Eicosanoid-induced Ca2+ release and sustained contraction in Ca(2+)-free media are mediated by different signal transduction pathways in rat aorta.

1. The effects of 12-O-tetradecanoyl 4 beta-phorbol 13-acetate (beta-TPA) on the inositol 1,4,5-trisphosphate (IP3) production, Ca2+ release from the intracellular Ca2+ stores and sensitization of contractile apparatus, induced by prostaglandin F2 alpha (PGF2 alpha) and U46619, a thromboxane A2-mimetic, were studied, using fura-2-loaded and -unloaded rat thoracic aortic strips. 2. Both eicosanoids had characteristic patterns of responses in Ca(2+)-free, 2 mM EGTA-containing solution (Ca(2+)-free solution). They induced transient increases in intracellular Ca2+ concentration ([Ca2+]i) without corresponding transient contraction, but produced delayed, sustained contraction, where [Ca2+]i was returned to the basal level. 3. Treatment with beta-TPA for 60 min reduced the eicosanoids-induced IP3 production, suggesting that the treatment inhibits PIP2 breakdown. 4. The treatment also attenuated [Ca2+]i transient induced by the eicosanoids, but not by caffeine (an IP3-independent releaser of stored Ca2+), in fura-2-loaded preparations incubated in Ca(2+)-free solution. 5. In contrast in the presence of beta-TPA, the sustained contractions evoked by the eicosanoids in Ca(2+)-free solution were potentiated, suggesting that the sites of actions of beta-TPA and the eicosanoids may differ from each other. 6. PGF2 alpha and U46619 utilize different and parallel signal transduction pathways to release Ca2+ by IP3 produced by PIP2 breakdown (beta-TPA-sensitive), and to increase the sensitivity of contractile apparatus, in which protein kinase C may not be involved (beta-TPA-insensitive).

Contrasting effects of tachykinins and guanethidine on the acetylcholine output stimulated by nicotine from guinea-pig bladder [corrected].

1. Contractile responses and acetylcholine release evoked by nicotine in guinea-pig detrusor strips were determined by isotonic transducer and radioimmunoassay, respectively. Nicotine stimulated acetylcholine release and a contractile response in guinea-pig detrusor strips treated with the cholinesterase inhibitor, methanesulphonyl fluoride (MSF). Both actions evoked by nicotine were antagonized by the nicotinic receptor antagonist, hexamethonium but were insensitive to tetrodotoxin. 2. A sympathetic nerve blocker, guanethidine and a tachykinin antagonist, [D-Arg1, D-Pro2, D-Trp7,9, Leu11]-substance P (rpwwL-SP) partially inhibited the acetylcholine release evoked by nicotine to much the same degree. The inhibitory effects of guanethidine and rpwwL-SP on acetylcholine release were significantly greater than corresponding effects on the contraction evoked by nicotine. 3. In preparations treated with rpwwL-SP to block the tachykinin receptors, guanethidine had no effect on the response to nicotine. Conversely, after treatment with guanethidine to block release of a mediator from sympathetic nerve endings, nicotine-induced responses were not affected by rpwwL-SP. 4. Nicotine-induced contraction was reduced to 30% by the muscarinic cholinoceptor antagonist, atropine and completely abolished after desensitization of P2-purinoceptors with alpha,beta-methylene ATP in the presence of atropine. 5. A concentration-contractile response curve to neurokinin A (NKA) was shifted to the left after cholinesterase inhibition with MSF. Atropine abolished the facilitatory effect of MSF and partially inhibited contractions induced by NKA at 100 nM to 1 microM. The contractile responses to substance P methyl ester (SPOMe) and Tyr0-neurokinin B (Tyr0-NKB) were not influenced by MSF or atropine. 6. After desensitization of NK, tachykinin receptors with SPOMe or preincubation with senktide, the cholinergic component of the nicotine-induced contraction was the same as the control value (100%). 7. Our findings give further support to our previous results: nicotine stimulates acetylcholine release in a tetrodotoxin-resistant manner in guinea-pig bladder and acetylcholine release evoked by nicotine is increased by the coordinated action of sympathetic nerves and tachykinin(s). It is suggested that the tachykinin receptor subtype involved in acetylcholine release is NK,.

Characterization of endothelin receptor subtypes mediating Ca2+ mobilization and contractile response in rabbit iris dilator muscle.

1. We investigated the characteristics of endothelin (ET)-induced contraction and changes in intracellular Ca2+ concentration ([Ca2+]i) using the fura-2-loaded and non-loaded rabbit iris dilator. ET-1 and ET-2 (3-100 nM) and ET-3 (30-100 nM) caused contraction in a concentration-dependent fashion. 2. The selective ETB-receptor agonists, IRL1620 and sarafotoxin S6c produced only a small contraction or no contraction at a concentration of 1 microM. The rank order of potencies for the contraction (pD2 value) was ET-1 = ET-2 > ET-3 >> sarafotoxin S6c = IRL1620. 3. The contractile response to ET-3 was antagonized by pretreatment with BQ-123 (10 nM), a selective ETA receptor antagonist. The contractile responses to ET-1 and ET-2 were antagonized by pretreatment with BQ-123 (10 microM), but not at a concentration of 10 nM. 4. ETs increased [Ca2+]i and sustained muscle contraction. ET-1 (100 nM), ET-2 (100 nM), and ET-3 (1 microM) induced an elevation of [Ca2+]i consisting of two components: first a rapid and transient elevation to reach a peak, followed by a second, sustained elevation; a sustained contraction was produced without a transient contraction. The ETB receptor-selective agonist, IRL1620 (1 microM) and sarafotoxin S6c (1 microM) also induced a rapid and transient elevation of [Ca2+]i to reach a peak and a sustained elevation, together with only a small contraction or no contraction. 5. ET-1 (100 nM) induced a transient increase in [Ca2+]i in a Ca(2+)-free, 2 mM EGTA-containing physiological saline solution (Ca(2+)-free PSS), and a small sustained contraction which was significantly different from that induced by ET-1 (100 nM) in normal PSS. The ET-1-induced increase in [Ca2+]i and sustained contraction were not affected by the voltage-dependent Ca2+ channel blocker, nicardipine (10 microM). The ET-1-induced transient increase in [Ca2+]i was significantly reduced by the sarcoplasmic reticulum (SR) Ca(2+)-ATPase inhibitor, cyclopiazonic acid (30 microM); however, the ET-1-induced sustained contraction was not affected by this agent. 6. The selective ETA receptor antagonist, BQ-123 (100 nM) reduced the ET-3 (100 nM)-induced contraction, but did not affect the transient increase or elevation of the second phase of [Ca2+]i. However, this antagonist at 1 microM did not affect the ET-1 (100 nM)- and ET-2 (100 nM)-induced elevation of [Ca2+]i and contractile response, or the IRL1620-induced elevation of [Ca2+]i. 7. The selective ETB receptor antagonist, BQ-788 (1 microM) reduced the transient increase in [Ca2+]i induced by ET-1 (30 nM), ET-2 (30 nM), ET-3 (100 nM) and IRL1620 (1 microM), but did not affect the sustained elevation of [Ca2+]i and contractile responses produced by ET-1, ET-2 and ET-3. 8. Pretreatment with IRL1620 (1 microM) reduced the increase in [Ca2+]i induced by IRL1620 (1 microM) and sarafotoxin S6c (1 microM), as well as the ET-1 (100 nM)-, ET-2 (100 nM)- and ET-3 (1 microM)-induced elevation of [Ca2+]i, whereas in the presence of IRL1620, ET-1-, ET-2- and ET-3-induced contractions were unaltered. 9. These results suggest that ETA and ETB receptor subtypes exist in the rabbit iris dilator muscle, and that the ETA receptor is divided into: (1) BQ-123-sensitive ETA subtypes activated by ET-1, ET-2 and ET-3, and (2) BQ-123-insensitive ETA subtypes activated by ET-1 and ET-2, which cause the sustained increase of [Ca2+]i and contraction; in contrast, ETB receptor subtypes are activated by ET-1, ET-2, ET-3, IRL1620 and sarafotoxin S6c and cause the transient and sustained increase in [Ca2+]i which is not able to contract the smooth muscle.

Tachykinin receptors of the NK2 type involved in the acetylcholine release by nicotine in guinea-pig bladder.

1. The effects of guanethidine and tachykinins on nicotine- and electrical stimulation-induced cholinoceptor responses were studied in isolated urinary bladder from the guinea-pig. 2. Acetylcholine release and the contractile response stimulated by nicotine were partially reduced by a sympathetic nerve blocker, guanethidine. Neurokinin A (but not substance P methyl ester or senktide) enhanced both acetylcholine release and contraction by nicotine in the presence of guanethidine. 3. Frequency-contraction curves (1 to 50 Hz) for electrical field stimulation (EFS) were partially reduced by atropine (1 microM), and after desensitization to alpha,beta-methylene adenosine 5'-triphosphate, the atropine-resistant contraction to EFS was completely abolished. Guanethidine, the tachykinin antagonist [D-Arg1, D-Pro2, Trp7,9, Leu11]-substance P and application of neurokinin A or substance P did not change the contractile response to EFS. Preganglionic nerve stimulation (5 Hz and 20 Hz) also evoked a similar response to EFS and was not influenced at all by guanethidine or neurokinin A. 4. We conclude that the ability of nicotine to release acetylcholine is enhanced both by endogenous tachykinins (probably released from sympathetic nerves) and by exogenously applied tachykinins as a result of interaction with NK2 receptors in the urinary bladder.

Ryanodine reveals multiple contractile and relaxant mechanisms in vascular smooth muscle: simultaneous measurements of mechanical activity and of cytoplasmic free Ca2+ level with fura-2.

1. The effects of ryanodine on changes in cytoplasmic Ca2+ level ([Ca2+]i) and muscle tension induced by maximum concentrations of phenylephrine (Phe; 1 microM), prostaglandin F2 alpha (PGF2 alpha, 10 microM), caffeine (Caf, 30 mM) and isoprenaline (Iso, 1 microM) were examined in rat aortic strips using fura-2. 2. In normal media, Phe and PGF2 alpha produced a phasic contraction, followed by a tonic one. Caf elicited only a transient contraction. When the preparation was treated with 10 microM ryanodine, an increase in [Ca2+]i was induced accompanied by a nicardipine (1 microM)-resistant contraction which was [Ca2+]o-dependent. 3. In Ca2(+)-free solution, the three stimulants elicited transient increases in [Ca2+]i. Transient contractions to Phe and Caf were accompanied by changes in [Ca2+]i. The transient increase in [Ca2+]i induced by PGF2 alpha was not accompanied by a corresponding contraction. 4. Sustained contractions were induced by Phe and PGF2 alpha in the absence of external Ca2+, while the increase in [Ca2+]i was reduced. A larger maximum contraction was induced by PGF alpha than by Phe. 5. Ryanodine abolished both the Caf- and Phe-induced [Ca2+]i transient increases and the corresponding contractions, but had no substantial effect on the PGF2 alpha-induced [Ca2+]i transient increase. Ryanodine had no influence on the sustained contractions induced by Phe and PGF2 alpha. 6. Iso relaxed both sustained contractions almost completely, without any detectable change in [Ca2+]i. Treatment of the preparation with ryanodine had no effect on the concentration-response curves for Iso in relaxing the 0.1 microM Phe- or 40 mM K(+)-induced precontraction. 7. It is suggested that Phe and Caf mobilize Ca2 + from a ryanodine-sensitive Ca2 + store and that PGF2 alpha. releases Ca2+ from a ryanodine-insensitive Ca2+ store. The former contributes to the transient contraction through a Ca2'-dependent process, while the latter seems not to be directly associated with the contraction. The sustained contraction under Ca2+-free conditions might involve a Ca2 '-independent process or a change in the sensitivity of the contractile filaments to Ca2 + 8. In addition to lowering cytoplasmic Ca2+ concentration, it is suggested that Iso counteracts the apparently Ca2 +-independent process. The ryanodine-sensitive Ca2+ store plays no substantial role in active relaxation by Iso, although it does play a major role in the maintenance of cytoplasmic Ca2+ in a quiescent muscle.

Mechanism of action of nicotine in isolated urinary bladder of guinea-pig.

1. Nicotine produced a transient contraction of isolated strips of guinea-pig urinary bladder. The response to nicotine was antagonized by the nicotinic receptor antagonist, hexamethonium but was insensitive to tetrodotoxin. 2. The nicotine-induced contraction was potentiated by the cholinesterase inhibitor, physostigmine, and was reduced to 50% and 70% by the muscarinic cholinoceptor antagonist, atropine and the sympathetic neurone blocking drug, guanethidine, respectively. Chemical denervation with 6-hydroxydopamine abolished the inhibitory effect of guanethidine. Simultaneous treatment with atropine and guanethidine did not abolish the response to nicotine, but the degree of inhibition was comparable to that obtained with atropine alone. 3. The nicotine-induced contraction was insensitive to bunazosin and yohimbine (alpha 1- and alpha 2-adrenoceptor antagonists, respectively), and exogenously applied noradrenaline did not cause a contraction even in the presence of blockade of noradrenaline uptake mechanisms with desipramine and normetanephrine and of beta-adrenoceptors with propranolol, suggesting a non-adrenergic nature of the sympathomimetic effect of nicotine in this tissue. 4. The nicotine-induced contraction in the presence of atropine was abolished after desensitization of P2-purinoceptors with alpha, beta-methylene adenosine 5'-triphosphate, a slowly degradable ATP analogue selective for P2-purinoceptors. By this desensitization, the response to ATP, but not to histamine, was also abolished. 5. A cyclo-oxygenase inhibitor flurbiprofen partially inhibited the nicotine-induced contraction. The degree of the inhibition was more pronounced in the presence of atropine than in its absence. Flurbiprofen antagonized the response to exogenously applied ATP in an unsurmountable manner, but not that to carbachol. 6. The present results suggest that nicotine might induce a contraction through an interaction with nicotinic receptors located on the terminals of, possibly, (i) parasympathetic cholinergic, (ii) sympathetic non-adrenergic and (iii) non-sympathetic purinergic nerves in guinea-pig detrusor preparations, and that a portion of the contraction due to the purine nucleotide released is possibly potentiated by intramural prostaglandin(s). Parasympathetic cholinergic output might be modulated by an unknown excitatory substance released by nicotine from sympathetic nerve. 7. Nicotine reveals a latent excitatory effect of the sympathetic hypogastric nerve which innervates guinea-pig detrusor.

Mechanism of action of nicotine in isolated iris sphincter preparations of rabbit.

1. Nicotine produced a transient contraction of rabbit isolated iris sphincter muscle, a parasympathetic ganglion-free tissue. The response to nicotine was antagonized by hexamethonium, but was insensitive to tetrodotoxin (TTX). While single treatments with atropine, capsaicin or [D-Arg1, D-Pro2, D-Trp7,9, Leu11]-substance P (rpwwL-SP) partially blocked the response, combined treatment abolished it. 2. Chronic treatment of animals with nicotine added to the drinking water (about 12 mg kg-1 per day) had no effect on the responsiveness to nicotine or the pharmacological properties of nicotine-induced contraction. 3. These results suggest that acetylcholine and tachykinin(s) released via sodium channel-independent mechanisms from nerve terminals of parasympathetic and primary sensory nerves, respectively, are involved in the nicotine-induced contractile response.

Effects of cyclic AMP and protein kinase on calcium uptake in a microsomal fraction from guinea pig taenia caecum.

A microsomal fraction was isolated from guinea pig taenia caecum by differential centrifugation. Activities of ouabain-sensitive (Na+, K+)-ATPase, 5'-nucleotidase and NADPH-cytochrome c reductase were enriched in the microsomal fraction. On the other hand, less cytochrome c oxidase and monoamine oxidase were contained in this fraction. These results suggest that the microsomal fraction used in this study was derived from both sarcolemma and sarcoplasmic reticulum. Ca2+ uptake by this fraction was strictly dependent on the presence of ATP and was facilitated by oxalate. An ATP-regenerating system was required for the determination of Ca2+ uptake, when a lower concentration of ATP (e.g. 0.25 mM) was used. Phosphorylation of the microsomal fraction was doubled when these membranes were incubated in the presence of cyclic AMP plus cyclic AMP-dependent protein kinase (protein kinase). When the microsomal fraction was pretreated with cyclic AMP plus protein kinase, Ca2+ uptake was stimulated. The increases in microsomal phosphorylation and Ca2+ uptake were significantly correlated (P less than 0.01). This stimulation of Ca2+ uptake by microsomal phosphorylation was observed only in the presence of protein kinase, oxalate, and low Ca2+ and Mg2+ concentrations. The results suggest that stimulation of Ca2+ uptake may be the mechanism by which cyclic AMP is involved in beta-adrenergic relaxation of smooth muscle.

Effect of tachykinins on the acetylcholine output stimulated by nicotine from guinea-pig bladder.

Effects of tachykinins on contractile responses and acetylcholine release evoked by nicotine were determined by isotonic transducer and radioimmunoassay. A sympathetic nerve blocker guanethidine and tachykinin antagonist, [D-Arg1,D-Pro2,D-Trp7,9,Leu11]-substance P (rpwwL-SP) partially inhibited the acetylcholine release evoked by nicotine to much the same degree. In preparations treated with rpwwL-SP to block the tachykinin receptors, guanethidine had no effect on the response to nicotine and vice versa, suggesting an exclusive contribution of the sympathetic nerve communications to the action of tachykinins. A concentration response curve to neurokinin A but not SP methyl ester or Tyr0-neurokinin B was shifted to the left after cholinesterase inhibition with methanesulphonyl fluoride (MSF) and was partially inhibited by atropine. In the presence of guanethidine, an acetylcholine output by nicotine was facilitated in the presence of neurokinin A (10 nM) but not SP methyl ester (10 nM) or senktide (10 nM). These results indicate that acetylcholine release evoked by nicotine is increased by the coordinated action of sympathetic nerves and tachykinins. Tachykinin receptor subtype involved in acetylcholine release is NK2.

Possible involvement of substance P immunoreactive nerves in the mediation of nicotine-induced contractile responses in isolated guinea pig bronchus.

Nicotine-induced contraction of the isolated guinea pig bronchial preparation was abolished by capsaicin and a substance P (SP) antagonist [( D-Arg1,D-Pro2,D-Trp7,9,Leu11]SP). Nicotine increased the release of immunoreactive SP from the preparations. The nicotine-evoked release of immunoreactive SP from the bronchial preparation was reduced by hexamethonium but not by tetrodotoxin. The results indicate that the responses to nicotine of the guinea pig bronchial preparation were mediated through the release of SP-like material(s), and that the nicotine-induced response may be produced through a process independent of the sodium action potential. In conclusion, the most likely site of action of nicotine in the isolated guinea pig bronchial preparation is the nicotinic receptor of SP immunoreactive nerves.

Effects of external calcium ions and strontium ions on interactions of isoprenaline and its competitive inhibitor with beta-adrenoceptors in the intestinal smooth muscle.

The effects were tested of Ca ions and Sr ions on interactions of isoprenaline and its competitive inhibitor with beta-adrenoceptors in the taenia caecum from the guinea pig. Ca ions were involved in the combination of isoprenaline with the beta-adrenoceptors but not involved in the combination of the competitive inhibitor with the beta-adrenoceptors. Sr ions could not substitute for Ca ions in the isoprenaline--receptor interaction.

Comparison of the muscarinic cholinoceptors in the rabbit ciliary body and the guinea-pig ileum.

Interactions of several muscarinic drugs with their receptors were studied in the ciliary body smooth muscle of rabbits and the ileal longitudinal muscle of guinea-pigs, using pharmacological and biochemical procedures. The dissociation constants of carbachol, pilocarpine, atropine and pirenzepine estimated by these procedures indicate no heterogeneity of muscarinic receptors in either tissue; thus both are probably of the M2 type. However, the density of the receptors in the ciliary body is lower than in the ileum. Pilocarpine, with lower intrinsic efficacy, demonstrated a pronounced organ selectivity when compared to carbachol as it was a potent full agonist in the ileum and a competitive antagonist in the ciliary body. These results suggest the importance of both receptor density and threshold as determinants of agonist potency.

Contractile response of rabbit dilator pupillae muscle to isotonic high K or norepinephrine, and Ca movement.

Isotonic 80 mM K and norepinephrine contracted the isolated dilator pupillae muscle of the rabbit. Norepinephrine-induced contraction was inhibited by papaverine, but not by Ca blockers, while K-induced contraction was abolished by removing Ca from the bathing fluid but there was a residual response to norepinephrine in Ca-free solution. These results suggest that, in the rabbit dilator pupillae muscle, K-induced contraction is mainly attributable to the increase of Ca influx, while norepinephrine-induced contraction is mainly due to the facilitation of release of intracellularly sequestered Ca.

Alpha 1A-adrenoceptors in rabbit bronchus.

In the presence of propranolol, bronchial preparations from rabbits were markedly contracted by norepinephrine and phenylephrine but only slightly by clonidine. The contractile responses to norepinephrine were inhibited by prazosin and by high concentrations of yohimbine. The concentration-response curve of norepinephrine was shifted in parallel by WB4101 and 5-methylurapidil but not by 60-min treatment with chloroethylclonidine. These results suggest strongly that norepinephrine-induced contraction of rabbit bronchus is mediated through alpha 1A-adrenoceptors. Furthermore, cyclooxygenase inhibitors did not influence the response to norepinephrine, so norepinephrine-induced contraction is considered not to be mediated through the release of prostaglandins.

Alpha 2-adrenoceptor mechanisms in guinea-pig trachea.

In the presence of propranolol tracheal strips from guinea-pigs were markedly contracted by norepinephrine and clonidine but only slightly by phenylephrine. The contractile responses to clonidine and norepinephrine were inhibited by yohimbine but not by prazosin. The specific binding of [3H]p-aminoclonidine to the microsomal fractions from guinea-pig tracheal muscle was much greater than that of [125I]BE2254. These results indicate that alpha 2-adrenoceptors are the predominant subtype of alpha-adrenoceptor in the guinea-pig trachea. The response to norepinephrine was abolished by cyclooxygenase inhibitors, suggesting the release of excitatory prostaglandins by norepinephrine. These results raise the possibility that norepinephrine interacts with alpha 2-adrenoceptors and releases prostaglandins to contract tracheal muscle.

Intrinsic activity and effects of guanyl-5'-yl imidodiphosphate, Gpp(NH)p on the affinity of partial agonists to the muscarinic receptor.

The effects of a GTP analogue, guanyl-5'-yl imidodiphosphate, Gpp(NH)p on the inhibition of [3H]3-quinuclidinyl benzilate (QNB) binding by some partial agonists were studied with a microsomal fraction of longitudinal muscle of guinea-pig ileum and compared with the intrinsic activity of the partial agonists which was estimated in the longitudinal muscle of guinea-pig ileum. There was a good correlation between the inhibitory effects of Gpp(NH)p on the affinity of the partial agonists to muscarinic receptors and their intrinsic activity.

Tetrodotoxin-resistant response to nicotine in rabbit bronchial preparation.

The mode of action of nicotine was studied in a rabbit bronchial preparation. Nicotine (3 X 10(-5)-10(-3) M) produced a phasic contraction. No inhibitory response to nicotine was observed. The contractile response to nicotine was inhibited by hexamethonium, pentolinium and atropine but not by tetrodotoxin. Nicotine increased the efflux of tritium from preparations which had been labelled with [3H]choline. Tetrodotoxin did not inhibit the nicotine-evoked tritium release from the bronchial preparation. The results indicate that the release of acetylcholine evoked by nicotine was not influenced by tetrodotoxin in this preparation, and that the nicotine-induced response may be produced mainly through a sodium action potential-independent process. We could not rule out a contribution by acetylcholine released from the acetylcholine store in the smooth muscle in the contractile mechanisms for nicotine.

A dual action of papaverine on calcium uptake by microsomal fraction isolated from rat uterus.

The effects of papaverine, cyclic AMP and MIX(3-isobutyl-1-methylxanthine, a phosphodiesterase inhibitor) on calcium uptake by a microsomal fraction from rat uterus were tested in the presence of 3 mM ATP. Papaverine potentiated calcium uptake in the presence of oxalate but inhibited it in the absence of oxalate. However, cyclic AMP and MIX did not influence calcium uptake, neither in the presence nor the absence of oxalate. These results suggest that calcium uptake by plasma membrane-derived vesicles in the absence of oxalate is inhibited by papaverine and that papaverine potentiated calcium uptake by the internal membranes in the presence of oxalate. They suggest also that the stimulatory action of papaverine involves a cyclic AMP-independent mechanism in addition to the mechanism via cyclic AMP.

Effects of aging on alpha 2-adrenoceptor mechanisms in isolated guinea-pig tracheal preparations.

The effects of aging on alpha 2-adrenoceptor mechanisms in tracheal preparations isolated from 3-, 6-, 10-, 40- and 100-week-old guinea-pigs were studied. The potency of noradrenaline decreased with age. The specific binding of [3H]p-aminoclonidine to microsomal fractions prepared from tracheal smooth muscles from 6- and 40-week-old guinea-pigs was analysed by Scatchard analysis. The capacity of maximum binding sites of [3H]p-aminoclonidine was greater in the preparations from 40-week-old guinea-pigs than in 6-week-old animals, whereas the dissociation constant did not change with age. The amount of prostaglandin F2 alpha released by noradrenaline and the contractile response to prostaglandin F2 alpha decreased with age. These results suggest that an age-related decrease in the potency of noradrenaline is due to the decrease in the amount of excitable prostaglandin F2 alpha released by noradrenaline and in the contractile response to prostaglandin F2 alpha, but not to a change in the total number of alpha 2-adrenoceptors or in the dissociation constant of a drug from alpha 2-adrenoceptors. Extraneuronal uptake also plays a role in the response to noradrenaline in the tracheal preparations from older guinea-pigs.

Possible mechanisms of beta-adrenoceptor-mediated relaxation induced by noradrenaline in guinea pig taenia caecum.

The mechanisms of the beta-adrenoceptor-mediated relaxation induced by noradrenaline in guinea pig taenia caecum were investigated. Noradrenaline caused graded relaxation of this preparation. However, the concentration-response curves for noradrenaline were unaffected by propranolol (approximately 10(-5) M) or phentolamine (approximately 10(-5) M). The responses to noradrenaline were antagonized in a concentration-dependent manner by bupranolol, and Schild plots of the data revealed a pA2 value of 5.53. Also, bupranolol antagonized responses to isoprenaline, and Schild plots of the data revealed the pA2 value to be 8.53. Noradrenaline significantly increased the cyclic AMP level in this preparation. Bupranolol (10(-4) M) significantly decreased the cyclic AMP response elicited by noradrenaline, whereas propranolol (10(-5) M) produced no effect. These results suggest that the relaxant response to noradrenaline in guinea pig taenia caecum is mainly mediated by beta 3-adrenoceptors (or atypical beta-adrenoceptors) and that in guinea pig taenia caecum noradrenaline behaves as a beta 3-selective adrenoceptor agonist.