skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Xylamine, a ligand for the catecholamine transporter

Abstract

Previous studies have established xylamine (N-2'-chloro-ethyl-N-ethyl-2-methylbenzylamine) irreversibly inhibits neuronal norepinephrine uptake with no concomitant effect on other neurotransmitter systems. Since xylamine is thought to alkylate transport-associated sites in the plasma membrane of noradrenergic neurons, so the loss of endogenous norepinephrine may be a consequence of neuronal membrane damage resulting from the alkylation of membrane components. In these studies, xylamine, under in vitro conditions, irreversibly inhibited both neuronal norepinephrine and dopamine uptake in the rat cortex and striatum, respectively. The efficacy of xylamine as a neuronal dopamine uptake inhibitor appeared to depend on its ability to access dopaminergic neurons during tissue exposure to the drug. In sympathetically innervated peripheral tissues, (/sup 3/H)xylamine was accumulated in noradrenergic neurons in a carrier-dependent manner. Although the data suggested that xylamine was interacting with the norepinephrine uptake carrier, (/sup 3/H)xylamine exposure to isolated synaptic membranes from superior cervical ganglia revealed a large proportion of (/sup 3/H)xylamine binding that was not associated with the noradrenergic transporter. For a closer characterization of xylamine binding in synaptic membranes, brain tissue was chosen as a more practical source of these membranes. While these experiments did not meet with great success, xylamine remains potentially useful as a ligand for identifyingmore » the catecholamine transporter, particularly in conjunction with procedures for protein purification and reconstitution.« less

Authors:
Publication Date:
Research Org.:
California Univ., Los Angeles (USA)
OSTI Identifier:
6990819
Resource Type:
Thesis/Dissertation
Resource Relation:
Other Information: Thesis (Ph. D.)
Country of Publication:
United States
Language:
English
Subject:
62 RADIOLOGY AND NUCLEAR MEDICINE; 59 BASIC BIOLOGICAL SCIENCES; CATECHOLAMINES; RADIORECEPTOR ASSAY; DOPAMINE; UPTAKE; TRITIUM COMPOUNDS; AMINES; BRAIN; INHIBITION; LIGANDS; NERVE CELLS; RATS; RECEPTORS; ANIMAL CELLS; ANIMALS; AROMATICS; AUTONOMIC NERVOUS SYSTEM AGENTS; BODY; CARDIOTONICS; CARDIOVASCULAR AGENTS; CENTRAL NERVOUS SYSTEM; DRUGS; HYDROXY COMPOUNDS; ISOTOPE APPLICATIONS; LABELLED COMPOUNDS; MAMMALS; MEMBRANE PROTEINS; NERVOUS SYSTEM; NEUROREGULATORS; ORGANIC COMPOUNDS; ORGANS; PHENOLS; POLYPHENOLS; PROTEINS; RODENTS; SOMATIC CELLS; SYMPATHOMIMETICS; TRACER TECHNIQUES; VERTEBRATES 550601* -- Medicine-- Unsealed Radionuclides in Diagnostics; 550201 -- Biochemistry-- Tracer Techniques

Citation Formats

Waggaman, L.A. Xylamine, a ligand for the catecholamine transporter. United States: N. p., 1985. Web.
Waggaman, L.A. Xylamine, a ligand for the catecholamine transporter. United States.
Waggaman, L.A. 1985. "Xylamine, a ligand for the catecholamine transporter". United States. doi:.
@article{osti_6990819,
title = {Xylamine, a ligand for the catecholamine transporter},
author = {Waggaman, L.A.},
abstractNote = {Previous studies have established xylamine (N-2'-chloro-ethyl-N-ethyl-2-methylbenzylamine) irreversibly inhibits neuronal norepinephrine uptake with no concomitant effect on other neurotransmitter systems. Since xylamine is thought to alkylate transport-associated sites in the plasma membrane of noradrenergic neurons, so the loss of endogenous norepinephrine may be a consequence of neuronal membrane damage resulting from the alkylation of membrane components. In these studies, xylamine, under in vitro conditions, irreversibly inhibited both neuronal norepinephrine and dopamine uptake in the rat cortex and striatum, respectively. The efficacy of xylamine as a neuronal dopamine uptake inhibitor appeared to depend on its ability to access dopaminergic neurons during tissue exposure to the drug. In sympathetically innervated peripheral tissues, (/sup 3/H)xylamine was accumulated in noradrenergic neurons in a carrier-dependent manner. Although the data suggested that xylamine was interacting with the norepinephrine uptake carrier, (/sup 3/H)xylamine exposure to isolated synaptic membranes from superior cervical ganglia revealed a large proportion of (/sup 3/H)xylamine binding that was not associated with the noradrenergic transporter. For a closer characterization of xylamine binding in synaptic membranes, brain tissue was chosen as a more practical source of these membranes. While these experiments did not meet with great success, xylamine remains potentially useful as a ligand for identifying the catecholamine transporter, particularly in conjunction with procedures for protein purification and reconstitution.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = 1985,
month = 1
}

Thesis/Dissertation:
Other availability
Please see Document Availability for additional information on obtaining the full-text document. Library patrons may search WorldCat to identify libraries that hold this thesis or dissertation.

Save / Share:
  • In primary cultures of bovine adrenomedullary cells, catecholamine uptake was found to be a saturable process exhibiting Michaelis-Menten kinetics with an apparent K/sub m/ for 1-norepinephrine of 0.5 ..mu..M. Radiolabeled catecholamines were employed to study the general characteristics and kinetic properties of catecholamine transport in cultured adrenomedullary cells. This process was found to be temperature, energy and Na/sup +/-dependent. In addition, uptake required the presence of extracellular Cl/sup -/, K/sup +/, and divalent cation such as Mn/sup 2 +/, Ca/sup 2 +/, or Mg/sup 2 +/. Agents that induce Ca/sup 2 +/-dependent, exocytotic secretion of catecholamines from these cells hadmore » significant inhibitory effects on catecholamine uptake. The secretagogues, nicotine, veratridine and elevated extracellular K/sup +/ concentrations, were all found to inhibit norepinephrine uptake. The inhibitory effects of the secretagogues could be fully demonstrated in the absence of catecholamine secretion. Investigation into the mechanism of catecholamine transport was pursued by measuring the effects of various catecholamine altering conditions or agents on the cellular membrane potential and/or the inwardly directed Na/sup +/ concentration gradient. Changes in the membrane potential were determined biochemically using tetraphenylphosphonium ion distribution, whereas changes in the Na/sup +/-gradient were assessed using /sup 22/Na/sup +/ distribution.« less
  • The catecholamine uptake system of PC12 was characterized. PC12 cells took up both norepinephrine (NE) and dopamine (DA) from the external medium. Uptake of both substances had Na/sup +/-dependent and Na/sup +/-independent portions. The Na/sup +/-dependent portion followed Michaelis-Menten kinetics. For NE the K/sub m/ was 0.5 ..mu..M, and the V/sub max/ was 2.7 pmol min/sup -1/ (mg protein)/sup -1/. For DA the K/sub m/ was 0.2 ..mu..M and the V/sub max/ was 3.8 pmol min/sup -1/ (mg protein)/sup -1/. The uptake of both substances was inhibited by desmethylimipramine with an IC/sub 50/ of 0.01 ..mu..M and by benztropine withmore » an IC/sub 50/ of 1 ..mu..M. These results suggest that NE and DA are transported by the same uptake system. Xylamine (N-2-chloroethyl-N-ethyl-2-methylbenzylamine) irreversibly inhibited the NE uptake (IC/sub 50/ = 15 ..mu..M). This inhibitions was Na/sup +/-dependent and was prevented by the coincubation of xylamine with cocaine or NE during the exposure of the cells to xylamine. These results indicate that xylamine must interact with the functioning NE uptake system to inhibit the uptake. PC12 accumulated (/sup 3/H)xylamine; this uptake had Na/sup +/-dependent and Na/sup +/-independent portions.« less
  • Dopamine and norepinephrine are inactivated by specific high affinity transport systems which mediate the recapture of the amines into presynaptic nerve terminals. (/sup 3/H)Maxindol labels neuronal dopamine uptake sites in corpus striatum membranes and neuronal norepinephrine uptake sites in cerebral cortex and submaxillary/sublingual gland membranes. The potencies of various inhibitors of biogenic amine uptake in reducing (/sup 3/H)mazindol binding in striatal membranes correlate with their potencies for inhibition of neurona (/sup 3/H)dopamine accumulation, whereas their potencies in reducing (/sup 3/H)mazindol binding to cortical and salivary gland membranes correlate with their potencies for inhibition of neuronal (/sup 3/H)norepinephrine accumulation. The associationmore » of (/sup 3/H)mazindol binding sites with neuronal dopamine uptake sites in the corpus striatum is further supported by the reduction of (/sup 3/H)mazindol binding sites in striatal membranes following destruction of dopaminergic neurons by 6-hydroxydopamine. Similarly, destruction of noradrenergic neurons by N-(2-chloro-ethyl)-N-ethyl-2-bromobenzylamine(DSP-4) decreases (/sup 3/H)mazindol binding to cortical membranes. Dopamine and norepinephrine uptake sites in rat brain have been differentially visualized using (/sup 3/H)mazindol autoradiography. N-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produces neuropathologic and clinical abnormalities in humans and animals that closely resemble idiopathic Parkinson disease. (/sup 3/H)MPTP binds with high affinity to brain membranes. The chemical specificity of the binding sites corresponds to structure-activity requirements for neurotoxicity.« less
  • The ability of catecholamines to modulate cell proliferation, differentiation and morphogenesis in other systems, and modulate adenylate cyclase activity in the developing palate during the period of cellular differentiation, made it of interest to determine their involvement in palatal ontogenesis. Catecholamines exert their physiologic effects via interaction with distinct membrane-bound receptors, one class being the B-adrenergic receptors which are coupled to stimulation of adenylate cyclase and the generation of cAMP. A direct radioligand binding technique utilizing the B-adrenergic antagonist (/sup 3/H)-dihydroalprenolol ((/sup 3/H)-DHA) was employed in the identification of B-adrenergic receptors in the developing murine secondary palate. Specific binding ofmore » (/sup 3/H)-DHA in embryonic (day 13) palatal tissue homogenates was saturable and of high affinity. The functionality of B-adrenergic receptor binding sites was assessed from the ability of embryonic palate mesenchmyal cells in vitro to respond to catecholamines with elevations of cAMP. Embryonic palate mesenchymal cells responded to various B-adrenergic catecholamine agonists with significant, dose-dependent accumulations of intracellular cAMP. Embryonic (day 13) maxillary tissue homogenates were analyzed for the presence of catecholamines by high performance liquid chromatography and radioenzymatic assay. Since normal palatal and craniofacial morphogenesis depends on proper temporal and spatial patterns of growth, the effect of B-adrenergic catecholamines on embryonic palate mesenchymal cell proliferation was investigated.« less
  • The discovery of a glycolate transporter in the pea (Pisum sativum) chloroplast envelope is described. Several novel silicone oil centrifugation methods were developed to resolve the initial rate kinetics of (/sup 14/C)glycolate transport by isolated, intact pea chloroplasts. Chloroplast glycolate transport was found to be carrier mediated. Transport rates saturated with increasing glycolate concentration. N-Ethylmaleimide (NEM) pretreatment of chloroplasts inhibited transport, an inhibition prevented by glycolate. Glycolate distributed across the envelope in a way which equalized stromal and medium glycolic acid concentrations, limiting possible transport mechanisms to facilitated glycolic acid diffusion, proton symport or hydroxyl antiport. The effects of stomalmore » and medium pH's on the K/sub m/ and V/sub max/ fit the predictions of mobile carrier kinetic models of hydroxyl antiport or proton symport (H/sup +/ binds first). The carrier mediated transport was fast enough to be consistent with in vivo rates of photorespiration. The 2-hydroxymonocarboxylates, glycerate, lactate and glyoxylate are competitive inhibitors of chloroplast glycolate uptake. Glyoxylate, D-lactate and D-glycerate cause glycolate counterflow, indicating that they are also substrates of the glycolate carrier. This finding was confirmed for D-glycerate by studies on glycolate effects on (1-/sup 14/C)D-glycerate transport.« less