract opiate-induced neuroplastic adaptations resulting in pain relief without analgesic tolerance.     

Is paradoxical pain induced by sustained opioid exposure an underlying mechanism of opioid antinociceptive tolerance?

Opiates are the primary treatment for pain management in cancer patients reporting moderate to severe pain, and are being increasingly used for non-cancer chronic pain. However, prolonged administration of opiates is associated with significant problems including the development of antinociceptive tolerance, wherein higher doses of the drug are required over time to elicit the same amount of analgesia. High doses of opiates result in serious side effects such as constipation, nausea, vomiting, dizziness, somnolence, and impairment of mental alertness. In addition, sustained exposure to morphine has been shown to result in paradoxical pain in regions unaffected by the initial pain complaint, and which may also result in dose escalation, i.e. 'analgesic tolerance'. A concept that has been gaining considerable experimental validation is that prolonged use of opioids elicits paradoxical, abnormal pain. This enhanced pain state requires additional opioids to maintain a constant level of antinociception, and consequently may be interpreted as antinociceptive tolerance. Many substances have been shown to block or reverse antinociceptive tolerance. A non-inclusive list of examples of substances reported to block or reverse opioid antinociceptive tolerance include: substance P receptor (NK-1) antagonists, calcitonin gene-related peptide (CGRP) receptor antagonists, nitric oxide (NO) synthase inhibitors, calcium channel blockers, cyclooxygenase (COX) inhibitors, protein kinase C inhibitors, competitive and non-competitive antagonists of the NMDA (N-methyl-D-aspartate) receptor, AMPA (alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid) antagonists, anti-dynorphin antiserum, and cholecystokinin (CCK) receptor antagonists. Without exception, these substances are also antagonists of pain-enhancing agents. Prolonged opiate administration indeed induces upregulation of substance P (SP) and calcitonin gene-related peptide (CGRP) within sensory fibers in vivo, and this is accompanied by an enhanced release of excitatory neurotransmitters and neuropeptides from primary afferent fibers upon stimulation. The enhanced evoked release of neuropeptides is correlated with the onset of abnormal pain states and opioid antinociceptive tolerance. Importantly, the descending pain modulatory pathway from the brainstem rostral ventromedial medulla (RVM) via the dorsolateral funiculus (DLF) is critical for maintaining the changes observed in the spinal cord, abnormal pain states and antinociceptive tolerance, because animals with lesion of the DLF did not show enhanced evoked neuropeptide release, or develop abnormal pain or antinociceptive tolerance upon sustained exposure to opiates. Microinjection of either lidocaine or a CCK antagonist into the RVM blocked both thermal and touch hypersensitivity as well as antinociceptive tolerance. Thus, prolonged opioid exposure enhances a descending pain facilitatory pathway from the RVM that is mediated at least in part by CCK activity and is essential for the maintenance of antinociceptive tolerance.     

Structure-activity relationships of bifunctional cyclic disulfide peptides based on overlapping pharmacophores at opioid and cholecystokinin receptors

Prolonged opioid exposure increases the expression of cholecystokinin (CCK) and its receptors in the central nervous system (CNS), where CCK may attenuate the antinociceptive effects of opioids. The complex interactions between opioid and CCK may play a role in the development of opioid tolerance. We designed and synthesized cyclic disulfide peptides and determined their agonist properties at opioid receptors and antagonist properties at CCK receptors. Compound 1 (Tyr-c[d-Cys-Gly-Trp-Cys]-Asp-Phe-NH(2)) showed potent binding and agonist activities at delta and mu opioid receptors but weak binding to CCK receptors. The NMR structure of the lead compound displayed similar conformational features of opioid and CCK ligands.     

Chronic morphine exposure increases the proportion of on-cells in the rostral ventromedial medulla in rats

Chronic opiate exposure produces tolerance and hypersensitivity to mechanical and thermal stimulation that involves increased pain facilitation from the rostral ventromedial medulla (RVM). The aim of the present study was to determine the effect of sustained systemic morphine exposure on RVM neurons. Three cell types in the RVM have been described: on-cells, off-cells and neutral cells. The activity of on-cells increases in response to noxious stimulation, whereas the activity of off-cells decreases following noxious stimulation. Neutral cells remain relatively unaffected. In lightly anesthetized rats, systematic exploration throughout the RVM using single-unit extracellular recordings was used to examine both the relative proportion and the neuronal properties of the different cell classes in chronic morphine and placebo treated animals. Seven days after implanting either morphine (150 mg, s.c.) or placebo pellets a total of four electrode penetrations through the RVM were made in each animal at identical coordinates along midline. Neuronal responses related to radiant heat-evoked paw withdrawals were recorded. When compared to placebo treated rats, chronic morphine increased the number of on-cells and decreased the number of neutral cells, while the number of off-cells remained unchanged. Chronic morphine exposure had no effect on the spontaneous or heat-evoked discharges in on-, off-, or neutral cells. These results indicate that chronic morphine may sensitize a subpopulation of RVM neurons to noxious stimulation, which would be expected to increase descending facilitation and promote tolerance and chronic morphine-induced paradoxical pain.     

Increased glutamate synaptic transmission in the nucleus raphe magnus neurons from morphine-tolerant rats

Currently, opioid-based drugs are the most effective pain relievers that are widely used in the treatment of pain. However, the analgesic efficacy of opioids is significantly limited by the development of tolerance after repeated opioid administration. Glutamate receptors have been reported to critically participate in the development and maintenance of opioid tolerance, but the underlying mechanisms remain unclear. Using whole-cell voltage-clamp recordings in brainstem slices, the present study investigated chronic morphine-induced adaptations in glutamatergic synaptic transmission in neurons of the nucleus raphe magnus (NRM), a key supraspinal relay for pain modulation and opioid analgesia. Chronic morphine significantly increased glutamate synaptic transmission exclusively in one class of NRM cells that contains μ-opioid receptors in a morphine-tolerant state. The adenylyl cyclase activator forskolin and the cAMP analog 8-bromo-cAMP mimicked the chronic morphine effect in control neurons and their potency in enhancing the glutamate synaptic current was significantly increased in neurons from morphine-tolerant rats. MDL12330a, an adenylyl cyclase inhibitor, and H89, a protein kinase A (PKA) inhibitor, reversed the increase in glutamate synaptic transmission induced by chronic morphine. In addition, PMA, a phorbol ester activator of protein kinase C (PKC), also showed an increased potency in enhancing the glutamate synaptic current in these morphine-tolerant cells. The PKC inhibitor GF109203X attenuated the chronic morphine effect. Taken together, these results suggest that chronic morphine increases presynaptic glutamate release in μ receptor-containing NRM neurons in a morphine-tolerant state, and that the increased glutamate synaptic transmission appears to involve an upregulation of both the cAMP/PKA pathway and the PKC pathway. This glutamate-mediated activation of these NRM neurons that are thought to facilitate spinal pain transmission may contribute to the reduced opioid analgesia during opioid tolerance.     

Association of enkephalin catabolism inhibitors and CCKB antagonists: A potential use in the management of pain and opioid addiction

The overlapping distribution of opioid and cholecystokinin (CCK) peptides and their receptors (μ and δ opioid receptors; CCK-A and CCK-B receptors) in the central nervous system have led to a large number of studies aimed at clarifying the functional relationships between these two neuropeptides. Most of the pharmacological studies devoted to the role of CCK and enkephalins have been focused on the control of pain. Recently the existence of regulatory mechanisms between both systems have been proposed, and the physiological antagonism between CCK and endogenous opioid systems has been definitely demonstrated by coadministration of CCK-B selective antagonists with RB 101, a systemically active inhibitor, which fully protects enkephalins from their degradation. Several studies have also been done to investigate the functional relationships between both systems in development of opioid side-effects and in behavioral responses. This article will review the experimental pharmacology of association of enkephalin-degrading enzyme inhibitors and CCK-B antagonists to demonstrate the interest of these molecules in the management of both pain and opioid addiction. Special issue dedicated to Dr. Eric J. Simon.     

Tolerance Effects Induced by NSAID Microinjections into the Central Nucleus of the Amygdala in Rats

Recent investigations have shown that microinjections of non-opioid analgesics, nonsteroidal anti-inflammatory drugs, NSAIDs, into some brain areas, particularly, into the midbrain periaqueductal gray matter (PAG) and rostral ventro-medial medulla (RVM), cause antinociception with some effects of tolerance. Our preliminary findings have also shown the same effects of tolerance after intraperitoneal injections. The present study was designed to examine whether microinjections of metamizole (Analgin), ketorolac, and xefocam into the central nucleus of the amygdala (Ce) lead to the development of tolerance in rats, and to ascertain whether this nucleus is the pain-modulating pathway through PAG. Our investigation revealed that microinjections of NSAIDs into the Ce both unilaterally (the left side) and bilaterally produced antinociception, as indicated by a latency increase in tail-flick reflex (TF) compared to controls with saline, on the first experimental day for Analgin (P < 0.001), ketorolac (P < 0.001), and xefocam (P < 0.001). However, when these drug microinjections were repeated during subsequent days, the antinociceptive effects progressively diminished so that on the fifth experimental day the TF latency was similar to that in the rats that received repeated injections of only saline. These results show that, alongside with PAG and RVM, the Ce is an important site of the endogenous antinociceptive system, which triggers the descending pain control mechanism and thus inhibits nociceptive transmission. On the other hand, our data confirm the results of other authors that NSAIDs are closely related to endogenous opioids, and tolerance to these non-opioid drugs probably depends on opioid tolerance.     

The mechanism of μ-opioid receptor (MOR)-TRPV1 crosstalk in TRPV1 activation involves morphine anti-nociception,tolerance and dependence

Initiated by the activation of various nociceptors, pain is a reaction to specific stimulus modalities. The μ-opioid receptor (MOR) agonists, including morphine, remain the most potent analgesics to treat patients with moderate to severe pain. However, the utility of MOR agonists is limited by the adverse effects associated with the use of these drugs, including analgesic tolerance and physical dependence. A strong connection has been suggested between the expression of the transient receptor potential vanilloid type 1 (TRPV1) ion channel and the development of inflammatory hyperalgesia. TRPV1 is important for thermal nociception induction, and is mainly expressed on sensory neurons. Recent reports suggest that opioid or TRPV1 receptor agonist exposure has contrasting consequences for anti-nociception, tolerance and dependence. Chronic morphine exposure modulates TRPV1 activation and induces the anti-nociception effects of morphine. The regulation of many downstream targets of TRPV1 plays a critical role in this process, including calcitonin gene-related peptide (CGRP) and substance P (SP). Additional factors also include capsaicin treatment blocking the anti-nociception effects of morphine in rats, as well as opioid modulation of TRPV1 responses through the cAMP-dependent PKA pathway and MAPK signaling pathways. Here, we review new insights concerning the mechanism underlying MOR-TRPV1 crosstalk and signaling pathways and discuss the potential mechanisms of morphine-induced anti-nociception, tolerance and dependence associated with the TRPV1 signaling pathway and highlight how understanding these mechanisms might help find therapeutic targets for the treatment of morphine induced antinociception, tolerance and dependence.     

The mechanism of μ-opioid receptor (MOR)-TRPV1 crosstalk in TRPV1 activation involves morphine anti-nociception,tolerance and dependence

Initiated by the activation of various nociceptors, pain is a reaction to specific stimulus modalities. The μ-opioid receptor (MOR) agonists, including morphine, remain the most potent analgesics to treat patients with moderate to severe pain. However, the utility of MOR agonists is limited by the adverse effects associated with the use of these drugs, including analgesic tolerance and physical dependence. A strong connection has been suggested between the expression of the transient receptor potential vanilloid type 1 (TRPV1) ion channel and the development of inflammatory hyperalgesia. TRPV1 is important for thermal nociception induction, and is mainly expressed on sensory neurons. Recent reports suggest that opioid or TRPV1 receptor agonist exposure has contrasting consequences for anti-nociception, tolerance and dependence. Chronic morphine exposure modulates TRPV1 activation and induces the anti-nociception effects of morphine. The regulation of many downstream targets of TRPV1 plays a critical role in this process, including calcitonin gene-related peptide (CGRP) and substance P (SP). Additional factors also include capsaicin treatment blocking the anti-nociception effects of morphine in rats, as well as opioid modulation of TRPV1 responses through the cAMP-dependent PKA pathway and MAPK signaling pathways. Here, we review new insights concerning the mechanism underlying MOR-TRPV1 crosstalk and signaling pathways and discuss the potential mechanisms of morphine-induced anti-nociception, tolerance and dependence associated with the TRPV1 signaling pathway and highlight how understanding these mechanisms might help find therapeutic targets for the treatment of morphine induced antinociception, tolerance and dependence.     

Tracking the opioid receptors on the way of desensitization

Opioid receptors belong to the super family of G-protein coupled receptors (GPCRs) and are the targets of numerous opioid analgesic drugs. Prolonged use of these drugs results in a reduction of their effectiveness in pain relief also called tolerance, a phenomenon well known by physicians. Opioid receptor desensitization is thought to play a major role in tolerance and a lot of work has been dedicated to elucidate the molecular basis of desensitization. As described for most of GPCRs, opioid receptor desensitization involves their phosphorylation by kinases and their uncoupling from G-proteins realized by arrestins. More recently, opioid receptor trafficking was shown to contribute to desensitization. In this review, our knowledge on the molecular mechanisms of desensitization and recent progress on the role of opioid receptor internalization, recycling or degradation in desensitization will be reported. A better understanding of these regulatory mechanisms would be helpful to develop new analgesic drugs or new strategies for pain treatment by limiting opioid receptor desensitization and tolerance.     

Opioid tolerance and the emergence of new opioid receptor-coupled signaling

Multiple cellular adaptations are elicited by chronic exposure to opioids. These include diminution of spare opioid receptors, decreased opioid receptor density, and G-protein content and coupling thereof. All imply that opioid tolerance is a manifestation of a loss of opioid function, i.e., desensitization. Recent observations challenge the exclusiveness of this formulation and indicate that opioid tolerance also results from qualitative changes in opioid signaling. In this article, Gintzler and Chakrabarti discuss the evidence that suggests that opioid tolerance results not only from impaired opioid receptor functionality, but also from altered consequences of coupling. Underlying the latter are fundamental changes in the nature of effectors that are coupled to the opioid receptor/G-protein signaling pathway. These molecular changes include the upregulation of adenylyl cyclase isoforms of the type II family as well as a substantial increase in their phosphorylation state. As a result, there is a shift in opioid receptor/G-protein signaling from predominantly G_iα inhibitory to G_βγ stimulatory following chronic in vivo morphine exposure. These adaptations to chronic morphine indicate the plasticity of opioid-signal transduction mechanisms and the ability of chronic morphine to augment new signaling strategies.     

Post-opioid receptor adaptations to chronic morphine; altered functionality and associations of signaling molecules

Opioid desensitization/tolerance mechanisms have largely focused on adaptations that occur on the level of the mu-opioid receptor (MOR) itself. These include opioid receptor phosphorylation and ensuing trafficking events. Recent research, however, has revealed additional adaptations that occur downstream from the opioid receptor, which involve covalent modification of signaling molecules and altered associations among them. These include augmented isoform-specific synthesis of adenylyl cyclase (AC) and their phosphorylation as well as augmented phosphorylation of the G(beta) subunit of G(beta gamma). The aggregate effect of these changes is to shift mu-opioid receptor-coupled signaling from predominantly G(i alpha) inhibitory to (G(i)-derived) G(beta gamma) stimulatory AC signaling. Most recently, chronic morphine has been shown to enhance the association (interaction) between MOR and G(s), which should provide an additional avenue for offsetting inhibitory MOR signaling sequelae. The unfolding complexity of chronic morphine-induced sequelae demands an evolving broader and more encompassing perspective on opioid tolerance-producing mechanisms. This should facilitate understanding tolerance within the context of physiological plasticity that is activated by chronic exposure to drugs of abuse. Additional research is required to integrate the various tolerance-producing adaptations that have been elucidated to date. Specifically, the relative contribution to opioid tolerance of identified adaptations is still unknown as is the extent to which they vary among different regions of the central nervous system.     

Descending antinociceptive mechanisms in the brainstem: Their role in the animal's defensive system

The identification of specialized mechanisms in the mammalian brainstem that function to inhibit the rostral transmission of nociceptive (pain-related) information in the spinal cord led to an explosion of research into the neuroanatomical and neurochemical substrates of these antinociceptive systems. As outlined in the present paper, most attention was directed at those mechanisms in the periaqueductal grey (PAG) and rostral ventromedial medulla (RVM). However, comparatively little attention has been paid to the functional role of these mechanisms in animal behaviour. The purpose of the present paper is to review research into the behavioural significance of those antinociceptive mechanisms in the PAG and RVM. It is concluded that these mechanisms function as part of the animal's fear or defensive system, serving to make a threatened animal insensitive to noxious stimulation and thereby allowing that animal to engage in defensive responses instead of recuperative activities. Further, it is argued that the organization of these antinociceptive circuits reflects the animal's increasing capacity for early detection of danger. Specifically, nociception itself is held to signify the presence of immediate threat, and consequently, nociceptive input directly activates antinociceptive circuits at either the spinal level (during intense noxious stimulation) or RVM (following exposure to moderate noxious stimuli). In contrast, events that are themselves innocuous but which signal threat (either learned or innate danger signals) activate fear and defensive systems in the amygdala and PAG which engage the descending antinociceptive projections in the RVM.     

Activation of ERK signaling in rostral ventromedial medulla is dependent on afferent input from dorsal column pathway and contributes to acetic acid-induced visceral nociception

Several lines of evidence from both animal and clinical studies have demonstrated that dorsal column (DC) pathway plays a critical role in visceral pain transmission from the spinal cord to supraspinal center. The descending pain modulation pathway from the rostral ventromedial medulla (RVM) area has been implicated in visceral nociceptive neurotransmission. Previous studies have demonstrated that the multiple protein kinase signaling transduction cascades in the RVM area contribute to the descending facilitation of inflammatory pain and neuropathic pain. However, whether these signaling transduction pathways in the RVM area are triggered by the afferent visceral input from the DC pathway during acute visceral pain remains elusive. Here, we have tested the hypothesis that the afferent visceral stimuli from the DC pathway might induce the activation of extracellular signal-regulated protein kinase (ERK) signaling in the RVM area and contribute to the descending facilitation of neurotransmission in a rat model of visceral pain. Our results showed that acetic acid-induced visceral nociception produced a persistent activation of ERK in the RVM area and a microinjection of a mitogen-activated ERK kinase (MEK) inhibitor, U0126, into the RVM area significantly inhibited the visceral noxious stimulation-induced behaviors in rats. A microinjection of lidocaine into the nucleus gracilis (NG) also inhibited the activation of ERK in the RVM area. The current study indicates that activated ERK signaling pathway in the RVM area is dependent on afferent input from dorsal column pathway and may contribute to acetic acid-induced visceral nociception.     

Increased level of cholecystokinin in cerebrospinal fluid is associated with chronic pain-like behavior in spinally injured rats.

Cholecystokinin (CCK) is a physiological antagonist of opioid-mediated antinociception and may be involved in some chronic pain states where opioids have reduced effect. We have previously shown in a rat model of central neuropathic pain after spinal cord injury that blockade of CCK-B receptors lead to marked pain relief. In the present study, we showed that spinally injured rats exhibiting chronic pain-like behaviors (aversive reaction to innocuous mechanical and cold stimulation) had significantly elevated level of CCK-like immunoreactivity in cerebrospinal fluid compared to normal rats or spinally injured rats which did not exhibit pain-like behaviors. The increased level of circulating CCK in the cerebrospinal fluid may thus contribute to the maintenance of chronic pain in these rats by reducing the endogenous inhibitory tone provided by opioid peptides and may be involved in the phenomenon of opioid insensitivity.     

病理性疼痛的分子机制

持续性或慢性疼痛是很多患者的主要描述症状。然而,现在的治疗手段还不能充分解决某些疼痛或会引起不能忍受的副作用。近来疼痛生物学者阐明了大量的参与疼痛发生和维持的细胞和分子活动。如何更好的理解这些分子活动的机制将有助于发展高效的,特异性的治疗手段。背根神经节中小细胞神经元向脊髓传递温觉和伤害性信息的感觉传递。这些神经元的外周突感受生理性和化学性刺激后,可以在脊髓背角的中枢突通过突触囊泡和大致密性囊泡释放兴奋性的神经递质和神经肽。这种兴奋性突触传递可以被一些抑制因子调控如脊髓中间神经元和下行系统中分泌的阿片肽、GABA、甘氨酸、5-羟色胺。本文将回顾脊髓抑制性系统所取得的一些研究进展,将重点介绍在阿片受体转运,阿片镇痛以及吗啡耐晋研究中的进展,这些发现可能的治疗前景也会一并讨论。     

Supraspinal circuitry mediating opioid antinociception: Antagonist and synergy studies in multiple sites

Supraspinal opioid antinociception is mediated by sensitive brain sites capable of supporting this response following microinjection of opioid agonists. These sites include the ventrolateral periaqueductal gray (vIPAG), the rostral ventromedial medulla (RVM), the locus coeruleus and the amygdala. Each of these sites comprise an interconnected anatomical and physiologically relevant system mediating antinociceptive responses through regional interactions. Such interactions have been identified using two pharmacological approaches: (1) the ability of selective antagonists delivered to one site to block antinociception elicited by opioid agonists in a second site, and (2) the presence of synergistic antinociceptive interactions following simultaneous administration of subthreshold doses of opioid agonists into pairs of sites. Thus, the RVM has essential serotonergic, opioid, cholinergic and NMDA synapses that are necessary for the full expression of morphine antinociception elicited from the vIPAG, and the vIPAG has essential opioid synapses that are necessary for the full expression of opioid antinociception elicited from the amygdala. Further, the vIPAG, RVM, locus coeruleus and amygdala interact with each other in synergistically supporting opioid antinociception.     

Lithium reverses the effect of opioids on eNOS/nitric oxide pathway in human umbilical vein endothelial cells

The main challenge of pain management with opioids is development of acute and chronic analgesic tolerance. Several studies on neuronal cells have focused on the molecular mechanisms involved in tolerance such as cyclic AMP (cAMP) activation, and nitric oxide (NO) pathway. However, the effects of opioids on non-neuronal cells and tolerance development have been poorly investigated. Lithium chloride is a glycogen synthase kinase 3β (GSK-3β) inhibitor and exert its effects through modulation of nitric oxide pathway. In this study we examined the effect of lithium on acute/chronic morphine and methadone administration in endothelial cells which express mu opioid receptors. Human umbilical vein endothelial cells (HUVECs) were treated with different doses of morphine, methadone, and lithium for six and 48 h. Then we evaluated cell viability, nitrite and cyclic AMP levels, as well as the expression of endothelial nitric oxide synthase (eNOS) protein using Immunocytochemistry (ICC) assay and phosphorylated GSK-3β enzyme by western blot analysis in cells. Both chronic morphine and methadone treatment increased NO level and eNOS expression in HUVECs. Morphine induced cAMP overproduction after 48 h exposure with cells. Lithium pretreatment (10 mM) in both morphine and methadone received groups significantly reduced nitrite and cAMP levels as well as eNOS expression as compared to the control. The decreased amount of phospho GSK-3β due to the opioid exposure was increased following lithium treatment. Tolerance like pattern may occur in non-neuronal cells with opioid receptors and this study clearly revealed the attenuation of morphine and methadone tolerance like behavior by lithium treatment in HUVECs.

     

Systemic morphine-induced release of serotonin in the rostroventral medulla is not mimicked by morphine microinjection into the periaqueductal gray

We used in vivo microdialysis in awake rats to test the hypothesis that intravenous morphine increases serotonin (5-HT) release within the rostral ventromedial medulla (RVM). We also injected morphine into various sites along the rostrocaudal extent of the periaqueductal gray (PAG), and examined the extent of its diffusion to the RVM. Intravenous morphine (3.0 mg/kg) produced thermal antinociception and increased RVM dialysate 5-HT, 5-hydroxyindole acetic acid (5-HIAA), and homovanillic acid (HVA) in a naloxone-reversible manner. As neither PAG microinjection of morphine (5 micro g/0.5 micro L) nor RVM administration of fentanyl or d-Ala(2),NMePhe(4),Gly-ol(5)]enkephalin (DAMGO) increased RVM 5-HT, we were unable to determine the precise site of action of morphine. Surprisingly, peak morphine levels in the RVM were higher after microinjection into the caudal PAG as compared to either intravenous injection or microinjection into more rostral sites within the PAG. Naloxone-precipitated withdrawal in morphine-tolerant rats not only increased extracellular 5-HT in the RVM, but also dopamine (DA) and HVA. We conclude that substantial amounts of morphine diffuse from the PAG to the RVM, and speculate that opioid receptor interactions at multiple brain sites mediate the analgesic effects of PAG morphine. Further studies will be required to elucidate the contribution of 5-HT and DA release in the RVM to opioid analgesia and opioid withdrawal.