Introduction
The κ-opioid receptor (κ-OR), that is distributed in the central and
peripheral nervous system mediates the diverse effects of opioids
ranging from pain perception, neurotransmitter release and respiratory
depression to the regulation of a variety of psychiatric disorders
including anxiety and addiction (1). The
κ-OR and its endogenous neuropeptide, dynorphin A, were found to play a
key role in modulating anxiety and stress-related behaviors. Thus,
stress has been shown to increase endogenous dynorphin levels and
up-regulate κ-OR signaling in the nucleus accumbens and the CA3 region
of the hippocampus (2). Ablation of κ-OR
from brain dopaminergic neurons produced anxiolytic effects, confirming
that the regulation of dopaminergic neurotransmission by κ-OR is
critical for manifestation of stress and anxiety
(3).
Recent results also suggest that
κ-ΟR antagonists possess promising antidepressant potential, indicating
that the κ-ΟR and its endogenous neuropeptide ligand, dynorphin A, are
critical mediators of stress and mood disorders with specific κ-ΟR
antagonists being currently tested in phase II clinical trials
(4-6). However, the signaling constituents
responsible for the neurobiological responses that regulate these
physiological phenomena have yet to deduced.
In the brain, the κ-OR is coupled to pertussis toxin sensitive Gi/o
proteins to regulate a variety of downstream effectors including
adenylyl cyclase, K+ and Ca2+channels, phospholipase C, and ERK1/2 phosphorylation
(1, 7). Such
diverse signaling events are mediated not only by interactions with G
proteins but also by other proteins that determine the generated signal
and alterations in the trafficking, targeting and fine tuning of this
receptor (1,
8, 9).
Macroautophagy, herein referred as autophagy, is a highly conserved
degradation process in which proteins and organelles are engulfed in
autophagic vesicles and subsequently targeted for degradation in
lysosomes (10). Autophagy plays an
important role in many organisms upon exposure to stress but is also
considered to be an important physiological mechanism in neuronal
homeostasis. In neurons, autophagy occurs constitutively under
physiological conditions, while impaired autophagy is implicated in many
neurodevelopmental and neurodegenerative disorders
(11). Recent evidence suggests that
autophagy regulates the development and function of axons, dendrites and
synapses, whereas aberrations in neuronal autophagy contribute to
pathological changes. Autophagy alters the kinetics of neurotransmitter
release and the density of synaptic vesicles and is also implicated in
the degradation of postsynaptic receptors such as the
GABAA and AMPA receptors
(11, 12).
Autophagy contributes to such alterations by degrading specific synaptic
proteins involved in spine remodeling and retraction suggesting a direct
link between autophagy and pruning of synaptic connections.
Consequently, the targeting of neuronal autophagy may have great
clinical implications in terms of treatment of various psychiatric
disorders (13,
14).
Additional findings also suggest that GPCRs are direct regulators of
autophagy (15). Previous studies have
shown that exposure of SH-SY5Y and endothelial cells to morphine or
methamphetamine respectively, induces autophagy through the involvement
of opioid receptors by as yet undefined mechanisms
(16, 17).
Moreover, other studies have shown that morphine dysregulates synaptic
balance in the hippocampus via a novel signaling pathway involving
reactive oxygen species, endoplasmic reticulum stress and autophagy
(18). Although opioid receptors and
interacting, Gi/o and Regulators of G protein signaling (RGS) proteins,
were shown to play key roles in neuronal signaling
(8, 19,
20), it is unknown whether κ-OR
activation by specific agonists can induce the autophagic machinery in
neuronal cells and whether these effects could result in synaptic
alterations implicated in stress-related behaviors.
The present study demonstrates a novel signaling pathway via which a
specific representative of opioid receptors, κ-ΟR, induces autophagy
resulting in synaptosomal integrity changes. In addition, we show that
administration of the κ-OR specific antagonist, norBNI to mice, during
acute stress exposure [daily forced swim test (FST)], prevents
autophagy induction and stress-induced degradation of synaptic proteins.
These results provide a novel insight to the role of this receptor in
the regulation of neuronal autophagy and demonstrate that κ-ΟR-mediated
autophagy is responsible for specific changes in stress-induced synaptic
alterations.