Faculty Bibliography

Amino acid uptake by slices of brain is greatly diminished by incubation of the slices in glucose-free media. Uptake can be restored by the addition of a number of substrates, e.g., lactate, citrate, or oxaloacetate. The mixture of succinate, malate, and pyruvate (SMP, 20, 5, and mM) restored amino acid uptake better than glucose after brain slices were incubated in glucose-free media to deplete endogenous energy stores. The degree of restoration of uptake was different with the various amino acid transport classes and was independent of the restoration of ATP levels in the tissue. After restoration of uptake with SMP the amino acid uptake was resistant to NaF, but was markedly more sensitive to arsenite and oligomycin. The results indicate the coupling of mitochondrial energy transducing systems to transport

The incorporation of tyrosine into proteins was measured after the subcutaneous implantation of a pellet of [14C]tyrosine in mice. This method keeps the specific radioactivity of free tyrosine fairly constant and makes it possible to follow incorporation up to a 10-day period. At the end of 10 days most of the protein-bound tyrosine was replaced (i.e. most protein turned over) in lung, liver, heart, kidney and spleen; about half was replaced in brain, one-quarter in muscle. The rate of protein turnover in myelin was approx. 40% of that of whole brain proteins; at 10 days one-fifth of the myelin proteins were replaced. All protein components of myelin measured were in a dynamic state; incorporation decreased in the following order, Wolfgram greater than DM-20 greater than basic greater than proteolipid proteins. The incorporation of tyrosine into each protein fraction was greater in the 0-5-day than in the 5-10-day period, indicating heterogeneity of metabolic rates. The results show that after myelination at least a portion of each protein component of myelin is undergoing significant metabolic turnover. In the adult, myelin components are not stable, but turnover is heterogeneous, and each protein may be compartmentalized. Turnover can be influenced by a variety of factors

The rate of appearance of label in the brain in mice following the intraperitoneal or intravenous injection of tracer doses of amino acids was measured in short time periods (1-8 min). Amino acid flux varied between 2 and 10 nmol/min per g brain for the amino acids used. Defining half-life as the uptake of labeled amino acid amounting to 50% of endogenous levels, a short half-life (between 3 and 30 min) was found for the essential amino acids. The half-life of the nonessential amino acids varied between 2 and 24 h, depending on their level in brain. Flux (exchange) of an amino acid was increased when the level of amino acids belonging to the same transport class was increased by intracerebral injection. Protein-free diet resulted in decrease in some amino acids, increase in others; flux was altered parallel to changes in brain levels in animals on this diet. The stercospecificity of exchange and the substrate specificity of effects of altered brain amino acids indicate that exchange occurs via mediated transport. Mediated exchange was present in immature brain. Heteroexchange (flow of one amino acid causing the counterflow of a related amino acid) may play an important part in cerebral homeostasis.

Calf brain prolidase covalently bound to CNBr-Sepharose 4B, retained about 32% of the activity of the uncoupled enzyme. The free enzyme showed slightly greater stability than the bound preparation when stored at 20 degrees C or at 0 degrees C. However, in either case the free and bound enzymes were more stable at the lower temperature. Greater thermal stability was shown by the free enzyme than by the bound preparation over a temperature range of 25 degrees C-60 degrees C. The free and bound prolidase, with and without Mn+2, had maximal activity at pH 4.0. Although the bound enzyme showed a single maximum, the free preparation exhibited three pH maxima of 4.0, 9.0, and 6.5, in decreasing order of activity. The ions Ag+, Cu+, Hg+2, and Zn+2 were strongly inhibitory on the free enzyme, whereas inhibition of the bound enzyme, with the exception of Zn+2 , was less. Unlike the coupled enzyme, a stimulatory effect was obtained on the free preparation with Co+3, Mg+2, and Mn+2. Various other compounds were studied and their effects were noted

Subcellular studies of choline uptake of rat striatum indicated a correspondence between the Na(+)-dependent uptake and choline acetyltransferase (ChAc), whereas there was a lack of correspondence between the Na(+)-independent uptake and ChAc. Subcellular studies also showed a correspondence between the Na(+)-dependent uptake and hemicholinium-3 inhibition, and more important, particles that accumulate choline were shown to consist of at least two subcellular populations. A comparison was made of kinetic data from three areas of the rat brain: corpus striatum, cerebral cortex, and hypothalamus. Taken together, our data on choline uptake give added support to the idea that the Na(+)-dependent choline transport is concentrated in the striatum and specifically related to cholinergic nerve endings. Morphine and methadone in vitro inhibited the Na(+)-dependent choline uptake. In vivo morphine induced a significant lowering of theV max in the rat cerebral cortex, but not in the striatum. This finding is consistent with the known action of morphine on acetylcholine turnover.

Pentoxifyline at 1 mM had no effect on [14C]isoleucine incorporation into mouse brain tissue suspension. At 5-20 mM, this compound inhibited incorporation. The inhibition was prompt, and it was reversible. Aminophyline at 3-12 mM produced inhibition, but theophyline at 2-16 mM had no effect. Pentoxifylline inhibited the incorporation of uridine into brain RNA to the same extent and with a similar time course as its effect on protein synthesis