High- and low-affinity uptake of amino acid transmitters in cultured neurones and muscle cells of the cockroach, Periplaneta americana

Bermúdez, Isabel; Botham, Roger P.; Beadle, David J.

doi:10.1016/0020-1790(88)90089-3

Cited by 13 publications

(8 citation statements)

References 30 publications

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“…In squid axons, the axoplasmic free glutamate concentration is high (25 mM; Deffner, 19611, and the efflux of 14C-L-glutamate from preloaded axons is relatively insensitive to the ionic gradient and to external glutamate concentration behaving like a simple passive leak (Baker and Carruthers, 1984). Electrical stimulation was reportedly without effect on 14C-Lglutamate efflux, which would argue against the hypothesis advanced here, However, in parts of the vertebrate and invertebrate nervous system where glutamate acts as a classical neurotransmitter, high-affinity Na-dependent uptake systems for glutamate are found on both neurons and glia (Balcar et al, 1977;Beadle et al, 1987;Bermudez et al, 1988;Henn et al, 1974;Hertz, 1979;Wheeler and Boyarsky, 1968) and are presumed to be important in the termination of transmitter action. High-affinity uptake systems are also found in the peripheral nerves of invertebrates (Evans, 1973) and in both the giant axon and the sheath (predominantly Schwann cells) of squid (Baker and Carruthers, 1984;Baker and Potashner, 1973).…”

Section: Discussionmentioning

confidence: 58%

Evidence that glutamate mediates Axon‐to‐Schwann cell signaling in the squid

Lieberman¹,

Abbott²,

Hassan³

1989

Glia

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High-frequency stimulation (100 Hz) of isolated giant axons of the small squid Alloteuthis subulata and the large squid Loligo forbesi caused the periaxonal Schwann cell resting potential (Em = -40 mV) to hyperpolarize up to 11 mV in direct proportion to train duration and action potential amplitude. In both species, the Schwann cell also hyperpolarized up to 17 mV with the application of L-glutamate (10(-9) to 10(-6) M), in a dose-dependent manner. By contrast, in the presence of 10(-8) M d-tubocurarine (d-TC) to block the cholinergic component of the Schwann cell response, Schwann cells depolarized 8-9 mV during electrical stimulation of the axon or application of L-glutamate. In the presence of 10(-5) M 2-amino-4-phosphonobutyrate (2-APB), the hyperpolarization to glutamate and to axon stimulation was blocked, whereas the cholinergic (carbachol-induced) hyperpolarization was unaffected. In experiments with Alloteuthis, L-aspartate (10(-7) M) also caused a Schwann cell hyperpolarization, but this was not blocked by 2-APB. In tests with glutamate receptor agonists and antagonists, quisqualate (10(-5) M) produced a hyperpolarization blocked by 10(-4) M L-glutamic acid diethylester (GDEE), which also blocked the response to axonal stimulation. Kainic acid (10(-4) M) also caused a hyperpolarization, but n-methyl-D-aspartate (NMDA; 10(-4) M), ibotenate (10(-5) M), alpha-amino-3-hydroxy-5-methyl-isoxazole proprionate (AMPA; (10(-4) M), and isethionate (10(-5) M) had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Discussionmentioning

confidence: 58%

Evidence that glutamate mediates Axon‐to‐Schwann cell signaling in the squid

Lieberman¹,

Abbott²,

Hassan³

1989

Glia

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“…High affinity uptake mechanisms (low Kin) have a low capacity for accumulating substrate, however. Embryonic insect neurons in culture, for instance, display a high-affinity (Km, 1.4 gmol.1-1) Na+-dependent glutamate uptake with low maximum capacity (Bermudez et al 1988). The epidermal medium-affinity uptake mechanism, however, has a high capacity for glutamate accumulation.…”

Section: Physiological Significance Of Co-transporter Kineticsmentioning

confidence: 98%

“…A 2:1 transport stoichiometry is common for Na § transport of L-glutamate and L-aspartate in arthropod neurons and muscle (Baker and Potashner 1971 ;Evans 1975;Bermudez et al 1988) and in neuronal (Erecinska 1987) and other vertebrate tissues (Fukuhara and Turner 1985), although a 1:1 stoichiometry is found in the peripheral nerves of Carcinus (Evans 1973). At neutral pH, dicarboxylic amino acids are present mostly in the anionic form -OOC(CH2)n CH(NH3+)COO -and are transported as anions (Gazzola et al 1981).…”

Section: Stoichiometry Of the Na+ /L-glutamate Co-transportermentioning

confidence: 99%

“…Glutamate is an excitatory neurotransmitter in skeletal muscle of the larval beetle (Yamamoto and Washio 1980) and single glutamate channels in this tissue are activated when 500 gmol. 1-1 glutamate is applied iontophoretically to the muscle surface (Saito and Kawai 1987 Bermudez et al (1988)], suggesting that the resting concentration of Lglutamate in the neuromuscular synaptic space is considerably lower than in the blood. High affinity uptake mechanisms (low Kin) have a low capacity for accumulating substrate, however.…”

Section: Physiological Significance Of Co-transporter Kineticsmentioning

confidence: 99%

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Na+-dependent medium-affinity uptake of L-glutamate in the insect epidermis

McLean

Caveney

1993

J Comp Physiol B

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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 gmol.1 1, and the maximum velocity of uptake (Fmax) was 12.1 pmol.mm -2 of epidermal sheet per minute. The corresponding values for L-aspartate were 191gmol'l 1 and 8.4pmol.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.1-1 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-cysteate_ threo-3-hydroxy-DL-aspartate > D-aspartate > L-aspartate > L-cysteine sulphinate > L-homocysteate ~> D-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 Abbreviations: ac, acetate; Ch, choline; CNS, central nervous system; cpm, counts per minute; CDTA, trans-l,2-diaminocyclohexane-N,N,N',N'-tetraacetic acids; HPLC, high performance liquid chromatography; Kin, Michaelis constant; napp, 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; Vmax, maximum velocity of uptake Correspondence to: H. McLean 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 Lglutamate/L-aspartate transporter is to keep the level of these excitatory amino acids in the blood below the postsynaptic activation thresholds.

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“…Dissociation resulted in the destruction of the glial cells, thus providing one way to obtain (almost) pure neuronal cultures. Since then, embryonic cockroach brain neurons in primary culture have been used for a variety of biophysical and pharmacological studies, many of which were devoted to the properties of the membrane ionic channels and receptors (Amar et al , 1992Beadle and Lees 1986;Beadle et al 1989;Bermudez and Beadle 1989;Christensen et al 1988;Shimahara et al 1988;Van Eyseren et al 1998). This problem was solved by Beadle et al (1982) with the use of a succession of two different culture media (Beadle and Hicks 1985).…”

Section: Introductionmentioning

confidence: 99%

Quantitative morphological analysis of embryonic cockroach ( Periplaneta americana ) brain neurons developing in vitro

Angevin¹,

Salecker²,

Vaillant³

et al. 2000

Cell and Tissue Research

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Neurons dissociated from the brain of embryonic cockroaches (Periplaneta americana) can be maintained in culture for several weeks. The survival as well as the progressive organization of the neurons into a complex network was studied during a 5-week period under different culture conditions. About 10% of the dissociated cells adhered to the culture dish. This figure remained constant throughout the culture. The cell diameter ranged from 10 to 20 µm and did not change significantly over time in culture. Whereas only a few cells exhibited neurites at the start of the culture, the number of cells exhibiting neurites increased to reach about 99% after 2 weeks. The different cells were then connected to each other, forming a network, which became more and more complex. The number of cells per cluster as well as the length and the diameter of the "connectives" that linked the different clusters were found to increase with time. The morphology of individual neurons within the network was visualized after intracellular injection of biocytin. Labeling with antibodies raised against serotonin or GABA indicated that neurons were able to differentiate and to acquire specific neurotransmitter fates. The serotonergic phenotype was found to appear progressively throughout the culture, in parallel with the formation of the network. Cell density, addition of fetal calf serum, and ecdysone were shown to influence the development of the network.

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High- and low-affinity uptake of amino acid transmitters in cultured neurones and muscle cells of the cockroach, Periplaneta americana

Cited by 13 publications

References 30 publications

Evidence that glutamate mediates Axon‐to‐Schwann cell signaling in the squid

Evidence that glutamate mediates Axon‐to‐Schwann cell signaling in the squid

Na+-dependent medium-affinity uptake of L-glutamate in the insect epidermis

Quantitative morphological analysis of embryonic cockroach ( Periplaneta americana ) brain neurons developing in vitro

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