Enhancement Of Acetylcholine Release Research Articles

Serotonin 5-HT4 receptors and forebrain cholinergic system: receptor expression in identified cell populations.

Activation of serotonin 5-HT4 receptors has pro-cognitive effects on memory performance. The proposed underlying neurochemical mechanism is the enhancement of acetylcholine release in frontal cortex and hippocampus elicited by 5-HT4 agonists. Although 5-HT4 receptors are present in brain areas related to cognition, e.g., hippocampus and cortex, the cellular localization of the receptors that might modulate acetylcholine release is unknown at present. We have analyzed, using dual label in situ hybridization, the cellular localization of 5-HT4 receptor mRNA in identified neuronal populations of the rat basal forebrain, which is the source of the cholinergic innervation to cortex and hippocampus. 5-HT4 receptor mRNA was visualized with isotopically labeled oligonucleotide probes, whereas cholinergic, glutamatergic, GABAergic and parvalbumin-synthesizing neurons were identified with digoxigenin-labeled oligonucleotide probes. 5-HT4 receptor mRNA was not detected in the basal forebrain cholinergic cell population. In contrast, basal forebrain GABAergic, parvalbumin synthesizing, and glutamatergic cells contained 5-HT4 receptor mRNA. Hippocampal and cortical glutamatergic neurons also express this receptor. These results indicate that 5-HT4 receptors are not synthesized by cholinergic cells, and thus would be absent from cholinergic terminals. In contrast, several non-cholinergic cell populations within the basal forebrain and its target hippocampal and cortical areas express these receptors and are thus likely to mediate the enhancement of acetylcholine release elicited by 5-HT4 agonists.

Natural Compounds Endowed with Cholinergic or Anticholinergic Activity. Enhancement of Acetylcholine Release by a Quaternary Derivative of l-Hyoscyamine

New compounds that target nicotinic receptors (nAChRs) have been sought to correct disorders affecting cholinergic transmission in central and peripheral synapses. A quaternary derivate of l-hyoscyamine, phenthonium (Phen), was shown by our group to enhance the spontaneous acetylcholine (ACh) release without altering the nerve-induced transmitter release at the neuromuscular junction. The effect was unrelated to membrane depolarization, and was not induced by an increase of calcium influx into the nerve terminal. Phen also presented a competitive antimuscarinic activity and blocked noncompetitively the neuromuscular transmission. In this work we re-examined the mechanisms underlying the facilitatory actions of Phen on [(3)H]-ACh release in isolated ganglia of the guinea pig ileal myenteric plexus. Exposure of the preparations to Phen (10-50 microM) increased the release of [(3)H]-ACh by 81 to 68% over the basal. The effect was not affected by the ganglionic nAChR antagonist hexamethonium (1 nM) at a concentration that inhibited the increase of [(3)H]-ACh release induced by the nicotinic agonist dimethylphenylpiperazinium (DMPP, 30 microM). Association of Phen (10 microM) with DMPP potentiated the facilitatory effect of Phen. [(3)H]-ACh release was not altered by the muscarinic antagonists atropine (1 nM) or pirenzepine (1 microM). However, both antagonists inhibited the release of [(3)H]-ACh induced by either the muscarinic M1 agonist McN-343 (10 microM) or Phen (20 microM). The facilitatory effect of Phen was not altered by CdCl(2) (50 mM), but it was potentiated in the presence of tetraethylammonium (40 mM). The results indicate that the facilitatory action of Phen appears to be mediated by an increase of the inwardly rectifying potassium channels conductance probably related to the compound antimuscarinic activity.

Effect of I.C.V. injection of AT 4 receptor ligands, NLE 1-angiotensin IV and LVV-hemorphin 7, on spatial learning in rats

Central administration of angiotensin IV (Ang IV) or its analogues enhance performance of rats in passive avoidance and spatial memory paradigms. The purpose of this study was to examine the effect of a single bolus injection of two distinct AT 4 ligands, Nle 1-Ang IV or LVV-haemorphin-7, on spatial learning in the Barnes circular maze. Mean number of days for rats treated with either Nle 1-Ang IV or LVV-haemorphin-7 to achieve learner criterion is significantly reduced compared with controls ( P<0.001 and P<0.05 respectively). This is due to enhanced ability of the peptide-treated rats to adopt a spatial strategy for finding the escape hatch. In all three measures of learning performance, (1) the number of errors made, (2) the distance travelled and (3) the latency in finding the escape hatch, rats treated with either 100 pmol or 1 nmol of Nle 1-Ang IV or 100 pmol LVV-haemorphin-7 performed significantly better than the control groups. As early as the first day of testing, the rats treated with the lower dose of Nle 1-Ang IV or LVV-haemorphin-7 made fewer errors ( P<0.01 and P<0.05 respectively) and travelled shorter distances ( P<0.05 for both groups) than the control animals. The enhanced spatial learning induced by Nle 1-Ang IV (100 pmol) was attenuated by the co-administration of the AT 4 receptor antagonist, divalinal-Ang IV (10 nmol). Thus, administration of AT 4 ligands results in an immediate potentiation of learning, which may be associated with facilitation of synaptic transmission and/or enhancement of acetylcholine release.

Sigma receptor binding profile of a series of analgesic tropane derivatives.

The binding profile on sigma receptor subtypes of a family of tropane derivatives, previously studied as analgesic and nootropic drugs, is reported. The results show that the sigma receptors are not involved in the enhancement of acetylcholine release and of the consequent analgesic activity of this class of drugs. Structure-affinity relationships for sigma receptor subtypes are discussed. They lead to the identification of a few useful leads for further development of potent sigma ligands within the series.

Nicotinic receptors modulating ACh release in rat cortical synaptosomes: role of Ca 2+ ions in their function and desensitization

Cholinergic nerve terminals in the central nervous system are endowed with both muscarinic and nicotinic autoreceptors, mediating inhibition, and enhancement of acetylcholine release, respectively. Exogenous acetylcholine inhibited the K +(15 mM)-evoked overflow of [ 3H]acetylcholine from superfused rat neocortical synaptosomes; however, in the presence of atropine, this muscarinic inhibition was reversed into a nicotinic potentiation when acetylcholine was added concomitantly with high-K +, but not before depolarization. Increasing concentrations of acetylcholine (plus atropine), nicotine and (+)-anatoxin-a produced elevations of the K +-evoked [ 3H]acetylcholine overflow resulting in bell-shaped concentration-response curves. Synaptosomes pretreated with different concentrations (10 μM to 0.001 μM) of acetylcholine or nicotine responded to a subsequent nicotinic stimulus (10 μM acetylcholine plus 0.1 μM atropine, in 15 mM K +) in a manner reflecting varying degrees of desensitization. This desensitization could be reversed by washings with standard medium and desensitization was attenuated when external Ca 2+ ([Ca 2+] e) was decreased. Lowering of [Ca 2+] e or chelation of internal Ca 2+ with 1,2-bis(2-aminophenoxy)ethone- N, N, N′, N′-tetracetic acid acetoxymethylester (BAPTA-AM) permitted the nicotinic response to acetylcholine alone (no atropine added) to prevail over the muscarinic response. Pretreatment with BAPTA-AM could however not prevent desensitization by acetylcholine (10 or 0.001 μM). The data indicate that Ca 2+ ions are involved in determining the balance between muscarinic and nicotinic autoreceptor function and in the desensitization of nicotinic autoreceptors.

Identification of SK-951, a Novel Benzofuran Derivative, as an Agonist to 5-HT4 Receptors

The pharmacological profile of SK-951 ((—)4-amino-N-[2-(1-azabicyclo[3.3.0]octan-5-yl) ethyl]-5-chloro-2,3-dihydro-2-methylbenzo[b]furan-7-carboxamide hemifumarate) was identified in relation to serotonin 5-HT3 and 5-HT4 receptors by the receptor binding assay and functional studies. The receptor binding assay showed that SK-951 bound to the 5-HT3 receptor with a high affinity, to the 5-HT4 receptor with relatively higher affinity and to the muscarinic M2 receptor with a low affinity, but not to dopamine D1 and D2 and serotonin 5-HT1 and 5-HT2 and muscarinic M1 and M3 receptors. SK-951 caused relaxations of tunica muscularis mucosae preparations from rat esophagus which were precontracted with carbachol, and the effects were antagonized by GR113808, a selective 5-HT4 antagonist. In the longitudinal muscle with myenteric plexus (LMMP) preparations from guinea pig ileum, SK-951 enhanced the electrically-stimulated contraction of preparations in which the 5-HT1, 5-HT2 and 5-HT3 receptors were blocked, and it enhanced the electrically-stimulated release of [3H]acetylcholine (ACh). These effects of SK-$51 were antagonized by GR113808. SK-951 inhibited the 5-HT3 receptor-mediated contractions. These results indicate that SK-951 possesses properties of an agonist for the 5-HT4 receptor and an antagonist for the 5-HT3 receptor. Thus, SK-951 is a new and potent 5-HT4-receptor agonist and causes contractions of guinea pig ileum mediated by enhancement of ACh release via the 5-HT4 receptor.

Gangliosides and sialylcholesterol as modulators of synaptic functions.

Gangliosides were shown to enhance the release of acetylcholine from synaptosomes on stimulation. The influx of calcium ion into synaptosomes on membrane depolarization was increased by gangliosides. This was hypothesized to be an underlying mechanisms for the enhancement of acetylcholine release. Studies using calcium channel blockers revealed that four distinct types of voltage-dependent calcium channels occurred in cerebrocortical synapses, and that the N-type was primarily responsible for the evoked release of acetylcholine. An additional result suggests that gangliosides may act mainly on the N-type calcium channel. Cholinergic-specific gangliosides, Chol-1 alpha, were assumed to participate in the mechanism of high-affinity choline uptake. These two different actions of gangliosides were found to be mimicked by synthetic ganglioside analogs. Calcium influx was increased by alpha-sialylcholesterol, and choline uptake was accelerated by beta-sialylcholesterol. Gangliosides and sialylcholesterol having these apparently beneficial effects were shown to ameliorate decreased functions of synapses from aged brains.

Control of the calcium concentration involved in acetylcholine release and its facilitation: an additional role for synaptic vesicles?

2,5-Diterbutyl-1,4-benzohydroquinone, a specific blocker of Ca 2+-ATPase pumps, increased acetylcholine release from an identified synapse of Aplysia, as well as from Torpedo and mouse caudate nucleus synaptosomes. Because 2,5-diterbutyl-1,4-benzohydroquinone does not change the presynaptic Ca 2+ influx, the enhancement of acetylcholine release could be due to an accumulation of Ca 2+ in the terminal. This possibility was further checked by studying the effects of 2,5-diterbutyl-1,4-benzohydroquinone on twin pulse facilitation, classically attributed to residual Ca 2+. While preventing the fast sequestration of Ca 2+ by presynaptic organelles, 2,5-diterbutyl-1,4-benzohydroquinone magnified both twin pulse facilitation observed under low extracellular Ca 2+ concentration and twin pulse dysfacilitation observed under high extracellular Ca 2+ concentration. Thus, it is concluded that 2,5-diterbutyl-1,4-benzohydroquinone, by preventing Ca 2+ buffering near transmitter release sites, modulates acetylcholine release. As 2,5-diterbutyl-1,4-benzohydroquinone was also shown to decrease by 50% the uptake of 45Ca 2+ by isolated synaptic vesicles, we propose that synaptic vesicles can control the presynaptic Ca 2+ concentration triggering the release of neurotransmitter.

Evidence for 5-HT 4 receptor involvement in the enhancement of acetylcholine release by p-chloroamphetamine in rat frontal cortex

The roles of endogenous serotonin (5-HT) and 5-HT receptor subtypes in regulation of acetylcholine (ACh) release in frontal cortex of conscious rats were examined using a microdialysis technique. Systemic administration (1 and 3 mg/kg, i.p.) of the 5-HT-releasing agent p-chloroamphetamine (PCA) elevated ACh output in a dose-dependent manner. Depletion of endogenous 5-HT by p-chlorophenylalanine significantly attenuated the facilitatory effect of PCA on ACh release. The PCA (3 mg/kg)-induced increase in ACh release was significantly inhibited by local application of the 5-HT 4 receptor antagonists RS23597 (50 μM) and GR113803 (1 μM), while the 5-HT 1A antagonist WAY-100135 (10 mg/kg, i.p.; 100 μM), 5-HT 1A/1B/β-adrenoceptor antagonists (−)-pindolol (8 mg/kg, i.p.) and (−)-propranolol (150 μM), 5-HT 2A/2C antagonist ritanserin (1 mg/kg, i.p.; 10 μM) and 5-HT 3 antagonist ondansetron (1 mg/kg, i.p.; 10 μM) failed to significantly modify the effect of PCA. These results suggest that PCA-induced enhancement of 5-HT transmission facilitates ACh release from rat frontal cortex at least in part through 5-HT 4 receptors.

Oral administration of KW-5092, a novel gastroprokinetic agent with acetylcholinesterase inhibitory and acetylcholine release enhancing activities, causes a dose-dependent increase in the blood acetylcholine content of beagle dogs

Acetylcholine (ACh) was detected in the blood and plasma of beagle dogs using a specific, sensitive radioimmunoassay. The mean basal ACh contents in the blood and plasma of beagle dogs were 451±65 and 83.5±12.3 pg/ml (±SEM, n=7), respectively, and were lower than the contents in humans reported previously by our laboratory. Oral administration of KW-5092 (10–30 mg/kg), a gastroprokinetic agent with acetylcholinesterase (AChE) inhibitory and ACh release enhancing activities, caused a dose-dependent increase in the ACh content of both the blood and plasma, as well as several behavioral side effects due to peripheral cholinergic stimulation. The size of the increase in the plasma ACh content at each dose of KW-5092 was greater than that in the blood, indicating that KW-5092 caused the increase in the blood ACh content through elevation of the plasma ACh content, by inhibition of AChE and facilitation of ACh release. These results demonstrate that the blood ACh of beagle dogs is present mainly in the blood cells and to a lesser degree in the plasma, and that KW-5092 increased the blood ACh content mainly by increasing the plasma ACh concentration.

A novel prolyl endopeptidase inhibitor, JTP-4819, with potential for treating Alzheimer's disease

The pharmacological actions of JTP-4819, a new prolyl endopeptidase (PEP) inhibitor targeted for the treatment of Alzheimer's disease, are reviewed with respect to its effects on PEP activity, brain neurotransmitters, and memory-related behaviour in rats. JTP-4819 was shown to be a very potent and specific inhibitor of PEP. At nanomolar concentration, JTP-4819 inhibited the degradation of substance P, arginine-vasopressin, and thyrotropin-releasing hormone by PEP in supernatants of the rat cerebral cortex and hippocampus. Repeated administration of JTP-4819 reversed the aging-induced decrease in brain substance P-like and thyrotropin-releasing hormone-like immunoreactivity, suggesting that this drug may be able to improve the imbalance of peptidergic neuronal systems that develops with senescense by inhibiting PEP activity. JTP-4819 increased acetylcholine release from the frontal cortex and hippocampus, regions closely associated with memory, in both young and aged rats. In addition, it improved performance in several memory and learning-related tests (e.g., the Morris water maze task in aged or MCA-occluded rats and the passive avoidance test). This memory-enhancing effect of JTP-4819 may result from prevention of the metabolic degradation of brain neuropeptides by PEP as well as from the enhancement of acetylcholine release. Taken together, these unique and potent pharmacological actions of JTP-4819 suggest that it may have the potential to be used for treating Alzheimer's disease.

Enhancement of acetylcholine release by homoanatoxin-a from Oscillatoria formosa

The strain NIVA-CYA 92 of Oscillatoria formosa Bory ex Gormont produces phycotoxins with neurotoxic properties. Chemical analysis by gas chromatography/mass spectrometry of a water extract of lyophilized material of the organism showed the presence of only homoanatoxin-a. The mechanism of action of homoanatoxin-a on peripheral cholinergic nerves is so far not known. The neurotoxicity of O. formosa containing homoanatoxin-a was investigated in rat bronchi, rat brain synaptosomes and in GH 4C 1 cells. The water extract of lyophilized material of the organism produced a concentration-dependent reversible increase in the release of [ 3H]acetylcholine from both K + (51 mM) depolarised and non-depolarised cholinergic nerves of the rat bronchial smooth muscle. The K +-evoked release of [ 3H]acetylcholine was enhanced by about 75% by a water extract from 15–20 mg/ml of lyophilized algal material. The enhanced release of [ 3H]acetylcholine was substantially reduced by the L-type Ca 2+-channel blocker verapamil (100 μM) and not by the N-type Ca 2+-channel blocker ω-conotoxin GVIA (1.0 μM) or the P-type Ca 2+-channel blocker ω-agatoxin IV-A (0.2 μM). Chelation of intra-cellular Ca 2+ by 1,2-bis-(aminofenoxi)etan- N,N,N′,N′-tetraacidic acid/acetoxymethyl (BAPTA/AM) (30 μM) had no effect on the phycotoxin-induced release of [ 3H]acetylcholine, indicating that an extracellular pool of Ca 2+ was important for the action of the phycotoxin on the release of [ 3H]acetylcholine from peripheral cholinergic nerves. In rat brain synaptosomes the algal extract enhanced the influx of 45Ca 2+ in a tetrodotoxin (1.0 μM) and ω-conotoxin MVIIC (blocker of N-, P- and Q-type Ca 2+ channels) (1.0 μM) insensitive manner. Patch-clamp studies showed that the phycotoxin opened endogenous voltage dependent L-type Ca 2+ channels in neuronal GH 4C 1 cells. These Ca 2+ channels and the effect of the toxin on the channels were blocked by the L-type Ca 2+-channel antagonist gallopamil (200 μM). The present results suggest, therefore, that the investigated strain of O. formosa contains homoanatoxin-a, which enhances the release of acetylcholine from peripheral cholinergic nerves through opening of endogenous voltage dependent neuronal L-type Ca 2− channels.

P-19-28 - Enhancement of acetylcholine release by SA4503, a novel σ1 agonist, in rat brain

P-19-28 - Enhancement of acetylcholine release by SA4503, a novel σ1 agonist, in rat brain

Enhancement of acetylcholine release by flumazenil in the hippocampus of rats chronically treated with diazepam but not with imidazenil or abecarnil.

The effects of long-term treatment (three times a day for 3 weeks) with pharmacologically active doses of the novel anxiolytics and anticovulsants abecarnil (0.5 mg/kg, IP) and imidazenil (0.5 mg/kg, IP) on basal hippocampal acetylcholine release in freely moving rats were compared with those of diazepam (3 mg/kg, IP). Challenge doses of diazepam, abecarnil, and imidazenil decreased the extracellular acetyl-choline concentration in the hippocampus by the same extent in animals chronically treated with the respective drug or vehicle. Moreover, the abrupt discontinuation of long-term treatment with diazepam, abecarnil, or imidazenil failed to affect hippocampal acetylcholine release during the first 5 days of withdrawal. In contrast, the acute administration of the benzodiazepine receptor antagonist flumazenil (1 mg/kg, IP) 2 days after diazepam withdrawal elicited a marked increase (65%) in acetylcholine release in the hippocampus. Flumazenil failed to induce the same effect 5 days after diazepam withdrawal or 2 or 5 days after discontinuation of long-term treatment with abecarnil or imidazenil. These results indicate that (i) the inhibitory effects of full (diazepam), partial (imidazenil), and selective (abecarnil) benzodiazepine receptor agonists on acetylcholine output in rat hippocampus are not affected by repeated drug administration; (ii) discontinuation of long-term treatment with each type of agonist does not affect hippocampal cholinergic mechanisms; and (iii) flumazenil increases acetylcholine release only in the hippocampus of rats chronically treated with diazepam. Together, these data further differentiate the pharmacology of benzodiazepine receptor full agonists from that of partial and selective agonists.

Adenosine A 2A receptors stimulate acetylcholine release from nerve terminals of the rat hippocampus

The nature of the adenosine receptors involved in the enhancement of acetylcholine release in the hippocampus was studied. The A 2A agonist, CGS 21680, increased the veratridine-evoked release of [ 3H]acetylcholine from hippocampal synaptosomes. This presynaptic effect of CGS 21680 was greater at 3–30 nM than at 100 nM. The excitatory effect of CGS 21680 was antagonised by the A 2 antagonist, DMPX (10μM), and by the A 2A antagonist, CSC (200 nM), but not by the A 1 antagonist, DPCPX (20 nM). We also found co-expression of A 2A and choline acetyltransferase mRNAs in the nucleus of the diagonal band and the medial septum, where the cholinergic cell bodies that project into the hippocampus are located. These results indicate that A 2A adenosine receptors are present in cholinergic nerve terminals in the hippocampus and that activation of these receptors enhances acetylcholine release.

Nitric oxide modulates the release of acetylcholine in the ventral striatum of the freely moving rat.

The influence of nitric oxide on acetylcholine release in the ventral striatum was investigated by the push-pull superfusion technique in the conscious, freely moving rat. Superfusion with the nitric oxide donors S-nitroso-N-acetylpenicillamine or with 3-morpholino-sydnonimine caused a pronounced increase in striatal acetylcholine release. This effect was prevented by superfusion with tetrodotoxin. Pre-superfusion with the guanylyl cyclase inhibitor methylene blue abolished the effect of 3-morpholino-sydnonimine. Superfusion of the ventral striatum with the guanylyl cyclase inhibitor LY83583 decreased acetylcholine release by 60% of basal release, whereas the less specific guanylyl cyclase inhibitor methylene blue was ineffective in this respect. Superfusion of the ventral striatum with inhibitors of nitric oxide synthase also led to different effects on basal acetylcholine release. Superfusion with L-NG-methylarginine did not influence basal acetylcholine release, whereas superfusion with L-NG-nitroarginine or with L-NG-nitroarginine methyl ester led to a substantial decrease in acetylcholine output, the latter compound being more effective. The effect of L-NG-nitroarginine was abolished by simultaneous superfusion with L-arginine. The effects of NO donors and of LY83583 suggest that NO increases acetylcholine release, probably by a cGMP-dependent mechanism. The effectiveness of nitric oxide synthase inhibitors shows that the activity of striatal neurons is under the permanent influence of nitric oxide, that leads, via a direct or indirect mechanism, to continuous enhancement of acetylcholine release. In conclusion, our findings suggest that NO synthesized in the ventral striatum acts as an intracellular messenger which modulates acetylcholine release.

Baclofen enhancement of acetylcholine release from amacrine cells in the rabbit retina by reduction of glycinergic inhibition.

1. The mechanism by which the GABAB-receptor agonist, baclofen, enhances the light-evoked release of [3H]acetylcholine (ACh) from cholinergic amacrine cells was studied using an eye-cup preparation in anaesthetized rabbits and isolated retinas. 2. When applied locally to the rabbit retina, baclofen increased the release of ACh evoked by a flickering light (3 Hz) by over 40%. 3. In isolated retinas, baclofen strikingly inhibited the K(+)-evoked release of glycine but had no effect on GABA release. 4. In the rabbit eye cup, strychnine enhanced the light-evoked release of ACh to a similar degree to that produced by baclofen. The effects of baclofen and strychnine on the light-evoked release of ACh were not additive. In contrast, bicuculline did not affect the enhancing action of baclofen on the light-evoked release of ACh. 5. In order to see whether the glycinergic amacrine cells might be stimulated by ACh, isolated rat and rabbit retinas were exposed to muscarine. This cholinergic agonist potentiated the K(+)-evoked release of glycine by 54%. 6. We suggest that baclofen enhances the light-evoked release of ACh from amacrine cells by inhibiting glycine release from glycinergic amacrine cells which are stimulated by ACh and form an inhibitory feedback loop to the cholinergic neurones.

Time course of decline of radiolabeled acetylcholine formed following intracerebroventricular administration of tritiated choline: Effects of oxotremorine and scopolamine

Rats were injected intracerebroventricularly with 5 microCi of [methyl-3H]choline. The time course of decline of the radiolabeled acetylcholine (ACh) formed was estimated in the ipsilateral cerebral cortex and striatum. The [3H]ACh levels declined biphasically from the cerebral tissue. The initial decline proceeded rapidly, after which labeled ACh declined more slowly. Scopolamine (1 mg/kg, i.v.) caused a significant increase in the rate of [3H]ACh disappearance, which can be interpreted as an enhancement of ACh release. By contrast, oxotremorine (0.8 mg/kg, i.v.) markedly reduced the [3H]ACh disappearance. The results show that drug-induced changes in cholinergic neuronal activities can be estimated from the disappearance of radioactive ACh after labeling the endogenous transmitter through intracerebroventricular administration of labeled choline.

Neural and myogenic effects of leukotrienes C4, D4, and E4 on canine bronchial smooth muscle

The leukotrienes (LTs), referred to as the slow-reacting substance of anaphylaxis (SRS-A), are reported to have little or no activity in the canine airway. The objective of this study was to determine whether LTC4, LTD4, and LTE4 (10(-10)-10(-7) M) play a role in neuromuscular control of third- to fifth-order canine bronchi. In the presence of 1 microM indomethacin (Indo), canine bronchial smooth muscle contracted and was depolarized in a concentration-dependent manner by LTC4 or LTD4 but not by LTE4. LTC4 and LTD4 concentration-response curves were not significantly affected when conducted in the presence of any of the following: 10(-7) M propranolol (beta-adrenoceptor antagonist), 10(-6) M chlorpheniramine (H1-receptor antagonist), 10(-6) M ketanserin (nonselective 5-hydroxytryptamine receptor antagonist), 10(-7) M atropine (muscarinic receptor antagonist), and 10(-6) M tetrodotoxin (sodium channel blocker). LTC4 and LTD4 also potentiated electrical field-stimulated (EFS) excitatory junction potentials (EJPs), suggesting a possible prejunctional enhancement of acetylcholine release. In the absence of Indo, no postjunctional responses to LTC4 and LTD4 occurred. Endogenous prostaglandin E2 (PGE2) and 6-keto-PGF1 alpha (a stable metabolite of PGI2) levels from canine bronchi were significantly reduced by Indo. In the presence of Indo, addition of > or = 10(-8) M of PGE2 suppressed contractions to LTC4 and LTD4. These data suggest that the decrease in PGE2 and PGI2 production by Indo is sufficient to unmask the excitatory postjunctional actions of LTC4 and LTD4 on bronchial smooth muscle. Serine borate (45 mM; an inhibitor of gamma-glutamyl transpeptidase, which prevents the conversion of LTC4 to LTD4) increased selectively the contractile activity of LTC4. L-Cysteine (3 mM; an inhibitor of an aminopeptidase, which prevents the conversion of LTD4 to LTE4) enhanced the contractile responses to LTD4. Serine borate increased the amplitude and duration of EFS contractions and potentiated the amplitude of EFS EJPs; the last effects were prevented by nordihydroguaiaretic acid. These and other studies suggest that LTs are synthesized by canine bronchi and have receptors on canine bronchial smooth muscle but that contractions to LTC4 and LTD4 in the canine airway are usually not observed because of the presence of inhibitory prostanoids (PGE2 and PGI2). We suggest that decreases in PGE2 and PGI2 in models of airway disease in combination with increases in LTC4, LTD4, and thromboxane A2 may contribute to airway hyperresponsiveness in vitro.

Enhancement of acetylcholine release in the insular gustatory cortex of rats in response to aversive taste stimulation

Enhancement of acetylcholine release in the insular gustatory cortex of rats in response to aversive taste stimulation