Spinal dopaminergic projections control the transition to pathological pain plasticity via a D1/D5-mediated mechanism.

The mechanisms that lead to the maintenance of chronic pain states are poorly understood, but their elucidation could lead to new insights into how pain becomes chronic and how it can potentially be reversed. We investigated the role of spinal dorsal horn neurons and descending circuitry in plasticity mediating a transition to pathological pain plasticity suggesting the presence of a chronic pain state using hyperalgesic priming. We found that when dorsal horn neurokinin 1 receptor-positive neurons or descending serotonergic neurons were ablated before hyperalgesic priming, IL-6- and carrageenan-induced mechanical hypersensitivity was impaired, and subsequent prostaglandin E2 (PGE2) response was blunted. However, when these neurons were lesioned after the induction of priming, they had no effect on the PGE2 response, reflecting differential mechanisms driving plasticity in a primed state. In stark contrast, animals with a spinally applied dopaminergic lesion showed intact IL-6- and carrageenan-induced mechanical hypersensitivity, but the subsequent PGE2 injection failed to cause mechanical hypersensitivity. Moreover, ablating spinally projecting dopaminergic neurons after the resolution of the IL-6- or carrageenan-induced response also reversed the maintenance of priming as assessed through mechanical hypersensitivity and the mouse grimace scale. Pharmacological antagonism of spinal dopamine D1/D5 receptors reversed priming, whereas D1/D5 agonists induced mechanical hypersensitivity exclusively in primed mice. Strikingly, engagement of D1/D5 coupled with anisomycin in primed animals reversed a chronic pain state, consistent with reconsolidation-like effects in the spinal dorsal horn. These findings demonstrate a novel role for descending dopaminergic neurons in the maintenance of pathological pain plasticity.

Development of highly potent protease-activated receptor 2 agonists via synthetic lipid tethering.

Protease-activated receptor-2 (PAR2) is a G-protein coupled receptor (GPCR) associated with a variety of pathologies. However, the therapeutic potential of PAR2 is limited by a lack of potent and specific ligands. Following proteolytic cleavage, PAR2 is activated through a tethered ligand. Hence, we reasoned that lipidation of peptidomimetic ligands could promote membrane targeting and thus significantly improve potency and constructed a series of synthetic tethered ligands (STLs). STLs contained a peptidomimetic PAR2 agonist (2-aminothiazol-4-yl-LIGRL-NH2) bound to a palmitoyl group (Pam) via polyethylene glycol (PEG) linkers. In a high-throughput physiological assay, these STL agonists displayed EC50 values as low as 1.47 nM, representing a ∼200 fold improvement over the untethered parent ligand. Similarly, these STL agonists were potent activators of signaling pathways associated with PAR2: EC50 for Ca(2+) response as low as 3.95 nM; EC50 for MAPK response as low as 9.49 nM. Moreover, STLs demonstrated significant improvement in potency in vivo, evoking mechanical allodynia with an EC50 of 14.4 pmol. STLs failed to elicit responses in PAR2(-/-) cells at agonist concentrations of >300-fold their EC50 values. Our results demonstrate that the STL approach is a powerful tool for increasing ligand potency at PAR2 and represent opportunities for drug development at other protease activated receptors and across GPCRs.