Faculty Bibliography

Nicotine is among the compounds that enter the brain very rapidly (blood-flow-limited). It also leaves the brain rapidly; by five minutes, 90% exits, an exit somewhat slower than that of water. In spite of rapid exit, brain levels remain higher than levels in blood over a wide range of blood concentrations. Nicotine enters the fetal circulation from the maternal blood; it enters fetal brain but to a smaller extent than adult brain. Nicotine entry is different from that of amino acid: No interaction of amino acid transport and nicotine could be detected. Most close analogs have no effect on nicotine uptake, but at higher concentrations nicotine uptake is saturable. Nicotine and morphine mutually inhibit each other's uptake. The results suggest an uptake compartment (lipid space) for nicotine shared by morphine

A sensitive and convenient method of endopeptidase assay using as substrate globin modified with pyridoxal-5-phosphate was used for determination of acid proteinases in bovine hypothalamus separated by isoelectric focusing. The soluble protein fraction of hypothalamus upon elution from Sephadex gave five peaks of proteinase activity at pH 3.2. The properties indicate that these peaks of endopeptidase activity are isoenzyme forms of cathepsin D

Rat liver phenylalanine hydroxylase (PheH) was covalently coupled to AH-Sepharose 4B, CH-Sepharose 4B, alginic acid and polygalacturonic acid. The activities of the bound enzyme from the ethanol and ammonium sulfate fractions were studied under a variety of conditions. The ethanol enzyme coupled to AH-Sepharose 4B showed the best thermal stability from 20 degrees to 50 degrees after heating for 15 min. It retained more than 15% of its initial activity after storage at 25 degrees for 9 days. The covalently linked enzymes generally had a broader range of optimal activity from pH 5.8 to 7.5. The presence of a positive or negative microenvironment on the matrix had no effect on the activity of the enzyme coupled to AH- or CH-Sepharose 4B. The failure to obtain hydroxylase activity with enzyme linked to alginic acid or polygalacturonic acid was attributed to the acidic microenvironment of the matrices

We determined the regional and subcellular distribution of prolidase in rat brain and its changes with development. The most rapid changes in enzyme activity occurred perinatally, with a maximum level of activity 2 days before birth and a minimum 1 day after birth. Of the 7 regions examined, cerebellum had the highest enzyme level, followed by medulla. The lowest levels were found in the hypothalamus in the adult and in the midbrain in the young. Polidase was mainly soluble; over 55% was recovered in the S2fraction, and the rest was released from the particulate fractions by hypotonic shock. Brains of male rats contained a slightly higher level of the enzyme

Optimal conditions have been determined for the coupling of rat liver phenylalanine hydroxylase (Phe H) to cyanogen bromide-Sepharose 4B. When 8 mg of ligand was reacted with 100 mg of matrix, 20 to 30 percent of the initial enzyme activity was covalently bound along with 90 percent of the protein. The coupled enzyme showed greater thermal stability from 40 degrees to 60 degrees, a broader base of optimal pH activity from 5.8 to 10.0, more resistance to proteolysis and less inhibition by various inhibitors. The uncoupled enzyme exhibited greater storage stability at 25 degrees after 24 hr and at 0 degrees after 18 days. Alteration of the microenvironment by introduction of sulfhydryl groups or of carriers having positive or negative charges had variable effects on the hydroxylase activity

The rate of protein degradation was estimated in several regions of rat brain at various ages by subtracting the rate of accumulation of protein from the rate of synthesis. The rate of degradation in cerebral hemisphere, which was 1.3%/h at 2 days of age, declined steadily with age, approaching the synthesis rate is about 30 days of age (0.8%/h). Degradation rates in the pons medulla, mid-brain and spinal cord were of a similar order to that in the cerebral hemisphere. The cerebellum had an exceptionally high rate of degradation in young rats, 1.9%/h at 2 days of age, which complemented its high rates of synthesis and accumulation. The degradation rate in the young was 2-2.5 times the rate in older rats and was approx. 65% of the synthesis rate during the more active phase of growth. The rapid accumulation of protein in the nervous system during the first week post partum was accompanied by high rates of breakdown, and was the result of a relatively small difference between that high rate of degradation and an even higher synthesis rate