Alter Acetylcholine Release Research Articles

Benzodiazepine site agonists differentially alter acetylcholine release in rat amygdala.

Agonist binding at the benzodiazepine site of γ-aminobutric acid type A receptors diminishes anxiety and insomnia by actions in the amygdala. The neurochemical effects of benzodiazepine site agonists remain incompletely understood. Cholinergic neurotransmission modulates amygdala function, and this study tested the hypothesis that benzodiazepine site agonists alter acetylcholine (ACh) release in the amygdala. Microdialysis and high-performance liquid chromatography quantified ACh release in the amygdala of Sprague-Dawley rats (n = 33). ACh was measured before and after IV administration (3 mg/kg) of midazolam or eszopiclone, with and without anesthesia. ACh in isoflurane-anesthetized rats during dialysis with Ringer's solution (control) was compared with ACh release during dialysis with Ringer's solution containing (100 μM) midazolam, diazepam, eszopiclone, or zolpidem. In unanesthetized rats, ACh in the amygdala was decreased by IV midazolam (-51.1%; P = 0.0029; 95% confidence interval [CI], -73.0% to -29.2%) and eszopiclone (-39.6%; P = 0.0222; 95% CI, -69.8% to -9.3%). In anesthetized rats, ACh in the amygdala was decreased by IV administration of midazolam (-46.2%; P = 0.0041; 95% CI, -67.9% to -24.5%) and eszopiclone (-34.0%; P = 0.0009; 95% CI, -44.7% to -23.3%), and increased by amygdala delivery of diazepam (43.2%; P = 0.0434; 95% CI, 2.1% to 84.3%) and eszopiclone (222.2%; P = 0.0159; 95% CI, 68.5% to 375.8%). ACh release in the amygdala was decreased by IV delivery of midazolam and eszopiclone. Dialysis delivery directly into the amygdala caused either increased (eszopiclone and diazepam) or likely no significant change (midazolam and zolpidem) in ACh release. These contrasting effects of delivery route on ACh release support the interpretation that systemically administered midazolam and eszopiclone decrease ACh release in the amygdala by acting on neuronal systems outside the amygdala.

Effects of memantine and donepezil on cortical and hippocampal acetylcholine levels and object recognition memory in rats

This preclinical study investigated the ability of memantine (MEM) to stimulate brain acetylcholine (ACh) release, potentially acting synergistically with donepezil (DON, an acetylcholinesterase inhibitor). Acute systemic administration of either MEM or DON to anesthetized rats caused dose-dependent increases of ACh levels in neocortex and hippocampus, and the combination of MEM (5 mg/kg) and DON (0.5 mg/kg) produced significantly greater increases than either drug alone. To determine whether ACh release correlated with cognitive improvement, rats with partial fimbria-fornix (FF) lesions were treated with acute or chronic MEM or DON. Acute MEM treatment significantly elevated baseline hippocampal ACh release but did not significantly improve task performance on a delayed non-match-to-sample (DNMS) task, whereas chronic MEM treatment significantly improved DNMS performance but only marginally elevated baseline ACh levels. Acute or chronic treatment with DON (in the presence of neostigmine to allow ACh collection) did not significantly improve DNMS performance or alter ACh release. In order to investigate the effect of adding MEM to ongoing DON therapy, lesioned rats pretreated with DON for 3 weeks were given a single intraperitoneal dose of MEM. MEM significantly elevated baseline hippocampal ACh levels, but did not significantly improve DNMS task scores compared to chronic DON-treated animals. These data indicate that MEM, in addition to acting as an NMDA receptor antagonist, can also augment ACh release; however, in this preclinical model, increased ACh levels did not directly correlate with improved cognitive performance.

Benzodiazepine receptor agonists cause drug-specific and state-specific alterations in EEG power and acetylcholine release in rat pontine reticular formation.

Benzodiazepine (BDZ) and non-benzodiazepine (NBDZ) hypnotics enhance GABAergic transmission and are widely used for the treatment of insomnia. In the pontine reticular formation (PRF), GABA inhibits rapid eye movement (REM) sleep and acetylcholine (ACh) release. No previous studies have characterized the effects of BDZ and NBDZ hypnotics on ACh release in the PRF. This study tested 2 hypotheses: (1) that microdialysis delivery of zolpidem, eszopiclone, and diazepam to rat PRF alters ACh release in PRF and electroencephalographic (EEG) delta power and (2) that intravenous (i.v.) administration of eszopiclone to non-anesthetized rat alters ACh release in the PRF, sleep, and EEG delta power. A within- and between-groups experimental design. University of Michigan. Adult male Crl:CD*(SD) (Sprague-Dawley) rats (n = 57). In vivo microdialysis of the PRF in rats anesthetized with isoflurane was used to derive the concentration-response effects of zolpidem, eszopiclone, and diazepam on ACh release. Chronically instrumented rats were used to quantify the effects of eszopiclone (3 mg/kg, i.v.) on ACh release in the PRF, sleep-wake states, and cortical EEG power. ACh release was significantly increased by microdialysis delivery to the PRF of zolpidem and eszopiclone but not diazepam. EEG delta power was increased by zolpidem and diazepam but not by eszopiclone administered to the PRF. Eszopiclone (i.v.) decreased ACh release in the PRF of both anesthetized and non-anesthetized rats. Eszopiclone (i.v.) prevented REM sleep and increased EEG delta power. The concentration-response data provide the first functional evidence that multiple GABA(A) receptor subtypes are present in rat PRF. Intravenously administered eszopiclone prevented REM sleep, decreased ACh release in the PRF, and increased EEG delta power. The effects of eszopiclone are consistent with evidence that ACh release in the PRF is lower during NREM sleep than during REM sleep, and with data showing that cholinergic stimulation of the PRF activates the cortical EEG.

Microdialysis Delivery of Morphine to the Hypoglossal Nucleus of Wistar Rat Increases Hypoglossal Acetylcholine Release

The medullary hypoglossal nucleus (XII) innervates the genioglossal muscles of the tongue, and opioid-induced alterations in tongue muscle tone contribute to airway obstruction. Previous studies have shown that morphine causes a significant decrease in acetylcholine (ACh) release in some brain regions, but the effects of morphine on ACh release in XII have not been quantified. A within-subjects design was used to test the hypothesis that morphine alters ACh release in XII of anesthetized Wistar rat. ACh release during microdialysis with Ringer's solution (control) was compared to ACh release during dialysis delivery of opioids. University of Michigan. N/A. Microdialysis delivery of opioids to XII. Morphine caused a statistically significant, concentration-dependent increase in XII ACh release. The increase in XII ACh release caused by 10 microM morphine was blocked by the mu opioid antagonist naloxone and not blocked by the kappa opioid antagonist norbinaltorphimine. The data comprise the first direct measures of ACh release in XII and support the conclusion that morphine depresses hypoglossal nerve activity, in part, by increasing ACh release in XII. Activation of mu opioid receptors on inhibitory neurons within XII likely disinhibits cholinergic terminals, causing increased ACh release. The results are consistent with previous studies showing that blocking the enzymatic degradation of ACh in XII significantly inhibited tongue muscle activity.

Morphine-Induced Acetylcholine Release at the Hypoglossal Motor Nucleus: Implications for Opiate-Induced Respiratory Suppression

Study Objectives: The medullary hypoglossal nucleus (XII) innervates the genioglossal muscles of the tongue, and opioid-induced alterations in tongue muscle tone contribute to airway obstruction. Previous studies have shown that morphine causes a significant decrease in acetylcholine (ACh) release in some brain regions, but the effects of morphine on ACh release in XII have not been quantified. Design: A within-subjects design was used to test the hypothesis that morphine alters ACh release in XII of anesthetized Wistar rat. ACh release during microdialysis with Ringer's solution (control) was compared to ACh release during dialysis delivery of opioids. Setting: University of Michigan Patients or Participants: N/A Interventions: Microdialysis delivery of opioids to XII. Measurements and Results: Morphine caused a statistically significant, concentration-dependent increase in XII ACh release. The increase in XII ACh release caused by 10 μM morphine was blocked by the mu opioid antagonist naloxone and not blocked by the kappa opioid antagonist norbinaltorphimine. Conclusions: The data comprise the first direct measures of ACh release in XII and support the conclusion that morphine depresses hypoglossal nerve activity, in part, by increasing ACh release in XII. Activation of mu opioid receptors on inhibitory neurons within XII likely disinhibits cholinergic terminals, causing increased ACh release. The results are consistent with previous studies showing that blocking the enzymatic degradation of ACh in XII significantly inhibited tongue muscle activity.

The TRPM7 Ion Channel Functions in Cholinergic Synaptic Vesicles and Affects Transmitter Release

A longstanding hypothesis is that ion channels are present in the membranes of synaptic vesicles and might affect neurotransmitter release. Here we demonstrate that TRPM7, a member of the transient receptor potential (TRP) ion channel family, resides in the membrane of synaptic vesicles of sympathetic neurons, forms molecular complexes with the synaptic vesicle proteins synapsin I and synaptotagmin I, and directly interacts with synaptic vesicular snapin. In sympathetic neurons, changes in TRPM7 levels and channel activity alter acetylcholine release, as measured by EPSP amplitudes and decay times in postsynaptic neurons. TRPM7 affects EPSP quantal size, an intrinsic property of synaptic vesicle release. Targeted peptide interference of TRPM7's interaction with snapin affects the amplitudes and kinetics of postsynaptic EPSPs. Thus, vesicular TRPM7 channel activity is critical to neurotransmitter release in sympathetic neurons.

Activation of 5-HT2 receptors enhances the release of acetylcholine in the prefrontal cortex and hippocampus of the rat.

The role of 5-HT2 receptors in the regulation of acetylcholine (ACh) release was examined in the medial prefrontal cortex and dorsal hippocampus using in vivo microdialysis. The 5-HT(2A/2C) agonist +/-1-(2,5-dimethoxy-4-iodophenyl) -2- aminopropane hydrochloride (DOI) (1 and 2 mg/kg, i.p.) significantly increased the extracellular concentration of ACh in both brain regions, and this response was attenuated in rats treated with the 5-HT(2A/2B/2C) antagonist LY-53,857 (3 mg/kg, i.p.). Treatment with LY-53,857 alone did not significantly alter ACh release in either brain region The 5-HT(2C) agonist 6-chloro-2-(1-piperazinyl)-pyrazine) (MK-212) (5 mg/kg, i.p.) significantly enhanced the release of ACh in both the prefrontal cortex and hippocampus, whereas the 5-HT2 agonist mescaline (10 mg/kg, i.p.) produced a 2-fold increase in ACh release only in the prefrontal cortex. Intracortical, but not intrahippocampal, infusion of DOI (100 microM) significantly enhanced the release of ACh, and intracortical infusion of LY-53,857 (100 microM) significantly attenuated this response. These results suggest that the release of ACh in the prefrontal cortex and hippocampus is influenced by 5-HT2 receptor mechanisms. The increase in release of ACh induced by DOI in the prefrontal cortex, but not in the hippocampus, appears to be due to 5-HT2 receptor mechanisms localized within this brain region. Furthermore, it appears that the prefrontal cortex is more sensitive than the dorsal hippocampus to the stimulatory effect of 5-HT2 agonists on ACh release.

Carbachol in the pontine reticular formation of C57BL/6J mouse decreases acetylcholine release in prefrontal cortex

The prefrontal cortex and brainstem modulate autonomic and arousal state control but the neurotransmitter mechanisms underlying communication between prefrontal cortex and brainstem remain poorly understood. This study examined the hypothesis that microdialysis delivery of carbachol to the pontine reticular formation (PRF) of anesthetized C57BL/6J (B6) mouse modulates acetylcholine (ACh) release in the frontal association cortex. Microdialysis delivery of carbachol (8.8 mM) to the PRF caused a significant ( P<0.01) decrease (−28%) in ACh release in the frontal association cortex, a significant ( P<0.01) decrease (−23%) in respiratory rate, and a significant ( P<0.01) increase (223%) in time to righting after anesthesia. Additional in vitro studies used the [ 35S]guanylyl-5′- O-(γ-thio)-triphosphate ([ 35S]GTPγS) assay to test the hypothesis that muscarinic cholinergic receptors activate guanine nucleotide binding proteins (G proteins) in the frontal association cortex and basal forebrain. In vitro treatment with carbachol (1 mM) caused a significant ( P<0.01) increase in [ 35S]GTPγS binding in the frontal association cortex (62%) and basal forebrain nuclei including medial septum (227%), vertical (210%) and horizontal (165%) limbs of the diagonal band of Broca, and substantia innominata (127%). G protein activation by carbachol was concentration-dependent and blocked by atropine, indicating that the carbachol-stimulated [ 35S]GTPγS binding was mediated by muscarinic cholinergic receptors. Together, the in vitro and in vivo data show for the first time in B6 mouse that cholinergic neurotransmission in the PRF can significantly alter ACh release in frontal association cortex, arousal from anesthesia, and respiratory rate.

Ketamine and MK-801 Decrease Acetylcholine Release in the Pontine Reticular Formation, Slow Breathing, and Disrupt Sleep

Ketamine induces a dissociated state of consciousness by binding to the phencyclidine binding site within the ion channel gated by the N-methyl-D-aspartate (NMDA) receptor. The brain regions and neurotransmitter systems mediating ketamine-induced alterations in arousal remain incompletely understood. This study used in vivo microdialysis to test the hypothesis that ketamine alters acetylcholine (ACh) release in the medial pontine reticular formation (mPRF). Acetylcholine (ACh) release, sleep, and breathing were quantified following systemic ketamine administration. Microdialysis was used to deliver the NMDA-channel blocker dizocilpine maleate (MK-801) and the R(-)-isomer of ketamine into the mPRF while measuring ACh release. N/A. N/A. N/A. Systemically administered ketamine disrupted normal sleep-cycle organization, reduced mPRF ACh release, and significantly slowed rate of breathing. Dialysis delivery of MK-801 to the mPRF significantly decreased respiratory rate and mPRF ACh release. Dialysis delivery to the mPRF of the R(-)-ketamine isomer significantly decreased mPRF ACh release. Decreased mPRF ACh release caused by systemically administered ketamine was mimicked by mPRF dialysis delivery of MK-801 and the R(-)-ketamine isomer. These data are consistent with the conclusion that systemically administered ketamine may alter arousal and breathing, in part, by altering cholinergic neurotransmission in the mPRF.

Age-related Effects of Chlorpyrifos on Acetylcholine Release in Rat Brain

Chlorpyrifos (CPF) is an organophosphorus insecticide that elicits toxicity through inhibition of acetylcholinesterase (AChE). Young animals are markedly more sensitive than adults to the acute toxicity of CPF. We evaluated acetylcholine (ACh) release and its muscarinic receptor-mediated regulation (i.e. muscarinic autoreceptor function, MAF) during maturation as a possible contributing factor to age-related differences in sensitivity. Cortical and striatal slices were prelabeled with [ 3H ]choline chloride, superfused in the presence or absence of the anticholinesterase physostigmine (PHY, 20 μM) and stimulated twice (S1 and S2) with a high concentration of potassium chloride (20 mM). Depolarization-stimulated ACh release (DSAR) was lowest in neonatal, intermediate in juvenile and markedly higher in adult tissues. MAF was not detectable in tissues from neonatal rats but was present in juvenile and adult tissues. ACh release and MAF were studied at 4, 24 and 96 h following oral exposure to CPF (0, 0.5 or 1×LD 10). In general, 40–60% and 80–90% maximal AChE inhibition followed exposure to the respective 0.5 and 1×LD 10 dosages. DSAR was decreased in neonatal cortex 1 day after LD 10 exposure but increased in juvenile striatum 1 day after LD 10 treatment. In adults, DSAR was reduced at 4 and 24 h after exposure, but increased 96 h after CPF exposure. In juveniles, MAF was reduced in both brain regions at 24 h after 0.5LD 10 exposure and at 24 and 96 h after LD 10 exposure in cortex. A later reduction in MAF was noted in adult tissues (i.e. only at 96 h after LD 10 treatment). Together, the results suggest that ACh release dynamics in brain vary markedly during postnatal maturation and that acute CPF exposure can alter ACh release in an age-related manner. The functional status of presynaptic processes regulating neurotransmitter release may contribute to age-related neurotoxicity elicited by high-dose exposures to chlorpyrifos.

D 1 Agonist Dihydrexidine Releases Acetylcholine and Improves Cognitive Performance in Rats

Dihydrexidine is a selective, full-efficacy dopamine D 1 receptor agonist that has displayed therapeutic potential in Parkinson’s disease by reversing motor deficits of MPTP-treated monkeys. The present study monitored the effects of dihydrexidine on acetylcholine release in rat brain by using in vivo microdialysis. Moderate doses of dihydrexidine [3 and 10 mg/kg, intraperitoneally (IP)] elevated extracellular concentrations of acetylcholine by 40–60% in rat striatum; higher doses did not significantly alter acetylcholine release. SCH 23390 blocked the dihydrexidine-induced increase, indicating a D 1 receptor-mediated action. A more robust stimulatory effect of dihydrexidine on acetylcholine release was observed in prefrontal cortex (to 300% of basal output) than in striatum. Dihydrexidine was also evaluated in a passive avoidance procedure in rats to determine if its neurochemical effects translated into cognition-enhancing activity; in this assay, dihydrexidine (0.3 mg/kg, IP) significantly improved the scopolamine-induced deficits. The results of these studies suggest that the acetylcholine-releasing properties of dihydrexidine and other D 1 agonists may underlie their cognition-enhancing activity and thus may have clinical value in the treatment of dementia.

Mechanism of action of β-bungarotoxin, a presynaptically acting phospholipase A 2 neurotoxin: Its effect on protein phosphorylation in rat brain synaptosomes

The snake venom phospholipase A 2 neurotoxin, β-bungarotoxin, acts presynaptically to alter acetylcholine release in both the peripheral and central nervous systems. In investigating the mechanism of this action, we found that β-bungarotoxin inhibited phosphorylation of synapsin I, GAP-43 and MARCKS in rat brain synaptosomes. This inhibition was not due to the inhibition of ATP synthesis, action of arachidonic acid metabolites, or stimulation of phosphatase activities. Furthermore, the activities of Ca 2+/calmodulin-kinase II, cAMP-kinase and protein kinase C were not altered by β-bungarotoxin in either synaptic plasma membranes or cytosol. When synaptic plasma membranes were treated with β-bungarotoxin, MARCKS phosphorylation was inhibited, and this inhibition was overcome by the addition of exogenous protein kinase C. These results suggest that the interaction between MARCKS and endogenous protein kinase C is altered by β-bungarotoxin. In contrast, Naja naja atra phospholipase A 2, a typical phospholipase A 2 enzyme, had effects on phosphorylation which were different from those of β-bungarotoxin: (1) inhibition of phosphorylation of synapsin I in intact synaptosomes was less potent than that by β-bungarotoxin; (2) it stimulated basal phosphorylation of GAP-43 and MARCKS; and (3) it increased the activity of protein kinase C. The inhibition of synapsin I phosphorylation by N. n. atra phospholipase A 2 in intact synaptosomes may be due to the inhibition of ATP synthesis. The stimulation of GAP-43 and MARCKS by N. n. atra phospholipase A 2 can be explained by the production of arachidonic acid, which stimulated protein kinase C activity to a similar extent as that caused by N. n. atra phospholipase A 2. Thus, the mechanism of action of β-bungarotoxin appears to be quite different from that of a phospholipase A 2 enzyme, suggesting that phospholipase A 2 activity of β-bungarotoxin may not be essential for its action. β-Bungarotoxin may be a useful tool to study the physiological role of phosphorylation of synaptosomal proteins in neurotransmitter release.

Differential effects of snake venom phospholipase A 2 neurotoxin (β-bungarotoxin) and enzyme ( Naja naja atra) on protein kinases

The phospholipase A 2 (PLA 2) neurotoxin, β-bungarotoxin (β-BuTX), presynaptically alters acetylcholine release. We previously found that β-BuTX inhibits protein phosphorylation in rat brain synaptosomes. This inhibition was not due to the inhibition of ATP synthesis, the action of arachidonic acid (AA) metabolites, or the stimulation of phosphatase activities. A typical PLA 2 enzyme from Naja naja atra ( N. n. atra) venom also inhibited phosphorylation but with lesser potency than that of β-BuTX. We now report the effects of β-BuTX and N. n. atra PLA 2 on the activities of protein kinases. Treatments of synaptic plasma membrane or cytosol with N. n. atra PLA 2 stimulated the activities of cAMP-dependent kinase, Ca 2+/calmodulindependent kinase II, and protein kinase C (PKC), whereas β-BuTX had no effect on these kinases. Calyculin A, a phosphatase-1 and -2A inhibitor, increased the stimulation of phosphorylation by N. n. atra PLA 2, indicating that the stimulation is not due to an inhibition of phosphatase activities. The stimulation of PKC by N. n. atra PLA 2 appears to be mediated by free fatty acids (FFAs) resulting from phospholipid hydrolysis by PLA 2, since (1) treatment of either synaptic plasma membrane or cytosol with N. n. atra PLA 2 produced large amounts of FFAs, and (2) AA, an exogenous FFA, stimulated PKC activity to an extent similar to that caused by N. n. atra PLA 2. Thus, the mechanisms of action of β-BuTX and N. n. atra PLA 2 appear quite different from each other although both agents inhibit phosphorylation in intact synaptosomes.

Acetylcholine release from airway cholinergic nerves in horses with heaves, an airway obstructive disease.

The present study was conducted to determine if acetylcholine (ACh) release from airway cholinergic nerves is increased and if modulation of ACh release by prejunctional receptors is altered in horses with heaves, an obstructive airway disease characterized by airway inflammation and bronchospasm. Trachealis strips and bronchial segments of normal horses and horses affected with heaves were suspended in 2-ml tissue baths. ACh release was induced by electrical field stimulation and the bath ACh content was measured by high performance liquid chromatography (HPLC) with electrochemical detection. In response to electrical field stimulation, the rate of ACh release from cholinergic nerves innervating the trachea was not significantly different between the two groups of horses. The nonselective muscarinic receptor antagonist atropine augmented ACh release to the same extent in both groups, indicating that there is no dysfunction of muscarinic autoreceptors on cholinergic nerves of large airways from horses with heaves. Compared with the data from normal horses, the inhibitory effect of the alpha 2-adrenoceptor agonist clonidine on ACh release was lacking in the bronchi and less potent in the trachea, suggesting that the prejunctional alpha 2-adrenoceptors are dysfunctional in horses with heaves. Neither exogenous prostaglandin E2 (PGE2) nor the cyclooxygenase inhibitor indomethacin influenced ACh release from the bronchi, suggesting that a decrease in airway mucosal PGE2 production reported previously in horses with heaves does not alter ACh release.

Acetylcholine release from airway cholinergic nerves in horses with heaves, an airway obstructive disease.

The present study was conducted to determine if acetylcholine (ACh) release from airway cholinergic nerves is increased and if modulation of ACh release by prejunctional receptors is altered in horses with heaves, an obstructive airway disease characterized by airway inflammation and bronchospasm. Trachealis strips and bronchial segments of normal horses and horses affected with heaves were suspended in 2-ml tissue baths. ACh release was induced by electrical field stimulation and the bath ACh content was measured by high performance liquid chromatography (HPLC) with electrochemical detection. In response to electrical field stimulation, the rate of ACh release from cholinergic nerves innervating the trachea was not significantly different between the two groups of horses. The nonselective muscarinic receptor antagonist atropine augmented ACh release to the same extent in both groups, indicating that there is no dysfunction of muscarinic autoreceptors on cholinergic nerves of large airways from horses with heaves. Compared with the data from normal horses, the inhibitory effect of the alpha 2-adrenoceptor agonist clonidine on ACh release was lacking in the bronchi and less potent in the trachea, suggesting that the prejunctional alpha 2-adrenoceptors are dysfunctional in horses with heaves. Neither exogenous prostaglandin E2 (PGE2) nor the cyclooxygenase inhibitor indomethacin influenced ACh release from the bronchi, suggesting that a decrease in airway mucosal PGE2 production reported previously in horses with heaves does not alter ACh release.

6-OHDA-induced lesion of the nigrostriatal dopaminergic neurons potentiates the inhibitory effect of 7-OHDPAT, a selective D 3 agonist, on acetylcholine release during striatal microdialysis in conscious rats

Using striatal microdialysis, we attempted to clarify whether or not locally infused 7-hydroxy- N, N-di-n-propyl-2-aminotetralin (7-OHDPAT), a selective D 3 agonist, modulates the basal acetylcholine (ACh) release in conscious rats sham-operated and lesioned with 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle. In the sham-operated rats, 7-OHDPAT at 1 μM decreased ACh release by 19% of control. In the 6-OHDA-lesioned rats, 7-OHDPAT (0.1–10 μM) decreased ACh release in a concentration-dependent manner. The decrease of 37% of control was seen at 1 μM 2 weeks after the lesion. The 7-OHDPAT (1 μM)-induced decrease was completely antagonized by 1 μM (−)-sulpiride, a D 2 and D 3 antagonist, while (−)-sulpiride at 1 μM alone failed to alter ACh release. There may exist intrastriatal D 3 receptors to inhibit ACh release, and supersensitization is evident in a 6-OHDA-lesioned rat Parkinson's model.

Human recombinant interleukin 1β suppresses acetylcholine release from rat myenteric plexus

Background: A marked suppression of acetylcholine (ACh) release from myenteric nerves in the inflamed intestine of rats infected by Trichinella spiralis has been shown. In this model, there is increased expression of interleukin 1β (IL-1β) in the myenteric plexus. Therefore, the ability of IL-1β to alter ACh release in longitudinal muscle-myenteric plexus (LMMP) preparations from noninfected rats was examined. Methods: LMMP preparations were loaded with [3H]choline before stimulation by KCl or electrical field stimulation. ACh release was recorded by measuring 3H in the superfusate. Experiments were performed in the presence or absence of human recombinant IL-1β. Results: IL-1β had no immediate effect on the basal or stimulated release of ACh. A marked suppression of ACh release was observed in tissues that had been preincubated with IL-1β for 60 minutes or more. The effect of IL-1β was concentration and time dependent with maximum suppression occurring with 10 ng/mL of the cytokine after a 90-minute incubation. The action of human recombinant (hr) IL-1β was abolished by boiling the cytokine for 20 minutes and was prevented by preincubating the cytokine with neutralizing antibody. The IL-1β effect was also blocked by cycloheximide and was spontaneously reversible after 60 minutes. Conclusion: It was concluded that IL-1β suppresses ACh release via the formation and release of a protein mediator that could be another cytokine, including IL-1. Based on these findings, we consider IL-1β a putative mediator of the changes in cholinergic nerve function observed in the inflamed rat intestine.

Inhibition of phosphorylation of synapsin I and other synaptosomal proteins by beta-bungarotoxin, a phospholipase A2 neurotoxin.

Some snake venom neurotoxins, such as beta-bungarotoxin (beta-BuTX), which possess relatively low phospholipase A2 (PLA2) activity, act presynaptically to alter acetylcholine (ACh) release both in the periphery and in the CNS. In investigating the mechanism of this action, we found that beta-BuTX (5 and 15 nM) inhibited phosphorylation, in both resting and depolarized synaptosomes, of a wide range of proteins, including synapsin I. Naja naja atra PLA2, which has higher PLA2 activity, also inhibited phosphorylation but was less potent than beta-BuTX. At 1 nM, beta-BuTX and N. n. atra PLA2 inhibited phosphorylation of synapsin I only in depolarized synaptosomes. Synaptosomal ATP levels were not affected by 5 or 15 nM beta-BuTX or by 5 nM N. n. atra PLA2. Limited proteolysis, using Staphylococcus aureus V-8 protease, indicated that beta-BuTX inhibited phosphorylation of synapsin I in both the head and the tail regions. The inhibition of phosphorylation was not antagonized by nordihydroguaiaretic acid or indomethacin, suggesting that arachidonic acid derivatives do not mediate this inhibition. Furthermore, inhibition of phosphorylation by beta-BuTX and N. n. atra PLA2 was not altered in the presence of the phosphatase inhibitor okadaic acid, suggesting that stimulation of phosphatase activity is not responsible for this inhibition. Inhibition of protein phosphorylation by PLA2 neurotoxins and enzymes may be associated with an inhibition of ACh release.

High‐Affinity Choline Transport Regulation by Drug Administration During Postnatal Development

High-affinity choline transport sites specifically bind [3H]hemicholinium-3. Hemicholinium-3 binding sites are regulated by in vivo drug treatments in the same manner as these drugs alter acetylcholine release and high-affinity choline transport. The current study examines regulation of binding sites by in vivo drug administration for adult, day 15, and day 5 rats. Drugs or saline were administered intraperitoneally, and striatal and cortical membrane preparations were assayed. Control [3H]hemicholinium-3 binding increases twofold between postnatal days 5 and 15 only in striatum. After day 15, binding increases 2.7-fold in cortex and striatum. Nicotine treatment increases striatal and cortical hemicholinium-3 binding at all three ages, with greater percent increases at day 5. Haloperidol increases binding only in striatum, again with larger effects at day 5. Both striatal and cortical binding are reduced by oxotremorine; however, the magnitude of this effect is unchanged during development. Pentobarbital reduces binding only in striatum, with no developmental change. Atropine and apomorphine do not change binding from control values. In summary, all drug treatments effective in adults were already effective by day 5. Cholinergic terminals present early in development are regulated by similar nicotinic and muscarinic cholinergic, dopaminergic, and sedative-hypnotic mechanisms as the adult. Changes in magnitude may be due to changes in drug metabolism or to developmental differences in regulation.

Evidence that Somatostatin Enhances Endogenous Acetylcholine Release in the Rat Hippocampus

The present experiments show that somatostatin (SS)-like immunoreactive material is present in the hippocampus and that its release can be increased by K+ stimulation of rat hippocampal slices, suggesting that SS-like peptides may be of significance to neurotransmission in the hippocampus. Exogenous SS-28 and SS-14 enhanced the K(+)-evoked release of endogenous acetylcholine (ACh) from rat hippocampal slices, whereas amino-terminal fragments of SS-28 did not. The increased ACh release in the presence of either peptide appeared to be mediated by an interaction with SS receptors because cyclo-SS, a putative SS antagonist, abolished the effects of both SS-28 and SS-14. In addition, the increase in ACh release induced by SS-14 or SS-28 was antagonized by the calcium channel antagonists omega-conotoxin GVIA, nifedipine, and cinnarizine, implicating voltage-sensitive calcium channels in this effect. Moreover, the effect was sensitive to tetrodotoxin, suggesting an indirect action of the peptides at a site distal to cholinergic nerve terminals. Cysteamine, which has been reported to deplete SS content and to increase SS release in brain, augmented the basal and evoked release of ACh from hippocampal slices, without affecting SS-like content and release. Finally, neuropeptide Y, which is colocalized with SS in many neurons of the hippocampal formation, did not alter ACh release, nor did it facilitate the SS-induced increase. The results suggest that in the rat hippocampus, both SS-28 and SS-14 interact with SS receptors to regulate ACh release indirectly by a mechanism that involves alterations of calcium influx during depolarization.