A man, a syndrome, a gene: Clouston's hidrotic ectodermal dysplasia (HED)

This paper presents a biographical sketch of Dr. H. R. Clouston, whose eponym is attached to a type of hidrotic ectodermal dystrophy, and a brief account of the mapping of the gene and its identification as the connexin gene, GJB6.     

Transfection of Lepidopteran insect cells with Baculovirus DNA

Summary Several protocols are presented for preparation and transfection of Baculovirus and plasmid DNAs into Lepidopteran insect cells using the calcium-phosphate co-precipitation technique. Important parameters for optimum efficiency include the inherent susceptibility of the recipient cell line for transfection, and the method of preparation of viral and plasmid DNAs. The protocols presented provide reproducible high efficiencies for transfection of several Lepidopteran cell lines.     

Pulmonary toxicity of trichloroethylene: induction of changes in surfactant phospholipids and phospholipase A2 activity in the mouse lung

Trichloroethylene (TCE) is a common organic solvent in use as a dry cleaning agent as well as an inhalant anesthetic. Nevertheless the effects of this material on the pulmonary surfactant which prevents alveolar collapse at maximal expiration is not known. Therefore, we have examined the effect of TCE on the intra- and extracellular surfactant pools and the activity of phospholipase A2, an enzyme which controls the remodeling of phosphatidylcholine to dipalmitoylphosphatidylcholine, the primary constituent of the pulmonary surfactant. Male CD-1 mice were treated ip with 2500 or 3000 mg/kg TCE. Twenty-four hours later mice were anesthetized and the lungs lavaged. Mice were then killed, the lungs perfused and excised, and subcellular fractions including lamellar bodies prepared. Some lungs were prepared for ultrastructural examination. Phospholipase A2 was assayed in all subcellular fractions. Phospholipid was assayed in the lavage (extracellular surfactant) and the lamellar bodies (intracellular surfactant). TCE (2500 mg/kg) caused selective exfoliation of Clara cells. However, only the dose of 3000 mg/kg TCE produced a significant decrease in the intracellular surfactant phospholipid. Minimal changes occurred in the phospholipid profiles. Phospholipase A2 specific activity was significantly decreased at both dosages within the lung microsomal fraction. In addition after treatment with 3000 mg/kg TCE the enzyme activity in the lamellar body fraction was significantly increased. These data suggest that inhalation of TCE may damage the enzymes which are responsible for synthesizing the pulmonary surfactant resulting in lower amounts of surfactant being stored and available for secretion into the alveolus.     

Evolution of neurotransmitter receptor systems

The presence of hormones, neurotransmitters, their receptors and biosynthetic and degradative enzymes is clearly not only associated with the present and the recent past but with the past several hundred million years. Evidence is mounting which indicates substantial conservation of protein structure and function of these receptors and enzymes over these tremendous periods of time. These findings indicate that the evolution and development of the nervous system was not dependent upon the formation of new or better transmitter substances, receptor proteins, transducers and effector proteins but involved better utilization of these highly developed elements in creating advanced and refined circuitry. This is not a new concept; it is one that is now substantiated by increasingly sophisticated studies. In a 1953 article discussing chemical aspects of evolution (Danielli, 1953) Danielli quotes Medawar, "... endocrine evolution is not an evolution of hormones but an evolution of the uses to which they are put; an evolution not, to put it crudely, of chemical formulae but of reactivities, reaction patterns and tissue competences." To also quote Danielli, "In terms of comparative biochemistry, one must ask to what extent the evolution of these reactivities, reaction patterns and competences is conditional upon the evolution of methods of synthesis of new proteins, etc., and to what extent the proteins, etc., are always within the synthetic competence of an organism. In the latter case evolution is the history of changing uses of molecules, and not of changing synthetic abilities." (Danielli, 1953). Figure 4 outlines a phylogenetic tree together with an indication of where evidence exists for both the enzymes that determine the biosynthesis and metabolism of the cholinergic and adrenergic transmitters and their specific cholinergic and adrenergic receptors. This figure illustrates a number of important points. For example, the evidence appears to show that the transmitters and their associated enzymes existed for a substantial period before their respective receptor proteins. While the transmitters and enzymes appear to exist in single cellular organisms, there is no solid evidence for the presence of adrenergic or cholinergic receptors until multicellular organisms where the receptors appear to be clearly associated with specific cellular and neuronal communication (Fig. 4). One can only speculate as to the possible role for acetylcholine and the catecholamine in single cell organisms.(ABSTRACT TRUNCATED AT 400 WORDS)     

Serine/threonine protein phosphatases and synaptic inhibition regulate the expression of cholinergic-dependent plateau potentials

We previously identified cholinergic-dependent plateau potentials (PPs) in CA1 pyramidal neurons that were intrinsically generated by interplay between voltage-gated calcium entry and a Ca(2+)-activated nonselective cation conductance. In the present study, we examined both the second-messenger pathway and the role of synaptic inhibition in the expression of PPs. The stimulation of m1/m3 cholinergic receptor subtypes and G-proteins were critical for activating PPs because selective receptor antagonists (pirenzepine, hexahydro-sila-difenidol hydrochloride, 4-diphenylacetoxy-N-methylpiperidine methiodide) and intracellular guanosine-5'-O-(2-thiodiphosphate) prevented PP generation in carbachol. Intense synaptic stimulation occasionally activated PPs in the presence of oxytremorine M, a cholinergic agonist with preference for m1/m3 receptors. PPs were consistently activated by synaptic stimulation only when oxytremorine M was combined with antagonists at both GABA(A) and GABA(B) receptors. These latter data indicate an important role for synaptic inhibition in preventing PP generation. Both intrinsically generated and synaptically activated PPs could not be elicited following inhibition of serine/threonine protein phosphatases by calyculin A, okadaic acid, or microcystin-L, suggesting that muscarinic-induced dephosphorylation is necessary for PP generation. PP genesis was also inhibited following irreversible thiophosphorylation by intracellular perfusion with ATP-gamma-S. These data indicate that the expression of cholinergic-dependent PPs requires protein phosphatase-induced dephosphorylation via G-protein-linked m1/m3 receptor(s). Moreover, synaptic inhibition via both GABA(A) and GABA(B) receptors normally prevents the synaptic activation of PPs. Understanding the regulation of PPs should provide clues to the role of this regenerative potential in both normal activity and pathophysiological processes such as epilepsy.     

Multiple Response Comparison of Parietal EEG and Frontalis EMG Biofeedback

In Study I parietal EEG and frontalis EMG were simultaneously recorded from 20 normal subjects while half of the subjects received 45 min of eyes-closed alpha EEG enhancement feedback and the other half received a similar amount of EMG suppression feedback. EMG feedback resulted in a significant reduction in frontalis EMG activity accompanied by a reliable increase in parietal alpha density, while EEG feedback produced only an increase in alpha without corresponding EMG reduction. In Study II, each of 8 subjects underwent four separate feedback contingencies in two 40-min sessions—one session with eyes open and the other with eyes closed. The four types of feedback were: a) alpha-up (alpha enhancement), b) alpha-down (alpha suppression), c) EMG-down (frontalis tension decrease), and d) EMG-up (frontalis tension increase). EMG feedback, up and down, resulted in the more consistent pattern of generalized arousal changes reflected in heart rate and respiratory rate as well as EEG and EMG activity. Within the constraints of a limited training period, the results suggest that frontalis EMG feedback is the more efficient procedure for producing a generalized relaxation response. However, since fingertip vasoconstriction accompanied all four types of feedback, caution must be exercised to avoid the oversimplification of generalized organism effects.