Hindlimb Scratching Research Articles

The protein kinase C activators, phorbol 12-myristate,13-acetate and phorbol 12,13-dibutyrate, are convulsant in the pico-nanomolar range in mice

Administration of phorbol 12-myristate,13-acetate (PMA, 10 fmol-10 nmol) orphorbol 12,13-dibutyrate (PDB, 0.2–495 nmol) (i.c.v.) to mice induced: hindlimb scratching, tremor, myoclonic jerks, hyperlocomotion, clonic seizure, followed by death or recovery. CD 50 values for clonic seizures for PMA and PDB were 1.0 pmol and 1.2 nmol. 4-α-Phorbol (68–686 nmol) was inactive. The effects of PDB (24–247 nmol) were reduced by pretreatment with staurosporine (30 nmol, i.c.v.). Protein kinase C activators are potent convulsants in vivo.

Peripheral 5-carboxamidotryptamine induces hindlimb scratching by stimulating 5-HT 1A receptors in rats

Treatment of rats with 5-carboxamidotryptamine (5-CT) or 5-methoxytryptamine (5-MeOT) induces a hindlimb scratch response. These compounds have high affinity for 5-HT 1A and 5-HT 1D receptors. The selective 5-HT 1A receptor agonist N,N-dipropyl-5-CT (DP-5-CT) also induced hindlimb scratching while the selective 5-HT 1D HD 1D receptor agonist, sumatriptan, did not. 5-CT-induced hindlimb scratching was inhibited dose-dependently by several 5-HT 1A antagonists (BMY 7378, NAN-190, MDL 73005EF and pindobind-5-HT 1A) as well as the non-selective 5-HT antagonist, methiothepin. Pretreatment of rats with the serotonin (5-HT) synthesis inhibitor, p- chlorophenylalanine (PCPA) or the 5-HT depleting agent, reserpine, markedly attenuated 5-CT-induced hindlimb scratching. These data suggest that hindlimb scratching induced by 5-HT agonists may not be centrally mediated but rather may be mediated by a neuronal 5-HT 1A receptor localized outside the blood-brain barrier.

The Behavioural Response to Intrathecal Serotonin is Changed by Acute but not by Repeated Treatment with Zimelidine or Metergoline

The behavioural response to intrathecal serotonin (5-HT) was examined in mice after acute and after withdrawal of repeated treatment with the 5-HT uptake inhibitor zimelidine or the 5-HT receptor antagonist metergoline. Intrathecal 5-HT elicits a response consisting of biting or licking of the lower part of abdomen and reciprocal hindlimb scratching, indicative of nociceptive stimulation. Acute injection of zimelidine (20 mg/kg) significantly increased the response to intrathecal 5-HT (0.25-1 micrograms) whereas a single dose of metergoline (5 mg/kg) completely blocked the response to intrathecal 5-HT (2 micrograms). The behavioural response to intrathecal 5-HT (0.25-2 micrograms) was not significantly changed 48 hr after withdrawal of repeated treatment with zimelidine (2 x 10 mg/kg/day for 14 days) or metergoline (2 x 2.5 mg/kg/day for 14 days). In the present experiments acute zimelidine appeared to increase nociceptive responsiveness, whereas metergoline had the opposite effect. This study does not provide evidence that long-term treatment with zimelidine or metergoline leads to adaptive changes in the response to spinal cord 5-HT receptor stimulation.

Different role of 5-HT 1A and 5-HT 2 receptors in spinal cord in the control of nociceptive responsiveness

The effects of the 5-hydroxytryptamine type-2 (5-HT 2) receptor agonist (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) and the 5-HT 1A agonist (+)-8-hydroxy-2-(di- n-propylamino)-tetralin [(+)-8-OH-DPAT] on nociceptive responsiveness were compared in mice. Intrathecal administration of DOI (5–20 μg) produced a dose-dependent behavioural syndrome, consisting of biting or licking, directed towards the caudal part of the body and reciprocal hindlimb scratching. However, (+)-8-OH-DPAT (5–20 μg) did not produce the biting and scratching behaviour. The response to DOI (20 μg) was reversed by treatment with the substance P receptor antagonist, [ d-Arg 1, d-Trp 7,9, Leu 11]-SP (Spantide) (8 μg). The tail-flick reflex was markedly depressed 5–20 min after administration of (+)-8-OH-DPAT; DOI did not change the tail-flick reflex after 5 min but significantly inhibited the reflex response 10–20 min after injection. The data show that stimulation of 5-HT 2 receptors, but not 5-HT 1A receptors, induced a behavioural syndrome, which may reflect activation of nociceptive pathways. The tail-flick reflex was more markedly inhibited by stimulation of 5-HT 1A than 5-HT 2 receptors. Accordingly, 5-HT 2 and 5-HT 1A receptors seem to have a different function in the modulation of nociceptive responsiveness in the mouse.

Age dependence of 5-methoxytryptamine-induced hindlimb scratching in rats

The influence of age on hindlimb scratching induced in rats by the serotonin agonist, 5-methoxytryptamine (5-MeOT), was studied. The 5-MeOT-induced scratching was strongest in 30-day-old rats and least in 90-day-old rats. Sex hormones do not play a role in these differences since treatment of young male rats with testosterone propionate did not change the scratching response. The same age dependence was found for female rats as for males.

A peripheral 5-HT 1D-like receptor involved in serotonergic induced hindlimb scratching in rats

The pharmacological characteristics of hindlimb scratching induced by serotonergic compounds were studied. We conclude that hindlimb scratching induced by serotonergic compounds is mediated by a serootonin 1D (5-HT 1D) or 5-HT 1D-like receptor outside the blood-brain barrier because hindlimb scratching could be induced by s.c. injection of 5-methoxytryptamine (5-MeOT), 5-carboxamidotryptamine (5-CT), bufotenine, 5-hydroxytryptamine (5-HT) and tryptamine. These compounds have high affinity for 5-HT 1A and 5-HT 1D receptors. The 5-HT 1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), the 5-HT 1C receptor agonist MK 212, and the mixed 5-HT 1C/5-HT 2 receptor agonists (dl)-1-(2,5 dimethoxy-4-iodophenyl)-2-aminopropane (DOI) and quipazine did not induce hindlimb scratching. Rather, the latter compounds attenuated 5-MeOT-induced hindlimb scratching. The 5-HT releasing compounds fenfluramine and p-chloroamphetamine (PCA) inhibited whereas the 5-HT re-uptake inhibitors fluvoxamine and indalpine potentiated 5-MeOT-induced hindlimb scatching, 5-MeOT-induced hindlimb scratching could be inhibited dose dependently by the α- 2-adrenoceptor blockers yohimbine and rauwolsince, which also have high affinity for 5-HT 1D receptors, whereas the α 2-adrenoceptor blocker piperoxan only weakly counteracted hindlimb scratching. Haloperidol, apomorphine, morphine, clonidine and methiothepin strongly attenuated hindlimb scratching, atropine, naloxone and ICS 205930 attenuated it weakly whereas domperidone, methylatropine and mepyramine were inactive in doses up to 10 mg/kg. Hindlimb scratching induced by 5-MeOT was potentiated by the 5-HT receptor antagonists metergoline, methysergide, mesulergine, mianserin, ritanserin and xylamidine. Hindlimb scratching was not induced by i.c.v. injection of 5-MeOT. This lack of i.c.v. activity of 5-MeOT and the effect of xylamidine suggests that the site of action for induction of hindlimb scratching is outside the blood-brain barrier.

Afferent control of central pattern generators: Experimental analysis of scratching in the decerebrate cat

Systematic quantitative analysis of changes in the spinal scratching generator motor activity evoked by tonic and phasic peripheral afferent signals during “fictitious” scratching was carried out in the cat. Correlations between the kinematics of hindlimb scratching movement, sensory inflow, and primary afferent depolarization were investigated. Reliable correlations between the parameters of generator motor activity during fictitious scratching were revealed: they depended on tonic peripheral afferent inflow. The functional role of these dependencies consists of providing stability for aiming the hindlimb to the itch site. It was shown that scratching generator reaction to a phasic sensory signal depended significantly on afferent input, signal intensity, and its arrival phase in the cycle of motor activity. Phase correction of “scratching” rhythm was performed by inhibition of the current stage of “scratching” cycle, the inhibition magnitude depending on the intensity of a sensory signal run along high threshold afferent fibers. The moments in the scratching cycle, in which the afferent signal caused no rearrangement in scratching generator activity, were discovered for all investigated afferent inputs. These moments corresponded to the transitions from one scratching cycle phase to another. Integral afferent activity was distributed unevenly in the cycle during real scratching. The main part of it was observed just in that scratching cycle part which included the above mentioned no rearrangement phase points. The data obtained allowed us to conclude that the scratching generator should be considered as a working program for the motor optimal control system containing the intrinsic model of the controlled object dynamics (e.g. hindlimb scratching movement dynamics), which produces an inner analog of peripheral flow. This inner flow interacts with peripheral afferent inflow just as one of the latter components. Centrally originated modulation of primary afferent depolarization is a result of affecting the depolarization generating system by this inner “sensory” activity. It is the model, with the aid of which the generator can work after deafferentation. The functional organization of a central pattern generator is considered.

Possible involvement of 5HT1D receptors in drug induced hindlimb scratching in rats

Possible involvement of 5HT1D receptors in drug induced hindlimb scratching in rats

Cutaneous dermatomes for initiation of three forms of the scratch reflex in the spinal turtle

The turtle spinal cord produces three forms of the hindlimb scratch reflex. Each scratch form is initiated in response to gentle mechanical stimulation of a distinct set of sites in the periphery, termed the receptive field for that scratch form. The turtle spinal cord consists of 8 cervical segments (C1-C8), 10 dorsal segments (D1-D10), 2 sacral segments (S1, S2), and about 16 caudal segments (Ca1-Ca16). First, we recorded cutaneous afferents in peripheral nerves to reveal the tactile dermatomes of segments D3-D8. These segments innervate regions of the body between the forelimb and hindlimb, directly lateral to their spinal cord segments. Adjacent segments innervate adjacent and partially overlapping regions of the periphery. Second, we used successive spinal cord transections combined with either a) behavioral analysis in turtles with limb movements or b) electroneurographic recordings in immobilized turtles, and mapped the zone of remaining sensibility after each transection to measure the borders of dermatomes D2-Ca2. This technique revealed that adjacent dermatomes are innervated by non-adjacent spinal segments in regions near the hip. Segments D8 and Ca1 innervate adjacent and partially overlapping regions ventral to the hip. There is a similar discontinuity in the innervation of the shell and skin dorsal to the hip. These discontinuities correlate with the innervation of the hindlimb skin by segments D8-Ca1. The rostral scratch receptive field is innervated by sensory afferents entering spinal segments D3-D6; the pocket scratch receptive field is innervated by D6-D8; the caudal scratch receptive field is innervated by S2, Ca1, and more caudal segments. The rostral-pocket transition zone is innervated mainly by one segment, D6; the ventral part of the caudal-pocket transition zone is innervated by two non-adjacent segments, D8 and Ca1. Thus the motor pattern blends elicited by stimulation of sites within the rostral-pocket transition zone must be produced in response to a very different distribution of sensory inputs than the blends elicited by stimulation of sites within the caudal-pocket transition zone.

Cardiovascular Responses Induced by Intrathecal Substance P in the Conscious Freely Moving Rat

Summary: In conscious freely moving rats, administration of 0.65 to 32.5-nmol doses of substance P (SP) into the intrathecal (i.th.) space at the T8-T10 level of the spinal cord increased both mean arterial pressure (MAP) and heart rate (HR) in a dose-related manner. Concomitant with these cardiovascular effects, behavioral responses were observed. Injection of the peptide produced agitation of the rat for ˜1 min as well as intermittent reciprocal hindlimb scratching which lasted for 3-6 min. The cardiovascular responses elicited by SP (6.5 nmol) were not blocked by systemic pretreatment with morphine (3-6 mg/kg), and the effect of SP on HR was potentiated by an i.th. dose of morphine (10 μg) given prior to the injection of SP. A systemic dose of phentolamine (1 mg/kg) blocked the pressor response and a depressor effect appeared, whereas propranolol (1 mg/kg) blocked the HR response. Although catecholamines mediate the spinal action of SP on MAP and HR, bilateral adrenalectomy of the rat 48 h prior to experimentation failed to affect the cardiovascular responses to SP. After transection of the spinal cord at the C3-C4 level, the hindlimb scratching behavior and the pressor response to SP were unaffected but the peak HR response was significantly reduced. These results demonstrate that the rise in MAP and HR evoked by SP is not secondary to perception of a noxious stimulus by the rat and that adrenal medullary catecholamines are not essential for these cardiovascular effects. The direct activation of sympathetic preganglionic fibers in the intermediolateral cell column can account for the pressor response; however, the cardiac response may result from the spinal activation of ascending fibers by SP which can counteract normal baro-reflex activity.

Cardiovascular Responses Induced by Intrathecal Substance P in the Conscious Freely Moving Rat

In conscious freely moving rats, administration of 0.65 to 32.5-nmol doses of substance P (SP) into the intrathecal (i.th.) space at the T8-T10 level of the spinal cord increased both mean arterial pressure (MAP) and heart rate (HR) in a dose-related manner. Concomitant with these cardiovascular effects, behavioral responses were observed. Injection of the peptide produced agitation of the rat for approximately 1 min as well as intermittent reciprocal hindlimb scratching which lasted for 3-6 min. The cardiovascular responses elicited by SP (6.5 nmol) were not blocked by systemic pretreatment with morphine (3-6 mg/kg), and the effect of SP on HR was potentiated by an i.th. dose of morphine (10 micrograms) given prior to the injection of SP. A systemic dose of phentolamine (1 mg/kg) blocked the pressor response and a depressor effect appeared, whereas propranolol (1 mg/kg) blocked the HR response. Although catecholamines mediate the spinal action of SP on MAP and HR, bilateral adrenalectomy of the rat 48 h prior to experimentation failed to affect the cardiovascular responses to SP. After transection of the spinal cord at the C3-C4 level, the hindlimb scratching behavior and the pressor response to SP were unaffected but the peak HR response was significantly reduced. These results demonstrate that the rise in MAP and HR evoked by SP is not secondary to perception of a noxious stimulus by the rat and that adrenal medullary catecholamines are not essential for these cardiovascular effects.(ABSTRACT TRUNCATED AT 250 WORDS)

Substance P(1–7) antagonizes substance P-induced aversive behaviour in mice

Substance P (SP) and its fragments were administered intrathecally into awake mice. SP and C-terminal fragments caused dose-dependent reciprocal hindlimb scratching responses. SP(5–11) was more potent than SP not only in inducing scratching response but also in inducing aversive behaviour including licking and biting. SP(1–7) induced no behavioural reactions. However, when low doses of SP(1–7) (1.0–4.0 pmol) were injected simultaneously with SP or SP(5–11) (0.1 nmol), aversive behaviours induced by SP or SP(5–11) were significantly reduced. These results indicate that SP(1–7) formed endogenously could modulate the actions of SP or SP(5–11) in the spinal cord.

Reciprocal hindlimb scratching and putative subtype-selective muscarinic agents

We have previously proposed that the reciprocal hindlimb scratching (RHS) of mice elicited by intrathecal injection of muscarinic agents is mediated by M 1 muscarinic receptors. In this study, benzhexol potently inhibited pilocarpine-induced RHS (ID 50 = 0.5 ng), methoctramine did not fully block RSH and RS 86 neither produced nor blocked RHS. These data (1) differentiate the three muscarinic agents using this in vivo endpoint, (2) substantiate binding studies characterizing benzhexol and methoctramine as putative M 2 and M 2 antagonists, respectively, and (3) further support using RHS for studying muscarinic agents.

Increased behavioural response to intrathecal serotonin after lesion of serotonergic pathways with 5,7-dihydroxytryptamine seems not to be due to depletion of serotonin.

The behavioural response to intrathecal i.th. serotonin (5-HT) was examined in mice pretreated with the neurotoxin 5,7-dihydroxytryptamine (5,7-DHT) or the 5-HT synthesis inhibitor p-chlorophenylalanine (PCPA) which produce similar extensive depletion of central 5-HT levels. Intrathecal 5-HT (0.4 micrograms) elicited a behavioural response consisting of reciprocal hindlimb scratching and biting or licking of the hindquarters indicative of nociceptive stimulation. The response to i.th. 5-HT was markedly increased 5 days after intracerebroventricular (i.c.v.) injection of 5,7-DHT (80 micrograms base per mouse). On the other hand, 24 h after the last pretreatment injection of PCPA (400 mg kg-1 for 6 consecutive days), the response to i.th. 5-HT was unaltered. These results indicate that i.c.v. 5,7-DHT produces supersensitivity to 5-HT. Since PCPA failed to alter the effect of 5-HT, the supersensitivity seems not to be due to depletion of 5-HT levels after the lesion.

Is substance P a primary afferent neurotransmitter for nociceptive input? IV.2-Amino-5-phosphonovalerate (APV) and [ d-Pro 2, d-Trp 7,9]-substance P exert different effects on behaviors induced by intrathecal substance P, strychnine and kainic acid

Intrathecal (i.t.) pretreatment with 2-amino-5-phosphovovalerate (APV), and NMDA receptor antagonist, diminished scratching with the hindlimbs, vocalization, biting or licking, and myoclonic twitches produced by i.t. strychnine, kainic acid, or morphine in rats. APV did not diminish these behaviors when produced by i.t. substance P (SP). The SP analogue [ d-Pro 2, d-Trp 7,9]-substance P (DPDT) failed to affect any behaviors produced by all of these compounds at non-paralytic doses. At these doses, neither APV nor DPDT changed the thermal and mechanical pain thresholds. These results indicate that SP does not produce scratching at the primary afferent synapse in the spinal cord, that hindlimb scratching in rats is neither necessary nor sufficient to infer the presence of pain in rats, and that SP may not be involved uniquely, if at all, in the transmission of nociceptive information at the primary afferent synapse.

The role of NK-3 and NK-4 receptors in the mediation of reciprocal hindlimb scratching induced by tachykinin receptor agonists

The role of NK-3 and NK-4 receptors in the mediation of reciprocal hindlimb scratching induced by tachykinin receptor agonists

Correlation between the kinematics of hindlimb movement and efferent activity in the decerebrate cat during scratching

Statistical dependence between parameters of hindlimb scratching movements and muscle electrical activity was studied in decerebrated cat during real scratching. Power of electrical signal to scratching and aiming muscles and duration of aiming muscle activity are determining factors of movement parameters (amplitude, velocity and phase duration). Experiments with limb deafferentation had shown that statistical dependences between movement parameters and muscle electrical activity reflect the peculiarities of the scratch generator functioning. Some investigated statistical dependences change under the afferent signal influence.

Evidence that reciprocal hindlimb scratching elicited in mice by intrathecal administration of muscarinic agonists is mediated through M 1 type receptors

Intrathecal (IT) administration of pilocarpine to mice produces a vigorous and dose-related reciprocal hindlimb scratching (RHS) response (ED 50 = 0.6 μg) that is potently blocked by simultaneous IT administration of atropine (ID 50 = 0.002 μg). We now report that RHS is (1) also elicited by the more selective M 1 agonist McN-A-343-11 (ED 50 = 11.6 μg), (2) blocked by the selective M 1 antagonist pirenzepine (ID 50 = 0.001 μg), and (3) is not blocked by the selective M 2 antagonist AF-DX 116 BS at a dose up to 100 times the ID 50 dose of pirenzepine. These results extend our earlier findings and suggest that the RHS elicited in mice by IT injection of muscarinic agonists is mediated through pirenzepine-sensitive (presumably M 1) receptors and that RHS may be a convenient in vivo centrally mediated M 1 endpoint.

Behavioural characterization of substance P-induced nociceptive response in mice

Intrathecal injection of substance P produced a behavioural syndrome, consisting of reciproacal hindlimb scratching and biting or fore- and hind-licking. Pretreatment with either an analogue of substance P, (D-Pro2, D-Trp7,9)-substance P (DPDT-SP) or (D-Arg1, D-Pro2,4, D-Trp7,9, Leu11)-substance P, given intrathecally, reduced the response to substance P in a dose-dependent manner. The behaviour induced by substance P was also inhibited by intrathecal, intracerebroventricular (i.c.v.) or intraperitoneal (i.p.) injection of morphine. Intrathecal or subcutaneous injection of naloxone showed a biphasic effect on substance P response; the substance P-induced nociceptive response was increased by a small dose of naloxone, while it was inversely decreased by a large dose of naloxone. The results with analogues of substance P support the hypothesis that substance P, injected intrathecally, acts directly on substance P receptors in the spinal cord. The nociceptive response induced by substance P appears to be controlled by endogenous opioids in the spinal cord.

Pilocarpine-induced reciprocal hindlimb scratching in mice

Intrathecal (IT) administration of pilocarpine (0.25–2.0 μg) to mice produced a vigorous and dose-related reciprocal hindlimb scratching response that lasted for 10–15 minutes. Neither the intracerebroventricular administration of pilocarpine at up to 10 times the intrathecal ED90 dose nor the subcutaneous administration of 10 mg/kg pilocarpine caused as robust an effects as IT administration. The reciprocal hindlimb scratching produced by the ED90 dose of pilocarpine (2 μg, IT) was antagonised in a dose-related manner by simultaneous IT administration of atropine (ID50=0.002 μg), methysergide (ID50=1.89 μg), the substance P antagonist [D-Pro 2, D-Trp 7,9]-SP (ID50=4.94 μg), and the putative neurokinin B antagonist [D-Pro 2, D-Trp 6,8, Nle 10]-NK (ID50=3.33 μg), but not by yohimbine (5 μg), phentolamine (2 μg), or naloxone (2.5 μg). These results suggest (a) that pilocarpine-induced reciprocal hindlimb scratching is mediated spinally, (b) that the effect is produced by an action of pilocarpine on muscarinic receptors in the spinal cord, and (c) that neurokinin, and perhaps 5-HT, mechanisms might also be involved.