Xiaoying Hua

Co-existence of substance P and calcitonin gene-related peptide-like immunoreactivities in sensory nerves in relation to cardiovascular and bronchoconstrictor effects of capsaicin.

Immunohistochemical studies showed that substance P (SP) and calcitonin gene-related peptide (CGRP) immunoreactivity co-exist in capsaicin-sensitive primary sensory neurons. Varicose SP- and CGRP-immunoreactive nerve fibres with a similar distribution pattern were seen in the lower airways and heart. The functional analysis revealed that CGRP caused cardiac stimulation and had, together with SP and neurokinin A, potent hypotensive effects. Vascular permeability was increased by SP and neurokinin A, and the bronchial smooth muscle was particularly sensitive to neurokinin A. Thus, multiple peptides stored in an possible released from the same nerve endings by capsaicin may exert differential effects in various target tissues.

Multiple tachykinins (neurokinin A, neuropeptide K and substance P) in capsaicin-sensitive sensory neurons in the guinea-pig

The occurrence of tachykinins in sensory neurons of the guinea-pig was studied by means of radioimmunoassay combined with ion-exchange and high-performance liquid chromatography as well as by immunohistochemistry. Antisera raised against kassinin (antiserum K12), neurokinin A (NKA) (antiserum NKA2) and substance P (SP) (antisera SP25 and SP2) were used. Antiserum K12 detected NKA, neuropeptide K (NPK) and a component eluting in the position of eledoisin (ELE) in extracts of the lung and ureter. Neurokinin B (NKB) was, however, not found. Neutral water extraction favored recovery of NKA and of the ELE-like component, while NPK was found only in acid extracts. The SP antisera detected two immunoreactive components of which the major form coeluted with synthetic SP. Capsaicin pretreatment depleted all these various forms of immunoreactivity in several peripheral organs including the ureter and lung. The immunoreactivity detected by antisera K12 or SP25 in radioimmunoassay had a similar regional distribution pattern in peripheral tissues. Immunohistochemical examination revealed that antiserum NKA2 stained the same spinal ganglion cells as the SP2 antiserum. The distribution of capsaicin-sensitive nerve fibers stained by these two antisera was also identical in peripheral organs such as the ureter, inferior mesenteric ganglion, heart and lung. It is concluded that multiple tachykinins, including SP, NKA, NPK and an ELE-like peptide, are present in capsaicin-sensitive sensory nerves in the guinea-pig. This finding can most likely be related to the origin of SP, NKA and NPK from the same precursor molecule, subsequent posttranslational tissue processing and axonal transport to terminal regions.

Capsaicin induced release of multiple tachykinins (substance P, neurokinin A and eledoisin-like material) from guinea-pig spinal cord and ureter.

The release of tachykinins from isolated slice preparations of the guinea-pig spinal cord and ureter was studied in vitro. Capsaicin (10 microM) caused release of substance P, neurokinin A and an eledoisin-like component from both the spinal cord and ureter. The release of tachykinins induced by capsaicin or potassium (60 mM) was calcium dependent. No detectable release of neurokinin B or neuropeptide K, an N-terminally extended form of neurokinin A, was induced by capsaicin. No detectable release of tachykinins could be demonstrated after exposure to agents which are known to activate C-fibre afferents, such as histamine, bradykinin, serotonin, prostaglandins E1, E2 or acetylcholine. Protein extravasation in the ureter, as determined by the Evans Blue extravasation technique was used as a functional correlate to the tachykinin release. Protein extravasation was induced in vivo by local intraluminal injections of capsaicin at several hundred-fold lower concentrations than those required to induce a detectable release of tachykinins in vitro. The difference may, however, partly depend on the experimental conditions and the detection limit of the tachykinin assay used. The protein extravasation response to capsaicin was absent after systemic capsaicin pretreatment, which causes a marked depletion of tachykinins in the ureter. In conclusion, capsaicin evokes release of several tachykinins from both central and peripheral endings of primary afferent neurons. The peptides released from sensory nerves in the periphery may induce effects such as protein extravasation and smooth muscle contraction.

Comparison of cardiovascular and bronchoconstrictor effects of substance P, substance K and other tachykinins.

SummaryThe effects of substance P (SP), substance K (SK), physalaemin, eledoisin, kassinin, neuromedin K and bombesin on blood pressure, heart rate, respiratory insufflation pressure and plasma extravasation were studied in the guinea-pig.2.All tachykinins except neuromedin K caused a fall in blood pressure with rather similar potency. The hypotensive response after physalaemin was comparatively more long-lasting.3.SK and eledoisin (2.5 nmol×kg−1 i.v.) caused an initial bradycardia which then changed into tachycardia. The other tachykinins induced a slowly developing tachycardia. Neuromedin K (up to 40 nmol×kg−1) did not influence heart rate.4.SK, kassinin and eledoisin were more potent than SP and physalaemin in increasing respiratory insufflation pressure. The effect of SK had a particularly long duration. Neuromedin K only induced a weak increase in insufflation pressure at a very high dose.5.All tachykinins except neuromedin K induced an increase in vascular permeability to plasma proteins in many visceral organs, as indicated by Evans blue extravasation. The trachea and ureter were the most sensitive organs with regard to this effect. Physalaemin and eledoisin were generally more potent in increasing vascular permeability in various organs than SP and SK. The maximal permeability-increasing effect of SK was smaller than that of SP, although the potency was similar.6.Bombesin increased insufflation pressure with no clearcut effects on vascular permeability.7.It is concluded that in the same species, i.e. guinea-pig, several tachykinins have rather similar hypotensive action, while the vascular permeability increase to plasma proteins is especially pronounced after physalaemin and eledoisin. SK, kassinin and eledoisin have prominent bronchoconstrictor effects. Neuromedin K, however, displays poor activity in the present models. The existence of novel tachykinins such as SK in addition to SP in mammalian tissues suggests that effects seen upon antidromic stimulation of sensory nerves may be caused by several structurally related peptides.

Leukotrienes C4, D4 and E4 cause widespread and extensive plasma extravasation in the guinea pig

Summary1.Intravenous injection of leukotriene C4 (1 nmol × kg−1) caused substantial plasma exudation in anesthetized guinea pigs, as evidenced by marked hemoconcentration 15% in 5 min) and significant extravasation of Evans blue.2.Fluorometric quantitation of Evans blue content in 38 selected tissues documented that leukotriene C4 caused significant plasma extravasation throughout the body, except for the brain, stomach, duodenum, colon and gonads.3.In particular, the respiratory and the uro-genital tracts, but also the conjunctiva, the esophagus, the bile ducts and the umbilical ligaments, were very sensitive to the edemapromoting effect of leukotriene C4.4.Intravenous injection of leukotriene D4 (1 nmol × kg −1) or E4 (5 nmol × kg−1) evoked plasma extravasation with a distribution and magnitude that was similar to that induced by leukotriene C4.5.It is concluded that the three major constituents of slow reacting substance of anaphylaxis (SRS-A), leukotrienes C4, D4 and E4, cause a generalized and extensive plasma exudation that is consistent with the proposal that these leukotrienes are important mediators of inflammation.

Co-localization of tachykinins and calcitonin gene-related peptide in capsaicin-sensitive afferents in relation to motility effects on the human ureter in vitro

Tachykinin- and calcitonin gene-related peptide (CGRP) immunoreactivities were localized by immunohistochemistry in the same nerves of the kidney, renal pelvis and ureter as well as in spinal ganglion cells of both the guinea-pig and man. The tachykinin and CGRP-immunoreactive nerves in the ureter were present within the smooth muscle layers, around blood vessels, close to and within the lining epithelium. The levels of neurokinin A-, substance P- and CGRP-like immunoreactivity per tissue weight, as determined by radioimmunoassay, were about 30-100-fold higher in the guinea-pig than in the human ureter, which was in good agreement with the relative density of immunoreactive nerve fibres, as seen by immunohistochemistry. Capsaicin treatment caused an almost total disappearance of both neurokinin A-, substance P- and CGRP-immunoreactive nerve fibres in the guinea-pig ureter and a 90% depletion of neurokinin A, substance P- and CGRP-like immunoreactivity, further supporting a sensory origin of these nerves. Reversed-phase high performance liquid chromatography of water extracts of the human ureter revealed the presence of neurokinin A- and eledoisin-like material using antiserum K12, which does not cross-react with substance P. Most of the CGRP-like immunoreactivity in human ureter extracts co-eluted with synthetic human CGRP. Capsaicin both caused inhibition of spontaneous motility of the human ureter in vitro and initiated contractions in some preparations. Neurokinin A and neuropeptide K potently initiated phasic contractions of the ureter, while substance P had only minor contractile effects. CGRP inhibited both spontaneous and neurokinin A-induced ureteric contractions. In conclusion, peptides with potent opposite motility effects are present in the same, presumably sensory nerves of the ureter in both the guinea-pig and man. It will be of importance to determine whether local release of neuropeptides can account for ureteric motility changes accompanying sensory nerve activation upon ureteral obstruction, by e.g. renal calculi.

Capsaicin-induced substance P release and sensory control of vascular permeability in the guinea-pig ureter

Substance P-immunoreactivity (SP-IR) in the guinea-pig ureter was found to be totally depleted after systemic capsaicin pretreatment. Removal of the inferior mesenteric ganglion (IMG) led to a total depletion of SP-IR from the rostral third of the ureter and to a partial depletion from the caudal third. Electrical stimulation of the IMG caused Evans blue extravasation mainly in the rostral third of both ureters, whereas stimulation of the right pelvic nerve caused Evans blue extravasation in the caudal third of the ureters on both sides. The responses to nerve stimulation were absent in capsaicin-pretreated animals. Furthermore, capsaicin caused release of SP-IR from ureter slices in vitro, this release was not inhibited by tetrodotoxin. Potassium (60 and 120 mM) also released SP-IR. It is concluded that SP-IR in the ureter is contained in capsaicin-sensitive sensory neurons reaching the ureter via both parasympathetic (caudal part) and sympathetic nerves (rostral part). Activation of these neurons by capsaicin leads to a peripheral release of SP-IR which most likely increases vascular permeability.

Inhibition of compound 48/80 - induced vascular protein leakage by pretreatment with capsaicin and a substance P antagonist

Summary1.Intravenous injection of compound 48/80 (1 mg×kg−1) induced an acute increase in vascular permeability to plasma proteins in various organs of rats.2.The compound 48/80 response was partly inhibited by histamine H1 and H2 receptor blockade in the urinary bladder and in the duodenum, but not in the trachea, the oesophagus, the ureter and the paw skin. Blockade of 5-hydroxytryptamine receptors with methysergide led to a reduction of the permeability response in the oesophagus and in the urinary bladder, leaving responses in other organs unchanged.3.Pretreatment of neonatal rats with capsaicin almost abolished the 48/80 response in all organs except in the duodenum. Pretreatment of rats with [D-Arg1, D-Trp7,9, Leu11]-substance P, a substance P antagonist, also caused a partial inhibition of the permeability response to compound 48/80 in several organs.4.Topical administration of compound 48/80 (1 mg×ml−1) onto the tracheal mucosa induced local Evans blue extravasation. This response was resistant to pretreatment with histamine receptor antagonists, but was largely inhibited after neonatal capsaicin pretreatment. Topical administration of compound 48/80 (1 mg×ml−1 or 10 mg×ml−1) into the eye did not cause visible Evans blue extravasation in the conjunctiva, nor any signs of pain reaction as indicated by the absence of the wiping response, usually seen upon noxious chemical stimuli in the eye.5.In guinea-pigs, 10 mg×kg−1 compound 48/80 i.v. were required to induce vascular protein leakage in different organs. This response was blocked by pretreatment with H1 and H2 receptor antagonists, but only slightly reduced after systemic capsaicin pretreatment of guinea-pigs.6.Pretreatment of rats with compound 48/80 for 3 consecutive days lead to an almost complete depletion of histamine in the paw skin and to a partial depletion of histamine in the trachea. In the lung and in the duodenum, however, histamine concentrations remained unchanged. Simultaneously, there was an about 25%–30% reduction of substance P concentrations in the paw skin and in the trachea, but not in the lung or in the spinal cord. In compound 48/80 pretreated rats, there was no change of the permeability response to intravenous substance P, nor any change of the behavioural nociceptive response upon topical capsaicin application in the eye.7.It is concluded that intravenous compound 48/80 administration induces an acute increase in vascular permeability in various organs of rats mainly via a histamine independent activation of capsaicin sensitive, most probably substance P containing sensory neurones. In guineapigs, the permeability response to intravenous compound 48/80 is mainly caused by histamine, acting both directly and indirectly via sensory neurones. Pretreatment of rats with compound 48/80 causes, besides histamine depletion from mast cells, a reduction of substance P in sensory nerves. This may reflect a long lasting stimulation of sensory nerve endings by mast cell constituents or a direct action of compound 48/80 on sensory nerves.

Action and metabolism of circulating leukotriene C

The microvascular permeability effects of intravenously administered leukotriene C4 were assessed by fluorometric determinations of extravasated Evans blue dye in various tissues of guinea pigs. The effects were widespread, with intensity differences between different vascular beds, which in part reflected the tissue distribution of tritium-labeled leukotriene C3 in mice. Local tissue specificity favored permeability effects in the urinary tract, pulmonary airways, esophagus, and distal colon. The permeability effects may contribute to the hypotensive actions of leukotriene C4 and enhance airway obstruction by edema formation in addition to smooth muscle contraction.