A novel alpha subunit in rat brain GABAA receptors

Two cDNAs (alpha 1 and alpha 4) from rat brain cDNA libraries encode isoforms of the alpha subunit of the GABA/benzodiazepine receptor, which differ at 30% of their amino acid residues. Northern blot analysis and in situ hybridization histochemistry show that alpha 1 and alpha 4 mRNAs have distinct sizes and distinct regional and cellular distributions in rat brain: both mRNAs are found in the cortex and hippocampus; however, only the alpha 1 mRNA is detected in the cerebellum. We injected RNA transcribed from alpha 1 and alpha 4 cDNAs into Xenopus oocytes, together with an RNA for a rat beta subunit. We obtained GABA-dependent inward currents that were reversibly blocked by picrotoxin. Picrotoxin alone, applied to oocytes producing the alpha and beta polypeptides, elicited an outward current. We suggest that these polypeptides together produce GABA-gated ion channels that can also open spontaneously.     

A hydrophobic, carboxy-proximal region of a light-harvesting chlorophyll a/b protein is necessary for stable integration into thylakoid membranes.

Proteins synthesized as soluble precursors in the cytoplasm of eukaryotic cells often cross organellar membrane barriers and then insert into lipid bilayers. One such polypeptide, the light-harvesting chlorophyll a/b-binding protein (LHCP), must also associate with pigment molecules and be assembled into the photosystem II light-harvesting complex in the chloroplast thylakoid membrane. A study of the import of mutant LHCPs into isolated chloroplasts has shown that a putative alpha-helical membrane-spanning domain near the carboxy terminus (helix 3) is essential for the stable insertion of LHCP in the thylakoid. Protease digestion experiments are consistent with the carboxy terminus of the protein being in the lumen. This report also shows that helix 3, when fused to a soluble protein, can target it to the thylakoids of isolated, intact chloroplasts. Although helix 3 is required for the insertion of LHCP and mutant derivatives into the thylakoid, the full insertion of helix 3 itself requires additionally the presence of other regions of LHCP. Thus, LHCP targeting and integration into thylakoid membranes requires a complex interaction involving a number of different domains of the LHCP polypeptide.     

Perceived contribution of children to marriage and its effects on family planning behavior.

The impact of the perceived consequences of having children on a couple's contraceptive practices and the tendency to plan births are examined. 2 separate measures of family planning were utilized, contraceptive pattern and proportion of unplanned pregnancies. It was found that the more a woman viewed children as an essential part of the marital experience the less likely she was to use reliable contraception early in marriage. The influence of the evaluation of children on contraceptive practices also was contingent upon the number of children a couple had had and the number of years they had been married. Differences in incidence of unplanned pregnancies among women was more a result of socioeconomic circumstance and the tendency to have a large family than attitude towards children. These 2 aspects of family planning have different implications for policy decisions. Changing the incidence of unplanned pregnancies would entail attacking those conditions which handicap couples in changing their life circumstances. It would involve educating them to the concept of family planning and the techniques of birth control and also increasing economic opportunity. Changing contraceptive patterns, on the other hand, might involve encouraging alternative adult roles, and creating differential incentives for childbearing.     

Search strategy of sex pheromone-stimulated male German cockroaches

Stimulation of the male German cockroach, Blattella germanica, with contact sex pheromone releases courtship behaviour and then area-restricted local search. Servosphere-recorded search patterns, compared to pre-stimulus pathways, are characterised by increased looping and path crossing and a relatively high rate of turning. Search behaviour can be performed in the absence of visual cues, including information relative to the site of stimulation. Information controlling search motor patterns is at least in part genetic; certain pattern components are generally similar from trial to trial, whereas specific pattern components vary widely.     

Comparison of nine brands of membrane filter and the most-probable-number methods for total coliform enumeration in sewage-contaminated drinking water.

Nine different brands of membrane filter were compared in the membrane filtration (MF) method, and those with the highest yields were compared against the most-probable-number (MPN) multiple-tube method for total coliform enumeration in simulated sewage-contaminated tap water. The water was chlorinated for 30 min to subject the organisms to stresses similar to those encountered during treatment and distribution of drinking water. Significant differences were observed among membranes in four of the six experiments, with two- to four-times-higher recoveries between the membranes at each extreme of recovery. When results from the membranes with the highest total coliform recovery rate were compared with the MPN results, the MF results were found significantly higher in one experiment and equivalent to the MPN results in the other five experiments. A comparison was made of the species enumerated by these methods; in general the two methods enumerated a similar spectrum of organisms, with some indication that the MF method was subject to greater interference by Aeromonas.     

Isolation of new promoter-mediated co-suppressed lines in Arabidopsis thaliana

Four new independent lines that exhibit co-suppression of an introduced cab140::tms2 gene and the native cab140 gene have been isolated in Arabidopsis thaliana. These lines are of particular interest because the homology shared between the introduced and native genes is 1.3 kb of promoter DNA that only contains 14 bp of transcribed region. Most other reported examples of co-suppression involve homologies between transcribed portions of genes. A similar line, lct, had been isolated previously from EMS-mutagenized seeds, and we concluded that this example of co-suppression was probably due to a mutation that mapped at or near the introduced cab140::tms2 gene [Brusslan JA, Karlin-Neumann GA, Huang L, Tobin EM: Plant Cell 5: 667–677 (1993)]. Our observations with these four new lines, however, suggest that an epigenetic event(s) rather than a mutation might be the cause of co-suppression in these and the lct line.     

Mechanism of adsorption of hard and soft metal ions to Saccharomyces cerevisiae and influence of hard and soft anions.

The applicability of the hard-and-soft principle of acids and bases in predicting metal adsorption characteristics in a biological context was investigated for metabolism-independent uptake of the metal ions Sr2+, Mn2+, Zn2+, Cu2+, Cd2+, and Tl+ by Saccharomyces cerevisiae. Metal adsorption increased with external metal concentration (5 to 50 microM), although some saturation of uptake of the harder ions examined, Sr2+, Mn2+, and Zn2+, was evident at the higher metal concentrations. Cation displacement experiments indicated that, with the exception of Tl+, relative covalent bonding (H+ displacement) of the metals was greater at low metal concentrations, while weaker electrostatic interactions (Mg2+ plus Ca2+ displacement) became increasingly important at higher concentrations. These results were correlated with curved Scatchard and reciprocal Langmuir plots of metal uptake data. Saturation of covalent binding sites was most marked for the hard metals, and consequently, although no relationship between metal hardness and ionic/covalent bonding ratios was evident at 10 microM metal, at 50 microM the ratio was generally higher for harder metals. Increasing inhibition of metal uptake at increasing external anion concentrations was partially attributed to the formation of metal-anion complexes. Inhibitory effects of the hard anion SO42(-) were most marked for uptake of the hard metals Sr2+ and Mn2+, whereas greater relative effects on adsorption of the softer cations Cu2+ and Cd2+ were correlated with complexation by the soft anion S2O32(-). Inhibition of uptake of the borderline metal Zn2+ by SO42(-) and that by S2O32(-) were approximately equal.(ABSTRACT TRUNCATED AT 250 WORDS)     

Muscarinic Receptor-Mediated Inositol 1,4,5-Trisphosphate Formation in SH-SY5Y Neuroblastoma Cells Is Regulated Acutely by Cytosolic Ca2+ and by Rapid Desensitization

Abstract: Stimulation of muscarinic receptors expressed in SH-SY5Y human neuroblastoma cells resulted in a complex profile of inositol 1,4,5-trisphosphate (InsP₃) accumulation, with a dramatic increase (six- to eightfold) over the first 10 s (the “peak” phase) and subsequently, from ～60 s onward, maintained at a lower but sustained level (the “plateau” phase). Chelation of extracellular Ca²⁺ with EGTA or inhibition of Ca²⁺ channels with Ni²⁺ showed that the plateau phase was dependent upon Ca²⁺ entry. Furthermore, use of thapsigargin and EGTA to discharge and sequester Ca²⁺ from intracellular stores revealed that Ca²⁺ from this source was capable of supporting the peak phase of the InsP₃ response. Carbachol-stimulated phosphoinositidase C activity in permeabilized SH-SY5Y cells was also shown to be highly dependent on free Ca²⁺ concentration (20–100 nM) and suggests that under normal conditions, InsP₃ formation is enhanced by increases in cytosolic free Ca²⁺ concentration that accompany muscarinic receptor activation. Measurement of carbachol-stimulated total inositol phosphate accumulation in the presence of Li⁺ indicated that the initial rate of phosphoinositide hydrolysis (from 0 to 30 s) was about fivefold greater than that from 30 to 300 s. This rapid but partial desensitization of receptor-mediated phosphoinositide hydrolysis provides strong evidence for the mechanism underlying the changes in InsP₃ accumulation over this time. Because very similar data were obtained in Chinese hamster ovary cells transfected with human m3 receptor cDNA, we suggest that although increases in cytosolic free Ca²⁺ concentration amplify InsP₃ formation during stimulation of m3 muscarinic receptors, the primary factor that governs the profile of InsP₃ accumulation is rapid, but partial, desensitization. Such desensitization does not appear to be mediated by changes in cytosolic Ca²⁺ or protein kinase C activity.