Molecular cloning and functional characterization of a gaba transporter from the CNS of the cabbage looper, Trichoplusia ni

Gao, Xiujuan; McLean, Heather; Caveney, Stanley; Donly, Cam

doi:10.1016/s0965-1748(99)00039-9

Cited by 19 publications

(14 citation statements)

References 61 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For example, DmCG1732 and its Anopheles homolog AgCP8499 are likely to function as GABA transporters as they are closely related to human GAT-1, C. elegans SNF11 [a recently identified GABA transporter (Mullen et al, 2006)], and the GABA transporters from Trichoplusia ni (Gao et al, 1999) and Manduca sexta (Mbungu et al, 1995). DmCG5549 and its Anopheles homolog AgCP10735 are related to the amino acid transporter subfamily, which includes the amino acid transporters for glycine and proline and a transporter with a broader substrate capacity, B O+ (Sloan and Mager, 1999).…”

Section: Drosophila Slc6 Transporters Segregate Into Four Previously mentioning

confidence: 99%

Comparative sequence analysis and tissue localization of members of the SLC6 family of transporters in adultDrosophila melanogaster

Thimgan

Berg

Stuart

2006

Journal of Experimental Biology

View full text Add to dashboard Cite

SUMMARY The SLC6 family comprises proteins that move extracellular neurotransmitters, amino acids and osmolytes across the plasma membrane into the cytosol. In mammals, deletion of SLC6 family members has dramatic physiologic consequences, but in the model organism Drosophila melanogaster, little is known about this family of proteins. Therefore,in this study we carried out an initial analysis of 21 known or putative SLC6 family members from the Drosophila genome. Protein sequences from these genes segregated into either well-defined subfamilies, including the novel insect amino acid transporter subfamily, or into a group of weakly related sequences not affiliated with a recognized subfamily. Reverse transcription-polymerase chain reaction analysis and in situhybridization showed that seven of these genes are expressed in the CNS. In situ hybridization revealed that two previously cloned SLC6 members, the serotonin and dopamine transporters, were localized to presumptive presynaptic neurons that previously immunolabelled for these transmitters. RNA for CG1732 (the putative GABA transporter) and CG15088 (a member of the novel insect amino acid transporter family)was localized in cells likely to be subtypes of glia, while RNA for CG5226, CG10804 (both members of the orphan neurotransmitter transporter subfamily) and CG5549 (a putative glycine transporter)were expressed broadly throughout the cellular cortex of the CNS. Eight of the 21 sequences were localized outside the CNS in the alimentary canal,Malpighian tubules and reproductive organs. Localization for six sequences was not found or not attempted in the adult fly. We used the Drosophilaortholog of the mammalian vesicular monoamine transporter 2, CG33528,to independently identify monoaminergic neurons in the adult fly. RNA for CG33528 was detected in a limited number of cells in the central brain and in a beaded stripe at the base of the photoreceptors in the position of glia, but not in the photoreceptors themselves. The SLC6 localization observations in conjunction with likely substrates based on phylogenetic inferences are a first step in defining the role of Na/Cl-dependent transporters in Drosophila physiology.

show abstract

Section: Drosophila Slc6 Transporters Segregate Into Four Previously mentioning

confidence: 99%

Comparative sequence analysis and tissue localization of members of the SLC6 family of transporters in adultDrosophila melanogaster

Thimgan

Berg

Stuart

2006

Journal of Experimental Biology

View full text Add to dashboard Cite

show abstract

“…The former was cloned from a cDNA library obtained from a 3-day-old egg, and the latter was isolated from a cDNA library of fifth-instar larval head. Both are high-affinity GABA transporters, having K m values of 16.4 and 9.4 M for MasGAT and TrnGAT, respectively, and have been found by Northern analysis to be abundantly expressed in the CNS (Mbungu et al, 1995;Gao et al, 1999;Sandhu and Gill, unpublished). Whereas the mammalian GATs can be distinguished from one another based on sensitivity to uptake inhibitors cis-1,3-aminocyclohexane carboxylic acid (ACHC) and ␤-alanine, neither MasGAT nor TrnGAT is inhibited by these compounds.…”

mentioning

confidence: 99%

“…Only two insect GABA transporters, from Manduca sexta (MasGAT) and Trichoplusia ni (TrnGAT), having 95% sequence identity to each other, have been cloned and functionally characterized to date (Mbungu et al, 1995;Gao et al, 1999). The former was cloned from a cDNA library obtained from a 3-day-old egg, and the latter was isolated from a cDNA library of fifth-instar larval head.…”

mentioning

confidence: 99%

Immunocytochemical localization of a Manduca sexta γ‐aminobutyric acid transporter

Umesh

Gill

2002

J of Comparative Neurology

View full text Add to dashboard Cite

Gamma-aminobutyric acid (GABA) is a major inhibitory neurotransmitter in insect central and peripheral nervous systems. Although much work has focused on the downstream targets of GABA, signal termination at insect GABAergic synapses has received very little attention. One of the major mechanisms of terminating synaptic transmission involves transport of the neurotransmitter molecules into presynaptic neurons or surrounding glia. Here we report the immunolocalization of a GABA transporter in the tobacco hornworm, Manduca sexta (MasGAT), using an affinity-purified antibody developed to the C-terminus. This is the first demonstration of an insect neurotransmitter transporter immunolocalization study. Results showed strong staining in the neuropil regions of embryonic, larval, and pharate adult central nervous system. Expression pattern in the pharate adult brain mostly mimicked that observed for GABA, with staining in parts of the optic and antennal lobes, mushroom body, lateral protocerebrum, and central complex. Certain longitudinal and lateral connectives of ganglia were observed to have immunostained fibers representing axons. These data support the view that GABA is involved in visual and olfactory processing in the insect brain.

show abstract

“…The first one was obtained from Manduca sexta embryos and designated Mas-GAT (Mbungu et al, 1995); it was functionally tested in Xenopus oocytes by measuring the [ 3 H]GABA transport. The other one, isolated from the cabbage lopper Trichoplusia ni and named TrnGAT (Gao et al, 1999), showed high identity with MasGAT as well as GAT-1; however, it was pharmacologically different from GAT-1 by the inability of cyclic GABA analogs, such as nipecotic acid, to inhibit [ 3 H]GABA uptake by TrnGAT.…”

mentioning

confidence: 99%

Expression and Functional Characterization of GABA Transporters in Crayfish Neurosecretory Cells

et al. 2002

View full text Add to dashboard Cite

The effect of GABA on membrane potential and ionic currents of X-organ neurons isolated from the crayfish eyestalk was investigated. Under voltage-clamp conditions, GABA elicited an inward Na ϩ current followed by a sustained outward chloride current. Sodium current was partially blocked in a dose-dependent manner by antagonists of GABA plasma membrane transporters such as ␤-alanine, nipecotic acid, 1-[2([(diphenylmethylene)imino]oxy)ethyl]-1,2,5,6-tetrahydro-3-pyridinecarboxylic acid hydrochloride (NO 711), and SKF89976-A at concentrations between 1 and 100 M. This current was totally blocked by the combined application of NO 711 (5 M) and ␤-alanine (50 M). We obtained an EC 50 of 5 M and a Hill coefficient of 0.97 for the GABA transport mediated response. These results together with studies of immunolocalization using antibodies against neuronal vertebrate GABA transporters (GATs) indicate the presence of GAT-1-and GAT-3-like proteins in X-organ neurons. To isolate the sustained outward Cl Ϫ current, extracellular free sodium solution was used to minimize the contribution of GAT activity. We concluded that this current was caused by the activation of GABA A -like receptors with an EC 50 of 10 M and a Hill number of 1.7.To assign a functional role to the GATs in the X-organ sinus gland system, we determine the GABA concentration (0.46-0.15 M) in hemolymph samples using HPLC.In summary, our results suggest that a sodium-dependent electrogenic GABA uptake mechanism has a direct influence on the excitability of the X-organ neurons, maintaining an excitatory tone that is dependent on the circulating GABA level.

show abstract

Molecular cloning and functional characterization of a gaba transporter from the CNS of the cabbage looper, Trichoplusia ni

Cited by 19 publications

References 61 publications

Comparative sequence analysis and tissue localization of members of the SLC6 family of transporters in adultDrosophila melanogaster

Comparative sequence analysis and tissue localization of members of the SLC6 family of transporters in adultDrosophila melanogaster

Immunocytochemical localization of a Manduca sexta γ‐aminobutyric acid transporter

Expression and Functional Characterization of GABA Transporters in Crayfish Neurosecretory Cells

Contact Info

Product

Resources

About