Functional relationship between brainstem putative pain-facilitating neurons and spinal nociceptfive neurons during development of inflammation in rats.

The so-called on- and off-cells of the rostral ventromedial medulla (RVM) send their axons to the spinal dorsal horn. Activation of on-cells precedes and coincides with a facilitation, and activation of off-cells coincides with an inhibition, of withdrawal reflexes elicited by noxious agents. Considerable evidence supports the notion that on- and off-cells modulate nocifensive reflexes during opioid and non-opioid action and also during normal circumstances and during peripheral neuropathy and inflammation. Yet it is unclear whether on- and off-cells act upon sensory spinal circuits that might lead to ascending projections and the experience of pain. Here, in deeply anesthetized rats we recorded single unit discharges from pairs of one on-like or off-like cell in RVM and a nociceptive neuron in the spinal dorsal horn with input from a hind paw. Both ongoing activity and responses to a calibrated noxious stimulus applied to the paw were documented during basal conditions and during development of paw inflammation. Probably due to the strong barbiturate anesthesia, off-like cells were depressed and did not yield interpretable results. However, we showed for the first time that during the increase in neuronal activity that results from paw inflammation the activity of spinal nociceptive neurons reflects the activity of their partner on-like cells in a highly correlated manner. This implies a tight relationship between spinal sensory and RVM modulatory functions that may underlie inflammation-induced hyperreflexia and clinically relevant hyperalgesia.

Activity correlations between on-like and off-like cells of the rostral ventromedial medulla and simultaneously recorded wide-dynamic-range neurons of the spinal dorsal horn in rats.

Considerable evidence supports the notion that on- and off-cells of the rostral ventromedial medulla (RVM) facilitate and depress, respectively, spinal nociceptive transmission. This notion stems from a covariation of on- or off-cell activities and spinal nocifensive reflexes. Such covariation could theoretically be due to their independently responding to a common source, or to an RVM-derived modulation of ventral horn neurons. Here, we tested whether on- and off-cells indeed modulate spinal nociceptive neurons. In deeply anesthetized rats, unitary recordings were simultaneously made from an RVM on-like or off-like cell and a spinal nociceptive neuron that shared a receptive field (RF) at a hind paw. Action potential firing in RVM/spinal neuron pairs was highly correlated, positively for on-like cells and negatively for off-like cells, both during ongoing activity and during application of calibrated noxious pressure to the RF. Microinjection of morphine into RVM induced a correlated decrease in on-like cell/spinal neuron ongoing activity and response to noxious stimulation. RVM morphine induced changes in off-like cell activity that were not correlated with spinal neuronal activity. These results suggest that on-cells exert a positive modulation upon spinal nociceptive neurons, upstream to ventral horn circuits and plausibly at the origin of nociceptive information that eventually reaches the cerebral cortex. On-cells may in this manner contribute to inflammation- and neuropathy-induced increases in withdrawal reflexes. Most significantly, on-cell modulation of nociceptive neurons may be a key factor in clinical pain conditions such as hyperalgesia and allodynia.

¿Siente dolor el feto?

Objetivo: Analizar una variedad de estudios publicados sobre el dolor fetal. Desde hace más de un siglo, investigadores clínicos y básicos han estudiado la posibilidad de que el feto sienta dolor. Este tema ha aumentado en relevancia debido al uso creciente de técnicas invasivas para diagnóstico y tratamiento fetal, y además por ser sustrato social de confrontación entre partidarios y adversarios del aborto. La pregunta fundamental es si el sistema nociceptivo del feto adquiere un desarrollo neurobiológico suficiente como para generar la experiencia mental que llamamos dolor. En el presente artículo, comenzamos con las etapas del desarrollo del sistema nociceptivo humano. Luego abordamos los aspectos éticos. Seguidamente discutimos si las manifestaciones conductuales, hormonales, hemodinámicas y electroencefalográficas del feto son evidencia de que ya tiene una mente y es capaz de sentir dolor. Conclusiones: No parece probable que el feto sienta dolor antes de la semana 20 de gestación; quizás pueda comenzar a sentir dolor entre las semanas 22 y 26. Sea cual sea la respuesta definitiva a la pregunta fundamental planteada más arriba, ante abordajes invasivos del feto, es recomendable usar analgesia o anestesia.

Objective: To analyze a variety of published studies on fetal pain. For a long time now, clinical and basic researchers have dealt with the question of whether the fetus is able to feel pain. The relevance of this subject has augmented in view of the increasing use of invasive techniques for fetal diagnosis and treatment, and also because of social strife between advocates and adversaries of abortion. The fundamental question here is whether the fetal nociceptive system has reached sufficient development for generating the mental experience that we call pain. We start with the developmental stages of the human nociceptive system. Then we approach the ethics of the problem. We further discuss whether the behavioral, hormonal, hemodynamic and electroencephalographic manifestations of a fetus are evidence for a mind capable of feeling pain. Conclusions: It seems unlikely that a fetus feels pain before week 20 of gestation; the fetus might be able to start feeling pain at weeks 22-26. We finally conclude that, whatever the answer to the fundamental question posed above, upon invasive approaches to the fetus the use of analgesia or anesthesia is recommended.

Enhanced spinal neuronal responses as a mechanism for the increased nociceptive sensitivity of interleukin-4 deficient mice.

Lack of the anti-inflammatory and analgesic cytokine interleukin-4 (IL-4) is associated with mechanical hypersensitivity in mice, and low systemic levels of IL-4 are associated with pain in humans. We investigated whether the firing properties of murine nociceptive neurons in the spinal dorsal horn are affected by IL-4 deficiency. Single unit recordings from lumbar dorsal horn wide-dynamic-range (WDR) neurons were performed in IL-4 knock out (ko) mice and wild type (WT) littermates (3, 9, and 22 months old). We measured neuronal responses to innocuous (1g) and noxious (26 g) von Frey mechanical stimulation at the plantar hind paw. Additionally, we induced secondary hyperalgesia by intraplantar injection of capsaicin and recorded discharges before and 5 and 10 min after injection. Baseline activity, activity upon innocuous stimulation, and postdischarges after noxious stimulation were not different between genotypes and ages. Responses to the noxious von Frey filament in 3 (34.5, 26.6-41.1 Hz) and 9 month old (49.7, 21.7-108.2 Hz) IL-4 ko mice were greater than in WT littermates (3 months, 18.1, 16.3-27.2 Hz, n.s.; 9 months, 33.6, 10.4-69.7 Hz; p<0.05). In contrast, 22 month IL-4 ko mice had lower discharges (22.4, 16.8-28.9 Hz) than 3 and 9 month IL-4 ko mice (p<0.01 each) and age-matched WT littermates (36.6, 10.4-59.4 Hz; n.s.). This pattern was also found 5 and 10 min after capsaicin injection. An enhanced excitability in the first segment of the nociceptive pathway may contribute to the increased behavioral responsiveness to painful stimuli of young IL-4 ko mice.

El dolor: los opioides: los anti-inflamatorios no esteroideos y los canabinoides / Pain: opioids: nonsteroidal antiinflammatory drugs and cannabinoids

Aquí exponemos un modelo que explica por qué, en el sistema nervioso central, los anti-inflamatorios no esteroideos, para ejercer su acción analgésica, deben interactuar con los opioides endógenos y los canabinoides endógenos. La sustancia gris del acueducto de Silvio es una estructura clave del llamado "sistema descendente de control nociceptivo". La activación de este sistema disminuye el flujo de mensajes nociceptivos hacia la corteza cerebral y, por lo tanto, el dolor. En la sustancia gris el ácido araquidónico es el elemento donde los opioides endógenos, los analgésicos opioides y los no-opioides (anti-inflamatorios no esteroideos) convergen para inducir analgesia. Las enzimas degradantes de los endocanabinoides son el punto donde estos y los analgésicos no-opioides convergen para inducir analgesia. Parece ventajoso el hecho de que los analgésicos que se compran libremente en la farmacia pueden aprovechar para su acción los mecanismos endógenos que todos nosotros poseemos

Here we present a model that explains why, in the central nervous system, the nonsteroidal antiinflammatory drugs, in order to induce analgesia, must interact with the endogenous opioids and the endocannabinoids. The periaqueductal gray matter is a key structure in the socalled "descending pain control system". Activations of this system diminishes the flow of nociceptive signals towards the cerebral cortex and, therefore, pain perception. In the periaqueductal gray matter, arachidonic acid is the elements where endogenous opioids analgesics and nonopioid analgesics converge to induce analgasia. The endocannabinoid metabolizing enzyme are the point at which endocannabinoids and nonsteroidal antinflammatory drugs converge to induce analgesia. There seems to be some advantage in that analgesics that can be bought over the counter can use for their action some endogenous mechanisms that we all possess

A aspirina, os opiáceos e a maconha no sistema endógeno de controle da dor / Aspirin, opiates and marijuana in the system of endogenous pain control

Tradução de Diego Molina e revisão do Prof. Dr. Luiz Roberto Giorgetti de Britto. O original em espanhol - "La aspirina, los opioides y la marijuana en el sistema endógeno de control del dolor" - encontra-se à disposição do leitor no Instituto de Estudos Avançados da USP para eventual consulta.

Preguntas de fisiología / Questions physiology

El presente manual es un compendio de preguntas de examen de fisiología normal para los temas de nervio, musculo, sinapsis, sistema nervioso central y sentidos especiales. Dentro de cada tema hay varios tipos de preguntas 1) las que tienen una (u) p dos (d) respuestas correctas entre cinco opciones posibles; 2) las de tipo hecho-razón (h), para indicar la veracidad de cada una de dos afirmaciones así como de la relación causal entre ellas; 3) las que requieren hacer una comparación (c) entre dos magnitudes; y 4) las que exigen al estudiante elaborar (e) una respuesta. Nuestro objetivo no es ayudar a los estudiantes a pasar en los exámenes, sino ayudarlos a aprender fisiología.

Spinal antinociceptive effects of cyclooxygenase inhibition during inflammation: Involvement of prostaglandins and endocannabinoids.

Both cyclooxygenase-1 and -2 are expressed in the spinal cord, and the spinal COX product prostaglandin E(2) (PGE(2)) contributes to the generation of central sensitization upon peripheral inflammation. Vice versa spinal COX inhibition is considered an important mechanism of antihyperalgesic pain treatment. Recently, however, COX-2 was shown to be also involved in the metabolism of endocannabinoids. Because endocannabinoids can have analgesic actions it is conceivable that inhibition of spinal COX produces analgesia not only by inhibition of PG synthesis but also by inhibition of endocannabinoid breakdown. In the present study, we recorded from spinal cord neurons with input from the inflamed knee joint and we measured the spinal release of PGE(2) and the endocannabinoid 2-arachidonoyl glycerol (2-AG) in vivo, using the same stimulation procedures. COX inhibitors were applied spinally. Selective COX-1, selective COX-2 and non-selective COX inhibitors attenuated the generation of spinal hyperexcitability when applied before and during development of inflammation but, when inflammation and spinal hyperexcitability were established, only selective COX-2 inhibitors reversed spinal hyperexcitability. During established inflammation all COX inhibitors reduced release of spinal PGE(2) almost equally but only the COX-2 inhibitor prevented breakdown of 2-AG. The reversal of spinal hyperexcitability by COX-2 inhibitors was prevented or partially reversed by AM-251, an antagonist at the cannabinoid-1 receptor. We conclude that inhibition of spinal COX-2 not only reduces PG production but also endocannabinoid breakdown and provide evidence that reversal of inflammation-evoked spinal hyperexcitability by COX-2 inhibitors is more related to endocannabinoidergic mechanisms than to inhibition of spinal PG synthesis.

NSAIDs, Opioids, Cannabinoids and the Control of Pain by the Central Nervous System.

Nonsteroidal anti-inflammatory drugs (NSAIDs) act upon peripheral tissues and upon the central nervous system to produce analgesia. A major central target of NSAIDs is the descending pain control system. The rostral structures of the descending pain control system send impulses towards the spinal cord and regulate the transmission of pain messages. Key structures of the descending pain control system are the periaqueductal gray matter (PAG) and the rostral ventromedial region of the medulla (RVM), both of which are critical targets for endogenous opioids and opiate pharmaceuticals. NSAIDs also act upon PAG and RVM to produce analgesia and, if repeatedly administered, induce tolerance to themselves and cross-tolerance to opioids. Experimental evidence shows that this is due to an interaction of NSAIDs with endogenous opioids along the descending pain control system. Analgesia by NSAIDs along the descending pain control system also requires an activation of the CB1 endocannabinoid receptor. Several experimental approaches suggest that opioids, NSAIDs and cannabinoids in PAG and RVM cooperate to decrease GABAergic inhibition and thus enhance the descending flow of impulses that inhibit pain.

Spinal endocannabinoids and CB1 receptors mediate C-fiber-induced heterosynaptic pain sensitization.

Diminished synaptic inhibition in the spinal dorsal horn is a major contributor to chronic pain. Pathways that reduce synaptic inhibition in inflammatory and neuropathic pain states have been identified, but central hyperalgesia and diminished dorsal horn synaptic inhibition also occur in the absence of inflammation or neuropathy, solely triggered by intense nociceptive (C-fiber) input to the spinal dorsal horn. We found that endocannabinoids, produced upon strong nociceptive stimulation, activated type 1 cannabinoid (CB1) receptors on inhibitory dorsal horn neurons to reduce the synaptic release of gamma-aminobutyric acid and glycine and thus rendered nociceptive neurons excitable by nonpainful stimuli. Our results suggest that spinal endocannabinoids and CB1 receptors on inhibitory dorsal horn interneurons act as mediators of heterosynaptic pain sensitization and play an unexpected role in dorsal horn pain-controlling circuits.

Joint pain.

Both inflammatory and degenerative diseases of joints are major causes of chronic pain. This overview addresses the clinical problem of joint pain, the nociceptive system of the joint, the mechanisms of peripheral and central sensitization during joint inflammation and long term changes during chronic joint inflammation. While the nature of inflammatory pain is obvious the nature and site of origin of osteoarthritic pain is less clear. However, in both pathological conditions mechanical hyperalgesia is the major pain problem, and indeed, both joint nociceptors and spinal nociceptive neurons with joint input show pronounced sensitization for mechanical stimulation. Molecular mechanisms of mechanical sensitization of joint nociceptors are addressed with an emphasis on cytokines, and molecular mechanisms of central sensitization include data on the role of excitatory amino acids, neuropeptides and spinal prostaglandins. The overview will also address long-term changes of pain-related behavior, response properties of neurons and receptor expression in chronic animal models of arthritis.

Tolerance to non-opioid analgesics in PAG involves unresponsiveness of medullary pain-modulating neurons in male rats.

Opiate analgesia can be hampered by a reduction in pharmacological effectiveness (tolerance), and this crucially depends on the periaqueductal gray matter (PAG). Non-opioids like metamizol (dipyrone) or aspirin also induce PAG-dependent analgesia and tolerance, but the neuronal bases of this tolerance are unknown. Metamizol is a pyrazolon derivative and cyclooxygenase inhibitor with widespread use as an analgesic in Europe and Latin America. Metamizol was microinjected into the PAG of awake male rats, and antinociception was assessed by the tail flick (TF) and hot plate (HP) tests. Microinjection twice daily for 2.5 days caused tolerance to metamizol. The rats were then anesthetized and recordings from pain-facilitating on-cells and pain-inhibiting off-cells of the rostral ventromedial medulla (RVM) were performed. PAG microinjection of morphine or metamizol depresses on-cells, activates off-cells and thus inhibits nociception, including TF and HP. In metamizol-tolerant rats, however, PAG microinjection of metamizol failed to affect on- or off-cells, and this is interpreted as the reason for tolerance. In metamizol-tolerant rats morphine microinjection into PAG also failed to affect RVM neurons or nociception (cross-tolerance). In naïve, non-tolerant rats the antinociceptive effect of PAG-microinjected metamizol or morphine was blocked when CTOP, a mu-opioid antagonist, was previously microinjected into the same PAG site. These results emphasize a close relationship between opioid and non-opioid analgesic mechanisms in the PAG and show that, like morphine, tolerance to metamizol involves a failure of on- and off-cells to, respectively, disfacilitate and inhibit nociception. Cross-tolerance between non-opioid and opioid analgesics should be important in the clinical setting.

Critical role of the rostral ventromedial medulla in early spinal events leading to chronic constriction injury neuropathy in rats.

UNLABELLED: Neuropathic pain is a major clinical problem, and several animal models have been developed to investigate its mechanisms and its treatment. In this report, the role of the rostral ventromedial medulla (RVM) in the early events of the chronic constriction injury (CCI) model was investigated in behavioral and electrophysiological experiments. Placing the 4 CCI ligatures around the sciatic nerve induced large discharges and residual ongoing activity in spinal nociceptive neurons. Two weeks after CCI ligation, the rats showed behavioral hyperalgesia and allodynia as well as increased ongoing activity and responsiveness of spinal nociceptive neurons to innocuous and noxious stimuli. Blockade of excitatory synapses in the RVM by a kynurenate microinjection (2 nmol in 0.5 muL) 5 minutes before placement of the sciatic ligatures had no immediate effect on spinal neuronal activity but largely prevented the activation of spinal neurons. In kynurenate microinjected rats, behavioral hyperalgesia and allodynia developed slowly and incompletely, which corresponded with an incompletely developed hyperexcitability of spinal neurons. To the best of our knowledge, these results show for the first time that the initial response to nerve damage requires facilitation from the RVM. PERSPECTIVE: The present and previous findings indicate that descending facilitation from brainstem nuclei critically contributes to the spinal hyperexcitability that underlies neuropathic pain. The present results indicate that this contribution begins at the very moment the nerve is damaged and should be prevented and treated accordingly.

A nonopioid analgesic acts upon the PAG-RVM axis to reverse inflammatory hyperalgesia.

Metamizol (dipyrone) and other nonsteroidal anti-inflammatory drugs (NSAIDs) induce antinociception by acting upon peripheral tissues and upon central nervous system structures, notably the periaqueductal grey matter (PAG) and the spinal cord. Inflammation-induced hyperalgesia is prevented by spinal application of NSAIDs before the inflammation, but once central sensitization is established the spinal effect of NSAIDs is uncertain. The present study examines whether the action upon the PAG contributes to the attenuation of inflammation-induced spinal hyperalgesia by NSAIDs. In deeply anaesthetized rats, responses of spinal multireceptive neurons to mechanical stimulation of the ipsilateral paw and leg were recorded. An inflammation in the paw was induced with carrageenan. Fifty minutes later, neuronal responses to innocuous and noxious stimulation had, respectively, increased to 206 and 304% for paw, and 160 and 190% for leg. When metamizol (150 microg in 0.5 microL) was microinjected into PAG before the inflammation, neuronal hyperexcitability was delayed for approximately 60 min and was much reduced by 215 min. More interestingly, microinjection of metamizol into PAG when hyperexcitability was fully developed depressed neuronal responses down to baseline for approximately 1 h. The effect of PAG metamizol was reversed by microinjection of a GABA(A) agonist into the rostral ventromedial medulla (RVM), which indicates that RVM relays the metamizol effect from PAG onto the spinal cord. These results suggest that, upon clinical administration of NSAIDs, a joint action upon PAG and spinal cord contributes to preventing the development of hyperalgesia but it is mainly the action upon PAG which contributes to reducing fully established hyperalgesia.

Antinociception induced by intravenous dipyrone (metamizol) upon dorsal horn neurons: involvement of endogenous opioids at the periaqueductal gray matter, the nucleus raphe magnus, and the spinal cord in rats.

Microinjection of dipyrone (metamizol) into the periaqueductal gray matter (PAG) in rats causes antinociception. This is mediated by endogenous opioidergic circuits located in the PAG itself, in the nucleus raphe magnus and adjacent structures, and in the spinal cord. The clinical relevance of these findings, however, is unclear. Therefore, in the present study, dipyrone was administered intravenously, and the involvement of endogenous opioidergic circuits in the so-induced antinociception was investigated. In rats, responses of dorsal spinal wide-dynamic range neurons to mechanical noxious stimulation of a hindpaw were strongly inhibited by intravenous dipyrone (200 mg/kg). This effect was abolished by microinjection of naloxone (0.5 microg/0.5 microl) into the ventrolateral and lateral PAG or into the nucleus raphe magnus or by direct application of naloxone (50 microg/50 microl) onto the spinal cord surface above the recorded neuron. These results show that dipyrone, a non-opioid analgesic with widespread use in Europe and Latin America, when administered in a clinically relevant fashion causes antinociception by activating endogenous opioidergic circuits along the descending pain control system.

Descending control of persistent pain: inhibitory or facilitatory?

The periaqueductal gray matter (PAG) and the nucleus raphe magnus and adjacent structures of the rostral ventromedial medulla (RVM), with their projections to the spinal dorsal horn, constitute the "efferent channel" of a pain-control system that "descends" from the brain onto the spinal cord. Considerable evidence has recently emerged regarding participation of this system in persistent pain conditions such as inflammation and neuropathy. Herein, this evidence is reviewed and organized to support the idea that persistent nociception simultaneously triggers descending facilitation and inhibition. In models of inflammation, descending inhibition predominates over facilitation in pain circuits with input from the inflamed tissue, and thus attenuates primary hyperalgesia, while descending facilitation predominates over inhibition in pain circuits with input from neighboring tissues, and thus facilitates secondary hyperalgesia. Both descending facilitation and inhibition mainly stem from RVM. The formalin-induced primary hyperalgesia, although considered a model for inflammation, is mainly facilitated from RVM. Also, formalin-induced secondary hyperalgesia is facilitated by RVM. Again, formalin triggers a concomitant but concealed descending inhibition. The (primary) hyperalgesia and allodynia of the neuropathic syndrome are also facilitated from RVM. Simultaneously, there is an inhibition of secondary neuronal pools that is partly supported from the PAG. Because in all these models of peripheral damage descending facilitation and inhibition are triggered simultaneously, it will be important to elucidate why inhibition predominates in some neuronal pools and facilitation in others. Therapies that enhance descending inhibition and/or attenuate descending facilitation are furthermore an important target for research in the future.

Involvement of cholecystokinin in the opioid tolerance induced by dipyrone (metamizol) microinjections into the periaqueductal gray matter of rats.

The analgesic effect of non-steroidal anti-inflammatory drugs (NSAIDs) is partly due to an action upon the periaqueductal gray matter (PAG), which triggers the descending pain control system and thus inhibits nociceptive transmission. This action of NSAIDs engages endogenous opioids at the PAG, the nucleus raphe magnus and the spinal cord. Repeated administration of NSAIDs such as dipyrone (metamizol) and acetylsalicylate thus induces tolerance to these compounds and cross-tolerance to morphine. Since cholecystokinin plays a key role in opioid tolerance, the present study in rats investigated whether PAG cholecystokinin is also responsible for tolerance to PAG-microinjected dipyrone. Microinjection of cholecystokinin (1 ng/0.5 microl) into PAG blocked the antinociceptive effect of a subsequent microinjection of dipyrone (150 microg/0.5 microl) into the same site, as evaluated by the tail flick and hot plate tests. Microinjection of proglumide (0.4 microg/0.5 microl), a non-selective cholecystokinin antagonist, into PAG prevented the development of tolerance to subsequent microinjections of dipyrone, as well as cross-tolerance to microinjection of morphine (5 microg/0.5 microl) into the same site. In rats tolerant to PAG dipyrone, a PAG microinjection of proglumide restored the antinociceptive effect of a subsequent microinjection of dipyrone or morphine. These results suggest that PAG-microinjected dipyrone triggers and/or potentiates local opioidergic circuits leading to descending inhibition of nociception, on the one hand, and to a local antiopioid action by cholecystokinin, on the other. Reiteration of these events would then result in an enhancement of cholecystokinin's antiopioid action and thus tolerance to opioids and dipyrone in the PAG.

Induction of opioid tolerance by lysine-acetylsalicylate in rats.

The analgesic effect of non-steroidal antiinflammatory drugs (NSAIDs) is due to their action upon the peripheral damaged tissues, the spinal cord, and brain stem structures of the 'descending pain-control system' such as the periaqueductal gray matter (PAG) and the nucleus raphe magnus (NRM). The NSAID dipyrone (metamizol) has been shown to engage opioidergic circuits at the PAG, the NRM and the spinal cord, but it is unknown whether this can be generalized to typical NSAIDs and to systemic administration. In the present study lysine-acetylsalicylate (LASA), an injectable form of the prototypical NSAID aspirin, was microinjected into the PAG (100 microg/0.5 microl) in freely moving rats to induce inhibition of tail flick and hot plate responses. This antinociception was reverted by naloxone (1 mg/kg i.p.). PAG microinjection of LASA twice daily for three days induced tolerance to LASA (i.e. a progressive loss of effectiveness) and cross-tolerance to PAG-microinjected morphine (5 microg/0.5 microl). The antinociceptive effect of systemically administered LASA (300 mg/kg i.p., equivalent to the 1000 mg analgesic dose for humans) was also abolished by naloxone. Intraperitoneal injection of LASA twice daily induced tolerance to LASA and cross-tolerance to i.p. morphine (1 or 5 mg/kg). LASA-tolerant rats showed opioid withdrawal signs when injected with naloxone. These findings support the notion that the contribution of the PAG and downstream pain-control structures to the analgesic effect of NSAIDs involves opioidergic mechanisms, and suggest that repeated therapeutic administration of NSAIDs may induce tolerance, cross-tolerance to opiates, and susceptibility to a withdrawal syndrome.

To the descending pain-control system in rats, inflammation-induced primary and secondary hyperalgesia are two different things.

The periaqueductal gray matter and the rostral ventromedial medulla (RVM), with its projections to the spinal dorsal horn, constitute the efferent channel of the 'descending pain-control system'. Noxious stimulation of a peripheral tissue causes more pain if this tissue is inflamed (primary hyperalgesia). In such cases, stimulation of neighboring but uninflamed tissues also becomes more painful (secondary hyperalgesia). In animal models of inflammation, the descending pain-control system sends down, simultaneously, inhibitory and facilitatory influences, but inhibition predominates for primary hyperalgesia while facilitation predominates for secondary hyperalgesia. Descending inhibition and facilitation during peripheral inflammation are due not only to previously existing descending modulation, but also to inflammation-induced changes in RVM which involve receptors for NMDA, AMPA, cholecystokinin and neurotensin, as well as synthesis of enkephalins and nitric oxide.

Involvement of local cholecystokinin in the tolerance induced by morphine microinjections into the periaqueductal gray of rats.

The ventrolateral periaqueductal gray (PAG) is a key structure for the development of opioid tolerance. An increased activity of 'anti-opioids' like cholecystokinin (CCK) has been proposed as a possible mechanism for opioid tolerance. The present study evaluates the role of PAG-located CCK in the opioid tolerance induced by repeated microinjections of morphine (MOR) into PAG. Male rats were implanted with chronic guide cannulae aimed at the PAG. Microinjection of MOR (0.5 microg in 0.5 microl) into PAG caused antinociception as quantified with the tail flick and the hot plate tests. When MOR microinjection was repeated twice daily, the antinociceptive effect disappeared within 2 days (tolerance). However, if each MOR microinjection was preceded (within 15 min) by a microinjection of the non-selective CCK receptor antagonist proglumide (PRO), (0.4 microg in 0.5 microl) into the same PAG site, the microinjections of MOR always produced antinociception and did not induce tolerance. If PRO microinjections were suspended, subsequent MOR microinjections induced tolerance. In MOR-tolerant rats, a single PRO microinjection into the same PAG site was enough to restore the antinociceptive effect of MOR. On the other hand, if CCK (1 ng in 0.5 microl) was microinjected into PAG, then MOR microinjection administered 15 min later into the same PAG site did not elicit antinociception. These results show that CCK has anti-opioid activity in PAG and that tolerance to MOR in PAG can be prevented or reversed if CCK receptors are blocked with PRO. Finally, opioid tolerance induced by repeated systemic MOR injections (5mg/kg intraperitoneal ) was reversed by a single microinjection of PRO into PAG. This emphasizes the central importance of PAG in the MOR/CCK interactions that lead to opioid tolerance.