Characterization and distribution of the neuronal glutamate transporter EAAC1 in rat brain

被引:67
作者
VelazFaircloth, M
McGraw, TS
Malandro, MS
Fremeau, RT
Kilberg, MS
Anderson, KJ
机构
[1] UNIV FLORIDA, HLTH SCI CTR, DEPT PHYSIOL SCI, GAINESVILLE, FL 32610 USA
[2] DUKE UNIV, DEPT PHARMACOL, DURHAM, NC 27710 USA
[3] DUKE UNIV, DEPT NEUROBIOL, DURHAM, NC 27710 USA
[4] UNIV FLORIDA, DEPT NEUROSCI, GAINESVILLE, FL 32610 USA
[5] UNIV FLORIDA, DEPT BIOCHEM & MOLEC BIOL, GAINESVILLE, FL 32610 USA
来源
AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY | 1996年 / 270卷 / 01期
关键词
transport; excitatory amino acid; aspartate; in situ hybridization;
D O I
10.1152/ajpcell.1996.270.1.C67
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
The extracellular concentration of glutamate and other related excitatory amino acids (EAA) is regulated by the action of transporter proteins located on either presynaptic terminals or adjacent astroglial processes. Recent molecular advances have led to the cloning of three separate cDNAs encoding for Na+-dependent glutamate transporters; two are thought to be primarily glial in origin (GLAST and GLT-1) and the third (EAAC1) is localized to neurons in the brain and other nonneural tissues. An EAAC 1 cDNA was initially cloned from rabbit small intestine (13). In this study, we report isolation and characterization of the homologous clone from rat brain. Northern blot hybridization revealed high levels of EAAC1 mRNA in rat brain and kidney and low levels in heart, lung, and skeletal muscle. Transient expression of EAAC 1 in HeLa cells resulted in an increase in Na-?+-dependent high-affinity L-[H-3]glutamate and D-[H-3] aspartate transport. The pharmacological profile of EAAC1 was very similar to that reported for the rabbit and human EAAC1 homologues. Transport activity was potently inhibited by D- and L-threo-beta-hydroxyaspartate and L-trans-pyrrolodine-2, 4-dicarboxylate. Dihydro-kainate and L-alpha-aminoadipate did not inhibit transport at concentrations below 1 mM. Oligonucleotide cDNA probes (45-mer) were constructed and labeled with S-35-ATP for film- and emulsion-based in situ hybridization of rat brain. EAAC1 mRNA had the highest density in the cerebellar granule cell layer, hippocampus, superior colliculus, and neocortex. Sections that were emulsion-dipped and counterstained with cresyl violet revealed EAAC1 labeling localized exclusively over neuronal cell bodies, including some nonglutamatergic neurons such as spinal cord ventral horn cells.
引用
收藏
页码:C67 / C75
页数:9
相关论文
共 30 条
[1]   ANATOMICAL AND PHARMACOLOGICAL HETEROGENEITY OF D-[H-3]ASPARTATE BINDING-SITES [J].
ANDERSON, KJ ;
VICKROY, TW .
EUROPEAN JOURNAL OF PHARMACOLOGY, 1990, 185 (01) :119-121
[2]  
ARRIZA JL, 1994, J NEUROSCI, V14, P5559
[3]   THE GLIAL-CELL GLUTAMATE UPTAKE CARRIER COUNTERTRANSPORTS PH-CHANGING ANIONS [J].
BOUVIER, M ;
SZATKOWSKI, M ;
AMATO, A ;
ATTWELL, D .
NATURE, 1992, 360 (6403) :471-474
[4]  
Chamberlin Richard, 1993, P231
[5]   SINGLE-STEP METHOD OF RNA ISOLATION BY ACID GUANIDINIUM THIOCYANATE PHENOL CHLOROFORM EXTRACTION [J].
CHOMCZYNSKI, P ;
SACCHI, N .
ANALYTICAL BIOCHEMISTRY, 1987, 162 (01) :156-159
[6]  
CURRIE DN, 1981, J EXP BIOL, V95, P181
[7]  
DANBOLT NC, 1992, NEUROSCIENCE, V51, P259
[8]   HETEROGENEITY OF SODIUM-DEPENDENT EXCITATORY AMINO-ACID-UPTAKE MECHANISMS IN RAT-BRAIN [J].
FERKANY, J ;
COYLE, JT .
JOURNAL OF NEUROSCIENCE RESEARCH, 1986, 16 (03) :491-503
[9]   REGIONAL HETEROGENEITY OF L-GLUTAMATE AND L-ASPARTATE HIGH-AFFINITY UPTAKE SYSTEMS IN THE RAT CNS [J].
FLETCHER, EJ ;
JOHNSTON, GAR .
JOURNAL OF NEUROCHEMISTRY, 1991, 57 (03) :911-914
[10]   GLUTAMATE - A NEUROTRANSMITTER IN MAMMALIAN BRAIN [J].
FONNUM, F .
JOURNAL OF NEUROCHEMISTRY, 1984, 42 (01) :1-11