Structure-activity analysis of dynorphin A toxicity in spinal cord neurons: intrinsic neurotoxicity of dynorphin A and its carboxyl-terminal, nonopioid metabolites.
Dynorphin A [dynorphin A (1-17)] is an endogenous opioid peptide that is antinociceptive at physiological concentrations. Levels of dynorphin A increase markedly following spinal cord trauma and may contribute to secondary neurodegeneration. Both kappa opioid and N-methyl-d-aspartate (NMDA) receptor antagonists can modulate the effects of dynorphin, suggesting that dynorphin is acting through kappa opioid and/or NMDA receptor types. Despite these findings, few studies have critically examined the mechanisms of dynorphin A neurotoxicity at the cellular level. To better understand how dynorphin affects cell viability, structure-activity studies were performed examining the effects of dynorphin A and dynorphin A-derived peptide fragments on the survival of mouse spinal cord neurons coexpressing kappa opioid and NMDA receptors in vitro. Time-lapse photography was used to repeatedly follow the same neurons before and during experimental treatments. Dynorphin A caused significant neuronal losses that were dependent on concentration (> or = 1 microM) and duration of exposure. Moreover, exposure to an equimolar concentration of dynorphin A fragments (100 microM) also caused a significant loss of neurons. The rank order of toxicity was dynorphin A (1-17) > dynorphin A (1-13) congruent with dynorphin A (2-13) congruent with dynorphin A (13-17) (least toxic) > dynorphin A (1-5) ([Leu(5)]-enkephalin) or dynorphin A (1-11). Dynorphin A (1-5) or dynorphin A (1-11) did not cause neuronal losses even following 96 h of continuous exposure, while dynorphin A (3-13), dynorphin A (6-17), and dynorphin A (13-17) were neurotoxic. The NMDA receptor antagonist MK-801 (dizocilpine) (10 microM) significantly attenuated the neurotoxic effects of dynorphin A and/or dynorphin-derived fragments except dynorphin A (13-17), suggesting that the neurotoxic effects of dynorphin were largely mediated by NMDA receptors. Thus, toxicity resides in the carboxyl-terminal portion of dynorphin A and this minimally includes dynorphin A (3-13) and (13-17). Our findings suggest that dynorphin A and/or its metabolites may contribute significantly to neurodegeneration during spinal cord injury and that alterations in dynorphin A biosynthesis, metabolism, and/or degradation may be important in determining injury outcome.
Hauser
K F
KF
Department of Anatomy and Neurobiology, University of Kentucky College of Medicine, Lexington, Kentucky 40536-0298, USA.
Knapp
P E
PE
Turbek
C S
CS
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D004399:Dynorphins / Q000737:chemistry / Q000633:toxicity*; D018691:Excitatory Amino Acid Antagonists / Q000633:toxicity; D005333:Fetus; D007700:Kinetics; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D010446:Peptide Fragments / Q000737:chemistry / Q000633:toxicity*; D016194:Receptors, N-Methyl-D-Aspartate / Q000502:physiology*; D017473:Receptors, Opioid, kappa / Q000502:physiology*; D013116:Spinal Cord / Q000166:cytology* / Q000196:embryology; D013329:Structure-Activity Relationship
D018691:Excitatory Amino Acid Antagonists; D010446:Peptide Fragments; D016194:Receptors, N-Methyl-D-Aspartate; D017473:Receptors, Opioid, kappa; D004399:Dynorphins