Excitability Of Flexor Reflex Research Articles

Pro-nociceptive effects of neuromedin u in rat

The neuropeptide neuromedin U (NMU) has been shown to have significant effects on cardiovascular, gastrointestinal and CNS functions. The peptide was first isolated from the porcine spinal cord and later shown to be present in spinal cords of other species. Little is known about the distribution of neuromedin U receptors (NMURs) in the spinal cord and the spinal action of the peptide. Here we report on the expression of NMURs and a potential role in nociception in the rat spinal cord using a combination of behavioral and electrophysiological studies. Receptor autoradiography showed that NMU-23 binding was restricted to the superficial layers of spinal cord, a region known to be involved in the control of nociception. In situ hybridization analysis indicated the mRNA of NMUR2 was located in the same region (laminae I and IIo) as NMU-23 binding, while the mRNA for NMU receptor 1 was observed in a subpopulation of small diameter neurons of dorsal root ganglia. Intrathecal (i.t.) administration of neuromedin U-23 (0.4–4.0 nmol/10 μl) dose-dependently decreased both the mechanical threshold to von Frey hair stimulation and the withdrawal latency to a noxious thermal stimulus. Mechanical allodynia was observed between 10 and 120 min, peaking at 30 min and heat hyperalgesia was observed 10–30 min after i.t. administration of NMU-23. A similar mechanical allodynia was also observed following i.t. administration of NMU-8 (0.4–4 nmol/10 μl). A significant enhancement of the excitability of flexor reflex was induced by intrathecal administration of NMU-23 (4 nmol/10 μl). Evoked responses to touch and pinch stimuli were increased by 439±94% and 188±36% ( P<0.01, n=6) respectively. The behavioral and electrophysiological data demonstrate, for the first time, a pro-nociceptive action of NMU. The restricted distribution of NMU receptors to a region of the spinal cord involved in nociception suggests that this peptide receptor system may play a role in nociception.

From spinal shock to spasticity

To investigate the adaptational changes in excitability of spinal neuronal circuits below the level of lesion from spinal shock to spasticity in patients with spinal cord injury (SCI). More than 6 months after an acute SCI, clinical follow-up examinations were paralleled by electrophysiologic recordings with tibial nerve stimulation (M-wave, F-wave, H-reflex, and flexor reflex). During spinal shock, the loss of tendon tap reflexes and flaccid muscle tone were associated with low persistence of F-waves and loss of flexor reflexes, whereas H-reflexes were already elicitable. During the transition to spasticity, the reappearance of tendon tap reflexes and muscle tone and the occurrence of spasms was associated with the recovery of F-waves and flexor reflex excitability, whereas the H-to-M ratio remained about stable over months. At later stages (2 to 6 months after SCI) when clinical signs of spasticity became established, the electrophysiologic measures showed little change. In paraplegic patients, in contrast to tetraplegic patients, M-wave and flexor reflex amplitudes even decreased. The late decrease in M-wave and flexor reflex amplitude in paraplegic patients suggests a secondary impairment/degeneration of premotoneuronal circuits and of motoneurons. The divergent course of clinical signs of spasticity and their probable neuronal correlates indicates the occurrence of non-neuronal changes contributing to spasticity.

On the role of galanin in mediating spinal flexor reflex excitability in inflammation

The effects of exogenous and endogenous galanin on spinal flexor reflex excitability was evaluated in rats one to eight days after the induction of inflammation by subcutaneous injection of carrageenan into the sural nerve innervation area. In normal rats, electrical stimulation of C-fibres in the sural nerve elicited a brisk reflex discharge. Conditioning stimulation of C-fibres (1/s) generated a gradual increase in reflex magnitude (wind-up), which was followed by a period of reflex hyperexcitability. Intrathecal galanin dose-dependently blocked reflex hyperexcitability induced by C-fibre conditioning stimulation whereas i.t. M-35, a high-affinity galanin receptor antagonist, moderately potentiated this effect. At one to three days after the injection of carrageenen, when inflammation was at its peak, the magnitude of the reflex was significantly increased and discharge duration became prolonged. However, wind-up and reflex hyperexcitability were significantly reduced. Furthermore, reduced reflex excitability during conditioning stimulation (“wind-down”) and depression of the reflex were sometimes present, which are rarely observed in normal rats. Intrathecal galanin reduced hyperexcitability during inflammation, although its potency was weaker than in normals. However, the galanin receptor antagonist M-35 strongly enhanced wind-up and reflex hyperexcitability, similarly as in normal rats. The baseline flexor reflex, wind-up and C-fibre conditioning stimulation-induced facilitation were normalized four to eight days after carrageenan injection when signs of inflammation were diminishing. Interestingly, intrathecal galanin and M-35 failed to influence spinal excitability. The results suggest a complex functional plasticity in the role of endogenous galanin in mediating spinal excitability during inflammation. There appears to be an enhanced endogenous inhibitory control by galanin on C-afferent input during the peak of inflammation, which may explain the relative ineffectiveness of exogenous galanin. During the recovery phase there may be a reduction in galanin receptors, which may impair the action of endogenous and exogenous galanin. These results further support the notion that galanin is an endogenous inhibitory peptide in nociception.

Subcutaneous carrageenan, but not formalin, increases the excitability of the nociceptive flexor reflex in the rat

The effects of subcutaneous (s.c.) administration of formalin or carrageenan to the rat hindpaw on the ipsilateral nociceptive flexor reflex was examined in decerebrate, spinalized, unanesthetized rats in order to assess the development of spinal hyperexcitability following peripheral inflammation. Carrageenan induced a weak and brief EMG discharge at the time of injection, followed by a gradual and delayed increase in flexor reflex magnitude, becoming significant 120 min postinjection. In addition, carrageenan induced a decrease in response threshold to peripheral mechanical stimulation as early as 30 min postinjection. Formalin induced an immediate and intense motor discharge which usually decreased after 5–10 min and was followed by a second phase of relatively weak motor discharge lasting for 20–70 min. However, the effect of s.c. formalin on the excitability of the flexor reflex was variable with either facilitation or inhibition. The flexor reflex was significantly depressed after formalin from 90 min post-injection. It is suggested that carrageenan induced prolonged spinal sensitization to nociceptive input, possibly through the induction of peripheral inflammation. In contrast, although formalin generated afferent input and evoked motor discharges, it did not induce significant spinal hyperexcitability. Thus, carrageenan, but not formalin, is useful to study central sensitization following peripheral chemical stimulation and/or inflammation.

Biological activities of two endogenously occuring N-terminally extended forms of galanin in the rat spinal cord

The occurence of two N-terminally extended forms of galanin in the porcine adrenal medulla was reported earlier by Bersani et al. (1991). We have synthesized and examined the ability of these two extended forms of galanin, galanin-( − 7 – 29) and galanin-(− 9–29), to bind to galanin receptors in the rat dorsal spinal cord. The effect of intrathecal (i.t.) injection of these peptides on spinal flexor reflex excitability in decerebrate, spinalized, unanesthetized rats was also studied. Both galanin-(− 7–29) and galanin-(−9 –29) fully displaced specific 125I-monoiodo-[Tyr 26]porcine galanin ( 125I-galanin) binding to membranes prepared from rat dorsal spinal cord, with IC 50 values 0.13 and 0.14 μM, respectively. The metabolic half-lives in spinal cord membranes for galanin-(1–29), galanin-(−7 –29) and galanin-(−9 –29) were 117 ± 17, 271 ± 23and185 ± 19 min, respectively. I.t. injection of galanin-(−7-29) and galanin-(−9-29) mimicked the biphasic facilitatory and inhibitory effect of i.t. galanin-(1–29) on flexor reflex excitability and antagonized C-fiber conditioning stimulus-induced spinal cord hyperexcitability, but with reduced potencies compared to galanin-(1–29). We suggest that the N-terminally extended forms of galanin act as endogenous ligands with low agonist activity.

The effect of intrathecal guanfacine and clonidine on the flexor reflex in rats with intact and sectioned sciatic nerves

We examined the effect of intrathecal (i.t.) administration of the non-selective α 2-adrenoceptor agonist clonidine and the α 2A- adrenoceptor selective agonist guanfacine on flexor reflex excitability in decerebrate, spinalized, unanesthetized rats before and after unilateral section of the sciatic nerve. Both guanfacine and clonidine dose dependently depressed the flexor reflex in rats with intact nerves. There was a dramatic increase in the sensitivity of the flexor reflex to the depressive effect of i.t. clonidine 4 to 18 days after sciatic nerve section. In contrast, the sensitivity to i.t. guanfacine increased only slightly. The present results suggest that the markedly increased sensitivity to the analgesic effect of the non-selective α 2-adrenoceptor agonist does not involve up-regulation of α 2A- adrenoceptors . Since radioligand binding data suggested that about 4% of spinal cord α 2- adrenoceptors are of the α 2C type, whereas α 2B- adrenoceptors are not detectable. α 2C- adrenoceptors seem to be a good candidate to mediate the up-regulated response to clonidine in axotomized rats. Thus, α 2C- adrenoceptor agonists may be useful in the treatment of neuropathic pain.

The depressive effect of intrathecal clonidine on the spinal flexor reflex is enhanced after sciatic nerve section in rats

The effect of intrathecal (i.t.) α 2-adrenoceptor agonist, clonidine, on the spinal nociceptive flexor reflex was studied in decerebrate, spinalized, unanesthetized rats with intact sciatic nerves or in rats in which the sciatic nerve had been ipsilaterally sectioned. In rats with intact nerves i.t. clonidine caused a dose-dependent biphasic effect on flexor reflex excitability. At low dose (10 ng) the effect of clonidine was purely facilitatory, whereas with 50–100 ng clonidine the initial facilitation was often followed by reflex depression. Long-lasting, strong reflex depression was observed after i.t. injection of high doses of clonidine (1 and 10 μg). Four to 18 days after sciatic nerve section, the depressive effect of clonidine on the flexor reflex was dramatically enhanced. Depression was frequently observed already with doses of 5 and 10 ng, and maximal depression was reached at 100 ng and 1 μg in axotomized rats. The facilitatory effect of low doses of clonidine on the reflex was also observed, although somewhat less frequently than in normals. The depressive effect of clonidine on the flexor reflex was reversed by the selective α 2-receptor antagonist, atipamezole (20 μg i.t.), in rats with both intact and sectioned sciatic nerves. The present results revealed an increased sensitivity and effectiveness of the depression of spinal reflex mechanisms by i.t. clonidine after sciatic nerve section, which is opposite to the decreased sensitivity to i.t. morphine after axotomy that we observed previously. Since the flexor reflex is evoked by the activation of afférents that may be the source of neuropathic pain after nerve lesion, the results support the usefulness of i.t. clonidine in treating pain associated with peripheral nerve injury.

Intrathecal CP-96,345 blocks reflex facilitation induced in rats by substance P and C-fiber-conditioning stimulation

We have examined the effects of intrathecally (i.t.) administered CP-96,345, a non-peptide NK 1 receptor ligand, on the spinal nociceptive flexor reflex and on the facilitation of this reflex evoked by i.t. substance P (SP), neurokinin A (NKA) and electrical conditioning stimulation of cutaneous C-afferents. CP-96,345 i.t. at 24 pmol-2.4 nmol had no significant effect on flexor reflex excitability. At the highest dose tested (24 nmol), CP-96,345 caused a brief facilitation of the flexor reflex, which was similar to the effect of the vehicle used at this drug concentration. CP-96,345 did not depress the flexor reflex at any dose. In rats with chronically implanted i.t. catheters, CP-96,345 at 24 nmol caused neither motor impairment nor morphological damage to the spinal cord. Pretreatment with CP-96,345 dose dependently and similarly antagonized facilitation of the flexor reflex induced by 7 pmol i.t. SP or by a 20-s, 1-Hz conditioning stimulus train applied to cutaneous C-fibers in the sural nerve innervation area. The vehicle had no effect. The antagonistic effect of CP-96,345 on the SP- and C-fiber reflex facilitation induced by conditioning stimulation became maximal only 20–30 min after the i.t. injection and lasted 3–4 h at the highest dose. CP-96,345 did not significantly block the facilitatory effect of 7 pmol i.t. NKA on the flexor reflex. These results demonstrate that CP-96,345 is a potent, long-lasting and selective antagonist of SP in rat spinal cord. Furthermore, facilitation of the flexor reflex (central sensitization) induced by conditioning stimulation of cutaneous C-afferents is mediated by NK 1 tachykinin receptors, but the NK 1 receptor may not be involved in the transmission of the flexor reflex. CP-96,345 is thus useful in experimental studies of the role of SP in the central nervous system.

Intrathecal galanin blocks the prolonged increase in spinal cord flexor reflex excitability induced by conditioning stimulation of unmyelinated muscle afferents in the rat

The effect of intrathecal (i.t.) galanin (GAL) on the prolonged increase of spinal reflex excitability produced by conditioning stimulation (CS) of unmyelinated muscle afferents was studied in decerebrate, spinalized, unanesthetized rats. A CS train (1 Hz, 20 s) applied to the unmyelinated fibers in the gastrocnemius muscle nerve facilitated the ipsilateral flexor reflex for about 1 h. Pretreatment with GAL (0.1–10 μg) decreased reflex facilitation induced by the gastrocnemius nerve CS. The present results indicate that GAL is capable of blocking the prolonged increase in spinal cord excitability after stimulation of unmyelinated muscle afferents, possibly by antagonizing the facilitatory effect of tachykinins and calcitonin gene-related peptide released at the intraspinal terminals of these fibers.

Intrathecal vasoactive intestinal polypeptide modulates spinal reflex excitability primarily to cutaneous thermal stimuli in rats

The effect of intrathecally (i.t.) injected vasoactive intestinal polypeptide (VIP) on spinal nocifensive flexor reflex excitability was examined in decerebrate, spinalized, unanesthetized rats. The hamstring reflex was evoked by intense cutaneous mechanical or thermal stimulation of the ipsilateral foot. I.t. VIP increased the excitability of the spinal cord to thermal much more than to mechanical stimuli. It is suggested that this neuropeptide is released by thermosensitive cutaneous afferents that respond poorly to mechanical stimuli.

Human flexor reflexes.

One type of flexor reflex, that recorded from the tibialis anterior muscle in response to electrical stimulation of the sole of the foot, was studied in normal subjects and patients with several neurological disorders. Normally this reflex consists of two components, the second of which is related to the actual withdrawal. The first component, normally of lower threshold, is difficult to evoke in patients with chronic spinal cord or discrete cerebral lesions, whereas it has an unusually low threshold and is very clearly seen in those with Parkinson's disease. In patients with spinal cord disease, the exaggerated flexor reflexes are seen at long latencies after relatively small stimuli. During the early phase of recovery from spinal transection, both components may be seen and are, therefore, spinal in origin. Studies of patients with the sensory neuropathy of Friedreich's ataxia suggest that the afferent fibres responsible for these flexor reflexes are the small myelinated fibres. Recovery curves demonstrate very long-lasting changes in flexor reflex excitability in normal subjects and patients with `spasticity' from spinal lesions. This differs in patients with `spasticity' from lesions rostral to the brain-stem. Examples in man of such physiological phenomena as reciprocal inhibition, local sign, habituation, temporal and spatial summation are discussed.