dynorphins and dermorphin

The opioid system in the central nervous system regulates depressive-like behavior in animals. Opioid receptors and their endogenous ligands have been focused on as novel therapeutic targets for depression. We synthesized dermorphin (DRM)-dynorphin (DYN) analogs (DRM-DYN001-004) using the message-address concept concerning interactions with opioid receptors. It has previously been reported that DRM-DYN001, 003, and 004 have shown high affinities for μ- and κ-opioid receptors, whereas all analogs had a lower affinity for the δ-opioid receptor than for other receptors using a receptor binding assay. However, it remains unknown whether these analogs show antidepressant-like effects in mice. We examined the effects of DRM-DYN analogs on the duration of immobile behavior in a tail suspension test. Intracerebroventricular administration of DRM-DYN001 in mice shortened the duration of immobile behavior, but did not affect locomotion. The DRM-DYN001-induced antidepressant-like effect was inhibited by co-administration of naloxone (non-selective opioid receptor antagonist), naloxonazine (selective μ

Topics: Animals; Antidepressive Agents; Dynorphins; Mice; Narcotic Antagonists; Opioid Peptides; Receptors, Opioid; Receptors, Opioid, kappa

Mild stress repeatedly applied to neonatal rodents induces alterations of central nervous system functions, persisting up to the adult age. Most alterations may be mediated through hormones and neuromediators active on the autonomic nervous system, therefore we tested the efficacy of selective opioid receptor agonists on the vas deferens of adult mice that, as neonates, had undergone daily mild stress until weaning (brief isolation and solvent injection). We found in the adult mouse (90 days old) decreased sensitivity of vas deferens to selective mu-, delta- and kappa-opioid receptor agonist drugs. The neonatal administration of an antisense oligodeoxynucleotide adrenocorticotropin-synthesis-inhibitor partly prevented these effects.

Topics: Adrenocorticotropic Hormone; Analgesics, Opioid; Animals; Animals, Newborn; Dose-Response Relationship, Drug; Dynorphins; Female; Humans; Male; Mice; Mutation; Oligonucleotides, Antisense; Oligopeptides; Opioid Peptides; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, kappa; Receptors, Opioid, mu; Stress, Physiological; Time Factors; Vas Deferens

Nerve injury-induced afferent discharge is thought to elicit spinal sensitization and consequent abnormal pain. Experimental neuropathic pain, however, also depends on central changes, including descending facilitation arising from the rostral ventromedial medulla (RVM) and upregulation of spinal dynorphin. A possible intersection of these influences at the spinal level was explored by measuring evoked, excitatory transmitter release in tissues taken from nerve-injured animals with or without previous manipulation of descending modulatory systems. Spinal nerve ligation (SNL) produced expected tactile and thermal hyperesthesias. Capsaicin-evoked calcitonin gene-related peptide (CGRP) release was markedly enhanced in lumbar spinal tissue from SNL rats when compared with sham-operated controls. Enhanced, evoked CGRP release from SNL rats was blocked by anti-dynorphin A(1-13) antiserum; this treatment did not alter evoked release in tissues from sham-operated rats. Dorsolateral funiculus lesion (DLF) or destruction of RVM neurons expressing mu-opioid receptors with dermorphin-saporin, blocked tactile and thermal hypersensitivity, as well as SNL-induced upregulation of spinal dynorphin. Spinal tissues from these DLF-lesioned or dermorphin-saporin-treated SNL rats did not exhibit enhanced capsaicin-evoked CGRP-IR release. These data demonstrate exaggerated release of excitatory transmitter from primary afferents after injury to peripheral nerves, supporting the likely importance of increased afferent input as a driving force of neuropathic pain. The data also show that modulatory influences of descending facilitation are required for enhanced evoked transmitter release after nerve injury. Thus, convergence of descending modulation, spinal plasticity, and afferent drive in the nerve-injured state reveals a mechanism by which some aspects of nerve injury-induced hyperesthesias may occur.

Topics: Afferent Pathways; Analgesics, Opioid; Animals; Calcitonin Gene-Related Peptide; Capsaicin; Disease Models, Animal; Dynorphins; Enkephalins; Hyperesthesia; Ligation; Lumbosacral Region; Male; Medulla Oblongata; Microinjections; N-Glycosyl Hydrolases; Nerve Compression Syndromes; Neuralgia; Neuronal Plasticity; Neurotransmitter Agents; Oligopeptides; Opioid Peptides; Pain Measurement; Protein Precursors; Rats; Rats, Sprague-Dawley; Receptors, Opioid, mu; Recombinant Fusion Proteins; Ribosome Inactivating Proteins, Type 1; Saporins; Sensory Thresholds; Spinal Cord; Spinal Nerves

Although injury-induced afferent discharge declines significantly over time, experimental neuropathic pain persists unchanged for long periods. These observations suggest that processes that initiate experimental neuropathic pain may differ from those that maintain such pain. Here, the role of descending facilitation arising from developing plasticity in the rostral ventromedial medulla (RVM) in the initiation and maintenance of experimental neuropathic pain was explored. Tactile and thermal hypersensitivity were induced in rats by spinal nerve ligation (SNL). RVM lidocaine blocked SNL-induced tactile and thermal hypersensitivity on post-SNL days 6-12 but not on post-SNL day 3. Lesion of RVM cells expressing mu-opioid receptors with dermorphin-saporin did not prevent the onset of SNL-induced tactile and thermal hypersensitivity, but these signs reversed to baseline levels beginning on post-SNL day 4. Similarly, lesions of the dorsolateral funiculus (DLF) did not prevent the onset of SNL-induced tactile and thermal hypersensitivity, but these signs reversed to baseline levels beginning on post-SNL day 4. Lesions of the DLF also blocked the SNL-induced increase in spinal dynorphin content, which has been suggested to promote neuropathic pain. These data distinguish mechanisms that initiate the neuropathic state as independent of descending supraspinal influences and additional mechanism(s) that require supraspinal facilitation to maintain such pain. In addition, the data indicate that these time-dependent descending influences can underlie some of the SNL-induced plasticity at the spinal level. Such time-dependent descending influences driving associated spinal changes, such as the upregulation of dynorphin, are key elements in the maintenance, but not initiation, of neuropathic states.

Topics: Animals; Behavior, Animal; Denervation; Dynorphins; Immunotoxins; Kinetics; Lidocaine; Ligation; Male; Medulla Oblongata; Microinjections; N-Glycosyl Hydrolases; Oligopeptides; Opioid Peptides; Pain; Plant Proteins; Rats; Rats, Sprague-Dawley; Ribosome Inactivating Proteins, Type 1; Saporins; Spinal Cord; Spinal Nerves

1. Single pulse electrical field stimulation (EFS, 0.5 ms pulse width, 60 V at a frequency of 0.05 Hz) induced twitch contractions of mucosa-free circular muscle strips from the guinea-pig proximal colon which were abolished by atropine (0.3 microM), tetrodotoxin (0.3 microM) or omega-conotoxin GVIA (0.1 microM). 2. Various opioid receptor agonist concentration-dependently inhibited twitches with the following rank order of potency (EC50 values in brackets): U 50488 (0.31 nM) > dermorphin (4.3 nM) = dynorphin A (1-13) (6.2 nM) > [D-Ala2, N-MePhe4, Gly5-ol]-enkephalin (DAMGO, 33.5 nM) = [D-Ala2, D-Leu5]-enkephalin (DADLE, 60 nM) > [D-Pen2, D-Pen2, D-Pen5]-enkepahlin (DPDPE, 1144 nM). 3. Peptidase inhibitors (captopril, thiorphan and bestatin, 1 microM each) did not modify the amplitude of twitches. In the presence of peptidase inhibitors the concentration-response curve to dynorphin A (1-13) was displaced to the left to yield an EC50 of 0.35 nM, comparable to that of the selective kappa receptor agonist, U50488. The curves to the other opioid receptor agonist were unaffected by peptidase inhibitors. 4. DPDPE, DADLE, dermorphin and DAMGO consistently induced a concentration-unrelated transient increase in basal tone and a small and transient facilitation of twitches before development of their inhibitory effect. These transient excitatory effects were not observed upon application of dynorphin A (1-13) or U 50488. The contraction produced by DPDPE (30 nM) was largely inhibited (> 80%) by 1 microM atropine. 5. Twitches suppression induced by dynorphin A (1-13) (30 nM) was partly reversed (46 +/- 8%, n = 6) by naloxone (0.3 microM). The potent and selective kappa opioid receptor antagonist nor-binaltorphimine (Nor-BNI, 3-100 nM)) did not affect the amplitude of twitches and potently antagonized (pKB 9.83 +/- 0.09, n = 10) the inhibitory effect of dynorphin. 6. Naloxone (1-300 nM) concentration-dependently depressed the cholinergic twitches: this depressant effect was largely counteracted in the presence of apamin (0.1 microM) and NG-nitro-L-arginine (30 microM) which potentiated cholinergic twitches on their own. 7. Dynorphin A (1-13) (10 nM, n = 6) did not affect the contractile response to exogenous acetylcholine (1 microM), indicating that depression of evoked twitches occurs prejunctionally. 8. We conclude that multiple opioid receptors modulate cholinergic twitches in the circular muscle of guinea-pig proximal colon. While mu and delta opioid receptor ag

Topics: Acetylcholine; Animals; Colon; Dynorphins; Electric Stimulation; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Leucine-2-Alanine; Enkephalins; Guinea Pigs; In Vitro Techniques; Male; Muscle, Smooth; Naloxone; Oligopeptides; Opioid Peptides; Receptors, Cholinergic; Receptors, Opioid, kappa

The effects of dynorphin-A, dermorphine and morphine on the secretion of transmitter from the toad (Bufo marinus) motor nerve terminal have been determined. Intracellular recordings of miniature end plate potentials (m.e.p.p.s) and evoked end plate potentials (e.p.p.s) were used to estimate quantal content (m) and binomial parameters p and n. Dynorphin-A, and to a lesser extent morphine, decreased (m) while dermorphine had no significant effect on m. Dynorphin-A (ED50 = 24 microM) was 21 times more potent then morphine (ED50 = 510 microM) in decreasing m. The decrease in m produced by dynorphin-A and morphine was accompanied by a greater decrease in the variance (S2) of number of quanta secreted per stimulation over the recording period. The decrease in m produced by dynorphin-A, and to a lesser extent by morphine, is probably mediated by the opiates acting on kappa-opioid receptors.

Topics: Animals; Bufo marinus; Dynorphins; Morphine; Motor Endplate; Naloxone; Narcotics; Oligopeptides; Opioid Peptides; Receptors, Opioid; Synaptic Transmission

The effects of substituting the enkephalin moiety of dynorphin with the dermorphin sequence were studied on the receptor preference, analgesic, and peripheral opioid potencies by using synthetic dermorphin-dynorphin hybrid peptides as the probe. Replacement of the enkephalin moiety of dynorphin with the dermorphin or dermorphin1-5 sequences caused a remarkable increase in analgesic potency, and a 3-6 fold increase in potency of binding against [3H]-dihydromorphine. The potency of receptor binding against [3H]-EKC was also increased by incorporation of the whole dermorphin sequence into the dynorphin molecule. In the presence of NaCl (100 mM), the effect of enhancing binding against [3H]-EKC due to dermorphin substitution disappeared, suggesting the contribution of opioid mu-receptor. Peripheral opioid activities assayed by various smooth muscle preparations showed that dermorphin incorporation caused a decreased in the potency of inhibition of the contractions of the guinea pig ileum and the rabbit vas deferens, no change in potency on the mouse vas deferens, and a marked increase in the inhibition of the rat vas deferens. Among the peripheral opioid activities only that assayed with the rat vas deferens appears to correlate approximately with the analgesic and the receptor binding activities. Judging from the relative potencies obtained from all assays, it is evident that the N-terminal dermorphin moiety, but not the C-terminal dynorphin fragment, dominates the opioid activity and receptor preference of the hybrid peptide.

Topics: Amino Acid Sequence; Analgesics; Animals; Chemical Phenomena; Chemistry; Dynorphins; Guinea Pigs; Mice; Molecular Sequence Data; Muscle, Smooth; Oligopeptides; Opioid Peptides; Rabbits; Rats; Receptors, Opioid; Recombinant Proteins

1. In the isolated electrically driven left atria from reserpine-pretreated guinea-pigs and in presence of 1 microM atropine, electrical field stimulation (EFS) at 10 Hz produces a delayed positive inotropic response (DPIR) involving activation of capsaicin-sensitive afferents. 2. Opioids inhibited the DPIR with the following order of potency: dermorphin greater than [D-Ala2,N-MePhe4, Gly5-ol]-enkephalin (DAGO) greater than or equal to [D-Ala2,D-Leu5]-enkephalin (DADLE) greater than morphine greater than dynorphin A (1-13) greater than [D-Pen2,D-Pen5]-enkephalin (DPDPE). U-50488 was ineffective up to 10 microM. 3. Opioids also inhibited resting inotropism (3 Hz) with the following rank order of potency: DADLE greater than DAGO greater than U-50488 = dynorphin A (1-13) = morphine = DPDPE. 4. Both inhibition of the DPIR and inhibition of resting inotropism were prevented by 10 microM naloxone. 5. Neither dermorphin (0.1 microM) nor DAGO (0.3 microM) or DADLE (1 microM) inhibit responses produced by capsaicin (30 nM) or calcitonin gene-related peptide (3 nM). 6. These findings indicate that capsaicin-sensitive nerves in the guinea-pig atrium are endowed with mu opioid receptors which inhibit transmitter release when sensory nerve terminals are activated by EFS but not by capsaicin.

Topics: Animals; Atropine; Calcitonin Gene-Related Peptide; Capsaicin; Dynorphins; Electric Stimulation; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Leucine; Enkephalin, Leucine-2-Alanine; Enkephalins; Guinea Pigs; Heart; In Vitro Techniques; Male; Myocardial Contraction; Naloxone; Neurons, Afferent; Oligopeptides; Opioid Peptides; Peptide Fragments; Receptors, Opioid; Receptors, Opioid, mu; Reserpine

Biological activity of synthetic beta-endorphin (beta-EP) analogs containing dermorphin or dynorphin-A-(1-13) structure has been investigated using the guinea pig ileum and the vas deferens of the mouse, rat and rabbit. Replacement of NH2-terminal 1-7 segment of camel beta-EP [beta c-EP-(1-7)] with dermorphin caused a great increase in opiate potency of the analog. [Dermorphin (1-7)]-beta c-EP was 120 times more potent than beta c-EP in the guinea pig ileum assay, 49 times more potent in the mouse vas deferens assay; and only 4 times more potent in the rat vas deferens assay. Replacement of NH2-terminal 1-13 segment of human beta-EP [beta h-EP-(1-13)] with dynorphin-A-(1-13) caused an increase in opiate potency in both the guinea pig ileum and rabbit vas deferens assays, a complete loss of potency in the rat vas deferens assay, and no change in the mouse vas deferens assay. In comparison with dynorphin-A-(1-13), the hybrid peptide was less potent in the guinea pig ileum assay as well as in mouse and rabbit vas deferens assay. It is suggested that beta c-EP-(8-31) facilitates the dermorphin moiety to act on opiate mu and delta receptors but not on the epsilon receptor, while beta h-(14-31) reduces the action of dynorphin on mu, delta and kappa receptors.

Topics: Amino Acid Sequence; Animals; beta-Endorphin; Chick Embryo; Dynorphins; Endorphins; Humans; Ileum; Male; Muscle, Smooth; Oligopeptides; Opioid Peptides; Structure-Activity Relationship; Vas Deferens