Opioid signaling in mast cells regulates injury responses associated with heterotopic ossification.

INTRODUCTION: Previous studies found that neuron specific enolase promoter (Nse-BMP4) transgenic mice have increased expression of the nociceptive mediator, substance P and exaggerated local injury responses associated with heterotopic ossification (HO). It is of interest great to know the pain responses in these mice and how the opioid signaling is involved in the downstream events such as mast cell (MC) activation. MATERIALS AND METHODS: This study utilized a transgenic mouse model of HO in which BMP4 is expressed under the control of the Nse-BMP4. The tactile sensitivity and the cold sensitivity of the mice were measured in a classic inflammatory pain model (carrageenan solution injected into the plantar surface of the left hind paw). The MC activation and the expression profiles of different components in the opioid signaling were demonstrated through routine histology and immunohistochemistry and Western blotting, in the superficial and deep muscle injury models. RESULTS: We found that the pain responses in these mice were paradoxically attenuated or unchanged, and we also found increased expression of both Methionine Enkephalin (Met-Enk), and the µ-opioid receptor (MOR). Met-Enk and MOR both co-localized within activated MCs in limb tissues. Further, Nse-BMP4;MOR(-/-) double mutant mice showed attenuated MC activation and had a significant reduction in HO formation in response to injuries. CONCLUSIONS: These observations suggest that opioid signaling may play a key role in MC activation and the downstream inflammatory responses associated with HO. In addition to providing insight into the role of MC activation and associated injury responses in HO, these findings suggest opioid signaling as a potential therapeutic target in HO and possibly others disorders involving MC activation.

Inhibiting roles of berberine in gut movement of rodents are related to activation of the endogenous opioid system.

Although Berberine (BER) is popular in treating gastrointestinal (GI) disorders, its mechanisms are not clear yet. In order to investigate the effects and possible mechanism of BER on GI motility in rodents, we first explored GI motility by recording the myoelectrical activity of jejunum and colon in rats, and upper GI transit with a charcoal marker in mice. Then, the plasma levels of gastrin, motilin, somatostatin and glucagon-like-peptide-1 (Glp-1) were measured by ELISA or radioimmunoassay (RIA). Furthermore, endogenous opioid-peptides (ß-endorphin, dynorphin-A, met-enkephalin) were detected by RIA after treatment with BER. Our results showed that BER concentration-dependently inhibited myoelectrical activity and GI transit, which can be antagonized by opioid-receptor antagonists to different extents. The elevated somatostatin and Glp-1, and decreased gastrin and motilin in plasma, which were caused by BER application, also could be antagonized by the opioid-receptor antagonists. Additionally, plasma level of ß-endorphin, but not dynorphin-A and met-enkephalin, was increased by applying BER. Taken together, these studies show that BER plays inhibiting roles on GI motility and up-regulating roles on somatostatin, Glp-1 and down-regulating roles on gastrin, motilin. The pharmacological mechanisms of BER on GI motility and plasma levels of GI hormones were discovered to be closely related to endogenous opioid system.

Autocrine regulation of human sperm motility by the met-enkephalin opioid peptide.

OBJECTIVE: To verify the presence of protein precursor pro-enkephalin (PENK) and met-enkephalin in human spermatozoa and to characterize the effects of exogenous and endogenous enkephalins on sperm motility. DESIGN: We carried out expression assays for met-enkephalin and its protein precursor PENK by reverse transcription-polymerase chain reaction (RT-PCR), Western blot, and immunofluorescence techniques in sperm cells and motility analysis after incubation of semen samples with met-enkephlin enzyme inhibitors and the opioid receptor antagonist naloxone. Met-enkephalin secretion was analyzed by flow cytometry. SETTING: Assisted reproduction unit and academic research laboratory. PATIENT(S): Semen from 50 normozoospermic healthy human donors. INTERVENTION(S): Spermatozoa isolated from semen on discontinuous Percoll gradient (40%-80%) followed by a swim-up was used for all techniques. MAIN OUTCOME MEASURE(S): Immunoblotting blots, indirect immunofluorescence antibody assays, RT-PCR blots, flow cytometry plots, and percentage of motile sperm. RESULT(S): We found by RT-PCR and immunofluorescence that met-enkephalin and its protein precursor PENK are present in the head of human sperm cells. Endogenous met-enkephalin increased sperm motility, whereas the addition of exogenous met-enkephalin had a biphasic effect on motility, likely due to the activation of distinct receptor subtypes. CONCLUSION(S): We provide evidence for a new role of met-enkephalin as an endogenous mediator of sperm motility. This autocrine regulation of sperm function by the opioid system represents a new mechanism of regulation of male factor fertility and could be useful as an emerging target for male contraception.

The opioid growth factor-opioid growth factor receptor axis: homeostatic regulator of cell proliferation and its implications for health and disease.

The opioid growth factor (OGF), chemically termed [Met(5)]-enkephalin, is an endogenous opioid peptide that interacts with the OGF receptor (OGFr) to delay the G(1)/S interface of the cell cycle by modulating cyclin-dependent inhibitory kinase (CKI) pathways. The OGF-OGFr axis is a tonically active, inhibitory pathway that is an important regulator during homeostasis and re-epithelialization, and plays a role in the onset and progression of autoimmune diseases and cancer. Modulation of the OGF-OGFr axis can be accomplished by a variety of pharmacological and molecular approaches including use of intermittent or continuous exposure to the opioid antagonist naltrexone, genetic manipulation of OGFr expression, and antibody neutralization of OGF. Clinically, OGF is a biological therapy that has potential application for treatment of cancer. Currently, naltrexone at low dosages is being evaluated for treatment of autoimmune diseases such as Crohn's and multiple sclerosis. High dosages of naltrexone are effective in reversing dry eye and accelerating the repair of corneal abrasions in normal and diabetic rats; these studies are under investigation in the clinical setting. Naltrexone also enhances full-thickness wound closure in animal models of Type 1 or Type 2 diabetes, and translation of this knowledge to the clinic is planned. In summary, understanding the OGF-OGFr axis as a homeostatic regulator of proliferation has substantial implications for maintaining human health and treatment of disease.

Endogenous endomorphin-2 contributes to spinal ĸ-opioid antinociception.

BACKGROUND/AIMS: Multiple opioid receptor (OR) types and endogenous opioid peptides exist in the spinal dorsal horn and there may be interactions among these receptor types that involve opioid peptides. In a previous study we observed that antinociceptive effects of the selective κ-opioid receptor (κOR) agonist, U50,488H, was attenuated in µ-opioid receptor (µOR) knockout mice as compared to wild-type mice when administered spinally. This suggests that an interaction between κORs and µORs exits in the spinal cord. The present study was aimed at investigating whether endogenous opioid peptides were involved in such interaction. METHODS: We examined whether the presence of antibodies to endogenous opioid peptides, endomorphin-2, met-enkephalin and dynorphin A affected the antinociceptive effects of spinal U50,488H in rats. The tail-flick test was used to assess pain thresholds. RESULTS: The increase in tail-flick latency after spinal U50,488H was attenuated when the rats were pretreated intrathecally with antiserum against endomorphin-2. Pretreatments with antisera against met-enkephalin and dynorphin A had no effect on U50,488H antinociception. The antisera alone did not affect pain threshold. CONCLUSION: The results suggest that endomorphin-2, an endogenous opioid peptide highly selective to the µOR, plays a role in antinociception induced by κOR activation in the spinal cord.

Plasma endogenous enkephalin levels in early systemic sclerosis: clinical and laboratory associations.

OBJECTIVE: Met- and leu-enkephalins are endogenous opioid neuropeptides with potent analgesic, vasoactive, immunomodulatory and anti-apoptotic properties. We hypothesised that clinical or immunological variables of early systemic sclerosis (SSc) might be correlated to plasma enkephalin levels. METHODS: Plasma samples were collected at study entry of the Genetics versus Environment in Scleroderma Outcomes Study (GENISOS) cohort (early SSc, n=116). Plasma met-enkephalin and leu-enkephalin levels (microg/ml) were measured by high performance liquid chromatography (HPLC) and correlated to clinical and laboratory parameters in the GENISOS database. Statistical analyses were performed by nonparametric Wilcoxon rank sum tests and Pearson correlation coefficients. RESULTS: Significantly lower plasma met-enkephalin levels were associated with anti-topoisomerase-I seropositivity (6+8.3 vs. 14.9+22.8 microg/ml, p=0.02). Plasma leu-enkephalin levels were significantly higher in SSc patients with digital pulp loss (95.6+130 vs. 64.9+101 microg/ml, p=0.02). Lower mean plasma met-enkephalin levels and inversely higher leu-enkephalin levels were noted in SSc patients with Raynaud's phenomena (p=NS). CONCLUSION: The associations of plasma enkephalin levels to immunologic or clinical pathologies may underscore their vasogenic or fibrogenic significance and potential as therapeutic targets in early SSc.

Regulation of Tenon's capsule fibroblast cell proliferation by the opioid growth factor and the opioid growth factor receptor axis.

PURPOSE: Glaucoma filtration surgery often fails because of the fibrotic reaction from Tenon's capsule fibroblasts (TCFs). This study examined whether the interaction of the opioid growth factor (OGF) [Met(5)]-enkephalin with its receptor (OGFr) is a regulator of TCF proliferation. METHODS: The presence of OGF and its receptor (OGFr) was determined in rabbit TCFs (RTCFs) by immunocytochemistry. The kinetics of OGFr were established in receptor binding assays. The ability of OGF to inhibit RTCF proliferation was assessed with dose-response, receptor mediation, and reversibility studies. Dependence on OGF and OGFr was ascertained by antibody neutralization and siRNA studies, respectively. The mechanism of action of the OGF-OGFr axis on survival (apoptosis, necrosis) and DNA synthesis of RTCFs was elucidated. RESULTS: OGF and OGFr were detected in RTCF cells, and specific and saturable binding to OGFr was recorded. Exogenous OGF had a dose-dependent, reversible, and receptor-mediated inhibitory effect on cell proliferation. Endogenous OGF was found to be constitutively produced and tonically active in cell replication, with neutralization of this peptide causing acceleration of cell proliferation. The silencing of OGFr by using siRNA technology stimulated cell replication, validating OGFr's integral role. The mechanism of OGF-OGFr action was not related to cell survival, but rather to DNA synthesis-specifically, the cyclin-dependent kinase inhibitory pathway. Knockdown of p16 or p21 eliminated OGF's inhibitory effect on growth. CONCLUSIONS: The OGF-OGFr system is a native biological regulator of cell proliferation in RTCFs and may offer a means of improving the success of glaucoma filtration surgery in a safe and nontoxic manner.

[Distribution of met-enkephalin expression in the visual pathway. Experimental study by inmunocytochemistry]. / Distribución de la metencefalina en la vía óptica. Estudio experimental inmunocitoquímico.

PURPOSE: The localization and distribution of neuropeptide expression in the cat visual pathway can provide information about the function of that pathway. METHOD: Study of optic pathway in eight cats. Following extraction of the brain, slices were prepared using a microkeratome. The slices were examined by indirect immunocytochemistry using anti-metenkephalin as antibody to determine the presence or absence of this pentapeptide in the visual pathway. RESULTS: Met-enkephalin receptors in both cortical and subcortical regions of the brain were detected. This suggests that met-enkephalin could be involved in the visual mechanism. CONCLUSIONS: The presence of met-enkephalin receptors in both cortical and subcortical regions of the brain suggests that this pentapeptide could be involved in the visual mechanism.

Two distinct mechanisms mediate acute mu-opioid receptor desensitization in native neurons.

Sustained stimulation of G-protein coupled receptors (GPCRs) leads to rapid loss of receptor function (acute desensitization). For many GPCRs including the mu-opioid receptor (MOR), an accepted mechanism for acute desensitization is through G-protein coupled receptor kinase (GRKs) mediated phosphorylation of the receptor, which facilitates the binding of beta-arrestins (betaarrs) to the receptor and then promotes endocytosis. However, the mechanism(s) that mediate acute desensitization have not yet been well defined in native neurons. This study used whole-cell patch clamp recording of G-protein coupled inward-rectifying potassium (GIRK) currents to assay MOR function and identify mechanisms of acute MOR desensitization in locus ceruleus (LC) neurons. The rate and extent of MOR desensitization were unaffected by beta(arr)-2 knock-out. Disruption of GRK2 function via inhibitory peptide introduced directly into neurons also failed to affect desensitization in wild type or beta(arr)-2 knock-outs. Inhibition of ERK1/2 activation alone had little effect on acute desensitization. However, when both GRK2-beta(arr)-2 and ERK1/2 functions were disrupted simultaneously, desensitization of MOR was nearly abolished. Together, these results suggest that acute desensitization of MOR in native LC neurons is determined by at least two molecular pathways, one involving GRK2 and beta(arr)2, and a parallel pathway mediated by activated ERK1/2.

Aspects of central and peripheral regulation of reproduction in mammals.

To increase our knowledge concerning the central and peripheral regulation of reproduction in mammals a series of studies were performed. In the first experiment, we found that exogenous leptin altered the activity of the hypothalmo-pituitary-gonadotropic axis in sheep during insufficient feeding. The action of leptin appears to be mediated by changes in GnRH and LH secretion as well as NPY immunoreactivity. The aim of the second experiment was to investigate the role of the adipoinsular axis hormones during pregnancy in rats. The elevated levels of plasma leptin as wells as the increased mRNAs expression of the leptin receptors in placenta indicate the significant role of the hormone in fetal growth and development. On the other hand, a decrease in leptin receptors mRNA content within hypothalamus and pituitary together with unchanged plasma insulin level may suggest that during rat pregnancy leptin resistance was developed in the hypothalamus, pituitary and pancreatic islets. The third experiment was carried out to establish the role of opioids and glucocorticoids in the regulation of the hypothalmo-pituitary-gonadal axis in ewes during natural or synchronized estrous cycle. Prolonged treatment with progesterone resulted in significant changes in plasma levels of Met-enkephalin, cortisol and steroids and altered the expression of proenkephalin mRNA in the hypothalamus, pituitary, ovary and adrenals. Injections of Met-enkephalin or naltrexone (blocker of opioid receptors) modulated the progesterone influence in tested tissues. The data clearly suggest that opioids are involved in the regulation of the estrous cycle at the hypothalamo-pituitary-gonadal/adrenal axes.

Endogenous signaling complexity in neuropeptides- leucine- and methionine-enkephalin.

1. In addition to his many fine contributions in furthering our understanding of the neurochemical action of ecosanoids, catchelomines, steroids, anandamines, cannabinoids, endorphins, and the many modifications made to these neural factors, twenty years ago Julius Axelrod published a noteworthy paper concerning the nature of neuropeptides and their potential for multiple neurophysiological effects (Redgate et al., 1986). 2. In that report, Axelrod and coworkers described the neurological actions of the then recently discovered leucine- and methionine-enkephalins, and their biological functions which were novel, atypical, and in possession of neurological effects that were significantly "much more than additive." 3. In this short communication I would like to expand on this observation concerning the "additive effects" contained within the amino acid sequence of the atypical neurotransmitter peptides leucine- and methonine-enkephalin.

Chimeric peptide of met-enkephalin and FMRFa: effect of chlorination on conformation and analgesia.

In our previous study YFa (YGGFMKKKFMRFa), a chimeric peptide of met-enkephalin and FMRFa, not only produced analgesia but also did not let the tolerance develop. In the continuation of the same study, Phe4 is chlorinated so as to assess the effect of chlorination on the conformation, lipophilicity and analgesia of chimeric peptide [p-Cl Phe(4)] YFa. Not only does the chlorination increase the lipophilicity but also enhances the propensity of [p-Cl Phe(4)] YFa to form alpha helix in comparison of YFa in presence of membrane mimicking solvent trifluoroethanol (TFE). This increase in lipophilicity and helix-forming ability results in more bioavailability and naloxone-reversible analgesia by [p-Cl Phe(4)] YFa. Though analgesia produced by [p-Cl Phe(4)] YFa is more than YFa at all doses, there is sudden decrease in analgesia at 45 and 60 min at 60 mg/kg. This sudden decrease of analgesia seems to be due to desensitization of opioid receptors.

Role of methionine-enkephalin on the regulation of carbohydrate metabolism in the rice field crab Oziotelphusa senex senex.

In the present study, the role of eyestalks and involvement of methionine-enkephalin in the regulation of haemolymph sugar level was studied. Bilateral eyestalk ablation significantly decreased the haemolymph sugar levels, whereas injection of eyestalk extract into ablated crabs significantly increased the haemolymph sugar levels. Total carbohydrate (TCHO) and glycogen levels were significantly increased in hepatopancreas and muscle of eyestalk-ablated crabs, with a decrease in phosphorylase activity. Injection of eyestalk extract into ablated crabs resulted in partial/complete reversal of these changes. Injection of methionine-enkephalin into intact crabs significantly increased the haemolymph sugar level in a dose-dependent manner. Total tissue carbohydrate and glycogen levels were significantly decreased, with an increase in phosphorylase activity in hepatopancreas and muscle tissues of intact crabs after methionine-enkephalin injection. Methionine-enkephalin injection did not cause any changes in haemolymph sugar, tissue total carbohydrate and glycogen levels and activity levels of phosphorylase in eyestalk-ablated crabs. These results suggest that the eyestalks are the main source of hyperglycaemic hormone and methionine-enkephalin induces hyperglycaemia through eyestalks.

Met-enkephalin involvement in morphine-modulated peritonitis in swiss mice.

Morphine coinjection with zymosan inhibits pain and leukocyte accumulation during peritonitis in several strains of mice, and affects systems of endogenous opioids. Present investigations focus on Met-enkephalin (Met-ENK) in the inflamed peritoneal cavity and brain centers of Swiss mice. Males of Swiss mice were IP injected with zymosan or zymosan supplemented with morphine. At the selected time points the peritoneal leukocytes were counted and the Met-ENK level was measured in exudatory fluid and leukocytes, striatum, hypothalamus, and pituitary gland. The Met-ENK content in peritoneal fluid rised sharply after zymosan injection, which corresponded with its decline in exudatory leukocytes, hypothalamus, and striatum. Morphine coinjection enhanced intraperitoneal accumulation of Met-ENK and its release from exudatory leukocytes, but inhibited its early fluctuations in hypothalamus and striatum. Effects of morphine-modulated inflammation on the Met-ENK system lasted longer than 7 days.

Particle-mediated gene transfer of opioid growth factor receptor cDNA regulates cell proliferation of the corneal epithelium.

PURPOSE: This study was designed to determine at the molecular level whether interactions between the opioid growth factor (OGF) and OGF receptor (OGFr) play a role in regulating DNA synthesis in the homeostasis of the corneal epithelium. METHODS: The plasmid pcDNA3.1+OGFr-HA, carrying the rat OGFr cDNA epitope-tagged with a C-terminal hemagglutinin (HA), or the empty-vector (pcDNA3.1+), was delivered twice by the Helios Gene Gun System at 300 psi to the cornea of anesthetized rats. The contralateral (untreated) cornea served as the naive specimen. BrdU was used to determine whether the recombinant OGFr was effective in regulating DNA synthesis in the rat peripheral corneal epithelium. RESULTS: Within 18 hours of transfection, positive HA staining was apparent in both the basal and suprabasal layers (efficiency > 90% of the cells) throughout the central and peripheral cornea. Quantitative immunohistochemistry with rhodamine-conjugated anti-OGFr antibodies revealed twofold more OGFr expression in the central and peripheral epithelium of transfected corneas relative to naive corneas. The number of BrdU-positive basal cells in the peripheral epithelium of the transfected cornea was one-third of that in the naive cornea. CONCLUSIONS: These data demonstrate the direct role of the OGF-OGFr system in determining cellular renewal in the mammalian corneal epithelium. Moreover, the successful establishment of a novel delivery system of cDNAs to the ocular surface suggests a therapeutic role for gene therapy in the eye.

Effects of met-enkephalin on cell proliferation in different models of adrenocortical-cell growth.

Met-enkephalin (met-Enk) is an opioid peptide that acts via three main subtypes of receptors referred to as mu (mu)-, delta (delta)- and zeta (zeta)-receptor. While the first two receptor subtypes mediate the classic opioid effects of met-Enk, zeta-receptors are reported to be involved in the non-opioid actions of the peptide, i.e. the inhibitory effect on the cell growth. Despite the fact that met-Enk is known to regulate the function of the hypothalamic-pituitary-adrenal axis acting on both its central and peripheral branches, none is known on the effects of met-Enk on adrenal growth. Hence, we have investigated the effects of met-Enk and its receptor agonists and antagonists on cell proliferation in three different models of rat adrenal growth: i) immature adrenal cortex, ii) regenerating adrenal cortex and iii) primary cultures of adrenocortical cells. In in vivo experiments, rats were given subcutaneous injections of 1 nmol/100 g of the peptides 28, 16 and 4 h before the sacrifice, and proliferative activity was assessed by counting the number of metaphase-arrested cells (after vincristine administration). In in vitro studies, cultured adrenocortical cells were exposed for 48 h to the peptides at a concentration of 10(-6) M, and proliferative activity was measured by the EZ4U method. The blockade of mu- and delta-receptors raised proliferative activity in immature adrenals and decreased it in regenerating glands, and the effects were reversed by mu- and delta-receptor agonists. Naltrexone-induced blockade of all met-Enk receptor subtypes decreased proliferative activity in immature adrenal and raised it in regenerating glands. The exposure to either mu- or delta-receptor agonists and antagonists evoked doubtful or no effects on the proliferative activity of cultured adrenocortical cells. In contrast, met-Enk exerted a marked antiproliferogenic effect that was reversed by naltrexone. Taken together, these findings allow us to draw the following conclusions: i) mu- and delta-receptor activation inhibits the growth of immature adrenals, stimulates adrenal regeneration, and does not affect proliferation of cultured adrenocortical cells; ii) zeta-receptors mediate the growth inhibitory effect of met-Enk on both regenerating adrenals and cultured adrenocortical cells, but unexpectedly their activation stimulates the growth of immature gland; and iii) the effects of mu- and delta-receptor activation in in vivo experiments are probably mediated by extra-adrenal indirect mechanisms.

Non-opioid actions of opioid peptides.

Beside the well known actions of opioid peptides on mu-, delta- and kappa-opioid receptors, increasing amount of pharmacological and biochemical evidence has recently been published about non-opioid actions of various opioid peptides. These effects are not abolished by naloxone treatments. Such non-opioid effects are observed both in nervous tissues and in the cellular elements of the immune system. Peptides exhibiting non-opioid effects include beta-endorphin, dynorphin A, nociceptin/OFQ, endomorphins, hemorphins and a number of Proenkephalin A derived peptides, such as Met-enkephalin, Met-enkephalin-Arg-Phe (MERF) and bovine adrenal medullary peptide (BAM22). Non-opioid actions are exerted through different neuronal receptors, e.g., dynorphin hyperalgesia through NMDA receptor, Met-enkephalin induced regulation of cell growth through zeta receptors, pain modulation by nociceptin through ORL-1 or NOP receptors, while BAM22 acts through sensory neuron specific G protein-coupled receptors (SNSR). We have investigated Met-enkephalin-Arg-Phe (MERF) and its analogues by the means of direct and indirect radioligand binding assays. It has been found that in addition to kappa(2) and delta-opioid receptors, MERF can act also through sigma(2)- or probably via FMRF-NH(2) receptors in rat cerebellum. A role of functionally assembling heterodimer receptors in mediating the non-conventional actions of these peptide ligands can not be excluded as well.

Signaling pathway involved in methionine enkephalin-promoted survival of lymphocytes infected by simian immunodeficiency virus in the early stage in vitro.

Methionine enkephalin, the endogenous opioid peptide, has a diversity of effects on the immune system. Although the biological effects of the pentapeptide have been well documented, little is known about the intracellular events involved in the effects of opioids on human immunodeficiency virus (HIV) infected immune cells. In the present investigation, the possible mechanism of apoptosis alleviated by exposure of methionine enkephalin at 1 micromol/l to CEM x 174 cells, the hybrid lymphocytes, infected with simian immunodeficiency virus (SIV) in vitro is elucidated. Apoptosis and cell cycle analysis is carried out by flow cytometry, the phosphorylation of mitogen-activated protein kinases (MAPK) ERK1 and ERK2 is detected by Western blotting assay, and changes of calcium concentration were analyzed using the calcium-sensitive dye Fluo-3 AM. The results exhibit that methionine enkephalin at the concentrations of 1 micromol/l increase remarkably the proportion of vital cells and decrease the apoptotic cells based on annexin V binding assay. In response to the treatment with methionine enkephalin, SIV-infected cells display a prolonged survival and are accumulated in G1 phase. Methionine enkephalin increase obviously the content of intracellular calcium in normal cells within 1-2 min and maintains a high level within monitoring time. However, the intracellular calcium reaches the highest level at 1 min and subsequently decline to background in SIV infected group. In addition, methionine enkephalin also elevates the levels of protein kinase C (PKC) activity and phosphorylated extracellular signal-regulated kinase (ERK) 1/2. It is proposed that calcium-PKC-MAPK cascade is involved in methionine enkephalin-prolonged survival of SIV-infected cells in the early stages of virus infection. The results provide a further evidence for potential use of methionine enkephalin on the therapy of Acquired Immunodeficiency Syndrome (AIDS).

Different mechanisms of intrinsic pain inhibition in early and late inflammation.

Neuroimmune interactions control pain through activation of opioid receptors on sensory nerves by immune-derived opioid peptides. Here we evaluate mechanisms of intrinsic pain inhibition at different stages of Freund's adjuvant-induced inflammation of the rat paw. We use immunohistochemistry and paw pressure testing. Our data show that in early (6 h) inflammation leukocyte-derived beta-endorphin, met-enkephalin and dynorphin A activate peripheral mu-, delta- and kappa-receptors to inhibit nociception. In addition, central opioid mechanisms seem to contribute significantly to this effect. At later stages (4 days), antinociception is exclusively produced by leukocyte-derived beta-endorphin acting at peripheral mu and delta receptors. Corticotropin-releasing hormone (CRH) is an endogenous trigger of these effects at both stages. These findings indicate that peripheral opioid mechanisms of pain inhibition gain functional relevance with the chronicity of inflammation.