Article Figures & Data
Figures
- Figure 1.
In naive control rats prostaglandin E2 (PGE2) 100 ng induces mechanical hyperalgesia (*p < 0.001 compared with baseline at 30 min (30′, n = 12) that lasts <4 h. When injected with PGE2, μ-opioid receptor agonist DAMGO (1 μg) significantly inhibited PGE2 hyperalgesia (DAMGO/PGE2, †p < 0.001 compared with PGE2 alone, n = 6). Following DAMGOx3 treatment, at the fourth hour, injected with PGE2, DAMGO no longer inhibited PGE2 hyperalgesia (DAMGOx3, DAMGO/PGE2, p = NS, n = 12). However, unlike in the naive rats, PGE2 hyperalgesia was still present at fourth hour in the DAMGOx3-pretreated rats (DAMGOx3, fourth hour DAMGO/PGE2 ¶p < 0.001, n = 12). After three hourly injections of DAMGO, PGE2 alone also produced prolonged (>4 h) hyperalgesia (DAMGOx3, PGE2, ‡p < 0.001, n = 6). The tolerance to DAMGO inhibition of PGE2 hyperalgesia and prolongation of PGE2 hyperalgesia was still observed 5 d after 3 hourly doses of DAMGO (DAMGOx3, day 5, DAMGO/PGE2, both #p < 0.001, n = 6). In all experiments, measurement of paw withdrawal threshold was done at 30 min (30′) and 4 h (4 h) after the administration of the test agents.
- Figure 2.
A, Spinal intrathecal injection of oligodeoxynucleotide (ODN) antisense (PKCε-AS) but not mismatch (PKCε-MM) for PKCε (20 μg in a volume of 20 μl, i.t.), daily for 3 d, prevented the development of tolerance and priming when on the fourth day 3 hourly injections of DAMGOx3 was followed at the fourth hour by DAMGO plus PGE2 (PKCε-ASx3, DAMGOx3, DAMGO/PGE2, *p < 0.001 compared with naive, n = 6 and PKCε-MMx3, DAMGOx3, DAMGO/PGE2, p = NS compared with naive, n = 6). B, Spinal intrathecal injection of PKCε-AS for 3 d prevented priming (hyperalgesia at the fourth hour), when on the fourth day whether PGE2 was injected with DAMGO, following 3 hourly injections of DAMGO (PKCε-ASx3, DAMGOx3, PGE2, p = NS, n = 6), or injected without DAMGO (PKCε-ASx3, PGE2, p = NS, n = 6); in both groups PGE2 hyperalgesia at 30 min was not affected. C, Intradermal injection of PKCεV1-2, a selective PKCε translocation inhibitor peptide (PKCεI, 1 μg), or pertussis toxin (PTX, 10 ng), a Gi-protein inhibitor, reversed (DAMGOx3-induced) tolerance and priming when injected 30 min (30′) before the fourth hourly injection of DAMGO plus PGE2 (DAMGOx3, PKCεI/DAMGO/PGE2, or DAMGOx3, PTX/DAMGO/PGE2, both *p < 0.001, n = 6/group).
- Figure 3.
A, DAMGO did not attenuate PGE2-induced hyperalgesia in carrageenan (CARR, 5 μl of 1% solution)- or protein kinase Cε activator (ψεRACK, 1 μg)-pretreated (5 d prior) rats (CARR, fifth day DAMGO/PGE2, p = NS, n = 6; ψεRACK, fifth day DAMGO/PGE2, p = NS, n = 6). PGE2 hyperalgesia was still present at the fourth hour in the CARR- and ψεRACK-pretreated rats. B, Spinal intrathecal injection of antisense (PKCε-AS) but not mismatch (PKCε-MM) ODN for PKCε (20 μg in a volume of 20 μl), for three d, prevented the development of tolerance and priming when CARR or ψεRACK were injected on the fourth day and the effect of DAMGO on PGE2 hyperalgesia was tested 5 d after the administration of CARR or ψεRACK. (PKCε-ASx3, CARR, DAMGO/PGE2 and PKCε-ASx3, ψεRACK, DAMGO/PGE2, both *p < 0.001, n = 6/group; PKCε-MMx3, CARR, DAMGO/PGE2 and PKCε-MMx3, ψεRACK, DAMGO/PGE2, both p = NS, n = 6/group). C, Intradermal injection of PKCεV1-2, a selective PKCε translocation inhibitor peptide (PKCεI, 1 μg), also reversed tolerance and priming when PKCεI was injected 30 min (30′) before DAMGO/PGE2 on the fifth day following the administration of CARR or ψεRACK (CARR, PKCεI/DAMGO/PGE2 and ψεRACK, PKCεI/DAMGO/PGE2, both *p < 0.001, n = 6/group). D, Intradermal injection of pertussis toxin, a Gi inhibitor (PTX, 10 ng), also reversed tolerance and priming when it was injected 30 min (30′) before DAMGO/PGE2 on the fifth day following the administration of CARR or ψεRACK (CARR, PTX/DAMGO/PGE2 and ψεRACK, PTX/DAMGO/PGE2, both *p < 0.001, n = 6/group).
- Figure 4.
A, Intradermal injection of CTOP (1 μg), a selective μ-opioid antagonist, at the fourth hour following three hourly injections of DAMGO produced hyperalgesia (DAMGOx3, CTOP, *p < 0.001 compared with baseline, n = 6) and this hyperalgesia was still present at the fourth hour after CTOP administration. Injection of PKCε inhibitor (PKCεV1-2) or the Gi-protein inhibitor PTX 30 min (30′) before CTOP reversed CTOP hyperalgesia (DAMGOx3, PKCεI/CTOP, †p < 0.001, n = 6; DAMGOx3, PTX/CTOP, ¶p < 0.001, n = 6). B, Intradermal injection of CTOP in CARR-pretreated rats (5 d prior) produced significant hyperalgesia (CARR, fifth day, CTOP, *p < 0.001, n = 6). Injection of PKCεV1-2 or PTX 30 min (30′) before CTOP reversed CTOP hyperalgesia (CARR, PKCεI/CTOP, †p < 0.001, n = 6; CARR, PTX/CTOP, ¶p < 0.001, n = 6). C, Intradermal injection of CTOP in ψεRACK-pretreated rats (5 d prior) produced significant hyperalgesia (ψεRACK, fifth day, CTOP, *p < 0.001, n = 6). Injection of PKCεV1-2 or the PTX 30 min (30′) before CTOP reversed CTOP hyperalgesia (ψεRACK, PKCεI/CTOP, †p < 0.001, n = 6; ψεRACK, PTX/CTOP, ¶p < 0.001, n = 6).
- Figure 5.
A proposed mechanism relating the transition to chronic pain and the loss of analgesic efficacy. A, In a nociceptor (purple) in the hindpaw of a naive animal, a proinflammatory cytokine (PGE2), causes acute pain (hyperalgesia lasting <4 h) that is mediated by activation of a Gs-protein-coupled receptor for prostaglandin (EP-R) causing increased adenylyl cyclase (AC) activity and activation of PKA, ultimately leading to increased membrane excitability and nerve activity, which underlies acute pain. In the naive animal, μ-opioids act at their receptor (μ-R) to inhibit the second messenger pathway mediating acute pain (μ-R activates a Gi/o protein which inhibits AC). B, The transition to chronic pain and loss of analgesic efficacy is due to a change in second messenger signaling in the nociceptor (priming). This change can be induced by exposure either to a proinflammatory cytokine or to an opioid analgesic. In this primed state, both the prostaglandin and opioid receptors become coupled to PKCε (via Gi/o) and can produce prolonged hyperalgesia, a model of chronic pain and opioid tolerance and dependence.
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