Opioid receptor antagonists increase hyperalgesia in humans and animals, indicating that endogenous activation of opioid receptors provides relief from acute pain; however, the mechanisms of long-term opioid inhibition of pathological pain have remained elusive. We found that tissue injury produced μ-opioid receptor constitutive activity (MORCA) that repressed spinal nociceptive signaling for months. Pharmacological blockade during the post-hyperalgesia state with MOR inverse agonists reinstated central pain sensitization, and precipitated hallmarks of opioid withdrawal (including cAMP overshoot and hyperalgesia) that required N-methyl-D-aspartate receptor activation of adenylyl cyclase type 1 (AC1). Thus, MORCA initiates both analgesic signaling as well as a compensatory opponent process that generates endogenous opioid dependence. Tonic MORCA suppression of withdrawal hyperalgesia may prevent the transition from acute to chronic pain.

Plasma membrane transporters belonging to the family of Na ϩ /Cl Ϫ -dependent neurotransmitter transporters play an important role in terminating the activity of the monoamine neurotransmitters and of ␥-aminobutyric acid (see Ref. 1). Thus, reuptake of dopamine (DA) 1 from the synaptic cleft by the dopamine transporter (DAT) serves as the major mechanism for terminating dopaminergic neurotransmission in the brain (2). Because the efficiency of DA removal depends on the number of DAT molecules expressed at the plasma membrane, trafficking processes that control transporter distribution in the cell represent a potentially important mechanism by which neurotransmission could be regulated. Newly synthesized DAT acquires glycosylation in the endoplasmic reticulum (ER) and Golgi complex and is then trafficked to the plasma membrane. Typically, a large pool of mature DAT molecules are found at the cell surface of dopaminergic neurons and when DAT is heterologously expressed in tissue culture cells. However, DAT localization can be altered rapidly. Acute exposure of cells to either phorbol esters or substrates reduces the number of plasma membrane DATs and thus DAT function, and this reduction is due to acceleration of DAT endocytosis through a dynamin-dependent mechanism (3-6). Recently, it has been shown that DAT can interact with the anchoring protein PICK 1 and the adaptor protein 8). However, in general, the molecular mechanisms controlling DAT trafficking are not yet well understood.The DAT molecule is predicted to have 12 membrane-spanning sequences with both amino and carboxyl termini oriented intracellularly. The specific function of DAT transmembrane (TM) motifs and termini are not well established. TM domains may play a role in intra-and intermolecular interactions. Many membrane receptors and other integral membrane proteins require dimerization or higher oligomerization for their activity. Several lines of evidence have suggested that monoamine transporters are also dimers or oligomers. Results with dominant negative forms of the serotonin transporter (SERT) and norepinephrine transporter were consistent with this idea (9, 10). Oligomerization of SERT was demonstrated directly by co-immunoprecipitation (11). The results of fluorescence resonance energy transfer (FRET) studies further confirmed that SERT and a ␥-aminobutyric acid transporter (GAT-1) are homo-oligomers (12, 13). Early radiation inactivation studies also suggested that DAT exists as a dimer or oligomer (14,15). Recently, the potential for DAT to exist as dimer or higherorder oligomer in the plasma membrane has been demonstrated using chemical cross-linking (16). More recently, Torres and co-workers (17) reported detection of DAT oligomerization by co-immunoprecipitation. In general, the mechanisms and functional roles of oligomerization of monoamine transporters remain to be defined. However, it has been proposed that SERT oligomerization may be important for its transport activity (9). Oligomerization may be also required for proper traffic...

No abstract

No abstract

In rat caudal artery, contraction to melatonin results primarily from activation of MT 1 melatonin receptors; however, the role of MT 2 melatonin receptors in vascular responses is controversial. We examined and compared the expression and function of MT 2 receptors with that of MT 1 receptors in male rat caudal artery. MT 1 and MT 2 melatonin receptor mRNA was amplified by reverse transcription-polymerase chain reaction from caudal arteries of three rat strains (i.e., Fisher, Sprague-Dawley, and Wistar). Antisense (but not sense) 33 P-labeled oligonucleotide probes specific for MT 1 or MT 2 receptor mRNA hybridized to smooth muscle, as well as intimal and adventitial layers, of caudal artery. In male Fisher rat caudal artery denuded of endothelium, melatonin was 10 times more potent than 6-chloromelatonin to potentiate contraction to phenylephrine, suggesting activation of smooth muscle MT 1 melatonin receptors. The MT 1 /MT 2 competitive melatonin receptor antagonist luzindole (3 M), blocked melatonin-mediated contraction (0.1-100 nM) with an affinity constant (K B value of 157 nM) similar to that for the human MT 1 receptor. However, at melatonin concentrations above 100 nM, luzindole potentiated the contractile response, suggesting blockade of MT 2 receptors mediating vasorelaxation and/or an inverse agonist effect at MT 1 constitutively active receptors. The involvement of MT 2 receptors in vasorelaxation is supported by the finding that the competitive antagonists 4-phenyl 2-acetamidotetraline and 4-phenyl-2-propionamidotetraline, at MT 2 -selective concentrations (10 nM), significantly enhanced contractile responses to all melatonin concentrations tested (0.1 nM-10 M). We conclude that MT 2 melatonin receptors expressed in vascular smooth muscle mediate vasodilation in contrast to vascular MT 1 receptors mediating vasoconstriction.Accumulating evidence indicates that the hormone melatonin regulates vascular tone; however, the nature of the response remains controversial. Both vasoconstrictor and vasodilator responses have been reported; however, data supporting the presence of melatonin receptors is found in some, but not all, vascular beds (Mahle et al., 1997). In isolated rat caudal arteries, nanomolar concentrations of melatonin potentiate contraction induced by either endogenous or exogenous vasoconstrictors (Viswanathan et al