Janice L.K. Hylden

The effects of a non-competitive NMDA receptor antagonist, MK-801, on behavioral hyperalgesia and dorsal horn neuronal activity in rats with unilateral inflammation

&NA; The involvement of NMDA receptors in rats with peripheral inflammation and hyperalgesia was evaluated by administration of the non‐competitive NMDA receptor antagonist, MK‐801. Inflammation and hyperalgesia was induced by intradermal injection of complete Freunds adjuvant (CFA) or carrageenan into the left hind paw. The latency of paw withdrawal from a thermal stimulus was used as a measure of hyperalgesia in awake rats. MK‐801 (1.6 mg/kg, i.p., or 31.5 &mgr;g, intrathecal) significantly attenuated thermal hyperalgesia and reduced its duration in comparison to saline‐injected rats (P < 0.05). The receptive field size of nociceptive‐specific and wide‐dynamic‐range neurons in the superficial and deep spinal dorsal horn recorded 24 h after injection of CFA was significantly reduced to 73 ± 6% (P < 0.05, n = 8) and 74 ± 4% (P < 0.05, n = 8) of control values, respectively, by a cumulative dose of 3 mg/kg of MK‐801 (i.v.). MK‐801 (2 mg/kg) prevented the expansion of the receptive fields of dorsal horn neurons recorded 5 ± 0.4 h (n = 5) after intradermal injection of CFA as compared to saline‐injected rats (P < 0.05). MK‐801 had no significant effect on receptive field size of dorsal horn neurons in rats without CFA‐induced inflammation but blocked a transient expansion of the receptive fields induced by 1 Hz, C‐fiber intensity electrical stimulation of the sciatic nerve. The background activity and noxious heat‐evoked response of dorsal horn neurons in rats with CFA‐induced inflammation were primarily inhibited and noxious pinch‐evoked activity was both facilitated and inhibited by the administration of MK‐801. These results support the hypothesis that NMDA receptors are involved in the dorsal horn neuronal plasticity and behavioral hyperalgesia that follows peripheral tissue inflammation.

Expansion of receptive fields of spinal lamina I projection neurons in rats with unilateral adjuvant-induced inflammation: the contribution of dorsal horn mechanisms

&NA; We have physiologically characterized the receptive field properties of lamina I projection neurons with cutaneous input in the lumbar spinal cords of control rats and rats with unilateral adjuvant‐induced inflammation of the hindlimb. The majority of cells recorded in rats with inflamed limbs demonstrated properties uncharacteristic of this cell population in control rats, including large receptive fields, discontinuous receptive fields, responsiveness to deep as well as cutaneous tissues and ongoing or bursting spontaneous activity. Cells with complex receptive fields were encountered from less than 6 h to 5 days after induction of inflammation. This time course correlates with the occurrence of hyperalgesia to thermal stimuli. The contributions of nociceptive afferent sensitization and alterations in the physical environment of peripheral receptors to the observed enlargement of receptive fields were examined by testing the responses of cells to localized electrical and thermal stimuli in the absence and presence of local anesthesia. Nociceptive primary afferents did not demonstrate enlarged receptive fields in this model of inflammation. The results imply that the enlargement of receptive fields cannot be accounted for by peripheral sensitization of peripheral nociceptors or physical changes in the environment of peripheral receptors and must therefore involve changes within the central nervous system.

The intrathecal administration of excitatory amino acid receptor antagonists selectively attenuated carrageenan-induced behavioral hyperalgesia in rats

A single unilateral injection of carrageenan (4.5-6.0 mg in 0.15-0.20 ml saline) into the rat hindpaw induced behavioral hyperalgesia as evidenced by a significant reduction in hindpaw withdrawal latency to a noxious thermal stimulus. The involvement of N-methyl-D-aspartate (NMDA) receptors in this model of hyperalgesia was examined by intrathecal administration of the selective excitatory amino acid (EAA) receptor antagonists: (+/-)-2-amino-5-phosphonopentanoic acid (AP-5), (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), ketamine hydrochloride (ketamine), 7-chlorokynurenic acid (7-Cl kynurenic acid), and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). The effects of dizocilpine maleate (MK-801) were studied under the same conditions and published previously (Ren et al., 1992) and the data are presented for comparison. While the withdrawal latencies of the non-injected paws and of the paws of naive rats were not significantly affected by application of the EAA receptor antagonists at doses tested, the paw withdrawal latencies of the carrageenan-injected paws were elevated dose dependently. The rank order of potency of these agents to reduce hyperalgesia was: MK-801 greater than or equal to AP-5 greater than or equal to CPP = 7-Cl kynurenic acid = ketamine much greater than CNQX greater than 0. In contrast, intrathecal injection of the opioid receptor agonists, [D-Ala2,MePhe4,Gly-ol5]enkephalin (DAMGO, mu-selective) and [D-Pen2,D-Pen5] enkephalin (DPDPE, delta-selective), produced antinociception in both injected and non-injected paws. DAMGO was much more potent, while DPDPE was less potent, than MK-801.(ABSTRACT TRUNCATED AT 250 WORDS)

Spinal opioid analgesic effects are enhanced in a model of unilateral inflammation/hyperalgesia : possible involvement of noradrenergic mechanisms

We have examined the spinal analgesic activity of opioid agonists and antagonists in a model of short term, unilateral, carrageenan-induced inflammation/hyperalgesia. Rats received a single s.c. injection of carrageenan (2-6 mg in saline) 3-24 h prior to testing hindpaw withdrawal latencies to noxious thermal stimuli. Dose-response curves for intrathecally administered agonists with mu- and/or delta-opioid activity were shifted to the left for inflamed hindpaws when compared to contralateral non-inflamed paws. The selective kappa-receptor agonist U-50,488H had no activity in this analgesic assay on either inflamed or non-inflamed paws when administered intrathecally. However, systemic administration of U-50,488H did produce significant elevations of paw withdrawal latencies in inflamed paws. The alpha 2-adrenoceptor agonist clonidine also produced dose-dependent antinociception in the paw withdrawal assay after systemic or intrathecal administration. Inflamed hindpaws were significantly more sensitive to the antinociceptive effect of morphine on inflamed hindpaws was blocked by the opioid antagonist naloxone or the alpha 2-adrenoceptor antagonist idazoxan. The effect of clonidine was only blocked by idazoxan. Antagonists alone had no significant effect on withdrawal latencies. The data indicate that the analgesic action of opioids during conditions of inflammation may depend on an interaction with spinal noradrenergic pathways.

Spinal lamina I projection neurons in the rat: Collateral innervation of parabrachial area and thalamus

A major ascending nociceptive pathway from spinal lamina I to the mesencephalon has previously been reported in the cat, rat and monkey. In the present paper, we have used single and double retrograde labeling techniques to describe this projection system and its collateralization to the thalamus in the rat. Injections of wheat germ agglutinin-horseradish peroxidase into the pontomesencephalic parabrachial area labeled cell bodies bilaterally in lamina I and deeper laminae of the spinal cord. Bilateral lesions of the dorsolateral funiculi at thoracic levels reduced labeling of lamina I neurons caudal to the lesions. Combined injections of fluorescent retrograde tracers into the lateral thalamus and parabrachial area resulted in double labeling of projection neurons in lamina I, lamina IV VIII and the lateral spinal nucleus of the cervical and lumbar enlargements. Double-labeled neurons were especially abundant in lamina I. Thus, we have demonstrated a major lamina I projection through the dorsolateral funiculi to the parabrachial area with significant collateralization to the thalamus. Moreover, since more than 80% of retrogradely labeled lamina I spinothalamic tract cells had collaterals to the parabrachial area we have indirectly demonstrated the presence of a dorsolateral funicular pathway for lamina I spinothalamic neurons in the rat. More lamina I neurons were retrogradely labeled from midbrain injections as compared to thalamic injections. The significance of these findings rest on previous work in this and other laboratories and concerns the understanding of spinal nociceptive mechanisms. Lamina I projection neurons are primarily nociceptive-specific in their response properties and have been shown to project to both the midbrain and thalamus via the dorsolateral funiculus in a number of species. The role of this projection system in nociceptive transmission may lie in its ability to distribute precise information to multiple brain stem sites which in turn activate autonomic or affective responses or descending pain modulatory mechanisms.

Effects of spinal kappa-opioid receptor agonists on the responsiveness of nociceptive superficial dorsal horn neurons.

&NA; Spinal cord application of the &kgr;‐opioid receptor agonists dynorphin (50 nmol) or (1S,2S)U‐50,488H (0.19–1.9 &mgr;mol) produced changes in the excitability of some superficial dorsal horn nociceptive neurons. One‐third of the cells exhibited expansion of their receptive fields as defined using mechanical stimuli following a spinal &kgr; agonist (dynorphin or U‐50,488H); receptive field expansions were of the same order as those observed immediately after a conditioning electrical stimulus applied to a peripheral nerve. In addition, spinal U‐50,488H produced changes in mechanical and thermal thresholds of the majority of superficial dorsal horn neurons. These changes were dose‐dependent. Facilitation of responses occurred at lower doses and inhibition occurred primarily at higher doses, but these effects were not reversed by subsequent administration of naloxone. The data are consistent with the hypothesis that one action of increases in spinal dynorphin levels due to peripheral inflammation, tissue injury or nerve damage, is to contribute to enhanced neuronal excitability in superficial dorsal horn neurons.

Spinal lamina I neurons projecting to the parabrachial area of the cat midbrain

We have observed a population of lamina I neurons in the cat that has projections to the parabrachial area (parabrachial and cuneiform nuclei). A subpopulation of these neurons also projects to the contralateral thalamus. The majority of projecting cells responded exclusively to noxious stimuli, a few wide-dynamic-range neurons were also observed. Conduction velocities for antidromic activation from the midbrain ranged from 1 to 18 m/s. We stained 14 cells intracellularly with horseradish peroxidase. These findings suggest that a major nociceptive projection pathway originating in lamina I and terminating in the parabrachial area exists in the cat.

Peripheral inflammation is associated with increased dynorphin immunoreactivity in both projection and local circuit neurons in the superficial dorsal horn of the rat lumbar spinal cord

The present study combined the retrograde transport of fluorescent tracers with the immunocytochemical identification of dynorphin A(1-8) in superficial dorsal horn neurons to examine whether peripheral inflammation-induced dynorphin increases are found in local circuit neurons only or also in neurons projecting at least to the caudal mesencephalon. Evidence is presented that complete Freunds adjuvant-induced inflammation produces a large increase in the number of lamina I dynorphin-containing projection and non-projection neurons, and in the number of lamina II dynorphin local circuit neurons.

Lamina I Spinomesencephalic Neurons in the Cat Ascend via the Dorsolateral Funiculi

Spinomesencephalic tract neurons in the cat spinal cord were retrogradely labeled following injections of wheatgerm agglutinin conjugated to horseradish peroxidase into the region of the midbrain parabrachial area. Labeled cell bodies were concentrated in lamina I, bilaterally. A more scattered distribution was observed in lamina V and deeper laminae. Bilateral lesions of the dorsolateral funiculus (DLF) at thoracic levels eliminated labeling of lamina I neurons below the lesions, but had no effect on the labeling of the neurons in deeper laminae. Injections of colchicine into the spinal white matter caused the label to accumulate intra-axonally and revealed labeled axons bilaterally in the DLF and ipsilaterally in the ventrolateral and ventral funiculi.

Serotonin innervation of physiologically identified lamina I projection neurons

Physiologically characterized lamina I projection neurons, including antidromically activated spinomesencephalic and spinothalamic tract cells, were intracellularly stained with HRP and then processed and examined for serotonin-immunoreactive contacts. We observed cells with both high and low densities of contacts from serotonergic axons. Serotonin contacts were found on both nociceptive-specific and wide-dynamic-range projection neurons. The density of contacts did not appear to correlate with any physiological characteristic.