Neurochemical Mechanisms in Disease P19 pps

Brain Edema in Neurological Disease 165 Rollins NK (2007) Clinical applications of diffusion tensor imaging and tractography in children. Pediatr Radiol 37:769–780 Romanic AM, White RF, Arleth AJ, Ohlstein EH, Barone FC (1998) Matrix metalloproteinase expression increases after cerebral focal ischemia in rats: inhibition of matrix metalloproteinase-9 reduces infarct size. Stroke 29:1020–1030 Rosenberg GA (1990) Brain fluids and metabolism. Oxford University Press, New York, NY Rosenberg GA (2002) Matrix metalloproteinases in neuroinflammation. Glia 39:279–291 Rosenberg GA (2009) Matrix metalloproteinases and their multiple roles in neurodegenerative diseases. Lancet Neurol 8:205–216 Rosenberg GA, Dencoff JE, Correa N Jr., Reiners M, Ford CC (1996b) Effect of steroids on CSF matrix metalloproteinases in multiple sclerosis: relation to blood-brain barrier injury. Neurology 46:1626–1632 Rosenberg GA, Estrada EY, Dencoff JE (1998) Matrix metalloproteinases and TIMPs are associated with blood-brain barrier opening after reperfusion in rat brain. Stroke 29: 2189–2195 Rosenberg GA, Estrada E, Kyner WT (1990) Vasopressin-induced brain edema is mediated by the V1 receptor. Adv Neurol 52:149–154 Rosenberg GA, Kyner WT, Estrada E (1980) Bulk flow of brain interstitial fluid under normal and hyperosmolar conditions. Am J Physiol 238:F42–F49 Rosenberg GA, Navratil M (1997) Metalloproteinase inhibition blocks edema in intracerebral hemorrhage in the rat. Neurology 48:921–926 Rosenberg GA, Navratil M, Barone F, Feuerstein G (1996a) Proteolytic cascade enzymes increase in focal cerebral ischemia in rat. J Cereb Blood Flow Metab 16:360–366 Rosenberg GA, Scremin O, Estrada E, Kyner WT (1992) Arginine vasopressin V1-antagonist and atrial natriuretic peptide reduce hemorrhagic brain edema in rats. Stroke 23: 1767–1774 Rosenberg GA, Yang Y (2007) Vasogenic edema due to tight junction disruption by matrix metalloproteinases in cerebral ischemia. Neurosurg Focus 22:E4 Roviezzo F, Tsigkos S, Kotanidou A, Bucci M, Brancaleone V, Cirino G, Papapetropoulos A (2005) Angiopoietin-2 causes inflammation in vivo by promoting vascular leakage. J Pharmacol Exp Ther 314:738–744 Schipke CG, Kettenmann H (2004) Astrocyte responses to neuronal activity. Glia 47: 226–232 Schleien CL, Eberle B, Shaffner DH, Koehler RC, Traystman RJ (1994) Reduced blood-brain barrier permeability after cardiac arrest by conjugated superoxide dismutase and catalase in piglets. Stroke 25:1830–1835 Schoch HJ, Fischer S, Marti HH (2002) Hypoxia-induced vascular endothelial growth factor expression causes vascular leakage in the brain. Brain 125:2549–2557 Schramm P, Schellinger PD, Klotz E, Kallenberg K, Fiebach JB, Kulkens S, Heiland S, Knauth M, Sartor K (2004) Comparison of perfusion computed tomography and computed tomography angiography source images with perfusion-weighted imaging and diffusion-weighted imaging in patients with acute stroke of less than 6 hours’ duration. Stroke 35:1652–1658 Shigemori Y, Katayama Y, Mori T, Maeda T, Kawamata T (2006) Matrix metalloproteinase-9 is associated with blood-brain barrier opening and brain edema formation after cortical contusion in rats. Acta Neurochir Suppl 96:130–133 Shuaib A, Xu WC, Yang T, Noor R (2002) Effects of nonpeptide V(1) vasopressin receptor antagonist SR-49059 on infarction volume and recovery of function in a focal embolic stroke model. Stroke 33:3033–3037 Sidel VW, Solomon AK (1957) Entrance of water into human red cells under an osmotic pressure gradient. J Gen Physiol 41:243–257 Simard JM, Chen M, Tarasov KV, Bhatta S, Ivanova S, Melnitchenko L, Tsymbalyuk N, West GA, Gerzanich V (2006) Newly expressed SUR1-regulated NC(Ca-ATP) channel mediates cerebral edema after ischemic stroke. Nat Med 12:433–440 166 E. Candelario-Jalil et al. Simard JM, Geng Z, Woo SK, Ivanova S, Tosun C, Melnichenko L, Gerzanich V (2009b) Glibenclamide reduces inflammation, vasogenic edema, and caspase-3 activation after sub- arachnoid hemorrhage. J Cereb Blood Flow Metab 29:317–330 Simard JM, Tsymbalyuk O, Ivanov A, Ivanova S, Bhatta S, Geng Z, Woo SK, Gerzanich V (2007) Endothelial sulfonylurea receptor 1-regulated NC Ca-ATP channels mediate progressive hemorrhagic necrosis following spinal cord injury. J Clin Invest 117:2105–2113 Simard JM, Woo SK, Bhatta S, Gerzanich V (2008) Drugs acting on SUR1 to treat CNS ischemia and trauma. Curr Opin Pharmacol 8:42–49 Simard JM, Yurovsky V, Tsymbalyuk N, Melnichenko L, Ivanova S, Gerzanich V (2009a) Protective effect of delayed treatment with low-dose glibenclamide in three models of ischemic stroke. Stroke 40:604–609 Sood R, Yang Y, Taheri S, Candelario-Jalil E, Estrada EY, Walker EJ, Thompson J, Rosenberg GA (2009) Increased apparent diffusion coefficients on MRI linked with matrix metalloproteinases and edema in white matter after bilateral carotid artery occlusion in rats. J Cereb Blood Flow Metab 29:308–316 Staub F, Baethmann A, Peters J, Weigt H, Kempski O (1990) Effects of lactacidosis on glial cell volume and viability. J Cereb Blood Flow Metab 10:866–876 Staub F, Winkler A, Peters J, Kempski O, Kachel V, Baethmann A (1994) Swelling, acidosis, and irreversible damage of glial cells from exposure to arachidonic acid in vitro. J Cereb Blood Flow Metab 14:1030–1039 Stoll G, Jander S (1999) The role of microglia and macrophages in the pathophysiology of the CNS. Prog Neurobiol 58:233–247 Suda S, Igarashi H, Arai Y, Andou J, Chishiki T, Katayama Y (2007) Effect of edaravone, a free radical scavenger, on ischemic cerebral edema assessed by magnetic resonance imaging. Neurol Med Chir (Tokyo) 47:197–202 Szmydynger-Chodobska J, Chung I, Kozniewska E, Tran B, Harrington FJ, Duncan JA, Chodobski A (2004) Increased expression of vasopressin v1a receptors after traumatic brain injury. J Neurotrauma 21:1090–1102 Taguchi Y, Takashima S, Hirano K, Dougu N, Toyoda S, Tanaka K (2007) Cerebral microbleeds identified by diffusion weighted MR imaging in a case of acute ischemic stroke. Intern Med 46:1275–1276 Tait MJ, Saadoun S, Bell BA, Papadopoulos MC (2008) Water movements in the brain: role of aquaporins. Trends Neurosci 31:37–43 Tang J, Liu J, Zhou C, Ostanin D, Grisham MB, Neil Granger D, Zhang JH (2005) Role of NADPH oxidase in the brain injury of intracerebral hemorrhage. J Neurochem 94:1342–1350 Terajima K, Igarashi H, Hirose M, Matsuzawa H, Nishizawa M, Nakada T (2008) Serial assess- ments of delayed encephalopathy after carbon monoxide poisoning using magnetic resonance spectroscopy and diffusion tensor imaging on 3.0T system. Eur Neurol 59:55–61 Thomas WE (1999) Brain macrophages: on the role of pericytes and perivascular cells. Brain Res Brain Res Rev 31:42–57 Thurston G (2002) Complementary actions of VEGF and angiopoietin-1 on blood vessel growth and leakage. J Anat 200:575–580 Trabold R , Krieg S, Scholler K, Plesnila N (2008) Role of vasopressin V(1a) and V(2) receptors for the development of secondary brain damage after traumatic brain injury in mice. J Neurotrauma 25:1459–1465 Trapp BD, Peterson J, Ransohoff RM, Rudick R, Mork S, Bo L (1998) Axonal transection in the lesions of multiple sclerosis. N Engl J Med 338:278–285 Trapp BD, Stys PK (2009) Virtual hypoxia and chronic necrosis of demyelinated axons in multiple sclerosis. Lancet Neurol 8:280–291 Unger E, Littlefield J, Gado M (1988) Water content and water structure in CT and MR signal changes: possible influence in detection of early stroke. AJNR Am J Neuroradiol 9:687–691 Vakili A, Kataoka H, Plesnila N (2005) Role of arginine vasopressin V1 and V2 receptors for brain damage after transient focal cerebral ischemia. J Cereb Blood Flow Metab 25:1012–1019 Brain Edema in Neurological Disease 167 Valable S, Montaner J, Bellail A, Berezowski V, Brillault J, Cecchelli R, Divoux D, Mackenzie ET, Bernaudin M, Roussel S, Petit E (2005) VEGF-induced BBB permeability is associated with an MMP-9 activity increase in cerebral ischemia: both effects decreased by Ang-1. J Cereb Blood Flow Metab 25:1491–1504 van Bruggen N, Thibodeaux H, Palmer JT, Lee WP, Fu L, Cairns B, Tumas D, Gerlai R, Williams SP, van Lookeren CM, Ferrara N (1999) VEGF antagonism reduces edema formation and tissue damage after ischemia/reperfusion injury in the mouse brain. J Clin Invest 104:1613–1620 Verkman AS (2005) More than just water channels: unexpected cellular roles of aquaporins. J Cell Sci 118:3225–3232 Verkman AS, Mitra AK (2000) Structure and function of aquaporin water channels. Am J Physiol Renal Physiol 278:F13–F28 Videen TO, Zazulia AR, Manno EM, Derdeyn CP, Adams RE, Diringer MN, Powers WJ (2001) Mannitol bolus preferentially shrinks non-infarcted brain in patients with ischemic stroke. Neurology 57:2120–2122 von Kummer R, Bourquain H, Bastianello S, Bozzao L, Manelfe C, Meier D, Hacke W (2001) Early prediction of irreversible brain damage after ischemic stroke at CT. Radiology 219: 95–100 Walder CE, Green SP, Darbonne WC, Mathias J, Rae J, Dinauer MC, Curnutte JT, Thomas GR (1997) Ischemic stroke injury is reduced in mice lacking a functional NADPH oxidase. Stroke 28:2252–2258 Wang W, Dentler WL, Borchardt RT (2001) VEGF increases BMEC monolayer permeability by affecting occludin expression and tight junction assembly. Am J Physiol Heart Circ Physiol 280:H434–H440 Wang X, Tsuji K, Lee SR, Ning M, Furie KL, Buchan AM, Lo EH (2004) Mechanisms of hemorrhagic transformation after tissue plasminogen activator reperfusion therapy for ischemic stroke. Stroke 35:2726–2730 Wang L, Zhang ZG, Zhang RL, Gregg SR, Hozeska-Solgot A, LeTourneau Y, Wang Y, Chopp M (2006) Matrix metalloproteinase 2 (MMP2) and MMP9 secreted by erythropoietin-activated endothelial cells promote neural progenitor cell migration. J Neurosci 26:5996–6003 Weed LH (1935) Certain anatomical and physiological aspects of the meninges and cerebrospinal fluid. Brain 58:383–397 Weed LH (1938) Meninges and Cerebrospinal Fluid. J Anat 72:181–215 Weisbrot-Lefkowitz M, Reuhl K, Perry B, Chan PH, Inouye M, Mirochnitchenko O (1998) Overexpression of human glutathione peroxidase protects transgenic mice against focal cerebral ischemia/reperfusion damage. Brain Res Mol Brain Res 53:333–338 Williams K, Alvarez X, Lackner AA (2001) Central nervous system perivascular cells are immunoregulatory cells that connect the CNS with the peripheral immune system. Glia 36:156–164 Winkler AS, Baethmann A, Peters J, Kempski O, Staub F (2000) Mechanisms of arachidonic acid induced glial swelling. Brain Res Mol Brain Res 76:419–423 Wintermark M et al (2008) Acute stroke imaging research roadmap. Stroke 39:1621–1628 Wolburg H, Lippoldt A (2002) Tight junctions of the blood-brain barrier: development, composi- tion and regulation. Vascul Pharmacol 38:323–337 Wong CH, Bozinovski S, Hertzog PJ, Hickey MJ, Crack PJ (2008) Absence of glutathione peroxidase-1 exacerbates cerebral ischemia-reperfusion injury by reducing post-ischemic microvascular perfusion. J Neurochem 107:241–252 Xia CF, Smith RS Jr., Shen B, Yang ZR, Borlongan CV, Chao L, Chao J (2006) Postischemic brain injury is exacerbated in mice lacking the kinin B2 receptor. Hypertension 47:752–761 Yamamoto N, Yokota K, Yamashita A, Oda M (1996) Effect of KBT-3022, a new cyclooxygenase inhibitor, on experimental brain edema in vitro and in vivo. Eur J Pharmacol 297: 225–231 Yancopoulos GD, Davis S, Gale NW, Rudge JS, Wiegand SJ, Holash J (2000) Vascular-specific growth factors and blood vessel formation. Nature 407:242–248 168 E. Candelario-Jalil et al. Yang G, Chan PH, Chen J, Carlson E, Chen SF, Weinstein P, Epstein CJ, Kamii H (1994) Human copper-zinc superoxide dismutase transgenic mice are highly resistant to reperfusion injury after focal cerebral ischemia. Stroke 25:165–170 Yang Y, Estrada EY, Thompson JF, Liu W, Rosenberg GA (2007) Matrix metalloproteinase- mediated disruption of tight junction proteins in cerebral vessels is reversed by synthetic matrix metalloproteinase inhibitor in focal ischemia in rat. J Cereb Blood Flow Metab 27:697–709 Yen MH, Lee SH (1987) Effects of cyclooxygenase and lipoxygenase inhibitors on cerebral edema induced by freezing lesions in rats. Eur J Pharmacol 144:369–373 Yong VW (2005) Metalloproteinases: mediators of pathology and regeneration in the CNS. Nat Rev Neurosci 6:931–944 Zador Z, Bloch O, Yao X, Manley GT (2007) Aquaporins: role in cerebral edema and brain water balance. Prog Brain Res 161:185–194 Zausinger S, Lumenta DB, Pruneau D, Schmid-Elsaesser R, Plesnila N, Baethmann A (2002) Effects of LF 16-0687 Ms, a bradykinin B(2) receptor antagonist, on brain edema formation and tissue damage in a rat model of temporary focal cerebral ischemia. Brain Res 950:268–278 Zeynalov E, Chen CH, Froehner SC, Adams ME, Ottersen OP, Amiry-Moghaddam M, Bhardwaj A (2008) The perivascular pool of aquaporin-4 mediates the effect of osmotherapy in postischemic cerebral edema. Crit Care Med 36:2634–2640 Zhang Z, Chopp M (2002) Vascular endothelial growth factor and angiopoietins in focal cerebral ischemia. Trends Cardiovasc Med 12:62–66 Zhang ZG, Zhang L, Jiang Q, Zhang R, Davies K, Powers C, Bruggen N, Chopp M (2000) VEGF enhances angiogenesis and promotes blood-brain barrier leakage in the ischemic brain. J Clin Invest 106:829–838 Zhang ZG, Zhang L, Tsang W, Soltanian-Zadeh H, Morris D, Zhang R, Goussev A, Powers C, Yeich T, Chopp M (2002) Correlation of VEGF and angiopoietin expression with disruption of blood-brain barrier and angiogenesis after focal cerebral ischemia. J Cereb Blood Flow Metab 22:379–392 Zhu Y, Lee C, Shen F, Du R, Young WL, Yang GY (2005) Angiopoietin-2 facilitates vascular endothelial growth factor-induced angiogenesis in the mature mouse brain. Stroke 36: 1533–1537 Zlokovic BV (2008) The blood-brain barrier in health and chronic neurodegenerative disorders. Neuron 57:178–201 Monoamine Transporter Pathologies Natalie R. Sealover and Eric L. Barker Abstract The monoamine neurotransmitters control a variety of functions including movement, appetite, mood, reward, and memory. The monoamine transporters are responsible for the termination of synaptic signaling by removing neurotransmitters from the synaptic cleft. Altered monoaminergic transporter function has been implicated in the pathology of disease states s uch as depression, anxiety, addiction, autism, Parkinson’s disease, and attention deficit hyperactivity disorder (ADHD). This review considers the mechanism of transporter action and regulation of transporter function. The implications of transporter polymorphisms are also addressed. Finally, a brief overview is presented that highlights important findings as well as existing problems that need to be considered in future studies. Keywords Dopamine transporter · Norepinephrine transporter · Serotonin transporter · Polymorphism · Attention deficit hyperactivity disorder · Parkinson’s disease · Addiction · Anxiety · Depression · Autism Abbreviations ADHD Attention deficit hyperactivity disorder β-CIT 2β-carbomethoxy-3β-(4-iodophenyl) tropane nor-β-CIT N-(2-fluoroethyl)-2β-carbomethoxy-3β-(4-iodophenyl)- nortropane DAT Dopamine transporter GAD Generalized anxiety disorder K m Substrate affinity LeuT Leucine transporter LeuT Aa Aquifex aeolicus leucine transporter E.L. Barker (B) Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, IN 47906, USA e-mail: barkerel@purdue.edu 169 J.P. Blass (ed.), Neurochemical Mechanisms in Disease, Advances in Neurobiology 1, DOI 10.1007/978-1-4419-7104-3_6, C  Springer Science+Business Media, LLC 2011 170 N.R. Sealover and E.L. Barker MAOI Monoamine oxidase inhibitor MAPK Mitogen-activated protein kinase MDMA, “ecstasy” 3,4-methylenedioxymethamphetamine MPP + 1-methyl-4-phenylpyridinium NET Norepinephrine transporter NSS Neurotransmitter/sodium symporter OCD Obsessive compulsive disorder PD Parkinson’s disease PKC Protein kinase C β-PMA Phorbol 12-myristate13-acetate PP2A Protein phosphatase 2A SERT Serotonin transporter SNP Single nucleotide polymorphism SPECT Single-photon emission computed tomography SSRI Selective serotonin reuptake inhibitor TAAR1 Trace amine-associated receptor 1 TCA Tricyclic antidepressant TMH Transmembrane helice VMAT Vesicular monoamine transporter V max Transport capacity VNTR Variable number tandem repeat Contents 1 Introduction to Monoamine Transporters 171 1.1 The Monoamine Transporter Family 171 1.2 Neuroanatomy 172 1.3 Physiological Functions 173 1.4 Structure and Transport Mechanism 173 1.5 Vesicular Monoamine Transporters 175 2 Regulation of Plasma Membrane Monoamine Transporters 175 3 Transporter Gene Polymorphisms 177 3.1 NET 177 3.2 DAT 178 3.3 SERT 179 4 Addiction 180 4.1 Psychostimulant Addiction 180 4.2 Alcoholism 181 5 Anxiety and Depression 182 6 Autism 184 7 Parkinson’s Disease (PD) 185 8 Important Findings and the Need for Future Studies 185 References 186 Monoamine Transporter Pathologies 171 1 Introduction to Monoamine Transporters 1.1 The Monoamine Transporter Family Synaptic transmission requires the release of neurotransmitters into the extracellular space to bind pre-or postsynaptic receptors, conveying a chemical message to nerve cells (Torres et al., 2003a). Termination of this signaling occurs rapidly by uptake of the released neurotransmitter into the presynaptic cell by high-affinity neurotransmitter transporters. The clearance of the monoamines dopamine, norepinephrine, and serotonin occurs via the dopamine transporter (DAT), norepinephrine transporter (NET), and serotonin transporter (SERT), respectively (Torres et al., 2003a) Fig. 1 General model of the release of vesicular neurotransmitter stores in response to cellular depolarization and the reuptake of the neurotransmitters by the monoamine transporters. Cytosolic neurotransmitters are taken into vesicles by VMAT and stored until the cell becomes depolarized, causing these vesicular stores to fuse with the plasma membrane and release the neurotransmitters into the synaptic cleft. Neurotransmitters in the synaptic cleft are available to bind pre- or postsynaptic receptors. Termination of signaling occurs when the neurotransmitters are taken back into the presynaptic cell by the monoamine transporters 172 N.R. Sealover and E.L. Barker (Fig. 1). These monoamine transporters belong to the SLC6 gene family of Na + -Cl – - coupled neurotransmitter transporters that is also referred to as the neurotransmitter sodium symporter (NSS) family (Chen et al., 2004). In addition to the monoamine transporters, the NSS family includes subfamilies of transporters for GABA, amino acids, creatine, and the osmolytes betaine and taurine (Chen et al., 2004). 1.2 Neuroanatomy In the brain, monoamine transporters are found on neurons that contain their respective neurotransmitter (Torres et al., 2003a). For example, neuronal cells that produce dopamine are localized in the substantia nigra, ventral tegmental area, and hypothalamus (Lin and Madras, 2006). The processes of dopaminergic neurons extend into the caudate nucleus, putamen, nucleus accumbens, and prefrontal cortex (Lin and Madras, 2006). Serotonergic neurons are located in the raphe nuclei of the brainstem and project into the cortex, thalamus, basal ganglia, hippocampus, and amygdala (Jacobs and Azmitia, 1992). Norepinephrine-producing neurons are found primarily in the locus coeruleus and raphe nuclei with moderate levels in the hypothalamus, midline thalamic nuclei, and the bed nucleus of the stria terminalis (Torres et al., 2003a; Donnan et al., 1991) (Fig. 2). Monoamine transporters are also located in peripheral areas of the body. Eisenhofer and colleagues demonstrated that DAT is present in the stomach, pan- creas, and kidney (Eisenhofer, 2001). NET is expressed in sympathetic peripheral neurons, the adrenal medulla, endothelial cells of the lung, and the placenta (Eisenhofer, 2001). SERT has been found in platelets (Talvenheimo and Rudnick, 1980), the intestinal tract (Wade et al., 1996), placenta (Padbury et al., 1997; Balkovetz et al., 1989), and in chromaffin cells of the adrenal gland (Schroeter et al., 1997). Reuptake by the monoamine transporters is the primary mechanism Fig. 2 (a) Location of serotonergic neurons and their projections in the human brain. (b) Location of noradrenergic neurons and their projections in the human brain. (c) Location of dopaminergic neurons and their projections in the human brain Monoamine Transporter Pathologies 173 of terminating monoaminergic neurotransmitter signaling in the central nervous system and periphery. 1.3 Physiological Functions The monoamine transporters are involved in the regulation of many physiological functions. DAT has been implicated in addiction and reward response, movement, cognition, and memory (Greengard, 2001). Altered dopaminergic regulation is involved in depression, suicide, anxiety, aggression, schizophrenia, attention defict hyperactivity disorder (ADHD), and Parkinson’s disease (Jayanthi and Ramamoorthy, 2005; Gainetdinov and Caron, 2003). SERT is involved in the regulation of appetite, libido, mood, anxiety, fear, reward, aggression, and memory (Barnes and Sharp, 1999). Disrupted serotonergic function has been implicated in depression, suicide, impulsive violence, autism, and alcoholism (Jayanthi and Ramamoorthy, 2005). NET plays an important role in arousal, mood, aggression, addiction, and attention, as well as in thermal and cardiac regulation (Jayanthi and Ramamoorthy, 2005; Howell and Kimmel, 2008). Alteration of the noradrengeric system can result in cardiac disease and psychiatric disorders including depression and anxiety (Jayanthi and Ramamoorthy, 2005). 1.4 Structure and Transport Mechanism The monoamine transporters contain 12 alpha helical transmembrane helices (TMHs) with a putative large extracellular loop between TMHs III and IV with potential glycosylation sites (Melikian et al., 1996, 1994) (Fig. 3). The amino and carboxy termini are located intracellularly and contain putative phosphorylation sites (Torres et al., 2003a). Uptake of the monoamines by their respective transporters utilizes an ion gradient generated by the plasma membrane Na + /K + ATPase (Torres et al., 2003a). NET and SERT are thought to translocate one Na + ion and one Cl – ion with the s ubstrate per transport cycle, whereas DAT is predicted to transport two Na + ions and one Cl – ion with its substrate (Torres et al., 2003a). The alternating access transport model has been used to describe the mechanism by which substrates are transported across the membrane via the monoamine transporters (Forrest et al., 2008). This model postulates that the transporter can exist in at least two conformations. These conformations include an extracellularly facing form that is open to the extracellular environment and can bind substrate and Na + and Cl – ions (Forrest et al., 2008). An intracellularly facing form allows the release of substrate into the cell and the binding of the countertransported K + ion to reverse the conformation of the transporter (Forrest et al., 2008). The alternating access model is supported by recent crystal structures of other transporters (Weyand et al., 2008; Faham et al., 2008). Two additional conformations of these transporters have also been described. A closed–closed conformation is predicted that prevents accessibility of substrate and ions from either side of the transporter. This closed– closed conformation was observed in the crystal structure of a leucine transporter 174 N.R. Sealover and E.L. Barker Fig. 3 Schematic representation of the predicted topology of the monoamine transporters based on the crystallization of LeuT Aa (Yamashita et al., 2005). The representation demonstrates how extracellular Na + ,Cl – , and substrate a re exchanged for intracellular K + . The putative phosphorylation sites on the N-terminus and C-terminus are shown along with predicted glycosylation sites between TMH III and TMH IV. This figure was adapted from Yamashita et al. (2005) from Aquifex aeolicus (LeuT Aa ), a bacterial homologue of the NSS transporter family (Yamashita et al., 2005). The closed–closed conformation has closed intra- and extracellular gates and may serve as an intermediate between the extracellularly and intracellularly facing states. Another conformation is predicted to have open intra- and extracellular gates. In this conformation the transporter is predicted to operate in a channel mode, allowing substrate molecules and ions to pass through the transporter quickly without an opening and closing of the gates for each transport cycle (Torres et al., 2003a). Comprehensive understanding of the mechanism of monoamine transport has been hampered by the lack of a crystal structure of these membrane transporters. As mentioned above, in 2005, LeuT Aa was crystallized (Yamashita et al., 2005). This structure and the cocrystallization of LeuT Aa with the tricyclic antidepressants (TCAs) have provided several clues about the putative structure of the monoamine transporters (Singh et al., 2007; Yamashita et al., 2005; Zhou et al., 2007). The LeuT Aa structures reveal binding sites for substrate and Na + ions located about halfway through the pore of the protein, interacting with TMHs III and VIII and the unwound regions of TMHs I and VI (Singh et al., 2007). The protein structure shows TMHs I through V are related to VI through X by a pseudo twofold axis in the membrane plane (Yamashita et al., 2005). Cocrystallization studies with the TCAs have identified a putative binding pocket in LeuT Aa that places the TCA binding . human brain. (c) Location of dopaminergic neurons and their projections in the human brain Monoamine Transporter Pathologies 173 of terminating monoaminergic neurotransmitter signaling in the central. Barker MAOI Monoamine oxidase inhibitor MAPK Mitogen-activated protein kinase MDMA, “ecstasy” 3,4-methylenedioxymethamphetamine MPP + 1-methyl-4-phenylpyridinium NET Norepinephrine transporter NSS. disorder PD Parkinson’s disease PKC Protein kinase C β-PMA Phorbol 12-myristate13-acetate PP2A Protein phosphatase 2A SERT Serotonin transporter SNP Single nucleotide polymorphism SPECT Single-photon