Matthias Pawlak

Increase of meningeal blood flow after electrical stimulation of rat dura mater encephali: mediation by calcitonin gene‐related peptide

1 The dura mater encephali of the rat was exposed and the blood flow around branches of the medial meningeal artery was monitored with a laser Doppler flowmeter. Changes in the meningeal blood flow (MBF) following electrical stimulation of the dura mater at a parasagittal site were registered. The effects of human calcitonin gene‐related peptide (h‐αCGRP) and the CGRP antagonist (h‐αCGRP8–37) on the MBF were tested.

Sensitisation of articular afferents in normal and inflamed knee joints by substance P in the rat

To examine whether substance P (SP) influences the response properties of fine articular afferents in normal and acutely inflamed joints, single units were recorded from the rat knee during normal and noxious joint rotations. Only three of 39 units were activated by a single bolus injection of 0.1 mM SP. However, 35% (7/20) of the nerve fibres from the normal joint and 21% (4/19) of the units from the inflamed joint significantly increased their responses to movements after the SP injection. This was most prominent during noxious movements in normal joints, whereas in inflamed joints increase of responses occurred mainly during normal movements. These data indicate that SP may also be involved in the process of sensitisation of primary afferents during an inflammation.

Inhibitory effect of somatostatin on the mechanosensitivity of articular afferents in normal and inflamed knee joints of the rat

&NA; The effect of somatostatin on the sensory activity of primary afferents was studied in normal and acutely inflamed rat knee joints. Fine afferent nerve fibers with conduction velocities of 0.9–18.0 m/s were recorded as single units. All nerve fibers tested responded to local mechanical stimulation, movements of the joint and i.a. injections of KCl (10−4 mol, 0.1 ml) close to the joint. Somatostatin (10−4 mol, 0.2 ml) caused no direct response of the units. In normal joints, somatostatin did not change the discharges evoked by non‐noxious movements but decreased the responses to noxious movements significantly to about 63% of the responses before the application. In acutely inflamed joints, somatostatin reduced the discharges of non‐noxious and of noxious movements to about 55% and 52%, respectively. Injections of somatostatin with lower concentrations (10−6 mol, 10−8 mol) i.a. close to inflamed joints revealed shorter and less pronounced reductions of the responses to noxious movements. In a proportion of afferents, substance P (10−4 mol) and bradykinin (10−4 mol) were able to increase these responses again. These data indicate that the mechanosensitivity of articular afferents in normal joints may also be regulated by several neuropeptides based on a balance of pro‐inflammatory peptides such as substance P, and anti‐inflammatory peptides such as somatostatin. In an inflamed joint, pro‐inflammatory peptides seem to predominate resulting in a sensitization of the peripheral nerve fibers. In this case, an application of somatostatin or its analogues could be used clinically to compensate this effect.

In vivo myograph measurement of muscle contraction at optimal length

BackgroundCurrent devices for measuring muscle contraction in vivo have limited accuracy in establishing and re-establishing the optimum muscle length. They are variable in the reproducibility to determine the muscle contraction at this length, and often do not maintain precise conditions during the examination. Consequently, for clinical testing only semi-quantitative methods have been used.MethodsWe present a newly developed myograph, an accurate measuring device for muscle contraction, consisting of three elements. Firstly, an element for adjusting the axle of the device and the physiological axis of muscle contraction; secondly, an element to accurately position and reposition the extremity of the muscle; and thirdly, an element for the progressive pre-stretching and isometric locking of the target muscle.Thus it is possible to examine individual in vivo muscles in every pre-stretched, specified position, to maintain constant muscle-length conditions, and to accurately re-establish the conditions of the measurement process at later sessions.ResultsIn a sequence of experiments the force of contraction of the muscle at differing stretching lengths were recorded and the forces determined. The optimum muscle length for maximal force of contraction was established. In a following sequence of experiments with smaller graduations around this optimal stretching length an increasingly accurate optimum muscle length for maximal force of contraction was determined. This optimum length was also accurately re-established at later sessions.ConclusionWe have introduced a new technical solution for valid, reproducible in vivo force measurements on every possible point of the stretching curve. Thus it should be possible to study the muscle contraction in vivo to the same level of accuracy as is achieved in tests with in vitro organ preparations.

Involvement of nitric oxide in the modulation of dural arterial blood flow in the rat

Nitric oxide (NO) has been proposed to be a key molecule in the pathogenesis of migraine pain and other headaches that are linked to vascular disorders. Several lines of evidence indicate that the meningeal vascularization is crucially involved in the generation of these headaches. In an experimental model in the rat a dominating role of calcitonin gene‐related peptide (CGRP) in causing neurogenic vasodilatation and increased blood flow has been shown. The aim of the present study was to clarify the role of NO in this model with regard to the meningeal blood flow. The blood flow in and around the medial meningeal artery (dural arterial flow) was recorded in the exposed parietal dura mater encephali of barbiturate anaesthetized rats using laser Doppler flowmetry. Local electrical stimulation of the dura mater (pulses of 0.5 ms delivered at 7.5–17.5 V and 5 or 10 Hz for 30 s) caused temporary increases in dural arterial flow for about 1 min that reached peaks of 1.6–2.6 times the basal flow. The effects of NO synthase (NOS) inhibitors on the basal flow and the electrically evoked increases in flow were examined. Systemic (i.v.) administration of Nω‐nitro‐L‐arginine methyl ester (L‐NAME) at cumulative doses of 10 and 50 mg kg−1 lowered the basal flow to 87 and 72%, respectively, of the control and reduced the evoked increases in blood flow to 82 and 44% on an average. Both these effects could partly be reversed by 300 mg kg−1 L‐arginine. The systemic arterial pressure was increased by L‐NAME at both doses. Injection of the stereoisomer D‐NAME at same doses did not change basal flow and evoked increases in flow. Topical application of L‐NAME (10−4–10−2 M) was effective only at the highest concentration, which caused lowering of the basal blood flow to 78% of the control; the evoked increases in flow were not changed. Topical application of 2‐amino‐5,6‐dihydro‐6‐methyl‐4H‐1,3‐thiazine (AMT), a specific inhibitor of the inducible NOS, at concentrations of 10−4–10−2 M lowered the basal flow to 89, 87.5 and 85%, respectively, but did not significantly change the evoked flow increases. Same concentrations of 7‐nitroindazole monosodium salt (7‐NINA), a specific inhibitor of the neuronal NOS, had no significant effects on basal flow and evoked increases in flow. It is concluded that NO is involved in the maintenance of the basal level of dural arterial blood flow as well as in the electrically evoked flow increases, which have been shown to be mainly mediated by CGRP released from dural afferent fibres. The most important source of NO is probably the endothelium of dural arterial vessels. The synergistic effect of NO and CGRP on the stimulated blood flow may be in part due to a NO mediated facilitation of the CGRP release.

Responses of fine primary afferent nerve fibres innervating the rat knee joint to defined torque.

In total 23 rat knee joint afferent units with conduction velocities of 1.2-17 m/s were recorded extracellularly during inward and outward joint rotations of defined torque using a newly developed torque-meter. There was not a linear relation between the torque and the obtained angle of the joint. The mechanical thresholds of the units ranged from 10 to 60 mNm. During 10 s of stimulation with 60 mNm they responded with 12-300 action potentials. Two mechanosensitive nerve fibres could not be activated via knee joint rotation at any torque at all. The units could be classified according to their response pattern during stimulation: (1) phasic-tonic response behaviour; (2) constant discharge rate; (3) delayed onset of activity. In conclusion, the novel torque-meter allows a precise characterisation of the mechanical threshold and the corresponding response of fine afferent joint units during knee joint rotation.

Galanin influences the mechanosensitivity of sensory endings in the rat knee joint

The aim of the present study was to examine the effect of galanin on group III and IV afferent nerve fibres (n = 53) innervating normal and acutely inflamed knee joints in rats. They responded to local mechanical stimulation, movements of the joint and i.a. injections of KCl close to the joint. Single i.a. bolus injections of galanin (0.1 mm, 0.2 mL) caused no direct responses of the units. In normal and acutely inflamed joints, about half of the units did not change the responses to knee joint rotation. A significant reduction of the responses to noxious movements was found in ∼ 40% of the units reaching a mean value of 57% in normal joints and 70% in inflamed joints compared with control movements. In ∼ 10% the responses increased to 143% in normal joints and 120% in inflamed joints. Injection of a galanin receptor antagonist (M35) doubled the responses to noxious movements in 36% of the units in normal joints and reduced it in 18% to 86% of the control movements, indicating a tonic release and influence on the mechanosensitivity of a proportion of primary afferents by galanin. In conclusion, these data further support the hypothesis that the mechanosensitivity of fine afferent nerve fibres is regulated by a mixture of different substances being released into the innervated tissue. Besides the action of several pro‐inflammatory peptides there seems to exist a tonic inhibitory system.

Effects of the 5-HT1 receptor agonists sumatriptan and CP 93,129 on dural arterial flow in the rat

The blood flow in and around the medial meningeal artery (dural arterial flow) was recorded in the exposed parietal dura mater encephali of the anesthetized rat using laser Doppler flowmetry. Local electrical stimulation of the dura mater (pulses of 0.5 ms delivered at 7.5-17.5 V and 5 or 10 Hz for 30 s) caused temporary increases in dural arterial flow. The effects of the 5-HT1 receptor agonists sumatriptan and CP 93,129 on the basal flow and the electrically evoked increases in flow were examined. Topical administration of undiluted sumatriptan (12 mg/ml) lowered the basal and the evoked flow by 20% on average. Systemic (i.v.) administration of sumatriptan (0.24, 0.72 and 3.6 mumol/kg) caused a short-lasting reduction of the evoked flow increases only at the higher doses while the basal flow was not significantly altered. Systemic administration of CP 93, 129 (0.46 and 4.6 mumol/kg) caused no significant changes of the basal and the evoked flow. At a dose of 23 mumol/kg CP 93,129 lowered the basal flow by 20% and the evoked flow by 30% for 20 min. The systemic arterial pressure was not significantly altered by sumatriptan and CP 93,129 within the whole range of doses. It is suggested that sumatriptan and CP 93,129 at high doses exert inhibitory effects on those fine afferent nerve fibers which release the calcitonin gene-related peptide, since this neuropeptide mediates the evoked increases in dural arterial flow.

Peripheral modulation of rat knee joint afferent mechanosensitivity by nociceptin/orphanin FQ.

The peripheral effects of nociceptin were examined in normal and acutely inflamed rat knee joints by analyzing single unit recordings from articular primary afferents in response to normal and extreme rotation of the knee. Bolus close intraarterial injection of nociceptin (0.01, 1 and 100 microM) caused a sensitization of normal and inflamed knee joint afferents in response to movements in the normal working range of the joint. When the joint was hyper-rotated, nociceptin had no significant effect on afferent discharge rate in normal knees, however, in inflamed joints the top dose of the neuropeptide caused a decrease in articular mechanosensitivity. These findings suggest that nociceptin seems to be involved in the control of peripheral nociceptive mechanisms, although the behaviour of the peptide is dependent upon the inflammatory status of the tissue.

Gabapentin reduces the mechanosensitivity of fine afferent nerve fibres in normal and inflamed rat knee joints

&NA; The antiepileptic drug gabapentin has been shown to have an antihyperalgesic effect following central administration. This electrophysiological investigation examined whether peripherally administered gabapentin could modulate the mechanosensitivity of primary afferents innervating normal and kaolin/carrageenan inflamed rat knee joints. Close intraarterial injection of gabapentin (0.01, 1 and 100 mg/kg) dose‐dependently reduced afferent firing rate in both normal and acutely inflamed rat knees in response to normal and hyper‐rotation of the joint. Thus, in addition to its central mode of action, peripheral administration of gabapentin reduces nociception locally and this may prove to be beneficial in the treatment of various pain syndromes including inflammatory arthritis.