Elevated plasma dopamine beta hydroxylase activity in rats with neurogenic hypertension.

Increased plasma dopamine beta hydroxylase, DBH, activity has been cited as evidence of increased sympathetic function in essential hypertension. Here-to-fore, experimental hypertension in animals has been associated with normal plasma DBH activity. This study shows that rats with neurogenic hypertension, induced by sinoaortic denervation, SAD, have elevated DBH activity; the mean increase in plasma DBH measured 3 days to 11 weeks after operation was 74% higher in the SAD group than in the sham-operated, control group. DBH activity showed a positive correlation with arterial pressure. Mesentery DBH activity was inversely related to plasma enzyme activity in SAD rats, indicating sympathetic nerve terminals in mesentery are a source of plasma DBH. We conclude that plasma DBH activity is an index of increased sympathetic function since it is consistently elevated in rats with neurogenic hypertension resulting from sustained central activation of the sympathetic nervous system.     

Regulation of retinal dopamine biosynthesis and tyrosine hydroxylase activity by light

Dopamine (DA)-containing neurons of the rat retina are apparently activated transsynaptically by photic stimulation. Exposure of dark-adapted rats to light increases retinal DA biosynthesis and metabolism. Associated with the light-evoked increase of DA biosynthesis is a rapid activation of tyrosine hydroxylase (TH), the rate-limiting enzyme of catecholamine biosynthesis. The activation of TH is characterized by an increased affinity of the enzyme for the pteridine cofactor. Because TH in dark-adapted retinas is apparently not saturated with cofactor, the light-evoked increase of affinity is probably responsible for the observed stimulation of DA biosynthesis. Cyclic AMP (cAMP)-dependent protein phosphorylation in vitro activates TH extracted from dark-adapted retinas, and phosphorylation-induced TH activation is very similar and not additive with light-evoked activation of the enzyme. Incubation of viable cell suspensions of dissociated retinas with 8-bromo cAMP also activates TH, which indicates the availability of sufficient cAMP-dependent protein kinase in the proper subcellular compartment to regulate the enzyme in situ. The DA-containing neurons of the rat retina are tonically inhibited in darkness, and evidence is presented that this tonic inhibition involves direct synaptic input to the DA neurons from gamma-aminobutyric acid-containing amacrine cells. The DA-containing neurons are also subject to feedback inhibition through DA receptors, and to modulation by alpha 2-adrenergic receptors.     

Short-term effect of stress on tyrosine hydroxylase activity

The short-term influences of stress on the activities of tyrosine hydroxylase in vivo and in vitro were examined in mice. The in vivo tyrosine hydroxylase activity was estimated by the rate of dopa accumulation which was measured at 30 min after the injection of NSD-1015 (100 mg kg), an aromatic l-amino acid decarboxylase inhibitor, intraperitoneally and was compared with tyrosine hydroxylase activity measured in vitro. For the in vivo assay, both the accumulation of dopa (tyrosine hydroxylase activity) and that of 5-hydroxytryptophan (tryptophan hydroxylase activity) and the levels of monoamines and the metabolites (noradrenalin, adrenalin, dopamine, normetanephrine, 3-methoxytyramine and serotonin) and those of precursor amino acids, tyrosine and tryptophan, were investigated in ten different brain regions and in adrenals. The amount of dopa accumulation in the brain as a consequence of decarboxylase inhibition, in vivo tyrosine hydroxylase activity, was significantly increased by stress, in nerve terminals (striatum, limbic brain, hypothalamus, cerebral cortex and cerebellum) and also in adrenals. The effect of stress on tyrosine hydroxylase activity in vitro at a subsaturating concentration of 6-methyltetrahydropterin cofactor was also observed in nerve terminals (striatum, limbic brain, hypothalamus, and cerebral cortex). The amount of 5-hydroxytryptophan accumulation, the in vivo tryptophan hydroxylase activity, was also significantly increased in bulbus olfactorius, limbic brain, cerebral cortex, septum and lower brain stem. The influence of stress was also observed on the levels of precursor amino acids, tyrosine and tryptophan and monoamines in specific brain parts. These results suggest that the stress influences both catecholaminergic neurons and serotonergic neurons in nerve terminals in the brain. This effect was also observed on tyrosine hydroxylase activity in vitro in nerve terminals. However, in adrenals, the influence by stress was not observed on the in vitro activity, although dopa accumulation was increased.     

Studies of the inhibition of dopamine beta hydroxylase by thiono-sulfur containing compounds.

The inhibitory effect of several thione and thioether compounds on dopamine β hydroxylase has been examined. Carbon disulfide and propylthioracil inhibited the enzyme competitively and showed a Ki of 1.1 × 10⁻⁴ and 1.0 × 10⁻⁴M, respectively. Dialysis reversed the inhibition but N-ethylmaleimide did not. The thioethers methionine, benzylmethyl sulfide, thiazole and 2-amino-thiazole had no inhibotory effect on enzymatic activity. No evidence of oxidative metabolism of the thione inhibitors by dopamine β hydroxylase could be detected.     

The effect of chronic iron deficiency on adrenal tyrosine hydroxylase activity.

Chronic nutritional iron deficiency of 2 to 5 weeks duration reduced the blood hemoglobin content to 30-50% of control values and resulted in an increase in rat adrenal tyrosine hydroxylase (TH) (EC 1.14.16.2) activity. Kinetic and mixing experiments indicated that this increase was due to an increase in enzyme protein. The body weight of iron-deficient rats ranged from 60 to 80% of control; this factor, however, was not responsible for the increase in adrenal TH as enzyme activity was directly proportional to final body weight. To determine whether the increase in adrenal TH in iron-deficient rats was due to increased sympathetic activity to the adrenal medulla, the splanchnic nerve was cut. The increased TH was still observed after adrenal denervation; this indicates that the mechanism of response to iron deficiency lies within the adrenal itself. Age of the rats is important in determining whether the increase in TH activity will occur.     

The effect of fasting time on plasma total cholesterol concentration in Mongolian gerbils

This study investigated the effect of length of fasting time on plasma total cholesterol response of male Mongolian gerbils (Meriones unguiculatus). Plasma cholesterol levels from fed and fasted gerbils were also compared with those reported for humans under similar metabolic states. Plasma total cholesterol response showed a significant quadratic relationship with time over a 15-hour period. Between 6 and 9 hours of fasting (the time during which plasma triglyceride concentration became relatively constant), the average plasma total cholesterol concentration was 178 mg/dl, compared with a zero hour (fed) cholesterol level of 265 mg/dl. The difference in plasma cholesterol levels observed in fed and fasted gerbils is unlike what has been reported for humans. Results from most human studies show no differences in plasma total cholesterol concentrations for fed and fasted subjects. Failure to consider species differences in metabolic responses may have implications when results from animal experiments are extrapolated to humans.     

Allosteric effect of tetrahydrobiopterin cofactors on tyrosine hydroxylase activity.

Allostery of tyrosine hydroxylase was found by kinetical studies of partially purified tyrosine hydroxylase from clonal rat pheochromocytoma PC12h cells. Positive cooperativity toward the cofactors, (6R)-L-erythro-5,6,7,8-tetrahydrobiopterin [(6R)BH4] and (6S)-L-erythro-5,6,7,8-tetrahydrobiopterin [(6S)BH4], was observed. It is indicated that biopterin might be the regulatory factor of the enzyme polymers, which changes the affinity for the cofactor itself. Moreover, the stereochemical structure of (6R)BH4, the naturally-occurring cofactor, took an important role on the kinetical properties of the enzyme in concern with L-tyrosine.     

The effect of post-heparin lipolytic activity on plasma lipoproteins

Summary Electron microscopy of negatively stained plasma shows that intravenous administration of heparin to healthy humans results in a fast degradation of very low density lipoproteins (VLDL), which are very rich in triglycerides (50% w/w). VLDL are also rapidly digested by post-heparin lipolytic activity when serum from non-heparinized subjects is incubated at 27°C in vitro with post-heparin serum of healthy controls.Abbreviations VLDL very low density lipoproteins - LDL low density lipoproteins - HDL high density lipoproteins - LP-X lipoprotein-X, abnormal lipoprotein characterizing obstructive jaundice     

The effect of fasting and physical exercise on plasma leptin concentrations in high-fat fed rats.

The aim of our study was to estimate the effect of fasting and physical exercise on a treadmill on plasma leptin concentrations in high-fat fed rats. Male Wistar rats were injected a low dose of streptozotocin (STZ) or buffer at 2 days of age and later fed a standard or high-fat diet (HFD). Plasma leptin was measured by RIA method in all the groups studied in basal conditions, after 48h fasting, a single bout of exhaustive exercise, and 4 weeks of exercise training. Plasma leptin concentrations were markedly elevated in the HFD and STZ/HFD groups compared to the control group. The significant correlation between plasma leptin and body weight was noted. Fasting and exercise training decreased plasma leptin in similar percentage in all the groups studied. The observed decrease was greater than expected from changes in body weight. We conclude that high-fat feeding results in an increase in plasma leptin levels in rats independently of plasma insulin or daily calorie intake. High-fat fed rats have maintained leptin response to fasting and exercise training. The reduction in plasma leptin after exercise training is partly independent on changes in body weight or plasma insulin.     

Tyrosine hydroxylase and regulation of dopamine synthesis

Tyrosine hydroxylase is the rate-limiting enzyme of catecholamine biosynthesis; it uses tetrahydrobiopterin and molecular oxygen to convert tyrosine to DOPA. Its amino terminal 150 amino acids comprise a domain whose structure is involved in regulating the enzyme’s activity. Modes of regulation include phosphorylation by multiple kinases at four different serine residues, and dephosphorylation by two phosphatases. The enzyme is inhibited in feedback fashion by the catecholamine neurotransmitters. Dopamine binds to TyrH competitively with tetrahydrobiopterin, and interacts with the R domain. TyrH activity is modulated by protein–protein interactions with enzymes in the same pathway or the tetrahydrobiopterin pathway, structural proteins considered to be chaperones that mediate the neuron’s oxidative state, and the protein that transfers dopamine into secretory vesicles. TyrH is modified in the presence of NO, resulting in nitration of tyrosine residues and the glutathionylation of cysteine residues.     

The stimulatory effect of platelet homogenate on prolyl hydroxylase activity in L-929 cells

We have found that the addition of platelet homogenate to confluent cultures of L-929 cells increases 2-3 times the activity of prolyl hydroxylase in these cells. Furthermore, it was found that the platelet homogenate potentiates the effect of ferrous ions and ascorbic acid, which are known activators of prolyl hydroxylase. The effect of the platelet homogenate is diminished by cycloheximide. It seems probable that some products present in the platelet homogenate may promote biosynthesis of the enzyme or they stimulate glycolysis and accumulation of lactic acid, an activator of the hydroxylase.     

The effect of population density on phenylalanine hydroxylase activity in rat-hepatoma cells in culture

The expression of phenylalanine hydroxylase activity in rat-hepatoma cells in culture (line H4-II-E-C3) is a function of culture density: under normal growth conditions in the presence or in the absence of exogenously added hydrocortisone, the levels of this enzyme are low in subconfluent cell populations, but increase steeply as cultures attain confluency. This phenomenon (i) is not an artifact of the subcultivation process and (ii) is not produced by some alteration in the growth medium effected by high-density cultures. The levels of phenylalanine hydroxylase in high-density cultures of H4-II-E-C3 cells in the presence of serum plus added hydrocortisone are at least 80% of those seen in adult-rat liver. It is concluded that this density-associated phenomenon is the result of an intrinsic property of H4-II-E-C3 cells and possibly constitutes a form of epigenetic control governing the sensitivity of these cells to stimulation by serum or by serum plus hydrocortisone.     

Localization of the human dopamine beta hydroxylase (DBH) gene to chromosome 9q34

A human cDNA clone for dopamine beta hydroxylase (DBH) has been isolated from a phaeochromocytoma library. In situ hybridization of this probe to replication-banded chromosomes has localized the gene to chromosome 9q34. The structural gene for the enzyme is therefore close to the ABO blood group locus. This suggests that the previously described activity variation in levels of serum DBH may reflect alterations in either the structure or regulation of the DBH coding sequences. Both biochemical and genetic evidence therefore indicate independence of DBH from the pterin-dependent aromatic amino acid hydroxylases of the neurotransmitter pathways.     

Influence of fasting and dopamine on the tissue catecholamines diurnal variations

To define the role of catecholamines (CA) in the metabolic adaptation to fasting we examined the effect of exogenous dopamine(DA) on heat production(HP) and CA content in the interscapular brown adipose tissue(IBAT) and adrenals of control-fed and 2-day fasted rats in the morning(M) and in the evening(E). DA stimulates HP in fed rats in the M by 45% but the thermogenic effect of this CA is markedly higher in the E. However, DA had no thermogenic effect in fasted rats. The tissue CA in fed rats fluctuates diurnally: in the IBAT noradrenaline(NA) was much higher in the E while adrenaline(A) in adrenals was lower. DA in fed rats did not change the adrenal A but reduced NA content both in the adrenals and in the IBAT all over the day. Fasting depleted A from adrenals but increased NA content both in the M and in the E. Unlike the adrenals in the IBAT fasting did not affect NA content. In the adrenal gland of fasted rats DA significantly increased the A content to the equal degree during the day, while this CA had no effect on NA content of the IBAT.