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Biomedical subjects

A L Miller

Publications and source records attributed to A L Miller.

At least 127 records · Page 7Linked to original sources
PubMed1987

Identification of polypeptides associated with sarcolemmal vesicles enriched in orthogonal arrays.

Electron microscopy of freeze-fracture replicas from the sarcolemmas of fast-twitch muscle fibers reveals orthogonal arrays of particles. The biochemical nature of macromolecules associated with the sarcolemmal orthogonal array was investigated using muscle fragments and isolated sarcolemmal vesicles. Muscle fragments incubated in vitro with the lectin concanavalin A exhibited a clustering of orthogonal arrays into local patches. Treatment with other lectins did not result in the clustering of arrays. Clustering was inhibited by the addition of alpha-methyl-D-mannoside, a ligand which also binds concanavalin A. These results suggest that the orthogonal arrays (or associated components) specifically bind concanavalin A. Sarcolemmal vesicles from rabbit sacrospinalis (SAC) and rat extensor digitorum longus (EDL) (both primarily fast-twitch) and rat soleus (SOL) (primarily slow-twitch) were obtained by a combination of low-salt fractionation and sucrose density gradient centrifugation. SDS-polyacrylamide gel electrophoresis (SDS-PAGE) of proteins and glycoproteins solubilized from these vesicles revealed several bands. Four of these bands were present in gels from both the rabbit and rat fast-twitch muscle sarcolemmal preparations (that contained arrays), yet were absent in gels from rat slow-twitch muscle sarcolemmal preparations (not bearing arrays). An enrichment in vesicles containing arrays was achieved by binding SAC sarcolemmal vesicles to Con A-Sepharose 4B beads. SDS-PAGE analysis of array-enriched vesicles from the concanavalin A beads revealed enrichment of three major bands at Mr 93,000, 54,000 and 49,000. These enriched bands correlate with three of the four bands common to fast-twitch EDL and SAC, yet absent in slow-twitch SOL sarcolemmal preparations. We conclude that at least one macromolecular component associated with the sarcolemmal orthogonal array is a concanavalin A binding glycoprotein. We further conclude that three candidates for this component co-purify with the morphological array, and have approximate molecular weights of 93,000, 54,000 and 49,000.

Adenosine Triphosphatases↗
PubMed1987

Disposition of homovanillic acid in the primate.

Prior studies have shown that homovanillic acid is the principal metabolite of dopamine in the primate central nervous system (CNS). In studies of primates given deuterated homovanillic acid systemically, however, only 50% of the administered amounts have been recovered in the urine over the next 4-48 hr. These findings have left it unclear whether there is a slowly turning-over compartment of homovanillic acid, conversion of homovanillic acid to another compound, or excretion of homovanillic acid from the body by a nonrenal route. We synthesized [3H]homovanillic acid and administered it intravenously to four rhesus monkeys. Over the subsequent 4 hr, 94.9 +/- 8.9% (SD) of the administered radioactivity was recovered in the urine, almost entirely as homovanillic acid. These results are consistent with the interpretation that, in primates, there is not a major body pool of homovanillic acid with slow turnover, nor is metabolism to other compounds significant, nor is there evidence for nonrenal excretion.

Animals↗
PubMed1987

Measurements of regional glucose metabolism in studies of motor control.

We used the incorporation of tracer quantities of radiolabeled glucose analogues into muscle and CNS tissues to determine, first, whether unilateral dorsal root deafferentation affects hindlimb flexor and extensor muscles of that side equally. For this we compared the incorporation of [3H]fluorodeoxyglucose into selected muscles in the intact and the deafferented side. The muscles were dissected out after exercise on a treadmill, and tracer incorporation was measured by scintillation counting. Second, we used the [14C]2-DG radioautographic method of Sokoloff to seek confirmation of the involvement of certain CNS structures in locomotor control. We found fascinating side to side asymmetries in glucose metabolism, in the cervical spinal cord, motor cortex, and caudate nucleus. There is still debate about the manner in which "neuronal function", regional blood flow, and glucose metabolism, may be coupled (or even on the existence of such coupling). However, these methods can be useful by their anatomical resolution, and the possibility of making multiple regional measurements from a single subject.

Animals↗
PubMed1986

Characterization of lysosomes and lysosomal enzymes from Chediak-Higashi-syndrome cultured fibroblasts.

Chediak-Higashi-syndrome cultured skin fibroblasts were used to study the possible involvement of lysosomal enzymes and lysosomal dysfunction in this disorder. Our evidence indicated that Chediak-Higashi fibroblasts displayed a significant decrease in the specific activity of the acidic alpha-D-mannosidase (pH 4.2) compared with normal controls. Additional studies revealed a small, but significant, decrease in the rate of degradation of 125I-labelled beta-D-glucosidase that had been endocytosed into Chediak-Higashi cells.

Cells, Cultured↗
PubMed1986

Heterogeneity of N-acetylglucosamine 1-phosphotransferase within mucolipidosis III.

The primary defect responsible for mucolipidosis III is a deficiency of UDP-N-acetylglucosamine:lysosomal enzyme N-acetylglucosamine 1-phosphotransferase activity (GlcNAc phosphotransferase). Genetic complementation analysis of cultured fibroblasts derived from 12 patients with mucolipidosis III identified complementation groups A, B, and C (Honey, N. K., Mueller, O. T., Little, L. E., Miller, A. L., and Shows, T. B. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 7420-7424). The GlcNAc phosphotransferase activity present in the cell lines comprising the complementation groups was characterized with respect to endogenous substrates and two exogenous acceptors, alpha-methyl-D-mannoside and high mannose glycopeptides. All group C cell lines and one group A cell line were found to have normal GlcNAc phosphotransferase activity levels at 37 degrees C when screened with these exogenous acceptors. The enzyme activity in group A cell lines was within normal range when assayed at 23 degrees C. Inhibition of the phosphorylation of alpha-methyl-D-mannoside in the presence of increasing amounts of endogenous substrate N-acetyl-beta-D-hexosaminidase B was demonstrated in normal cell lines at 23 and 37 degrees C and in group A cells at 23 degrees C. However, group C cell lines did not show any inhibition at either temperature. This suggests that the alteration of the GlcNAc phosphotransferase from individuals in group C affects the recognition site for the protein portion of lysosomal enzymes, whereas group A individuals have mutations which result in a temperature-sensitive enzyme.

Cell Line↗
PubMed1986

Regional glucose and beta-hydroxybutyrate use by developing rat brain.

Rates of glucose and D-beta-hydroxybutyrate use were determined in five brain regions of 20-day-old rats. The regions studied were cerebral cortex, thalamus, striatum, cerebellum, and brain stem. The tracers for determining rates of substrate use were [3H]fluorodeoxyglucose and [3-14C]-D-beta-hydroxybutyrate. Two or five minutes after isotope administration the animals were sacrificed in a 6-kW, 2450-MHz focused microwave device. Ten minutes prior to isotope administration the animals were injected intraperitoneally with normal saline or DL-beta-hydroxybutyrate (10 mmol/kg). Blood D-beta-hydroxybutyrate levels averaged 0.21 mumol/ml in saline-injected and 3.13 mumol/ml in hyperketonemic rats. Rates of glucose utilization were significantly heterogeneous between regions in both groups: thalamus greater than cerebral cortex greater than or equal to striatum greater than brain stem greater than cerebellum. These rates were 20-35% lower in hyperketonemic rats. Rates of D-beta-hydroxybutyrate use varied significantly between regions only in the saline group, with the brain stem rate being significantly lower than that in cortex or cerebellum. Regional rates of D-beta-hydroxybutyrate use did not correlate significantly with regional rates of glucose use in either the saline or the hyperketonemic groups. Regional rates of glucose use were strongly and positively correlated between conditions, as were regional rates of D-beta-hydroxybutyrate use. Thus, in 20-day-old rats, the regional heterogeneity of brain glucose use is similar to that in adult rats. D-beta-Hydroxybutyrate use is much less regionally heterogeneous.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Hydroxybutyric Acid↗
PubMed1986

Cerebral metabolic effects of organophosphorus anticholinesterase compounds.

Rats treated intravenously with an organophosphorus anticholinesterase compound, paraoxon or soman, were sacrificed 2 to 131 min later, using 0.7 sec of focused microwave irradiation (25 kW at 915 MHz). Brain regional rates of glucose utilization during 3-min intervals were determined with labeled glucose and fluorodeoxyglucose as tracers. Levels of glucose, lactate, ATP, and creatine phosphate were assayed in the same samples. The two compounds differed markedly in their effects on brain metabolism. Paraoxon (0.8 LD50) depressed rates of glucose use in all brain regions, without causing consistent changes in brain metabolite levels. This depressant effect was most pronounced during the first 30 min after toxin exposure and had largely disappeared by 2 hr. Soman (0.8-0.95 LD50) was variable in its effects. Animals that showed seizure-like behavior had marked increases in glucose use in diencephalon and cerebrum but no changes in cerebellum or brain stem. Rapid rates of glucose use were associated with high levels of lactic acid and lower levels of creatine phosphate. In cerebrum, but not diencephalon, levels of ATP fell by as much as 50% in strongly affected animals by 30-130 min after soman. All of these effects were reversible with atropine. Soman-treated animals that did not have seizure-like activity did not exhibit these brain metabolic changes. These results and those of others show that cholinergic compounds vary greatly in their effects on brain glucose and energy metabolism. Although noncholinergic mechanisms are a possibility, the most parsimonious explanation for these findings is that cholinesterase inhibitors vary in their affinity for different central nervous system (CNS) acetylcholine receptor populations.

Adenosine Triphosphate↗
PubMed1986

Evaluation of strategy for testing thyroid function applied to hypothyroidism.

A strategy for testing thyroid function with discretionary assay of thyroid stimulating hormone (TSH) following initial measurement of total plasma thyroxine (tT4) was retrospectively evaluated in relation to the diagnosis of primary hypothyroidism. Over a two year period 14 641 tT4 assays were done and 6887 TSH assays, of which 29% were initiated in the laboratory. The percentage of raised TSH values (over 5 mU/1) was similar to those for clinician and pathologist initiated requests (23.2% and 23.6%). Some TSH requests (1620) were cancelled; when 212 of these were subsequently analysed the TSH value was raised in 5.7% The incidence of raised TSH values in 188 patients with a tT4 below 100 nmol/1 (7.8 micrograms/100 ml), in whom TSH was not requested, was 5.3% Most of these raised TSH results could be readily explained by information available to the requesting clinician but not given on the request form.

Adult↗
PubMed1985

I-cell disease and pseudo-Hurler polydystrophy: heterozygote detection and characteristics of the altered N-acetyl-glucosamine-phosphotransferase in genetic variants.

The human disorders I-cell disease and pseudo-Hurler polydystrophy (also known as mucolipidosis II and III, respectively) are caused by an inherited deficiency of UDP-GlcNAc: lysosomal enzyme precursor GlcNAc-P transferase activity. The most common genetic variants of these diseases (complementation group A) can be identified in homozygotes and heterozygotes using a GlcNAc-P transferase assay with artificial acceptors and commercially available radiochemicals. The kinetic characteristics of the residual GlcNAc-P transferase activity in complementation group A fibroblasts indicates that the low activity is due to a low Vmax. The measured Michaelis-Menten constants for the substrates UDP-GlcNAc and alpha-methyl mannoside are in the normal range. Homozygotes and heterozygotes of another less common variant of pseudo-Hurler polydystrophy (complementation group C) have normal activity and normal kinetic characteristics with this assay using alpha-methyl mannoside as the acceptor substrate. Several PHP variants with unusual characteristics are discussed.

Cells, Cultured↗
PubMed1985

Effects of sensory deafferentation on glucose metabolism of muscles during locomotion.

We measured the uptake of a radiolabeled analogue of glucose ([3H]FDG) into muscles during treadmill walking so as to determine whether sensory deafferentation of a limb affects the muscles' metabolic response to the exercise. The muscles on the deafferented side took up less tracer than those in the intact side with exercise, but not at rest. Extensor muscles were more affected than flexors by deafferentation. This agrees with the proposal that "reflex" inputs adjust the level of recruitment of motor units by central pattern generators.

Afferent Pathways↗
PubMed1985

Rat liver alkaline phosphatases. Evidence hepatocyte and portal triad enzymes differ.

Using biochemical and electron microscopic histochemical techniques, we studied membrane-bound alkaline phosphatase activities of rat hepatocytes and portal triads. Activity in portal triads was localized to capillaries surrounding bile ducts (peribiliary plexus) and arterioles. Despite the reputation of alkaline phosphatase as a "biliary enzyme," activity was not observed in bile ducts. Livers were separated into hepatocyte and portal triad fractions with collagenase. Enzyme from hepatocytes migrated faster during electrophoresis and eluted later during anion-exchange chromatography than that from portal triads. Thus, hepatocyte enzyme is more negatively charged (and also possibly smaller) than portal triad enzyme. Twelve hours after bile duct obstruction, new activity appeared on lateral and sinusoidal membranes of hepatocytes; appearance of portal triads did not change with obstruction. Electrophoretic mobilities of the two forms were not altered by obstruction. We conclude that two distinct liver alkaline phosphatases exist, one in hepatocytes, the other in portal triad blood vessels.

Alkaline Phosphatase↗
PubMed1985

Lithium and impairment of renal concentrating ability.

In 19 patients on lithium maintenance therapy, impairment of renal concentrating ability was found to correlate with daily lithium dose. The correlation between dose and impaired renal concentrating ability was strongest for patients on regular lithium preparations and not significant for patients on a slow release preparation. In 2 patients with severe polyuria, renal concentrating ability increased markedly with thiazide treatment. No relationship between lithium dosing parameters and other side effects was found. These results emphasize the importance of dose and type of preparation of lithium in the production of renal side effects.

Adult↗
PubMed1985

Intestinal function and intestinal blood supply: a 20 year surgical study.

The experience of The Middlesex Hospital in the investigation and management of chronic intestinal arterial occlusion (IAO) over a 20 year period is presented. Of some 100 patients referred only 28 were found to have IAO and these were critically evaluated in terms of insorptive and exsorptive function. No relation was found between intestinal performance and the degree of potential ischaemia suggested by angiography. Twenty two patients underwent reconstruction of the coeliac axis and mesenteric arteries, with three perioperative deaths. The remainder were followed for periods ranging between six months and 10 years. Of these, 15 are asymptomatic, one is unchanged, and one is subjectively worse. The role of elective surgery in relation to the visceral arteries is critically discussed in the light of these results, and in the perspective of the world experience.

Adult↗
PubMed1985

Factors affecting amnesia, seizure duration, and efficacy in ECT.

Twenty-nine patients given unilateral ECT were tested for memory with each treatment. Forgetting of nonverbal material correlated positively with seizure duration and with anesthetic dose. Seizure duration did not correlate with forgetting of verbal material or with changes in Hamilton depression ratings. Seizure duration was inversely related to succinylcholine and methohexital doses. These findings suggest that muscle relaxant and anesthetic doses can be adjusted to lessen the amnestic effects of ECT. There are, however, insufficient data on the relationship between seizure length and ECT efficacy to specify a minimum duration for seizures, individually or cumulatively.

Amnesia↗
PubMed1985

Lysosomal cystine storage in cystinosis and mucolipidosis type II.

Cultured fibroblasts from mucolipidosis II (ML-II) patients demonstrated an elevated cystine content which increased with time in culture compared to fibroblasts from cystinotic patients or normal controls under the same conditions. In both cystinotic and ML-II cells the increased levels of cystine could be derived either from endogenous proteolysis or from in vitro supplementation of the cultured cells with cysteine-glutathione mixed disulfide. Cystine was depleted from both cell types by cysteamine. When cysteamine was replaced with complete medium, the cystine reaccumulated in both cystinotic and ML-II cells within 24 h, although a lag of 4 h was seen with ML-II cells. The intracellular location of the increased cystine in cultured fibroblasts was examined utilizing free-flow electrophoresis and found to be in the purified population of secondary lysosomes of both cystinotic and ML-II cells. White blood cell and hepatic cystine, which was greatly increased in cystinotic patients, was not elevated in ML-II patients. Compared to normal control fibroblasts the efflux of cystine from isolated granular fractions was virtually absent in cystinotic fibroblasts and considerably reduced in ML-II fibroblasts. The examination of such similarities and differences in cystine accumulation and transport in tissues from cystinotic and ML-II patients has provided some insight into the defects in these diseases.

Cells, Cultured↗