Faculty Bibliography

Des-tyrosine-gamma-endorphin (DT gamma E) has been reported to alleviate symptoms of schizophrenia. Attempting to replicate those reports, we administered 1 mg of DT gamma E, i.m., for 8 consecutive days to nine patients meeting the DSM-III criteria for schizophrenia. Patients in this double-blind, crossover, and placebo-controlled study showed a statistically significant, but clinically modest improvement. The improvement was detectable during the first several days of the DT gamma E treatment; the symptoms then returned to baseline level in spite of continued doses of DT gamma E. Testing the metabolism of DT gamma E in the patients' plasma, we found a high rate of formation and of degradation, but the metabolic rates were not related to clinical symptoms

Using synaptosomal rat brain membranes, the degradation of Met-enkephalin in both free and receptor-bound form was measured, together with the dissociation of Met-enkephalin from the receptors. The results show that the degradation rate of initially receptor-bound Met-enkephalin is significantly smaller than either the rate of dissociation from the receptors or that of the degradation of free Met-enkephalin. These data suggest that intact Met-enkephalin dissociates from the receptors and then it becomes a target for the same membrane-associated peptidases that split free Met-enkephalin. This view is supported by the good fitness of the measured degradation of initially receptor-bound Met-enkephalin to the predicted degradation curve calculated from the dissociation rate of receptor-bound Met-enkephalin and the degradation rate of free Met-enkephalin

The microbial peptides amastatin and bestatin as well as several dipeptide analogues of the latter exerted little or no inhibitory effect on enkephalin hydrolysis by an aminopeptidase purified from the thermophiles Thermomonospora fusca, ATCC 27730 (Tf) and Thermus thermophilus, ATCC 27634 (Tt). The enzyme catalyzes the cleavage of the tyrosyl-glycyl bond of leucine- and methionine-enkephalin. Intermediate compounds having the same amino acid sequence as the parent substrate disclosed that the residual tetrapeptide can be further degraded to its constituent parts. Each preparation also hydrolyzes to varying extents neutral dipeptides, tripeptides, tetrapeptides, can be further degraded to its constituent parts. Each preparation also hydrolyzes to varying extents neutral dipeptides, tripeptides, tetrapeptides, and larger molecules containing the Met-enkephalin sequence. The Tf enzyme has a pH optimum of 7.5, Km of 667 microM and Vmax of 92 nmol/min/mg of protein; the Tt enzyme, with a pH optimum of 7.2 has a Km of 400 microM and Vmax of 33 nmol/min/mg of protein. Activated by dithiothreitol (DTT) and inactivated by p-chloro- and p-hydroxymercuribenzoate, both are sulfhydryl enzymes. The activity lost by hydrolysis against EDTA can be restored, wholly or in part, by Co+2, Mg+2, and Mn+2; ions with an inhibitory effect were A1+3, Cd+2, Cu+2, Hg+2, and Zn+2. The enzymes are not glycoproteins since they pass unretained through a Con A-Sepharose column

[3H]Imipramine was accumulated by rat hypothalamic slices mostly by a passive low-affinity process. Accumulated [3H]imipramine was released by 60 mM potassium, but the release was small and was not Ca2+-dependent. Imipramine induced some release of [3H]norepinephrine. None of these events were abolished by pretreatment of the rats with reserpine. These results do not favor the view that imipramine acts as a false transmitter in the rat hypothalamus. Rather, imipramine appears to be accumulated in an extragranular pool from which it releases extragranular norepinephrine

Three lines of evidence are brought forward in support of an association in the brain cortex of some, but not all, of the cocaine binding sites with serotonergic nerve terminals. The first is based upon the significant correlation observed between the inhibition of cocaine binding by various drugs and the inhibition of neuronal uptake of serotonin in the mouse cerebral cortex. The second is based upon the demonstration of cocaine binding in human blood platelets, a model system for central serotonergic neurons. The third comes from experiments in which rats were treated with p-chloroamphetamine and 5,7-dihydroxytryptamine (serotonin neurotoxins), 6-hydroxydopamine (catecholamine neurotoxin), or p-chlorophenylalanine (inhibitor of tryptophan hydroxylase). Only the serotonin neurotoxins decreased the binding of [3H]cocaine in the rat cerebral cortex, but to a lower extent than the binding of [3]imipramine, which is known to be associated with serotonergic terminals. In contrast to the cocaine binding in the mouse cerebral cortex, the binding in the rat cerebral cortex included a considerable portion of low-affinity binding that was relatively unaffected by lesions of serotonergic neurons

A membrane-bound aminopeptidase was purified from rat brain, and its activity was assayed by high-pressure liquid chromatography with Met-enkephalin as the substrate. The enzyme was extracted with 1% Triton X-100 and purified by chromatography, successively on DEAE-Sepharose CL-6B, Bio-Gel HTP, and Sephadex G-200 columns. The overall purification was about 1200-fold, with 25% yield. The purified enzyme showed one band on disc gel electrophoresis and two bands on sodium dodecyl sulfate electrophoresis with molecular weights of 62 000 and 66 000. The aminopeptidase has a pH optimum of 7.0, a Km of 0.28 mM, and a Vmax of 45 mumol (mg of protein)-1 min-1 for Met-enkephalin. It releases tyrosine from Met-enkephalin, but it does not split the byproduct. It does not hydrolyze gamma- or beta-endorphin, or dynorphin, but it does hydrolyze neutral and basic aminoacyl beta-naphthylamides. The enzyme is inhibited by the aminopeptidase inhibitors amastatin, bestatin, and bestatin-Gly. Its properties, such as its subcellular localization, substrate specificity, pH optimum, and molecular weight, distinguish it from leucine aminopeptidase, aminopeptidase A, aminopeptidase B, aminopeptidase M, and the soluble aminopeptidase for enkephalin degradation

The anticonvulsive effects of GABA, taurine, and glycine were investigated on several chemically-induced and genetic seizure models. Intravenous injections of either GABA, taurine, or glycine provided protection against 3-mercaptopropionic acid (MPA)-induced convulsions in adult Swiss mice. GABA was partially effective against isonicotinic acid hydrazide and was without effect against bicuculline-induced convulsions. Prolonged administration of glycine prevented MPA-induced convulsions but not electrically induced seizures or seizures induced by strychnine or metrazol. Intragastric glycine protected young audiogenic seizure-susceptible DBA/2 mice against all three phases of sound-induced convulsions (wild running, clonic and tonic seizure), but GABA and taurine provided little or no protection. With increase of glycine, the cerebral levels of glutamine and serine also increased, but that of glutamic acid decreased. The endogenous glutamic and glycine levels were slightly higher in the brains of the audiogenic seizure-susceptible DBA/2 mice than in that of the resistant BALB/Cy strain

The specific [3H]spiperone binding by sheep caudate nucleus homogenate is increased by divalent cations. The effect of Ca2+ or Mn2+ (5 mM) is temperature-dependent, and it is optimal at about 37 degrees, but is relatively low below 15 degrees and above 50 degrees. In the absence of added Ca2+ or Mn2+, the maximal specific [3H]spiperone binding is observed at about 25 degrees, and the cations shift the optimum to about 37 degrees. Under the experimental conditions used, the KD is about 0.6 nM and is not influenced by Ca2+ or Mn2+, or by temperature (25 and 37 degrees). In addition to Ca2+ and Mn2+, Mg2+ and Zn2+ also increase the specific [3H]spiperone binding, but to a smaller extent. At the concentrations of Ca2+, Mn2+, Mg2+ and Zn2+ which produce a maximal increase in the [3H]spiperone binding, the membranes are nearly saturated with the cations which bind about 100 nmoles of Ca2+ or Mg2+/mg of protein, 170 nmoles Zn2+/mg of protein and at least 300 nmoles Mn2+/mg of protein. It is suggested that the cations increase the [3H]spiperone binding by either exposing more binding sites, by preventing denaturation or by increasing the solubility of [3H]spiperone in the membrane phase, or by a combination of these processes

Binding of [3H]imipramine in mouse cerebral cortex was found to be nonhomogeneous. Competition experiments, Scatchard analysis, and Hill plots are compatible with the existence of binding with high (nanomolar) and low (micromolar) affinity. Low-affinity binding could be eliminated by the use of low concentrations of imipramine as the competing ligand. In contrast to the high-affinity binding, the low-affinity binding was found to be unrelated to the neuronal uptake system for serotonin

Thyroid hormones and their derivatives were found to inhibit [3H]flunitrazepam binding stereospecifically and in a monophasic manner. Among the compounds tested, D-thyroxine was the most potent inhibitor (IC50 = 0.5 microM). The naturally occurring L-thyroxine was about 40-fold less potent (IC50 = 20 microM). The structure-activity relationships seem to imply that the thyronine base has the principal role in the inhibition of benzodiazepine receptor binding. The type of inhibition was examined with the most potent inhibitor, D-thyroxine, by Scatchard analysis. The apparent dissociation constant (KD) of the [3H]flunitrazepam binding increased and the receptor density (Bmax) decreased as a function of D-thyroxine concentration; this is characteristic of mixed-type inhibition