Pauline Lerner

Modification of the tyrosine hydroxylase assay. Increased enzyme activity in the presence of ascorbic acid.

A modification of the tyrosine hydroxylase assay is described in which ascorbate, rather than 2-mercaptoethanol or dihydropteridine reductase with NADPH, is used as the reductant. Enzyme activity is 3–4 times higher with ascorbate than with the other reducing agents. Low blanks are obtained with the ascorbate system provided that catalase is also included. The tissue distribution and kinetic activation of the enzyme have been studied with the ascorbate assay. The results obtained are consistent with the biological and regulatory properties of the enzyme which have been determined with the other reducing systems.

Dopamine-β-hydroxylase and norepinephrine in human cerebrospinal fluid: Effects of monoamine oxidase inhibitors

Abstract The effect of the monoamine oxidase inhibitors, clorgyline and pargyline, on central noradrenergic function in depressed patients was assessed by measuring dopamine beta hydroxylase (DBH) and norepinephrine in cerebrospinal fluid (CSF). These drugs caused a significant decrease in DBH in CSF. This decrease is consistent with a reduction in the release of DBH into CSF as noradrenergic firing is slowed by the drugs. Levels of DBH and norepinephrine in CSF were highly correlated. The ratio of CSF norepinephrine to CSF DBH increased when the patients were treated with monoamine oxidase inhibitors, consistent with a drug-induced increase in brain norepinephrine.

The role of reductants in the tyrosine hydroxylase reaction.

Abstract The role of several reducing systems in the tyrosine hydroxylase reaction has been studied. A significant dependence upon the reducing systems beyond that required to regenerate the oxidized cofactor has been observed. 2-Mercaptoethanol, NADPH, and ascorbate are each effective at reducing the cofactor, but their abilities to stimulate tyrosine hydroxylase vary over a threefold range. NADPH is a suitable reductant for the tyrosine hydroxylase reaction, even in the absence of pteridine reductase. A reducing system containing ascorbate, ferrous ion, and catalase gives unusually high enzyme activity and low blanks. This ascorbate system, in addition to being useful for in vitro enzyme assays, may serve as a model for the in vivo reaction. Ascorbate may play an important role in the hydroxylation of tyrosine in catecholaminergic tissues. This study demonstrates that an efficient reductant for the tyrosine hydroxylase reaction must, in addition to reducing the pterin cofactor, also interact effectively with the enzyme itself.

Dopamine-β-hydroxylase in cerebrospinal fluid and plasma: Effects of α-adrenergic agents

Dopamine-beta-hydroxylase (DBH) and norepinephrine are both localized in noradrenergic storage vesicles. When noradrenergic nerves fire, both norepinephrine and DBH are released by exocytosis. DBH released from the peripheral nervous system and the adrenal medullae is found in blood, while DBH in cerebrospinal fluid (CSF) is presumably of central origin. This study was designed to: (1) investigate the effect of drugs which alter central noradrenergic activity on DBH activity in CSF; and (2) compare the effects of these drugs on DBH in CSF and plasma in cats. Phenoxybenzamine was given subcutaneously at 6 mg/kg and DBH was measured 8 h later. This treatment significantly increased DBH activity in CSF (n = 10,P less than 0.005). There were no consistent changes in plasma DBH, although there was a tendency for DBH to increase from low basal levels and to decrease from high basal levels. Clonidine was administered in 4 subcutaneous injections (100, 50, 50, 50 microgram/kg) over a 19-h period, and blood and CSF were taken 5 h after the last injection. This treatment caused a significant decrease in CSF DBH activity (P less than 0.05, n = 8). The effect of clonidine on plasma DBH was strongly dependent on the basal enzyme level. The 3 lowest DBH values increased and the 5 highest DBH values decreased on drug treatment. These results are discussed with respect to the theory that changes in CSF DBH may reflect central noradrenergic activity.

The central noradrenergic system and affective response to MAO inhibitors

1. In humans, norepinephrine (NE) has been postulated to be involved in the regulation of mood and behavior and to be altered in patients with manic-depressive illness. 2. Recent methodological advances have made possible a more direct assessment of central noradrenergic activity by the accurate measurement of the small amounts of NE and of the enzyme responsible for the conversion of dopamine to NE, dopamine-beta-hydroxylase (DBH), found in cerebrospinal fluid (CSF). 3. Cerebrospinal fluid samples were obtained from depressed patients both before and after treatment with two monoamine oxidase-inhibiting antidepressant drugs, clorgyline and pargyline. 4. Patients were rated twice daily by nursing staff on a modified 15-point scale for severity of global depression and anxiety. Patients were also rated using the Hamilton depression rating scale. 5. High negative correlations were observed between the drug-related changes in CSF NE and the changes in depression ratings on both the global ratings (r = -.95, p less than .001) and the Hamilton rating scale (r = -.81, p less than .01). Changes in NE were also highly correlated with changes in global anxiety ratings (r = -.85, p less than .01) calculated on the basis of changes from baseline for each measurement. Drug-related changes in CSF DBH similarly showed negative correlations with clinical response (r = -.79, r = -.38, r = -.68 respectively). In contrast, no significant correlations were found when drug-related changes in CSF MHPG were compared to changes in clinical state.

Central and peripheral dopamine beta hydroxylase: responses to long term treatment with monoamine oxidase inhibitors.

Abstract The effect of long-term (3 weeks) treatment with monoamine oxidase inhibitors on dopamine beta hydroxylase (DBH) in cats was studied. The DBH was measured in blood and CSF drawn before treatment began and after 2, 8, 14, and 22 days of injections. Animals were killed after 24 days of injections, and DBH was measured in brain cortex and pons-medulla. Both clorgyline and pargyline decreased DBH activity in CSF after 22 days of treatment. These drugs decreased plasma DBH after 8 days of treatment, and the decrease persisted throughout the study. Brain DBH levels in the drug-treated animals were not significantly different from enzyme levels in saline-treated controls. The decrease in CSF DBH occurred 2 weeks after the decrease in the plasma enzyme, suggesting that CSF DBH levels do not simply reflect plasma DBH levels. The decrease in DBH in CSF was not caused by a large-scale depletion of the enzyme in brain. It is likely that the observed change in CSF DBH is due to a decrease in central noradrenergic activity, which has been demonstrated in rats. It is of interest that this change is seen only after three weeks of drug treatment, at the time when the therapeutic effects of these drugs become evident in patients.

Central noradrenergic adaptation to long-term treatment with imipramine in rhesus monkeys

Rhesus monkeys were treated with the antidepressant drug imipramine; cerebrospinal fluid norepinephrine and dopamine-beta-hydroxylase were measured to assess central noradrenergic activity. Large changes occurred after short-term, but not long-term, treatment. Biochemical stabilization occurs at the time when therapeutic effects are seen in patients.

CEREBROSPINAL FLUID NOREPINEPHRINE AND DOPAMINE-BETA-HYDROXYLASE IN AFFECTIVE ILLNESS AND SCHIZOPHRENIA

Publisher Summary A variety of studies have utilized cerebrospinal fluid (CSF) methodology to measure norepinephrine metabolites in patients with affective illness. Preliminary studies in animals and man have suggested that there is a diurnal variation in norepinephrine in CSF. This chapter describes a study where it was found that both baseline levels of norepinephrine and those achieved following 18 hours of probenecid administration were significantly higher in manic compared to depressed patients or neurological controls. While levels of CSF norepinephrine were significantly elevated in manic patients compared to other patient populations and control groups, DBH levels were decreased in these manic patients. CSF norepinephrine was elevated in manic patients as compared to depressed or normal controls. CSF DBH was not different in schizophrenic patients as a group compared to controls, but it was lower in a subgroup of patients with good premorbid interpersonal functioning.

Dopamine-β-Hydroxylase in Cerebrospinal Fluid

Dopamine-β-hydroxylase (DBH), the biosynthetic enzyme for norepinephrine, is unique among enzymes in that it is localized with a neurotransmitter in storage vesicles and is released with the transmitter during nerve firing. After release, norepinephrine binds to a receptor on a postsynaptic or effector cell, but the released DBH has no known physiological function. Although the exocytotic release of DBH has no readily apparent value to the cell, its release is potentially of great value for neuroscientists. DBH released from noradrenergic tissue can be measured in plasma and cerebrospinal fluid (CSF), and such measurements may give a chemical evaluation for noradrenergic activity in vivo.80a This chapter reviews and evaluates studies on DBH in CSF, with special reference to the theory that DBH activity in CSF may reflect central noradrenergic activity.