Josef Donnerer

Increased content and transport of substance P and calcitonin gene-related peptide in sensory nerves innervating inflamed tissue: evidence for a regulatory function of nerve growth factor in vivo.

The responses of sensory neuropeptides during unilateral, Freunds adjuvant-induced, paw inflammation in the rat were examined. After five days of inflammation, the substance P and calcitonin gene-related peptide content in the sciatic nerve supplying the inflamed paw were increased by 60-75% when compared with the contralateral side. At this time-point, there was also a 30-40% increase in the substance P and calcitonin gene-related peptide content of the dorsal root ganglia (L4-L6), and a 40% increase in the calcitonin gene-related peptide content of the L4-L6 segments of the dorsal spinal cord on the inflammation side. In the dorsal root ganglia, calcitonin gene-related peptide content was also increased as early as 12 h and 48 h after induction of paw inflammation. On day 5 of inflammation, the axonal transport of both sensory neuropeptides towards the inflamed paw, as determined after sciatic nerve ligation, was also markedly increased as compared with the control side. Despite this increased transport, the amount of substance P and calcitonin gene-related peptide present in the inflamed paw itself was either reduced or remained unchanged from day 1 through to day 5 of inflammation pointing towards reduced storage and increased release of the peptides in the inflamed tissue. Nerve growth factor content was markedly increased in the sciatic nerve of the inflamed paw with a peak of +136% at time-point 24 h after induction of inflammation. When rats were systemically treated with anti-nerve growth factor serum, the increase in neuropeptide content in the sciatic nerve of the inflamed paw (day 5) was prevented. On the other hand, local injections of nerve growth factor for 5 days into a noninflamed paw were able to induce an increase in substance P and calcitonin gene-related peptide content in the supplying sciatic nerve. These findings point towards a regulatory function for nerve growth factor in vivo in the stimulation of sensory neuropeptide synthesis during prolonged inflammatory processes.

Inhibition of neurogenic vasodilatation and plasma extravasation by substance P antagonists, somatostatin and [D-Met2, Pro5]enkephalinamide

The substance P (SP) analogues [D-Pro2, D-Phe7, D-Trp9]SP and [D-Pro2, D-Trp7,9]SP, which have been reported to be SP antagonists, inhibited the vasodilation and plasma extravasation induced by antidromic stimulation of the saphenous nerve or by i.a. infusion of SP. Somatostatin inhibited the vasodilatation and plasma extravasation induced by saphenous nerve stimulation, but had no effect on the vascular responses to i.a. infused SP. The opiate agonist [D-Met2, Pro5]enkephalinamide inhibited the vasodilation evoked by antidromic nerve stimulation in a naloxone reversible manner, but did not change the effect of i.a. infusion of SP. Calcitonin and caerulein had no effect on neurogenic vasodilatation. These results further support the concepts that neurogenic vasodilatation and plasma extravasation are mediated by SP, and that somatostatin and opiates inhibit the release of SP from peripheral sensory nerve endings.

Effects of capsaicin on inflammation and on the substance P content of nervous tissues in rats with adjuvant arthritis

Capsaicin (20-80 mg/kg, s.c.) reduced the inflammatory response to inoculation with Mycobacterium butyricum in the rat. The effect was apparent within 24 h, was partial, persisted for well over 20 days, and occurred irrespective of whether capsaicin was administered before or after the onset of inflammation, or at the time when the pathology reached peak. Capsaicin also attenuated the increase in substance P content in sciatic nerve, saphenous nerve, dorsal root ganglia, dorsal roots, and dorsal spinal cord (L4, L5) which occurs in rats with adjuvant arthritis. The data are consistent with a possible role of substance P in the peripheral manifestations of adjuvant arthritis.

Increase of substance P in primary afferent nerves during chronic pain

Abstract Substance P was found to be increased in the sciatic nerve of rats suffering chronic pain due to adjuvant-induced polyarthritis. This increase of substance P in primary afferents is assumed to reflect adaptive changes initiated by chronic noxious events such as inflammation.

The non-peptide tachykinin antagonist, CP-96,345, is a potent inhibitor of neurogenic inflammation

1 Release of the tachykinin, substance P, from the peripheral terminals of polymodal afferent C‐fibres is thought to be largely responsible for the vasodilatation and plasma protein extravasation described as neurogenic inflammation. The effects of CP‐96,345, a non‐peptide antagonist at the substance P (NK1) receptor, on these vascular reactions were investigated in the rat. 2 Intravenously (i.v.) injected CP‐96,345 (0.4–3.0 μmol kg−1) prevented the drop in blood pressure, a measure of the peripheral vasodilatation, evoked by substance P and neurokinin A in a dose‐ and time‐dependent manner, but did not affect that elicited by the non‐tachykinin peptides calcitonin gene‐related peptide and vasoactive intestinal polypeptide. 3 Plasma protein extravasation evoked by i.a. infusion of substance P, antidromic stimulation of the saphenous or the vagus nerve, and stimulation of cutaneous afferent nerves with mustard oil, were each significantly inhibited by CP‐96,345 (3.0–9.0 μmol kg−1, i.v.). Furthermore, CP‐96,345 was orally active in blocking mustard oil‐induced plasma extravasation with an ED50 of 10 μmol kg−1. 4 The inhibition of substance P‐induced vasodilatation and of neurogenic plasma extravasation by CP‐96,345 was stereospecific as the inactive isomer CP‐96,344 (2R, 3R enantiomer of CP‐96,345) had no effect. 5 Thus CP‐96,345 is a specific, highly potent, long‐acting and orally active inhibitor of tachykinin‐mediated neurogenic inflammation.

Release of histamine by substance P.

Summary1.The basic peptide substance P causes histamine release from peritoneal mast cells of the rat in vitro whereas the closely related neutral peptides eledoisin and physalaemin do not.2.Infusion of substance P (7.4 nmol min−1), but not of eledoisin (8.4 nmol min−1) or physalaemin (7.9 nmol min−1), into the rat hindquater preparation caused a more than 4-fold increase of the histamine content in the venous outflow. The outflow of 5-HT remained unchanged under infusion of all three peptides.3.No histamine depletion in the skin of the rat hind paw was observed following antidromic stimulation of the saphenous nerve or cutaneous application of mustard oil. Infusion of substance P (7.4 nmol min−1) caused a 47% depletion of histamine in the paw skin although only a small proportion of the infused substance P seemed to enter the tissue from the blood vessels.4.The results further substantiate the view that substance P upon release from peripheral nerve endings induces release of histamine from cutaneous mast cells, a mechanism which contributes to neurogenic vasodilatation and plasma extravasation.

Opioid control of the function of primary afferent substance P fibres.

Lofentanil, a very potent and long-acting opiate agonist, was used to evaluate the opioid control of substance P release from primary afferents. Substance P release from the central terminals of primary afferents was studied in the superfused isolated dorsal half of the rat spinal cord. Substance P release as initiated by electrical field stimulation and by capsaicin was found to be diminished by 50% by lofentanil (1 microM) in a naloxone-reversible manner. Substance P release from peripheral terminals of primary afferents was induced by antidromic saphenous nerve stimulation. Release was measured indirectly by its effect on blood flow and plasma extravasation in the rat hind paw. Both antidromic vasodilatation and plasma extravasation were dose dependently inhibited by lofentanil. The inhibition of antidromic vasodilatation by 10 micrograms X kg-1 lofentanil i.p. was completely prevented by 1 mg X kg-1 naloxone. On the other hand, vasodilatation and plasma extravasation induced by infusion of 3.7 pmol X min-1 substance P remained unaffected by lofentanil. It is concluded that lofentanil inhibits the release of substance P from central as well as peripheral terminals of substance P-containing primary afferent neurons.

Intraplantar injection of nerve growth factor into the rat hind paw: local edema and effects on thermal nociceptive threshold

&NA; Nerve growth factor (NGF) is known to produce hyperalgesia as well as to stimulate synthesis of neuropeptides in dorsal root ganglia (DRG). In the present study, we wanted to determine the effects of local NGF administration and assess to which extent mast cell‐dependent factors are mediating NGF responses. Rats received 1 daily unilateral intraplantar injection for 3 days. Local edema (days 1–3), changes in thermal nociceptive threshold (days 1–4), and the content of calcitonin gene‐related peptide (CGRP) and substance P (SP) in the sciatic nerve (day 4), were determined. NGF injection caused edema which was absent in rats pretreated with compound Symbol as well as in rats treated neonatally with capsaicin (‘capsaicin denervation’). NGF‐induced edema was not reduced by the neurokinin‐1 receptor antagonist SR140333, but attenuated by the CGRP receptor antagonist CGRP[8–37]. Symbol. No caption available On each day, NGF injection caused a decrease in thermal nociceptive threshold which lasted for less than 3 h. Capsaicin denervation, but not treatment with indomethacin, abolished NGF‐induced thermal hyperalgesia. Treatment with compound Symbol attenuated hyperalgesia produced by the first, but not by subsequent, NGF injections. Symbol. No caption available On day 4, 24 h after the last of 3 NGF injections, thermal nociceptive threshold was not different from control values, but at that time, CGRP and SP were elevated in the sciatic nerve. We suggest therefore that NGF‐induced local edema was caused by mast cell‐derived vasoactive compounds which act together with afferent neuron‐derived CGRP to increase vascular permeability. NGF‐induced thermal hyperalgesia most likely was caused by an increased sensitivity of peripheral endings of capsaicin sensitive afferents. This effect of NGF was not mediated by products of the cyclooxygenase pathway, and was also observed in mast cell‐depleted rats.

Time course of capsaicin-induced functional impairments in comparison with changes in neuronal substance P content.

Summary1.Changes in the content of substance P (dorsal spinal cord, dorsal roots, dorsal root ganglia, saphenous nerve, skin) and functional changes (neurogenic plasma extravasation, chemosensitivity of the cornea) were measured in the rat from 10 min to 4 days after the s.c. injection of a single dose of 50 mg kg−1 capsaicin.2.The substance P content in dorsal roots, saphenous nerve and hind paw skin progressively declined to about 60–70% of control 4 days after treatment, whereas that of the dorsal root ganglia rose, after an initial decline, to 140% after 1–4 days.3.After denervation, impairment of neurogenic plasma extravasation could be observed not earlier than after one day, thus being comparable in time course to the depletion of substance P in the skin and saphenous nerve.4.Neurogenic plasma extravasation and the chemosensitivity of the cornea were greatly diminished already 10 min after systemic capsaicin treatment, i.e. at a time when the substance P content of the peripheral nerve was still unchanged. These early effects of systemic capsaicin treatment are therefore caused by actions other than depletion of substance P.

Analgesic effect of antagonists of substance P

[D-Pro2,D-Phe7,D-Trp9]-SP and [D-Pro2,D-Trp7,9]-SP havebeen shown to be antagonists of substance P. The hindlimb scratching syndrome of mice, known to be caused by substance P was absent when these peptides were injected into substance P-treated mice. Substance P shortens “tail withdrawal time” from hot water; the two peptides greatly prolonged tail withdrawal time. Antidromic stimulation of the saphenous nerve (rat), known to release substance P and to induce vasodilatation plasma extravasation, was also greatly inhibited by [D-Pro2,D-Phe7,D-Trp9]-SP. These peptides presumably cause anti-nociceptor effects (analgesia) by inhibition of substance P at receptors and favor the concept that substance P is a sensory neurotransmitter of nociceptive messages.