William E. Wilson

Effect of Acrylamide on Neurotransmitter Metabolism and Neuropeptide Levels in Several Brain Regions and Upon Circulating Hormones

The effect of acute and subchronic acrylamide treatment on levels of dopamine, serotonin, and their metabolites was determined in several brain regions of the rat. Concentrations of several neuropeptides and circulating hormones were also measured. Both a single and repeated doses of acrylamide resulted in elevated levels of 5-hydroxyindolacetic acid in all regions studied (frontal cortex, striatum, hippocampus, brain stem, and hypothalamus). Changes in regional content of other monoamines were much less pronounced. Turnover studies following pargyline blockage of monoamine oxidase, suggested results were due to increased rates of serotonin turnover in acrylamide-treated rats.Changes in neuropeptide levels were only detected in the hypothalamus where a single acrylamide treatment caused elevated levels of β-endorphin and substance P, and in frontal cortex where met-enkephalin levels were higher after repeated acrylamide injection. Such repeated injection caused a major depression in plasma levels of testosterone and prolactin.

Flow cytometric immunophenotyping of cerebrospinal fluid.

Leptomeningeal disease is an important adverse complication occurring in patients with B and T cell lymphomas and acute leukemias of lymphoid and myeloid origin. Recent reports suggest that multiparameter flow cytometry immunophenotypic assessment of spinal fluid samples could improve the efficiency of detection of CNS involvement, due to its high specificity and greater sensitivity. However, spinal fluid samples are frequently paucicellular with a rapidly decreasing cell viability. Staining of spinal fluid therefore requires dedicated sample storage/transport, staining, and preparation protocols. The Basic Protocol in this unit outlines a consensus multiparameter (3‐ to 8‐color) flow cytometry immunophenotypic protocol for the evaluation of CNS involvement of cerebrospinal fluid (CSF) samples by neoplastic cells. A Support Protocol describing the simultaneous assessment of surface and cytoplasmic antigens is also provided. Finally, in the Alternate Protocol, we describe a method to calculate absolute numbers of both normal and pathological cell subpopulations by adding counting beads to the assay. Curr. Protocol. Cytom. 45:6.25.1‐6.25.16.

Interaction of deltorphin with opioid receptors: molecular determinants for affinity and selectivity.

Opioid receptor analyses of deltorphin A (H-Tyr-D-Met-Phe-His-Leu-Met-Asp-NH2) analogues indicated the following: (a) increased negativity differentially affected affinities (Ki) and selectivity (Ki mu/Ki delta); (b) shifted sequence heptapeptides, [Asp5,Leu6,Met-NH2(7)] and [Asp4,His5,Leu6,Met-NH2(7)], reversed selectivity (delta-->mu); (c) substitutions at positions 4, 5, and 6 diminished selectivity, with changes in residue 5 being the most detrimental; (d) C-terminal deletions differentially effected Ki. These are the first data to demonstrate a reversal of delta selectivity in heptapeptides containing a negative charge and indicate that modifications in affinity occur through changes in both anionic and hydrophobic properties of residues at specific positions in the peptide. Deltorphin analogues might also be applied to differentiate between opioid receptor subsites.

Effects of chlordecone exposure on brain neurotransmitters: Possible involvement of the serotonin system in chlordecone-elicited tremor

The purpose of this study was to correlate the chlordecone-elicited tremor activity with alterations of brain neurotransmitters. A single injection of chlordecone (80 mg/kg, ip) significantly increased the brain levels of 5-hydroxyindoleacetic acid (5-HIAA) but did not affect the concentrations of dopamine, dihydroxphenylacetic acid, aspartate, taurine, glutamate, glycine, and gamma-aminobutyric acid (GABA). There was a dose- and time-related correlation between the increases in striatal 5-HIAA levels and tremor after chlordecone treatment. A subsequent study with pargyline indicated that the increase in striatal 5-HIAA level represented an increase in the turnover of serotonin. This study plus the previous finding that pizotifen (BC-105), a serotonin receptor blocker, attenuated chlordecone-elicited tremor strongly suggests a possible involvement of the serotonin system in mediating the tremor elicited by this insecticide.

Brain Regional Levels of Neurotransmitter Amines as Neurochemical Correlates of Sex-Specific Ontogenesis in the Rat

Brain regional levels of three neurotransmitter amines - serotonin (5-HT), norepinephrine (NE), dopamine (DA) - were measured in young rats prior to weaning to determine the extent to which modifications in levels of amines might reflect alterations in the sex steroid hormonal environment during the first postnatal week in the life of the rat. Sex-related levels of DA, NE, and 5-HT were found in some brain regions of the 12-day-old rat. Male midbrain DA exceeded the corresponding female value while female hypothalamic NE levels were greater than those of the male. Levels of 5-HT in the corpus striatum and the midbrain of males were greater than those of the female. Castration of the male on day 1 or testosterone propionate (TP) administration to the newborn female resulted in modifications of levels of midbrain 5-HT which reflected feminization of the castrated males and masculinization of the TP-treated females. Castration on day 1, or diethylstilbestrol given on days 2, 4, and 6, resulted in apparent feminization of NE levels in the hypothalamus of 12-day-old male rats. Thus, it appears that regional levels of hypothalamic NE and midbrain 5-HT in the 12-day-old rat may reflect the course of brain organizational activity which becomes recognizable in the adult as sex-specific behavior.

Evolutionary relationship between nonmammalian and mammalian peptides

An hypothesis has been developed to rationalize the evolution of regulatory peptides. In order to account for critical relationships involving peptide regulators, their receptors, and peptide processing enzymes, the following generalizations will be supported: (1) peptides arose from protein precursors as proteolytic digestion by-products and acquired hormonal status during the course of natural selection; (2) initially, peptides served primarily nutritional roles, thereby permitting increased growth rates and reproductive advantages for recipient cells; (3) specific peptide sequences were conserved during evolution and were associated with biological activities which were essential for survival of species as divergent as unicellular organisms, amphibians, and mammals; and (4) regulatory peptides probably arose simultaneously with their membrane-oriented, macromolecular receptor sites. In support of the conservation of sequence information or function, or both, during evolutionary development, evidence has been obtained to indicate that peptide sequences which occur in two classes of amphibian peptides appear to be extensively conserved in mammals. Studies with an antiserum directed against the N-terminal sequence of amphibian physalaemin have permitted the recognition of a mammalian octapeptide which exhibits 80% homology with residues 1-5 in that region. Another study with an antiserum directed against the midregion (sequence 5-8) of amphibian bombesin has indicated the existence of milk peptides which mimic bombesin in several pharmacological bioassays. These studies indicate that radioimmunoassays can be powerful tools in facilitating recognition of peptide sequences conserved throughout evolution.

Correlation of neurochemical and behavioral effects of triethyl lead chloride in rats

Adult male Fischer-344 rats were dosed sc with 1 or 2.5 mg/kg of triethyl lead chloride (TEL) for 5 consecutive days. One week after the last dose, TEL-exposed rats had decreased Met-enkephalin in the hypothalamus, septum, and frontal cortex, while substance P was decreased in the hippocampus and frontal cortex. Dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) in the caudate nucleus were not altered by TEL nor were serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) in the caudate nucleus, hypothalamus, hippocampus, or frontal cortex. In a second experiment, rats were dosed with 1.75 mg/kg sc for 5 days. Subsequent assay of brain tissue indicated that TEL decreased met-enkephalin levels in the septum of rats one and seven days after cessation of dosing; effects on substance P were not observed. TEL-induced decreases in Met-enkephalin in the septum were temporally associated with increased hot plate latencies. One day after cessation of dosing with TEL, concentration of 5-HIAA in the caudate nucleus, hippocampus, frontal cortex, and brain stem, and 5-HT in the hippocampus and brain stem were increased. Biogenic amine concentrations were not affected in any other region or at any other time postdosing. A third experiment indicated that TEL-induced analgesia could be attenuated by 10 mg/kg chlordiazepoxide or 10 mg/kg of naloxone. The present results suggest that TEL-induced analgesia may be due to alterations in emotionality or reactivity to noxious stimuli, which may be associated with the alteration in delta opiate mechanism in the limbic system, such as the change of septal enkephalin neuronal activities.

Complex equilibria involving DDT and model compounds for phospholipid and protein

Abstract Equilibrium constants and values of thermodynamic parameters for complex formation between 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (the pesticide p,p′-DDT) and compounds containing biologically significant functional groups were obtained by proton magnetic resonance. The results imply that phospholipid phosphate and ester groupings, and aromatic substituents in protein are the most likely important binding sites for DDT.

Unique sequence in deltorphin C confers structural requirement for δ opioid receptor selectivity

Abstract A series of deltorphin C (H-Tyr- d -Ala-Phe-Asp-Val-Val-Gly-NH 2 ) analogues were synthesized to assess the consequences of changing anionic and hydrophobic residues on δ receptor selectivity. Analogues with altered C-terminal groups, inverted sequences, or esterified with tert -butyl, benzyl, or ethyl groups revealed that high δ selectivity required an unmodified amino acid sequence. Shifts of Asp and hydrophobic residues decreased δ selectivity due to loss in δ affinity (5- to ≈ 700-fold); μ affinity was unchanged or increased 14-fold. Suppression of charge or deamidation diminished δ selectivity through reduced δ and modified μ affinities. Data provide evidence that a negative charge does not a priori guarantee high selectivity and specific alignment of anionic and hydrophobic residues might facilitate optimum spatial configuration which complements the δ receptor binding site.

A comparison of Na-K-ATPases obtained from brains of adult and fetal rat

1. 1. An examination of several properties of partially purified preparations of Na-K-ATPases from brain tissue microsomes of adult and fetal rats has revealed several property differences other than specific activities. 2. 2. Those properties which differ include (a) apparent Km values for ATPase activation by potassium; (b) apparent Km values for ATP hydrolysis by the Na-K-ATPases; (c) activation of the adult enzyme by the surfactant, Tergitol NPX; (d) activation of the fetal enzyme by phosphatidyl choline; and (e) relative extents of inhibition by 10−5 M ouabain. 3. 3. Both adult and fetal Na-K-ATPases appear to require acidic phospholipids, such as phosphatidyl serine, for stabilization within the plasma membrane. 4. 4. These results provide further evidence that the relative prominence of multiple forms of the Na-K-ATPases varies with brain development.