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Biomedical subjects

A Freese

Publications and source records attributed to A Freese.

At least 19 recordsLinked to original sources
PubMed1992

Basic fibroblast growth factor protects striatal neurons in vitro from NMDA-receptor mediated excitotoxicity.

Basic fibroblast growth factor (bFGF) promotes the survival and outgrowth of neurons. In this study the neuroprotective effects of bFGF were examined in 12-18-day-old cultured striatal neurons exposed to glutamic acid, kainic acid (KA), and quinolinic acid (QA), an N-methyl-D-aspartate (NMDA)-receptor agonist. Results showed that preincubation with bFGF (6 pM) from the day of plating significantly increased the survival of striatal neurons treated for 3 h with glutamate (3 mM) or QA (1 mM), but had little effect on KA (1 mM) induced toxicity. Moreover, maximum protection by bFGF against glutamate neurotoxicity was observed in cultures treated as little as 2 h before glutamate exposure. These results show that bFGF markedly protects striatal neurons from NMDA-receptor induced neurotoxicity.

Animals↗
PubMed1992

Biochemical and behavioral recovery in a rodent model of Parkinson's disease following stereotactic implantation of dopamine-containing liposomes.

We have developed and characterized dopamine-containing liposomes which exhibited in vitro sustained release of dopamine for over 40 days. These liposomes were stereotactically implanted into the partially denervated corpus striatum of rats subjected to unilateral lesions of the substantia nigra. In vivo release of dopamine into striatal extracellular fluid was monitored by microdialysis and behavior was assessed by quantifying apomorphine-induced asymmetric rotation. Extracellular dopamine levels in the partially denervated striatum of the dopamine liposome-treated rats were greater than the levels in the lesioned rats which received control liposomes and these levels remained elevated for 25 days. In parallel, those rats which received dopamine liposomes exhibited partial behavioral recovery, with attenuation of asymmetric rotation following systemic apomorphine administration. These results suggest that dopamine-containing liposomes can partially ameliorate the deficits associated with a rodent model of Parkinson's disease, and demonstrate the potential of this technology as a method for the controlled delivery of therapeutic agents into discrete areas of the brain.

Animals↗
PubMed1991

Infection of cultured striatal neurons with a defective HSV-1 vector: implications for gene therapy.

Several neurological diseases which affect the corpus striatum are candidates for gene therapy. We have developed a defective Herpes Simplex Virus (HSV-1) vector system to introduce genes into postmitotic cells, such as neurons. The prototype vector, pHSVlac, contains a transcription unit which places the E. coli Lac Z gene under the control of the HSV-1 immediate early (IE) 4/5 promoter, a constitutive promoter. We now demonstrate that a HSV-1 vector can deliver a gene into striatal neurons. Infection of cultured rat striatal neurons with pHSVlac virus resulted in stable expression of beta-galactosidase for at least two weeks, without cell death. The potential to replace the Lac Z gene with other genes of interest, such as the gene responsible for Huntington's Disease, once it is isolated, may lead to insights about the pathogenesis of this genetic neurodegenerative disease, and may provide a method for performing gene therapy on this disease. Similarly, introduction of the tyrosine hydroxylase gene, which encodes the rate-limiting enzyme in the conversion of tyrosine to dopamine, into striatal neurons might provide a novel gene therapy approach towards treating Parkinson's Disease.

Animals↗
PubMed1990

Characterization and mechanism of glutamate neurotoxicity in primary striatal cultures.

Excitatory amino acids may play a role in the pathogenesis of cell death in neurodegenerative diseases, including Huntington's disease (HD). In an attempt to develop a tissue culture model for HD, the toxicity of glutamate was examined in primary striatal cultures derived from newborn rats. Morphological criteria were used to determine the toxic effects of glutamate in 6-, 12-, and 18-day-old cultures which were examined before and after 1-3 h of exposure to glutamate. Although younger cultures demonstrated little susceptibility to glutamate relative to controls, the number of neurons in older cultures was significantly depleted in the presence of glutamate. Glutamate toxicity was dose-dependent, with an ED50 of approximately 300 microns glutamate, and a maximal effect was observed within 3 h of initial exposure. Affected neurons demonstrated somal swelling within 1 h of glutamate exposure and disruption of neuritic processes and somal integrity within 3 h. Cell death was significantly increased by raising the extracellular calcium concentration and could be decreased by the addition of magnesium to the incubation medium. Moreover, the N-methyl-D-aspartate (NMDA) receptor agonist, quinolinic acid, showed a toxicity profile similar to that of glutamate. The NMDA receptor competitive antagonist, 2-amino-5-phosphonovalerate (APV) significantly reduced toxicity, albeit incompletely. An additional component of glutamate mediated toxicity in striatal cultures could be explained by activation of non-NMDA receptor subtypes. These in vitro studies indicate that glutamate is toxic to a subset of mature striatal neurons in the absence of a glutamatergic afferent input, and that this toxicity is mediated partially by the NMDA receptor, with an additional component due to non-NMDA receptors.

2-Amino-5-phosphonovalerate↗
PubMed1990

The correlation between excitatory amino acid-induced current responses and excitotoxicity in striatal cultures.

Neuronal excitotoxic injury is initiated by activation of specific excitatory amino acid receptors and is mediated by ion flow through associated ion channels. In a companion paper we defined the age dependence of glutamate toxicity in striatal cultures. In this study we examined the correlation between age-dependent changes in the electrophysiologic responses to glutamate and N-methyl-D-aspartate (NMDA) and the development of excitotoxicity. Currents were recorded in voltage-clamped neurons in response to the pressure application of glutamate or NMDA. We found that 1-week-old cultures were resistant to excitotoxic effects of 1 or 3 mM glutamate (Freese et al., companion paper). Neurons from cultures of this age often did not exhibit current responses when exposed to NMDA (10 microM or 100 microM). However, most neurons were responsive to 10 microM glutamate. After 12 days in culture the current responses induced by NMDA were more prevalent and shifted toward higher amplitudes. These changes in NMDA current responses coincided temporally with major increases in excitotoxic sensitivity. However, we then found that excitotoxic susceptibility doubled between 12 and 18 days in culture without any parallel increase in glutamate- or NMDA-induced currents. Additionally, neurons which survived 1 mM glutamate exposure for 3 h were found to exhibit glutamate and NMDA responses. These data show that the expression of excitatory amino acid-induced currents is a prerequisite for excitotoxicity, but that additional factors must also contribute to the full maturation of in vitro neuronal vulnerability.

Animals↗
PubMed1990

Extended levodopa release from a subcutaneously implanted polymer matrix in rats.

It is well recognized that plasma fluctuations resulting from oral levodopa therapy may cause an unstable clinical response in parkinsonian patients. We have therefore developed a slow-release polymer matrix system that can deliver levodopa continuously for extended periods of time (at least 225 days) after subcutaneous implantation in rats. Advantages of this approach include (1) the elimination of levodopa plasma fluctuations and (2) the possibility of reducing the required dose due to constant plasma levels and because the gastrointestinal tract is circumvented. The peripheral implantation of polymer systems containing levodopa, dopamine receptor agonists, or other anti-Parkinson agents may constitute a novel technology of drug delivery to improve the care of patients with Parkinson's disease.

Animals↗
PubMed1990

Infection of cultured central nervous system neurons with a defective herpes simplex virus 1 vector results in stable expression of Escherichia coli beta-galactosidase.

We have developed a defective herpes simplex virus (HSV) vector system that permits the introduction of virtually any gene into mammalian central nervous system neurons. The prototype vector, pHSVlac, contains a transcription unit that places the Escherichia coli lacZ gene under the control of the HSV-1 immediate early 4/5 promoter. pHSVlac was propagated using the HSV-1 temperature-sensitive mutant ts K as helper virus. Infection of rat neurons in primary culture derived from various regions throughout the central nervous system, including spinal cord, cerebellum, thalamus, basal ganglia, hippocampus, occipital cortex, temporal cortex, and frontal cortex, resulted in stable expression of high levels of beta-galactosidase for at least 2 weeks, without cell damage. Since other genes can be expressed from pHSVlac, HSV-1 vectors may prove useful for delivery of genes into central nervous system neurons for studies on nervous system physiology or to perform gene therapy for neurological conditions.

Animals↗
PubMed1990

Kynurenine metabolites of tryptophan: implications for neurologic diseases.

Over the past 2 decades, a number of studies have demonstrated that amino acids act as precursors for the biosynthesis of a variety of neuroactive compounds, including catecholamines and indoleamines. For example, the aromatic amino acid L-tryptophan is a precursor for serotonin biosynthesis. Based on this observed precursor relationship, dietary tryptophan supplementation is used to treat a number of neurologic disorders attributed to alterations in serotoninergic neurotransmission. Recent studies have revealed that, in addition to serotonin, a number of neuroactive compounds, the kynurenines, are metabolities of tryptophan. Of these, perhaps the most important is quinolinic acid, a neurotoxin that acts at the N-methyl-D-aspartate (NMDA) receptor and whose precursor responsiveness to tryptophan far exceeds that of serotonin. In the central nervous system, kynurenines, and in particular quinolinic acid, may modulate excitatory amino acid transmission, and may act as neurotoxic agents implicated in the pathogenesis of several neurologic diseases.

Animals↗
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↗