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Effects of clonixin on the transmitter release at the mouse neuromuscular junction
C A L C I U M CHANNEL BLOCKERS POTENTIATE THE A N A L G E S I C E F F E C T OF A C E T A M I N O P H E N 1N MICE M. Koleva and S. Dimova Institute of Physiology, Bulg. Acad. Sci., Acad. G. Bonchev St., BL 23, 1113 Sofia, Bulgaria. Calcium channel blockers (CCBs) have been used for the treatment of angina and hypertension, and as calcium channels exist in the brain, the action of the blockers on the CNS are also of interest. The possibility that CCBs might produce antinociception and increase acetaminophen (APAP) analgesia was studied using the acetic acid writhing test in mice. CCBs were administered orally at the following doses: nifedipine (NF) - 50 mg/kg, verapamil (V) - 20 mg/kg and diltiazem (DL) - 70 mg/kg. APAP was applied orally at a dose of 100 mg/kg, alone or lh after NF, V and DL. The CCBs studied showed a significant antinociceptive effect compared to the controls. Antinociception was more pronounced in NF treated mice: NF, V and DL applied lh before APAP significantly potentiated its analgesic effect, which was more pronounced in NF pretreated mice. The antinociceptive effect of V and APAP was additive. NF increased cytochrome P-450 content, NADPH-cytochrome c reductase and ethylmorphine-N-demethylase activities. V and DL did not change the drug metabolizing enzyme activities studied, suggesting that their effect on APAP analgesia is not associated with changes in the APAP metabolism. These results well documented the antinociceptive effect of NF, V and DL and their ability to potentiate APAP analgesia, which could be explained by the calcium channel blocking action of the drugs.
Dexamethasone reduced clonidine-induced antinociception in mice Stefano Pieretti 1 Amalia Di Giannuario 1, Ludovico Sorrentino 2 Anna Capasso2 and Alberto Lolzzo 1 1Laboratorio di Farmacologia, Istituto Superiore di.Sanit&, Roma and 2School of Pharmacy, University of Salerno, Salerno, Italy. The effects of dexamethasone pretreatment on clonidine-induced antinociception were investigated in mice. In the hot plate and tail flick tests dexamethasone administered intraperitoneally (1 mg/kg) 15 min before clonidine did not change clonidine antinociception, whereas administered 30 or 60 min before clonidine, it reduced clonidine antinociception in the hot plate and tail flick tests. A lower dexamethasone dose (0.1 mg/kg) administered 30 min before clonidine was not able to change clonidine-induced effects, while higher dexamethasone doses (0.5 and 10 mg/kg) induced the same effect observed at t h e dose of 1 mg/kg. Dexamethasone administered centrallyi(1, 5 and 10 ng) 30 min before clonidine was also able to reduce clonidine-induced antinociception. Cycloheximide administered at the dose of 10 mg/kg 2 h before clonidine did not change clonidine-induced effects in the hot plate test and in the tail flick test, but was able to prevent dexamethasone effects on clonidine-induced antinociception. The glucocorticoid receptor antagonist RU38486 administered centrally at the dose of 1 ng did not change clonidine-induced effects, whereas it was able to block dexamethasone effects on clonidineinduced antinociception. These results may suggest that dexamethasone effects on clonidine-induced antinociception depend on the stimulating effects that dexamethasone exert on the protein synthesis via the glucocorticoid receptor in the brain.
HISTAMINE HI-ANTAGONISTS ANTINOCICEPTIONIN MICE AND INHIBITION OF HISTAM1NE-N-METHYLTRANSFERASE C. Lamberti, A. Bartolini, C. Ghelardini and P. Malmberg-Aiello Dept. of Pharmacology, V.le Morgagni 65, 1-50134-Florence It is known that histamine Hi-receptor antagonists are able to raise the pain threshold in both laboratory animals and humans. Histamine itself and histamine-N-methyltransferase (HMT) inhibitors have also been reported to be antinociceptive, and since some Hi-antagonists are known to be able to inhibit histamine catabolism in vitro in the presence of low histamine concentrations (1), it seemed worthwhile to investigate whether their antinociception is due to HMT inhibition by means of the mouse hot plate (52.5°C) and acetic acid abdominal constriction tests. For this purpose the H3-receptor agonist, (R)-cx-methylhistamine 2HC1 (RAMH), was used at a dose (10 mg/kg i.p.) which did not alter the pain threshold, but was still able to block histamine release and synthesis. The two Hi-receptor antagonists used (diphenhydramine HC1, DIP and pyrilamine maleate, PYR) induced a dose-dependent antinociception in both tests. The dose which was significantly antinociceptive but did not impair animal performance in the rota rod test (24 r.p.m.) (20 mg/kg s.c. for bot h substances) was also tested in combination with RAMH, administered 15 min before. RAMH was not able to antagonize the antinociception induced by the two HI-blockers in either of the two tests. Since RAMH at this dose has been previously described to antagonize the antinociception yielded by a potent HMT inhibitor, metoprine (2), it can be suggested that the antinociception caused by DIP and PYR is not due to inhibition of HMT. 1) Taylor & Snyder, Mol. Pharmacol., 8, 300, 1972. 2) Malmberg-Aiello et al., Br. J. Pharmacol. 111, 1269, 1994.
EFFECTS OF CLONIXlN ON THE TRANSMITTER RELEASE AT THE MOUSE NEUROMUSCULAR JUNCTION L. Re ,C. Concettoni, H. Ponce Monter:l:, MA. Morales:[: IMSC-Pharmacology, Via Ranieri, 2, 60131 Ancona, Italy and ¢IMSS, Siglo XXI, Mexico DF, Mexico. Clonixin (CIx) is a non-steroid analgesic used in humans in oral and parenteral form. Many Authors have suggested that although there is a strong resemblance between the analgesic effect of CIx and opiates, CIx shows a central action that is not mediated by tH, 8 or ~-opioid receptors and that the mechanism of action of CIx remains unclear. Studies using vas deferens have shown that CIx should produce a significant reduction of muscular twitch induced by transmural neurogenic stimulation. It has been proposed that norepinephdne release should be altered probably through calcium channel blockade. However, direct proofs in support of a modulation of neurosecretion by CIx are still lacking. In this work the loose patch clamp technique is used to further elucidate CIx action. The neuromuscular preparation, in which the acetylcholine (ACh) release is linked to calcium channels, should be a very useful model to this study, In the mouse neuromuscular junction Clx induces a very weak effect on the evoked and spontaneous release of ACh. The end-plate currents and the spontaneous currents were reduced at 100 I~M CIx of 31.5 and 8.3 % respectively. These effects indicate that probably the calcium channel type modulated by CIx could be of the N-type. Indeed, recent studies carded out using the blocker (o-conotoxin, an N-type specific blocker, demonstrate that this drug fails to block release at mouse junction being effective only on the neuromuscular junction of different species such as frog or snake.
Acknowledgements The support of the Italian CNR and MURST 40% given to L. Re is gratefully acknowledged.
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Dexamethasone reduced clonidine-induced antinociception in mice Stefano Pieretti 1 Amalia Di Giannuario 1, Ludovico Sorrentino 2 Anna Capasso2 and Alberto Lolzzo 1 1Laboratorio di Farmacologia, Istituto Superiore di.Sanit&, Roma and 2School of Pharmacy, University of Salerno, Salerno, Italy. The effects of dexamethasone pretreatment on clonidine-induced antinociception were investigated in mice. In the hot plate and tail flick tests dexamethasone administered intraperitoneally (1 mg/kg) 15 min before clonidine did not change clonidine antinociception, whereas administered 30 or 60 min before clonidine, it reduced clonidine antinociception in the hot plate and tail flick tests. A lower dexamethasone dose (0.1 mg/kg) administered 30 min before clonidine was not able to change clonidine-induced effects, while higher dexamethasone doses (0.5 and 10 mg/kg) induced the same effect observed at t h e dose of 1 mg/kg. Dexamethasone administered centrallyi(1, 5 and 10 ng) 30 min before clonidine was also able to reduce clonidine-induced antinociception. Cycloheximide administered at the dose of 10 mg/kg 2 h before clonidine did not change clonidine-induced effects in the hot plate test and in the tail flick test, but was able to prevent dexamethasone effects on clonidine-induced antinociception. The glucocorticoid receptor antagonist RU38486 administered centrally at the dose of 1 ng did not change clonidine-induced effects, whereas it was able to block dexamethasone effects on clonidineinduced antinociception. These results may suggest that dexamethasone effects on clonidine-induced antinociception depend on the stimulating effects that dexamethasone exert on the protein synthesis via the glucocorticoid receptor in the brain.
HISTAMINE HI-ANTAGONISTS ANTINOCICEPTIONIN MICE AND INHIBITION OF HISTAM1NE-N-METHYLTRANSFERASE C. Lamberti, A. Bartolini, C. Ghelardini and P. Malmberg-Aiello Dept. of Pharmacology, V.le Morgagni 65, 1-50134-Florence It is known that histamine Hi-receptor antagonists are able to raise the pain threshold in both laboratory animals and humans. Histamine itself and histamine-N-methyltransferase (HMT) inhibitors have also been reported to be antinociceptive, and since some Hi-antagonists are known to be able to inhibit histamine catabolism in vitro in the presence of low histamine concentrations (1), it seemed worthwhile to investigate whether their antinociception is due to HMT inhibition by means of the mouse hot plate (52.5°C) and acetic acid abdominal constriction tests. For this purpose the H3-receptor agonist, (R)-cx-methylhistamine 2HC1 (RAMH), was used at a dose (10 mg/kg i.p.) which did not alter the pain threshold, but was still able to block histamine release and synthesis. The two Hi-receptor antagonists used (diphenhydramine HC1, DIP and pyrilamine maleate, PYR) induced a dose-dependent antinociception in both tests. The dose which was significantly antinociceptive but did not impair animal performance in the rota rod test (24 r.p.m.) (20 mg/kg s.c. for bot h substances) was also tested in combination with RAMH, administered 15 min before. RAMH was not able to antagonize the antinociception induced by the two HI-blockers in either of the two tests. Since RAMH at this dose has been previously described to antagonize the antinociception yielded by a potent HMT inhibitor, metoprine (2), it can be suggested that the antinociception caused by DIP and PYR is not due to inhibition of HMT. 1) Taylor & Snyder, Mol. Pharmacol., 8, 300, 1972. 2) Malmberg-Aiello et al., Br. J. Pharmacol. 111, 1269, 1994.
EFFECTS OF CLONIXlN ON THE TRANSMITTER RELEASE AT THE MOUSE NEUROMUSCULAR JUNCTION L. Re ,C. Concettoni, H. Ponce Monter:l:, MA. Morales:[: IMSC-Pharmacology, Via Ranieri, 2, 60131 Ancona, Italy and ¢IMSS, Siglo XXI, Mexico DF, Mexico. Clonixin (CIx) is a non-steroid analgesic used in humans in oral and parenteral form. Many Authors have suggested that although there is a strong resemblance between the analgesic effect of CIx and opiates, CIx shows a central action that is not mediated by tH, 8 or ~-opioid receptors and that the mechanism of action of CIx remains unclear. Studies using vas deferens have shown that CIx should produce a significant reduction of muscular twitch induced by transmural neurogenic stimulation. It has been proposed that norepinephdne release should be altered probably through calcium channel blockade. However, direct proofs in support of a modulation of neurosecretion by CIx are still lacking. In this work the loose patch clamp technique is used to further elucidate CIx action. The neuromuscular preparation, in which the acetylcholine (ACh) release is linked to calcium channels, should be a very useful model to this study, In the mouse neuromuscular junction Clx induces a very weak effect on the evoked and spontaneous release of ACh. The end-plate currents and the spontaneous currents were reduced at 100 I~M CIx of 31.5 and 8.3 % respectively. These effects indicate that probably the calcium channel type modulated by CIx could be of the N-type. Indeed, recent studies carded out using the blocker (o-conotoxin, an N-type specific blocker, demonstrate that this drug fails to block release at mouse junction being effective only on the neuromuscular junction of different species such as frog or snake.
Acknowledgements The support of the Italian CNR and MURST 40% given to L. Re is gratefully acknowledged.
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