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D L Martin

Publications and source records attributed to D L Martin.

At least 127 records · Page 7Linked to original sources
PubMed1984

Cl- transport in a glioma cell line: evidence for two transport mechanisms.

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.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1984

Activation of beta-adrenergic receptors stimulates taurine release from glial cells.

Activation of adrenergic receptors in primary cultures of astrocytes and LRM55 glial cells stimulates taurine release. Release is inhibited by the beta-adrenergic antagonists alprenolol and propanolol but not by the alpha-adrenergic antagonist phentolamine. Receptor-mediated taurine release occurs via a specific mechanism, since efflux of other amino acids and metabolites is not affected by the beta-adrenergic agonist isoproterenol.

Alprenolol↗
PubMed1984

Evidence for feedback regulation of glutamate decarboxylase by gamma-aminobutyric acid.

Although feedback control mechanisms for regulating the synthesis of various neurotransmitters have been demonstrated no such mechanism has been described for gamma-aminobutyric acid (GABA) in mammalian brain. Physiological concentrations of GABA inactivated glutamate decarboxylase, the enzyme responsible for GABA synthesis, by converting it to apoenzyme. This inactivation was opposed by the cofactor, pyridoxal 5'-phosphate (pyridoxal-P), and was promoted by ATP. GABA also competitively inhibited the enzyme, and the Ki for inhibition was essentially the same as the concentration of GABA giving the half-maximal rate of inactivation (16 mM). These results provide a mechanism for direct feedback control of presynaptic GABA synthesis and provide further support for the regulation of glutamate decarboxylase in vivo by a cycle of inactivation and reactivation.

Adenosine Triphosphate↗
PubMed1984

Binding of ATP to brain glutamate decarboxylase as studied by affinity chromatography.

The interactions of two forms of porcine brain glutamate decarboxylase (beta-GAD and gamma-GAD) with the effector ATP were studied by affinity chromatography. A third form, alpha-GAD, was only slightly retarded by the affinity matrix and was eluted in the buffer wash. The interaction of GAD with the ATP affinity matrix was qualitatively similar to its interaction with free ATP as reported in previous kinetic studies. The rank order of adenine nucleotides as eluting agents and affinity ligands was ATP greater than ADP greater than AMP. GAD was also eluted by its cofactor, pyridoxal 5'-phosphate, and this was enhanced by 1 mM Pi. In contrast, a high concentration (140 mM) of Pi by itself was required to elute the enzyme. GAD remained active while bound to the affinity column and was eluted in the holoenzyme form by ATP, indicating that the affinity ligand did not bind in the active site and did not displace catalytically active cofactor from the enzyme.

Adenosine Diphosphate↗
PubMed1984

Characterization of L-glutamic acid transport by glioma cells in culture: evidence for sodium-independent, chloride-dependent high affinity influx.

The transport of radiolabeled L-glutamic acid by LRM55 glioma cells in culture was examined. Time course studies indicated that L-[3H]glutamic acid is rapidly accumulated, and then 3H is lost from the cell, presumably in the form of glutamate metabolites. Kinetic analysis of L-glutamate uptake provided evidence for two components of transport. A low affinity component was found to persist at 0 to 4 degrees C and was not saturable, influx being proportional to the substrate concentration. A high affinity component, resolved by subtraction of the influx at 0 to 4 degrees C, followed Michaelis-Menten kinetics having a Km of 123 microM and a Vmax of 2.99 nmol/min/mg of protein. The transport system was highly substrate-specific: At least 27-fold larger concentrations of the most potent analogues--cysteic acid, cysteine sulfinic acid, and L-aspartic acid--were required to compete effectively with glutamate. Second, the system was not severely affected by exposure to inhibitors of oxidative phosphorylation or gamma-glutamyltranspeptidase. Third, only 65% of the high affinity uptake was dependent upon the presence of sodium, the other 35% being dependent upon chloride. These observations were supported by the findings that uptake was only partially inhibited by ouabain and quite effectively reduced by several inhibitors of chloride transport. The results of this study provide information on the properties of low affinity glutamate transport, as well as the first description of sodium-independent, chloride-dependent high affinity glial transport. The high affinity component of influx is stimulated by elevated potassium and inhibited by several pharmacological agents. The sodium independence of a significant proportion of high affinity glutamate transport suggests that glutamate binding studies done in sodium-free medium with intact cells may be confounded by a considerable amount of intracellular uptake.

Animals↗
PubMed1983

Cholera on a Gulf Coast oil rig.

A single case of severe diarrhea on a floating Texas oil rig was followed two days later by what proved to be the largest outbreak of cholera in the United States in over a century. After isolation of toxigenic Vibrio cholerae El Tor Inaba of the typical United States phage type from the index patient's stool, the ensuing investigation detected 14 additional cases of cholera and one asymptomatic infection serologically. Infection was associated with eating rice on the oil rig on a particular day (P = 0.03) when an open valve permitted the rig's drinking-water system to be contaminated by canal water containing sewage (including that from the index patient) discharged from the rig. The rice had been rinsed in the contaminated water after cooking, and before being served it had been maintained at a temperature that allows V. cholerae 01 to multiply. Toxigenic V. cholerae 01 is persisting in the United States, and large common-source outbreaks of cholera can occur if proper sanitation is not maintained.

Adult↗
PubMed1983

Selective inhibition of glial versus neuronal uptake of L-glutamic acid by SITS.

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.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1983

Inactivation of brain glutamate decarboxylase and the effects of adenosine 5'-triphosphate and inorganic phosphate.

The substrate-promoted inactivation of glutamate decarboxylase from hog brain was studied. Inactivation was a slow process that was dependent on the concentration of glutamate. Glutamate-dependent inactivation was not first order but was best described as the sum of two exponential decay processes. At 10 mM glutamate, the half-lives at 30 degrees C were about 6 min for the fast component and 70 min for the slow component. Glutamate-dependent inactivation appeared to be due to the formation of apoenzyme since the rate and extent of inactivation were greatly reduced by the presence of pyridoxal 5'-phosphate (the cofactor, pyridoxal-P). Also, inactivated enzyme could be reactivated by adding pyridoxal-P (Meeley and Martin, 1983). Micromolar concentrations of ATP enhanced glutamate-promoted inactivation in the absence of pyridoxal-P. ATP also enhanced inactivation in the presence of 10 microM pyridoxal-P, but somewhat higher concentrations were required for an equal effect. ATP had little or no direct effect on the enzyme in the absence of glutamate. In the absence of pyridoxal-P, Pi reduced the enhancement of inactivation by 10 microM but not by 750 microM ATP. Glutamate-promoted inactivation, its enhancement by ATP, and the opposition to inactivation by pyridoxal-P and Pi appear to be important in the regulation of glutamate decarboxylase.

Adenosine Triphosphate↗
PubMed1983

Reactivation of substrate-inactivated brain glutamate decarboxylase.

The effects of ATP and inorganic phosphate (Pi) on the reactivation of glutamate apodecarboxylase by its cofactor pyridoxal-5'-phosphate (pyridoxal-P) was studied. Apoenzyme was prepared by preincubation with glutamate. Apoenzyme prepared with glutamate alone was reactivated slowly and incompletely by adding a saturating concentration of pyridoxal-P (20 microM). Reactivation was slightly enhanced by 1-10 mM Pi. Reactivation by pyridoxal-P plus Pi was greatly enhanced by the presence of low concentrations (less than 100 microM) of ATP during the preparation of apoenzyme with glutamate. Reactivation was much lower if Pi was omitted. Enhancement of reactivation by ATP was due to its effect during apoenzyme formation, since ATP did not enhance reactivation if added only during reactivation and since the enhancing effect persisted after the removal of free ATP by chromatography on Sephadex G-25 after apoenzyme preparation and before reactivation. Reactivation was inhibited by high concentrations of ATP (greater than 100 microM), possibly by competition of ATP for the cofactor binding site. Four factors (glutamate, pyridoxal-P, ATP, and Pi) control a cycle of inactivation and reactivation that appears to be important in the regulation of brain glutamate decarboxylase.

Adenosine Triphosphate↗
PubMed1983

Inhibitory effect of triethyltin on taurine transport by glioma cells.

The effects of triethyltin (TET) on the transport of taurine, glutamate, lysine, Na+, K+ (using 86Rb+ as tracer), and Cl- by LRM55 glioma cells were examined. Taurine transport was inhibited by TET at much lower concentrations (IC50 = 2.5 microM) than either glutamate or lysine transport (135 and 110 microM, respectively). TET had no significant effect on Na+, Cl-, or 86Rb+ influx at the low concentrations greater than 100 microM. The failure of low concentrations (less than or equal to 10 microM) of TET to affect ion transport indicated that inhibition of taurine transport was not secondary to effects of TET on ion movements or gradients. This conclusion was supported by the observation that neither ouabain nor furosemide, which do affect ion movement and gradients, strongly inhibited taurine transport. Uncouplers and inhibitors of oxidative phosphorylation (cyanide, 2,4-dinitrophenol, and carbonyl cyanide-m-chlorophenyl hydrazone) also had only small effects on taurine transport, suggesting that inhibition by TET was not secondary to possible effects on oxidative phosphorylation. TET had no effect on the efflux of taurine from LRM55 cells at the low concentrations that inhibit uptake, but it induced a nonspecific increase in membrane permeability at much higher concentrations (greater than 100 microM). Tri-n-propyltin and tri-n-butyltin were also potent inhibitors of taurine transport (IC50 = 2.3 and 11 microM, respectively), but trimethyltin was much less potent (144 microM).

Biological Transport↗
PubMed1983

Multiple forms of glutamate decarboxylase in porcine brain.

Three forms of glutamate decarboxylase from hog brain (termed alpha-, beta-, and gamma-GAD) were separated by hydrophobic interaction chromatography on phenyl-Sepharose, by isoelectric focusing, and by polyacrylamide gel electrophoresis. When rechromatographed on phenyl-Sepharose, each form migrated as a single entity, indicating that the forms are not readily interconvertible. The three forms are not different-sized aggregates of one form, since all three have the same approximate molecular weight (100,000) as determined by Sephadex G-200 chromatography. The pIs of the three forms separated by phenyl-Sepharose were determined by isoelectric focusing. The values obtained (5.3, 5.5, and 5.8 for alpha-, beta-, and gamma-GAD, respectively) were comparable to the pIs of the three peaks of activity observed upon focusing of enzyme that had been subjected to phenyl-Sepharose chromatography. These results indicate that phenyl-Sepharose chromatography and isoelectric focusing separate the same three components. When synaptosomal extracts were analyzed by phenyl-Sepharose chromatography without intervening purification steps, all three forms were present, but the proportion of beta-GAD was somewhat higher and that of gamma-GAD somewhat lower than in the usual preparations.

Animals↗