Calcitonin gene-related peptide in the regulation of urinary tract motility.
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Animals↗
Biomedical subjects
P Santicioli
Publications and source records attributed to P Santicioli.
Cyclophosphamide cystitis in rats: involvement of capsaicin-sensitive primary afferents.
The involvement of capsaicin-sensitive primary afferent neurons in cyclophosphamide (CYP)-induced cystitis has been investigated in rats. CYP (150 mg/kg) was administered 48 h before testing in both vehicle- and capsaicin- (50 mg/kg s.c., 4 days before) treated rats. Some experiments were also performed 96 h after bilateral removal of pelvic ganglia to produce bladder denervation. CYP administration produced a marked detrusor hyperreflexia which was abolished by capsaicin pretreatment, demonstrating that it is mediated through stimulation of capsaicin-sensitive afferent neurons. CYP administration was followed by a marked increase in bladder weight and plasma protein extravasation (measured by the Evans blue leakage technique). The latter effect was largely prevented by ganglionectomy but was aggravated by capsaicin pretreatment. The effect of capsaicin was suppressed by ganglionectomy. Isolated strips of detrusor muscle from CYP-treated animals developed less tension in response to various stimuli as compared to strips from vehicle-treated animals; however, when contractile responses were expressed as percentage of an internal standard (carbachol-induced contraction) no difference was evident between the two groups. The bladder content of calcitonin gene-related peptide, used as a marker of the bladder afferent fibres that are capsaicin-sensitive in adult rats, was slightly reduced as compared to controls, but the difference can be accounted for by the increased bladder weight. We conclude that CYP-induced cystitis is not accompanied by a toxic effect on bladder nerves and that the decrease in bladder capacity is entirely mediated through stimulation of capsaicin-sensitive afferent fibres, presumably linked to the formation of the irritant metabolite of CYP, acrolein.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Activity of peptide and non-peptide antagonists at peripheral NK1 receptors.
We investigated the affinity of several tachykinin antagonists reportedly selective for NK1 receptors at various tachykinin receptors and NK2 receptors subtypes. The four antagonists tested were: L 668,169, Spantide II, Ac-Thr-DTrp(for)-Phe-NMeBzl (FR 113680) and the novel nonpeptide antagonist (+/-)-CP-96,345. The four antagonists were found to be effective against NK1 receptor-mediated responses in the guinea-pig ileum with the following rank order of potency (pKB values in parentheses): (+/-)-CP-96,345 (8.11) greater than Spantide II (7.08) greater than FR 113680 (6.61) greater than or equal to L 558,169 (6.44). (+/-)-CP-96,345, Spantide II and FR 113680 were distinctly more potent at NK1 receptors than at NK2 receptors (NK2A in the rabbit pulmonary artery, NK2B in the hamster trachea). L 668,169 antagonized neurokinin A-induced contractions in the hamster trachea with an affinity similar (pKB value 6.16) to that found in the guinea-pig ileum for NK1 receptors (pKB value 6.44). All antagonists were inactive at NK3 receptors of the rat portal vein. In a second series of experiments, the affinities of test antagonists for NK1 receptors in the guinea-pig ileum were compared to those for NK1 receptors in the guinea-pig vas deferens, the rabbit jugular vein and the rat urinary bladder. For each antagonist, the affinity measured in the guinea-pig vas deferens and the rabbit jugular vein was comparable to that found in the guinea-pig ileum. In the rat urinary bladder, (+/-)-CP-96,345 was about 100 times less potent in blocking NK1 receptor-mediated contractions than in the guinea-pig ileum.(ABSTRACT TRUNCATED AT 250 WORDS)
Amino Acid Sequence↗
Release of calcitonin gene-related peptide like-immunoreactivity induced by electrical field stimulation from rat spinal afferents is mediated by conotoxin-sensitive calcium channels.
Electrical field stimulation (EFS, 10 V, 50 mA/cm2, 5-50 Hz, 1 ms pulse duration, 10 s train every 20 s for 5 min) produced a rapid and reproducible outflow of calcitonin gene-related peptide like-immunoreactivity (CGRP-LI) from superfused slices from the dorsal half of the rat spinal cord which is abolished by tetrodotoxin (TTX, 0.3 microM), in vitro capsaicin desensitization (10 microM for 30 min) and in Ca-free medium. The response was unaffected by ruthenium red (10 microM), indomethacin (10 microM) and nifedipine (1 microM) while it was abolished by omega-conotoxin (omega-CTX, 0.1 microM) and, in a naloxone-sensitive manner, by morphine (3 microM). Since CGRP release from capsaicin-sensitive afferents evoked by EFS in rat peripheral tissues is conotoxin-resistant, these findings provide direct evidence for a qualitative difference between central and peripheral endings of capsaicin-sensitive primary afferents in the mechanisms regulating transmitter release in the same species.
Animals↗
Tachykinin antagonists inhibit nerve-mediated contractions in the circular muscle of the human ileum. Involvement of neurokinin-2 receptors.
The effects of some newly developed tachykinin antagonists that are selective for the neurokinin (NK)-1 (L 668,169) or the NK-2 (MEN 10,207, L 659,877 and R 396) tachykinin receptor on the cholinergic and noncholinergic contraction and on the nonadrenergic noncholinergic relaxation produced by electrical field stimulation (50 Hz) were investigated in mucosa-free circular strips of the human ileum. The strips were contracted by substance P and neurokinin A as well as by selective NK-2-receptor ligands, [beta Ala8]neurokinin A(4-10), and MDL 28,564, the latter peptide being capable of discriminating between NK-2-receptor subtypes. The selectivity of the antagonists for NK-1 or NK-2 receptors was confirmed in pharmacological experiments using substance P, neurokinin A, and [beta Ala8]neurokinin A(4-10) as stimulants. Among the NK-2-selective antagonists, MEN 10,207 displayed the highest affinity, followed by L 659,877 and R 396. The antagonists MEN 10,207 and L 659,877 inhibited the noncholinergic contraction to electrical stimulation in a concentration-dependent manner; L 668,169 and R 396 were poorly effective. Thus the potency of antagonists toward the noncholinergic response closely paralleled their rank order of potency at NK-2 receptors. The cholinergic contraction and nonadrenergic noncholinergic relaxation were not inhibited by the antagonists. Both substance P- and neurokinin A-like immunoreactivities were detected in extracts of the human ileum, and the identity of the corresponding peptides was confirmed by reverse-phase high-performance liquid chromatography. It was concluded that in addition to NK-1 receptors, the circular muscle of the human ileum also contains NK-2 receptors. Activation of the latter is chiefly responsible for the noncholinergic contraction to nerve stimulation.
Aged↗
Capsaicin-like activity of N-ethylmaleimide in rat stomach.
1. N-Ethylmaleimide (NEM) and capsaicin induced a release of calcitonin gene-related peptide-like immunoreactivity (CGRP-li) in superfusates from rat gastric corpus. 2. The prior application of capsaicin completely blocked CGRP-li release by NEM while the exposure to NEM reduced the capsaicin-evoked CGRP-li outflow by about 51%. 3. These findings provide evidence that NEM might exert capsaicin-like activity in rat stomach and the release of CGRP-li from capsaicin-sensitive afferent fibres might influence the properties exerted by NEM in this tissue.
Animals↗
Tachykinins and calcitonin gene-related peptide as co-transmitters in local motor responses produced by sensory nerve activation in the guinea-pig isolated renal pelvis.
Electrical field stimulation of circular muscle strips from the guinea-pig isolated renal pelvis produces a frequency-dependent positive inotropic effect of the spontaneous contractions which is unaffected by atropine and guanethidine and abolished by tetrodotoxin or in vitro capsaicin desensitization. Omega conotoxin fraction GVIA markedly inhibited the response to low frequencies of stimulation but had only a partial or minor inhibitory effect at higher frequencies. Tachykinins produce a concentration-dependent positive inotropic effect, neurokinin A being more potent than substance P. On the other hand, rat alpha calcitonin gene-related peptide (CGRP) inhibited spontaneous contractions of the renal pelvis. MEN 10,376 a neurokinin A (4-10) analog, antagonized the positive inotropism produced by neurokinin A, without affecting the response to KCl, and suppressed the positive inotropic response produced by electrical field stimulation. In the presence of MEN 10,376, a negative inotropic response was produced by electrical field stimulation which was antagonized by the C-terminal fragment (8-37) of human alpha calcitonin gene-related peptide (hCGRP). hCGRP (8-37) antagonized the negative inotropic effect of exogenously administered CGRP without affecting inhibition by isoprenaline. Application of capsaicin (10 microM) produced a marked increase in the outflow of substance P-, neurokinin A- and CGRP-like immunoreactivities from the superfused guinea-pig renal pelvis. Substance P-, neurokinin A- and CGRP-like immunoreactivities were also detected in tissue extracts of the renal pelvis by radioimmunoassay. These experiments indicate that peptide release from peripheral endings of capsaicin-sensitive primary afferents represents the major type of nerve-mediated response affecting motility of the guinea-pig isolated renal pelvis. Tachykinins and CGRP act as physiological antagonists and the excitatory action of tachykinins prevails over the inhibitory action of CGRP. Local modulation of renal pelvis motility by sensory nerves could facilitate removal of irritants present in the urine, protecting the kidney during obstruction and ureteral antiperistalsis.
Animals↗
Effect of the GABAB antagonist, phaclofen, on baclofen-induced inhibition of micturition reflex in urethane-anesthetized rats.
The effect of intrathecal or intracerebroventricular administration of the GABAB receptor agonist, baclofen, on rhythmic contractions induced by distension of the urinary bladder (micturition reflex) was evaluated in urethane-anesthetized rats. Baclofen inhibited bladder motility acting at central nervous system sites (spinal and supraspinal) with a comparable potency. The inhibitory effect of i.t. baclofen (0.1-10 nmol) was blocked by i.t. phaclofen (200 nmol) while i.c.v. phaclofen did not affect i.c.v. baclofen (0.1-1 nmol). The inhibition of the micturition reflex induced by bladder distension observed after i.t. administration of baclofen was unaffected by systemic capsaicin pretreatment (50 mg/kg s.c., four days before). On the other hand, i.t. baclofen suppressed, in a phaclofen-sensitive manner, the reflex bladder contraction evoked by chemical stimulation (topical capsaicin) of capsaicin-sensitive bladder afferents. Intrathecal baclofen did not affect the hexamethonium-resistant tonic contraction produced by topical application of capsaicin on to the urinary bladder, which is ascribable to local peptide release from sensory nerves. Bladder motility inhibition by i.t. or i.c.v. baclofen (1 nmol) was unchanged by previous administration of p-chlorophenylalanine, indicating that the serotonergic pathways do not play a role in its action. Baclofen (100 microM) suppressed the release of calcitonin gene-related peptide-like immunoreactivity evoked by electrical field stimulation from the dorsal half of the rat spinal cord. This response was also abolished by in vitro capsaicin desensitization or tetrodotoxin, indicating that baclofen suppresses transmitter release from central endings of capsaicin-sensitive primary afferents. The present findings indicate that baclofen acts at both spinal and supraspinal sites to inhibit, with different mechanisms, the micturition reflex.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Local motor responses to bradykinin and bacterial chemotactic peptide formyl-methionyl-leucyl-phenylalanine (FMLP) in the guinea-pig isolated renal pelvis and ureter.
The local motor response to bradykinin and the bacterial chemotactic peptide, formyl-methionyl-leucyl-phenylalanine (FMLP) was investigated in the guinea-pig isolated renal pelvis and ureter in relation to possible activation of capsaicin-sensitive primary afferent nerves and release of sensory neuropeptides. Both bradykinin (1 nM-10 microM) and FMLP (10 nM-10 microM) produced a concentration-dependent positive inotropic effect in the isolated renal pelvis which was unaffected by in vitro capsaicin desensitization. The response to bradykinin was antagonized by HOE 140, a bradykinin receptor antagonist, while it was unaffected by MEN 10,376, a tachykinin receptor antagonist, hCGRP(8-37) a calcitonin gene-related peptide (CGRP) receptor antagonist and N-t-BOC-Phe-DLeu-Phe-DLeu-Phe (BPLPLP), an FMLP antagonist. The response to FMLP was blocked by BPLPLP while it was unaffected by HOE 140, MEN 10,376 or hCGRP(8-37). Indomethacin (10 microM) enhanced the response to both bradykinin and FMLP. Bradykinin transiently activated rhythmic contractions in the isolated ureter. The response to bradykinin was blocked by HOE 140 and was unaffected by in vitro capsaicin desensitization, indomethacin, MEN 10,376 or BPLPLP. FMLP had no motor effect on the resting ureter but when rhythmic background contractions were evoked by the addition of 100 nM endothelin 1, it produced a transient suppression of ureteral motility. This inhibitory effect was unchanged by in vitro capsaicin desensitization or HOE 140 while it was abolished by indomethacin or BPLPLP pretreatment. Both bradykinin and FMLP evoked the release of CGRP-like immunoreactivity in the renal pelvis. The effect of bradykinin but not that of FMLP was abolished by indomethacin. By contrast neither bradykinin nor FMLP did evoke a significant CGRP-LI release in the ureter. It is concluded that bradykinin and FMLP affect pyeloureteral motility through specific and independent pathways. The local motor responses produced by these chemical stimulants are independent from the release of sensory neuropeptides from capsaicin-sensitive primary afferent neurons. Direct neurochemical evidence was obtained for activation of capsaicin-sensitive primary afferents in the renal pelvis: such a mechanism could be involved in the genesis of ureteral pain whenever bradykinin or FMLP come into contact with sensory nerves in the pyeloureteral wall.
Animals↗
Tachykininergic transmission to the circular muscle of the guinea-pig ileum: evidence for the involvement of NK2 receptors.
1. The effect of newly developed, receptor-selective tachykinin antagonists (GR 71,251 for NK1 receptors, MEN 10,376 and L 659,877 for NK2 receptors) on noncholinergic transmission to the circular muscle of the guinea-pig ileum has been investigated. 2. In circular muscle strips of the ileum, electrical field stimulation in the presence of atropine (2 microM) and apamin (0.1 microM) evoked a complex motor response. The tonic primary contraction in this response was reduced by GR 71,251 (10 microM) and MEN 10,376 (3-10 microM) but not by L 659,877 (up to 10 microM). The presence of apamin was necessary in this experimental arrangement to unmask an atropine-resistant primary contraction, sensitive to tachykinin antagonists. The motor response was abolished by tetrodotoxin. 3. In circular strips of the ileum GR 71,251 (10 microM) inhibited the tonic contraction produced by [Sar9] substance P sulphone, a selective NK1 receptor agonist but not that produced by [beta Ala8] neurokinin A (4-10), a selective NK2 receptor agonist. By contrast, MEN 10,376 antagonized the effect of the NK2 agonist while leaving the response to the NK1 agonist unaffected. 4. In whole segments of the ileum, distension of the gut wall by an intraluminal balloon placed at about 1 cm from the point of recording of mechanical activity of the circular muscle produced atropine-sensitive phasic contractions (ascending enteric reflex). In the presence of atropine (2 microM), a noncholinergic response was elicited, which required larger volumes of distension that the cholinergic one. The atropine-resistant ascending enteric reflex was enhanced by apamin (0.1 microM) and abolished by tetrodotoxin, either in the presence or absence of apamin.5. MEN 10,376 (3-lOmicroM) inhibited the atropine-resistant ascending enteric reflex in the presence of apamin while GR 71,251 or L 659,877 (10 microM each) were ineffective. MEN 10,376 inhibited the atropine-resistant ascending enteric reflex to a larger extent in the absence than in the presence of apamin and also slightly inhibited the ascending enteric reflex in the absence of atropine.6. These findings provide evidence for an involvement of NK2 tachykinin receptors in excitatory transmission to the circular muscle of the guinea-pig ileum. NK2 receptors are also involved in the physiological-like circular muscle activation produced by stimulation of intramural neuronal pathways which subserve the atropine-resistant ascending enteric reflex.
Animals↗
Involvement of 5-hydroxytryptamine1A receptors in the modulation of micturition reflexes in the anesthetized rat.
Intravenous administration of the selective 5-hydroxytryptamine (5-HT)1A receptor agonist 8-hydroxy-2-(di-N-propylaminotetralin (8-OH-DPAT) and of a low doses of buspirone elicited the supraspinal micturition reflex (SMR) in urethane-anesthetized rats when the urinary bladder was filled with just a subthreshold volume of saline (threshold conditions). The effect of i.v. 8-OH-DPAT was abolished by hexamethonium or spiroxatrine. When SMR was elicited by bladder distension (suprathreshold conditions), i.v. 8-OH-DPAT increased the frequency of bladder contractions. In threshold conditions, stimulation of SMR was also induced by i.c.v. or by i.t. administration of 8-OH-DPAT and 5-HT but not by topical application of 8-OH-DPAT onto the bladder. Guanethidine pretreatment, which produced detrusor hyperreflexia, antagonized the effect of both i.c.v. and i.t. 8-OH-DPAT. In rats treated with capsaicin as adults, the response to 8-OH-DPAT was unchanged. In rats treated with capsaicin as newborns, instead, the response to i.t. 8-OH-DPAT was abolished and that to i.c.v. 8-OH-DPAT was shifted to higher doses. Pretreatment with 5,7-dihyroxytryptamine did not affect the response to i.t. 8-OH-DPAT but shifted to higher doses the response to i.c.v. 8-OH-DPAT. Intravenous administration of spiroxatrine, methysergide, NAN-190 [1-(2-methoxyphenyl)-4-[4-(2-phtalimido)butyl] piperazine] or high doses of buspirone but not of 1-sulpiride inhibited SMR in suprathreshold conditions. The inhibitory effect of spiroxatrine, NAN-190 and buspirone was not reduced by guanethidine pretreatment. In chronically spinalized animals, i.v. 8-OH-DPAT increased the amplitude of the reflex bladder contractions induced by bladder distension.(ABSTRACT TRUNCATED AT 250 WORDS)
8-Hydroxy-2-(di-n-propylamino)tetralin↗
Different pathways by which extracellular Ca2+ promotes calcitonin gene-related peptide release from central terminals of capsaicin-sensitive afferents of guinea pigs: effect of capsaicin, high K+ and low pH media.
Different modes by which Ca2+, entering the nerve terminal, promotes transmitter secretion as well as the ability of protons to release neuropeptides, have been shown in peripheral endings of capsaicin-sensitive afferents. We have studied these two aspects in the central endings of these neurons by measuring the release of calcitonin-gene related peptide-like immunoreactivity (CGRP-LI) from slices of the dorsal half of the guinea pig spinal cord. Although capsaicin (1 microM) released both CGRP-LI and substance P-like immunoreactivity (SP-LI), CGRP-LI was chosen as the sole suitable marker of peptides released from central terminals of capsaicin-sensitive afferents, since after in vitro desensitization to capsaicin (1 microM capsaicin for 20 min), high K+ (80 mM) failed to evoke CGRP-LI release, whereas SP-LI release was still observed. The capsaicin (1 microM)-evoked CGRP-LI release was entirely dependent on extracellular Ca2+. It was unaffected by 0.3 microM tetrodotoxin (TTX), slightly reduced by 0.1 microM omega-conotoxin (CTX) and blocked by 10 microM Ruthenium red (RR). The Ca(2+)-dependent K+ (80 mM)-evoked CGRP-LI release was unaffected by TTX, markedly reduced by CTX and only moderately inhibited by RR. Low pH (pH 5) produced a remarkable increase in CGRP-LI outflow that was abolished after exposure to capsaicin, reduced by about 50% in Ca(2+)-free medium and unaffected by TTX (0.3 microM). The Ca(2+)-dependent component of the proton-evoked CGRP-LI release was abolished in the presence of RR (10 microM) and slightly inhibited by CTX (0.1 microM).(ABSTRACT TRUNCATED AT 250 WORDS)
Afferent Pathways↗
Intracerebroventricular administration of endothelins: effects on the supraspinal micturition reflex and blood pressure in the anaesthetized rat.
Intracerebroventricular (i.c.v.) administration of endothelin-1 (ET-1, 10 and 100 pmol/rat) or endothelin-3 (ET-3, 100 pmol/rat) inhibited the supraspinal micturition reflex (SMR) in urethane-anesthetized rats. Guanethidine pretreatment, vagotomy or forced ventilation did not modify the effect of i.c.v. ET-1 on the SMR. Intravenous (i.v.) administration of ET-1 (350 pmol/rat) also blocked the SMR with a delay of about 8 min. Guanethidine pretreatment revealed a stimulatory effect of i.v. ET-1 on the frequency of the SMR and the late inhibition was also reduced. In the isolated rat urinary bladder ET-1, ET-3 and ET-(16-21) induced a dose-related contraction and a small enhancement of the neurogenic contraction caused by electrical field stimulation. Inhibition of bladder motility by i.c.v. administration of ET-1 or ET-3 was paralleled by an increase in blood pressure. Both ET-1 and ET-3, at the same doses which evoked these autonomic effects, also killed some animals. The ET-1-induced increase in blood pressure was not prevented by guanethidine pretreatment or by spinal cord transection. Guanethidine pretreatment also did not modify the pressor response induced by i.v. ET-1. Bilateral cervical vagotomy enhanced the pressor response to i.c.v. ET-1. Forced ventilation prevented the death of some animals following i.c.v. administration of ET-1.
Animals↗
Tachykinin receptors in the guinea-pig isolated bronchi.
The aim of the study was to assess which tachykinin receptors mediate the contractile response in the guinea-pig isolated bronchi. Experiments with natural tachykinins and receptor-selective tachykinin agonists were performed in the absence or presence of peptidase inhibitors and in bronchi pretreated with phenoxybenzamine. Both NK-1 (substance P, substance P methylester and septide) and NK-2 (neurokinin A, [beta-Ala8]neurokinin A-(4-10) and MDL 28,564) receptor agonists produced concentration-dependent contraction. NK-3 agonists (senktide and [MePhe7]neurokinin B) were active only at high concentrations. Phenoxybenzamine pretreatment reduced the maximal response to NK-1 agonists and produced a rightward shift of the curve to NK-2 agonists, without depression of the maximum. Five tachykinin antagonists selective for the NK-1 (L 668,169) or the NK-2 (MEN 10,207, MEN 10,376, L 659,877 and R 396) receptor were tested against substance P methylester and [beta-Ala8]neurokinin A-(4-10). The results indicated that these receptor-selective antagonists maintain their characteristic even when tested in a multireceptor assay such as the guinea-pig bronchus. The rank order of potency of NK-2 antagonists against [beta-Ala8]neurokinin A-(4-10) was MEN 10,207 = MEN 10,376 greater than L 659,877 much greater than R 396. This pattern, with the observation of the full agonist activity of MDL 28,564, indicates that in addition to NK-1 receptors, NK-2 receptors also are present in the guinea-pig bronchi and belong to the same subtype (NK-2A) as present in the rabbit pulmonary artery.
Amino Acid Sequence↗
Hypertonic media produce Ca(2+)-dependent release of calcitonin gene-related peptide from capsaicin-sensitive nerve fibres in the rat urinary bladder.
Superfusion of slices of the rat urinary bladder with hypertonic NaCl produced a remarkable and concentration-dependent (150-280 mM) increase in the outflow of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI). This effect was completely abolished by pre-exposure of the tissue to capsaicin (10 microM for 20 min) or by superfusion with a Ca(2+)-free medium. Capsaicin (10 microM) was still able to release a consistent amount of CGRP-LI from tissue pre-exposed (20 min) to 280 mM NaCl. Similarly, hypertonic sucrose (160 mM added to the physiological salt solution) induced a consistent release of CGRP-LI that was abolished by capsaicin-pretreatment or in a Ca(2+)-free medium. The experiments demonstrate that hypertonic solutions activate the efferent function of capsaicin-sensitive neurons and suggest that this event may have some relevance in pathophysiological conditions of the lower urinary tract in which hypertonic urine may diffuse to submucosal layers.
Animals↗
Vasoactive intestinal polypeptide (VIP) and the specific motor response to capsaicin of the human isolated ileum.
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Calcitonin Gene-Related Peptide↗
Tachykinin receptors in the longitudinal and circular muscle of the human ileum.
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Aged↗
Sensory denervation with capsaicin reduces the liver collagen deposition induced by common bile duct obstruction in rats.
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Animals↗