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M K Stachowiak,
A Kucinski,
R Curl,
C Syposs,
Y Yang,
S Narla,
C Terranova,
D Prokop,
I Klejbor, M Bencherif,
B Birkaya,
T Corso,
A Parikh,
E S Tzanakakis,
S Wersinger,
E K Stachowiak
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ABSTRACT: Schizophrenia is a neurodevelopmental disorder featuring complex aberrations in the structure, wiring, and chemistry of multiple neuronal systems. The abnormal developmental trajectory of the brain appears to be established during gestation, long before clinical symptoms of the disease appear in early adult life. Many genes are associated with schizophrenia, however, altered expression of no one gene has been shown to be present in a majority of schizophrenia patients. How does altered expression of such a variety of genes lead to the complex set of abnormalities observed in the schizophrenic brain? We hypothesize that the protein products of these genes converge on common neurodevelopmental pathways that affect the development of multiple neural circuits and neurotransmitter systems. One such neurodevelopmental pathway is Integrative Nuclear FGFR1 Signaling (INFS). INFS integrates diverse neurogenic signals that direct the postmitotic development of embryonic stem cells, neural progenitors and immature neurons, by direct gene reprogramming. Additionally, FGFR1 and its partner proteins link multiple upstream pathways in which schizophrenia-linked genes are known to function and interact directly with those genes. A th-fgfr1(tk-) transgenic mouse with impaired FGF receptor signaling establishes a number of important characteristics that mimic human schizophrenia - a neurodevelopmental origin, anatomical abnormalities at birth, a delayed onset of behavioral symptoms, deficits across multiple domains of the disorder and symptom improvement with typical and atypical antipsychotics, 5-HT antagonists, and nicotinic receptor agonists. Our research suggests that altered FGF receptor signaling plays a central role in the developmental abnormalities underlying schizophrenia and that nicotinic agonists are an effective class of compounds for the treatment of schizophrenia.
Biological Psychiatry 12/2012; · 8.28 Impact Factor
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ABSTRACT: Genetic and post mortem evidence has implicated the α7 neuronal nicotinic receptor (NNR) in the etiology of schizophrenia and related disorders. In schizophrenia, enhanced subcortical dopamine (DA) correlates with positive and cognitive of the disease, including impairments in sensorimotor gating. We measured the levels of extracellular DA and DA metabolites during an acoustic test session of prepulse inhibition (PPI) of the startle response, a measure of sensorimotor gating, by microdialysis and HPLC-EC in a transgenic mouse model of schizophrenia. In th-fgfr1(tk-) mice, blockade of fibroblast growth factor receptor 1 (FGFR1) signaling during development in catecholaminergic neurons results in reduced size and density of midbrain DA neurons of the substantia nigra pars compacta (SNc) and ventral tegmental area (VTA). These mice displayed reduced PPI and enhanced startle response relative to control mice as well as a potentiation of DA release in the dorsal striatum during a 30 minute PPI test session. Acute administration of a partial α7 NNR agonist TC-7020 (1.0 mg/kg) normalized PPI and startle deficits and attenuated increases of DA release during acoustic PPI testing. These results provide direct evidence of elevated striatal dopaminergic transmission with impaired sensorimotor gating that may underlie cognitive and positive symptoms and motor deficits in schizophrenia and related disorders. Also, systemic targeting of alpha7 NNRs may ameliorate these deficits by functionally suppressing striatal DA activity.
Biological Psychiatry 04/2012; 136(1-3):82-7. · 8.28 Impact Factor
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T A Hauser,
A Kucinski,
K G Jordan,
G J Gatto,
S R Wersinger,
R A Hesse,
E K Stachowiak,
M K Stachowiak,
R L Papke,
P M Lippiello, M Bencherif
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ABSTRACT: A growing body of evidence suggests that the alpha7 neuronal nicotinic receptor (NNR) subtype is an important target for the development of novel therapies to treat schizophrenia, offering the possibility to address not only the positive but also the cognitive and negative symptoms associated with the disease. In order to probe the relationship of alpha7 function to relevant behavioral correlates we employed TC-5619, a novel selective agonist for the alpha7 NNR subtype. TC-5619 binds with very high affinity to the alpha7 subtype and is a potent full agonist. TC-5619 has little or no activity at other nicotinic receptors, including the alpha4beta2, ganglionic (alpha3beta4) and muscle subtypes. The transgenic th(tk-)/th(tk-) mouse model that reflects many of the developmental, anatomical, and multi-transmitter biochemical aspects of schizophrenia was used to assess the antipsychotic effects of TC-5619. In these mice TC-5619 acted both alone and synergistically with the antipsychotic clozapine to correct impaired pre-pulse inhibition (PPI) and social behavior which model positive and negative symptoms, respectively. Antipsychotic and cognitive effects of TC-5619 were also assessed in rats. Similar to the results in the transgenic mice, TC-5619 significantly reversed apomorphine-induced PPI deficits. In a novel object recognition paradigm in rats TC-5619 demonstrated long-lasting enhancement of memory over a wide dose range. These results suggest that alpha7-selective agonists such as TC-5619, either alone or in combination with antipsychotics, could offer a new approach to treating the constellation of symptoms associated with schizophrenia, including cognitive dysfunction.
Biochemical pharmacology 06/2009; 78(7):803-12. · 4.25 Impact Factor
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CNS Drug Reviews 09/2006; 3(4):325 - 345. · 4.92 Impact Factor
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ABSTRACT: TC-2559 [(E)-N-Methyl-4-[3-(5-ethoxypyridin)yl]-3-buten-1-amine] is a novel nicotinic agonist markedly more selective than recently reported novel nicotinic receptor ligands (selectivity ratio for central nervous system (CNS) to peripheral nervous system (PNS)>4000). TC-2559 competes effectively with [3H]-nicotine binding (K(i)=5 nM) but not with [125I]-bungarotoxin (>50,000 nM). Dopamine release from striatal synaptosomes and ion flux from thalamic synaptosomes indicate that TC-2559 is potent and efficacious in the activation of CNS receptors and significantly reduced glutamate-induced neurotoxicity in vitro. TC-2559 has no detectable effects on muscle and ganglion-type nicotinic acetylcholine receptors at concentrations up to 1 mM. TC-2559 significantly attenuates scopolamine-induced cognitive deficits in a step-through passive avoidance task. Acute and repeated oral dosing of TC-2559 enhances performance in a radial arm maze task. In contrast to the effects of equimolar concentrations of (-) nicotine, TC-2559 does not induce hypothermia and locomotor activity is not enhanced following repeated daily administration of 14 days. TC-2559 has a markedly enhanced CNS-PNS selectivity ratio and an intra-CNS selectivity as evidenced by the improved cognition without increased locomotor activity. The in vitro and in vivo studies in the present study suggest that TC-2559 has the desired profile to be further evaluated as a potential therapeutic agent for neurodegenerative diseases.
European Journal of Pharmacology 01/2001; 409(1):45-55. · 2.52 Impact Factor
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ABSTRACT: Human nicotinic acetylcholine (ACh) receptor subtypes expressed in Xenopus oocytes were characterized in terms of their activation by the experimental agonist RJR-2403. Responses to RJR-2403 were compared with those evoked by ACh and nicotine. These agonists were also characterized in terms of whether application of the drugs had the effect of producing a residual inhibition that was manifest as a decrease in subsequent control responses to ACh measured 5 min after the washout of the drug. For the activation of alpha4beta2 receptors, RJR-2403 had an efficacy equivalent to that of ACh and was more potent than ACh. RJR-2403 was less efficacious than ACh for other human receptor subtypes, suggesting that it is a partial agonist for all these receptors. Nicotine activated peak currents in human alpha4beta2 and alpha3beta2 receptors that were 85 and 50% of the respective ACh maximum responses. Nicotine was an efficacious activator of human alpha7 receptors, with a potency similar to ACh, whereas RJR-2403 had very low potency and efficacy for these receptors. At concentrations of <1 mM, RJR-2403 did not produce any residual inhibition of subsequent ACh responses for any receptor subtype. In contrast, nicotine produced profound residual inhibition of human alpha4beta2, alpha3beta2, and alpha7 receptors with IC(50) values of 150, 200, and 150 microM, respectively. Co-expression of the human alpha5 subunit with alpha3 and beta2 subunits had the effect of producing protracted responses to ACh and increasing residual inhibition by ACh and nicotine but not RJR-2403. In conclusion, our results, presented in the context of the complex pharmacology of nicotine for both activating and inhibiting neuronal nicotinic receptor subtypes, suggest that RJR-2403 will be a potent and relatively selective activator of human alpha4beta2 receptors.
Journal of Neurochemistry 07/2000; 75(1):204-16. · 4.06 Impact Factor
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ABSTRACT: Nicotinic acetylcholine receptors (nAChRs) exist as a diverse family of physiologically important ligand-gated ion channels active in classic, excitatory neurotransmission and perhaps in more novel forms of neurochemical signaling. Because of their critical functional roles centrally and peripherally, nAChRs are ideal targets for the regulation of nervous system function. nAChRs also are targets of nicotine, which acts acutely like acetylcholine to stimulate nAChR function. Here, we report studies using model cell culture systems testing the general hypothesis that more chronic nicotine exposure has unique effects on nAChRs. Chronic nicotine treatment induces increases in numbers of human muscle-type nAChRs containing alpha-1, beta-1, gamma and delta subunits, a human ganglionic nAChR subtype containing alpha-3 and beta-4 subunits and a human ganglionic nAChR containing alpha-7 subunits in intracellular and (except for alpha-7 nAChRs) in cell surface pools. However, the half-maximal potency with which nicotine has these effects differs across these nAChR subtypes, as do rates and magnitudes of the "nicotine-induced nAChR up-regulation." These changes in nAChR numbers are not attributable to either transient or sustained changes in nAChR subunit mRNA levels. Nicotine exposure more potently, more rapidly, and with nAChR-subtype specificity, induces two phases of losses in functional responsiveness of muscle-type nAChRs and alpha-3 beta-4 nAChRs, including a "persistent inactivation" that is distinct from classicly defined "desensitization." Based on these results, we hypothesize that chronic nicotine treatment induces persistent functional inactivation and numerical up-regulation of all nAChR subtypes via distinct post-transcriptional mechanisms and with potencies, at rates and with magnitudes that are nAChR-subtype specific. We also hypothesize that chronic nicotine exposure produces long-lasting changes in nervous system function, at least in part, by disabling rather than activating nicotinic cholinergic signaling.
Journal of Pharmacology and Experimental Therapeutics 09/1998; 286(2):825-40. · 3.83 Impact Factor
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ABSTRACT: The present report describes in vitro studies demonstrating that the heterocyclic substituted pyridine compound (+/-)-2-(3-pyridinyl)-1-azabicyclo[2.2.2]octane (RJR-2429) is extremely potent in activating human muscle nicotine ACh receptor (nAChR) (EC50 = 59 +/- 17 nM; Emax = 110 +/- 09% vs. nicotine). RJR-2429 is markedly less potent in activating nAChRs in the clonal cell line PC12, with EC50 = 1100 +/- 230 nM and Emax = 85 +/- 20% when compared with nicotine. The activation of a putative alpha 3 beta 4-containing nAChR in PC12 by RJR-2429 reveals a potency intermediate between nicotine and epibatidine (EC50 of 20,000 nM for nicotine and 30 nM for epibatidine). Dose-response curves for agonist-induced ileum contraction indicate that RJR-2429 is equipotent with nicotine, having an EC30 of approximately 2 microM. RJR-2429 binds with high affinity to alpha 4 beta 2 receptor subtype (Ki = 1.0 +/- 0.3 nM), and chronic exposure results in significant up-regulation of the high-affinity [3H]nicotine binding sites. In addition, RJR-2429 does not activate nAChRs present in rat thalamic preparations but is a potent inhibitor of this receptor subtype. It antagonizes nicotine-stimulated ion flux in thalamic synaptosomes with an IC50 of 154 +/- 37 nM. It also is a potent partial agonist at nAChRs mediating dopamine release from rat synaptosomal preparations (EC50 = 2 +/- 1 nM; Emax = 40%; epibatidine and nicotine have EC50 values of 0.4 and 100 nM, respectively). A model for the structure-activity profile of RJR-2429, nicotine and epibatidine was derived by molecular forcefield and quantum mechanics calculations and may provide important clues for the development of ligands selective for nAChR subtypes as probes in the life sciences or as potential therapeutic tools.
Journal of Pharmacology and Experimental Therapeutics 04/1998; 284(3):886-94. · 3.83 Impact Factor
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ABSTRACT: We have evaluated the physiological and behavioral effects of the CNS-selective nicotinic agonist (E)-N-methyl-4-(3-pyridinyl) -3-butene-1-amine (RJR-2403) using a number of different methods, including 1) reversal of pharmacologically induced amnesia in a step-through passive avoidance paradigm, 2) radial arm maze performance in rats with chemically induced brain lesions, 3) changes in HR and blood pressure in rats and 4) changes in body temperature, Y-maze activity, acoustic startle response and respiration in mice. Our results indicate that RJR-2403 is equal to or better than nicotine on measures of CNS function and cognitive enhancement. Specifically, RJR-2403 significantly improved passive avoidance retention after scopolamine-induced amnesia and enhanced both working and reference memory in rats with ibotenic acid lesions of the forebrain cholinergic projection system in an 8-arm radial maze paradigm. By comparison, RJR-2403 was 15 to 30-fold less potent than nicotine in decreasing body temperature, respiration, Y-maze rears and crosses and acoustic startle response. RJR-2403 also demonstrated greatly reduced cardiovascular effects. RJR-2403 was approximately 10-fold less potent than nicotine in increasing HR and 20-fold less potent in increasing blood pressure. These results are consistent with in vitro data indicating this compound's high selectivity for CNS nicotinic ACh receptor subtypes relative to peripheral ganglionic and muscle-type nicotinic ACh receptors. Therefore, RJR-2403 may be a valuable tool for understanding the central and peripheral pharmacology of nicotinic cholinergic systems as well as a potential lead compound for the development of nicotinic therapeutics to treat neurological diseases where cholinergic neurotransmission has been compromised.
Journal of Pharmacology and Experimental Therapeutics 01/1997; 279(3):1422-9. · 3.83 Impact Factor
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ABSTRACT: Increasing evidence for an involvement of nicotinic cholinergic systems in neurodegenerative disorders has stimulated the search for compounds with selectivity for CNS nicotinic ACh receptors (nAChRs). To this end, we have evaluated a number of nicotinic agonists for their ability to 1) bind to and up-regulate high-affinity nAChRs, 2) release [3H]-dopamine or induce 86Rb+ efflux in synaptosomes, 3) activate nAChRs in PC12 cells, 4) activate muscle-type nAChRs in human TE671/RD cells and 5) induce contraction of guinea pig ileum. Our results indicate that (E)-N-methyl-4-(3-pyridinyl)-3-butene-1-amine (RJR-2403) binds with high affinity to rat brain cortex (Ki = 26 +/- 3 nM). Functional studies show that RJR-2403 is comparable to nicotine in activating rat thalamic synaptosomes (EC50 = 732 +/- 155 nM and Emax = 91 +/- 8% for RJR-2403; EC50 = 591 +/- 120 nM and Emax = 100 +/- 25% for nicotine) but is one-tenth as potent in inducing dopamine release (EC50 = 938 +/- 172 nM and Emax = 82 +/- 5% for RJR-2403; EC50 = 100 +/- 25 nM and Emax = 100 +/- 13% for nicotine). At concentrations up to 1 mM, RJR-2403 does not significantly activate nAChRs in PC12 cells, muscle type nAChRs or muscarinic receptors. Dose-response curves for agonist-induced ileum contraction indicate that RJR-2403 is less than one-tenth as potent as nicotine with greatly reduced efficacy. RJR-2403 does not antagonize nicotine-stimulated muscle or ganglionic nAChR function (IC50 > 1 mM). Chronic exposure of M10 cells to RJR-2403 (10 microM) results in an up-regulation of high-affinity nAChRs phenomenologically similar to that seen with nicotine. These results suggest that RJR-2403 interacts with higher potency at CNS nAChR sub-types than at muscle, ganglionic or enteric nAChRs and has higher selectivity for CNS vs. muscle or ganglionic nAChRs than does nicotine.
Journal of Pharmacology and Experimental Therapeutics 12/1996; 279(3):1413-21. · 3.83 Impact Factor
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ABSTRACT: The agonist properties of acetylcholine (ACh), tetramethylammonium, ethyl-trimethylammonium and choline were evaluated for muscle and neuronal nicotinic receptors in Xenopus oocytes. The only essential feature for a neuronal receptor agonist appears to be the charged nitrogen. For specific receptor subtypes, other structural elements appear permissive (neither increasing nor decreasing activity) or non-permissive (decreasing activity). Choline was a full agonist for alpha 7, but a hydroxyl group was strongly non-permissive for other receptor subtypes (alpha 1 beta 1 gamma delta, alpha 3 beta 4, alpha 3 beta 2, and alpha 4 beta 2). The binding of these ligands to brain membranes is consistent with the electrophysiological results. Physiological concentrations of choline desensitize alpha 7 receptors to ACh suggesting that, in vivo, choline may regulate both the activation and inactivation of this receptor.
Neuroscience Letters 09/1996; 213(3):201-4. · 2.11 Impact Factor
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ABSTRACT: [3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester] (TMB-8) has seen wide use as an "intracellular Ca2+ antagonist." However, this study shows that TMB-8 acts as a noncompetitive, functional antagonist at diverse nicotinic acetylcholine receptor (nAChR) subtypes with potencies that exceed those for other reported effects of TMB-8, including inhibition of intracellular Ca2+ mobilization. TMB-8 is a potent inhibitor (IC50 approximately 400 nM) of agonist-stimulated ion flux mediated by functional human muscle nAChR or ganglionic alpha 3 beta 4-nAChR subtypes expressed by TE671/RD or SH-SY5Y cells. TMB-8 is also a potent inhibitor (IC50 approximately 500 nM) of a functional, central nervous system nAChR subtype that mediates nicotinic agonist-stimulated [3H]dopamine release from rat brain synaptosomes. TMB-8 is much less potent (IC50 approximately 30-200 microM) as an inhibitor of high-affinity 3H-labeled acetylcholine or 125I-labeled alpha-bungarotoxin binding to human muscle nAChR, ganglionic alpha 3 beta 4-nAChR, or ganglionic alpha 7-nAChR subtypes. Moreover, functional inhibition by TMB-8 of muscle-type nAChR is due to a reduction in agonist efficacy, but not potency, and is proportionately stronger with increasing agonist concentration, thereby suggesting that TMB-8 acts as a noncompetitive inhibitor. Similar effects are observed for local anesthetics such as tetracaine and procaine (functional IC50 values of approximately 5 and approximately 50 microM, respectively), although TMB-8 is the most potent of these agents. Studies with TMB-8 or BAPTA [1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid] analogues indicate that the amino group of TMB-8 is essential and that Ca2+ chelation is not required for inhibition of nAChR function.(ABSTRACT TRUNCATED AT 250 WORDS)
Journal of Pharmacology and Experimental Therapeutics 01/1996; 275(3):1418-26. · 3.83 Impact Factor
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ABSTRACT: Smokers are reported to have a higher density of central nicotinic acetylcholine receptors (nAChRs) that non-smokers at autopsy. Whether this increased receptor density is a response to smoking or a result of genetic variability is not known. While sub-chronic treatment of rats and mice with nicotine results in upregulation of central nAChRs, changes in receptor density in response to cigarette smoke have not been studied previously. In this study, male Sprague-Dawley rats were exposed nose-only for 13 weeks to mainstream cigarette smoke followed by assessment of [3H]nicotine binding in five brain regions of smoke- and sham-exposed animals. In smoke-exposed animals, there was a significant increase in nAChR density in the cortex, striatum, and cerebellum (35, 25, and 31% increases, respectively), while there was no significant change in receptor density in the thalamus and hippocampus. Smoke exposure did not alter markedly the affinity of the receptor for nicotine in these brain regions. Furthermore, up-regulation of nAChRs did not alter the biphasic binding properties by which nicotine binds to its receptor. There were no changes in the association (fast phase) or isomerization (slow phase) rate constants, and the percent contribution of slow and fast phase binding to nAChRs was not altered in the up-regulated receptor population compared with control. Similar results were observed following chronic nicotine exposure of cultured cortical cells from fetal rat brain or cells transfected with the alpha 4 beta 2 nAChR subtype. These results show that the up-regulation following smoke exposure in the rat is phenomenologically similar to that observed in vitro. These data provide preliminary evidence for a relationship between cigarette smoking and nAChR up-regulation in vivo and suggest that similar mechanisms of upregulation may underlie chronic smoke exposure of live animals and nicotine exposure of artificially expressed alpha 4 beta 2 receptors in vitro.
Biochemical Pharmacology 01/1996; 50(12):2001-8. · 4.70 Impact Factor
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ABSTRACT: There is a consensus that high-affinity [3H]-L-nicotine binding sites in the mammalian brain, which are thought to represent a predominant form of central nervous system nicotinic acetylcholine receptor (nAChR) composed of alpha 4 and beta 2 subunits, are increased in number after chronic nicotine exposure. However, mechanisms responsible for this effect have not yet been elucidated. To evaluate this issue, we have used, as models, primary cell cultures of fetal rat brain cortex, in which high-affinity [3H]-L-nicotine binding sites are naturally expressed, and clonal cell cultures of fibroblasts stably transfected to express nAChR composed of transgenic chick alpha 4 and beta 2 subunits under control of dexamethasone-inducible promoters. Chronic nicotine exposure induced an approximately 2.5-fold increase in high-affinity [3H]-L-nicotine binding sites in M10 cells maintained in the presence of dexamethasone or in primary fetal rat brain cortical cultures. Up-regulation of [3H]-L-nicotine binding sites was evident for M10 cells treated at nicotine concentrations as low as 10 nM (EC50 and EC100 values were 100 nM and 10 microM, respectively). Scatchard analyses of [3H]-L-nicotine binding data in M10 cells indicated a change in Bmax with no significant change in affinity for radioligand (KD = 2.5 +/- 0.5 nM in control cells vs. 2.0 +/- 0.4 nM in nicotine-treated cells). Northern blot analyses indicated that nicotine treatment alone had no direct effect on the promoter driving transgenic nAChR subunit gene transcription in M10 cells and that steady state levels of fetal rat brain cortical cell or M10 cell nAChR alpha 4 or beta 2 mRNAs were unaffected under conditions of chronic nicotine treatment that produced up-regulation of high-affinity [3H]-L-nicotine binding sites.(ABSTRACT TRUNCATED AT 250 WORDS)
Journal of Pharmacology and Experimental Therapeutics 12/1995; 275(2):987-94. · 3.83 Impact Factor
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ABSTRACT: Previous studies have shown that cells of the TE671/RD human clonal line express muscle-type nicotinic acetylcholine receptors (nAChR) and m3-type muscarinic acetylcholine receptors (mAChR) whose numbers and function are regulated by agonist treatment and second messenger modulation. Here we show that cytochalasin treatment, which causes disruption of actin networks, induces marked changes in the numbers and distribution of nAChR, but not mAChR. Moreover, whereas cytochalasin treatment fails to alter nAChR function significantly, it acutely potentiates mAChR-mediated phosphoinositide hydrolysis. Treatment of TE671/RD cells with different cytochalasin analogues (rank order efficacy at 5 micrograms/ml is H > J = B = C = D > A = E) produces a two- to fourfold increase in numbers of membrane-bound nAChR (Bmax in units of specific 125I-labeled alpha-bungarotoxin binding per milligram of membrane protein). nAChR up-regulation is evident after 1-2 days of cytochalasin B exposure, is maximal after 3-6 days of drug treatment, and is dominated by an approximately 10-fold increase (per cell) in an intracellular nAChR pool. Cytochalasin-induced nAChR up-regulation is similar in magnitude to, but not additive with, up-regulation of nAChR following chronic exposure to nicotine or phorbol ester. Northern blot analysis shows a four- to five-fold coordinate increase in levels of mRNA that encode nAChR alpha, beta, gamma, or delta subunits in cytochalasin-treated cells, suggesting that nAChR up-regulation has a possible transcriptional basis. Studies done using a 86Rb+ efflux assay indicate that cytochalasin treatment has no significant effect on nAChR function. By contrast, cytochalasin treatment has no effect on the numbers of mAChR as assessed by binding studies with the radioantagonist 3H-labeled quinuclidinyl benzilate, but it induces marked enhancement of carbachol-stimulated, but not basal, phosphoinositide hydrolysis. These studies suggest that presumed modulation by cytochalasin treatment of cytoskeletal microfilament integrity can differentially influence expression and function of mAChR (a prototype of the metabotropic receptor superfamily) and nAChR (a prototype of the ligand-gated ion-channel superfamily). The results also suggest possible new roles for the cytoskeleton in regulation of membrane receptor expression, function, and cross talk.
Journal of Neurochemistry 09/1993; 61(3):852-64. · 4.06 Impact Factor
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International Review of Neurobiology 02/1992; 34:25-131. · 2.35 Impact Factor
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ABSTRACT: Lithium ion, which inhibits hydrolytic degradation of inositol monophosphates, is the most common therapeutic agent used in the control of bipolar disorder. There exists evidence that elevated elemental vanadium levels may play an etiological role in at least some forms of manic-depression. Here we demonstrate that vanadate treatment of intact cells from several different clonal lines synergistically induces substantial augmentation in neurotransmitter receptor-mediated or growth factor receptor-triggered inositol trisphosphate accumulation in situ. Furthermore, studies done using cellular extracts indicate that effects of vanadate treatment in situ may be due to its ability to inhibit hydrolysis of inositol 1,4,5-trisphosphate inositol 1,3,4-trisphosphate, and inositol 1,3,4,5-tetrakisphosphate in vitro. These results suggest that vanadate treatment may facilitate characterization of inositol phosphate metabolism and intracellular signaling.
Neuroscience Letters 02/1992; 134(2):157-60. · 2.11 Impact Factor
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ABSTRACT: Exposure to sodium fluoride (NaF) resulted in an increased accumulation (up to 10-fold) of total [(3)H]inositol phosphates (T-InsP) in rat PC 12 cells. The magnitude of the NaF effect was comparable to that for muscarinic acetylcholine receptor-mediated stimulation of T-InsP accumulation in the presence of saturating concentrations of carbachol, but effects of NaF and muscarinic agonists were additive at subsaturating concentrations. The NaF effect was atropine insensitive; was not mimicked by effects of NaCl (10 mM), aluminum fluoride (1 to 100 muM), forskolin (up to 100 muM), or dibutyryl cyclic AMP (1 mM); and was not altered by treatment with pertussis or cholera toxins (1 mug/ml for 24 h). By contrast, the carbachol response was fully sensitive to atropine and partly sensitive to pertussis toxin. Chelation of extracellular calcium ion following 10 min of pretreatment with EDTA or EGTA (3 mM) inhibited carbachol-stimulated T-InsP accumulation by 50%, but resulted in an enhancement of NaF-stimulated T-InsP accumulation. By contrast, inhibition of the mobilization of intracellular calcium ion with 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate inhibited NaF stimulation of T-InsP accumulation by more than 50% but inhibited carbachol-stimulated TInsP accumulation to a much lower extent. Enhanced calcium influx and cell depolarization stimulated by high extracellular concentrations of KCl markedly potentiated carbachol, but not NaF, stimulation of T-InsP accumulation. This differential sensitivity to muscarinic antagonists, cell depolarization, and manipulation of intra- and extracellular calcium ion indicates that different mechanisms underly NaF and carbachol stimulation of T-InsP accumulation. However, stimulation of T-InsP accumulation in the presence of carbachol alone, NaF alone, or carbachol plus NaF was inhibited to a similar extent in the presence of the phorbol ester, phorbol 12-myristate13-acetate. Taken together, these observations suggest that NaF and carbachol effects are mediated through distinct mechanisms but share a common target, perhaps a GTP-binding protein and/or phospholipase C, whose activity is known to be influenced by protein kinase C.
Molecular and Cellular Neuroscience 10/1991; 2(5):377-83. · 3.66 Impact Factor
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ABSTRACT: Cells of the TE671/RD human clonal line express a finite number (Bmax) of about 350 fmol/mg of membrane protein) of apparently noninteracting, high-affinity binding sites (KD of 0.07 nM and a Hill coefficient close to unity, nH = 0.94) for the muscarinic acetylcholine receptor (mAChR) radio antagonist, tritium-labeled quinuclidinyl benzilate [( 3H]QNB). The rank order potency of selective antagonists that inhibit specific [3H]QNB binding is: atropine greater than 4-DAMP (4-diphenylacetoxy-N-methylpiperidine methiodide) greater than pirenzepine greater than methoctramine greater than AFDX-116 (11-2[[2-[(diethylamino)methyl]-1-[piperidinyl] acetyl]-5,11-dihydro-6H-pyrido[2,3-b][1,4]benzodiazepin-6-one). Functional studies indicate that phosphoinositide (PIns) hydrolysis in TE671/RD cells is increased by carbachol (EC50 of 10 microM), but not by nicotine (to concentrations as high as 1 mM). Agonist-stimulated PIns metabolism is inhibited by antagonists with the same rank order potency as for inhibition of [3H]QNB binding. Functional responses are augmented in the presence of a nonhydrolyzable GTP analog, are strongly inhibited after 24-hr exposure to cholera toxin, but are only slightly inhibited after long-term exposure to pertussis toxin or forskolin. These studies identify a pharmacologically-defined M3-subtype of mAChR strongly coupled via a cholera toxin-sensitive mechanism to PIns hydrolysis in these cells. Within 1 hr of treatment of TE671/RD cells with 1 mM dibutyryl cyclic AMP or with 10 microM phorbol-12-myristate-13-acetate (PMA), there is a 30 to 50% decrease in carbachol-stimulated PIns responsiveness that recovers to control values after 5 days of continued drug treatment. However, a comparable and more persistent inhibition of mAChR function is observed on cell treatment with 20 nM PMA.(ABSTRACT TRUNCATED AT 250 WORDS)
Journal of Pharmacology and Experimental Therapeutics 07/1991; 257(3):946-53. · 3.83 Impact Factor
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ABSTRACT: Effects of second messenger system modulation and sodium butyrate (NaBu) treatment on nicotinic acetylcholine receptor (nAChR) expression by cells of the TE671/RD human clone were established. Treatment with dibutyryl cyclic adenosine 3':5'-monophosphate (dbcAMP) or other substances that increase cellular cAMP content induces a 70% loss of nAChR per unit of membrane protein as assessed by binding studies using (125)I-labeled alpha-bungarotoxin (I-Bgt). By contrast, phorbol 12-myristate-13-acetate (PMA) treatment induces an initial 50% decrease, and then a later two- to threefold increase, in I-Bgt binding sites. These PMA effects are temporally distinct from a PMA treatment-induced 50% downregulation of membrane-bound phorbol ester binding sites, are blocked by treatment of cells with the putative protein kinase C inhibitors H7 or trifluoroperazine, and are sensitive to the protein synthesis inhibitor cycloheximide and the topoisomerase inhibitor novobiocin. Treatment with both PMA and dbcAMP induces a threefold increase in nAChR expression, whereas treatment with NaBu alone or with PMA induces an 80% decrease in I-Bgt binding site expression. All of these effects are dose and time dependent and reflect changes in the number of binding sites rather than changes in nAChR afnity for I-Bgt. These data indicate involvement of both cAMP and C-kinase pathways in the regulation of nAChR expression in ways that are not simply additive, possibly due to cross-talk between second messenger pathways. In addition, transcriptional and/or translational events are implicated in PMA and NaBu effects. The results indicate a multiplicity in the effects and mechanisms involved in regulation of nAChR expression.
Molecular and Cellular Neuroscience 02/1991; 2(1):52-65. · 3.66 Impact Factor
