Absence of 6-Hydroxydopamine in the Rat Brain After Treatment with Stimulants and Other Dopaminergic Agents: A Mass Fragmentographic Study

Abstract: Formation of 6-hydroxydopamine (6-OHDA) from dopamine has been hypothesized to mediate neuro-degeneration induced by some psychostimulants. Although the emergence of a 6-OHDA-like substance was reported in the striatum of methamphetamine-treated rats, this substance has not been identified by a direct approach. We used mass fragmentography to search for 6-OHDA in the rat frontal cortex and striatum after the administration of a number of drugs including 3,4-dihy-droxyphenyl-L-alanine, methamphetamine, amphetamine, and cocaine, all of which increase synaptic dopamine. No 6-OHDA was detected after the acute systemic administration of these agents. Intraventricular administration of 6-OHDA (10 μg/rat.) produced measurable concentrations of 6-OHDA that were higher in the striatum than in the frontal cortex. Intraventricular administration of 2,4,5-trihydroxy-phenyl-D,L-alanine (6-OHDOPA; 10 μg/rat) produced similar concentrations of 6-OHDA in both regions. Pargyline, but not carbidopa (α-methyldopahydrazine), enhanced the effect of intraperitoneal 6-OHDOPA administration (80 mg/kg). We conclude that (1) 6-OHDOPA can cross the blood-brain barrier and is converted to 6-OHDA in the brain, (2) 6-OHDA is a substrate for monoamine oxidase(s) and therefore a search for its purported deaminated metabolite is warranted, and (3) acute treatment with the above stimulants either does not lead to the formation of 6-OHDA or produces concentrations below the detection limit of the assay (<34 pg/mg of protein).     

Enumeration of stressed cells of Escherichia coli.

Cells of Escherichia coli K-12 were stressed by heating at 48 degrees C or by acid treatment at pH 4.2 for periods up to 1h. The addition of catalase to the selective medium increased the count of heat-stressed cells by 2.3-fold and acid-stressed cells by 4.8-fold. However, these values represented only a small percentage (3% for heat-stressed and 6% for acid-stressed cells respectively) of the population of injured but still viable cells. The addition of mannitol to the selective medium used to count acid-stressed cells did not increase the count. Whilst the presence of H2O2 in media may cause significant errors in the estimation of E. coli in certain situations these errors are unlikely to be significant in physiological studies of populations of cells injured by stress.     

Antibiotic decontamination of the dog and its consequences.

An isolation-decontamination regimen was developed which effectively reduced the numbers of resident flora of the dog. Bacterial counts in four dogs before treatment were 3.8 X 10(9) per gram of feces; no organisms were detectable in these same dogs after treatment, however, the intestinal flora had returned to slightly above normal levels 1 week after treatment. Decontamination was accomplished in a laminar air flow system designed to minimize the area that had to be under controlled conditions. By determining the antibiotic sensitivities of 67 isolated organisms representing eight species or groups of bacteria recovered from the four dogs, a standardized antibiotic regimen was developed consisting of bacitracin and neomycin administered as a dry powder in the food. The decontamination treatment apparently did not affect host metabolism because no alterations in serum levels of urea nitrogen, glucose, phosphate, total protein, chloride, sodium, potassium, serum glutamic oxalacetic transaminase, or serum glutamic pyruvic transaminase were found in the antibiotic-treated dogs. The decontamination process did, however, reduce normal granulopoietic stimulation.     

A micro-enzymatic method to measure cholesterol and triglyceride in lipoprotein subfractions separated by density gradient ultracentrifugation from 200 microliters of plasma or serum

A micro-enzymatic method was developed to measure total cholesterol (CHOL) and triglyceride (TG) in lipoproteins and their subfractions separated by density gradient ultracentrifugation. This method had a detection limit and sensitivity below 2 mg/dl and accuracy (bias to reference sera) and imprecision (coefficient of variation) of less than 3% between 2 and 30 mg/dl for both CHOL and TG. In addition, the method was in good agreement with standardized Abell-Kendall CHOL (r = 0.98) and enzymatic TG (r = 0.99) methods. Lipoproteins from 200 microliters of plasma or serum were separated by either equilibrium (EQ)- or rate zonal (RZ)-density gradient ultracentrifugation and the resulting fractions were analyzed for CHOL and TG by the micro-enzymatic method. Lipoprotein measurements by these micro-enzymatic/density gradient methods were highly correlated with standardized Lipid Research Clinic (LRC) procedures and preparative ultracentrifugation. The EQ-density gradient procedure also allowed determination of CHOL and TG in LDL and HDL subfractions within any desired density interval. These methods will facilitate the measurements and study of lipoproteins and their subfractions especially in infants, children, the elderly, and small animals. In addition, the micro-enzymatic method may be adapted to other modes of lipoprotein separation such as liquid chromatography, electrophoresis, and precipitation. CHOL or TG determinations could be made on approximately 500 density gradient fractions per hour.     

Plasma aldosterone, plasma lipoproteins, obesity and insulin resistance in humans.

Aldosterone production in vitro can be affected by many hormones, autacoids, ions, and lipids, but regulation in humans is incompletely understood. We measured plasma aldosterone in adult subjects with a wide range of obesity and insulin resistance. Aldosterone levels correlated with measures of visceral obesity in one predominantly male cohort and in the women of a second cohort. In the same subjects, aldosterone correlated with insulin resistance. Aldosterone also correlated with plasma cortisol in men and women, and with DHEA-S in women. The data suggested that visceral fat stimulates adrenal steroidogenesis. We found that certain fatty acids stimulated aldosterone production in vitro by rat adrenal cells incubated with rat hepatocytes, but not adrenal cells alone. The results suggested that fatty acids from visceral adipocytes induce hepatic formation of an adrenal secretagogue. This may explain the correlation of plasma steroids with visceral obesity. Aldosterone may contribute to vascular diseases that complicate obesity.