Development of an odd-Z-projectile reaction for heavy element synthesis: 208Pb(64Ni,n)271Ds and 208Pb(65Cu,n)(272)111.

Seven 271Ds decay chains were identified in the bombardment of 208Pb targets with 311.5 and 314.3 MeV 64Ni projectiles using the Berkeley Gas-filled Separator. These data, combined with previous results, provide an excitation function for this reaction. From these results, an optimum energy of 321 MeV was estimated for the production of (272)111 in the new reaction 208Pb(65Cu,n). One decay chain was observed, resulting in a cross section of 1.7(+3.9)(-1.4) pb. This experiment confirms the discovery of element 111 by the Darmstadt Group who used the 209Bi(64Ni,n)(272)111 reaction.

Chemical investigation of hassium (element 108).

The periodic table provides a classification of the chemical properties of the elements. But for the heaviest elements, the transactinides, this role of the periodic table reaches its limits because increasingly strong relativistic effects on the valence electron shells can induce deviations from known trends in chemical properties. In the case of the first two transactinides, elements 104 and 105, relativistic effects do indeed influence their chemical properties, whereas elements 106 and 107 both behave as expected from their position within the periodic table. Here we report the chemical separation and characterization of only seven detected atoms of element 108 (hassium, Hs), which were generated as isotopes (269)Hs (refs 8, 9) and (270)Hs (ref. 10) in the fusion reaction between (26)Mg and (248)Cm. The hassium atoms are immediately oxidized to a highly volatile oxide, presumably HsO(4), for which we determine an enthalpy of adsorption on our detector surface that is comparable to the adsorption enthalpy determined under identical conditions for the osmium oxide OsO(4). These results provide evidence that the chemical properties of hassium and its lighter homologue osmium are similar, thus confirming that hassium exhibits properties as expected from its position in group 8 of the periodic table.

Effects of the CRF(1) receptor antagonist, CP 154,526, in the separation-induced vocalization anxiolytic test in rat pups.

CRF(1) receptor antagonists have been proposed as novel pharmacological treatments for depression, anxiety and stress disorders. The primary goal of the present study was to evaluate the effects of the CRF(1) receptor antagonist, CP 154,526, in the separation-induced vocalization (SIV) model of anxiety. Nine- to 11-day-old rat pups were separated from their litter and the effects of intraperitoneally administered test compounds on the elicited ultrasonic vocalizations were measured. Side-effect potential was assessed using a modified inclined plane test ('time on an inclined plane', or TIP), and using negative geotaxis. SIV was reduced by CP 154,526 at doses that did not affect TIP or negative geotaxis, a profile like that of the 5-HT(1A) partial agonist buspirone. The benzodiazepine anxiolytic, diazepam, decreased SIV but also produced significant side effects at one to three-fold higher doses. Additional pharmacological characterization of SIV demonstrated anxiolytic-like effects of the atypical antipsychotic, clozapine, but not the typical antipsychotic, haloperidol, and of the serotonin reuptake inhibitor, zimelidine, but not the norepinephrine reuptake inhibitor, desipramine. In summary, the data support the conclusion that selective CRF(1) receptor antagonists may have utility in anxiety and stress disorders. The data further support the use of separation-induced vocalizations for identifying mechanistically diverse compounds with anxiolytic actions in man.

The allele-specific suppressor sup-39 alters use of cryptic splice sites in Caenorhabditis elegans.

Mutations in the Caenorhabditis elegans sup-39 gene cause allele-specific suppression of the uncoordination defect of unc-73(e936). e936 is a point mutation that changes the canonical G at the 5' end of intron 16 to a U. This mutation activates three splice donors, two of which define introns beginning with the canonical GU. Use of these two cryptic splice sites causes loss of reading frame; interestingly these messages are not substrates for nonsense-mediated decay. The third splice donor, used in 10% of steady-state e936 messages, is the mutated splice donor at the wild-type position, which defines an intron beginning with UU. In the presence of a sup-39 mutation, these same three splice donors are used, but the ratio of messages produced by splicing at these sites changes. The percentage of unc-73(e936) messages containing the wild-type splice junction is increased to 33% with a corresponding increase in the level of UNC-73 protein. This sup-39-induced change was also observed when the e936 mutant intron region was inserted into a heterologous splicing reporter construct transfected into worms. Experiments with splicing reporter constructs showed that the degree of 5' splice site match to the splicing consensus sequence can strongly influence cryptic splice site choice. We propose that mutant SUP-39 is a new type of informational suppressor that alters the use of weak splice donors.

A novel acidophilic RNA motif that recognizes coenzyme A.

Specific recognition of nucleotide cofactors by RNA may be important in engineering new RNA enzymes (ribozymes). Although in vitro selections (SELEX) have identified nucleic acid motifs ("aptamers") that bind a variety of adenosine cofactors, none of these recognizes coenzyme A (CoA), the primary biological cofactor used in acyltransfer reactions. We used SELEX experiments with two random RNA pools to identify aptamers that bind CoA. Functional boundary determination and extensive comparative sequence analysis (including reselection of a mutagenized, circularly permuted RNA) led to the identification of a 52 nucleotide minimal aptamer ("min52"). The RNA structural motif contains a large internal loop with 26 unpaired nucleotides flanked by helices of any base-paired sequence. Twenty loop nucleotides are specifically required for binding activity, 12 of which are derived from the original primer binding sequences. Specificity studies with CoA analogues demonstrated that the aptamer recognizes many adenosine analogues, including ATP, and that recognition is predominantly through the Höogsteen face of adenine. Binding activity is greatest at acidic pH (optimum near 5.0), in low or no monovalent salt, and at high concentrations of either Mg2+ or Mn2+. Strong binding activity (86% of maximum) is observed at pH 4.0, suggesting that at least some extreme conditions (acidic pH) may be compatible with RNA World theories of the origin and early evolution of life. In the presence of 10 mM Mg2+, binding is unaffected by the addition of 1 mM Ca2+, but it is mildly inhibited by 1 mM Zn2+ or Co2+ or by 0.1 mM Cu2+ or Ni2+. The dissociation constant (Kd) for the association of min52 RNA with ATP in solution was measured to be 2.4 +/- 0.4 microM under the conditions of the selection and 0.5 +/- 0.1 microM under optimized conditions. Finally, we show that the selected CoA aptamer populations contain other RNAs at low frequencies that preferentially recognize intact CoA and are not eluted from the resin by AMP alone.

Phylogenetic relationships among hypotrichous ciliates determined with the macronuclear gene encoding the large, catalytic subunit of DNA polymerase alpha.

The complete macronuclear DNA polymerase alpha gene, previously sequenced in Oxytricha nova, has been cloned from a genomic macronuclear library and sequenced for the hypotrich O. trifallax. Macronuclear DNA clones of DNA polymerase alpha encoding approximately 1000 amino acids, or approximately two-thirds of the open reading frame, have been obtained by PCR and sequenced for Halteria grandinella, Holosticha species, Paraurostyla viridis, Pleurotricha lanceolata, Stylonychia lemnae Teller, Sty. mytilus, Uroleptus gallina, and Urostyla grandis. Phylogenetic relationships inferred from DNA polymerase alpha amino acid sequences have been used to clarify taxonomic relationships previously determined by morphology of the cell cortex. Hypotrich phylogenies based on DNA polymerase alpha amino acid sequences are incongruent with morphological and other molecular phylogenies. Based upon these data, we assert that, contrary to morphological data, O. nova and O. trifallax are different species, and we propose that the oligotrich Halteria grandinella be reclassified as a hypotrich. This work also extends the available data base of eukaryotic DNA polymerase alpha sequences, and suggests new amino acid sequence targets for mutagenesis experiments to continue the functional dissection of DNA pol alpha biochemistry at the molecular level.

Evolution of internal eliminated segments and scrambling in the micronuclear gene encoding DNA polymerase alpha in two Oxytricha species.

To learn about the evolution of internal eliminated segments (IESs) and gene scrambling in hypotrichous ciliates we determined the structure of the micronuclear (germline) gene encoding DNA polymerasealpha(DNA polalpha) in Oxytricha trifallax and compared it to the previously published structure of the germline DNA polalphagene in Oxytricha nova . The DNA polalphagene of O.trifallax contains 51 macronuclear-destined segments (MDSs) separated by 50 IESs, compared to 45 MDSs and 44 IESs in the O.nova gene. This means that IESs and MDSs have been gained and/or lost during evolutionary divergence of the two species. Most of the MDSs are highly scrambled in a similar non-random pattern in the two species. We present a model to explain how IESs, non-scrambled MDSs and scrambled MDSs may be added and/or eliminated during evolution. Corresponding IESs in the two species differ totally in sequence, and junctions between MDSs and IESs are shifted by 1-18 bp in O.trifallax compared to the O.nova gene. In both species a short region of the gene is distantly separated from the main part of the gene. Comparison of the gene in the two species shows that IESs and scrambling are highly malleable over evolutionary time.

RNA aptamers to the peptidyl transferase inhibitor chloramphenicol.

BACKGROUND: The problem of how macromolecules adopt specific shapes to recognize small molecules in their environment is readily addressed through in vitro selections (the SELEX protocol). RNA-antibiotic interactions are particularly attractive systems for study because they provide an opportunity to expand our understanding of molecular recognition by RNA and to facilitate ribosomal modeling. Specifically, the antibiotic chloramphenicol (Cam) naturally binds bacterial ribosomes in the 'peptidyl transferase loop' of 23S ribosomal RNA to inhibit peptide bond formation. RESULTS: We identified Cam-binding RNA molecules ('aptamers') from two independent initial random RNA populations. Boundary determinations, ribonuclease S1 sensitivity analyses and the activity of truncated minimal RNAs identified a structural motif that is shared by sequences from both selections. The pseudosymmetric motif consists of a highly conserved central helix of five to six base pairs flanked by A-rich bulges and additional helices. Addition of Cam prior to ribonuclease S1 protected nucleotides in the conserved cores from cleavage. Reselection from a pool of mutated variants of the minimal aptamer further refined the sequence requirements for binding. Finally, we used proton nuclear magnetic resonance (NMR) to establish a 1:1 RNA: Cam stoichiometry of the complex. Both the protection and NMR data both show that Cam stabilizes the active fold of this aptamer. CONCLUSIONS: There are many different RNA sequences that can bind Cam. The Cam aptamers that we examined have a well-defined secondary structure with a binding pocket that appears to be stabilized by Cam. This RNA motif superficially resembles the Cam-binding site in 23S rRNA, although further work is needed to establish the significance of these similarities.

The germline gene encoding DNA polymerase alpha in the hypotrichous ciliate Oxytricha nova is extremely scrambled.

We report the structure of the micronuclear (germline) gene encoding the large catalytic subunit of DNA polymerase alpha (DNA pol alpha) in the ciliate Oxytricha nova. It contains 44 internal eliminated segments (IESs) that divide the gene into 45 macronuclear-destined segments (MDSs) that are in a non-randomly scrambled order with an inversion near the gene center. Odd numbered MDSs 29-43, containing 230 bp out of a total of 4938 bp of macronuclear sequence, are missing from the 14 kb cloned gene. The missing MDSs have not been located but are at least several kilobases from the main body of the gene. The remarkably scrambled DNA pol alpha gene must be extensively cut, re-ordered and spliced and an inversion must occur to produce an unscrambled, functional version of the gene during development of a new macronucleus. Unscrambling is hypothesized to occur by a homologous recombination mechanism guided by repeat sequences at MDS ends.

Catalepsy as a rodent model for detecting antipsychotic drugs with extrapyramidal side effect liability.

The predictive validity of catalepsy as a rodent model for detecting the extrapyramidal side effects (EPS) of antipsychotic drugs was recently questioned when the novel antipsychotic savoxepine produced little catalepsy in rodents while producing significant EPS in schizophrenic patients. Because catalepsy is viewed as an important model for predicting EPS, we decided to re-evaluate the effects of savoxepine. Savoxepine, clozapine, haloperidol, olanzapine, ORG 5222, raclopride, and risperidone were examined in two tests for catalepsy (grid and bar tests) in male Sprague-Dawley rats. The ability to antagonize amphetamine-induced hypermotility was also examined, since this measure is believed to predict clinical efficacy. With the exception of clozapine, all drugs produced dose-dependent catalepsy in both tests. For each drug, the minimum effective dose for producing catalepsy was greater than or equal to the ED50 for antagonizing amphetamine-induced hyperactivity (defined as the dose producing a 50% reduction in hyperactivity). Clozapine resulted in the widest separation of effective doses in the catalepsy and activity models. Raclopride produced the next largest separation while the remaining drugs resulted in only a one- or two-fold dose separation between the two behavioral tests. The results with haloperidol and clozapine are consistent with the clinical effects of these drugs (severe versus mild EPS). The ratios of effective doses in catalepsy and activity for the remaining novel drugs are also consistent with preliminary clinical findings indicating some EPS with each of these compounds. Thus, catalepsy remains a suitable rodent model for detecting compounds with EPS liability in humans.

Macronuclear gene-sized molecules of hypotrichs.

The macronuclear genome of hypotrichous ciliates consists of DNA molecules of gene-sized length. A macronuclear DNA molecule contains a single coding region. We have analyzed the many hypotrich macronuclear DNA sequences sequenced by us and others. No highly conserved promoter sequences nor replication initiation sequences have been identified in the 5' nor in the 3' non-translated regions, suggesting that promoter function in hypotrichs may differ from other eukaryotes. The macronuclear genes are intron-poor; approximately 19% of the genes sequenced to date have one to three introns. Not all macronuclear DNA molecules may be transcribed; some macronuclear molecules may not have any coding function. Codon bias in hypotrichs is different in many respects from other ciliates and from other eukaryotes.

A 10-point strategic checklist for rural health care systems.

A checklist format is used to provide a framework for rural hospital executives and community members for gauging the health and stability of rural hospitals and rural hospital systems. Benchmarks are provided for financial and operational performance and emphasis is placed on medical staff size and physician recruitment. Physician/hospital organizations and regional partnerships are used as examples of strategies available to rural providers. The importance of market knowledge and regional strategic alliances also is stressed. In an era of dwindling resources and tight reimbursement, rural providers are encouraged to consider cooperative clinical programming and technology consolidation.

D1 and D2 dopamine receptor antagonists reverse prepulse inhibition deficits in an animal model of schizophrenia.

The amplitude of the acoustic startle response is decreased if the startle stimulus is preceded by a nonstartle eliciting stimulus. This sensorimotor gating phenomenon, known as prepulse inhibition, is diminished in schizophrenic individuals. In rats, the dopamine agonist apomorphine disrupts prepulse inhibition and this disruption is reversed by classical and atypical antipsychotics. Furthermore, the ability of antipsychotics to reverse the apomorphine disruption is correlated with clinical potency and D2 receptor affinity. In the present study, the role of the D1 receptor in prepulse inhibition of the acoustic startle response was studied; the effects of the D1 receptor antagonist SCH 23390 were examined and compared to the effects of the D2 receptor antagonist eticlopride. Male Sprague-Dawley rats were placed into a startle chamber and presented with auditory stimuli consisting of either 95 or 105 dB noise bursts presented alone or preceded by a 75 dB noise burst. Trials consisting of no stimulus and the 75 dB prepulse stimulus alone were also included. These six trial types (ten each) were randomly presented within a 35-min session. Rats treated with 2.0 mg/kg apomorphine (SC) demonstrated a significant disruption of prepulse inhibition compared to vehicle controls. Pretreatment with the D1 antagonist SCH 23390 (0.01, 0.05, 0.1 mg/kg SC) or the D2 antagonist eticlopride (0.01, 0.05, 0.1 mg/kg SC) attenuated the disruptive effects of apomorphine. These results indicate that selective blockade of either the D1 or D2 receptor subtype is sufficient in reversing the sensorimotor gating deficits produced by apomorphine.(ABSTRACT TRUNCATED AT 250 WORDS)

A gene-sized DNA molecule encoding the catalytic subunit of DNA polymerase alpha in the macronucleus of Oxytricha nova.

We have isolated a gene-sized molecule encoding the catalytic subunit of DNA polymerase alpha from a macronuclear genomic library of Oxytricha nova, by using a 0.7-kb fragment of the corresponding human gene as a hybridization probe. Two different versions of the gene are present in the macronucleus, one with an EcoRI site (RI+) and one without an EcoRI site (RI-). The cloned RI- version has been characterized. It is 4938 bp in length, excluding telomeres. It consists of a 329-bp 5' leader, a 4479-bp coding region and a 130-bp 3' trailer. The deduced amino-acid sequence shares conserved regions with the yeast and human polypeptides. We also demonstrate by Southern analysis that gene-sized molecules of similar size, homologous to the isolated O. nova gene are present in the mac genome of closely and distantly related hypotrichs.

The noncompetitive NMDA antagonist MK-801 fails to block amphetamine-induced place conditioning in rats.

The noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 prevents the development of sensitization to the locomotor-activating effects of amphetamine. In the present study, the possibility that the NMDA receptor might also play a role in the rewarding effects of amphetamine (as measured in the conditioned place preference paradigm) was investigated. Male Sprague-Dawley rats received amphetamine (2.0 mg/kg IP) paired with one side of a two-compartment box and saline paired with the other side. During these pairings locomotor activity was measured. On the test day, the amount of time drug-free rats spent in each compartment was determined. Rats trained with amphetamine alone showed a significant increase in time spent on the drug-paired side from pre- to postconditioning, indicating a place preference. When rats were injected with MK-801 (0.03, 0.1, or 0.3 mg/kg SC) prior to amphetamine, no significant effects on amphetamine place conditioning were observed. Rats treated with MK-801 alone showed significant place conditioning, but only at the intermediate dose. On conditioning days, MK-801 produced a dose-dependent enhancement of amphetamine-induced locomotor activity; however, MK-801 alone caused a similar increase in activity. The preferential D2 dopamine receptor antagonist eticlopride (0.01, 0.05, or 0.1 mg/kg SC) significantly reduced amphetamine locomotor activity, and the highest dose blocked place conditioning. These data suggest that the NMDA receptor is not involved in either the rewarding or locomotor-activating effects of amphetamine.

The effect of arthroscopic surgery on mandibular range of motion.

Limited range of motion of the mandible secondary to intra-articular problems has become an accepted indication for temporomandibular joint (TMJ) arthroscopy. The authors have looked at 297 patients over a 2 1/2 year follow-up period. The patients were subdivided by their initial opening measurement (10-20 mm or 20-30 mm). The post-operative improvement of these patients was then evaluated. Furthermore, multiple variables were used to look at the effect of surgical procedures, diagnosis and other associated clinical findings on outcome. In conclusion, the authors found that the range of motion significantly improved to a close to normal opening in almost all patient groupings. This finding was unrelated to a variety of variables and supports the hypothesis that TMJ arthroscopic surgery has a definite role in treating restricted range of motion secondary to intra-articular pathology.

Lack of cross-sensitization between the locomotor-activating effects of bromocriptine and those of cocaine or heroin.

Rats were given daily injections of bromocriptine (5.0 mg/kg IP) or vehicle either in the home cage or in a test box equipped with photocells to measure locomotion. The animals were then tested in the photocell boxes for their locomotor response to cocaine (10.0 mg/kg IP), heroin (0.5 mg/kg IP), or quinpirole (0.1 mg/kg IP). Repeated bromocriptine in the test box but not in the home cage caused progressive increases in sensitivity to the locomotor-stimulating effects of bromocriptine and increases in the subsequent sensitivity to quinpirole but caused only trivial signs of cross-sensitization to cocaine or heroin. Cross-sensitization to quinpirole was temporary; responsiveness to quinpirole decreased with further quinpirole injections. Lack of significant cross-sensitization between bromocriptine and either cocaine or heroin and lack of permanence of the cross-sensitization between bromocriptine and quinpirole raise questions as to the biological basis of psychomotor stimulant sensitization.