BACKGROUND AND PURPOSEWe hypothesized that proteinase-activated receptor-2 (PAR2)-mediated vasorelaxation in murine aorta tissue can be due in part to the release of adipocyte-derived relaxing factors (ADRFs). EXPERIMENTAL APPROACHAortic rings from obese TallyHo and C57Bl6 intact or PAR2-null mice either without or with perivascular adipose tissue (PVAT) were contracted with phenylephrine and relaxation responses to PAR2-selective activating peptides (PAR2-APs: SLIGRL-NH2 and 2-furoyl-LIGRLO-NH2), trypsin and to PAR2-inactive peptides (LRGILS-NH2, 2-furoyl-OLRGIL-NH2 and LSIGRL-NH2) were measured. Relaxation was monitored in the absence or presence of inhibitors that either alone or in combination were previously shown to inhibit ADRF-mediated responses: L-NAME (NOS), indomethacin (COX), ODQ (guanylate cyclase), catalase (H2O2) and the K + channel-targeted reagents, apamin, charybdotoxin, 4-aminopyridine and glibenclamide. KEY RESULTSEndothelium-intact PVAT-free preparations did not respond to PAR2-inactive peptides (LRGILS-NH2, LSIGRL-NH2, 2-furoyl-OLRGIL-NH2), whereas active PAR2-APs (SLIGRL-NH2; 2-furoyl-LIGRLO-NH2) caused an L-NAME-inhibited relaxation. However, in PVAT-containing preparations treated with L-NAME/ODQ/indomethacin together, both PAR2-APs and trypsin caused relaxant responses in PAR2-intact, but not PAR2-null-derived tissues. The PAR2-induced PVAT-dependent relaxation (SLIGRL-NH2) persisted in the presence of apamin plus charybdotoxin, 4-aminopyridine and glibenclamide, but was blocked by catalase, implicating a role for H2O2. Surprisingly, the PAR2-inactive peptides, LRGILS-NH2 and 2-furoyl-OLRGIL-NH2 (but not LSIGRL-NH2), caused relaxation in PVAT-containing preparations from both PAR2-null and PAR2-intact (C57Bl, TallyHo) mice. The LRGILS-NH2-induced relaxation was distinct from the PAR2 response, being blocked by 4-aminopyridine, but not catalase. CONCLUSIONSDistinct ADRFs that may modulate vascular tone in pathophysiological settings can be released from murine PVAT by both PAR2-dependent and PAR2-independent mechanisms.
Neurophysin has been separated into seven distinct protein fractions. One of these components had no hormone-binding activity. The fractions that had hormone-binding activity were similar in amino acid composition: their cystine content was in the range 11.5-14.5%. The major component, neurophysin-M, was distinguished from the protein isolated by van Dyke by the presence of methionine and the absence of histidine. Neurophysin-M binds both oxytocin and vasopressin with similar affinities.
We studied the effect of pertussis toxin (PT) treatment on the ability of insulin to inhibit lipolysis and to stimulate glucose oxidation in isolated rat adipocytes. In cells maximally modified by PT (100% ADP ribosylation of a 41-kdalton protein in membranes), the ability of insulin to inhibit lipolysis stimulated either by PT alone or in combination with a catecholamine was abolished. In cells wherein ADP ribosylation was submaximal (about 67% modification), a small but variable antilipolytic action of insulin could still be detected. In cells maximally modified by PT, both basal and insulin-stimulated glucose oxidation were markedly reduced (to 10-15% of control levels). However, relative to the basal oxidation level, the fold stimulation by insulin in PT-treated cells was equivalent to the fold stimulation in control cells. Nonetheless, PT treatment caused a rightward shift in the dose-response curve for insulin-stimulated glucose oxidation as well as a small reduction in insulin binding. Our results point strongly not only to a link between the inhibitory guanine nucleotide regulatory complex (Gi) and the antilipolytic action of insulin but also to a link between the Gi complex and the overall regulation of glucose metabolism in adipocytes.
BACKGROUND AND PURPOSEThe PAR2 receptors are involved in chronic arthritis by mechanisms that are as yet unclear. Here, we examined PAR2 activation in the rat knee joint. EXPERIMENTAL APPROACHPAR2 in rat knee joint dorsal root ganglia (DRG) cells at L3-L5, retrogradely labelled with Fluoro-gold (FG) were demonstrated immunohistochemically. Electrophysiological recordings from knee joint nerve fibres in urethane anaesthetized Wistar rats assessed the effects of stimulating joint PAR2 with its activating peptide, 2-furoyl-LIGRLO-NH2 (1-100 nmol·100 mL -1 , via close intra-arterial injection). Fibre firing rate was recorded during joint rotations before and 15 min after administration of PAR2 activating peptide or control peptide. Leukocyte kinetics in the synovial vasculature upon PAR2 activation were followed by intravital microscopy for 60 min after perfusion of 2-furoyl-LIGRLO-NH2 or control peptide. Roles for transient receptor potential vanilloid-1 (TRPV1) or neurokinin-1 (NK1) receptors in the PAR2 responses were assessed using the selective antagonists, SB366791 and RP67580 respectively. KEY RESULTSPAR2 were expressed in 59 Ϯ 5% of FG-positive DRG cells; 100 nmol 2-furoyl-LIGRLO-NH2 increased joint fibre firing rate during normal and noxious rotation, maximal at 3 min (normal; 110 Ϯ 43%, noxious; 90 Ϯ 31%). 2-Furoyl-LIGRLO-NH2 also significantly increased leukocyte rolling and adhesion over 60 min. All these effects were blocked by pre-treatment with SB366791 and RP67580 (P < 0.05 compared with 2-furoyl-LIGRLO-NH2 alone). CONCLUSIONS AND IMPLICATIONSPAR2 receptors play an acute inflammatory role in the knee joint via TRPV1-and NK1-dependent mechanisms involving both PAR2-mediated neuronal sensitization and leukocyte trafficking.
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