4-acetamido-4--isothiocyanatostilbene-2-2--disulfonic-acid and Glioma

Previously, we reported on the distinct effects of bioactive lysophospholipids, including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), and sphingosylphosphorylcholine (SPC), on membrane potentials in rat C6 glioma cells. In the present report we have tested lysophosphatidylserine (LPS), another bioactive lysophospholipid, on membrane potentials in the same cell line. Membrane potentials were estimated by measuring the fluorescence changes of DiBAC-loaded glioma cells. LPS largely increased membrane potentials in a concentration-dependent manner. The LPS-induced membrane potential increases were not affected by treatment with pertussis toxin, implying no involvement of Gi/o proteins. In contrast to other lysophospholipids, the LPS-induced membrane potential increase was not diminished by a Na(+)-free media but was enhanced by suramin. Furthermore, this change was blunted by EIPA, an inhibitor of Na(+)/H(+) exchanger, but not by SITS, a specific inhibitor of bicarbonate transporter. Our observations suggest that LPS acts on membrane potentials in a unique manner in the C6 glioma cells, although the precise action mechanism requires additional investigation.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; Amiloride; Animals; Cell Line, Tumor; Glioma; GTP-Binding Proteins; Lysophospholipids; Membrane Potentials; Rats; Receptors, G-Protein-Coupled

Studies with brain slices have provided evidence that synthesis of kynurenic acid (KYNA) from kynurenine (KYN), which occurs in astrocytes, is modulated by changes in the ionic composition of the medium and the presence of depolarizing agents or the excitatory amino acid glutamate (Glu). The present study analyzed the effects of changes in incubation medium on KYNA synthesis in cultured C6 glioma cells. The synthesis was not affected by omission of Na(+) and raising K(+) concentration to 50 mM, conditions that in brain slices stimulate or inhibit KYNA formation, respectively. KYNA synthesis in C6 cells was inhibited by the absence of Ca(2+), which contrasts with its Ca(2+) independence in brain slices. Also, lack of Mg(2+) and addition of a chloride channel blocker, 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonate (SITS), did not affect the synthesis. KYNA synthesis in C6 cells was dose dependently inhibited by Glu. The inhibitory effect of Glu was not affected by GDPbetaS, an antagonist of metabotropic Glu receptors, the receptor class prevailing in C6 cells, suggesting that Glu acted intracellularly. NH(4)Cl and veratridine decreased KYNA production, mirroring the effects noted in brain slices. KYNA synthesis was strongly reduced in the presence of leucine (Leu), and the uptake of [(14)C]Leu was inhibited by the KYNA precursor KYN, which points to Leu as a potential endogenous modulator of KYNA formation in CNS cells.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; Animals; Buffers; Calcium; Glioma; Glutamic Acid; Guanosine Diphosphate; Kynurenic Acid; Kynurenine; Leucine; Magnesium; Potassium; Rats; Receptors, Metabotropic Glutamate; Sodium; Thionucleotides; Tumor Cells, Cultured; Veratridine

Cl- transport was studied in the glioma cell line LRM55 . Our results indicate that LRM55 cells contain two major Cl- transporters, an anion exchanger and a K+/Cl- cotransporter , and that these are similar to the Cl- porters found in primary cultures of astrocytes. The exchanger was studied by measuring fluxes of Cl- (as 36Cl-). 36Cl- flux was trans-stimulated by Cl- or HCO-3 and was inhibited by SITS or furosemide. The K+/Cl- cotransporter was studied by measuring fluxes of 36Cl- and K+ (as 86Rb+). External K+ stimulated 36Cl- influx, and external Cl- stimulated 86Rb+ influx. Furosemide, but not SITS, inhibited the K+/Cl- cotransporter . As in primary cultures of astrocytes, the steady-state concentration of Cl- in LRM55 cells was higher than that predicted from passive equilibration according to the membrane potential. LRM55 cells appear to be a good in vitro model for glial Cl- transport.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; Animals; Astrocytes; Bicarbonates; Cell Line; Chlorides; Furosemide; Glioma; Ion Channels; Kinetics; Membrane Potentials; Potassium; Rats; Sodium; Spinal Cord Neoplasms; Temperature

SITS, an inhibitor of anion exchange, was found to be a potent and selective inhibitor of L-glutamic acid uptake by cultured LRM55 glioma cells and rat brain astrocytes. Synaptosomal uptake of glutamate was relatively insensitive to inhibition by SITS. This differential effect indicates that the glutamate transport system in glia differs from that in neurons and that SITS may provide a tool for investigating the exclusive neuronal transport and metabolism of L-glutamic acid.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; Animals; Astrocytes; Brain; Cell Line; Glioma; Kinetics; Neoplasms, Experimental; Neuroglia; Neurons; Rats; Rats, Inbred Strains; Spinal Cord Neoplasms; Stilbenes; Synaptosomes