d-ala(2)-mephe(4)-met(0)-ol-enkephalin and deltakephalin

We have previously reported that met-enkephalin has dual immunomodulatory properties in vitro. We have continued this investigation using an in vivo system. In this study, Alzet miniosmotic pumps were loaded with either met-enkephalin, DTLET or FK 33-824 and were surgically implanted into BAF1/J mice. Twenty-four hours after pump implantation, mice were challenged with sub-optimal, optimal or supraoptimal immunizing doses of antigen. The immune response was assessed 4 or 5 days after primary immunization. FK 33-824, a met-enkephalin analogue, had no effect on the response of mice challenged with a suboptimal antigen dose. However, FK 33-824, at a pump concentration of 10(-3) M, suppressed the response against optimal challenge doses of antigen. At a pump concentration of 10(-8) M, FK 33-824 suppressed, enhanced or had no effect on the supraoptimal antigen dose-induced immune response. The suppressive effect of FK 33-824 in mice immunized with either optimal or supraoptimal doses of antigen was blocked by naloxone. Met-enkephalin and its delta opioid receptor specific analogue, DTLET, had no effect on the immune response to optimal antigen immunization. These results indicate that FK 33-824 has in vivo immunomodulatory activity and provide evidence that opioid peptides may either upregulate or downregulate the in vivo immune response depending on the strength of the response.

Topics: Amino Acid Sequence; Animals; Antibody Formation; Corticosterone; D-Ala(2),MePhe(4),Met(0)-ol-enkephalin; Dose-Response Relationship, Drug; Enkephalin, Methionine; Female; Immunization; Immunologic Tests; Infusion Pumps, Implantable; Mice; Mice, Inbred Strains; Molecular Sequence Data; Oligopeptides; Receptors, Opioid

Superfusion of slices from the dorsal half of the lumbar enlargement of rat spinal cord with Krebs-Henseleit medium supplemented with 30 microM bacitracin allowed the collection of substance P-like immunoreactive material (SPLI), which was released at a rate of approximately 10 pg/4 min. Tissue depolarization by an excess of K+ (30-60 mM) or veratridine (50 microM) induced a marked increase in SPLI outflow, provided that Ca2+ was present in the superfusing fluid. K+- or veratridine-induced SPLI overflow could be modulated in opposite directions by mu and delta opioid receptor agonists. Thus, the two preferential mu agonists Tyr-D-Ala-Gly-MePhe-Gly-ol (DAGO; 10 microM) and Tyr-D-Ala-Gly-MePhe-Met(O)5-OH (FK-33824; 0.1 microM) enhanced SPLI overflow from depolarized tissues, whereas the selective delta agonists Tyr-D-Thr-Gly-Phe-Leu-Thr (deltakephalin; 3 microM) and [2-D-penicillamine, 5-D-penicillamine]enkephalin (50 microM) reduced it. The effect of DAGO was antagonized by a low concentration (1 microM) of naloxone but not by the selective delta antagonist ICI-154129 (50 microM). In contrast, the latter drug prevented the inhibitory influence of delta agonists on K+-induced SPLI release. Complementary experiments with morphine (10 microM) and [2-D-alanine, 5-D-leucine]enkephalinamide (3 microM), in combination with 1 microM naloxone or 50 microM ICI-154129 for the selective blockade of mu or delta receptors, respectively, confirmed that the stimulation of mu receptors increased, whereas the stimulation of delta receptors reduced, SPLI overflow. The results suggest that, at the spinal level, and antinociceptive action of delta but not mu agonists might involve a presynaptic inhibition of substance P-containing primary afferent fibers.

Topics: Animals; D-Ala(2),MePhe(4),Met(0)-ol-enkephalin; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, Leucine; Enkephalins; In Vitro Techniques; Male; Morphine; Naloxone; Oligopeptides; Potassium; Rats; Rats, Inbred Strains; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, mu; Spinal Cord; Substance P; Tetrodotoxin; Veratridine

Using an in-vitro superfusion system, the relative importance of three distinct subtypes of the opiate receptor in the control of the secretion of LHRH from the mediobasal hypothalamus of the cockerel was investigated. Basal release of LHRH was increased by the antagonist naloxone, which shows some mu-receptor selectivity, in a manner which was reversed by the mu-receptor specific agonist [D-Ala2, N-Phe4-Gly-ol5]-enkephalin (DAGO) and the mu- and delta-specific agonist [D-Ala2,N-Phe4,Met(0)ol5]-enkephalin (FK 33-824). The delta-specific agonist [D-Thr2,L-Leu5]-enkephalyl-Thr (DTLET) and the kappa-specific agonist 1-methyl-2(3-thienylcarbonyl)-aminomethyl-5-(2-fluorophenyl)-H-2, 3-dihydro-1,4-benzodiazepine (KC 6128; (+)-titfluadom) did not reverse the effect of naloxone. The delta-specific antagonist N,N-diallyl-Tyr-alpha-aminoisobutyricacid-Phe-Leu-OH (ICI 174,864) failed to influence basal release. Release of LHRH stimulated by increasing the potassium ion concentration of the superfusate to 48 mmol/l was reduced by DAGO in a manner which was reversed by naloxone, and by FK 33-824 in a manner which was reversed by both naloxone and ICI 174,864. The agonists DTLET and titfluadom did not affect stimulated release of LHRH. These results support the proposal that spontaneous release of LHRH is tonically inhibited by agonists acting through the mu-receptor whilst, in response to a stimulus, the delta-receptor, in addition to the mu-receptor, may be involved.

Topics: Animals; Benzodiazepines; Chickens; D-Ala(2),MePhe(4),Met(0)-ol-enkephalin; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, Leucine; Enkephalins; Gonadotropin-Releasing Hormone; Hypothalamus; In Vitro Techniques; Male; Naloxone; Oligopeptides; Receptors, Opioid