Methionine Enkephalinamide Research Articles

Effects of opioid receptor activation on cardiovascular responses and extracellular monoamines within the rostral ventrolateral medulla during static contraction of skeletal muscle.

During static muscle contraction, activation of opioid receptors alters the extracellular glutamate concentrations within the rostral ventrolateral medulla (RVLM). In addition, microdialysis of glutamate in the ventrolateral medulla (VLM) increases the release of norepinephrine (NE), dopamine (DA), and serotonin (5-HT). Therefore, we hypothesized that extracellular concentrations of these monoamines as well as cardiovascular responses during static skeletal muscle contraction would be modulated following administration of [D-Ala(2)]methionine enkephalinamide (DAME), an opioid receptor agonist, into the RVLM. Microdialysis of 100 microM DAME into the RVLM of 10 rats significantly (P<0.01) decreased extracellular levels (in pg/10 microl) of NE (from 3.3+/-0.3 to 1.9+/-0.3), DA (from 5.5+/-0.2 to 3.7+/-0.3), and 5-HT (from 6.1+/-0.8 to 3.6+/-0.2) during static exercise. After microdialysis of DAME, the exercise pressor reflex also significantly (P<0.01) decreased mean arterial pressure (MAP) by 13+/-3 mmHg and heart rate (HR) by 16+/-6 bpm, compared with control (MAP=22+/-4 mmHg and HR=31+/-7 bpm). Subsequently, after 30 min microdialysis of naloxone, an opioid receptor antagonist, muscle contraction increased the extracellular monoamine levels (in pg/10 microl, 3.8+/-0.3 NE; 5.2+/-0.3 DA; and 5.5+/-0.4 5-HT) similar to the control groups and evoked a reversal of cardiovascular responses. Similarly, 30 min of microdialyzing naloxone, added to the perfusing medium containing DAME, reversed the attenuating effects of DAME on monoamines, MAP, and HR during a muscle contraction. Furthermore, microdialysis of 100 microM naloxone alone for 30 min potentiated cardiovascular responses and monoamine levels during a muscle contraction. In summary, the present data demonstrates that microdialysis of DAME into RVLM attenuates the exercise pressor reflex mediated increases in MAP, HR and extracellular levels of biogenic monoamines. A subsequent microdialysis of naloxone reversed the effects suggesting that an opioidergic mechanism within RVLM modulates the exercise pressor reflex. Overall, the present study provides further insights into the opioidergic modulation of the exercise pressor reflex.

Rostral ventrolateral medulla opioid receptor activation modulates glutamate release and attenuates the exercise pressor reflex

We previously reported that the administration of [ d-Ala 2]methionine enkephalinamide (DAME), an opioid receptor agonist, into the rostral (RVLM) but not into the caudal ventrolateral medulla (CVLM), attenuated increases in mean arterial pressure (MAP) and heart rate (HR) during static muscle contraction that had been blocked by prior microdialysis of the opioid receptor antagonist, naloxone [Am. J. Physiol. 274 (1998) H139–H146]. In this study, we determine whether this RVLM-mediated opioidergic-modulation of cardiovascular responses is associated with localized changes in extracellular concentrations of glutamate, an excitatory amino acid, using microdialysis techniques in anesthetized rats. Muscle contraction increased MAP and HR by 37±5 mmHg and 23±3 bpm, respectively. Extracellular glutamate concentrations, determined using HPLC–ECD, increased from 0.8±0.2 to 6.6±1.2 ng/5 μl in the bilateral RVLM areas. Microdialysis of DAME (100 μM) for 30 min attenuated the contraction-evoked increases in MAP, HR, and glutamate levels (20±4 mmHg, 10±2 bpm, and 1.8±0.2 ng/5 μl, respectively). After microdialysis of naloxone (100 μM) for 30 min into the RVLM, muscle contraction blocked the attenuations (35±5 mmHg, 26±4 bpm, and 5.8±1.0 ng/5 μl, respectively). Developed muscle tensions were similar throughout the protocol (676±38, 678±37 and 687±37 g, respectively). These results suggest that an opioidergic receptor-mediated mechanism within the RVLM attenuates cardiovascular responses during static exercise via modulating extracellular concentrations of glutamate in the RVLM.

Enzymatic and permeation barrier of [ d-Ala 2]-Met-enkephalinamide in the anterior membranes of the albino rabbit eye

Enzymatic and physical barrier properties of anterior ocular membranes were characterized. The permeation and metabolic degradation of [ d-Ala 2]-methionine enkephalinamide (DAMEA) in the albino rabbit cornea, conjunctiva and sclera were studied in vitro. DAMEA was administered with and without peptidase inhibitors bestatin (aminopeptidase inhibitor) and SCH 39370 (enkephalinase inhibitor). The modified Ussing chambers were used to study the peptide permeation and the samples were analyzed with a novel HPLC method using UV and EC detectors. Sclera was the most permeable membrane to DAMEA, while cornea was almost impermeable to DAMEA. Without inhibitors, the permeability coefficients of DAMEA were 2.7×10 −8 cm/s, 3.1×10 −6 cm/s and 12.5×10 −6 cm/s in the cornea, conjunctiva and sclera, respectively. DAMEA was partly metabolized to tyrosine (Tyr) and tyrosine- d-alanine-glycine (Tyr- d-Ala-Gly). When inhibitors were co-administered with DAMEA, the corneal permeability of intact DAMEA increased 15 times, while conjunctival permeability increased 5.5 times and scleral permeability remained practically unaltered. The formation of metabolites decreased markedly, when the inhibitors were used. Interestingly, when the permeability of DAMEA was compared to permeabilities of polyethylene glycols in different membranes, the permeation was in the same range suggesting that DAMEA permeates through cornea via a paracellular pathway. Both enzymatic and physical barriers were more prominent in the cornea than in the conjunctiva and sclera. Non-corneal pathway of absorption and combined with inhibition of peptidases may be the most viable pathway for ocular peptide administration.

Gradient high-performance liquid chromatographic analysis of enkephalin peptides, their metabolites, and enzyme inhibitors using combined ultraviolet and electrochemical detection: II. Application to ocular permeability studies in vitro

A gradient HPLC method with combined ultraviolet (UV) and electrochemical detection (ED) was used to study the ocular permeability of [ d-Ala 2]-methionine enkephalinamide (MEA) in vitro. Coulometric ED was selective for MEA and its tyrosine-containing metabolites with quantitation limits between 20 and 60 n M (1.0–3.0 pmol per 50-μl injection), whereas UV detection at 205 nm allowed the determination of several aromatic metabolites and enzyme inhibitors with quantitation limits between 40 and 500 n M (2.0–25.0 pmol). This method was capable of detecting permeability of MEA and metabolite formation in the cornea and conjunctiva in vitro. Furthermore, effects of aminopeptidase inhibitor bestatin and enkephalinase inhibitor SCH 39370 on permeation and metabolism of MEA could be determined.

Effect of the opioid methionine enkephalinamide on signal transduction in human T-lymphocytes

T cell receptor ( TCR CD3 ) induced fluctuations in intracellular free ionizied calcium, [Ca 2+] i, was analysed in the human T leukemia cell clone, Jurkat, cultured in the presence of the opioid methionine enkephalinamide (Met-Enk) in titrated concentrations (10 −7 to 10 −15 M) or saline (PBS). In the majority of individual experiments, the activation-induced fluctuations in [Ca 2+] i were similar in cells cultured in the presence of Met-Enk and PBS, respectively. However, when all the experimental data from 101 separate TCR CD3 -activation experiments with Met-Enk were compared with the 67 separate control experiments, we found that a fraction (20–40%) of the individual sets of Met-Enk experiments responded significantly different when compared to PBS-controls. In this fraction of experiments the increase in [Ca 2+] i after ligation of the TCR CD3 complex was extremely slow compared to controls. Moreover, the levels of [Ca 2+] i in this particular fraction were lower than control levels prior to ligation of the TCR CD3 complex. The data support the idea that signal transduction in T cells can be influenced by endogenous opioid. The data therefore give credit to the evolving hypothesis of a functional relationship between the neuroendocrine system and the immune system.

Rostral ventrolateral medullary opioid receptor activation modulates pressor response to muscle contraction

The effects of an opioid agonist, [D-Ala2]methionine enkephalinamide (DAME), administered into the rostral ventrolateral medulla (rVLM) or caudal ventrolateral medulla (cVLM) on cardiovascular responses to isometric muscle contraction were determined in anesthetized rats. A 30-s contraction evoked by tibial nerve stimulation increased mean arterial pressure (MAP) and heart rate (HR) by 34 +/- 6 mmHg and 40 +/- 7 beats/min, respectively, with a developed tension of 322 +/- 30 g, after bilateral insertion of microdialysis probes into the rVLM. Thirty-minute dialysis of DAME (10 and 100 microM) attenuated the contraction-evoked cardiovascular changes dose dependently (10 microM: MAP = 25 +/- 4 mmHg, HR = 27 +/- 3 beats/min, tension = 333 +/- 25 g; 100 microM: MAP = 14 +/- 4 mmHg, HR = 16 +/- 5 beats/min, tension = 330 +/- 34 g). Preadministration of an opioid antagonist, naloxone (100 microM), augmented contraction-evoked MAP and HR responses and blocked effects of 100 microM DAME. Microdialysis of DAME into the cVLM produced no changes in the pressor response to contraction. At end of each experiment, tibial nerve stimulation after neuromuscular blockade evoked no MAP or HR change. Results demonstrate that opioid receptor activation within the rVLM modulates cardiovascular responses to isometric muscle contraction.

Gradient high-performance liquid chromatographic analysis of enkephalin peptides, their metabolites and enzyme inhibitors using combined ultraviolet and electrochemical detection.: I. Optimization of separation and detection

Coulometric electrochemical detection (ED) was compatible with gradient HPLC analysis of enkephalin peptides, their metabolites and several enzyme inhibitors with combined UV detection. The compounds were separated using a reversed-phase octylsilane column with 35 m M phosphate buffer (pH 2.1)–acetonitrile eluent. Hydrodynamic voltammograms were determined for [ d-Ala 2]–methionine enkephalinamide (MEA) and its metabolites, and the response of MEA consisted of the oxidation of the tyrosine and methioninamide residues with a total transfer of five electrons at the highest potential used. Based on the voltammograms, ED was optimized for the selective determination of MEA and other tyrosine-containing peptides with detection limits between 0.4 and 0.6 pmol per 50 μl injection. In addition, simultaneous UV detection at 205 nm allowed the determination of all the aromatic peptides and enzyme inhibitors with detection limits between 1.5 and 30.0 pmol.

Inhibition of respiratory burst in mice macrophages by opioid peptide [D-ALA 2]methionine enkephalinamide

Inhibition of respiratory burst in mice macrophages by opioid peptide [D-ALA 2]methionine enkephalinamide

Inhibition of the respiratory burst in mouse macrophages by ultra-low doses of an opioid peptide is consistent with a possible adaptation mechanism

The respiratory burst induced by phorbol myristate acetate in mouse macrophages was inhibited by ultra-low doses (10 −15 −10 −13 M) of an opioid peptide [ d-Ala 2] methionine enkephalinamide. The effect disappeared at concentrations above and below this range. The inhibition approached 50% and was statistically significant ( P < 0.001). Increasing the time of the opioid incubation with cells brought about a shift in the maximal effect to lower concentrations of the opioid (from 10 −13 to 5 · 10 −15 M) and led to a decrease in the value of the effect, fully in accord with the previously proposed adaptation mechanism of the action of ultra-low doses.

Effects of (d-Ala2) methionine enkephalinamide in PMA-induced mouse ear edema

The synthetic opioid peptide (D-Ala2) methionine enkephalinamide (DAMEA) was reported to be an extremely potent inhibitor of the "respiratory burst" of polymorphonuclear neutrophils (PMNs). In the present paper the activity of DAMEA was investigated in the PMA-induced mouse ear edema since there is strong evidence that reactive oxygen species (ROS) are involved in the pathogenesis of this inflammatory reaction. DAMEA inhibited PMA-induced mouse ear edema in a dose-dependent manner between 10(-6) and 10(-9) mol/ear. The well-known anti-inflammatory activity of narcotic analgesics could be explained partly by an inhibition of ROS production.

Mesoaccumbens dopamine-opiate interactions in the control over behaviour by a conditioned reinforcer

These experiments examined the role of dopamine-opiate interactions in the ventral tegmental area (VTA) and nucleus accumbens in the mediation of reinforcement-related behaviour. It has been shown previously that opiates induce a dopamine-dependent increase in locomotor activity in rats when infused into the VTA, and a dopamine-independent hyperactivity when infused into the nucleus accumbens. The present study investigated the generality and significance of these two findings, by examining dopamine-opiate interactions in the control over behaviour exerted by a conditioned reinforcer (CR), an arbitrary stimulus which gains control by association with primary reinforcement. Rats were trained to associate a light/noise stimulus with sucrose reinforcement, and the efficacy of the CR in controlling behaviour was assessed by measuring its ability to support a new lever pressing response. Responding on one lever (CR lever) produced the CR, responding on the other lever had no programmed consequences. In experiment 1, intra-accumbens infusions of d-amphetamine (10 micrograms), the D1 dopamine receptor agonist SKF-38393 (0.1 microgram), the D2 dopamine receptor agonist LY-171555 (quinpirole; 0.1 microgram) or the opiate receptor agonist [D-Ala2]-methionine enkephalinamide (DALA; 1 microgram) selectively increased responding on the CR lever. Infusion with DALA intra-VTA had no effect. However, pretreatment with DALA intra-VTA (10 x 1 microgram/day) subsequently reduced the selectivity of the response to infusions intra-accumbens with d-amphetamine or SKF-38393, and blocked the response to LY-171555 or DALA. Pretreatment also shifted to the right the dose-response function for DALA intra-accumbens. In experiment 2, intra-accumbens infusions of d-amphetamine, SKF-38393, LY-171555 or DALA again increased responding on the CR lever only. Pretreatment with intra-accumbens d-amphetamine (5 x 1 microgram/day) reduced the selectivity of the response subsequently to d-amphetamine, and blocked the response to SKF-38393, LY-171555 or DALA. In experiment 3, intra-accumbens infusions of the mu-opiate receptor agonist [D-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (0.003-0.1 microgram), or the delta-opiate receptor agonist [D-Pen2,5]-enkephalin (0.03-1 microgram) enhanced selectively responding on the CR lever. Thus, the dopamine-dependent locomotor-stimulant properties of intra-VTA infusions of opiates are associated with impaired conditioned reinforcer efficacy. Finally, repeated stimulation of the mesoaccumbens dopamine pathway may compromise the dopamine-independence of the opiate system within the nucleus accumbens.

Respiratory burst inhibition in human neutrophils by ultra-low doses of [D-Ala 2]methionine enkephalinamide

An ultra-low dose (10 −14 M) of opioid peptide [D-Ala 2]methionine enkephalinamide (DAMEA) is found to exert an inhibitory effect on the production of reactive oxygen species (respiratory burst) in human neutrophils. The validity of this phenomenon has been verified in a series of studies that comprised 30 experiments. The inhibition has proved to be statistically significant (P<0.001). The dose-response dependence of the effect (10 −15−10 −9 M) followed a characteristic biphasic pattern (with the maximum effect at ultra-low doses). An opioid antagonist, naloxone partially blocks the inhibitory effect, which indicates that the DAMEA action is at least partially mediated by opioid receptors.

Regulation of Dopamine Release in vitro from the Posterior Pituitary by Opioid Peptides

Opioid peptides are present in both the posterior pituitary (PP) and stalk-median eminence (SME). Their effects on the dopaminergic neurons in the SME are well documented, but little is known concerning their role in the regulation of dopamine (DA) release from the PP. The objectives of this study were (1) to develop an in vitro method suitable for examining the regulation of endogenous DA release from PP and SME, and (2) to describe and compare the effects of selected opioid peptides on potassium-evoked DA release from these tissues. Tissues were dissected from ovariectomized rats and incubated in media. After equilibration, two pulses of 28 mM potassium (K+), 3 min each, were delivered 30 min apart. Test substances were administered 20 min before the second K+ stimulus. DA in the media was determined by high-performance liquid chromatography. Potassium at 28 and 56 mM elicited a marked increase in DA release from the PP and SME; this was abolished by the removal of calcium. The opioid receptor antagonist, naloxone, significantly increased the release of DA from both PP and SME by 55%. Dynorphin A elicited a significant inhibition of DA release from PP and SME by 33 and 50%, respectively. In contrast, methionine enkephalinamide decreased DA release from the SME by 50%, but was without effect in the PP. The release of DA from both PP and SME was significantly inhibited by beta-endorphin, and this was reversed by naloxone. However, beta-endorphin was fourfold more effective in the SME. N-acetyl-beta-endorphin did not alter DA release. (1) we have developed a simple and sensitive in vitro method for studying the effects of hormones and drugs on the release of endogenous DA from PP and SME; (2) tuberoinfundibular dopaminergic and tuberohypophyseal dopaminergic nerve terminals are subjected to a similar inhibitory control by endogenous opioid peptides, and (3) exogenously applied opioid peptides exert differential effects on the release of DA from SME and PP which could be attributable to a dissimilar distribution of opioid receptor subtypes in these two tissues.

Effects of [D-Ala 2] Met-enkephalinamide, a Met-enkephalin analog, on delayed response by squirrel monkeys

Squirrel monkeys were tested on an indirect spatial delayed response task after subcutaneous injections of either physiological saline, 100, or 500 microgram/kg [D-Ala 2] methionine enkephalinamide (DAME). During Experiment 1 monkeys completed two-choice delayed response problems with 0-, 6-, 12- and 18- sec delays; DAME significantly enhanced performance at 18-sec delays but not for the shorter delay. During Experiment 2, monkeys performed a nine-choice spatial delayed response task with 0-, 4- and 8-sec delays. In Experiment 2, low but not high doses of DAME impaired performance. Memory functions represented by a bias-free performance measure paralleled percentage correct in both experiments. Therefore, DAME effects, in enhancing and in impairing performance, were not attributable to changes in the relative number of systematic spatial errors (positional win-shift; lose-stay, position perseveration, and position preference).

Effects of opioids on acetylcholine output of perfused human placental cotyleda

1. (1) Output of acetylcholine (ACh) from fetal vessels of Krebs-perfused human placental cotyleda was estimated by bioassay using the rat stomach strip. 2. (2) Infusion of high concentrations of the opioids pethidine, morphine or ethylketocyclazocine, but not [ d-Ala 2]-methionine enkephalinamide (0.1–300 μmol/l), reduced ACh output. Fifty per cent inhibition of output occurred in the presence of concentrations of ⩾ 300 μmol/l (morphine and ethylketocyclazocine) and 338 (317, 353; 95 per cent c.l., n = 6) μmol/l (pethidine). 3. (3) ACh output was inhibited by infusion of 100 μmol/l naloxone or 10 μmol/l naltrexone, but was not affected by lower concentrations of either antagonist. 4. (4) These results suggest that the therapeutic concentrations of morphine and pethidine likely to occur in vivo would not affect placental ACh output into fetal vessels. The finding that high concentrations of synthetic opioids or opioid antagonists were required to inhibit output suggests that they may not be acting specifically, and provides no evidence for the hypothesis that endogenous opioids play a role in control of ACh release into fetal vessels of human placentae.

Antitumor activity of enkephalin analogues in inhibiting PYB6 tumor growth in mice and immunological effects of methionine enkephalinamide

Recent evidence has implicated enkephalins as immunomodulators. Several studies have reported the regulation of tumor growth by methionine enkephalin (ME). However, there has been little effort to relate the immunological significance of enkephalins to the development of anticancer drugs. The present study had three aims: first, to compare the antitumor activity of the synthetic peptide, D-[Ala 2]methionine enkephalinamide (MEA), with endogenous enkephalins on PYB6 fibrosarcoma tumor growth; second, to determine whether tumor growth inhibition was mediated by an opiate receptor; and third, to investigate the effects of MEA on selected immune responses. Female B6C3F1 mice were injected i.p. daily for 7 days with 50–4000 μg/kg of ME, MEA, leucine enkephalin (LE) or D-[Ala 2]leucine enkephalinamide (LEA), beginning 1 day after PYB6 inoculation. ME and MEA, but not LE or LEA, decreased the PYB6 growth rate. The dose of 50 μg/kg MEA exerted the maximum inhibition of tumor growth (nearly 72% on day 15 post tumor transplantation). MEA was not directly toxic to PYB6 tumor cells, as evaluated by the measurement of DNA synthesis and cellular ATP levels of PYB6 cells exposed to MEA in vitro. No 3H-etorphine specific bindings were detected on the cell membrane or sonicates of splenic lymphocytes or PYB6 cells. Therefore, the antitumor activity by MEA is likely mediated by an indirect mechanism. Immunological studies indicated that MEA selectively enhanced the lymphoproliferative response to the T-cell mitogen, concanavalin A, but not to the B-cell mitogen, lipopolysaccharide. Natural killer cell activity was unaffected both after in vivo and in vitro exposure to MEA and the number of IgM antibody-forming cells in response to the T-dependent antigen, sheep red blood cells, was unaffected after in vitro exposure to MEA.

Characterization of the effects of neurokinins on canine antral muscle

The excitation of longitudinal antral muscle by substance P (SP) involves both a myogenic and a cholinergic effect. To examine if these responses are mediated by different neurokinin receptors, we studied the mechanical response and the release of [3H]acetylcholine from antral muscle strips in response to SP, substance P methylester (SPME), neurokinin A (NKA), neurokinin B (NKB), and several non-mammalian tachykinins. All peptides studied showed a dose-dependent inotropic and chronotropic effect on spontaneous phasic contractions. This ionotropic effect in longitudinal muscle was partially atropine sensitive for SPME, SP, and NKB but not for NKA, whereas neither atropine nor tetrodotoxin had an effect in circular muscle. In longitudinal muscle, all three neurokinins were equipotent. In longitudinal muscle treated with atropine and in circular muscle, the rank order of potency for the inotropic response was NKA greater than NKB greater than SP greater than SPME. For the chronotropic response the rank order was SPME, SP greater than NKA greater than NKB. NKA, NKB, and SP caused a dose-dependent, tetrodotoxin-sensitive increase in [3H]acetylcholine release from strips preincubated with [3H]choline. NKA was significantly more potent to release [3H]acetylcholine than either NKB or SP. The stimulated release was inhibited by [D-Ala2,D-Met5]methionine enkephalinamide and the SP antagonist, spantide. These results are consistent with the hypothesis that NKA is the natural ligand mediating the myogenic inotropic response in both muscle layers and the cholinergic response in longitudinal muscle.

Effects of intrathecal antagonists on the antinociception, hypotension, and bradycardia produced by intravenous administration of [D-Ala2]-methionine enkephalinamide (DALA) in the rat.

DALA is a synthetic pentapeptide that produces inhibition of the tail-flick reflex evoked by radiant heat, as well as hypotension and bradycardia. Two experiments examined the effects of administration of various receptor antagonists into the subarachnoid space of the lumbar spinal cord on the antinociception produced by IV administration of DALA. Experiment 1 showed that intrathecal administration of 30 micrograms of phentolamine produced a significant reduction in the antinociceptive effect of DALA, while naloxone (30 micrograms), methysergide (30 micrograms), or vehicle control had no effect. Experiment 2 showed that intrathecal administration of combinations of either phentolamine and methysergide, or phentolamine and naloxone, were no more effective in reducing the antinociceptive effect of DALA than administration of phentolamine alone. These data demonstrate the involvement of descending noradrenergic systems in the production of antinociception by IV DALA. Further, they indicate that the antinociception produced by DALA is independent of a direct spinal action of the drug.

Modulation of Human Neutrophil Function by Endogenous Opiate Enkephalins

On the basis of recent evidence, the natural opiate enkephalins, which previously were believed to be confined to the central nervous system, are now known, in fact, to be released from the adrenal glands by sympathetic activation or trauma. To determine if enkephalins (EKs) affect peripheral function, the influence of synthetic leucine and methionine enkephalin (leuEK and metEK) on several relevant functions of human polymorphonuclear leukocytes was evaluated. Initial attempts to detect interaction of leuEK and metEK with neutrophils yielded inconsistent results. Further studies were done using protease-resistant methionine enkephalin-amide (metEKamide). MetEKamide was able to induce degranulation when present at 10(-3) and 10(-4) mmol/L as determined by release of beta-glucuronidase and lysozyme. Using the under-agarose chemotaxis method, treatment with metEKamide resulted in no change of the neutrophil's chemotactic response to an optimal concentration of the chemotactic peptide formyl-methionyl-leucyl-phenyl-alanine (FMLP). However, responsiveness to low levels of FMLP increased in cells treated with 10(-3)-10(-5) mmol/L metaEKamide. This appeared to be a result of increased chemokinesis of the treated cells. Scanning electron microscopic studies of cells exposed to metEKamide revealed that treatment resulted in changes in neutrophil morphology. When metEKamide itself was tested as a potential chemotactic agent, 10(-2) mmol/L metEKamide in an opposing well served to induce chemotaxis. Our results, along with those of recent studies of EKs as immunomodulators of T cell function, suggest that neurohormones can function as regulators of the immune response.

Interaction between enkephalin and dopamine in the avian retina

Biochemical and pharmacological techniques were utilized to investigate the interaction between the enkephalinergic and dopaminergic systems in the chicken retina. Exogenously applied enkephalin and its analogues were observed to inhibit the release of preloaded dopamine from the retina. This inhibition was concentration-dependent and was suppressed by the opiate antagonist, naloxone. The relationship between enkephalinergic and dopaminergic amacrine cells was studied in retinas which were subjected to 6-hydroxydopamine (6-OHDA) treatments. 6-OHDA degenerated approximately 80–90% of those cells which exhibit high affinity uptake of [ 3H]dopamine. In 6-OHDA-treated retinas, the capacity of 3H-labelled [ d-Ala 2]methionine enkephalinamide to bind specifically to opiate receptors was substantially reduced (only 70–75% of the control). Scatchard analyses and ligand displacement studies indicated that this decrease in binding was due to a reduction in the number of opiate receptors. Taken together, these observations strongly indicate that a fraction of the opiate receptors in the chicken retina (25–30%) are closely associated with the population of dopaminergic amacrine cells.