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Biomedical subjects

A Freese

Publications and source records attributed to A Freese.

54 records · Page 3Linked to original sources
PubMed1990

Cerebral synthesis and release of kynurenic acid: an endogenous antagonist of excitatory amino acid receptors.

Excitatory amino acid (EAA)-mediated synaptic transmission is the most prevalent excitatory system within the mammalian brain. Activation of EAA receptors has been postulated to contribute to neuronal cell death in stroke, epilepsy, hypoglycemia, and Huntington's disease. Kynurenic acid is an endogenous substance that inhibits EAA receptors and may therefore influence important physiologic and pathologic processes. The release of intracerebrally synthesized kynurenic acid into the extracellular fluid (ECF), where it may act at EAA receptors, has not been established in vivo. We studied the synthesis and release of kynurenic acid in the rat striatum using intracerebral microdialysis coupled with high performance liquid chromatography and fluorescence detection. The basal ECF concentration of kynurenic acid in the rat corpus striatum was 17.1 +/- 1.1 nM. Peripheral administration of the immediate biosynthetic precursor of kynurenic acid, L-kynurenine, resulted in marked dose-dependent increases in striatal ECF concentrations of kynurenic acid, peaking at 2-2.5 hr. The highest dose of L-kynurenine (100 mg/kg), administered peripherally, resulted in a 108-fold increase in plasma kynurenic acid levels and a 37-fold increase in cerebral ECF levels. Peripheral administration of kynurenic acid, at a dose that caused plasma levels to increase 430-fold, resulted in only 4-fold increases in striatal ECF concentrations. The precursor responsiveness of striatal ECF kynurenic acid to peripherally infused L-kynurenine was blocked by the central application (via the dialysis probe) of aminooxyacetic acid, an inhibitor of the immediate synthetic enzyme for kynurenic acid, kynurenine aminotransferase. Administration of L-tryptophan was less effective than L-kynurenine in increasing ECF kynurenic acid concentrations and did so at a considerably later time interval (6 hr).(ABSTRACT TRUNCATED AT 250 WORDS)

Aminooxyacetic Acid↗
PubMed1990

Levodopa delivery from controlled-release polymer matrix: delivery of more than 600 days in vitro and 225 days of elevated plasma levels after subcutaneous implantation in rats.

Parkinson's disease is commonly treated with orally applied levodopa (l-dopa). However, because this treatment modality is associated with a number of undesirable side effects, some due to plasma fluctuations, we have developed a slow-release polymer system that can be used to deliver l-dopa continuously for extended periods of time in vitro (greater than 600 days) and in vivo (at least 225 days) in rats. In vitro l-dopa release was evaluated using polymer matrices with appropriately selected parameters (loading and geometry), and zero-order (linear) release of l-dopa was observed for more than 600 days (in highly loaded, noncoated material first-order kinetics), in some instances in mg quantities per day. This was achieved even in polymer matrices, which did not possess a dissolution limiting barrier. Scanning electron-microscopic analysis suggests that the mechanism of release is dissolution through channels and pores within the polymer matrix. To assess in vivo release, l-dopa was quantified in plasma from rats given s.c. implants of l-dopa polymer matrices using high-performance liquid chromatography. We observed release of l-dopa for a period of at least 225 days after an initial burst of release. Continuous release of l-dopa from s.c. implanted slow-release polymer matrices has several advantages over oral delivery: 1) l-dopa plasma fluctuations are eliminated, 2) patient compliance issues are reduced and 3) the gastrointestinal tract is circumvented, thus requiring a lower dose.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Neuroactive metabolites of L-tryptophan, serotonin and quinolinic acid, in striatal extracellular fluid. Effect of tryptophan loading.

Extracellular fluid levels of the neurotoxin quinolinic acid in the corpus striatum of rats, measured by in vivo microdialysis, were increased in a dose-dependent manner following the intraperitoneal administration of tryptophan. The lowest dose of tryptophan (12.5 mg/kg), equivalent to about 5% of the normal daily intake, increased peak quinolinic acid levels nearly 3-fold. At higher doses of tryptophan (up to 250 mg/kg), concentrations of quinolinic acid increased over 200-fold and exceeded potentially neurotoxic levels (10 microM). In contrast, the increase in extracellular serotonin following even the highest tryptophan dose was small (less than 2-fold). These data indicate that quinolinic acid is present in the extracellular fluid where it may function as a neuromodulator and that it is very responsive to physiological changes in precursor availability.

Animals↗
PubMed1989

Quinolinic acid concentrations in striatal extracellular fluid reach potentially neurotoxic levels following systemic L-tryptophan loading.

Following a systemic tryptophan load, striatal extracellular fluid levels of quinolinic acid in the rat were quantified using intracerebral microdialysis. After an intraperitoneal dose of L-tryptophan (250 mg/kg), quinolinic acid levels in striatal perfusates increased by 230 fold. Peak concentrations of quinolinic acid exceeded 10(-5)M, a concentration previously shown to be neurotoxic in vitro. These results indicate that quinolinic acid is markedly precursor responsive and that its concentration in striatal extracellular fluid may reach neurotoxic levels following an acute tryptophan load.

Animals↗
PubMed1989

Controlled release of dopamine from a polymeric brain implant: in vivo characterization.

Intracerebral microdialysis was used to evaluate the long-term in vivo release of dopamine from ethylene-vinyl acetate (EVAc)-dopamine copolymer matrix discs for up to 65 days following striatal implantation. Dopamine release occurred through a single cavity present on one side of the disc, which was otherwise fully coated with an additional, impermeable layer of EVAc. At 20 days following implantation of the device, extracellular concentrations of dopamine within the striatum reached micromolar levels, over 200-fold greater than control values. Release of dopamine was shown to be stable and maintained for the 2-month duration of the experiment. Histological examination confirmed the biocompatible nature of the implant. There are potential applications of this technology to the treatment of Parkinson's disease and other neurological and psychiatric disorders.

Animals↗
PubMed1989

Controlled release of dopamine from a polymeric brain implant: in vitro characterization.

A biocompatible polymeric matrix system for the long-term controlled release of dopamine has been developed. Solid particles of this bioactive agent were encapsulated in ethylene-vinyl acetate copolymer (EVAc). Following immersion in an aqueous buffer solution, the release rate of dopamine from the polymer matrix was found to depend on the initial concentration of dopamine in the polymer. After coating the matrix devices with an additional impermeable layer of EVAc, constant rates of release were obtained by creating a cavity in this impermeable layer. The observed experiments are consistent with a diffusion-limited model of dopamine release; all the in vitro experimental results were therefore correlated by the effective diffusion coefficient of dopamine through the porous polymer network. These results are discussed in terms of potential design modifications to achieve desired release characteristics for a variety of neuroactive substances, including neurotransmitters or their precursors.

Brain↗
PubMed1987

Stimulation of choline release from NG108-15 cells by 12-O-tetradecanoylphorbol 13-acetate.

The effects of the potent tumour-promoting phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) on phosphatidylcholine (PtdCho) metabolism were investigated in the neuroblastoma X glioma hybrid cell line NG108-15. TPA (100 nM) stimulated by 150-200% the release into the medium of 3H radioactivity from cells that had been pre-labelled with [3H]choline. H.p.l.c. analysis of the medium revealed that TPA stimulated the release of only free [3H]choline (212 +/- 11% of control), without affecting such other labelled metabolites as [3H]phosphocholine and [3H]glycerophosphocholine. This effect was concentration-dependent, with a half-maximal effect obtained at 27.5 +/- 6.8 nM, and was observable as early as 5-10 min after exposure to TPA. The TPA-induced release of [3H]choline into the medium was accompanied by a small and variable decrease in cellular [3H]PtdCho (to 93 +/- 4% of control). However, the radioactivity associated with water-soluble cellular choline metabolites (mainly [3H]phosphocholine and [3H]glycerophosphocholine) remained unchanged. TPA also stimulated the release of [3H]choline derived from [3H]PtdCho that had been produced via the methylation pathway from [3H]methionine. These data suggest that phosphatidylcholine may serve as the source of free choline released from the cells in response to TPA. The possible enzymic mechanisms underlying this response are discussed.

Cell Line↗
PubMed1986

Phosphatidylcholine biosynthesis in the neuroblastoma-glioma hybrid cell line NG108-15: stimulation by phorbol esters.

We have examined the effects of phorbol esters on phosphatidylcholine (PtdCho) metabolism in the neuroblastoma-glioma hybrid cell line NG108-15. 12-O-Tetradecanoylphorbol-13-acetate (TPA), 100 nM, stimulated twofold the incorporation of [3H]choline into PtdCho during 2 h of incubation at 37 degrees C. This effect of TPA was concentration dependent, exhibiting an EC50 of 24.5 +/- 4.4 nM. The effect of TPA was also time dependent and became apparent only after a lag period of 15-30 min. TPA also decreased the incorporation of [3H]choline into water-soluble cellular constituents in a manner whose concentration and time-dependence paralleled the changes observed in PtdCho content. HPLC analysis of this pool revealed that the levels of its major (85-95%) constituent, [3H]phosphocholine, were decreased by 29 +/- 5%, whereas those of [3H]glycerophosphocholine (0.5-2% of the pool) were increased by 84 +/- 4%. PtdCho labeling was also stimulated when cells were pulse labeled with [3H]choline and chased in the presence of TPA. The incorporation of [3H]inositol, [14C]ethanolamine, or [14C]serine into phospholipids was not affected by TPA. The non-tumor-promoting compounds phorbol and 4 alpha-phorbol-12,13-didecanoate (at 100 nM) were completely ineffective in modulating choline incorporation, whereas the biologically active analogs 4 beta-phorbol-12,13-didecanoate and 4 beta-phorbol-12,13-dibutyrate were as effective as TPA. We conclude that tumor-promoting phorbol esters can modulate PtdCho metabolism in neural-derived cells. The mechanisms mediating this effect and the possible involvement of PtdCho metabolism in normal signal transduction events and in the biological actions of tumor promoters are discussed.

Animals↗
PubMed1985

High-performance liquid chromatography of water-soluble choline metabolites.

We have developed a new method for the separation of [3H]choline metabolites by high-performance liquid chromatography. Using this method it is possible to separate, in one step, all of the known major water-soluble choline metabolites present in crude acid extracts of cells that have been incubated with [3H]choline, with baseline or near-baseline resolution. We use a gradient HPLC system with a normal-phase silica column as the stationary phase, and a linear gradient of increasing polarity and ionic strength as the mobile phase. The mobile phase is composed of two buffers: Buffer A, containing acetonitrile/water/ethyl alcohol/acetic acid/0.83 M sodium acetate (800/127/68/2/3), and buffer B (400/400/68/53/79), pH 3.6. A linear gradient from 0 to 100% buffer B, with a slope of 5%/min, is started 15 min after injection. At a flow rate of 2.7 ml/min and column temperature of 45 degrees C, typical retention times for the following compounds are (in min): betaine, 10; acetylcholine, 18; choline, 22; glycerophosphocholine, 26; CDP-choline, 31; and phosphorylcholine, 40. This procedure has been applied in tracer studies of choline metabolism utilizing the neuronal NG108-15 cell line and rat hippocampal slices as model systems. While the compounds labeled in the NG108-15 cells were primarily phosphorylcholine and glycerophosphocholine, reflecting high rates of phospholipid turnover, in the hippocampal slices choline and acetylcholine were the major labeled species. Identification of individual peaks was confirmed by comparing the elution profiles of untreated cell extracts with extracts that had been treated with hydrolyzing enzymes of differing specificities. This HPLC method may be useful in studies of acetylcholine and phosphatidylcholine metabolism, and of the possible interrelationships of these compounds in cholinergic cells.

Acetylcholine↗