Evidence for Functional Activity of Up-Regulated Nicotine Binding Sites in Rat Striatal Synaptosomes

A number of studies have found that the chronic administration of nicotine causes an increase in the density of nicotinic binding sites in the brain, but it is not known whether these additional binding sites are functionally active receptors. In this study, the effects of 1-week administration of the potent nicotinic agonist, (+)-anatoxin-a (96 nmol/day via osmotic minipumps), was assessed on [3H]nicotine binding and [3H]dopamine uptake and release in rat striatal synaptosomes. Chronic (+)-anatoxin-a treatment resulted in a 32% increase in the Bmax of [3H]nicotine binding in anatoxin-treated animals compared to control. There was a 43% increase in the activity of 3 microM nicotine to release [3H]dopamine from synaptosomes of anatoxin-treated animals, but the release induced by 20 mM K+ depolarization was unaffected. There was no effect of chronic (+)-anatoxin-a treatment on the uptake of [3H]dopamine. A strong positive correlation (r = 0.64) was found between the density of [3H]nicotine binding sites and the nicotine-induced stimulation of [3H]dopamine release in individual animals. These results indicate that (+)-anatoxin-a, like nicotine, produces an up-regulation of nicotine binding sites following chronic administration, and that these additional sites are functional receptors capable of mediating the release of dopamine from striatal synaptosomes.     

Presymptomatic testing for Huntington's disease

Summary Presymptomatic testing for Huntington's disease (HD) is possible through the use of restriction fragment length polymorphisms (RFLPs) at the closely linked D4S10 locus. Recombination between the HD and D4S10 loci will occur in 4%–5% of meioses, and is a well-recognised complication of predictive testing. Recombination between RFLPs within the D4S10 locus is a rare event and can usually be ignored. We report a case where such an intra-locus recombination frustrated attempts to predict the chance of a high-risk individual inheriting the HD gene.     

The expression of a highly expressed Bacillus subtilis gene is not reduced by introduction of multiple codons normally not present in such genes

A recombinant DNA Proteus mirabilis L-form expression system, LVI (pJS127), was used to synthesize human fusion interferon alpha 1 (f-IFN-alpha 1). In the expression plasmid used, the complete coding sequence of IFN-alpha 1 was linked to the streptococcal speA promoter and the 5' end of the speA structural gene including its signal sequence coding region. LVI (pJS127) was capable of complete secretion into the culture medium of biologically active f-IFN-alpha 1 whose identity was confirmed by immunological and chemical evidence. In particular, bacterial L-forms were for the first time shown to be capable of correct signal peptide processing, as determined by N-terminal sequencing of the secreted f-IFN.     

Experimental investigations of benthic reentry by migrating meiobenthic copepods

The resettlement behavior of meiobenthic copepods, which actively migrated from sediments in a seagrass bed, was investigated in a shallow subtidal area in Tampa Bay, Florida, U.S.A. Experimental studies were conducted to determine whether meiobenthic copepods after emerging from sediments at sunset reenter the sedimentary substratum or select other subhabitats, water and seagrass blades. Migrating copepods were collected with emergence traps and transferred to experimental aquaria in the field which contained sediment, seagrass-blade and water treatments. Settlement into each type of treatment was measured in separate 2-h and 9-h experiments. Differences in densities of copepod taxa retrieved from emergence traps and introduced into experimental aquaria were recorded as were differing relative proportions of each copepod species returning to the substratum treatments. Settlement patterns of total copepods and three dominant copepod species, Zausodes arenicolus, Halicyclops sp. and Robertsonia hamata, departed from those expected by chance. The populations of R. hamata and Halicyclops sp. which settled were generally skewed towards males and a close matching of males and copepodites within treatment dishes was evident. Similar to nighttime-emergence patterns, timing and magnitude of postmigration reentry differs among copepod taxa and such reentry may be linked to reproductive events. Complex behavioral processes previously noted for fish and macrofaunal organisms in seagrass beds may also be important in recruitment and reassortment of meiobenthic copepods.     

Heat treatment of ripening apples: Differential effects on physiology and biochemistry

Apples ( Malus domestica Borkh.) were heated for 4 days at 38°C immediately after harvest and then placed at 20°C for 7–10 days. Protein synthesis, ethylene production and fruit softening were reversibly inhibited by the heat treatment. Fruit respiration, membrane permeability and chlorophyll degradation in the fruit peel were enhanced during the treatment. The heat-treated apples ripened normally but more slowly than untreated apple We hypothesize that heat treatment differentially affects processes which normally increase simultaneously during fruit ripening, by inhibiting those processes which require tie novo protein synthesis and enhancing those that do not.     

Control of Continuous Irradiation Injury on Potatoes with Daily Temperature Cycling

Two controlled-environment experiments were conducted to determine the effects of temperature fluctuations under continuous irradiation on growth and tuberization of two potato (Solanum tuberosum L.) cultivars, Kennebec and Superior. These cultivars had exhibited chlorotic and stunted growth under continuous irradiation and constant temperatures. The plants were grown for 4 weeks in the first experiment and for 6 weeks in the second experiment. Each experiment was conducted under continuous irradiation of 400 micromoles per square meter per second of photosynthetic photon flux and included two temperature treatments: constant 18°C and fluctuating 22°C/14°C on a 12-hour cycle. A common vapor pressure deficit of 0.62 kilopascal was maintained at all temperatures. Plants under constant 18°C were stunted and had chlorotic and abscised leaves and essentially no tuber formation. Plants grown under the fluctuating temperature treatment developed normally, were developing tubers, and had a fivefold or greater total dry weight as compared with those under the constant temperature. These results suggest that a thermoperiod can allow normal plant growth and tuberization in potato cultivars that are unable to develop effectively under continuous irradiation.     

Immunogold localization of ribulose-1,5-bisphosphate carboxylase/oxygenase in the nitrogen-fixing cyanobacterium,Plectonema boryanum

Summary Antiserum against the Calvin cycle enzyme, ribulose-1,5-bisphosphate carobxylase/oxygenase (RuBisCO), was used in conjunction with colloidal gold to localize RuBisCO in nitrogen-fixing (fix+) and nonfixing (fix–)Plectonema boryanum cells. RuBisCO antiserum consistently labeled the cytoplasm and polyhedral bodies (carboxysomes) in both fix+ and fix– cells. Through morphometry, it was determined that significantly less gold label (indicative of RuBisCO) was present in fix+ cells. This decreased RuBisCO content correlated with a decrease in net photosynthetic oxygen evolution also observed in fix+P. boryanum.Abbreviations RuBisCO Ribulose-1,5-bisphosphate carboxylase/oxygenase - fix+ nitrogen-fixing - fix– nonfixing     

Mutation analysis of the cystic fibrosis transmembrane regulator gene in native American populations of the southwest

We report DNA and clinical analyses of cystic fibrosis (CF) in two previously unstudied, genetically isolated populations: Pueblo and Navajo Native Americans. Direct mutation analysis of six mutations of the CFTR gene--namely, delta F508, G542X, G551D, R553X, N1303K, and W1282X--was performed on PCR-amplified genomic DNA extracted from blood samples. Haplotype analyses with marker/enzyme pairs XV2c/TaqI and KM19/PstI were performed as well. Of the 12 affected individuals studied, no delta F508 mutation was detected; only one G542X mutation was found. None of the other mutations was detected. All affected individuals have either an AA, AC, or CC haplotype, except for the one carrying the G542X mutation, who has the haplotype AB. Clinically, six of the affected individuals examined exhibit growth deficiency, and five (all from the Zuni Pueblo) have a severe CF phenotype. Four of the six Zunis with CF are also microcephalic, a finding not previously noted in CF patients. Our DNA data have serious implications for risk assessment of CF carrier status for these people.     

Evidence for a composite phylogenetic origin of the plastid genome of the brown alga Pylaiella littoralis (L.) Kjellm

The nucleotide sequence and the 5 flanking region of the rbcL gene coding for the large subunit of ribulose bisphosphate-1,5-carboxylase/oxygenase of Pylaiella littoralis, a brown alga, has been determined and the deduced amino-acid sequence has been compared to those of various photosynthetic and chemoautotrophic Eubacteria, of a red alga and of green plastids (Euglena gracilis, green algae and higher plants). Unlike the rbcL genes of green plastids which are more closely related to those of cyanobacteria the P. littoralis rbcL gene is more closely related to that of a -purple bacterium, as was found for the rbcS gene of another chromophytic alga [Boczar et al., Proc Natl Acad Sci USA 86: 4996–4999, 1989]. Matrix data of homology between the rbcL gene of P. littoralis and the same gene of other organisms are presented. Based on our previous report, the gene coding for the 16S rRNA from P. littoralis is closely related to that of E. gracilis (Markowicz et al., Curr Genet 14: 599–608, 1988). We suggest that the large plastid DNA molecule of P. littoralis is a phylogenetically composite genome which probably resulted from mixed endosymbiosis events, or from a horizontal transfer of DNA.     

DNA fingerprinting of lactococci and streptococci used in dairy fermentations

Summary A DNA fingerprinting procedure was developed for strains of Lactococcus lactis subsps. lactis and cremoris, biovar. diacetylactis, and Streptococcus salivarius subsp. thermophilus, used in dairy fermentations. Total cellular DNA was extracted and digested with restriction endonucleases, HindIII or HaeIII, followed by separation of the fragments using agarose gel electrophoresis. L. lactis C2 was used as a representative strain for examining the effect of growth phase and cell concentration, cell washing conditions prior to lysis, type and concentration of the enzyme used to digest the cell wall, composition of the lysis buffer, and gel electrophoresis conditions. Following optimization of the fingerprinting procedure, electrophoretic migration of fragments from 23 strains produced reproducible gel patterns. L. lactis subsp. lactis strains ML3 and C2 appeared to be identical when restrricted with either Hind III or HaeIII. Similarly, S. salivarius subsp. thermophilus strains 19987 and 19258, and L. lactis subsp. cremoris strains 134 and C3, appeared to have identical DNA fingerprints following digestion with HindIII. To determine the usefulness of this technique for monitoring population changes during fermentation, various ratios of two closely related strains were inoculated into milk and allowed to grow for 16 h at 32° C. The initial inoculum ratios were determined by standard plate counts, and the final ratio was deterimined by DNA fingerprinting. DNA fingerprinting will be useful in the identification, characterization, and comparison of food fermentation microorganisms.Published as paper No. 17,803 of the contribution series of the Minnesota Agricultural Experiment Station Offprint requests to: S. K. Harlander