Faculty Bibliography

Some moderate and extreme thermophilic bacteria grew well on media other than the recommended basic media. Attempts to induce phenylalanine hydroxylase (Phe H) in the various thermophiles as well as mild thermophily in the mesophiles Pseudomonas sp ATCC 11299a and Chromobacterium violaceum ATCC 12540 were unsuccessful. Evidence is presented indicating that the enzyme in the latter two organisms may be membrane bound. The level of Phe H activity induced was not always consistent with the level of the inducer phenylalanine (Phe) in the growth medium

The effects of lithium and sodium were studied in the corpus striatum and cerebral cortex of rats. Lithium was inhibitory at low concentrations but at 20 mM it increased the binding of [G-3H]naloxone (specific activity 15.6 Ci/mmol). Sodium stimulated the high-affinity binding of this compound. Membranes obtained from the rats treated with lithium showed lower specific binding of both [3H]naloxone and [3H]DHM. Binding of [3H]d-alanine Leu-enkephalin was not changed in the brains of lithium-treated rats, but that of [3H]-spiroperidol was lowered. Cerebral cortex and striatum of lithium-treated rats had a decreased apparent dissociation constant and a lower receptor concentration of naloxone binding sites

Protein degradation rates were measured in brain slices prepared from rats of various ages. This was done by adding the protein synthesis rate, determined by incorporation of a labeled precursor, and the net protein degradation rate, determined by measuring the changes with time of total free amino acids. These rates are about 30% higher than those previously calculated from data on protein synthesis rates and protein accumulation rates in vivo. The protein degradation rates in brain slices diminish with age; i.e., 2-day cerebellum greater than 2-day cerebral hemisphere greater than 12-day cerebral hemisphere greater than young adult cerebral hemisphere. Protein degradation rates in slices from young brain are initially slightly higher than protein synthesis rates, resulting in a small net degradation with time. Unlike slices from adult brain, the protein degradation rates in slices from young brain decline only modestly with time for as much as 100 min of incubation. The characteristics of protein degradation in brain slices from young animals are roughly similar to some of the data calculated for protein degradation in vivo, suggesting that this system may prove useful for studying factors which control or affect brain protein degradation

Kinetic experiments indicate that association of [3H]-cocaine to its binding site in brain occurs rapidly (seconds). Dissociation of membrane-bound cocaine is also rapid, with a dissociation half-life in seconds; this raises the question of whether membrane-bound cocaine is released during the time required for rapid filtration of tissue-containing filters. Results from experiments with increasing numbers of filter-washes indicate that there is no significant loss of [3H]-cocaine saturably bound to brain membranes within the time-scale of the rapid filtration procedure. In addition, saturation analysis of binding data obtained with the filtration procedure and with the centrifugation method give similar estimates of the affinity (dissociation constant: 0.8 microM) and the maximal binding (5 pmol/mg of protein) of cocaine. However, the nonspecific binding and the experimental error in the saturable binding are considerably greater in centrifugation assays than in filtration assays

(+/-)-[3H]Nicotine was bound saturably to crude particulate, and synaptosomal-mitochondrial fraction from mouse brain. Scatchard and Hill plots of the binding data are in agreement with the existence of two independent classes of binding sites with high (Kd of 0.1-0.4 microM) and low(Kd is 20 microM) affinities, although negative cooperativity or a two-step model of ligand-receptor interaction cannot be ruled out. Nicotinic or muscarinic agonists and antagonists had little or no affinity for the nicotine binding sites, suggesting that nicotine binds in brain to noncholinergic sites. The binding did not display stereospecificity; this is consistent with the similarity in the pharmacological effects of (-)- and (+)-nicotine. Our results indicate that binding studies with [3H]nicotine should be interpreted with extreme caution

Naloxone strongly inhibited the breakdown of Met- and Leu-enkephalin when the substrate was incubated with brain homogenate, supernatant, or partially purified soluble aminopeptidase, but it had no effect on the mitochondrial fraction. Arylamidase was also inhibited by naloxone. The inhibitory effect of naloxone on the soluble aminopeptidase was in a linear relationship with concentration in the range of 8-500 microM. The KI for Met-enkephalin is 0.6 mM and for Leu-enkephalin is 2.0 mM. When the naloxone was administered intraperitoneally to the mice, its effects were somewhat different from its effects in vitro. It inhibited the brain catabolism of Met-enkephalin but not of Leu-enkephalin or Tyr-beta NA in vitro. In vivo, acute morphine in mice decreased the Met-enkephalin hydrolysis but increased the Leu-enkephalin hydrolysis. Chronic morphine (by morphine pellet implantation) increased the breakdown rate of enkephalin and of Tyr-beta NA. Naloxone (1 mg/kg) could not reverse the effects of morphine, although abstinence syndrome and stereotypical jumping were precipitated. Naloxone decreased the Leu-enkephalin level in the control mice but did not affect it in the addicted mice

The effects of insulin, hydroxybutyrate, deoxypyridoxine, chlorpromazine, codeine, morphine, puromycin, and cycloheximide on the composition of the free amino acids in mouse and rat brain were tested. Significant changes occurred in a number of amino acids with most compounds tested; the largest was of alanine (a 50% increase with glucose, a 50% decrease with drugs); histidine was often increased, and the nonessential amino acids were mostly decreased. The pattern of changes was somewhat different in the mouse brain from that in the rat brain. Changes of amino acid levels may participate in the pharmacological action of a number of compounds