Search

Opioid-induced constipation (OIC) is a common side effect of opioid pharmacotherapy for the management of pain because opioid agonists bind to µ -opioid receptors in the enteric nervous system (ENS). Naloxegol, a polyethylene glycol derivative of naloxol, which is a derivative of naloxone and a peripherally acting µ -opioid receptor antagonist, targets the physiologic mechanisms that cause OIC. Pharmacologic measures of opioid activity and pharmacokinetic measures of central nervous system (CNS) penetration were employed to characterize the mechanism of action of naloxegol. At the human µ -opioid receptor in vitro, naloxegol was a potent inhibitor of binding ( K i = 7.42 nM) and a neutral competitive antagonist (p A 2 - 7.95); agonist effects were <10% up to 30 μ M and identical to those of naloxone. The oral doses achieving 50% of the maximal effect in the rat for antagonism of morphine-induced inhibition of gastrointestinal transit and morphine-induced antinociception in the hot plate assay were 23.1 and 55.4 mg/kg for naloxegol and 0.69 and 1.14 mg/kg by for naloxone, respectively. In the human colon adenocarcinoma cell transport assay, naloxegol was a substrate for the P-glycoprotein transporter, with low apparent permeability in the apical to basolateral direction, and penetrated the CNS 15-fold slower than naloxone in a rat brain perfusion model. Naloxegol-derived radioactivity was poorly distributed throughout the rat CNS and was eliminated from most tissues within 24 hours. These findings corroborate phase 3 clinical studies demonstrating that naloxegol relieves OIC-associated symptoms in patients with chronic noncancer pain by antagonizing the µ -opioid receptor in the ENS while preserving CNS-mediated analgesia., (Copyright © 2017 The Author(s).)

Purinergic receptor P2X3 has been linked to analgesia in a number of pre-clinical models of pain, and is expressed in the human pain perception pathway. Only few P2X3-selective antagonists have been reported to date. This Letter describes the SAR and in vivo analgesic profile of a novel scaffold of selective P2X3 antagonists., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

An oral, peripherally restricted CB1/CB2 agonist could provide an interesting approach to treat chronic pain by harnessing the analgesic properties of cannabinoids but without the well-known central side effects. γ-Carbolines are a novel class of potent mixed CB1/CB2 agonists characterized by attractive physicochemical properties including high aqueous solubility. Optimization of the series has led to the discovery of 29, which has oral activity in a rat inflammatory pain model and limited brain exposure at analgesic doses, consistent with a lower risk of CNS-mediated tolerability issues., (Copyright © 2012. Published by Elsevier Ltd.)

Neuromedin U (NMU), through its cognate receptor NMUR2 in the central nervous system, regulates several important physiological functions, including energy balance, stress response, and nociception. By random screening of our corporate compound collection with a ligand binding assay, we discovered (R)-5'-(phenylaminocarbonylamino)spiro[1-azabicyclo[2.2.2]octane-3,2'(3'H)-furo[2,3-b]pyridine] (R-PSOP), a highly potent and selective NMUR2 antagonist. R-PSOP is a nonpeptidic small-molecule with the chemical composition C(20)N(4)O(2)H(22). In competition binding experiments, this compound was found to bind to NMUR2 with high affinity; the K(i) values were determined to be 52 and 32 nM for the human and rat NMUR2, respectively. Moreover, in functional assays measuring phosphoinositide turnover or intracellular calcium mobilization, R-PSOP strongly inhibited the responses stimulated by peptide agonists NMU-25, NMU-23, and NMU-8 in human embryonic kidney 293 cells expressing NMUR2. From Schild analyses, the functional K(b) values for R-PSOP were determined to be 92 and 155 nM at human and rat NMUR2, respectively. Highly selective for NMUR2, R-PSOP exhibited low affinity to the other subtype of NMU receptor, NMUR1, with a K(i) value >10 microM. R-PSOP in vivo attenuated NMU-23-evoked nociceptive responses in a rat spinal reflex preparation. To our knowledge, this is the first antagonist ever reported for NMU receptors. This compound could serve as a valuable tool for further understanding the physiological and pathophysiological roles of NMU system, while providing a chemical starting point that may lead to development of new therapeutics for treatment of eating disorders, obesity, pain, and stress-related disorders.

A series of 1-aminotetralin scaffolds was synthesized via metal-catalyzed ring-opening reactions of heterobicyclic alkenes. Small libraries of amides and amines were made using the amino group of each scaffold as a handle. Screening of these libraries against human opioid receptors led to the identification of (S)-(S)-5.2a as a high-affinity selective mu ligand (IC(50)mu=5 nM, kappa=707 nM, delta=3,795 nM) displaying mu-agonist/antagonist properties due to its partial agonism (EC(50)=2.6 microM; E(max)=18%).

The preparation and evaluation of a novel class of CB2 agonists based on a benzimidazole moiety are reported. They showed binding affinities up to 1nM towards the CB2 receptor with partial to full agonist potencies. They also demonstrated good to excellent selectivity (>1000-fold) over the CB1 receptor.

Several peptide fragments are produced by proteolytic cleavage of the opioid peptide precursor proenkephalin A, and among these are a number of enkephalin fragments, in particular bovine adrenal medulla peptide 22 (BAM22). These peptide products have been implicated in diverse biological functions, including analgesia. We have cloned a newly identified family of 'orphan' G protein--coupled receptors (GPCRs) and demonstrate that BAM22 and a number of its fragments bind to and activate these receptors with nanomolar affinities. This family of GPCRs is uniquely localized in the human and rat small sensory neuron, and we called this family the sensory neuron--specific G protein--coupled receptors (SNSRs). Receptors of the SNSR family are distinct from the traditional opioid receptors in their insensitivity to the classical opioid antagonist naloxone and poor activation by opioid ligands. The unique localization of SNSRs and their activation by proenkephalin A peptide fragments indicate a possible function for SNSRs in sensory neuron regulation and in the modulation of nociception.

Galanin is a 29-aa neuropeptide with a complex role in pain processing. Several galanin receptor subtypes are present in dorsal root ganglia and spinal cord with a differential distribution. Here, we describe a generation of a specific galanin R2 (GalR2) agonist, AR-M1896, and its application in studies of a rat neuropathic pain model (Bennett). The results show that in normal rats mechanical and cold allodynia of the hindpaw are induced after intrathecal infusion of low-dose galanin (25 ng per 0.5 microl/h). The same effect is seen with equimolar doses of AR-M1896 or AR-M961, an agonist both at GalR1 and GalR2 receptors. In allodynic Bennett model rats, the mechanical threshold increased dose-dependently after intrathecal injection of a high dose of AR-M961, whereas no effect was observed in the control or AR-M1896 group. No effect of either of the two compounds was observed in nonallodynic Bennett model rats. These data indicate that a low dose of galanin has a nociceptive role at the spinal cord level mediated by GalR2 receptors, whereas the antiallodynic effect of high-dose galanin on neuropathic pain is mediated by the GalR1 receptors. Thus, a selective GalR1 agonist may be used to treat neuropathic pain.

Nonpeptide delta opioid agonists are analgesics with a potentially improved side-effect and abuse liability profile, compared to classical opioids. Andrews analysis of the NIH nonpeptide lead SNC-80 suggested the removal of substituents not predicted to contribute to binding. This approach led to a simplified lead, N, N-diethyl-4-[phenyl(1-piperazinyl)methyl]benzamide (1), which retained potent binding affinity and selectivity to the human delta receptor (IC(50) = 11 nM, mu/delta = 740, kappa/delta > 900) and potency as a full agonist (EC(50) = 36 nM) but had a markedly reduced molecular weight, only one chiral center, and increased in vitro metabolic stability. From this lead, the key pharmacophore groups for delta receptor affinity and activation were more clearly defined by SAR and mutagenesis studies. Further structural modifications on the basis of 1 confirmed the importance of the N, N-diethylbenzamide group and the piperazine lower basic nitrogen for delta binding, in agreement with mutagenesis data. A number of piperazine N-alkyl substituents were tolerated. In contrast, modifications of the phenyl group led to the discovery of a series of diarylmethylpiperazines exemplified by N, N-diethyl-4-[1-piperazinyl(8-quinolinyl)methyl]benzamide (56) which had an improved in vitro binding profile (IC(50) = 0.5 nM, mu/delta = 1239, EC(50) = 3.6 nM) and increased in vitro metabolic stability compared to SNC-80.

The design, synthesis, and pharmacological evaluation of a novel class of delta opioid receptor agonists, N, N-diethyl-4-(phenylpiperidin-4-ylidenemethyl)benzamide (6a) and its analogues, are described. These compounds, formally derived from SNC-80 (2) by replacing the piperazine ring with a piperidine ring containing an exocyclic carbon carbon double bond, were found to bind with high affinity and exhibit excellent selectivity for the delta opioid receptor as full agonists. 6a, the simplest structure in the class, exhibited an IC(50) = 0.87 nM for the delta opioid receptors and extremely high selectivity over the mu receptors (mu/delta = 4370) and the kappa receptors (kappa/delta = 8590). Rat liver microsome studies on a selected number of compounds show these olefinic piperidine compounds (6) to be considerably more stable than SNC-80. This novel series of compounds appear to interact with delta opioid receptors in a similar way to SNC-80 since they demonstrate similar SAR. Two general approaches have been established for the synthesis of these compounds, based on dehydration of benzhydryl alcohols (7) and Suzuki coupling reactions of vinyl bromide (8), and are herewith reported.

The neuropeptide galanin is a 29- or 30-residue peptide whose physiological functions are mediated by G-protein-coupled receptors. Galanin's agonist activity has been shown to be associated with the N-terminal sequence, galanin(1-16). Conformational investigations previously carried out on full-length galanin have, furthermore, indicated the presence of a helical conformation in the neuropeptide's N-terminal domain. Several cyclic lactam analogues of galanin(1-16)-NH2 were prepared in an attempt to stabilize an N-terminal helix in the peptide. Here we describe and compare the solution conformational properties of these analogues in the presence of SDS micelles as determined by NMR, CD, and fluorescence spectroscopy. Differences in CD spectral profiles were observed among the compounds that were studied. Both c[D4, K8]Gal(1-16)-NH2 and c[D4,K8]Gal(1-12)-NH2 adopted stable helical conformations in the micelle solution. On the basis of the analyses of their respective alpha H chemical shifts and NOE patterns, this helix was localized to the first 10 residues. The distance between the aromatic rings of Trp2 and Tyr9 in c[D4, K8]Gal(1-16)-NH2 was determined to be 10.8 +/- 3 A from fluorescence resonance energy transfer measurements. This interchromophore spacing was found to be more consistent with a helical structure than an extended one. Removal of the Gly1 residue in compounds c[D4,K8]Gal(1-16)-NH2 and c[D4, K8]Gal(1-12)-NH2 resulted in a loss of helical conformation and a concomitant reduction in binding potency at the GalR1 receptor but not at the GalR2 receptor. The nuclear Overhauser enhancements obtained for the Gly1 deficient analogues did, however, reveal the presence of nascent helical structures within the N-terminal sequence. Decreasing the ring structure size in c[D4, K8]Gal(1-16)-NH2 by replacing Lys8 with an ornithine residue or by changing the position of the single lysine residue from eight to seven was accompanied by a complete loss of helical structure and dramatically reduced receptor affinity. It is concluded from the data obtained for the series of cyclic galanin(1-16)-NH2 analogues that both the ring structure size and the presence of an N-terminal glycine residue are important for stabilizing an N-terminal helix in these compounds. However, although an N-terminal helix constitutes a predominant portion of the conformational ensemble for compounds c[D4,K8]Gal(1-16)-NH2 and c[D4, K8]Gal(1-12)-NH2, these peptides nevertheless are able to adopt other conformations in solution. Consequently, the correlation between the ability of the cyclic galanin analogues to adopt an N-terminal helix and bind to the GalR1 receptor may be considered as a working hypothesis.

AR-M100613 ([I]-Dmt-c[-D-Orn-2-Nal-D-Pro-D-Ala-]) is the iodinated analog of a cyclic casomorphin previously shown to be a potent antagonist at the delta opioid receptor. Specific [125I]AR-M100613 binding to rat whole brain membranes was saturable, reversible, and best fit to a one-site model (Kd = 0.080 +/- 0.008 nM, Bmax = 45.2 +/- 4.4 fmol/mg protein). [125I]AR-M100613 binding was displaced with high affinity by the delta opioid receptor ligands SNC-80, Deltorphin II and DPDPE but not the mu or kappa-selective receptor ligands DAMGO and U69593. Residual non-selective binding of [125I]AR-M 100613 to mu opioid receptors is blocked by the addition of CTOP to the assay buffer. [35S]GTPgammaS binding assays indicate that AR-M100613 is a potent, selective, and reversible antagonist for delta opioid receptors in rat brain membranes. The high-affinity, high specific activity, low nonspecific binding and antagonist profile of [125I]AR-M100613 favor its use as a radiochemical probe for delta opioid receptors.

Gene-knockout studies of melanin-concentrating hormone (MCH) and its effect on feeding and energy balance have firmly established MCH as an orexigenic (appetite-stimulating) peptide hormone. Here we identify MCH as the ligand for the orphan receptor SLC-1. The rat SLC-1 is activated by nanomolar concentrations of MCH and is coupled to the G protein G alpha i/o. The pattern of SLC-1 messenger RNA expression coincides with the distribution of MCH-containing nerve terminals and is consistent with the known central effects of MCH. Our identification of an MCH receptor could have implications for the development of new anti-obesity therapies.

We have identified a novel subtype of galanin receptor (GALR-2) in rat dorsal root ganglia and spinal cord. The open reading frame of GALR-2 is 1116 nucleotides long, encoding a protein of 372 amino acids with a theoretical molecular mass of 40.7 kD. Membranes prepared from stable pools of 293 cells expressing GALR-2, but not wild-type 293 cells, demonstrated high affinity galanin binding sites. Rat galanin and galanin-related peptides M40, C7, M15, and galanin effectively competed for binding; peptide C7 demonstrated a lower affinity for rGALR-2, and all these peptides were agonists at rGALR-2 when assessed on a microphysiometer. Studies on the expression of GALR-2 in various tissues by Northern and in situ hybridization analyses suggest a low abundance but wide distribution of GALR-2 mRNA, including several discrete areas in brain and spinal cord and a high abundance in the dorsal root ganglia.

Pharmacological study of Phe-Met-Leu-Phe-amide (FMRFa) receptors is hindered by the lack of selective ligands. The classification of these selective ligands is further hampered by the limited availability of functional assays. In this study, we evaluated several synthetic FMRFa analogs for agonist and antagonist activity by measuring their abilities to produce [35-S]-GTP-gamma-S stimulation or to inhibit FMRFa-induced [35S]-GTP-gamma-S binding in squid optic lobes. Analogs included acetyl-Phe-norLeu-Arg-Phe-amide (acFnLRFa), desamino-Tyr-Phe-Leu-Arg-amide (daYFLRa), desamino Tyr-Phe-norLeu-Arg-Phe-amide (daYFnLRFa), desamino Tyr-Phe-norLeu-Arg-[TIC]-amide (daYFnLR[TIC]a), desamino Tyr-Trp-norLeu-Arg-amide (daYWnLRa), (D)-Tyr-Phe-norLeu-Arg-Phe-amide (D)-YFnLRFa), Phe-Leu-Arg-Phe-amide (FLRFa), and the D-amino acid analogs of FMRFa (D-FMRFa, F-(D)-MRFa and FM-(D)-RFa). For agonist studies, full dose-response curves were generated and analyzed for potency and efficacy (maximal percent effect). FMRFamide as well as analogs ac-FnLRFa, daYFnLRFa, daYFnLR[TIC]a, D-YFnLRFa, FLRFa, and (D)-FMRFa stimulated [35S]-GTP-gamma-S binding. Analogs daYWnLRa, daYFLRa, F-(D)-MRFa, and FM-(D)-RFa failed to stimulate either [35S]-GTP-gamma-S binding or to inhibit FMRFa-induced [35S]-GTP-gamma-S binding. The rank order of potency was daYFnLRFa > or = daYFnLRF[TIC]a > acFnLRFa > (D)YFnLRFa > FLRFa > or = FMRFa >> (D)-FMRFa. The order of efficacy was daYFnLRFa = acFnLRFa = (D)-YFnLRFa > FLRFa = FMRFa > or = (D)-FMRFa > or = daYFnLRF[TIC]a. Peptide analog daYFnLR[TIC]a was less efficacious (59% maximal stimulation) than analogs daYFnLRFa, acFnLRFa, and (D)-YFnLRFa (113-146% maximal stimulation). A maximal concentration of daYFnLR[TIC]a (10 microM) reduced daYFnLRFa, acFnLRFa, and (D)-YFnLRFa induced [35S]-GTP-gamma-S stimulation, indicating that daYFnLR[TIC]a is a partial agonist at the receptor stimulated by the FMRFamide analogs. Analysis of the structural requirements needed for promoting [35S]-GTP-gamma-S binding show that elongation (i.e., daYFnLRFa, D-YFnLRFa) or modification of Phe1 (ac-FnLRFa) leads to increased efficacy and potency. Moreover, elimination of the C-terminal Phe (daYWnLRa, daYFLRa,) leads to a loss of biological activity. However, substitution with L-1,2,3,4 tetrahydroisoquinoline-3-carboxylic acid, a rigid analog of the C-terminal Phe (daYFnLR[TIC]a), leads to decreased efficacy but not loss of potency. The data suggest that immobilization or modification of the C-terminal Phe may produce highly selective and potent FMRFamide antagonists. These results agree with published receptor radioligand studies and indicate that the [35S]GTP-gamma-S assay may be useful in classifying novel FMRFamide-selective ligands.

In an attempt to attenuate specifically identified receptors through photolysis, a four-step synthesis is of a useful tethered derivative of Azure-B (Az) was developed After characterization, this derivative was covalently attached to CFMRFamide, CFMRF, and CLRFamide (i.e., three different neuropeptide analogues of the putative neurotransmitter FMRFamide. This resulted in the formation of three dye-neuropeptide conjugates: Az-CFMRFamide, Az-CFMRF, and Az-CLRFamide.

Using a radioligand binding assay, we examined ionic modulation and G protein coupling of neuropeptide FF (NPFF) receptors in membranes of rat brain and spinal cord. We found that NaCl (but not KCl or LiCl) and MgCl2 increased specific 125I-YLFQPQRFamide (125I-Y8Fa) binding to NPFF receptors in both tissues in a dose-dependent manner, with optimal conditions being 60 mM NaCl and 1 mM MgCl2. Guanine nucleotides dose-dependently inhibited specific 125I-Y8Fa binding to rat brain and spinal cord membranes with maximal effects of 64 +/- 6 and 71 +/- 2%, respectively. The order of potency was nonhydrolyzable GTP analogues > GTP > or = GDP >> GMP, ATP. The guanine nucleotide inhibition was observed in the absence and presence of NaCl and MgCl2. The mechanism of inhibition in spinal cord membranes appeared to be a reduction in the number of NPFF receptors; in one experiment, control KD and Bmax values were 0.068 nM and 7.2 fmol/mg of protein, respectively, and with 0.1 microM guanylylimidodiphosphate the respective values were 0.081 nM and 4.9 fmol/mg, a 32% reduction in receptor number. Similar results were obtained with guanosine 5'-O-(3-thiotriphosphate). Our data suggest that 125I-Y8Fa binding sites in rat CNS are G protein-coupled NPFF receptors regulated by GTP and cations.

Cytoplasmic calcium concentration was measured in neurosecretory nerve terminals (neurosecretosomes) isolated from rat neurohypophyses by fura-2 fluorescence measurements and digital video microscopy. Hormone release and cytoplasmic calcium concentration were measured during depolarizations induced by elevated extracellular potassium concentration. During prolonged depolarizations with 55 mM [K+]o, the cytoplasmic calcium concentration remained elevated as long as depolarization persisted, while secretion inactivated after the initial sharp rise. The amplitude and duration of the increase in [Ca2+]i was dependent on the degree of depolarization such that upon low levels of depolarizations (12.5 mM or 25 mM [K+]o), the calcium responses were smaller and relatively transient, and with higher levels of depolarization (55 mM [K+]o) the responses were sustained and were higher in amplitude. Responses to low levels of depolarization were less sensitive to the dihydropyridine calcium channel blocker, nimodipine, while the increase in [Ca2+]i induced by 55 mM [K+]o became transient, and was significantly smaller. These observations suggest that these peptidergic nerve terminals possess at least two different types of voltage-gated calcium channels. Removal of extracellular sodium resulted in a significant increase in [Ca2+]i and secretion in the absence of depolarizing stimulus, suggesting that sodium-calcium exchange mechanism is operative in these nerve terminals. Although the [Ca2+]i increase was of similar magnitude to the depolarization-induced changes, the resultant secretion was 10-fold lower, but the rate of inactivation of secretion, however, was comparable.

The octapeptide FLFQPQRFamide (neuropeptide FF or F8Fa) may play a role in opiate dependence and subsequent abstinence syndrome. Previously, NPFF precipitated opiate abstinence syndrome, while IgG from NPFF antiserum attenuated subsequent naloxone-precipitated abstinence signs in dependent rats. The peptide desamino YFLFQPQRamide (daY8Ra) was synthesized as a possible NPFF antagonist. At a dose of 600 ng ICV, daY8Ra significantly attenuated (p less than 0.001) the number of abstinence-like signs subsequently induced by 10 micrograms NPFF ICV, suggesting that daY8Ra does have antagonist activity against NPFF. Pretreatment of morphine-dependent rats with the same dose of daY8Ra also significantly attenuated (p less than 0.001) the abstinence signs subsequently precipitated by 10 micrograms naloxone ICV. Pretreatment with 600 ng of NPFF itself, or of NPFF modified at the N-terminal only (daY9Fa), failed to attenuate subsequent naloxone-precipitated abstinence, suggesting that the C-terminal modification is critical for NPFF antagonist activity. It should be noted, however, that higher doses of daY8Ra (2 micrograms or more) can precipitate some abstinence signs in a manner similar to NPFF.

Neurosecretory terminals (neurosecretosomes, NSS) were isolated from rat neurohypophyses. High [K+]o or veratridine stimulated secretion of vasopressin and oxytocin by up to approximately 100-fold. Stimulated secretion was dependent on calcium and temperature, and could be elicited from NSS maintained in culture for 4 days. After overnight culture of the NSS, secretion was still inhibited by calcium channel blockers (cobalt, dihydropyridines, omega-conotoxin, D 600) and kappa opiates (dynorphin and U50488). Ionomycin evoked dose- and calcium-dependent hormone release, with a Hill coefficient for calcium of 1.74. High [K+]o enhanced the 5 microM ionomycin-induced secretion, apparently through calcium entry rather than depolarization, as the increase in secretion was abolished by 100 microM D 600. During prolonged depolarization the hormone secretion peaked within 2 min, then declined to near basal levels. Depolarization for 25 min without calcium neither activated secretion nor prevented subsequent secretion on readdition of calcium, suggesting that the decline in secretion was not due to membrane depolarization. Indeed, the rates of decline in secretion were similar for different levels of depolarization (0.070 +/- 0.003 and 0.081 +/- 0.003 min-1 for 25 and 45 mM [K+]o, respectively). Four minutes after the onset of continuous depolarization (45 mM [K+]o) in the presence of calcium, the declining secretion was still dependent on voltage-activated calcium influx through channels sensitive to D 600 and nitrendipine. The results presented here suggest that the decline in secretion during prolonged depolarizing stimuli may be due to exhaustion, inactivation, or desensitization of a calcium-triggered event.

1. We studied the effects of extracellular sodium on the secretion of vasopressin (VP) and oxytocin (OT) and the efflux of 45Ca from isolated, perfused nerve endings of the rat neurohypophysis (neurosecretosomes). 2. Upon removal of sodium from the perfusing medium, basal release of VP and OT increased by 3.95 +/- 0.23- and 3.71 +/- 0.22-fold, respectively, followed by a decline to about double the levels in normal (150 mM) sodium (P less than or equal to 0.1). 3. Compared to neurosecretosomes perfused in normal (150 mM) sodium, omission of sodium from the medium augmented ionomycin-induced VP and OT secretion by 66 +/- 5- and 20 +/- 3-fold, respectively, and A23187-induced secretion was increased 1.3 +/- 0.4- and 1.3 +/- 0.1-fold (P less than or equal to 0.01 for both ionophores). 4. The inhibition of ionomycin-induced secretion by sodium was concentration dependent (P less than or equal to 0.01 for sodium greater than or equal to 5 mM); the IC50 was about 10 mM sodium for both hormones, and the Hill slope was close to -1. 5. The rate of 45Ca efflux from neurosecretosomes showed 2.7 +/- 0.1-fold stimulation upon increasing sodium from 4.5 to 150 mM (P less than or equal to 0.01). 6. Our results suggest that sodium inhibits basal and stimulated secretion at the nerve terminal, possibly by reducing intraterminal calcium through sodium/calcium exchange.