Na+-dependent medium-affinity uptake of L-glutamate in the insect epidermis |
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Authors: | H McLean S Caveney |
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Institution: | (1) Department of Zoology, University of Western Ontario, N6A 5B7 London, Ontario, Canada |
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Abstract: | It is proposed that the activity of an epidermal cotransport system for Na+ and dicarboxylic amino acids accounts for the small amounts of L-glutamate and L-aspartate in the otherwise amino-acid-rich blood plasma of insects. This Na+-dependent transport system is responsible for more than 95% of the uptake of these amino acids into the larval epidermis of the beetle Tenebrio molitor. Kinetic analysis of uptake showed that the Na+-dependent co-transporter has medium affinity for L-glutamate and L-aspartate. The K
m for L-glutamate uptake was 146 mol·l-1, and the maximum velocity of uptake (V
max) was 12.1 pmol·mm-2 of epidermal sheet per minute. The corresponding values for L-aspartate were 191 mol·l-1 and 8.4 pmol·mm-2·min-1. The Na+/L-glutamate co-transporter has a stoichiometry of at least two Na+ ions for each L-glutamate-ion transported (n=217). The co-transporter has an affinity for Na+ equivalent to a K
m of 21 mmol · l-1 Na+. Na+ is the only external ion apparently required to drive L-glutamate uptake. Li+ substitutes weakly for Na+. Removal of external K+ or addition of ouabain decreases uptake slowly over 1 h, suggesting that these treatments dissipate the Na+/K+ gradient by inhibiting epidermal Na+/K+ ATPase. Several structural analogues of L-glutamate inhibit the medium-affinity uptake of L-glutamate. The order of potency with which these competitive inhibitors block glutamate uptake is L-cysteatethreo-3-hydroxy-Dl-aspartate >
D-aspartateL-aspartate>
L-cysteine sulphinate >
L-homocysteateD-glutamate. L-trans-Pyrrolidine-2,4-dicarboxylate, a potent inhibitor of L-glutamate uptake in mammalian synaptosomes, is a relatively weak blocker of epidermal uptake. The epidermis takes up substantially more L-glutamate by this Na+-dependent system than tissues such as skeletal muscle and ventral nerve cord. The epidermis may be a main site regulating blood L-glutamate levels in insects with high blood Na+]. Because L-glutamate and L-aspartate stimulate skeletal muscle in insects, a likely role for epidermal L-glutamate/L-aspartate transporter is to keep the level of these excitatory amino acids in the blood below the postsynaptic activation thresholds.Abbreviation ac
acetate
- Ch
choline
- CNS
central nervous system
- cpm
counts per minute
- CDTA
trans-1,2-diaminocyclohexane-N,N,N,N-tetraacetic acids
- HPLC
high performance liquid chromatography
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K
m
Michaelis constant
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n
app
apparent number
- NMG
N-methyl-D-glucamine
- Pipes
Piperazine-N,N-bis-2-ethanesulfonic acid]
- SD
standard deviation
- TEA
tetraethyl-ammonium
-
V
velocity of uptake
-
V
max
maximum velocity of uptake |
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Keywords: | L-glutamate co-transport" target="_blank">Na+/L-glutamate co-transport Neurotransmitter amino acids Insect epidermis Beetle Tenebrio molitor |
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