Opiates produce significant and persistent changes in synaptic transmission; knowledge of the
proteins involved in these changes may help to understand the molecular mechanisms underlying
opiate dependence. Using an integrated quantitative proteomics and systems biology approach, we explored changes in the presynaptic
protein profile following a paradigm of chronic
morphine administration that leads to the development of dependence. For this, we isolated presynaptic fractions from the striata of rats treated with saline or escalating doses of
morphine, and analyzed the
proteins in these fractions using differential isotopic labeling. We identified 30
proteins that were significantly altered by
morphine and integrated them into a
protein-
protein interaction (PPI) network representing potential
morphine-regulated
protein complexes. Graph theory-based analysis of this network revealed clusters of densely connected and functionally related
morphine-regulated clusters of
proteins. One of the clusters contained
molecular chaperones thought to be involved in regulation of neurotransmission. Within this cluster,
cysteine-string protein (CSP) and the
heat shock protein Hsc70 were downregulated by
morphine. Interestingly, Hsp90, a
heat shock protein that normally interacts with CSP and Hsc70, was upregulated by
morphine. Moreover, treatment with the selective Hsp90 inhibitor,
geldanamycin, decreased the somatic signs of
naloxone-precipitated
morphine withdrawal, suggesting that Hsp90 upregulation at the presynapse plays a role in the expression of
morphine dependence. Thus, integration of proteomics, network analysis, and behavioral studies has provided a greater understanding of
morphine-induced alterations in synaptic composition, and identified a potential novel therapeutic target for
opiate dependence.