Faculty Bibliography

The uptake of seven amino acids, alpha-aminoisobutyric acid (AIB), cyclo-leucine (cyclo-Leu), gamma-aminobutyric acid (GABA), glycine (Gly), glutamic acid (Glu), lysine (Lys), and taurine (Tau), representatives of different amino acid transport systems, was studied in slices of brain from Tokay lizards and White Leghorn chicks. In descending order, the rate of the initial uptake of the amino acids in both species was Glu greater than Gly greater than GABA greater than Cyclo-Leu greater than AIB greater than Lys greater than Tau. The substrate specificities and the differences in sodium and temperature dependence of the uptake of the amino acids indicate the presence of several distinct amino acid transport systems, some sodium-dependent and some sodium-independent. The structural specificity of amino acid transport classes in the brain of these species is similar to that in other vertebrate brain preparations

Changes in the activity of proteases (cathepsin D and calpains) caused by 48-h food withdrawal were studied in the brain, liver, kidney, spleen, and heart of 3-, 12-, and 24-month-old Fischer rats. Cathepsin D activity was similar in brain, liver, and heart of control animals; in kidney it was 5-fold higher and in spleen about 10-fold higher. With age, activity increased in all organs tested except spleen. Brief starvation caused no change of cathepsin D activity in brain, but caused an increase in liver and a decrease in spleen. Neutral proteolytic activity in control was highest in the pons-medulla-cerebellum fraction of brain, and activity in liver and heart was below that in brain. Activity increased with age in brain and decreased in other organs. Brief starvation in young animals caused an increase in activity in brain, and a decrease in liver and spleen. Isolated calpain II activity was high in control brain. It increased with age in the cerebrum. Brief starvation resulted in a decrease in the brain. The results indicate that the protease content of the brain is altered with age and in malnutrition, with changes not being the same for all proteases, and changes in brain being different from those in other organs

Using the developing chick embryo as a model and a very sensitive micromethod for amino acid analysis, a complete analysis is presented of the developmental changes in free amino acid concentration in the blood, in the CSF, and in two different brain regions (optic lobe and frontal lobe) of the chick embryo (from day 4 of incubation, until day 5 post hatching). The developmental profile of Lys is the only one that is almost identical in all three compartments. The developmental profiles of the serum and of the brain are very similar for Arg and Phe, less so for Leu and Gly, and towards the end of the embryonic period, similar also for Val, Ile, Trp, and Met. The amino acid concentrations in the CSF are either much lower than in serum and brain already at the earliest stages, or they progressively decline to levels lower than those in brain and serum, most rapidly between day 6 and 8 of embryonic life. The concentrations of neuroactive amino acids (Gln, Glu, Asp, GABA, Tau, and Gly) in both brain regions begin to increase very early, and continue to rise, except Tau, which goes through a maximum at day 8. Comparative analysis of the developmental profiles of each amino acid in serum, brain, and CSF reveals that the blood supply and the cellular uptake, retention, and metabolism by neural cells are the major determinants of the free amino acid pool of the developing brain

The carrier-mediated efflux of [3H]1-methyl-4-phenylpyridine (MPP+) and [3H]dopamine was examined in mouse striatal synaptosomal P2 fractions. Although the two compounds are transported by the same carrier, the translocation of the carrier-ligand complex is more rapid with MPP+ than with dopamine. With dopamine-stimulated efflux of preloaded [3H]dopamine, externally present dopamine at a concentration of 1.3 microM reduced the intrasynaptosomal concentration of [3H]dopamine by 50% (the ECR value) with 8 min of incubation. The ECR value of dopamine in promoting the efflux of [3H]MPP+, however, was only 0.15 microM. Similarly, ascorbic acid was weaker in enhancing the efflux of [3H]dopamine (ECR greater than 2000 microM) than that of [3H]MPP+ (ECR = 567 microM). This effect of ascorbic acid on the efflux of [3H]MPP+ was attenuated by mazindol, a blocker of dopamine uptake. It is proposed that ascorbic acid has a neuromodulatory role involving changes at the level of carrier-membrane translocation and/or orientation

The effect of chronic nicotine on dopamine-agonist-mediated locomotor activity response was measured in BALB/cBy and C57BL/6 mice. Mice were injected twice daily for 10 days with 1.2 mg/kg SC (-)-nicotine di-(+)tartrate. Subsequent locomotor activity response to apomorphine (1 mg/kg SC) was measured. Apomorphine induced hypomotility in both strains of mice, with the BALB/cBy mice showing a greater hypomotility compared to the C57BL/6 mice. The response to apomorphine was attenuated in both strains of mice that were treated previously with repeated injections of nicotine. The results suggest that chronic nicotine may induce changes in the dopaminergic system, which is reflected in altered behavioral response to a dopamine agonist

Brain homogenate, cerebral microvessels, and endothelial cells (ECs) were prepared from 15-18-week-old human fetuses and analyzed biochemically for the presence of elements of the cholinergic system [acetylcholinesterase (AChE), choline acetyltransferase (ChAT), and butyrylcholinesterase]. The ECs were cultured, and their purity was checked by light microscopic immunohistochemistry with the application of anti-human factor VIII and glial fibrillary acidic protein. The highest activity of ChAT was found in the brain homogenate and the lowest in the microvessel fraction. No ChAT activity could be detected in the cultured ECs, despite the presence of high AChE activity. It is suggested that human brain ECs may be under the control of acetylcholine released from cholinergic nerve terminals but that the cells do not produce the transmitter itself. In coculture experiments, when ECs were plated on the upper surface of a polycarbonate filter and glial cells were seeded on the lower surface, the electric resistance was measured. During the culture period, the resistance first increased up to 5 days in vitro (297 +/- 17 ohm.cm2) but later gradually declined. These results demonstrate that human ECs cocultured with glial cells provide a useful model for study of the function of the blood-brain barrier in vitro

Adult male mice, rats, and guinea pigs were subjected to intense sound stimulation of an electric bell (100 dB, 12 kHz for 60 s) after a single intraperitoneal (i.p.) injection of metaphit (1-(1-(3 isothiocyanatophenyl)-cyclohexyl)piperidine) (50 mg/kg). When the animals were tested 24 h after administration of metaphit, audiogenic seizures were observed. None of the control saline-injected animals had convulsions. EEG recordings demonstrated the appearance of paroxysmal activity and spike-wave complexes in the trace from cortical and hippocampal electrodes, with frequency and amplitude increasing with time. Behaviorally, myoclonic jerks of facial muscles, ears, and neck appeared, but no correlation was noted between EEG and the motor phenomena. Auditory stimulation was necessary to elicit the full-blown sequence of seizure responses consisting of wild running followed by clonic and then tonic extension. At the time of seizures, repetitive high-amplitude spikes and waves appeared in the EEG, followed by profound EEG and behavioral depression. None of the animals died during or immediately after seizures. The seizure response to sound stimulation of mice, rats, and guinea pigs was phenomenologically similar, with minor differences in quantitative pattern of convulsive components, which suggests that all three animal species share the common property of extreme susceptibility to audiogenic stimulation caused by metaphit administration

We studied the effect of acetyl-L-carnitine (ALCAR) on dopamine release and the effect of long-term acetyl-L-carnitine treatment on age-related changes in striatal dopamine receptors and brain amino acid levels. In striatal tissue that had been incubated with [3H]dopamine, acetyl-L-carnitine increased the release of [3H]dopamine evoked by electrical stimulation. In striatal tissue from aged mice administered acetyl-L-carnitine for 3 months, the release of [3H]dopamine evoked by electrical stimulation was higher than that of its aged control; the release after a second stimulation was similar in the two groups. There was a significant decline in the number of D1 striatal dopamine receptors with age. The Bmax was 51% lower in 1.5-year-old mice than in 4-month-old animals. Administration of acetyl-L-carnitine for 3 months diminished the reduction in the binding of [3H]SCH-23390. [3H]Spiperone binding to D2 receptors was not decreased with age and was not affected by acetyl-L-carnitine treatment. Age-related decreases in levels of several amino acids were observed in several brain regions. Acetyl-L-carnitine lessened the reduction in the level of taurine only in the striatum. The findings confirm the multiple effects of acetyl-L-carnitine in brain, and suggest that its administration can have a positive effect on age-related changes in the dopaminergic system

To explore the possible therapeutic use of electric convulsive treatment in Parkinson's disease (PD), the authors examined the biochemical effects of electroconvulsive shock (ECS) on dopaminergic systems in a rodent model of PD, induced with the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MPTP increased dopamine turnover, as indicated by an increase in the ratio of the dopamine metabolites dihydroxyphenylacetic acid and homovanillic acid to dopamine. [3H]Spiperone binding to the D2 site increased after lesioning of striatal dopamine terminals. With ECS alone, no changes were found in monoamine levels, brain monoamine oxidase activity, or the D2-labeled sites measured 24 hours after the last treatment. [3H]SCH-23390 binding to the D1 site increased after ECS. In MPTP-treated mice, ECS also increased [3H]SCH-23390 binding to the D1 site, whereas [3H]spiperone binding to the D2 site was unchanged compared to control or to only ECS-treated animals, and decreased compared to the MPTP-treated group that did not receive ECS. ECS appears to selectively modify both the D1 and D2 sites when given after MPTP, increasing the binding of a D1 radioligand and decreasing the binding of a D2 radioligand

The effects of nicotine and dimethylphenylpiperazinium (DMPP) on resting and stimulation-evoked release of [3H]-acetylcholine ([3H]ACh) from cholinergic interneurons and neuro-effector neurons of the ileal longitudinal muscle and the responses of the smooth muscle to nicotinic agonists were studied. (-)-Nicotine was 15 times more effective than (+)-nicotine in releasing ACh. Since tetrodotoxin (1 microM) completely antagonized the effect of nicotinic agonists, the site of action of the nicotinic agonists studied was on the somatodendritic nicotinic receptors. The electrical field stimulation-evoked release was not affected by nicotinic agonists and antagonists, indicating that the axon terminals of cholinergic interneurons are not equipped with nicotinic receptors. This preparation proved to be useful to study the effect of nicotinic agonists on somatodendritic receptors, to determine the affinity constants of nicotinic antagonists, and to characterize these receptors. The rank order of antagonists was d-tubocurarine = mecamylamine greater than pipecuronium greater than pancuronium greater than vecuronium greater than hexamethonium; the apparent affinity constants (KD) were 1.15, 1.55, 3.06, 3.98, 13.59 and 32.88 microM, respectively. alpha-Bungarotoxin had no antagonistic activity at all. This finding indicates that nicotine and the endogenous ligand ACh act via a postsynaptic, somatodendritic nicotinic receptor that is pharmacologically similar to those located on the axon terminals of sympathetic neurons or in ganglions, but is dissimilar to those located at the postsynaptic site of the neuromuscular junction