Roy B. McCauley

Selective inhibition of MAO-A but not MAO-B activity increases rat pineal melatonin

Clorgyline, a selective MAO-A inhibitor, increased (5 times) rat pineal melatonin and N-acetyl-serotonin (NAS) content, and decreased 5-HIAA level by 80%. Deprenyl, a selective MAO-B inhibitor, did not change melatonin or other pineal indoles content. The data obtained show that inhibition of MAO-A but not B enzyme is responsible for pineal melatonin increase caused by MAO inhibitors. It is suggested that the stimulation of melatonin synthesis caused by MAO inhibitors may contribute to their antidepressive effect.

Cardiac release of urocortin precedes the occurrence of irreversible myocardial damage in the rat heart exposed to ischemia/reperfusion injury

This study evaluates whether cardiac ischemia induces release of urocortin, before and independently from myocyte cell death. Urocortin levels rose after 5‐min ischemia and peaked after 10‐min ischemia, when cell death was not detected. However, myocyte apoptosis and/or necrosis occurred following 20‐ and 30‐min ischemia, which paralleled a fall in urocortin levels, suggesting that urocortin expression and release are mainly sustained by metabolically challenged, though still viable myocytes. Hence, since cardiac release of urocortin, unlike that of conventional biomarkers, occurs before and apart from cell death, urocortin levels may be clinically useful in the diagnosis of sublethal myocardial ischemia.

Single dose of tranylcypromine increases human plasma melatonin

From the Psychoendocrine Research Unit, Lafayette Clinic (G.F.O., I.M.McI.), and the Departments of Psychiatry (G.F.O., I.M.McI., R.B., A.K.J., D.A.), and Pharmacology (R.B.McC., G.F.O.), Wayne State University School of Medicine, Detroit, MI. Supported in part by NIH Biomedical Research Support Grant RROS384-24 and a research award from Wayne State University. Presented at ACNP meetina, December 1985. Address reprint requests to-Dr. G. F. Oxenkrug, Psychoendocrine Research Unit, Lafayette Clinic, 951 E. Lafayette Avenue, Detroit, MI 48207. Received February 6, 1986; revised March 28, 1986 volved in depression (serotonin and noradrenaline) (Pare and Sandler 1959). However, no clear correlation between antidepressant effect of MAO inhibitors and alterations of monoamine metabolism produced by these drugs has been found (Murphy et al. 1983). Depression has been associated with the increased melatonin sensitivity to the inhibitory effect of bright light (Lewy et al. 1981). The disturbed melatonin circadian rhythm (Mendlewicz et al. 1979) and the combination of decreased nighttime melatonin levels with increased plasma cortisol (Branchey et al. 1982) and abnormal Dexamethasone Suppression Test (Wetterberg 1983) have been also observed in depressed patients. MAO inhibitors have been shown to stimulate melatonin synthesis in rat pineal (Klein and Rowe 1969; Deguchi and Axelrod 1972).

Monoamine oxidase A and monoamine oxidase B activities are catalyzed by different proteins.

Monoamine oxidases A and B (amino: oxygen oxidoreductase (deaminating) (flavin-containing), EC 1.4.3.4) have been identified in the outer membranes of rat liver mitochondria by their covalent reaction with the inhibitor, [3H]pargyline. On analysis by polyacrylamide gel electrophoresis under denaturing conditions. Monoamine oxidase A was found to migrate more slowly that monoamine oxidase B. Proteins which correspond to monoamine oxidases A and B (as identified by the electrophoretic distribution of covalently bound [3H]pargyline) were excised from the gels. Subsequent analysis showed that both monoamine oxidase A and monoamine B had been highly purified by this procedure. Electrophoretic analysis of the peptides produced by limited proteolysis with bovine trypsin, alpha-chymotrypsin, Staphylococcus aureus V8 proteinase and cyanogen bromide indicate that monoamine oxidases A and B have different amino acid sequences.

Activation of Src protein tyrosine kinase plays an essential role in urocortin-mediated cardioprotection

Urocortin is a 40 amino acid peptide of the corticotrophin-releasing factor (CRF) family that is synthesized and released by cardiac myocytes. Endogenous urocortin expression is increased during ischemia/reperfusion (I/R) and addition of exogenous urocortin reduces cell death caused by I/R injury. Studies have also showed that the protective action of urocortin is mediated by the activation of ERK1/2. We discovered that a non-receptor tyrosine kinase, Src, is involved in the urocortin-induced activation of ERK1/2 in mouse atrial HL-1 myocytes. The selective Src family kinase inhibitor, PP2, reduced the urocortin-induced phosphorylation of ERK1/2, and so did the expression of a dominant-negative mutant of Src in transfected HL-1 cells. Inhibition of Src by PP2 also reduced urocortins protective effects in HL-1 cells after hypoxia/reoxygenation (H/R), as assessed by flow cytometry and caspase-3 activation assay. Titration studies indicated that as little as 10(-8)M urocortin was sufficient to induce Src activation. Maximal phosphorylation/activation of Src and ERK1/2 were both detected after 5 min incubation with urocortin. These effects of urocortin were largely mediated by CRF receptor-1, although a minor contribution of CRF receptor-2 cannot be excluded. Here we report for the first time that short-term treatment with urocortin causes rapid phosphorylation of Src, and that the urocortin-activated Src kinase serves as an upstream modulator of ERK1/2 activation, playing an essential role in urocortin-mediated cardioprotection.

Possible melatonin involvement in the hypotensive effect of MAO inhibitors

Effect of selective inhibitors of MAO-A and B isoenzymes on pineal melatonin (and related indoles), arterial blood pressure and brain MAO-A and B activities has been evaluated in intact, pinealectomized and shamoperated rats. Selective inhibition of MAO-A but not MAO-B activity stimulated pineal melatonin synthesis and decreased arterial blood pressure in intact and sham-operated animals. Pinealectomy attenuated the hypotensive effect of MAO-A inhibition. The possible melatonin contribution to both antidepressive and hypotensive effects of MAO inhibitors is discussed.

Properties of a monoamine oxidase from rat liver mitochondrial outer membranes

Abstract A relatively simple and reproducible method for the purification of rat liver monoamine oxidase (MAO) is described. The purified enzyme is capable of oxidizing 0.8 μmol of benzylamine/min/mg of protein, and has the same apparent K m value for benzylamine as MAO in the mitochondrial outer membrane. As judged by cofactor content and polyacrylamide gel electrophoresis, the minimum molecular weight is about 65,000. Both purified and membrane-bound MAOs are sensitive to pargyline, but unlike MAO in the outer membrane, the purified enzyme will not oxidize serotonin, norepinephrine, or octopamine, and is only poorly inhibited by clorgyline. Catalytically the purified enzyme resembles the MAO B isoenzyme and seems to be devoid of MAO A activity; however, comparisons between the recoveries of MAO B flavin (as estimated by pargyline binding) and total flavin indicate that catalytically inactive MAO A is also present.

Effect of Selective Monoamine Oxidase Inhibitors on Rat Pineal Melatonin Synthesis In Vitro

Freshly cultured pineal glands respond to the monoamine oxidase A (MAO A) inhibitor clorgyline and to high concentrations of the MAO B inhibitor, deprenyl, with an increase in serotonin N‐acetyltransferase activity and N‐acetylated indoles and a fall in 5‐hydroxylated serotonin degradation products. Glands cultured for 48 hours before challenge respond less. Response is absent in glands cultured for 72 hours and in glands from ganglionectomized animals cultured for 48 hours before challenge. These data are consistent with the hypothesis that these MAO inhibitors stimulate melatonin synthesis by protecting norepinephrine from degradation.

The in vitro insertion of monoamine oxidase B into mitochondrial outer membranes

Bovine monoamine oxidase (MAO) B has been synthesized in vitro using a reticulocyte lysate translation system directed by bovine liver poly(A)+ RNA. The newly synthesized enzyme apparently lacks a cleavable N‐terminal extension, but MAO B is readily incorporated into mitochondria or isolated mitochondrial outer membranes prepared from rat liver. ATP is not required for the binding of the newly synthesized enzyme to the outer membranes, but is necessary for the insertion of MAO B into these membrane vesicles. The ATP is not required to generate a mitochondrial membrane potential as assembly occurs under conditions that preclude either the formation or the maintenance of the potential. MAO B will bind to but not become incorporated into outer membrane vesicles which have been treated with trypsin, suggesting that the insertion of MAO B also depends on protein factors present on the outer membranes.

Carbidopa effect on rat brain monoamine oxidase and pineal melatonin

A number of investigators have observed a diurnal rhythm in brain monoamine oxidase (MAO) activities (Chevillard et al. 198 l; Bhaskaran and Radha 1984), and it has been suggested that the pineal gland may influence pituitary MAO-A activity (Oxenkrug et al. 1984a). The substrates of MAO-A (serotonin and noradrenaline) are involved in the regulation of melatonin synthesis, whereas MAO-B apparently does not participate in melatonin metabolism (Lewy 1983). In accordance with this suggestion, selective inhibition of MAO-A but not MAO-B has been found to stimulate melatonin synthesis (Oxenkrug et al. 1984a). However, the negative correlation between age-associated changes in rat pineal melatonin and brain MAOB suggests the possibility of melatonin influence on MAO-B activity (Filipowicz et al. submitted). This report represents the attempt to explore such a possibility. In these experiments, rats were treated with a dose of the aromatic amino acid decarboxylase inhibitor, carbidopa, that was sufficient to inhibit the synthesis of pineal serotonin and melatonin but not brain serotonin (Oxenkrug et al. 1984b). This differential effect can be obtained because carbidopa does not readily penetrate the blood-brain barrier. MAO activities were then measured in the