Changes in gene expression and neuronal phenotype in brain stem pain modulatory circuitry after inflammation. J Neurophysiol 87: 750 -760, 2002; 10.1152/jn.00534.2001. Recent studies indicate that descending pain modulatory pathways undergo time-dependent changes in excitability following inflammation involving both facilitation and inhibition. The cellular and molecular mechanisms of these phenomena are unclear. In the present study, we examined N-methyl-Daspartate (NMDA) receptor gene expression and neuronal activity in the rostral ventromedial medulla (RVM), a pivotal structure in pain modulatory circuitry, after complete Freund's adjuvant (CFA)-induced hindpaw inflammation. The reverse transcription polymerase chain reaction analysis indicated that there was an upregulation of mRNAs encoding NMDA receptor subunits in the RVM after inflammation. The increase in the NR1, NR2A, and NR2B receptor mRNAs started at 5 h, maintained for 1-7 days (P Ͻ 0.05-0.001) and returned to the control level at 14 days after inflammation. Western blot analysis indicated that the protein translation products of the NR2A subunit were also increased (P Ͻ 0.01). In single-unit extracellular recordings, we correlated RVM neuronal activity with the paw withdrawal response in rats with inflammation. We describe these RVM cells as on-, off-, and neutral-like cells because of their similarity to previous studies in which neuronal responses were correlated with tail-flick nocifensive behavior in the absence of inflammation. In contrast to previous studies in the absence of inflammation, using tail flick as a behavioral correlate, fewer off-like cells in naïve animals exhibited a complete pause before the paw withdrawal to a noxious thermal stimulus. The percentage of cells showing a pause of activity after noxious stimulation was further reduced after inflammation ( 2 P Ͻ 0.0001 vs. naïve rats). Continuous neuronal recordings (3-6.5 h) revealed a phenotypic switch of RVM neurons during the development of inflammation: 11/15 neutral-like cells initially unresponsive to noxious stimuli exhibited and maintained response profiles characteristic of pain modulatory neurons (became off-like: n ϭ 5; became on-like: n ϭ 6). Neutral-like cells recorded in noninflamed animals did not show response profile changes during continuous recordings (5-5.5 h, n ϭ 7). A population study (n ϭ 165) confirmed an increase in on-and off-like cells and a decrease in neutral-like cells at 24 h after inflammation as compared with naïve rats (P Ͻ 0.001). These results suggest that enhanced NMDA receptor activation mediates time-dependent changes in excitability of RVM pain modulatory circuitry. The functional phenotypic switch of RVM neurons provides a novel mechanism underlying activity-dependent plasticity and enhanced net descending inhibition after inflammation.

Treatment of pain in newborns is associated with problematic drug side effects. Previous studies demonstrate that an intraoral infusion of sucrose and other sweet components of mother's milk are effective in alleviating pain in infant rats and humans. These findings are of considerable significance, as sweet tastants are used in pain and stress management in a number of clinical procedures performed in human infants. The ability of sweet stimuli to induce analgesia is absent in adult rats, suggesting that this is a developmentally transient phenomenon. However, the age range over which intraoral sucrose is capable of producing analgesia is not known. We investigated the effects of intraoral sucrose (7.5%) on nocifensive withdrawal responses to thermal and mechanical stimuli in naive and inflamed rats at postnatal days (P) P0-21. In some rats, Complete Freund's adjuvant (CFA) was injected in a fore- or hindpaw to produce inflammation. In non-inflamed animals, for noxious thermal stimuli, sucrose-induced analgesia emerged at P3, peaked at P7-10, then progressively declined and was absent at P17. For mechanical forepaw stimuli, sucrose-induced analgesia emerged, and was maximal at approximately P10, then declined and was absent at P17. By contrast, maximal sucrose-induced analgesia for mechanical hindpaw stimuli was delayed (P13) compared to that for the forepaw, although it was also absent at P17. In inflamed animals, sucrose reduced hyperesthesia and hyperalgesia assessed with mechanical stimuli. Sucrose-induced analgesia in inflamed animals was initially present at P3 for the forepaw and P13 for the hindpaw, and was absent by P17 for both limbs. Intraoral sucrose produced significantly greater effects on responses in fore- and hindpaws in inflamed rats than in naive rats indicating that it reduces hyperalgesia and allodynia beyond its effects on responses in naive animals. These findings support the hypothesis that sucrose has a selective influence on analgesic mechanisms and that an enhanced sucrose effect takes place in hyperalgesic, inflamed animals as compared to naive animals. Taken together, these results indicate that intraoral sucrose alleviates transient pain in response to thermal and mechanical stimuli, and also effectively reduces inflammatory hyperalgesia and allodynia. Sucrose-induced analgesia is age-dependent and limited to the pre-weaning period in rats. The age-dependency of sucrose-induced analgesia and its differential maturation for the fore- and hindpaw may be due to developmental changes in endogenous analgesic mechanisms and developmental modulation of the interaction between gustatory and pain modulatory pathways.

Brain stem descending pathways modulate spinal nociceptive transmission. In a lightly anesthetized rat preparation, we present evidence that such descending modulation undergoes time-dependent changes following persistent hindpaw inflammation. There was an initial decrease and a subsequent increase in the excitability of neurons in the rostral ventromedial medulla (RVM) involving facilitation and inhibition. These changes were most robust after stimulation of the inflamed paw although similar findings were seen on the non-inflamed paw and tail. The enhanced descending modulation appeared to be mediated by changes in the activation of the NMDA excitatory amino acid receptor. These findings demonstrate the dynamic plasticity of the pain modulating pathways in response to persistent tissue injury.

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The role for excitatory amino acids (EAAs) in the rostral ventromedial medulla (RVM) in descending pain modulation after persistent noxious input is unclear. In an animal model of inflammatory hyperalgesia, we examined the effects of intra-RVM microinjection of EAA receptor agonists and antagonists on paw withdrawal and tail-flick responses in lightly anesthetized rats. N-Methyl-D-aspartate (NMDA) produced effects that depended upon the postinflammatory time period. At 3 h postinflammation, NMDA induced facilitation at a lower dose (10 pmol) and inhibition at a higher dose (1000 pmol). At 24 h postinflammation, NMDA (0.1-1000 pmol) produced a dosedependent inhibition. The facilitation and inhibition, respectively, were attenuated significantly by the preadministration of an NMDA receptor antagonist, DL-2-amino-5-phosphonovaleric acid (APV) (10 pmol, P Ͻ 0.05), to the same site. Intra-RVM microinjection of ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) (0.1-100 pmol) produced dose-dependent inhibition at both 3 and 24 h postinflammation that was blocked by the preadministration of an AMPA/kainate receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline (100 pmol, P Ͻ 0.05). Unexpectedly, AMPA-produced inhibition was also significantly attenuated by preadministration of APV (10 pmol, P Ͻ 0.05). Compared with 3 h postinflammation, both NMDA and AMPA showed a leftward shift in their dose-response curves at 24 h postinflammation. These results demonstrate that NMDA and AMPA receptors in the RVM are involved in the descending modulation after inflammatory hyperalgesia. There is a time-dependent increase in EAA neurotransmission in the RVM after inflammation and NMDA receptors play an important role in AMPA-produced inhibition.