Succinic semialdehyde dehydrogenase deficiency: an inborn error of gamma-aminobutyric acid metabolism

Gamma-hydroxybutyric aciduria is a disorder of gamma-aminobutyric acid metabolism in which a compound of known neuropharmacologic activity accumulates. We have studied two patients in whom high levels of gamma-hydroxybutyric acid were found in blood, urine and cerebrospinal fluid. A coupled assay has been developed which estimates succinic semialdehyde dehydrogenase activity in isolated human lymphocytes. The mean activity of succinic semialdehyde dehydrogenase in a control and the four parents and two healthy siblings of these patients was 8.8 +/- 1.9 pmol . min-1 . mg-1 protein. In the patients the activities were 0.8 and 1.1 pmol . min-1 . mg-1 protein, approximately 9-13% of control. In the presence of saturating amounts of NAD+, lymphocyte sonicates, derived from the patients accumulated a significant amount of 14C-succinic semialdehyde from 14C-gamma aminobutyric acid, whereas none could be detected in controls. The data suggest a deficiency of succinic semialdehyde dehydrogenase in these patients, the first documented defect of the metabolism of gamma-aminobutyric acid in man.     

Serum high density lipoprotein cholesterol in diabetes mellitus: An analysis of factors which influence its concentration

The study assesses whether diabetes has an effect on serum high density lipoprotein (HDL) cholesterol concentration independent of other factors known to influence serum HDL cholesterol concentration. Concentrations of serum HDL cholesterol, serum cholesterol, serum M particles and blood sugar, and the proportion of haemoglobin in the form of HbA₁ were measured in a diabetic and a non-diabetic population. Relative body weight, alcohol and cigarette consumption, age, a clinical estimate of diabetic control and duration of diabetes were also recorded. The diabetic patients were divided into those in whom insulin treatment was clinically indicated and those in whom it was not.Serum HDL cholesterol concentration was significantly higher in diabetic men treated with insulin than in normal men and also higher in diabetic women treated with insulin than in normal women. In the diabetic men and women not treated with insulin, serum HDL cholesterol concentration was not significantly different from normal. There were differences between the diabetic and non-diabetic populations in terms of factors known to influence serum HDL cholesterol and also in the degree of correlation between these and the serum HDL cholesterol. Multivariate analysis revealed that in both male and female diabetic patients treated with insulin, diabetes was a highly significant influence on serum HDL cholesterol concentration, but in the non-insulin-treated diabetic patients the influence was absent in women and only marginal in men. The proportion of HbA₁ influenced serum HDL cholesterol concentration negatively in insulin-treated diabetes but not in diabetes treated without insulin.     

3-Hydroxy-3-methylglutaryl-CoA lyase in human skin fibroblasts: study of its properties and deficient activity in 3-hydroxy-3-methylglutaric aciduria patients using a simple spectrophotometric method

In this paper we studied the properties of 3-hydroxy-3-methylglutaryl-CoA lyase (HMG-CoA lyase) in human skin fibroblasts. The enzyme was found to exhibit an absolute requirement for divalent cations such as magnesium. Furthermore, dithiothreitol was necessary for full activity. The enzyme was found to be maximally active at pH 9.25. When measured at this pH in the presence of magnesium and dithiothreitol enzyme activity was high enough to be determined by simple spectrophotometry by measuring the amounts of acetoacetate produced. The results obtained suggest that the large variation in the values reported in literature for the activity of HMG-CoA lyase in human skin fibroblasts is due to the fact that the enzyme shows little activity at pH values below 8.     

Aspirin clearly depresses responses of ventrobasal thalamus neurons to joint stimuli in arthritic rats

This study dealt with the effect of aspirin upon activities of 17 ventrobasal thalamic neurons recorded in 17 rats rendered arthritic by injection of Freund's adjuvant into the tail. These neurons presented reproducible responses to mobilization and/or mild lateral pressure on a joint and were recorded for at least 40 min after aspirin administration. After intravenous injection of aspirin at the dose of 50 mg/kg (13 neurons tested), there was a progressive decrease in the number of spikes in the discharges. The maximum effect occurred at 30 min where the mean value of the response expressed as a percentage of the control was m = 34.62 +/- 7.5% (n = 13, p less than or equal to 0.001). Recovery was progressive and could be considered as complete at 60 min. By contrast, no significant modification of the spontaneous firing has been observed. With lower doses of aspirin (12.5 or 25 mg/kg tested with 4 neurons) there was respectively no clear depressive effect or only a transient decrease of the response.     

Behavioral treatment of chronic pain: variables affecting treatment efficacy

Behavioral approaches to pain management have been shown to be efficacious in many instances but many of the variables indicating or contra-indicating such treatment remain unspecified. The data reported here demonstrated that while patients referred by both Disability and Medical Subspecialties showed improvement in pain report, affect, and feelings of control, those patients referred by Disability improved reliably less, remain hospitalized longer and were reliably less compliant. These data can be understood within an operant conditioning conceptualization of illness behavior.     

Determination of noradrenaline and adrenaline in plasma by a radioenzymatic assay using high pressure liquid chromatography for the separation of the radiochemical products.

Plasma catecholamines are determined by a radioenzymatic assay using high pressure liquid chromatography for the separation of the labelled radiochemical products. Noradrenaline and adrenaline are converted to their O-methylated analogues, normethanephrine and metanephrine, by the enzyme catechol O-methyltransferase in the presence of tritiated S-adenosyl-L-methionine. Non-radioactive carriers are added, whereupon the metanephrines are extracted in an organic phase and re-extracted in an aqueous phase. The aqueous phase is dried, the residue taken up in a phosphate solution and chromatographed. The normetanephrine and metanephrine peaks are collected, converted to vanillin and assayed for radioactivity. The detection limit for noradrenaline is 32 ng/l plasma and for adrenaline 16 ng/l plasma. Intra- and interassay variables are respectively 6.2 and 10% for noradrenaline and 5.3 and 8.8% for adrenaline. The mean supine levels of noradrenaline and adrenaline in normal subjects are 355 and 61 ng/l plasma, respectively. This highly specific procedure takes two days for one person to assay 45 blood samples and can be reduced to one day if the chromatography step is automated.     

Potentiation of morphine antinociception by monoamine reuptake inhibitors in the rat spinal cord

Potentiation of the antinociceptive effects of morphine by the tricyclic antidepressants was assayed in awake restrained rats using the tail-flick test. Intrathecally administered amitriptyline, desipramine or sertraline at doses that had no effect themselves (25-30 micrograms) potentiated a subthreshold parenteral dose of morphine (0.5 mg/kg). The morphine potentiating effect of amitriptyline was prevented by prior administration of parachlorophenylalanine (PCPA). This effect of PCPA was not restored by 5-hydroxytryptophan (5-HTP) but was restored when the animals were left for 14 days to replete. The morphine potentiating effects of amitriptyline, desipramine and sertraline were blocked by intrathecal administration of low doses of the serotonin antagonist methysergide and the alpha-adrenergic antagonists yohimbine and phentolamine but not by the beta-adrenergic antagonist propranolol. The results are consistent with the hypothesis that the potentiation of morphine's antinociceptive effect by tricyclic antidepressants depends on activation of both spinopetal serotonin and adrenergic neurons.     

Myeloma immunoglobulin interferes with serum thyroxine analysis by homogeneous enzyme immunoassay

Myeloma immunoglobulin paraproteins interfere with a homogeneous enzyme immunoassay (EMIT) for serum thyroxine. The EMIT assay failed to detect any hormone in three hyperproteinemic sera from multiple myeloma patients, although thyroxine in these sera was accurately measured by our competitive protein-binding radio-assay on small, re-usable Sephadex columns. The interference was due to turbidity of the paraproteins in the EMIT reaction mixture, resulting in an increased absorbance and a marked underestimation of hormone concentration. Thyroxine was detected (82 to 107% recovery) by the EMIT assay in ethanol extracts of myeloma sera. With 82 other sera there was an excellent correlation (r = 0.985, SLOPE = 0.912, Y intercept (EMIT) = 6.8) of the EMIT assay with our competitive radioassay. Thus, although the EMIT thyroxine assay possesses many desirable features and it is an attractive alternative method to competitive radioassays, its susceptibility to interferences by immunoglobulin paraproteins is a troublesome liability.     

Determination of naproxen and its desmethyl metabolite in human plasma or serum by high performance liquid chromatography.

1. A liquid chromatographic method has been developed for the simultaneous determination of the anti-inflammatory agent naproxen and its metabolite, 6-0-desmethyl-naproxen, in human plasma or serum. 2. After addition of p-chlorowarfarin as the internal stardard, plasma is acidified and extracted with either chloroform (for assay of naproxen only) or ethyl acetate (for assay of naproxen and desmethyl-naproxen), the organic solvent is evaporated, the residue is dissolved in acetonitrile and injected onto a reverse phase column coupled with an ultra-violet absorbance detector. 3. Drug and metabolite can be detected readily in concentrations above 2 micrograms per ml in 0.5 ml samples. Salicylic acid and its metabolites and a number of other durgs were found not to interfere with the assay.     

3-Mercaptopyruvate sulfurtransferase (EC 2.8.1.2): a simple assay adapted to human blood cells

A product of cysteine catabolism, 3-mercaptopyruvate, is enzymatically degraded to sulfur and pyruvate by a sulfurtransferase (EC 2.8.1.2.) present in human red and white blood cells. A simple sulfurtransferase assay is reported which takes advantage of the fact that pyruvate is conveniently measured enzymatically by lactate dehydrogenase (LDH) after the elimination of 3-mercaptopyruvate, also a substrate of LDH, by addition of N-ethylmaleimide. Sulfite is employed as sulfur acceptor, and conditions for a reproducible assay including data for preparation and storage of reactants, their assay, and their optimal concentrations are given. The apparent K_m for sulfite is 6.5 · 10^?3 M and for 3-mercaptopyruvate is 1.9 · 10^?3 M. A reducing agent, in this assay dithiothreitol (Cleland's reagent), is essential for active transsulfuration. A normal metabolite, 3-mercaptopyruvate is reported elsewhere to have the capacity of producing polyploidy and chromosomal endoreduplication, a feature rendering it of interest in tumor metabolism.     

Specific radioactivity of radioimmunoassay tracer determined by self-displacement: a re-evaluation.

A gas Chromatographie procedure with flame ionisation detection has been developed to measure 2-phenylethylamine in human urine as the trifluoroacetyl derivative. The amine is purified by an ether extraction procedure and derivatised with trifluoroacetic anhydride. Quantification is obtained by comparing the peak height with that for an internal standard added to the urine before extraction.The normal 24-h excretion value for 2-phenylethylamine in human urine is found to be 6.8 ± 2.9 Mg (mean ± S.D.).