a-85380 and Pain

A-85380 [3-(2(s)-azetidinylmethoxy) pyridine] is a neuronal nicotinic acetylcholine receptor (nAChR) agonist that has been a useful tool in the investigation of the function of nAChRs in both preclinical and clinical studies. Amongst nAChR subtypes, A-85380 shows selectivity for the alpha(4)beta(2) vs. the alpha(7) or alpha(1)beta(1)deltagamma nAChRs. In functional in vitro cation flux assays, A-85380 is a potent and full agonist. A-85380 has a broad-spectrum analgesic profile with efficacy in acute, persistent, and neuropathic pain models. As demonstrated using selective nAChR antagonists or alpha(4) antisense, the alpha(4)beta(2) nAChR mediates the analgesic effects of A-85380. Interestingly, the site of action depends upon the type of pain as antinociception is mediated by descending inhibition into the spinal cord whereas anti-allodynia in neuropathic pain is mediated at both central and peripheral sites. Radiolabelled forms of A-85380 have been developed and shown to be safe for use in vivo in humans. In clinical studies using positron and photon emission tomography, marked decreases in alpha(4)beta(2) nAChRs have been seen in patients with Parkinson's and Alzheimer's disease. Although not developed as a therapeutic agent, A-85380 has proven to be an important component in the development of novel nAChR ligands for the treatment of pain and other disorders.

Topics: Animals; Azetidines; Humans; In Vitro Techniques; Mental Disorders; Nicotinic Agonists; Pain; Pharmacology, Clinical; Radioligand Assay; Receptors, Nicotinic

Recent evidence has suggested that the anti-allodynic effect of neuronal acetylcholine receptor (nAChR) agonists may have a peripheral component [L.E. Rueter, K.L. Kohlhaas, P. Curzon, C.S. Surowy, M.D. Meyer, Peripheral and central sites of action for A-85380 in the spinal nerve ligation model of neuropathic pain, Pain 103 (2003) 269-276]. In further studies of the peripheral anti-allodynic mechanisms of nAChR agonists, we investigated the function of nAChRs in acutely isolated dorsal root ganglion (DRG) neurons from allodynic [L5-L6 spinal nerve ligation (SNL)] and naive adult rats. Following determination of cell diameter and membrane capacitance, responses to rapid applications of nAChR agonists were recorded under whole cell patch clamp. nAChR inward currents were observed in approximately 60% of naive neurons, across small, medium, and large diameter cells. Evoked nAChR currents could be clustered into three broad classes: fast transient, biphasic, and slow desensitizing currents, consistent with multiple subtypes of nAChR expressed in DRG [J.R. Genzen, W. Van Cleve, D.S. McGehee, Dorsal root ganglion neurons express multiple nicotinic acetylcholine receptor subtypes, J. Neurophysiol. 86 (2001) 1773-1782]. In contrast, in neurons from allodynic animals, the occurrence and amplitude of responses to nAChR agonists were significantly reduced. Reduced responsiveness to nAChR agonists covered the range of DRG neuron sizes. The decrease in the responsiveness to nAChR agonists was not seen in neighboring uninjured L4 neurons. The significant decrease in the number of cells with nAChR agonist responses, compounded with the significant decrease in response amplitude, indicates that there is a marked down regulation of functional nAChRs in DRG somata associated with SNL.

Topics: Acetylcholine; Animals; Azetidines; Cell Count; Cells, Cultured; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Ganglia, Spinal; Ligation; Lumbosacral Region; Male; Mecamylamine; Membrane Potentials; Neurons; Nicotinic Agonists; Nicotinic Antagonists; Pain; Patch-Clamp Techniques; Rats; Rats, Sprague-Dawley; Receptors, Nicotinic; Spinal Cord Injuries

Pharmacological activation of neuronal nicotinic acetylcholine receptors can produce non-opioid antinociception in rodents. However, multiple nAChR subtypes exist, the most abundant of which contain alpha4 and beta2 subunits. The purpose of the present study was to investigate the role of alpha4-containing nAChRs in mediating nicotinic antinociception using an in vivo antisense strategy. Both i.c.v. infusion and repeated bolus injections into the cerebral aqueduct of an antisense oligonucleotide against the alpha4 subunit significantly attenuated the antinociceptive effects of the nAChR agonist A-85380 in the paw withdrawal test of acute thermal pain. Rats treated with a scrambled oligonucleotide displayed a full antinociceptive response to A-85380, while discontinuing antisense treatment restored the antinociceptive effects of the nicotinic agonist. Double immunohistochemical labeling revealed near-complete overlap of expression of the serotonin marker tryptophan hydroxylase and the alpha4 nAChR subunit in the dorsal raphe nucleus. The expression of alpha4-containing nAChRs by serotonergic neurons in the dorsal raphe offered a means to address nonspecific alpha4 knock-down, i.e., oligonucleotide-induced neurotoxicity. Immunohistochemical detection of alpha4 expression was reduced by nearly 50% in the dorsal raphe of antisense-treated rats as compared to either saline or missense-treated controls. In contrast, the expression of tryptophan hydroxylase, as well as, the alpha7 nAChR subunit in antisense-infused rats was similar to that observed in saline- and missense-treated controls. The results of these studies suggest that alpha4-containing nAChRs, possibly expressed by serotonergic neurons, are involved in nicotinic-mediated analgesia. However, these data do not eliminate the possibility that other nicotinic subunit combinations may also play a role in antinociception produced by nAChR activation.

Topics: Animals; Azetidines; Cerebral Aqueduct; Cerebral Ventricles; Edema; Injections, Intraventricular; Male; Microinjections; Nicotinic Agonists; Oligodeoxyribonucleotides, Antisense; Pain; Raphe Nuclei; Rats; Rats, Sprague-Dawley; Reaction Time; Receptors, Nicotinic; Thionucleotides

Systemic administration of nicotinic receptor (nAChR) agonists is antinociceptive in models of acute pain whereas their intrathecal (i. t.) administration has been reported to be antinociceptive, nociceptive or without effect. It has been hypothesized that the action induced is dependent upon the subtype and location of the nAChR activated. In addition, there is considerable evidence that nAChR ligand-induced antinociception is mediated by other neurotransmitter systems via descending pathways from the brainstem to the spinal cord. The present study investigated the effects of i. t. and systemic administration of A-85380, a novel nAChR agonist, in the paw withdrawal model of acute thermal pain in the rat. Given i.t. , A-85380 (1 and 10 nmol/rat) decreased the latency to paw withdrawal by 2-4 s. This pronociception was accompanied by a spontaneous flinching behavior. Both of these effects were differentially blocked by i.t. pretreatment with the nAChR antagonists mecamylamine (10 nmol)>MLA (100 nmol)>DHbetaE (50% with 1000 nmol) but not by alpha-bungarotoxin (0% at 0.63 nmol). Given systemically, A-85380 (0.56 micromol/kg, i.p.) induced antinociception as indicated by an increased latency to paw withdrawal, an effect differentially altered by i.t. pretreatment with monoaminergic antagonists (100 nmol/rat). While mecamylamine and prazosin had no effect, scopolamine, methysergide and MDL 72222 partially antagonized and idazoxan completely antagonized A-85380-induced antinociception. Finally, as measured by in vivo microdialysis, levels of 5-HT, but not NE, in the i.t. space of the lumber region of the spinal cord were significantly increased following the systemic administration of A-85380. Together these data suggest that the nociceptive properties of spinally administered nAChR agents are not mediated by either an alpha(4)beta(2) or an alpha(7) subtype nAChR, whereas the antinociceptive properties of systemically-administered nAChR agents are mediated by descending noradrenergic, serotonergic and muscarinic inhibitory pathways.

Topics: Adrenergic alpha-Agonists; Animals; Azetidines; Behavior, Animal; Bridged Bicyclo Compounds, Heterocyclic; Bungarotoxins; Clonidine; Dose-Response Relationship, Drug; Hot Temperature; Injections, Intraperitoneal; Injections, Spinal; Lumbosacral Region; Male; Mecamylamine; Microdialysis; Muscarinic Antagonists; Nicotinic Agonists; Nicotinic Antagonists; Norepinephrine; Pain; Pain Measurement; Pyridines; Rats; Rats, Sprague-Dawley; Reaction Time; Serotonin; Serotonin Antagonists; Spinal Cord

New members of a previously reported series of 3-pyridyl ether compounds are disclosed as novel, potent analgesic agents acting through neuronal nicotinic acetylcholine receptors. Both (R)-2-chloro-5-(2-azetidinylmethoxy)pyridine (ABT-594, 5) and its S-enantiomer (4) show potent analgesic activity in the mouse hot-plate assay following either intraperitoneal (i.p.) or oral (p.o.) administration, as well as activity in the mouse abdominal constriction (writhing) assay, a model of persistent pain. Compared to the S-enantiomer and to the prototypical potent nicotinic analgesic agent (+/-)-epibatidine, 5 shows diminished activity in models of peripheral side effects. Structure-activity studies of analogues related to 4 and 5 suggest that the N-unsubstituted azetidine moiety and the 2-chloro substituent on the pyridine ring are important contributors to potent analgesic activity.

Topics: Administration, Oral; Analgesics, Non-Narcotic; Animals; Azetidines; Diastole; Female; Humans; Injections, Intraperitoneal; Kinetics; Mice; Molecular Structure; Muscle Contraction; Neuroblastoma; Neurons; Nicotinic Agonists; Oocytes; Pain; Pain Measurement; Pyridines; Rats; Receptors, Nicotinic; Recombinant Proteins; Stereoisomerism; Structure-Activity Relationship; Tumor Cells, Cultured; Xenopus