Taurine prevents the neurotoxicity of -amyloid and glutamate receptor agonists: activation of GABA receptors and possible implications for Alzheimer’s disease and other neurological disorders PAULO ROBERTO LOUZADA, ANDRE´A C. PAULA LIMA, DAYDE L. MENDONC ¸ A-SILVA,* ¨ L,* FERNANDO G. DE MELLO,† AND SE´RGIO T. FERREIRA1 FRANC¸OIS NOE Departamento de Bioquı´mica Me´dica and *Departamento de Farmacologia Ba´sica e Clı´nica, Instituto de Cieˆncias Biome´dicas, †Instituto de Biofı´sica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-590, Brasil Alzheimer’s disease (AD) and several other neurological disorders have been linked to the overactivation of glutamatergic transmission and excitotoxicity as a common pathway of neuronal injury. The -amyloid peptide (A) is centrally related to the pathogenesis of AD, and previous reports have demonstrated that the blockade of glutamate receptors prevents A–induced neuronal death. We show that taurine, a -amino acid found at high concentrations in the brain, protects chick retinal neurons in culture against the neurotoxicity of A and glutamate receptor agonists. The protective effect of taurine is not mediated by interaction with glutamate receptors, as demonstrated by binding studies using radiolabeled glutamate receptor ligands. The neuroprotective action of taurine is blocked by picrotoxin, an antagonist of GABAA receptors. GABA and the GABAA receptor agonists phenobarbital and melatonin also protect neurons against A-induced neurotoxicity. These results suggest that activation of GABA receptors decreases neuronal vulnerability to excitotoxic damage and that pharmacological manipulation of the excitatory and inhibitory neurotransmitter tonus may protect neurons against a variety of insults. GABAergic transmission may represent a promising target for the treatment of AD and other neurological disorders in which excitotoxicity plays a relevant role.—Louzada, P. R., Lima, A. C. P., Mendonc¸a-Silva, D. L., Noe¨l, F., de Mello, F. G., Ferreira, S. T. Taurine prevents the neurotoxicity of -amyloid and glutamate receptor agonists: activation of GABA receptors and possible implications for Alzheimer’s disease and other neurological disorders. FASEB J. 18, 511–518 (2004) ABSTRACT
Key Words: amyloid 䡠 neuroprotection Excitotoxicity (the neuronal damage caused by overstimulation of excitatory receptors) has been implicated in several neurological disorders that affect millions of people worldwide (1, 2). Glutamate, the major excitatory neurotransmitter in the central ner0892-6638/04/0018-0511 © FASEB
vous system (CNS), plays important physiological roles in brain development and in processes such as learning, memory, sensory activity, movement control, and modulation of synaptic transmission (3). However, the overstimulation of both ionotropic and metabotropic glutamate receptors has been clearly implicated in the neuronal injury observed in several neurodegenerative disorders, including Alzheimer’s disease (AD), Huntington’s disease, amyotrophic lateral sclerosis, AIDS dementia complex, and Parkinson’s disease (1, 2, 4 –7). Other acute insults leading to massive brain cell death that have been related to excitatory imbalance include hypoglycemia, neurologic trauma, stroke, and epilepsy (1, 2). AD is histopathologically characterized by the deposition of a 39-43 amino acid residue peptide, known as -amyloid (A), in senile plaques in the brains of affected individuals (8). Early evidence of the involvement of A in the neuronal death in AD came from studies showing that A itself was neurotoxic in vitro (9) and in vivo (10). However, the mechanisms of cell death induced by A are not yet fully elucidated. The possible involvement of glutamate excitotoxicity in the pathophysiology of AD has been supported by a number of studies (for examples, see refs 11–16). Recently, we have shown that the blockade of ionotropic and/or metabotropic glutamate receptors prevents the neurotoxicity of A in retinal neuronal cultures (16). The retina has been used as a model system to investigate several aspects of neurochemical signaling in the CNS at the cellular level. Much is known about the morphology, ontogeny, biochemistry, and physiology of the retina. One of the most interesting features of this tissue is that its neurotransmitters participate in neural circuits that are entirely confined within the 1
Correspondence: Departamento de Bioquı´mica Me´dica, Instituto de Cieˆncias Biome´dicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-590, Brasil. E-mail:
[email protected] doi: 10.1096/fj.03-0739com 511
retina. Retinas obtained from chick embryos can be easily maintained in culture either as explants or as dissociated cells. Under these conditions, most neurochemical markers of tissue development and proper neurochemical communication are established among the different cell types. The retinas of different species have been used to study excitotoxicity mediated by excitatory amino acids. Recently, cultured retinal neurons have been shown to be highly sensitive to the neurotoxicity of A (16, 20). However, the mechanisms through which A mediates cell death still are not fully understood. Taurine (2-aminoethanesulfonic acid) is present at high concentrations in the mammalian brain (17), with several proposed roles in neurotransmission, neuromodulation, osmoregulation, control of calcium influx, and cell excitability. Early studies have shown that taurine increases Cl– conductance in excitable tissues and binds to GABAA receptors (18, 19). Here, we provide evidence supporting the notion that the neurotoxicity of A to retinal neurons is mediated by glutamate receptors. We also show for the first time that taurine prevents the neurotoxicity of A and that the neuroprotection is related to the activation of GABAA receptors. Similar neuroprotective actions of taurine were observed against the excitotoxicity of various glutamate receptor agonists. GABA and the pharmacological agonist phenobarbital had similar neuroprotective actions. These results suggest that selective pharmacological modulation of glutamate and GABA receptors could offer an interesting therapeutic approach in many neurological disorders, including AD.
MATERIALS AND METHODS Materials Kainic acid (KA) was purchased from Research Biochemicals International (Natick, MA, USA) and [3H]-MK-801, [3H]AMPA, and [3H]-Kainic acid were from New England Nuclear (Boston, MA, USA). Tris, PPO, POPOP, PMSF, EDTA, glutamate, NMDA, GABA, thiobarbituric acid, trypan blue, taurine, TFE, deferoxamine, BHT, and phenobarbital were from Sigma Chemical Co. (St. Louis, MO, USA). 〈 peptide (A1-42) was from Bachem Inc. (Torrance, CA, USA). All other reagents were of the highest analytical grade available. Retinal cultures Neuronal cultures of 8- or 9-day-old chick embryo retinas were used. 24-Well plates were incubated with poly-l-lysine hydrobromide (20 g/mL, 500 L/well) for 24 h. The wells were then washed three times with sterile distilled water (500 L) to remove excess polylysine. In each experiment, retinas were dissected under sterile conditions in a calcium/magnesium-free solution. Trypsinization (0.05% trypsin) was carried out for 10 min at 37°C in calcium/magnesium-free solution and the dissociated tissue was centrifuged for 2 min at 2000 rpm. The supernatant was discarded, and the pellet was carefully resuspended in BME supplemented with 5% (v/v) fetal calf serum and mechanically dissociated by gently pipetting the tissue 10 –20 times. Final cell concentration was ⬃106 512
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cells/mL for low-density cultures and 5 ⫻ 106 cells/mL for dense cultures. 500 L of the cell suspension was added to poly-l-lysine- wells. Cultures were kept at 37°C in a humidified atmosphere of 92% air/8% CO2 and the medium was changed 24 h after plating. For neuroprotection assays, taurine, phenobarbital, or GABA was added to the medium, as indicated in Results. After an incubation of 1 h, A was added to a final concentration of 44 M. Picrotoxin, when used, was added to the culture 1 h before other additions to ensure complete blockade of GABA receptors. Cell viability assay Cell survival was evaluated 48 h after various types of treatment of the cultures using the trypan blue exclusion method as described previously (16, 21). Binding assay Membrane preparations were obtained from retinal cells after 6, 24, 48, or 96 h in culture, as described in Results. Briefly, retinal cells were homogenized in ice-cold 50 mM Tris-acetate buffer containing 5 mM EDTA and 1 mM PMSF, pH 7.2, with a Dounce homogenizer at 4°C. The homogenate was centrifuged at 100,000 g for 1 h at 4°C. The pellet was resuspended in 50 mM Tris-acetate, pH 7.2, and stored at –70°C until use. Protein concentration was determined by the method of Bradford (22), using bovine serum albumin as standard. For kainate binding assays, membranes (80 –100 g of protein) were incubated for 1 h at 4°C in 0.5 mL of 50 mM Tris-acetate, pH 7.2, containing 5 nM [3H]-KA (58.0 Ci/ mmol). Bound and free [3H]-KA were separated by rapid filtration under vacuum on glass fiber filters (Whatman GF/C) followed by two 3 mL washes with ice-cold 50 mM Tris-acetate buffer. After drying, the filters were placed in vials containing a scintillation mixture (0.1 g/L POPOP and 4.0 g/L PPO, in toluene) and radioactivity was measured in a Tri-Carb Packard liquid scintillation counter. The specific binding was calculated by subtracting nonspecific binding, determined in the presence of 100 M unlabeled KA, from the total binding. Essentially the same protocol was used for NMDA receptor binding assays, except that 10 nM [3H]-MK801 (28.9 Ci/mmol), a noncompetitive NMDA antagonist, was used in 5 mM Tris-HCl buffer (pH 7.2). In this case, the filters were rinsed twice with 4 mL of ice-cold Tris-HCl buffer and the nonspecific binding was determined in the presence of 3 mM ketamine, a noncompetitive antagonist of the NMDA receptor. For AMPA receptor binding assays, the same protocol was used except that 5 nM [3H]-AMPA (40.6 Ci/mmol) was used in 50 mM Tris-acetate, 100 mM KSCN buffer (pH 7.2) and the filters were rinsed twice with 3 mL of ice-cold Tris-acetate/KSCN buffer. Nonspecific binding was determined in the presence of 1 mM l-glutamate. Amyloid aggregation measurements A aggregation was assayed by diluting A1-42 from a stock solution in 50% (v/v) trifluoroethanol (TFE) into PBS at a final concentration of 6.6 M peptide (0.75% residual TFE). When indicated, 5 mM taurine was also included in the PBS buffer. Right-angle light scattering measurements at 400 nm were carried out in an ISS PC1 spectrofluorometer (ISS Inc., Champaign, IL, USA). Aggregation was complete after 24 h incubation, as indicated by control measurements (data not shown). ThT fluorescence measurements were carried out in the same instrument using excitation at 450 nm and emission at 485 nm.
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Statistical analysis Values presented are means ⫾ se of at least three independent assays. Statistical significances were evaluated using unpaired Student’s t test.
RESULTS Figure 1 shows the effects of A on low-density chick embryo retinal cultures at different ages. Figure 1A shows a control 6-day-old (C6) neuronal culture. When A (44 M) was added on the fourth day of culture (C4) and maintained in the medium for 2 days, a marked neurotoxicity was apparent (Fig. 1B), with a sharp decrease in cell viability and extensive neurite retraction. By contrast, no neurotoxicity was observed when A was added to the culture 6 h after cell dissociation and maintained in the medium for 2 days (Fig. 1C, D). On the other hand, cultures to which A
Figure 1. Neurotoxicity of A and glutamate receptor agonists to chick embryo retinal neurons. A) Control 6-day-old (C6) retinal culture. B) 44 M A was added to a 4-day-old (C4) culture and kept in the medium 2 additional days. C) Control 2-day-old (C2) retinal culture. Note different overall morphology of the neurons, with fewer, shorter, and thinner neurites compared with a C6 culture (A). D) C2 culture to which 44 M A was added 6 h after trypsinization. E) Mean neuronal viabilities in cultures treated with A (44 M), glutamate (400 M), NMDA (400 M) or KA (400 M) at different times after trypsinization (gray bars: additions 6 h after trypsinization; black bars: additions at C4). Control cultures (at both C2 and C6) are shown as open bars. The bars represent means sd of 3 experiments. NEUROPROTECTION BY TAURINE
was added 6 h after cell dissociation and maintained in the medium for 6 days exhibited extensive neuronal death, similar to those to which A was added at C4 (Fig. 1E). These results show that neurons recently subjected to trypsin treatment for cell dissociation were not susceptible to A toxicity, whereas marked neurotoxicity was observed when aged neurons in culture (i.e., at C4) were exposed to A. Neuronal cultures treated with various glutamate receptor agonists (kainate, NMDA, and glutamate) showed a similar pattern of age-dependent sensitivity to excitotoxicity (Fig. 1E). The age-dependent sensitivity of retinal neurons to excitotoxic insults was further investigated by examining the expression of glutamate receptors on the cell surface as a function of time after trypsinization (Fig. 2). Very low kainate binding (⬃15 fmol/mg ptn) was observed in membrane preparations from cells that had recently been trypsinized. [3H]-Kainate binding increased as a function of time and approached saturation (⬃60 fmol/mg ptn) 48 –96 h after trypsinization. The parallelism between the onset of neurotoxicity and the increase in kainate binding (Figs. 1 and 2) suggests that cell surface expression of glutamate receptors is necessary for the neurotoxicity of A. Binding of radiolabeled AMPA and MK-801 to neuronal membranes were also examined. No difference was observed when the binding of MK-801 was measured immediately or 96 h after trypsinization (⬃220 fmol/mg ptn). This likely reflects the more deeply buried location of the binding site of MK-801 on the NMDA receptor, making it barely accessible or inaccessible to trypsin. Binding of AMPA was found to be uniformly low (⬃13 fmol/mg ptn) in all preparations examined, suggesting a very low expression of AMPA receptors in the avian retina. Figure 3 shows dense retinal cultures treated at C4 with A (44 M) and examined after 2 days of incubation. Remarkably, A was not toxic to dense cultures (Fig. 3A, B), in sharp contrast to the results described above for low-density cultures. However, dense cultures treated with D(⫹)-threo-3-hydroxyaspartic acid (DT) were very sensitive to the toxicity of A (Fig. 3C, D). DT is a potent competitive inhibitor of the uptake of l-glutamate by neurons and glia, leading to an increase in the extracellular glutamate pool (23). These results further implicate the excitotoxicity mediated by glutamate receptors in the process of cell death induced by A. Given the indication of the involvement of glutamatergic overactivation in A-induced neurotoxicity, we sought to counteract the neurotoxicity by activation of GABAergic neurotransmission. We initially investigated the effect of taurine, a known physiological GABA receptor agonist (19), in neuronal cultures. Remarkably, 100 M taurine strongly protected neurons against the neurotoxicity of A (Fig. 4A–D). Control cultures exhibited a mean viability of 78%, and the viability decreased to 26% in cultures treated with A vs. 67% in cultures treated with A in the presence of 100 M taurine (Fig. 4E). Taurine also protected retinal neurons against the toxicity of glutamate, 513
Figure 2. [3H]-Kainate binding to neuronal membranes. Membrane preparations were obtained from retinal cultures as described in Materials and Methods. [3H]-Kainate binding was measured at different times after trypsinization of retinal explants. Symbols correspond to means ⫾ sd of 2 independent experiments done in quadruplicate.
NMDA, and kainate (Fig. 4E). Further increases in the concentration of taurine (up to 1 mM) had no additional protective effect (data not shown). Control experiments showed that taurine (1 mM) did not interfere with binding of the glutamate receptor agonists AMPA, NMDA, and kainic acid to neuronal membrane preparations (data not shown). In addition, light scattering and thioflavine T fluorescence measurements showed that taurine did not interfere with A aggregation (data not shown). To determine whether the neuroprotection by taurine could be explained by its antioxidant properties or by activation of GABAA receptors (19), we examined the effects of picrotoxin, an antagonist of GABAA receptors, in retinal cultures. Picrotoxin abolished the neuroprotective action of taurine (Fig. 5). Picrotoxin
Figure 4. Taurine prevents the neurotoxicity of A and glutamate receptor agonists. A) Control C6 culture. B) Culture exposed to 44 M A at C4 and inspected at C6. C) Culture exposed to 44 M A ⫹ 100 M taurine (additions at C4 and inspection at C6). D) Culture exposed to 44 M A ⫹ 1 mM taurine. E) Mean cell viabilities under different conditions. The bars represent means ⫾ sd of 3 independent experiments done in triplicate. The concentrations of A, glutamate, KA, NMDA, and taurine were 44 M, 600 M, 800 M, 100 M, and 100 M, respectively.
also blocked the neuroprotective action of taurine against the excitotoxicity of kainic acid (data not shown). These results suggest the participation of GABAA receptors in the protection against neuronal injury induced by both A and kainate. Similar neuroprotective actions were observed when retinal neurons were challenged with A in the presence of GABA or the GABA agonists phenobarbital (Fig. 6) and melatonin (data not shown).
DISCUSSION
Figure 3. Neurotoxicity of A in dense neuronal cultures. A) Control C6 dense retinal culture (5⫻106 cells). B) 44 M A was added at C4 and kept in the medium for 48 h. C) Control C6 dense retinal culture maintained in the presence of 25 M D-(⫹)-threo-3-hydroxyaspartic acid (DT). D) C4 dense retinal culture exposed to 44 M A for 48 h in the presence of 25 M DT. 514
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In this work, we describe for the first time the neuroprotective action of the -amino acid taurine against the neurotoxicity of A. Pharmacological characterization of this effect indicated a major role of the activation of GABAA receptors in the mechanism of neuroprotection. Taurine is involved in several physiological actions in the brain, including osmoregulation (24) and neurotransmission (25). Taurine has been shown to protect against the excitotoxicity of glutamate in other cellular
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Figure 5. Picrotoxin blocks the neuroprotective action of taurine. Cell viabilities were evaluated at C6. A) Control culture. B) Culture exposed to 44 M A at C4. C) culture exposed to A ⫹ 1mM taurine at C4. D) Culture exposed to A ⫹ taurine ⫹ 50 M picrotoxin at C4. E) Mean cell viabilities. The results are means ⫾ se of 4 independent experiments.
models, although the mechanisms of such protection have not been fully elucidated. For example, a decrease in calcium overload in cerebellar granule and hippocampal cells has been implicated in the protective effects of taurine against glutamate toxicity (26, 27). By contrast, Lima et al. (28) suggested that taurine improves neurite outgrowth in the goldfish retina by increasing calcium influx into the cell. The trophic properties of taurine in the retina appear to be linked to the modulation of protein phosphorylation (29). Induction of protein phosphorylation by taurine has also been demonstrated in the brain and heart (30). The release of taurine in hippocampal slices is maximized under conditions associated with neuronal damage, such as ischemia (31), suggesting a possible pathophysiological role of taurine in those conditions. The well-accepted notion that overactivation of glutamatergic transmission could mediate cell death in several neurological diseases (1, 2) has prompted considerable effort to develop drugs to attenuate or abolish the excitotoxicity. Excitotoxicity has been related to a host of chronic and acute neuronal insults, including Parkinson’s, Huntington’s and Alzheimer’s diseases, AIDS dementia complex, brain trauma, stroke (1, 2, 3–7), and spinal cord injury (32). Our observation that NEUROPROTECTION BY TAURINE
soon after trypsinization, retinal neurons are resistant to the neurotoxic actions of both A and glutamatergic agents (KA, NMDA, and glutamate) whereas aged neuronal cultures (i.e., on or after C4) are very sensitive to those agents (Fig. 1), suggests that expression of functional receptors in the cell membrane is required for neurotoxicity. Indeed, cells at the early stages of culture showed reduced [3H]-KA binding; increased binding was observed after 48 –96 h in culture (Fig. 2). Therefore, the onset of the toxicity elicited by EAAs and A is coincident with the expression of glutamate receptors at the cell surface after trypsinization. The effects of A on dense, mixed retinal cultures containing neurons and glia were quite different from those observed in low-density, primarily neuronal cultures. In dense cultures, neurons were resistant to A toxicity (Fig. 3), a phenomenon observed with other EAAR agonists (33). The resistance of dense cultures to A is probably related to the efficient uptake of glutamate by Mu¨ ller cells (retinal glia), which are present in large numbers in dense cultures. In the presence of the inhibitor of glutamate uptake, DT, A was highly toxic even to dense neuronal cultures, indicating that accumulation of extracellular glutamate is necessary for A neurotoxicity. The fact that most, if not all, of the toxic effects of A appear to be mediated by glutamate excitotoxicity suggests that blockade of glutamate receptors could be considered a neuroprotective strategy. However, longterm blockade of glutamatergic transmission has been shown to decrease cell viability in mature brains undergoing slowly progressing neurodegeneration (34). NMDA receptor inhibition by MK-801 has been shown to cause necrotic neuronal death (35, 36). Furthermore, phencyclidine and ketamine, other NMDA antagonists, were shown to cause a schizophrenic-like psychosis (37, 38). This seems to be because controlled glutamatergic transmission is necessary for the proper availability of trophic factors important in maintaining neuronal health (3). Moreover, pharmacological inhi-
Figure 6. GABA receptor agonists protect retinal neurons against the neurotoxicity of A. Cultures were maintained in BME plus 5% fetal calf serum and cell viabilities were evaluated at C6. A (44 M) was added at C4. The results are means ⫾ se of 3 independent experiments. Concentrations of phenobarbital and GABA were 16 g/mL and 3 M, respectively. 515
bition of EAARs is not selective and the use of EAAR antagonists may affect indiscriminately various brain circuits (39). Taurine has been shown to be neuroprotective against the excitotoxicity of glutamate (27). This effect, however, is not mediated by blockade of glutamate receptors (data not shown). Taurine potently protected neurons in culture against the toxicity of A, glutamate, kainate, and NMDA (Fig. 4). Oxidative stress and lipid peroxidation have been implicated in glutamate- and A-induced neurotoxicity (for recent examples, see refs 40, 41). On the other hand, Tadolini et al. (42) have shown that taurine and hipotaurine decrease lipid peroxidation in liposomes. This raised the possibility that the neuroprotective action of taurine could be related to its antioxidant properties. Indeed, we found that taurine blocked lipid peroxidation induced by A and by glutamate receptor agonists in retinal cell cultures (data not shown). However, the fact that picrotoxin abolished the protective action of taurine (Fig. 5) clearly indicates that neuroprotection is in fact mediated by interaction with GABAA receptors and is not related to the antioxidant properties of taurine. This conclusion is reinforced by the fact that GABA and phenobarbital protected neurons from the neurotoxic effects of A (Fig. 6) and that their effects were abolished by picrotoxin (data not shown). These results are also in line with previous report showing that taurine activates GABAA receptors in rat hippocampal slices (19). We have recently shown that the neuroprotective action of melatonin against the toxicity of A is partly blocked by picrotoxin (43). This suggests that, in addition to the well-known antioxidant properties of melatonin, its protective action is partly mediated by activation of GABAA receptors. The notion that GABAA receptor activation by GABA may be associated with neuroprotection has been proposed (44, 45). Moreover, activation of GABAergic transmission has also been proposed as a possible treatment for acute ischemic stroke (46, 47). This agrees with the available evidence indicating that anticonvulsants such as phenobarbital (48) counteract the excitotoxicity of glutamatergic agonists. Furthermore, GABA agonists carbamazepine, phenytoin, and valproic acid have been shown to attenuate the neurotoxic effects of A by stabilization of intracellular calcium levels (49). A possible caveat in the use of the latter compounds results from their side effects. A clear advantage of the use of taurine as a GABA agonist resides in its safety and tolerability. In humans, it has been reported that taurine in amounts as high as 2 g per day is well tolerated, with no signs of side effects (50). The use of up to 12 g of taurine per day, as an adjunct therapy for hepatic disease was also well tolerated by most patients (51). The use of taurine as a possible therapeutic agent in several non-neurological disorders has been proposed (50). The fact that taurine crosses the blood– brain barrier (52, 53) opens the possibility of using taurine in the therapy of neurological diseases. Increasing taurinergic tonus using drugs 516
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such as lamotrigine, which has been shown to elevate taurine levels in the CNS (54), may also constitute an alternative clinical approach to neuroprotection. Rodrigues et al. (55) recently suggested a protective role of the bile salt tauroursodeoxycholate against lipid peroxidation induced by A. O’Byrne and Tipton (56) reported a protective effect of taurine against the neurotoxicity induced by MPTP. Taken together, these previous findings and our present results suggest that taurine should be considered as a possible therapeutic tool to treat AD and other neurological disorders characterized by overstimulation of glutamatergic transmission. This possibility is reinforced by an early study by Pomara et al. (57), who reported low levels of taurine and high levels of glutamate in the brain of AD patients. This could mean that at least part of the AD pathophysiology could result from a chronic imbalance between taurinergic and glutamatergic tonus in the CNS. To our knowledge, no clinical trial so far has aimed at investigating the efficacy of taurine in the treatment of AD patients. Based on the present results, we suggest that taurine or related compounds with GABAergic activity should be explored as neuroprotectants against the toxicity of A and other neurological insults characterized by hyperactivation of glutamatergic transmission. Supported by grants from Howard Hughes Medical Institute, the John Simon Guggenheim Memorial Foundation, Conselho Nacional de Desenvolvimento Cientı´fico e Tecnolo´gico, and Fundac¸ a˜ o de Amparo a` Pesquisa do Estado do Rio de Janeiro. S.T.F. is a Howard Hughes Medical Institute International Scholar.
REFERENCES 1. 2. 3. 4.
5. 6.
7. 8. 9.
Lipton, S. A., and Rosenberg, P. A. (1994) Excitatory amino acids as a final common pathway for neurologic disorders. N. Engl. J. Med. 330, 613– 622 Mattson, M. (2003) Excitotoxic and excitoprotective mechanisms: abundant targets for the prevention and treatment of neurodegenerative disorders. Neuromolec. Med. 3, 65–94 Gasic, G. P., and Hollmann, M. (1992) Molecular neurobiology of glutamate receptors. Annu. Rev. Physiol. 54, 507–536 Behrens, P. F., Franz, P., Woodman, B., Lindenberg, K. S., and Landwehrmeyer, G. B. (2002) Impaired glutamate transport and glutamate-glutamine cycling: downstream effects of the Huntington mutation. Brain 125, 1908 –1922 Cluskey, S., and Ramsden, D. B. (2001) Mechanisms of neurodegeneration in amyotrophic lateral sclerosis. J. Clin. Pathol. 54, 386 –392 Kieburtz, K. D., Epstein, L. G., Gelbard, H. A., and Greenamyre, J. T. (1991) Excitotoxicity and dopaminergic dysfunction in the acquired immunodeficiency syndrome dementia complex. Therapeutic implications. Arch. Neurol. 48, 1281–1284 Difazio, M. C., Hollingsworth, Z., Young, A. B., and Penney, J. B., Jr. (1992) Glutamate receptors in the substantia nigra of Parkinson’s disease brains. Neurology 42, 402– 406 Selkoe, D. J. (1999) Translating cell biology into therapeutic advances in Alzheimer’s disease. Nature (London) 399, A23– A31 Pike, C. J., Walencewicks, A. J., Glabe, C. G., and Cotman, C. W. (1991) In vitro aging of -amyloid causes peptide aggregation and neurotoxicity. Brain Res. 563, 311–314
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10. 11.
12.
13.
14. 15.
16.
17. 18. 19. 20.
21.
22. 23.
24.
25. 26.
27. 28. 29. 30.
Kowall, N. W., McKee, A. C., Yankner, B. A., and Beal, M. F. (1992) In vivo neurotoxicity of -amyloid [ (1-40)] and the  (25-35) fragment. Neurobiol. Aging 13, 537–542 Mattson, M. P., Cheng, B., Davis, D., Bryant, K., Lieberburg, I., and Ryde, R. E. (1992) -Amyloid peptides destabilize calcium homeostasis and render human cortical neurons vulnerable to excitotoxicity. J. Neurosci. 12, 376 –389 Launderback, C. M., Harris-White, M. E., Wang, Y., Pedigo, N. W., Jr., Carney, J. M., and Butterfield, D. A. (1999) Amyloid beta peptide inhibits Na⫹-dependent glutamate uptake. Life Sci. 650, 1977–1981 Arias, C., Arrieta, I., and Tapia, R. (1995) -Amyloid peptide fragment 25-35 potentiates the calcium-dependent release of excitatory amino acids from depolarized hippocampal slices. J. Neurosci. Res. 41, 561–566 Noda, N., Nakanishi, H., and Akaike, N. (1999) Glutamate release from microglia via glutamate transporter is enhanced by amyloid-beta peptide. Neuroscience 92, 1465–1474 Harkany, T., Abraham, I., Timmerman, W., Laskay, G., Toth, B., Sasvari, M., Konya, C., Sebens, J. B., Korf, J., Kyakas, C., et al. (2000) -Amyloid neurotoxicity is mediated by a glutamatetriggered excitotoxic cascade in rat nucleus basalis. Eur. J. Neurosci. 12, 2735–2745 Louzada, P. R., Lima, A. C. P., Mello, F. G., and Ferreira, S. T. (2001) Dual role of glutamatergic transmission on amyloid 1-42 aggregation and neurotoxicity in embryonic avian retina. Neurosci. Lett. 301, 59 – 63 Guidotti, A., Badiani, G., and Pepeu, G. (1972) Taurine distribution in cat brain. J. Neurochem. 19, 431– 435 Okamoto, K., Kimura, H., and Sakai, Y. (1983) Taurine-induced increase of the Cl-conductance of cerebellar Purkinje cell dendrites in vitro. Brain Res. 259, 319 –323 del Olmo, N., Bustamante, J., del Rı´o, R. M., and Solis, J. M. (2000) Taurine activates GABA(A) but not GABA(B) receptors in rat hippocampal CA1 area. Brain Res. 864, 298 –307 Alvarez, A., Alarco´ n, R., Opazo, C., Campos, E. O., Mun ˜ oz, F. J., Caldero´n, F. H., Dajas, F., Gentry, M. K., Doctor, B. P., Mello, F. G., et al. (1998) Stable complexes involving acetylcholinesterase and amyloid- peptide change the biochemical properties of the enzyme and increase the neurotoxicity of Alzheimer’s fibrils. J. Neurosci. 18, 3213–3223 De Felice, F. G., Houzel, J. C., Garcia-Abreu, J., Louzada, P. R., Jr., Afonso, R. C., Meirelles, M. N., Neto, V. M., and Ferreira, S. T. (2001) Inhibition of Alzheimer’s disease beta-amyloid aggregation, neurotoxicity, and in vivo deposition by nitrophenols: implications for Alzheimer’s therapy. FASEB J. 7, 1297– 1299 Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248 –254 Johnston, G. A., Lodge, D., Bornstein, J. C., and Curtis, D. R. (1980) Potentiation of l-glutamate and l-aspartate excitation of cat spinal neurones by the stereoisomers of threo-3 hydroxyaspartate. J. Neurochem. 34, 241–243 Nagelhus, E. A., Lehmann, A., and Ottersen, O. P. (1993) Neuronal-glial exchange of taurine during hypo-osmotic stress: a combined immunocytochemical and biochemical analysis in rat cerebellar cortex. Neuroscience 54, 615– 631 Mc Bride, W. J., and Frederickson, R. C. (1980) Taurine as a possible inhibitory transmitter in the cerebellum. Federation Proc. 39, 2701–2705 Zhao, P., Huang, Y. L., and Cheng, J. S. (1999) Taurine antagonizes calcium overload induced by glutamate or chemical hypoxia in cultured rat hippocampal neurons. Neurosci. Lett. 268, 25–28 El Idrissi, A., and Trenkner, E. (1999) Growth factors and taurine protect against excitotoxicity by stabilizing calcium homeostasis and energy metabolism. J. Neurosci. 19, 9459 –9468 Lima, L., Matus, P., and Drujan, B. (1993) Taurine-induced regeneration of goldfish retina in culture may involve a calcium mediated mechanism. J. Neurochem. 60, 2153–2158 Lima, L., and Cubillos, S. (1998) Taurine might be acting as a trophic factor in the retina by modulating phosphorylation of cellular proteins. J. Neurosci. Res 53, 377–384 Lombardini, J. B., and Props, C. (1996) Effects of kinase inhibitors and taurine analogues on the phosphorylation of
NEUROPROTECTION BY TAURINE
31. 32. 33.
34. 35.
36.
37.
38. 39. 40.
41.
42. 43.
44.
45.
46.
47.
48.
49.
50.
specific proteins in mitochondrial fractions of rat heart and retina. Adv. Exp. Med. Biol. 403, 343–350 Saransaari, P., and Oja, S. S. (1997) Enhanced taurine release in cell-damaging conditions in the developing and ageing mouse hippocampus. Neuroscience 79, 847– 854 Mills, C. D., Johnson, K. M., and Hulsebosch, C. E. (2001) Role of group II and group III metabotropic glutamate receptors in spinal cord injury. Exp. Neurol. 173, 153–167 Loureiro-Dos-Santos, N. E., Reis, R. A., Kubrusly, R. C., de Almeida, O. M., Gardino, P. F., de Mello, M. C., and de Mello, F. G. (2001) Inhibition of choline acetyltransferase by excitatory amino acids as a possible mechanism for cholinergic dysfunction in the central nervous system. J. Neurochem. 77, 1136 –1144 Ikonomidou, C., Stefovska, V., and Turski, L. (2000) Neuronal death enhanced by N-methyl-d-aspartate antagonists. Proc. Natl. Acad. Sci. USA 97, 12885–12890 Fix, A. S., Horn, J. W., Wightman, K. A., Johnson, C. A., Long, G. G., Storts, R. W., Farber, N., Wozniak, D. F., and Olney, J. W. (1993) Neuronal vacuolization and necrosis induced by the noncompetitive N-methyl-d-aspartate (NMDA) antagonist MK(⫹)801 (dizocilpine maleate): a light and electron microscopic evaluation of the rat retrosplenial cortex. Exp. Neurol. 123, 204 –215 Jevtovic-Todorovic, V., Wozniak, D. F., Powell, S., and Olney, J. W. (2001) Propofol and sodium thiopental protect against MK-801-induced neuronal necrosis in the posterior cingulate/ retrosplenial cortex. Brain Res. 913, 185–189 Balla, A., Koneru, R., Smiley, J., Sershen, H., and Javitt, D. C. (2001) Continuous phencyclidine treatment induces schizophrenia-like hyperreactivity of striatal dopamine release. Neuropsychopharmacology 25, 157–164 Abel, K. M., Allin, M. P. G., Hemsley, D. R., and Geyer, M. A. (2003) Low dose ketamine increases prepulse inhibition in health men. Neuropharmacology 44, 729 –737 Holden, C. (2003) Excited by glutamate. Science 300, 1866 –1868 Keller, J. N., Mark, R. J., Bruce, A. J., Blanc, E., Rothstein, J. D., Uchida, K., Waeg, G., and Mattson, M. P. (1997) 4-Hydroxynonenal, an aldehydic product of membrane lipid peroxidation, impairs glutamate transport and mitochondrial function in synaptosomes. Neuroscience 80, 685– 696 Butterfield, A. D., Castegna, A., Lauderback, M., and Drake, J. (2002) Evidence that amyloid beta-peptide-induced lipid peroxidation and its sequelae in Alzheimer's disease brain contribute to neuronal death. Neurobiol. Aging 23, 655– 664 Tadolini, B., Pintus, G., Pinna, G. G., Bennardini, F., and Franconi, F. (1995) Effects of taurine and hypotaurine on lipid peroxidation. Biochem. Biophys. Res. Commun. 213, 820 – 826 Paula Lima, A. C., Louzada, P. R., Mello, F. G., and Ferreira, S. T. (2003) Neuroprotection against A and glutamate toxicity by melatonin: are GABA receptors involved? Neurotox. Res. 5, 323–328 Forloni, G., Lucca, E., Angeretti, N., Chiesa, R., and Vezzani, A. (1997) Neuroprotective effect of somatostatin on nonapoptotic NMDA-induced neuronal death: role of cyclic GMP. J. Neurochem. 68, 319 –327 Velasco, I., and Tapia, R. (2002) High extracellular gammaaminobutyric acid protects cultured neurons against damage induced by the accumulation of endogenous extracellular glutamate. J. Neurosci. Res. 67, 406 – 410 Green, A. R., Hainsworth, A. H., and Jackson, D. M. (2000) GABA potentiation: a logical pharmacological approach for the treatment of acute ischaemic stroke. Neuropharmacology 39, 1483–1494 Schwartz-Bloom, R. D., Miller, K. A., Evenson, D. A., Crain, B. J., and Nadler, J. V. (2000) Benzodiazepines protect hippocampal neurons from degeneration after transient cerebral ischemia: an ultrastructural study. Neuroscience 98, 471– 484 Frandsen, A., Quistorff, B., and Schousboe, A. (1990) Phenobarbital protects cerebral cortex neurones against toxicity induced by kainate but not by other excitatory amino acids. Neurosci. Lett. 111, 233–238 Mark, R. J., Ashford, W., Goodman, Y., and Mattson, M. P. (1995) Anticonvulsants attenuate amyloid beta-peptide neurotoxicity, Ca2⫹ deregulation, and cytoskeletal pathology. Neurobiol. Aging 16, 187–198 Birdsall, T. C. (1998) Therapeutic applications of taurine. Altern. Med. Rev. 3, 128 –136
517
51.
52.
53.
54. 55.
518
Matsuyama, Y., Morita, T., Higuchi, M., and Tisujii, T. (1983) The effect of taurine administration on patients with acute hepatitis. Prog. Clin. Biol. Res. 125, 461– 468 Benrabh, H., Bourre, J. M., and Lefauconnier, J. M. (1995) Taurine transport at the blood– brain barrier: an in vivo brain perfusion study. Brain Res. 692, 57– 65 Tamai, I., and Tisuji, A. (1995) Na(⫹)- and Cl(–)-dependent transport of taurine at the blood3H brain barrier. Biochem. Pharmacol. 50, 1783–1793 Hassel, B., Tauboll, E., and Gjerstad, L. (2001) Chronic lamotrigine treatment increases rat hippocampal GABA shunt activity and elevates cerebral taurine levels. Epilepsy Res. 43, 153–163 Rodrigues, C. M. P., Sola´ , S., Brito, M. A., Brondino, C. D., Brites, D., and Moura, J. J. G. (2001) Amyloid beta-peptide
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March 2004
disrupts mitochondrial membrane lipid and protein structure: protective role of tauroursodeoxycholate. Biochem. Biophys. Res. Commun. 281, 468 – 474 56. O’Byrne, M. B., and Tipton, K. F. (2000) Taurine-induced attenuation of MPP⫹ neurotoxicity in vitro: a possible role for the GABAA subclass of GABA receptors. J. Neurochem. 74, 2087– 2093 57. Pomara, N., Rajkumar Singh, M. D., Deptula, D., Chou, J. C. Y., Schwartz, M. B., and LeWitt, P. A. (1992) Glutamate and other CSF amino acids in Alzheimer's disease. Am. J. Psychiatry 149, 251–254
The FASEB Journal
Received for publication August 18, 2003. Accepted for publication November 12, 2003.
LOUZADA ET AL.
