M. Satoh

Separate involvement of the spinal noradrenergic and serotonergic systems in morphine analgesia: the differences in mechanical and thermal algesic tests

Experiments using 3 analgesic tests, the tail-pinch, hot-plate and tail-flick methods, were done to evaluate the roles of the spinal noradrenergic and serotonergic systems in the production of morphine analgesia in rats. To deplete noradrenaline or serotonin in the spinal cord, 6-hydroxydopamine or 5,6-dihydroxytryptamine was given intrathecally. 6-Hydroxydopamine suppressed the antinociceptive effects of morphine injected systemically or intracerebrally (into the nuclei reticularis gigantocellularis and paragigantocellularis or into the periaqueductal gray matter) in the tail-pinch test, but not significantly in the hot-plate and tail-flick tests. Conversely, 5,6-dihydroxytryptamine suppressed the antinociceptive effects of systemically given morphine in the hot-plate test, but not significantly in the tail-pinch and the tail-flick tests. The results not only provide further evidence for the involvement of the descending inhibitory systems in morphine antinociception, but also show that the extent of participation of the spinal noradrenergic and serotonergic systems in the effects of morphine has to be carefully assessed as different analgesic tests (tail-pinch, tail-flick and hot-plate) yield different results.

Stimulus specificity of peripherally evoked substance P release from the rabbit dorsal horn in situ

Characteristics of in situ substance P release from the lumbar dorsal horn were investigated in decerebrated rabbits. Noxious mechanical stimuli produced by pinching the skin of a hind leg ipsilateral to the perfusion site remarkably and significantly increased the release of immunoreactive substance P, which was identified as substance P itself, using separation with high-performance liquid chromatography and radioimmunoassay. The noxious pinch did not affect the release of immunoreactive substance P, when applied to the contralateral hind leg. Both the basal and pinch-evoked release of immunoreactive substance P were largest in the dorsolateral part of the dorsal horn. The pinch-evoked release of immunoreactive substance P was abolished when the dorsal horn was perfused with a Ca2+-free medium containing 7 mM Mg2+ or with a medium with 10 microM tetrodotoxin added. The evoked release of immunoreactive substance P was also abolished following pretreatment of a stimulated region with the local anesthetic dibucaine, a procedure which inhibited the pinch-evoked aversive behavior in freely-moving rabbits. Among a variety of natural stimuli applied to the hind leg, noxious pinch and a subcutaneous injection of formaldehyde solution significantly evoked the release of immunoreactive substance P from the dorsal horn. The most intense heat or scalding stimulation increased the immunoreactive substance P release in two out of five experiments. However, other natural stimuli such as ice-cold, warm, noxious heat and innocuous mechanical stimuli produced no apparent changes in the release of immunoreactive substance P. These results suggest that among the noxious stimuli, only mechanical and inflammatory but not thermal stimuli lead to a release of substance P from the primary afferent terminals in the dorsal horn. The present findings suggest that, at least in rabbits, substance P-containing primary afferents have high-threshold mechanoreceptors. Substance P may participate in the transmission of information related to noxious mechanical and inflammatory stimulation from the periphery to the dorsal horn.

Antinociception induced in rats by intrathecal administration of antiserum against calcitonin gene-related peptide

To determine whether or not endogenous calcitonin gene-related peptide (CGRP) participates in pain transmission in the spinal dorsal horn, effects of an intrathecal injection of anti-CGRP antiserum on nociceptive threshold in the paw-pressure test was examined in non-arthritic and adjuvant arthritic rats. An intrathecal injection of the antiserum increased the nociceptive threshold in non-arthritic animals, while 0.9% saline, pre-immune serum and antiserum, previously absorbed by synthetic CGRP, were without effect. Adjuvant arthritic rats showed a hyperalgesia, and improvement occurred with intrathecal injection of the antiserum. Saline and absorbed antiserum were without effect on the hyperalgesia. These results suggest that the endogenous CGRP present in primary afferents probably has a facilitating function in nociceptive transmission in the spinal dorsal horn.

Effects of morphine on noxious stimuli-induced release of substance P from rabbit dorsal horn in vivo

We investigated the effects of morphine on the noxious mechanical stimuli-induced release of immunoreactive substance P (iSP) from the rabbit dorsal horn in vivo. Systemic morphine in a dose of 10 mg/kg, but not 1 mg/kg, inhibited the iSP release induced by noxious stimuli, although both dosages inhibited the nociceptive responses of rabbits to the stimuli. The inhibitory effect of morphine (10 mg/kg) on the iSP release was partially or completely antagonized by the local application of naloxone or prazosin, respectively, to the dorsal horn. The local application of methysergide did not affect the morphine action. The inhibitory effect of morphine (10 mg/kg) on the iSP release was diminished in acute spinal animals. These results suggest that the inhibition of the evoked release of substance P participates only in the action of the larger dose of morphine, and that this inhibition of the substance P release may be mediated by opioid receptors and the noradrenergic system, but not the serotonergic system, in the spinal cord.

Antagonism by phenoxybenzamine of the analgesic effect of morphine injected into the nucleus reticularis gigantocellularis of the rat

Abstract Effects of some monoaminergic blockers on the analgesia induced by morphine microinjection into the nucleus reticuloris gigantocellularis were studied in rats. Pretreatment with phenoxybenzamine (0.1–1 mg/kg i.p.), but neither propranolol (5 mg/kg i. p.) nor methysergide (i mg/kg i.p.), suppressed the analgesic effect of morphine. The morphine analgesia was also inhibited by intrathecal administration of phenoxybenzamine (10 μg) at the level of the lumbar spinal cord. These results emphasise the important role of the spinal noradrenergic system, mediated by α-adrenoceptors, in the production of analgesia by morphine injected into the nucleus reticularis gigantocellularis.

Involvement of medullary opioid-peptidergic and spinal noradrenergic systems in the regulation of formalin-induced persistent pain

To confirm that endogenous opioid-peptidergic systems and monoaminergic systems participate in the regulation of pain, the effects of a narcotic antagonist naloxone, inhibitors of enkephalin-degrading enzymes and monoaminergic blockers on persistent pain induced by formalin were investigated. Rats were given formalin into the plantar region of the left hind-paw and the duration of licking responses was measured at periods of 0-10 min (period I), 10-30 min (period II), 30-60 min (period III) and 60-120 min (period IV). Naloxone was administered systemically through mini-osmotic pumps and there was an enhanced licking response at period III and a tendency toward enhancement at period IV. The duration of the licking response at period III was increased when naloxone was given into the fourth ventricle in a dose of 30 nmol/rat, but not when it was injected into the lumbo-sacral subarachnoid space in doses of 30 and 300 nmol/rat. Thiorphan and bestatin injected simultaneously into the fourth ventricle (50 micrograms/rat) depressed the licking response at period III. Intrathecal injection of phentolamine significantly enhanced the licking response at periods I-III and produced a similar, but weaker effect at period IV. Intrathecal injection of propranolol and methysergide did not affect the response. These results suggest that opioid-peptidergic systems in the brain stem and noradrenergic systems in the spinal cord (via alpha-adrenoceptors) participate in the regulation of pain.

Met-enkephalin and morphine but not dynorphin inhibit noxious stimuli-induced release of substance P from rabbit dorsal horn in situ.

The effects of locally applied opioids on the release of immunoreactive Substance P (iSP), induced by mechanical stimuli, from the dorsal horn of the rabbit in situ, were investigated. Morphine and met-enkephalin (met-enk), but not dynorphin A (1-17) (DYN), in a concentration of 10 microM, significantly inhibited the evoked release. These inhibitory effects of morphine and met-enkephalin were antagonized by the local application of naloxone (10 microM) to the dorsal horn. These results suggest that the inhibition of the release of Substance P induced by noxious mechanical stimuli may be mediated by mu and delta, but not by kappa opioid receptors.

Opioidergic inhibition of capsaicin-evoked release of glutamate from rat spinal dorsal horn slices

We investigated the effects of opioid agonists on the capsaicin-evoked release of glutamate from nociceptive primary afferent fibers of the rat (6-8 weeks) using a fluorometric on-line continuous monitoring system for glutamate. In the presence of 0.3 microM tetrodotoxin, the application of 3 microM capsaicin to spinal dorsal horn slices produced an evoked glutamate release (55.9 +/- 4.02 pmol.mg-1 protein, n = 15). DAMGO ([D-Ala2,N-Me-Phe4,Gly5-ol]enkephalin; 0.3-10 microM) and morphine (1-30 microM), mu-opioid agonists, produced a concentration-dependent reduction (approximately 85 and approximately 77% reduction, respectively) in the capsaicin (3 microM)-evoked release of glutamate. These inhibitory effects were significantly antagonized by naloxone (1 microM). DPDPE ([D-Pen2,5]enkephalin; 1-10 microM), a delta-opioid agonist, also reduced the capsaicin-evoked release in a concentration-dependent manner (approximately 59% reduction). Naltrindole (1 microM), a selective delta-antagonist, significantly antagonized the inhibitory effect of DPDPE (10 microM). In contrast, neither U-50,488H (1-10 microM) nor U-69,593 (10 microM), kappa-opioid agonists, had any effects on the evoked release of glutamate. These results suggest that mu-, and delta-opioid agonists modulate pain transmission in the spinal dorsal horn, at least in part, by inhibiting the release of glutamate from capsaicin-sensitive primary afferents.

Effect of morphine on bradykinin-induced unitary discharges in the spinal cord of the rabbit

Abstract In the rabbit, bradykinin (1–2 μg) injected iinto the femoral artery induced a marked increase in the firing rates of about half of the dorsal horn neurons which responded to noxious stimulation, such as a squeeze of the skin. This bradykinin-indduced response was markedly inhibited by small doses (2 mg/kg) of morphine in intact rabbits, but was not inhibited in spinal rabbits. These results suggest that morphine, in small doses, inhibits the sensory transmission of bradykinin-induced pain at the dorsal horn of the spinal cord, through its facilitatory action on the descending inhibitory mechanism.

Effects of tyrosyl-arginine (kyotorphin), a new opioid dipeptide, on single neurons in the spinal dorsal horn of rabbits and the nucleus reticularis paragigantocellularis of rats

The effects of a new endogenous opioid dipeptide (Tyr-Arg), kyotorphin, on single unit activities recorded from the lamina V type neurons in the spinal dorsal horn and the neurons in nucleus reticularis paragigantocellularis (NRPG) of the medulla oblongata were investigated in the rabbit and rat, respectively. Microelectrophoretically applied kyotorphin predominantly depressed the lamina V type neurons but excited the NRPG neurons. Such predominant effects were antagonized by naloxone. These results suggest that kyotorphin has qualitatively similar actions to those of enkephalins in both central regions examined.