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W2061458168 abstract "Valproate is an important anticonvulsant currently in clinical use for the treatment of seizures. We used electrophysiological and tracer uptake methods to examine the effect of valproate on a γ-aminobutyric acid (GABA) transporter (mouse GAT3) expressed in Xenopus laevis oocytes. In the absence of GABA, valproate (up to 50 mm) had no noticeable effect on the steady-state electrogenic properties of mGAT3. In the presence of GABA, however, valproate enhanced the GABA-evoked steady-state inward current in a dose-dependent manner with a half-maximal concentration of 4.6 ± 0.5 mm. Maximal enhancement of the GABA-evoked current was 275 ± 10%. Qualitatively similar observations were obtained for human GAT1 and mouse GAT4. The valproate enhancement did not alter the Na+or Cl− dependence of the steady-state GABA-evoked currents. Uptake experiments under voltage clamp suggested that the valproate enhancement of the GABA-evoked current was matched by an enhancement in GABA uptake. Thus, despite the increase in GABA-evoked current, ion/GABA co-transport remained tightly coupled. Uptake experiments indicated that valproate is not transported by mouse GAT3 in the absence or presence of GABA. Valproate also enhanced the rate of the partial steps involved in transporter presteady-state charge movements. We propose that valproate increases the turnover rate of GABA transporters by an allosteric mechanism. The data suggest that at its therapeutic concentration, valproate may enhance the activity of neuronal and glial GABA transporters by up to 10%. Valproate is an important anticonvulsant currently in clinical use for the treatment of seizures. We used electrophysiological and tracer uptake methods to examine the effect of valproate on a γ-aminobutyric acid (GABA) transporter (mouse GAT3) expressed in Xenopus laevis oocytes. In the absence of GABA, valproate (up to 50 mm) had no noticeable effect on the steady-state electrogenic properties of mGAT3. In the presence of GABA, however, valproate enhanced the GABA-evoked steady-state inward current in a dose-dependent manner with a half-maximal concentration of 4.6 ± 0.5 mm. Maximal enhancement of the GABA-evoked current was 275 ± 10%. Qualitatively similar observations were obtained for human GAT1 and mouse GAT4. The valproate enhancement did not alter the Na+or Cl− dependence of the steady-state GABA-evoked currents. Uptake experiments under voltage clamp suggested that the valproate enhancement of the GABA-evoked current was matched by an enhancement in GABA uptake. Thus, despite the increase in GABA-evoked current, ion/GABA co-transport remained tightly coupled. Uptake experiments indicated that valproate is not transported by mouse GAT3 in the absence or presence of GABA. Valproate also enhanced the rate of the partial steps involved in transporter presteady-state charge movements. We propose that valproate increases the turnover rate of GABA transporters by an allosteric mechanism. The data suggest that at its therapeutic concentration, valproate may enhance the activity of neuronal and glial GABA transporters by up to 10%. γ-aminobutyric acid GABA transporter mouse GAT human GAT current-voltage charge-voltage maximum transporter-mediated charge γ-Aminobutyric acid (GABA)1 is the most abundant inhibitory neurotransmitter in the central nervous system. Transport of GABA into cells is accomplished by Na+-dependent and Cl−-facilitated GABA transporters (GATs) found in the plasma membrane of neurons and glia (1Kavanaugh M.P. Arriza J.L. North R.A. Amara S.G. J. Biol. Chem. 1992; 267: 22007-22009Abstract Full Text PDF PubMed Google Scholar, 2Keynan S. Suh Y.-J. Kanner B.I. Rudnick G. Biochemistry. 1992; 31: 1974-1979Crossref PubMed Scopus (121) Google Scholar, 3Mager S. Naeve J. Quick M. Labarca C. Davidson N. Lester H.A. Neuron. 1993; 10: 177-188Abstract Full Text PDF PubMed Scopus (276) Google Scholar, 4Clark J.A. Amara S.G. Mol. Pharmacol. 1994; 46: 550-557PubMed Google Scholar, 5Borden L.A. Neurochem. Int. 1996; 29: 335-356Crossref PubMed Scopus (518) Google Scholar, 6Nelson N. J. Neurochem. 1998; 71: 1785-1803Crossref PubMed Scopus (321) Google Scholar, 7Matskevitch I. Wagner C.A. Stegen C. Bröer S. Noll B. Risler T. Kwon H.M. Handler J.S. Waldegger S. Busch A.E. Lang F. J. Biol. Chem. 1999; 274: 16709-16716Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar, 8Lu C.-C. Hilgemann D.W. J. Gen. Physiol. 1999; 114: 429-444Crossref PubMed Scopus (101) Google Scholar, 9Loo D.D.F. Eskandari S. Boorer K.J. Sarkar H.K. Wright E.M. J. Biol. Chem. 2000; 275: 37414-37422Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar, 10Sacher A. Nelson N. Ogi J.T. Wright E.M. Loo D.D.F. Eskandari S. J. Membr. Biol. 2002; 190: 57-73Crossref PubMed Scopus (28) Google Scholar). Thus, the GABA transporters regulate synaptic and extra-synaptic concentrations of GABA and, in this capacity, are partly responsible for the regulation of inhibitory neurotransmission in the nervous system. Because of the inhibitory role of GABA, potentiation of GABAergic neurotransmission via inhibition or reversal of the GABA transporters is believed to have therapeutic value in treating epileptic seizures and stroke (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 12Roettger V.R. Amara S.G. Adv. Neurol. 1999; 79: 551-560PubMed Google Scholar, 13Green A.R. Hainsworth A.H. Jackson D.M. Neuropharmacology. 2000; 39: 1483-1494Crossref PubMed Scopus (186) Google Scholar). Indeed, inhibitors of the GABA transporters are known to increase GABA levels in the brain (14Fink-Jensen A. Suzdak P.D. Swedberg M.D. Judge M.E. Hansen L. Nielsen P.G. Eur. J. Pharmacol. 1992; 220: 197-201Crossref PubMed Scopus (174) Google Scholar, 15Richards D.A. Bowery N.G. Neurochem. Res. 1996; 21: 135-140Crossref PubMed Scopus (33) Google Scholar, 16Dalby N.O. Neuropharmacology. 2000; 39: 2399-2407Crossref PubMed Scopus (126) Google Scholar). These agents exhibit anticonvulsant activity in animal models, and one (tiagabine) that preferentially targets the most abundant GABA transporter isoform in the brain (GAT1) has been in clinical use since 1997 (16Dalby N.O. Neuropharmacology. 2000; 39: 2399-2407Crossref PubMed Scopus (126) Google Scholar, 17Yunger L.M. Fowler P.J. Zarevics P. Setler P.E. J. Pharmacol. Exp. Ther. 1984; 228: 109-115PubMed Google Scholar, 18Nielsen E.B. Suzdak P.D. Andersen K.E. Knutsen L.J. Sonnewald U. Braestrup C. Eur. J. Pharmacol. 1991; 196: 257-266Crossref PubMed Scopus (215) Google Scholar, 19Swinyard E.A. White H.S. Wolf H.H. Bondinell W.E. Epilepsia. 1991; 32: 569-577Crossref PubMed Scopus (23) Google Scholar, 20Suzdak P.D. Frederiksen K. Andersen K.E. Sorensen P.O. Knutsen L.J. Nielsen E.B. Eur. J. Pharmacol. 1992; 224: 189-198Crossref PubMed Scopus (148) Google Scholar, 21Dalby N.O. Thomsen C. Fink-Jensen A. Lundbeck J. Søkilde B. Man C.M. Sørensen P.O. Meldrum B. Epilepsy Res. 1997; 28: 51-61Crossref PubMed Scopus (45) Google Scholar, 22Morimoto K. Sato H. Yamamoto Y. Watanabe T. Suwaki H. Epilepsia. 1997; 38: 966-974Crossref PubMed Scopus (56) Google Scholar). Several other clinically used antiepileptic drugs are reported to act, at least in part, via potentiating GABA-mediated inhibition in the brain (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 23Kwan P. Sills G.J. Brodie M.J. Pharmacol. Ther. 2001; 90: 21-34Crossref PubMed Scopus (230) Google Scholar); however, little is known regarding the potential effect of these drugs on the GABA transporters (24Eckstein-Ludwig U. Fei J. Schwarz W. Br. J. Pharmacol. 1999; 128: 92-102Crossref PubMed Scopus (65) Google Scholar).Valproate (2-propylpentanoate) has been in clinical use since 1967 and is effective against many types of epileptic seizures (both partial and generalized seizures). Although the exact mechanism of valproate action is not clear, its effectiveness as a broad-spectrum anticonvulsant is usually attributed to a combination of actions at multiple molecular targets (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 25Johannessen C.U. Neurochem Int. 2000; 37: 103-110Crossref PubMed Scopus (325) Google Scholar). A preponderance of evidence suggests that valproate potentiates GABAergic neurotransmission by increasing GABA levels in the brain (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 26Godin Y. Heiner L. Mark J. Mandel P. J. Neurochem. 1969; 16: 869-873Crossref PubMed Scopus (372) Google Scholar, 27Iadarola M.J. Raines A. Gale K. J. Neurochem. 1979; 33: 1119-1123Crossref PubMed Scopus (52) Google Scholar, 28Löscher W. Siemes H. Lancet. 1984; 2: 225Abstract PubMed Scopus (55) Google Scholar, 29Löscher W. Siemes H. Epilepsia. 1985; 26: 314-319Crossref PubMed Scopus (76) Google Scholar, 30Löscher W. Brain Res. 1989; 501: 198-203Crossref PubMed Scopus (74) Google Scholar, 31Biggs C.S. Pearce B.R. Fowler L.J. Whitton P.S. Brain Res. 1992; 594: 138-142Crossref PubMed Scopus (70) Google Scholar, 32Rowley H.L. Marsden C.A. Martin K.F. Eur. J. Pharmacol. 1995; 294: 541-546Crossref PubMed Scopus (46) Google Scholar). The effect may be attributed both to enhanced GABA synthesis (stimulation of glutamic acid decarboxylase) and decreased GABA degradation (inhibition of GABA trans-aminase and succinic semialdehyde dehydrogenase) (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 33van der Laan J.W. de Boer T. Bruinvels J. J. Neurochem. 1979; 32: 1769-1780Crossref PubMed Scopus (131) Google Scholar, 34Larsson O.M. Gram L. Schousboe I. Schousboe A. Neuropharmacology. 1986; 25: 617-625Crossref PubMed Scopus (75) Google Scholar, 35Taylor C.P. Vartanian M.G. Andruszkiewicz R. Silverman R.B. Epilepsy Res. 1992; 11: 103-110Crossref PubMed Scopus (92) Google Scholar). Correspondingly, many studies have examined valproate-induced GABA release from neurons and glia, although vesicular versus non-vesicular (i.e. transporter-mediated) release mechanisms have not been definitively addressed. These studies as well as those examining the effect of valproate on GABA uptake by neurons and/or glial cells have not provided entirely consistent results; however, the available evidence favors valproate-induced GABA release from nerve terminals in selected brain regions (e.g. 31, 32, 36–47; for review see Refs. 11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar and 25Johannessen C.U. Neurochem Int. 2000; 37: 103-110Crossref PubMed Scopus (325) Google Scholar). In light of this evidence, it is of interest to characterize the effect of valproate on the GABA transporters expressed in an expression system where any putative interaction may be examined by using sensitive biophysical tools.Here, we have combined electrophysiological and tracer uptake methods to examine the effect of valproate on the GABA transporters expressed in Xenopus laevis oocytes. Our results suggest that valproate enhances the turnover rate of these transport proteins via an allosteric mechanism. The interaction of valproate with GATs opens new experimental avenues for probing the mechanism of Na+/Cl−/GABA co-transport.DISCUSSIONWe propose that valproate leads to an increase in the turnover rate of GABA transporters and that the effect involves an important rate-limiting step in the transport cycle. Our data do not allow us to identify the partial step in the transport cycle that is altered by valproate; however, we have shown that valproate increases the turnover rate for the forward mode of the transporter (Fig.11A) as well as increases the rates of conformational changes of the empty carrier (presteady-state relaxations) (Fig. 11B, shaded steps). Valproate interaction with the transporter appears to be allosteric involving a site other than the GABA binding site, because valproate is not a transported substrate of GATs and valproate interaction does not compete with GABA. Indeed, valproate enhances GABA trans-location across the plasma membrane in the forward transport mode. Despite enhancement of GABA transport across the plasma membrane, the ion/GABA coupling ratio remains the same, suggesting that the transport cycle remains tightly coupled. Valproate interaction with the GABA transporters is specific as other short chain fatty acids such as butanoic acid and pentanoic acid were without effect (data not shown). Furthermore, valproate did not enhance the rate of transport for the Na+/iodide symporter, a Na+-coupled transporter with mechanistic features similar to those of the GABA transporters. Examination of transporter presteady-state kinetics suggests that valproate increases the apparent affinity of the empty transporter for Na+. The presteady-state charge movements further suggest that valproate can interact with the transporter in the absence of GABA and Cl−. However, it cannot be determined whether valproate can interact with the transporter in the absence of Na+, because no charge movements are induced by valproate in the absence of external Na+. Finally, valproate-induced enhancement is rapid and limited only by the speed of the perfusion system, and moreover, the enhancement is fully reversible.At least three observations suggest that valproate interaction with mGAT3 is not at the GABA binding site. (i) Valproate alone neither evokes an inward current nor is it itself transported across the plasma membrane alone or in the presence of GABA. (ii) Increasing concentrations of valproate enhance both the GABA-evoked current and GABA uptake, whereas interaction at the GABA binding site would be expected to lead to competitive inhibition of GABA uptake. (iii) Valproate had no effect on the maximal charge moved in response to voltage pulses (Qmax).Qmax is a measure of the total number of functional transporters available to bind substrate (56Eskandari S. Kreman M. Kavanaugh M.P. Wright E.M. Zampighi G.A. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 8641-8646Crossref PubMed Scopus (109) Google Scholar). As transported substrates lead to a concentration-dependent reduction in Qmax (Fig. 10C) (10Sacher A. Nelson N. Ogi J.T. Wright E.M. Loo D.D.F. Eskandari S. J. Membr. Biol. 2002; 190: 57-73Crossref PubMed Scopus (28) Google Scholar,51Eskandari S. Loo D.D.F. Dai G. Levy O. Wright E.M. Carrasco N. J. Biol. Chem. 1997; 272: 27230-27238Abstract Full Text Full Text PDF PubMed Scopus (376) Google Scholar), the results suggest that interaction of valproate with the GABA transporters occurs in an entirely different fashion from that of a transported substrate. Because of the moderate membrane permeability of valproate, it is not possible to know whether this drug acts at the extracellular, intracellular, or membrane-spanning region of the transporter.The effect of valproate cannot be attributed to an increase in the total number of transporters caused by vesicle trafficking to the plasma membrane. Four observations support this view. (i) The total number of transporters in the plasma membrane (as determined fromQmax) was not changed by valproate. (ii) The effect was fully reversible and repeatable in the same cell, persisting for as long as the experiment was continued. It seems unlikely that such mGAT3-containing vesicle insertion could be balanced rapidly and precisely by recruitment of mGAT3 in retrieved vesicles. (iii) The effect was specific to the GABA transporters as the transport rate for the Na+/iodide symporter was not altered. (iv) There was no change in the whole-cell capacitance in the presence of valproate. This is shown clearly in Fig. 3, A and B. We have shown previously that heterologous membrane proteins expressed in oocytes are targeted to the plasma membrane in 100-nm diameter vesicles containing 5–40 copies of the expressed protein (57Zampighi G.A. Loo D.D.F. Kreman M. Eskandari S. Wright E.M. J. Gen. Physiol. 1999; 113: 507-523Crossref PubMed Scopus (98) Google Scholar). In the oocyte of Fig. 3A, the GABA-evoked current was enhanced by ≈50 nA. If it is assumed that the enhancement was because of insertion of new mGAT3 copies in the plasma membrane, it can be estimated that a total of ≈7 × 1010 transporters were newly inserted into the membrane (I = NRze, where I is current,N is the number of transporters, R is the turnover rate (2 s−1 at −60 mV; see Ref. 10Sacher A. Nelson N. Ogi J.T. Wright E.M. Loo D.D.F. Eskandari S. J. Membr. Biol. 2002; 190: 57-73Crossref PubMed Scopus (28) Google Scholar),z is the net charge trans-located across the membrane per transport cycle (2.2; see legend to Fig. 2B), ande is the elementary charge). Assuming that there were 5–40 copies of mGAT3 per vesicle, 2 × 109 to 14 × 109 vesicles would have been expected to fuse with the plasma membrane, leading to a 2–16-fold increase in the total surface area of the oocyte. Clearly, with a resolution of ≈10 nanofarads, such an increase in capacitance (260 nanofarads to at least 520 nanofarads) would have been detected (for example see Ref. 58Hirsch J.R. Loo D.D.F. Wright E.M. J. Biol. Chem. 1996; 271: 14740-14746Abstract Full Text Full Text PDF PubMed Scopus (170) Google Scholar).A prominent effect of valproate was an increase in the rate of the presteady-state ON and OFF relaxations. Upon application of depolarizing voltage pulses, the ON transients represent the release of Na+ and Cl− followed by reorientation of the empty carrier (Fig. 11B, shaded steps). Upon return from the test voltage to the holding voltage, the OFF transients represent the return of the binding sites to the external medium, Na+ and Cl− entry into the membrane electric field, binding, and subsequent ligand-induced conformational changes (Fig. 11B) (3Mager S. Naeve J. Quick M. Labarca C. Davidson N. Lester H.A. Neuron. 1993; 10: 177-188Abstract Full Text PDF PubMed Scopus (276) Google Scholar, 52Loo D.D.F. Hazama A. Supplisson S. Turk E. Wright E.M. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 5767-5771Crossref PubMed Scopus (203) Google Scholar, 53Mager S. Kleinberger-Doron N. Keshet G.I. Davidson N. Kanner B.I. Lester H.A. J. Neurosci. 1996; 16: 5405-5414Crossref PubMed Google Scholar, 54Lu C.-C. Hilgemann D.W. J. Gen. Physiol. 1999; 114: 445-457Crossref PubMed Scopus (61) Google Scholar, 55Li M. Farley R.A. Lester H.A. J. Gen. Physiol. 2000; 115: 491-508Crossref PubMed Scopus (44) Google Scholar). As presteady-state charge movements are obtained in the absence of GABA, the data suggest that valproate also increases the rates associated with voltage-induced conformational changes of the empty carrier.Valproate also led to a small reduction in the apparent affinity for GABA. Both the half-maximal concentration for steady-state GABA-evoked inward current and the GABA concentration for 50% reduction inQmax were increased by valproate. The values increased from ≈6 μm in the absence of valproate to ≈20 μm in the presence of 10 mm valproate. Apparently, valproate reversibly transforms mGAT3 into a lower affinity, higher capacity transporter. Interestingly, valproate was also reported to decrease the GABA affinity of the astroglial GABA uptake system (44Nilsson M. Hansson E. Rönnbäck L. Neurochem. Res. 1992; 17: 327-332Crossref PubMed Scopus (28) Google Scholar), although maximal transport rates were not altered.At saturating Na+ and Cl− concentrations, valproate did not alter the maximal charge moved in response to voltage pulses (see Figs. 7, 8, and 10), suggesting that valproate entry/exit into/out of the membrane electric field does not contribute to presteady-state charge movements. Valproate did not alter the apparent valence of the moveable charge (zδ), suggesting that the same number of charges moved the same distance within the membrane electric field in the absence or presence of valproate. Valproate led to a decrease in the half-maximal concentration for Na+activation (43 versus 22 mm) of the charge movements but had no effect on the half-maximal concentrations for Cl− enhancement (15 mm) of the charge movements. Therefore, the results suggest that valproate increases the apparent affinity of the empty transporter for Na+. Thus, by increasing the affinity of the empty carrier for Na+, significantly more charge can be moved at lower Na+ and Cl− concentrations. This effect may contribute to the enhancement of transporter turnover rate induced by valproate. In contrast, valproate does not alter the apparent Na+affinity of the GABA-loaded transporter (see Fig. 5A). As the presteady-state transitions (Fig. 11B, shaded steps) represent only a subset of those of the entire transport cycle (Fig. 11A), the data suggest that the effect of valproate is complex and may also involve partial steps other than those responsible for the voltage-induced presteady-state transitions. The data do not allow us to specify the steps altered by valproate.Our results are in contrast to those obtained by Eckstein-Ludwiget al. (24Eckstein-Ludwig U. Fei J. Schwarz W. Br. J. Pharmacol. 1999; 128: 92-102Crossref PubMed Scopus (65) Google Scholar) who report the inhibition of mouse GAT1-mediated GABA uptake by valproate, although it was also reported that valproate did not alter the steady-state or presteady-state currents of mouse GAT1. The authors propose a valproate-induced dissociation of GABA uptake and GABA-induced currents. We were unable to observe such effects under our experimental conditions. Indeed, we have provided direct demonstration of tight coupling between GABA uptake and steady-state GABA-induced currents in the absence and presence of valproate. Differences in the GAT isoform and/or valproate concentration used may be responsible for the observed discrepancy between our results and those of Eckstein-Ludwig et al.(24Eckstein-Ludwig U. Fei J. Schwarz W. Br. J. Pharmacol. 1999; 128: 92-102Crossref PubMed Scopus (65) Google Scholar).In humans, the therapeutic concentration of valproate in the plasma ranges from 0.28 to 0.69 mm (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar). Because of valproate breakdown to metabolic byproducts as well as carrier-mediated efflux from the brain, valproate concentration in the cerebrospinal fluid is a fraction of that in plasma and ranges from 0.042 to 0.19 mm(11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar). The half-maximal concentration for valproate enhancement of the GAT turnover rate is ≈4.5 mm, and maximal enhancement is ≈275%. Therefore, we predict that at therapeutic levels of valproate, the activity of the GABA transporters may be enhanced by up to 10%. Valproate has been shown to lead to a 15–40% elevation of GABA levels in the cerebrospinal fluid (25Johannessen C.U. Neurochem Int. 2000; 37: 103-110Crossref PubMed Scopus (325) Google Scholar). Although our data do not allow us to speculate on the role played by the GABA transporters, at least in principle, it is possible that valproate enhancement of the GABA transporters may contribute to the observed elevation of GABA levels in the cerebrospinal fluid. By stimulating GABA synthesis and inhibiting GABA degradation, valproate increases the nerve ending GABA pool (36Iadarola M.J. Gale K. Eur. J. Pharmacol. 1979; 59: 125-129Crossref PubMed Scopus (63) Google Scholar, 41Löscher W. Vetter M. Biochem. Pharmacol. 1985; 34: 1747-1756Crossref PubMed Scopus (65) Google Scholar), which may favor GABA release via the reversal of the GABA transporters (8Lu C.-C. Hilgemann D.W. J. Gen. Physiol. 1999; 114: 429-444Crossref PubMed Scopus (101) Google Scholar). Therefore, valproate enhancement of the GABA transporter turnover rate may add to its effectiveness in facilitating GABA release from cells, leading to GABA potentiation in the brain. These findings add support to the notion that the effectiveness of valproate as an anticonvulsant results from its combined action at multiple molecular targets in the brain.CONCLUSIONSince its fortuitous discovery as an anticonvulsant in 1962, valproate has become one of the most widely used drugs of its class, perhaps because of its wide spectrum of anticonvulsant activity against different types of seizures. The wide spectrum of activity most certainly arises from diverse molecular actions. Here, we have presented data for an additional role of valproate in the central nervous system, the enhancement of the turnover rate of the GABA transporters. We predict that at therapeutic concentrations, valproate may enhance the activity of neuronal and glial GABA transporters by up to 10%. γ-Aminobutyric acid (GABA)1 is the most abundant inhibitory neurotransmitter in the central nervous system. Transport of GABA into cells is accomplished by Na+-dependent and Cl−-facilitated GABA transporters (GATs) found in the plasma membrane of neurons and glia (1Kavanaugh M.P. Arriza J.L. North R.A. Amara S.G. J. Biol. Chem. 1992; 267: 22007-22009Abstract Full Text PDF PubMed Google Scholar, 2Keynan S. Suh Y.-J. Kanner B.I. Rudnick G. Biochemistry. 1992; 31: 1974-1979Crossref PubMed Scopus (121) Google Scholar, 3Mager S. Naeve J. Quick M. Labarca C. Davidson N. Lester H.A. Neuron. 1993; 10: 177-188Abstract Full Text PDF PubMed Scopus (276) Google Scholar, 4Clark J.A. Amara S.G. Mol. Pharmacol. 1994; 46: 550-557PubMed Google Scholar, 5Borden L.A. Neurochem. Int. 1996; 29: 335-356Crossref PubMed Scopus (518) Google Scholar, 6Nelson N. J. Neurochem. 1998; 71: 1785-1803Crossref PubMed Scopus (321) Google Scholar, 7Matskevitch I. Wagner C.A. Stegen C. Bröer S. Noll B. Risler T. Kwon H.M. Handler J.S. Waldegger S. Busch A.E. Lang F. J. Biol. Chem. 1999; 274: 16709-16716Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar, 8Lu C.-C. Hilgemann D.W. J. Gen. Physiol. 1999; 114: 429-444Crossref PubMed Scopus (101) Google Scholar, 9Loo D.D.F. Eskandari S. Boorer K.J. Sarkar H.K. Wright E.M. J. Biol. Chem. 2000; 275: 37414-37422Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar, 10Sacher A. Nelson N. Ogi J.T. Wright E.M. Loo D.D.F. Eskandari S. J. Membr. Biol. 2002; 190: 57-73Crossref PubMed Scopus (28) Google Scholar). Thus, the GABA transporters regulate synaptic and extra-synaptic concentrations of GABA and, in this capacity, are partly responsible for the regulation of inhibitory neurotransmission in the nervous system. Because of the inhibitory role of GABA, potentiation of GABAergic neurotransmission via inhibition or reversal of the GABA transporters is believed to have therapeutic value in treating epileptic seizures and stroke (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 12Roettger V.R. Amara S.G. Adv. Neurol. 1999; 79: 551-560PubMed Google Scholar, 13Green A.R. Hainsworth A.H. Jackson D.M. Neuropharmacology. 2000; 39: 1483-1494Crossref PubMed Scopus (186) Google Scholar). Indeed, inhibitors of the GABA transporters are known to increase GABA levels in the brain (14Fink-Jensen A. Suzdak P.D. Swedberg M.D. Judge M.E. Hansen L. Nielsen P.G. Eur. J. Pharmacol. 1992; 220: 197-201Crossref PubMed Scopus (174) Google Scholar, 15Richards D.A. Bowery N.G. Neurochem. Res. 1996; 21: 135-140Crossref PubMed Scopus (33) Google Scholar, 16Dalby N.O. Neuropharmacology. 2000; 39: 2399-2407Crossref PubMed Scopus (126) Google Scholar). These agents exhibit anticonvulsant activity in animal models, and one (tiagabine) that preferentially targets the most abundant GABA transporter isoform in the brain (GAT1) has been in clinical use since 1997 (16Dalby N.O. Neuropharmacology. 2000; 39: 2399-2407Crossref PubMed Scopus (126) Google Scholar, 17Yunger L.M. Fowler P.J. Zarevics P. Setler P.E. J. Pharmacol. Exp. Ther. 1984; 228: 109-115PubMed Google Scholar, 18Nielsen E.B. Suzdak P.D. Andersen K.E. Knutsen L.J. Sonnewald U. Braestrup C. Eur. J. Pharmacol. 1991; 196: 257-266Crossref PubMed Scopus (215) Google Scholar, 19Swinyard E.A. White H.S. Wolf H.H. Bondinell W.E. Epilepsia. 1991; 32: 569-577Crossref PubMed Scopus (23) Google Scholar, 20Suzdak P.D. Frederiksen K. Andersen K.E. Sorensen P.O. Knutsen L.J. Nielsen E.B. Eur. J. Pharmacol. 1992; 224: 189-198Crossref PubMed Scopus (148) Google Scholar, 21Dalby N.O. Thomsen C. Fink-Jensen A. Lundbeck J. Søkilde B. Man C.M. Sørensen P.O. Meldrum B. Epilepsy Res. 1997; 28: 51-61Crossref PubMed Scopus (45) Google Scholar, 22Morimoto K. Sato H. Yamamoto Y. Watanabe T. Suwaki H. Epilepsia. 1997; 38: 966-974Crossref PubMed Scopus (56) Google Scholar). Several other clinically used antiepileptic drugs are reported to act, at least in part, via potentiating GABA-mediated inhibition in the brain (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 23Kwan P. Sills G.J. Brodie M.J. Pharmacol. Ther. 2001; 90: 21-34Crossref PubMed Scopus (230) Google Scholar); however, little is known regarding the potential effect of these drugs on the GABA transporters (24Eckstein-Ludwig U. Fei J. Schwarz W. Br. J. Pharmacol. 1999; 128: 92-102Crossref PubMed Scopus (65) Google Scholar). Valproate (2-propylpentanoate) has been in clinical use since 1967 and is effective against many types of epileptic seizures (both partial and generalized seizures). Although the exact mechanism of valproate action is not clear, its effectiveness as a broad-spectrum anticonvulsant is usually attributed to a combination of actions at multiple molecular targets (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 25Johannessen C.U. Neurochem Int. 2000; 37: 103-110Crossref PubMed Scopus (325) Google Scholar). A preponderance of evidence suggests that valproate potentiates GABAergic neurotransmission by increasing GABA levels in the brain (11Löscher W. Prog. Neurobiol. 1999; 58: 31-59Crossref PubMed Scopus (446) Google Scholar, 26Godin Y. Heiner L. Mark J. Mandel P. J. Neurochem. 1969; 16: 869-873Crossref Pub" @default.
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W2061458168 title "The Anticonvulsant Valproate Increases the Turnover Rate of γ-Aminobutyric Acid Transporters" @default.
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