Abstract
This report further characterizes the intermediate metabolic effects of the psychotropic amphetamine derivative, 3,4-methylenedioxymethamphetamine (MDMA or “ecstasy”), on the activity of second messenger-dependent kinases. Previous work has demonstrated that two injections of MDMA (20 mg/kg) elicits a prolonged translocation of the calcium and phospholipid-dependent enzyme, protein kinase C (PKC) in rats. However, because MDMA has actions at the 5-HT transporter and 5-HT2A/2C receptors, our experiments were directed at uncovering which of these many sites may be involved in this second messenger-dependent response. A single injection of MDMA produced a time- and dose-dependent increase in the density of cortical and hippocampal PKC (as measured by 3H-phorbol 12,13-dibutyrate (PDBu)) binding sites. MDMA-mediated PKC translocation was long-lasting and remained above control (saline-treated rats) for up to 24 h after injection. This effect was mimicked by another substituted amphetamine, p-chloroamphetamine (pCA), but with a temporal-response curve that was to the left of MDMA’s. However, pure uptake inhibitors like fluoxetine, cocaine, and the selective 5-HT2A/2C agonist, DOB, were unable to produce a long-lasting translocation of PKC binding sites in rat cortex. Fluoxetine, a selective serotonin uptake inhibitor (SSRI) and ketanserin, a 5-HT2A antagonist, attenuated PKC translocation by MDMA with differing efficacies; however, both compounds completely prevented the loss of 5-HT uptake sties after multiple doses of MDMA. These results suggest that MDMA increases PKC translocation by two interrelated mechanisms that involve 5-HT2A/2C receptors and the 5-HT transporter. This pathway appears to include: (1) the drug binding to the 5-HT transporter, (2) the release of cytosolic 5-HT stores into the extracellular space, and (3) the activation of post-synaptic 5-HT2A/2C receptors linked to G-protein-mediated phospholipid hydrolysis.
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Appel NM, Contrera J, DeSouza EB (1989): Fenfluramine selectively and differentially decreases the density of serotonergic nerve terminals in rat brain: Evidence from immunocytochemical studies. J Pharmacol Exp Ther 249:928–943
Azmitia EC (1989): Nimodipine attenuates NMDA- and MDMA-induced toxicity of fetal serotonergic neurons: Evidence for a generic model of calcium toxicity. In Gispen WH, Traber J (eds), Nimodipine and Central Nervous System Functions: New Vistas. Stuttgart, Springer-Verlag, pp 141–159
Azmitia EC, Murphy RB, Whitaker-Azmitia PM (1990): MDMA (Ecstasy) effects on cultured serotonergic neurons: Evidence for Ca2+-dependent toxicity linked to release. Brain Res 510:97–103
Azmitia EC, Kramer HK, Kim-Park WK (1993): Nimodipine blocks the efflux of 45Ca2+ and enhances the depolarization-induced release of 3H-5-HT from CNS synaptosomes. In Scriabine A, Janis RA, Triggle DJ (eds), Drugs in Development. Branford, Neva Press, pp 437–446
Battaglia G, Yeh SY, O’Hearn E, Molliver ME, Kuhar MJ, DeSouza EB (1987): 3,4-Methylenedioxymethamphetamine and 3,4-methylenedioxyamphetamine destroy serotonin terminals in the rat brain: Quantification of neurodegeneration by measurement of [3H]paroxetine-labeled serotonin uptake sites. J Pharmacol Exp Ther 242:911–916
Battaglia G, Brooks BP, Kulsakdinun C, DeSouza EB (1988): Pharmacological profile of MDMA (3,4-methylenedioxymethamphetamine) at various brain recognition sites. Eur J Pharmacol 149:159–163
Berger UV, Grzanna R, Molliver ME (1989): Depletion of serotonin using p-chlorophenylalanine (PCPA) and reserpine protects against the neurotoxic effects of p-chloroamphetamine (PCA) in the brain. Exp Neurol 103:111–115
Berger UV, Grzanna R, Molliver ME (1992a): The neurotoxic effects of p-chloroamphetamine in rat brain are blocked by prior depletion of serotonin. Brain Res 578:177–185
Berger UV, Gu XF, Azmitia EC (1992b): The substituted amphetamines 3,4-methylenedioxymethamphetamine, methamphetamine, p-chloroamphetamine and fenfluramine induce 5-hydroxytryptamine release via a common mechanism blocked by fluoxetine and cocaine. Eur J Pharmacol 215:153–160
Berridge MJ (1984): Inositol triphosphate and diacylglycerol as second messengers. Biochem J 220:345–360
Broening HW, Bacon L, Slikker Jr. W (1994): Age modulates the long-term but not the acute effects of the serotonergic neurotoxicant 3,4-methylenedioxymethamphetamine. J Pharmacol Exp Ther 271:285–293
Castanga M, Takai Y, Kaibuchi K, Sano K, Kikkawa U, Nishizuka Y (1982): Direct activation of calcium-activated, phospholipid-dependent protein kinase by tumor-promoting phorbol esters. J Biol Chem 257:7847–7851
Cho AK, Hiramatsu M, DiStefano EW, Chang AS, Jenden DJ (1990): Stereochemical differences in the metabolism of 3,4-methylenedioxymethamphetamine in vivo and in vitro: A pharmacokinetic analysis. Drug Metab Disp 18:686–691
Commins DL, Vosmer G, Virus RM, Wollverton WL, Schuster CR, Seiden LS (1987): Biochemical and histological evidence that methylenedioxymethamphetamine (MDMA) is toxic to the rat brain. J Pharmacol Exp Ther 241:338–345
Conn PJ, Sanders-Bush E (1985): Serotonin-stimulated phosphoinositide turnover: Mediation by the S2 binding site in rat cerebral cortex but not in subcortical regions. J Pharmacol Exp Ther 234:195–203
Conn PJ, Sanders-Bush E (1986): Regulation of serotonin-stimulated phosphoinositide hydrolysis: Relation to the serotonin 5-HT2 binding site. J Neurosci 6:3669–3675
Edwards E, Ashby CR, Wang RY (1993): Further characterization of 5-HT and 5-HT3 receptor agonists-stimulated phosphoinositol phosphates accumulation. Brain Res 617:113–119
Farfel GM, Vosmer GL, Seiden LS (1992): The N-methyl-D-aspartate antagonist MK-801 protects against serotonin depletions induced by methamphetamine, 3,4-methylenedioxymethamphetamine and p-chloroamphetamine. Brain Res 595:121–128
Favaron M, Manev H, Alho H, Bertolino M, Ferret B, Guidotti A, Costa E (1988): Gangliosides prevent glutamate and kianate neurotoxicity in primary neuronal cultures of neonatal rat cerebellum and cortex. Proc Natl Acad Sci 85:7351–7355
Fischer C, Hatzidimitriou G, Wlos J, Katz J, Ricaurte G (1995): Reorganization of ascending 5-HT axon projections in animals previously exposed to the recreational drug 3,4-methylenedioxymethamphetamine (MDMA, “Ecstasy”). J Neurosci 15:5476–5485
Fisher JF, Cho AK (1979): Chemical release of dopamine from striatal homogenates: Evidence for an exchange diffusion model. J Pharmacol Exp Ther 208:203–209
Fuller RW, Perry KW, Molloy BB (1975): Reversible and irreversible phases of serotonin depletion by 4-chloroamphetamine. Eur J Pharmacol 33:119–124
Giambalvo CT (1992a): Protein kinase C and dopamine transport. 1. Effects of amphetamine in vivo. Neuropharmacology 31:1201–1210
Giambalvo CT (1992b): Protein kinase C and dopamine transport. 2. Effects of amphetamine in vitro. Neuropharmacology 31:1211–1222
Gott AL, Mallon BS, Paton A, Groome N, Rumsby MG (1994): Rat brain glial cells in primary culture and subculture contain the delta, epsilon, and zeta subspecies of protein kinase C as well as the conventional subspecies. Neurosci Lett 171:117–120
Gu XF, Azmitia EC (1993): Integrative transporter-mediated release from cytoplasmic and vesicular 5-hydroxytryptamine stores in cultured neurons. Eur J Pharmacol 235:51–57
Gudelsky GA, Yamamoto BK, Nash JF (1994): Potentiation of 3, 4-methylenedioxymethamphetamine-induced dopamine release and serotonin neurotoxicity by 5-HT2 agonists. Eur J Pharmacol 264:325–330
Harvey JA, McMaster SE, Yunger LM (1975): p-Chloroamphetamine: Selective neurotoxic action in the brain. Science 187:841–843
Hekmatpanah CR, Peroutka SJ (1990): 5-hydroxytryptamine uptake blockers attenuate the 5-hydroxytryptamine-releasing effect of 3,4-methyldioxymethamphetamine and related agents. Eur J Pharmacol 177:95–98
Hrdina PD, Vu TB (1993): Chronic fluoxetine treatment upregulates 5-HT uptake sites and 5-HT2 receptors in rat brain: An autoradiographic study. Synapse 14:324–331
Huff RA, Vaughan RA, Kuhar MJ, Uhl GR (1997): Phorbol esters increase dopamine transporter phosphorylation and decrease transport Vmax. J Neurochem 68:225–232
Hyde CE, Bennett BA (1994): Similar properties of fetal and adult amine transporters in the rat brain. Brain Res 646:118–123
Insel TR, Battaglia G, Johannessen JN, Marra S, DeSouza EB (1989): 3,4-methylenedioxymethamphetamine (“Ecstasy”) selectively destroys brain serotonin terminals in rhesus monkeys. J Pharmacol Exp Ther 249:713–720
Johnson MP, Hoffman AJ, Nichols DE (1986): Effects of the enantiomers of MDA, MDMA, and related analogues on [3H]serotonin and [3H]dopamine release from superfused rat brain slices. Eur J Pharmacol 132:269–276
Johnson MP, Conarty PF, Nichols DE (1991): [3H]Monoamine releasing and inhibition properties of 3,4-methylenedioxymethamphetamine and p-chloramphetamine analogues. Eur J Pharmacol 200:9–16
Kagaya A, Mikuni M, Kusumi I, Yamamoto H, Takahashi K (1990): Serotonin-induced acute desensitization of serotonin2 receptors in human platelets via a mechanism involving protein kinase C. J Pharmacol Exp Ther 255:305–311
Kendall DA, Nahorski SR (1985): 5-HT-stimulated inositol phospholipid hydrolysis in rat cerebral cortex slices: Pharmacological characterization and effects of antidepressants. J Pharmacol Exp Ther 233:473–479
Kimelberg HK (1986): Occurrence and functional significance of serotonin and catecholamine uptake by astrocytes. Biochem Pharmacol 35:2273–2281
Kokotos Leonardi ET, Azmitia EC (1994): MDMA (Ecstasy) inhibition of MAO type A and type B: Comparisons with fenfluramine and fluoxetine (Prozac). Neuropsychopharmacology 10:231–238
Kramer HK, Azmitia EC (1994): Is protein kinase C activation a key step in MDMA-induced toxicity of serotonergic neurons? CPDD Abstr 144:273
Kramer HK, Poblete JC, Azmitia EC (1995): 3,4-methylenedioxymethamphetamine (“Ecstasy”) promotes the translocation of protein kinase C (PKC): Requirement of viable serotonin nerve terminals. Brain Res 680:1–8
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951): Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275
Malberg JE, Malis RW, Sabol KE, Seiden LS (1994): Both ketanserin and alpha-methyl-para-tyrosine may protect against MDMA neurotoxicity by producing a hypothermic response in the rat. CPDD Abstr 56:213
Manev H, Costa E, Wroblewski JT, Guidotti A (1990): Abusive stimulation of excitatory amino acid receptors: A strategy to limit neurotoxicity. FASEB 4:2789–2797
Masliah E, Yoshida K, Shimohama S, Gage FH, Saitoh T (1991): Differential expression of protein kinase C isozymes in rat glial cell cultures. Brain Res 549:106–111
Melloni E, Pontremolli S, Michetti M, Sacco O, Sparatore B, Salamino F, Horrecker BL (1985): Binding of protein kinase C to neutrophil membranes in the presence of Ca2+ and its activation by a Ca2+-requiring proteinase. Proc Natl Acad Sci 82:6435–6439
Miller HH, Shore PA, Clarke DE (1980): In vivo monoamine oxidase inhibition by amphetamine. Biochem Pharmacol 29:1347–1354
Miller KJ, Hoffman BJ (1994): Adenosine A3 receptors regulate serotonin transport via nitric oxide and cGMP. J Biol Chem 269:27351–27356
Nash JF (1990): Ketanserin pretreatment attenuates MDMA-induced dopamine release in the striatum as measured by in vivo microdialysis. Life Sci 47:2401–2408
Nash JF, Brodkin J (1991): Microdialysis studies on 3,4-methylenedioxy-methamphetamine-induced dopamine release: Effect of dopamine uptake inhibitors. J Pharmacol Exp Ther 259:820–825
Nash JF, Arora RC, Schreiber MA, Meltzer HY (1991): Effect of 3,4-methylenedioxymethamphetamine on 3H-paroxetine binding in the frontal cortex and blood platelets of rats. Biochem Pharmacol 41:79–84
Nichols DE, Lloyd DH, Hoffman AJ, Nichols MB, Yim GK (1982): Effects of certain hallucinogenic amphetamine analogues on the release of [+3]H-serotonin from rat brain synaptosomes. J Med Chem 25:530–535
Nishizuka Y (1986): Studies and perspectives of protein kinase C. Science 233:305–312
Novotney S, Lowy MT (1995): Short-term and long-term effects of p-chloroamphetamine on hippocampal serotonin and corticosteroid receptor levels. Brain Res 684:19–25
O’Hearn E, Battaglia G, DeSouza EB, Kuhar MJ, Molliver ME (1988): Methylenedioxyamphetamine (MDA) and methylenedioxymethamphetamine (MDMA) cause selective ablation of serotonin axon terminals in forebrain: Immunocytochemical evidence for neurotoxicity. J Neurosci 8:2788–2803
Onodera H, Araki T, Kogyre K (1989): Protein kinase C activity in the rat hippocampus after forebrain ischemia: Autoradiographic analysis by 3Hphorbol 12,13 dibutyrate. Brain Res 481:1–7
Park WP, Azmitia EC (1991): 5-HT, MDMA (Ecstasy), and nimodipine effects on 45Ca2+-uptake into rat brain synaptosomes. Ann NY Acad Sci 635:438–442
Poblete JCP, Azmitia EC (1995): Activation of glycogen phosphorylase by 3,4-methylenedioxymethamphetamine in astrocyte-rich primary glial cultures: Role of the 5-HT2A receptor. Brain Res 680:9–16
Pranzatelli MR (1991): Regulation of 5-HT2 receptors in rat cortex. Studies with a putative selective agonist and antagonist. Biochem Pharmacol 42:1099–1105
Pu C, Vorhees CV (1995): Protective effects of MK-801 on methamphetamine-induced depletion of dopaminergic and serotonergic terminals and striatal astrocytic response: An immunohistochemical study. Synapse 19:97–104
Ricaurte GA, DeLanney LE, Weiner SG, Langston JW (1988): Toxic effect of MDMA on central serotonergic neurons in the primate: Importance of route and frequency of drug administration. Brain Res 446:165–168
Rudnick G, Wall SC (1992a): The molecular mechanism of “ecstasy” [3,4-methylenedioxymethamphetamine (MDMA)]: Serotonin transporters are targets for MDMA-induced serotonin release. Proc Natl Acad Sci 89:1817–1821
Rudnick G, Wall SC (1992b): p-chloroamphetamine induces serotonin release through serotonin transporters. Biochem 31:6710–6718
Sanders-Bush E, Strenaka LR (1978): Immediate and long-term effects of p-chloroamphetamine on brain amines. Ann NY Acad Sci 305:208–221
Schmidt CJ (1987): Neurotoxicity of the psychedelic amphetamine, methylenedioxymethamphetamine. J Pharmacol Exp Ther 240:1–7
Schmidt CJ, Taylor VL (1987): Depression of rat brain tryptophan hydroxylase activity following the acute administration of methylenedioxymethamphetamine. Biochem Pharmacol 36:4095–4102
Schmidt CJ, Black CK, Taylor VL (1990): Antagonism of the neurotoxicity due to the single administration of methylenedioxymethamphetamine. Eur J Pharmacol 181:59–70
Schmidt CJ, Taylor VL, Abbate GM, Nieduzak TR (1991): 5-HT2 antagonists stereoselectively prevent the neurotoxicity of 3,4-methylenedioxymethamphetamine by blocking the acute stimulation of dopamine synthesis: Reversal by I-dopa. J Pharmacol Exp Ther 256:230–235
Silver PJ, Sigg EB, Moyer JA (1986): Antidepressants and protein kinases: Inhibition of Ca2+-regulated myosin phosphorylation by fluoxetine and iprindole. Eur J Pharmacol 121:65–71
Steele TD, McAnn UD, Ricaurte GA (1994): 3,4-methylenedioxymethamphetamine (MDMA, “Ecstacy”): Pharmacology and toxicology in animals and humans. Addiction 89:539–551
Wang H-Y, Friedman E (1990): Central 5-hydroxytryptamine receptor-linked protein kinase C translocation: A functional postsynaptic signal transduction system. Mol Pharmacol 37:75–79
Wolf M, Cuatrecasas P, Sahyoun N (1985): Interaction of protein kinase C with membranes is regulated by Ca++, phorbol ester, and ATP. J Biol Chem 260:15718–15722
Yang CM, Yo YL, Hsieh JT, Ong R (1994): 5-Hydroxytryptamine receptor-mediated phosphoinositide Hydrolsysis in canine cultured tracheal smooth muscle cells. Br J Pharmacol 111:777–786
Yau JLW, Kelly PAT, Sharkey J, Seckl JR (1994): Chronic 3,4-methylenedioxymethamphetamine administration decreases glucocorticoid and mineralocorticoid receptor, but increases 5-HT-1C receptor gene expression in the rat hippocampus. Neuroscience 61:31–40
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Kramer, H., Poblete, J. & Azmitia, E. Activation of Protein Kinase C (PKC) by 3,4-Methylenedioxymethamphetamine (MDMA) Occurs Through the Stimulation of Serotonin Receptors and Transporter. Neuropsychopharmacol 17, 117–129 (1997). https://doi.org/10.1016/S0893-133X(97)00026-2
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DOI: https://doi.org/10.1016/S0893-133X(97)00026-2
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