[Regulation of spinal nociceptin expression by neuropathic pain]. / Regulation der spinalen Expression von Nociceptin unter neuropathischen Schmerzen.

Nociceptin is the endogenous ligand of a new opioid receptor, the opioid receptor-like-1 (ORL1) receptor. Chronic inflammatory pain causes an increase in the expression of nociceptin and the ORL1 receptor in the dorsal horn of rat spinal cord, thus indicating an involvement of the endogenous nociceptin/ORL1 system in mechanisms of pathological pain. This study investigates the influence of neuropathic pain on the expression of nociceptin using immunohistochemistry. To induce neuropathic pain, a ligation of the sciatic nerve was performed in 12 rats under general anesthesia. A sham operation was performed in 12 rats of the control group. Nerve ligation caused a significant ipsilateral thermal hyperalgesia, a typical sign of neuropathic pain. The paw withdrawal latency was decreased by 45.7+/-4.9% ( p<0.05) at day 5 and by 37.3+/-1.8% ( p<0.05) at day 10. Although hyperalgesia was fully present after 5 days, no changes in nociceptin immunoreactivity in the lumbar spinal cord were detected at this time point. Ten days after nerve ligation, there was a 2.46+/-0.38 fold ( p<0.05) bilateral increase in nociceptin immunoreactivity in the lamina superficiales (I and II), with a notable increase in the inner lamina II at the level of L4. Further investigations are necessary to elucidate the relationship between neuropathic pain, the nociceptin-ORL1 receptor system and potential therapeutic options.

Inhibition of synovial plasma extravasation by preemptive administration of an antiinflammatory irrigation solution in the rat knee.

UNLABELLED: Inflammation and hyperalgesia during surgical procedures are caused by the local release of multiple inflammatory mediators. We used a rat knee joint model of acute inflammation (synovial plasma extravasation) to determine whether preemptive intraarticular irrigation of the antiinflammatory drugs ketoprofen, amitriptyline, or oxymetazoline, alone or in combination, can reduce inflammatory soup-induced plasma extravasation. These three drugs were selected because of their abilities to collectively inhibit the inflammatory effects of biogenic amines, eicosanoid production, and the release of neuropeptides from C-fiber terminals. Synovial perfusion of each one of the three drugs 10 min before, and then in combination with, the inflammatory soup (bradykinin, 5-hydroxytryptamine, and mustard oil) did not reduce plasma extravasation. Similarly, two-drug combinations did not significantly reduce inflammatory soup-induced plasma extravasation. The combination of all three drugs (amitriptyline, ketoprofen, and oxymetazoline) produced a dramatic inhibition of plasma extravasation and was more effective than any of the two-drug combinations. A comparison between the preemptive (10 min before inflammatory soup perfusion) and postinflammatory administration (10 min after inflammatory soup perfusion) showed that the postinflammatory administration of the three-drug solution lost all ability to inhibit inflammatory soup-induced plasma extravasation. We conclude that acute synovial inflammation, which is induced and maintained by multiple mediators, can be substantially inhibited only by the preemptive administration of a drug combination that targets multiple inflammatory mediators. IMPLICATIONS: Preemptive, intraarticular irrigation of a combination of multiple antiinflammatory drugs is a novel and potentially effective method for reducing the synovial inflammatory response, such as that during arthroscopy. In this study, a three-drug combination infusion was statistically superior to one- or two-drug infusions in a rat model.

Bi-directional modulation of 5-hydroxytryptamine-induced plasma extravasation in the rat knee joint by nociceptin.

The role of nociceptin, the endogenous ligand for the opioid receptor-like (ORL1) receptor, in nociceptive processing is controversial. Most studies demonstrate hyperalgesia following supraspinal administration, analgesia following intrathecal and peripheral administration at higher doses, and hyperalgesia following intrathecal and peripheral application at lower doses. The present study investigates the effect of nociceptin on synovial plasma extravasation and its ability to modulate 5-hydroxytryptamine-induced synovial plasma extravasation using the rat knee joint model of inflammation. Nociceptin alone does not alter synovial plasma extravasation from baseline. Nociceptin at concentrations up to 1 nM enhances 5-hydroxytryptamine-induced synovial plasma extravasation (up to 50%) and nociceptin at concentrations above 100 nM inhibits 5-hydroxytryptamine-induced synovial plasma extravasation (down to 45%). The novel, selective ORL1 receptor antagonist J-113397 potently inhibits the pro-inflammatory effect of nociceptin, but only partly inhibits, at higher concentrations, the anti-inflammatory effects of nociceptin.These findings demonstrate a dose-dependent bi-directional effect of nociceptin on inflammatory processes and may indicate a target for novel therapeutics.

An alternatively spliced transcript of the rat nociceptin receptor ORL1 gene encodes a truncated receptor.

Opioid receptor-like protein ORL1, the receptor for the neuropeptide nociceptin (also named orphanin FQ), has two alternatively spliced isoforms in the rat. This alternative splicing event is generated by retaining of intron 3, 81 bases in length, in the mRNA region encoding the second extracellular loop of ORL1. A full-length rat ORL1 receptor has 367 amino acid residues. However, as revealed by sequencing of rat ORL1 genomic DNA and cDNA, the insertion of the unspliced intron 3 brings in an in-frame stop codon and, therefore, creates a truncated open-reading frame encoding only the N-terminal half of ORL1 (from the N-terminus to an alternate extracellular tail C-terminal to the fourth transmembrane domain). The two alternatively spliced transcripts are differentially expressed in tissues. In transfected mammalian cells, the full-length ORL1 displays high-affinity and selective binding for nociceptin, and inhibits the production of cyclic AMP. In contrast, the truncated ORL1 binds nociceptin and other opioid peptides very poorly and non-selectively (affinity in micromolar range), and it does not mediate any inhibitory effects on cyclic AMP production. Apparently, this truncated ORL1 does not function as a receptor for nociceptin or other ligands tested. Such alternative splicing to create a truncated ORL1 receptor might be an endogenous mechanism to negatively regulate nociceptin/ORL1 functions.

Multiple splice patterns of cyclic AMP response element-binding protein mRNA in the central nervous system of the rat.

The alternative splicing pattern of cyclic AMP response element-binding protein (CREB) in the central nervous system (CNS) of the rat has been investigated by an exon-flanking polymerase chain reaction (PCR) strategy. A series of RT-PCR studies with primer pairs flanking all possible alternative splicing sites (corresponding to a genomic region with at least one full exon and two flanking introns) has revealed multiple splice patterns in nine regions of the rat CNS. These include some novel transcripts that lack the phosphorylation site and a segment of the leucine zipper region which is crucial for dimerization and DNA binding. Some isoforms previously reported as testis-specific were also detected in the rat CNS. The findings from this study, which include differential splicing patterns among CNS regions, suggest a complex expression and functional regulation of CREB in the CNS.