Release Of Calcitonin Gene-related Peptide-like Immunoreactivity Research Articles

Modulation of Capsaicin-Sensitive Nerve Activation by Low pH Solutions in Guinea-Pig Lung

We have studied the stimulation of airways sensory nerves by low pH solutions and concomitantly induced bronchoconstriction. The effect of low pH buffer and lactic acid solutions at the same pH (5 and 6) were compared and the influence of low pH on the capsaicin effect was recorded. We have used the isolated guinea-pig perfused lung model taking the insufflation pressure as an indicator of bronchial smooth muscle tone while the calcitonin gene-related peptide-like immunoreactivity measured in the lung perfusate represented sensory nerves activation. Low pH buffer and lactic acid solution (3 and 4.1 mM) at the same pH of 5 and 6 induced pH-dependent bronchoconstriction and peptides release which were completely abolished after systemic pretreatment with capsaicin. Both responses were significantly inhibited after Ca2+-free infusion. Capsazepine (10(-6) M), a selective capsaicin antagonist, significantly reduced the calcitonin gene-related peptide-like immunoreactivity overflow evoked by all the solutions studied. Diclofenac (10(-5) M), a cyclooxygenase blocker, inhibited pH 5, pH 6 and lactic acid 3 mM (pH 6)-evoked peptide release, but not lactic acid 4.1 mM (pH 5). The functional response was not significantly modified after diclofenac while only the lactic acid 3 mM response was significantly reduced by capsazepine. There was a synergistic interaction between capsaicin and low pH buffer on calcitonin gene-related peptide-like immunoreactivity release and an additive effect on bronchoconstriction. It is concluded that in the isolated perfused guinea-pig lung, lactic acid and low pH buffer induced calcitonin gene-related peptide-like immunoreactivity release and bronchoconstriction by stimulation of capsaicin-sensitive C fibres via a pathway partly dependent of extracellular Ca2+. The mechanism of calcitonin gene-related peptide-like immunoreactivity release seems to be the same at pH 6, while differences are evident at pH 5 between low pH buffer and lactic acid. Our results also suggest that proton activity could exert a modulatory role on the capsaicin-sensitive sensory nerves by a mechanism which remains to be clarified.

Involvement of an increased spinal TRPV1 sensitization through its up-regulation in mechanical allodynia of CCI rats

Vanilloid receptor 1 (TRPV1) antagonists are known to attenuate the neuropathic pain symptoms in peripheral nerve injury models, but the mechanism(s) of their effect remains unclear. At the same time, the role of spinal TRPV1 in pain transduction system has not been fully understood. In this study, the role of spinal TRPV1 in mechanical allodynia in rat chronic constriction injury (CCI) model was investigated. Intrathecal administration of a selective TRPV1 antagonist, N-(4-tertiarybutylphenyl)-4-(3-cholorphyridin-2-yl)tetrahydropryazine-1(2H)-carbox-amide (BCTC) significantly attenuated mechanical allodynia in CCI rats at 100 and 300 nmol. In vitro, BCTC inhibited capsaicin (300 nM)-induced releases of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) and substance P-like immunoreactivity (SP-LI) from the rat spinal cord slice preparations with IC 50s of 37.0 and 36.0 nM, respectively, confirming that BCTC potently inhibits TRPV1 function in the rat spinal cord. TRPV1 expression levels in the spinal cord following CCI were quantified in by Western blot analysis. TRPV1 protein levels were significantly increased in the ipsilateral side of the lumbar spinal cord at 7 and 14 days following CCI surgery, but not in the contralateral side. Furthermore, capsaicin (300 nM)-evoked release of CGRP-LI was significantly higher in the ipsilateral spinal cord of CCI rats (14 days after surgery) than that of sham-operated rats. These findings suggest that an increased sensitization of the spinal TRPV1 through its up-regulation is involved in the development and/or maintenance of mechanical allodynia in rat CCI model.

Different ion channel control pH6-induced bronchoconstriction and calcitonin gene-related peptide release in the guinea-pig lung.

We have studied the bronchoconstriction and the release of calcitonin gene-related peptide-like immunoreactivity induced by perfusion of pH6 buffer in the isolated guinea-pig perfused lung. Both bronchoconstriction and peptide release were completely abolished after systemic capsaicin pretreatment. Ca(2+)-free pH6 buffer infusion also completely inhibited the bronchial response, whereas the calcitonin gene-related peptide-like immunoreactivity overflow was significantly reduced. omega-Conotoxine and omega-agatoxin IVA known as N-, L- and P-type Ca2+ channel blocker, respectively, and the Na+ channel blocker tetrodotoxin decreased significantly the pH6-induced bronchial response and calcitonin gene-related peptide like immunoreactivity overflow. Nifedipine was without influence suggesting the involvement of both P- and N-type Ca2+ channel as well as the activation of an axon reflex. Ruthenium red had a more pronounced reduction effect on the functional response than on the peptide release. Ryanodine and caffeine are both agents known to influence Ca2+ release from sarcoplasmic reticulum. Ryanodine significantly reduced both bronchoconstriction and calcitonin gene-related peptide-like immunoreactivity overflow. Caffeine as well as theophylline and the Na(+)-H+ blocker, dimethylamiloride, largely depressed the functional response while producing a significant increase of calcitonin gene-related peptide-like immuno-reactivity basal value. The pH6-induced peptide overflow was slightly inhibited after caffeine and dimethylamiloride pre-treatment whereas no significant change was observed after theophylline. It is concluded that multiple ion channels including different type of Ca2+ channels appear to participate in pH6-induced bronchoconstriction and calcitonin gene-related peptide-like immunoreactivity release in the guinea-pig lung.

Effects of diabetes on tissue content and evoked release of calcitonin gene-related peptide-like immunoreactivity from rat sensory nerves

We measured the evoked release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) from sensory nerve terminals in tracheas from control and diabetic rats using an in vitro perfusion system and also the CGRP-LI content of the vagus nerve and trachea. Diabetes caused a 29% ( P<0.05) reduction in CGRP-LI content of the vagus nerve and a decrease in CGRP-LI release from nerve endings in the trachea evoked by either capsaicin (30% decrease, P<0.05) or electrical field stimulation (50% decrease: P<0.05). In contrast, there was a 50% increase in the CGRP-LI content of the unstimulated trachea. Thus, diabetes induces an impairment in neuropeptide release from peripheral terminals of sensory nerves that corresponds to decreased levels in the supplying nerve but is not reflected in tissue measurements that incorporate nerve terminals. Impaired neuropeptide release may contribute to peripheral and central sensory dysfunction in diabetic rats.

Calcitonin gene-related peptide content, basal outflow and electrically-evoked release from monoarthritic rat spinal cord in vitro

In this study, Freund's adjuvant-induced monoarthritis in the rat hind paw was used to induce chronic pain and inflammation. In order to compare the basal outflow, electrically-evoked release and total content of calcitonin gene-related peptide like immunoreactivity (CGRP-LI) with previously reported changes in substance P (SP-LI), the lumbar enlargement of monoarthitic (complete Freund's adjuvant-treated, CFA rat) and control (incomplete Freund's adjuvant-treated, IFA rat) spinal cords were used. During the 4-wk period after injection, neither the basal nor the evoked release of CGRP-LI from CFA cords differed from controls. By contrast, we have previously reported that SP-LI release from CFA rat spinal cords was significantly higher than from controls, 21 days after inoculation with Freund's adjuvant. Electrically-evoked CGRP-LI release from 21-day CFA rat spinal cord slices was not modified by superfusion with a GABAB antagonist, CGP 36742 (100 μM) which could greatly increase SP-LI release. However, the release of both peptides was significantly increased to the same extent in IFA and normal tissue but to a lesser extent in CFA cords, by superfusion with the opioid antagonist naloxone (1 μM). In conclusion, CGRP-LI, unlike SP-LI, did not appear to be susceptible to any changes in the lumbar enlargement of the rat spinal cord during inflammation of the hind paw. In addition, CGRP-LI release was increased by antagonism of opiate but not GABAB receptors, suggesting that during chronic inflammation of one hind paw, the GABABergic system, unlike the opioid system, might be activated to selectively inhibit the enhanced SP-LI release but not CGRP-LI release which is not changed.

Effect of interleukin-1β on the release of substance P from rat isolated spinal cord

Superfusion of rat spinal cord slices with rat interleukin-1β resulted in a significant enhancement of electrically evoked substance P-like immunoreactivity with a maximal effect (> 2-fold increase) at 0.1 ng/ml, whereas higher concentration (10–50 ng/ml) of the cytokine inhibited (≈ 50%) the release of the neuropeptide. Interleukin-1β (0.1 ng/ml) potentiation of substance P-like immunoreactivity release was abrogated by co-perfusion with interleukin-1 receptor antagonist (10–100 ng/ml) or with indomethacin (1 μM). Superfusion of spinal cord with interleukin-1β inhibited electrically evoked calcitonin gene-related peptide-like immunoreactivity release. Modulation of substance P-like immunoreactivity release from the spinal cord by interleukin-1β may represent a mechanism responsible for the hyperalgesic action of the cytokine characteristic of the inflammatory response.

Effect of capsazepine on the release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) induced by low pH, capsaicin and potassium in rat soleus muscle.

1. We have determined the effect of the competitive antagonist capsazepine at the capsaicin receptor on the release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) from rat isolated soleus muscle induced by capsaicin (1 microM), by superfusion with low pH medium (pH 5) or by KCl (80 mM). 2. Each one of the three stimuli tested produced a marked CGRP-LI release. Total evoked release (fmol g-1) was 482 +/- 69, 169 +/- 20 and 253 +/- 43 for capsiacin, low pH medium and KCL, respectively. 3. Prior application of capsiacin (10 microM for 30 min followed by 30 min of washout) to produce capasaicin desensitization in vitro abolished CGRP-LI release induced by the three stimuli. 4. Capsazepine (1-100 microM, 45 min preincubation) inhibited the evoked CGRP-LI release. Capsaicin-induced release was significantly inhibited by 77, 92 and 96% with 10, 30 and 100 microM capsazepine, respectively. Low pH-induced release was inhibited by 78, 84, 88 and 93% with 3, 10, 30 and 100 microM capsazepine, respectively. KCl-induced release was significantly inhibited by 55 and 93% with 30 and 100 microM (but not with 10 microM) capsazepine, respectively. 5. These findings demonstrate that capsazepine prevents low pH- and capsaicin-induced CGRP-LI release from rat soleus muscle at concentrations which do not affect the release evoked by KCl. These findings imply a relationship between the action of low pH and activation of the capsaicin receptor. At high concentrations, capsazepine produces a nonspecific inhibitory effect on CGRP-LI release from peripheral endings of the capsaicin-sensitive primary afferent neurone.

Effect of bradykinin and prostaglandins on the release of calcitonin gene-related peptide-like immunoreactivity from the rat spinal cord in vitro.

1. The release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) from the dorsal horn of the rat spinal cord in vitro in response to dorsal root stimulation was measured by radioimmunoassay. 2. Stimulation of the dorsal roots (3 or 4 roots on each side) at 10 Hz for 5 min evoked a mean release (R1) of 134.3 +/- 17.5 (n = 10) fmol CGRP-LI; the release (R2) evoked by a second stimulation period 30 min later under control conditions was 77 +/- 10% (n = 10) of R1. Test compounds were applied to the preparation following release R1, and their effect calculated from the value of R2/R1. 3. Bradykinin (0.01-10 microM) had no significant effect on the basal release of CGRP-LI, but at 0.1-10 microM it increased 2-3 fold the release evoked by dorsal root stimulation. 4. This effect of bradykinin was prevented by indomethacin (10 microM), or by the B2-receptor antagonist, Hoe140 (1-10 microM). In the presence of Hoe140, bradykinin significantly reduced R2/R1; the explanation for this is not clear. 5. The B1-receptor agonist, Des-Arg9-bradykinin (10 microM), did not affect CGRP-LI release nor was the effect of bradykinin blocked by the B1-receptor antagonist, Des-Arg9-Leu8-bradykinin (10 microM). 6. Various prostaglandins were found to mimic the effect of bradykinin on CGRP-LI release. Their approximate order of potency was prostaglandin D2 (PGD2) = PGE1 > PGF2 alpha = PGE2; PGI2 was ineffective at 10 microM.7. Forskolin (30 muM) and 3-isobutyl l-methylxanthine (IBMX; 10 fM) also increased the evoked release of CGRP-LI.8. It is concluded that bradykinin acts on B2-receptors in the spinal cord, causing the formation ofprostanoids, which in turn cause an enhancement of neuropeptide release from primary afferent nerve terminals in the dorsal horn. This effect may be secondary to activation of adenylate cyclase. Because B2-receptors are mainly associated with primary afferent nerve terminals, it is likely that prostanoid production is also a function of these structures. Whether this action of bradykinin has any physiological function in nociceptive transmission remains unclear..

Adenosine A 1 receptors mediate the presynaptic inhibition of calcitonin gene-related peptide release by adenosine in the rat spinal cord

Electrical field stimulation evoked a reproducible outflow of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) from the dorsal half of the rat spinal cord, an effect which was abolished by prior application of capsaicin, tetrodotoxin or removal of extracellular Ca 2+. Adenosine (EC 50 3.2 μM) and the selective adenosine A 1 receptor agonist N 6-cyclohexyladenosine (EC 50 8.2 nM) inhibited evoked CGRP-LI outflow, while the selective adenosine A 2 receptor agonist CGS-21680 was ineffective up to 10 μM. The action of adenosine was prevented by the adenosine A 1 receptor selective antagonist 8-cyclopentyl-1,3-dipropylxanthine (30 μM), which did not affect CGRP-LI release on its own.

Veratridine evokes release of calcitonin gene-related peptide from capsaicin-sensitive nerves of rat urinary bladder

The effect of superfusion with veratridine on the release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) was studied in slices of rat urinary bladder. Exposure to veratridine (1–200 μM) produced a concentration-related release of CGRP-LI. Veratridine (50 μM)-evoked CGRP-LI release was abolished in slices pre-exposed to capsaicin (10 μM for 40 min) or superfused in a Ca 2+-free medium containing 1 mM EDTA. After exposure to veratridine (50 μM for 40 min), capsaicin (10 μM) was still able to release CGRP-LI. CGRP-LI release evoked by veratridine (50 μM) was inhibited by about 60% by tetrodotoxin (0,3 μM), attenuated (30%) by nifedipine (1 μM), and not affected by ω-conotoxin (0.1 μM). The capsaicin antagonist ruthenium red (10 μM) did not affect veratridine (50 μM)-evoked CGRP-LI release. The present results indicate that depolarization by veratridine induces CGRP-LI release from capsaicin-sensitive nerve fibres, an effect that is entirely dependent on extracellular Ca 2+. The Ca 2+ influx that promotes CGRP-LI release is mediated mostly by nifedipine-, ω-conotoxin- and ruthenium red-insensitive channels.

Capsaicin-like activity of N-ethylmaleimide in rat stomach

1. 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. 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. 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.

Different pathways by which extracellular Ca 2+ 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 Ca 2+, 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. Altough capsaicin (1 μM) 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 (1μM capsaicin for 20 min), high K + (80 mM) failed to evoke CGRP-LI release, whereas SP-LI release was still observed. The capsaicin (1 μM)-evoked CGRP-LI release was entirely dependent on extracellular Ca 2+. It was unaffected by 0.3 μM tetrodotoxin (TTX), slightly reduced by 0.1 μM ω-conotoxin (CTX) and blocked by 10 μM Ruthenium red (RR). The Ca 2+-dependent K + (80 mM)-evoked CGRP-LI release was unaffected by TTX, markedly reduced by CTX and only moderatedly 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 μM). The Ca 2+-dependent component of the proton-evoked CGRP-LI release was abolished in the presence of RR (10 μM) and slightly inhibited by CTX (0.1 μM). The mode by which capsaicin or high K + promote Ca 2+ entry into the central endings of capsaicin-sensitive afferents, and hence promote neuropeptide release may be distinguished on a pharmacological basis. Protons release CGRP in the spinal cord by a mechanism that shares a common pathway with that activated by capsaicin.

Calcitonin gene-related peptide in rat arterial and venous vessels: Sensitivity to capsaicin, bradykinin and FMLP

Sensitivity of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) to capsaicin was investigated in different arterial and venous tissues (mesenteric, renal and femoral artery and vein and carotid artery and jugular vein) of the rat. In vivo capsaicin pre-treatment depleted or reduced tissue CGRP-LI in all the vessels examined with the exception of the carotid artery. Likewise, in vitro exposure to capsaicin evoked release of CGRP-LI from all vessels, excluding the carotid artery. Remarkable amount of CGRP-LI sensitive to capsaicin was present in both mesenteric artery and vein as compared to the other vascular tissues. Endogenous (bradykinin) or exogenous (N-formyl-methionyl-leucyl-phenylalanine) proinflammatory agents, were found to release CGRP-LI from mesenteric veins.

Selectivity of ruthenium red in inhibiting bronchoconstriction and CGRP release induced by afferent C‐fibre activation in the guinea‐pig lung

In the present study we evaluated the effects of ruthenium red, a blocker of transmembrane Ca2+ fluxes, on bronchoconstriction and the release of calcitonin gene-related peptide-like immunoreactivity induced by different stimuli in the isolated perfused guinea-pig lung. Vagal stimulation (1 Hz, 1 min), capsaicin (10(-8) M, 10(-6) M), resiniferatoxin (3 x 10(-10) M), nicotine (10(-4) M), bradykinin (5 x 10(-6) M) and histamine (10(-5) M) evoked bronchoconstriction and calcitonin gene-related peptide-like immunoreactivity overflow. Ruthenium red (5 x 10(-6) M) almost completely inhibited the bronchoconstriction and calcitonin gene-related peptide-like immunoreactivity overflow induced by capsaicin and resiniferatoxin but did not influence the effects induced by vagal nerve stimulation, nicotine, bradykinin or histamine. The 20-deacetylated derivative of resiniferatoxin (ROPA), which lacks the homovanillyl ester group, did not evoke release or bronchoconstriction. Ruthenium red (3 x 10(-4) M) aerosol attenuated the cough induced by nebulized citric acid in conscious guinea-pigs. Citric acid-induced coughing is mediated via capsaicin-sensitive neurons. However, cigarette smoke-induced coughing, which involves capsaicin-resistant mechanisms, was not affected by ruthenium red. In conclusion, ruthenium red selectively inhibits the capsaicin, resiniferatoxin and citric acid-induced excitation of the sensory nerves as revealed by calcitonin gene-related peptide-like immunoreactivity release, bronchoconstriction and coughing, suggesting that these agents share a common mechanism of action.

The effect of thiorphan on release of sensory neuropeptides from guinea-pig cerebral venous sinuses

The effect of peptidase inhibitors on neuropeptide release from peripheral endings of capsaicin-sensitive sensory neurons was studied in cerebral superior sagittal and transverse sinuses of guinea-pig. Capsaicin (1 microM)-evoked release of substance P-like immunoreactivity (SP-LI) was increased in a concentration-dependent manner by thiorphan (0.1-10 microM). Captopril (10 microM) or a mixture of bestatin (10 microM), leupeptin (10 microM) and bacitracin (10 microM) did not affect the capsaicin-evoked SP-LI release. Thiorphan (10 microM) increased also the capsaicin-evoked release of neurokinin A-like immunoreactivity (TK-LI) and calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) by 228% and 172%, respectively, while captopril (10 microM) was without effect. Thiorphan (10 microM), but not captopril (10 microM), enhanced by 239% CGRP-LI release induced by bradykinin (10 microM). In the cerebral venous vessels neutral endopeptidase (EC 3.4.24.11, NEP)-like activity was 58.8 +/- 6.1 pmol/mg protein/min, while angiotensin converting enzyme-like activity was below the detection limit of the assay. A thiorphan-sensitive mechanism, putatively attributable to NEP, plays a major role in the inactivation of peptides released from or acting on capsaicin-sensitive sensory fibres of cerebral venous sinuses of guinea-pig.

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 μM for 20 min) or by superfusion with a Ca 2+-free medium. Capsaicin (10 μM) 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.

Neurochemical evidence of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) release from capsaicin-sensitive nerves in rat mesenteric arteries and veins

1. 1. The ability of capsaicin and antidromic stimulation of perivascular nerve fibers to release sensory neuropeptides (SP-LI and CGRP-LI) have been investigated in rat mesenteric arteries and veins. 2. 2. Both in mesenteric arteries and veins substantial SP-LI and CGRP-LI tissue levels were measured. A significant reduction in sensory neuropeptides levels was observed in tissues obtained from capsaicin-pretreated animals. 3. 3. Superfusion of isolated vessels with capsaicin (1 μM) produced a prompt and remarkable release of both SP-LI and CGRP-LI, which can be evoked only once in each preparation. 4. 4. Electrical field stimulation (EFS, 20 Hz, 50 V, 0.5 msec, trains of 10 sec every 20 sec for 15 min) of isolated vessels resulted in a significant release of CGRP-LI. This release was significantly greater in veins as compared to arteries. EFS-induced CGRP-LI release was unaffected by atropine or guanethidine and absent in preparations obtained from capsaicin-pretreated rats. 5. 5. These neurochemical findings further suggest that the local release of sensory neuropeptides from capsaicin-sensitive nerve endings might be important in the regulation of mesenteric circulation.

GABA A and GABA B receptors modulate the K +-evoked release of sensory CGRP from the guinea pig urinary bladder

Superfusion of mucosa-free muscle slices of guinea-pig urinary bladder with 40 mM K + produced a remarkable increase in calcitonin gene-related peptide-like immunoreactivity (CGRP-LI), that in this organ is entirely contained in capsaicin-sensitive nerves. GABA (1 mM) did neither affect the basal nor the 40 mM K + evoked CGRP-LI release. Baclofen (0.1 mM) or muscimol (1 mM) did not affect the basal CGRP-LI outflow. However, baclofen (0.1 mM) significantly reduced by 32% and muscimol (0.1-1 mM) significantly increased by 60% and 70%, respectively the K +- evoked CGRP-LI release. These findings add neurochemical evidence to the functional data suggesting the existence of GABA A and GABA B receptors which modulate the efferent function of capsaicin-sensitive afferents.

A spinal mechanism of action is involved in the antinociception produced by the capsaicin analogue NE 19550 (olvanil)

We have studied the effect of NE 19550 (olvanil, N-(4-hydroxy-3-methoxyphenyl) methyl-9Z-octadecenamide), a capsaicin analogue with approximately equipotent antinociceptive activity in vivo compared with capsaicin, on nociceptive responses recorded from spinal dorsal horn neurones in vivo and from a spinal ventral root in vitro. In adult rats anaesthetized with halothane, antinociceptive doses of olvanil (20–40 μmol/kg, s.c.) reduced C-fibre responses evoked in wide dynamic range, lumbar dorsal horn neurones, by peripheral transcutaneous electrical stimulation. Intradermal injection of olvanil, localized to a discrete region of the peripheral receptive field, did not activate C-fibres nor change C-fibre evoked activation of dorsal horn neurones. Spinal intrathecal administration of olvanil attenuated C-fibre evoked responses and, at the highest concentration, significantly reduced Aβ-fibre evoked activity. In the neonatal rat spinal cord/tail preparation maintained in vitro, superfusion of the cord with olvanil (500 nM-5 μ M) did not evoke a depolarization but responses to peripheral noxious stimulation were attenuated. In a similar in vitro preparation of the neonatal rat spinal cord, the release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) was measured in spinal cord superfusates. Capsaicin (2–10 μM) evoked a large release of CGRP-LI but olvanil (2–10 μM) produced only a small or undetectable release. Following the administration of each substance, however, the release of CGRP-LI evoked by a depolarizing potassium stimulus was significantly attenuated. These data indicate that C-fibre input to the dorsal horn was attenuated by acute systemic doses of olvanil that were antinociceptive in behavioural tests. This effect was unlikely to be due to impairment of C-fibre function by a peripheral locus of action but was more consistent with an action in the spinal cord in which the reduced release of a neurotransmitter substance from afferent nerve terminals may play a prominent role.

Desensitization of capsaicin-evoked neuropeptide release — Influence of Ca2+ and temperature

Capsaicin-induced stimulation and desensitization of neuropeptide release from primary afferent neurons was investigated in the rat urinary bladder in-vitro. The capsaicin (5 min contact time)-evoked release of calcitonin gene-related peptide-like immunoreactivity (CGRP-IR) was dose-dependent; threshold to produce detectable release was 0.1 mumol/l, the EC50 was 0.17 mumol/l. Pre-exposure of tissues to capsaicin (0.1-1.0 mumol/l, 5 min contact time) caused a dose-dependent reduction of the amount of CGRP-IR which was released by a second exposure to capsaicin. At 0.1 and 0.3 mumol/l, capsaicin was less effective to inhibit the subsequent K(+)-evoked release than that evoked by a second capsaicin exposure. Pre-exposure to 1 mumol/l capsaicin completely prevented subsequent K(+)- or capsaicin-evoked release of CGRP-IR. Exposure of the preparation to capsaicin (0.3 mumol/l) in a Ca2(+)-free, EDTA-containing medium did not produce release of CGRP-IR. A subsequent stimulation with capsaicin in a 2.5 mmol/l Ca2(+)-containing superfusion solution was not less effective to release CGRP-IR than in tissues which had not been pre-exposed to capsaicin. At 18 degrees C, the capsaicin-evoked release of CGRP-IR was reduced to 20% of the value obtained by the same dose (0.3 mumol/l for 5 min) of capsaicin at 37 degrees C. Comparison of the desensitizing effect of 0.3 and 0.1 mumol/l capsaicin at 18 degrees C and 37 degrees C, respectively, showed significant inhibition of desensitization at 18 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)