Depletion of brainstem epinephrine stores by alpha-methyldopa: possible relation to attenuated sympathetic outflow

The antihypertensive effect of alpha-methyldopa (MD) is believed to be critically dependent on its ability to deplete endogenous catecholamines or cause the synthesis of false neurotransmitters. We used liquid chromatography with electrochemical detection (LCEC) and negative chemical ionization gas chromatography-mass spectrometry (GC-MS) for quantitation of catecholamines and MD metabolites in rat. MD intraperitoneally (100 mg/kg q12 hr X 12 days), significantly increased alpha-methylnorepinephrine (MNE) in brain (1.02 +/- 0.33 micrograms/g), heart (1.67 +/- 0.57 micrograms/g) and adrenal glands (114.93 +/- 50.47 micrograms/g) Endogenous norepinephrine (NE), epinephrine (E) and dopamine (DA) were reduced. ME levels were 2.19 +/- 0.44 micrograms/g (n = 6) in the adrenal gland but only 99 +/- 26 pg/g (n = 3) in the brainstem. MD-induced endogenous brainstem NE depletion was more than compensated by MNE production, but brainstem E depletion was not compensated for by a stoichiometric production of brainstem ME. We conclude (1) although ME is a metabolite of MD, it is present in extremely low concentrations in brainstem and (2) central epinephrine-containing neurons are depleted of neurotransmitter by MD therapy. If this selective epinephrine depletion occurs in the bulbospinal tract neurons responsible for maintaining sympathetic tone, then this effect could contribute to the antihypertensive effect of MD.     

Chorusing Behaviour,a Density-dependent Alternative Mating Strategy in Male Common Toads (Bufo bufo)

Male chorusing behaviour was studied in a population of common toads (Bufo bufo) on the island of Öland south Sweden, and the functional role of male advertisement calling in this species was experimentally examined. Calling males were larger and heavier than non-calling males (t = 2.41, p < 0.025 and t = 2.36, p < 0.025, respectively). However, small males were also found to call. This indicates that large males persisted in calling for longer and/or called more often. The proportion of calling males decreased as population size increased during the breeding season, indicating that calling is a low density strategy. Females responded more readily to calls than males. There were insufficient data to determine if the dominant frequencies of advertisement calls were inversely correlated with male body size, however, this relationship was found for the similar release calls. Females were found not to discriminate between high and low frequency calls, but when given a choice between two calls of different sound pressure levels (SPL), females were attracted to the louder calls. Thus, the function of chorusing is to advertise the position of males to mate-seeking females when the population density is low.     

Survival of Cryptosporidium parvum oocysts under various environmental pressures.

The survival of various isolates of Cryptosporidium parvum oocysts under a range of environmental pressures including freezing, desiccation, and water treatment processes and in physical environments commonly associated with oocysts such as feces and various water types was monitored. Oocyst viability was assessed by in vitro excystation and by a viability assay based on the exclusion or inclusion of two fluorogenic vital dyes. Although desiccation was found to be lethal, a small proportion of oocysts were able to withstand exposure to temperatures as low as -22 degrees C. The water treatment processes investigated did not affect the survival of oocysts when pH was corrected. However, contact with lime, ferric sulfate, or alum had a significant impact on oocyst survival if the pH was not corrected. Oocysts demonstrated longevity in all water types investigated, including seawater, and when in contact with feces were considered to develop an enhanced impermeability to small molecules which might increase the robustness of the oocysts when exposed to environmental pressures.     

Isolation and Characterization of a Cold-Active Xylanase Enzyme from Flavobacterium sp.

Xylan is the major component of hemicellulose, and xylan should be fully utilized to improve the efficiencies of a biobased economy. There are a variety of industrial reaction conditions in which an active xylanase enzyme would be desired. As a result, xylanase enzymes with different activity profiles are of great interest. We isolated a xylanase gene (xyn10) from a Flavobacterium sp. whose sequence suggests that it is a glycosyl hydrolase family 10 member. The enzyme has a temperature optimum of 30°C, is active at cold temperatures, and is thermolabile. The enzyme has an apparent K_m of 1.8 mg/ml and k_cat of 100 sec⁻¹ for beechwood xylan, attacks highly branched native xylan substrates, and does not have activity against glucans.     

Recent trends in the intrinsic water-use efficiency of ringless rainforest trees in Borneo

Stable carbon isotope (δ(13)C) series were developed from analysis of sequential radial wood increments from AD 1850 to AD 2009 for four mature primary rainforest trees from the Danum and Imbak areas of Sabah, Malaysia. The aseasonal equatorial climate meant that conventional dendrochronology was not possible as the tree species investigated do not exhibit clear annual rings or dateable growth bands. Chronology was established using radiocarbon dating to model age-growth relationships and date the carbon isotopic series from which the intrinsic water-use efficiency (IWUE) was calculated. The two Eusideroxylon zwageri trees from Imbak yielded ages of their pith/central wood (±1 sigma) of 670 ± 40 and 759 ± 40 years old; the less dense Shorea johorensis and Shorea superba trees at Danum yielded ages of 240 ± 40 and 330 ± 40 years, respectively. All trees studied exhibit an increase in the IWUE since AD 1960. This reflects, in part, a response of the forest to increasing atmospheric carbon dioxide concentration. Unlike studies of some northern European trees, no clear plateau in this response was observed. A change in the IWUE implies an associated modification of the local carbon and/or hydrological cycles. To resolve these uncertainties, a shift in emphasis away from high-resolution studies towards long, well-replicated time series is proposed to develop the environmental data essential for model evaluation. Identification of old (greater than 700 years) ringless trees demonstrates their potential in assessing the impacts of climatic and atmospheric change. It also shows the scientific and applied value of a conservation policy that ensures the survival of primary forest containing particularly old trees (as in Imbak Canyon and Danum).     

Estimated feeding rates and energy requirements of gentoo penguins,Pygoscelis papua,at Macquarie Island

Summary Water and sodium turnovers of 6–7 week old gentoo penguin chicks and breeding adults were measured using isotopically labelled water and sodium. Influx rates for chicks averaged 188 ml·kg^-1·day^-1 and 13.9 mmol·kg^-1·day^-1 for water and sodium, respectively. Chicks consumed an estimated 228 g·kg^-1·day^-1 fresh food or 886 kJ kg^-1 day. These values correspond to 761 g·day^-1 or 2945 kJ·day^-1 for a gentoo chick mid-way through the growth period. Flux rates for adults attending chicks ranged from 199 to 428 ml·kg^-1·day^-1 for water and from 15 to 36 mmol·kg^-1·ay^-1 for sodium.     

Long-pore Electrostatics in Inward-rectifier Potassium Channels

Inward-rectifier potassium (Kir) channels differ from the canonical K⁺ channel structure in that they possess a long extended pore (~85 Å) for ion conduction that reaches deeply into the cytoplasm. This unique structural feature is presumably involved in regulating functional properties specific to Kir channels, such as conductance, rectification block, and ligand-dependent gating. To elucidate the underpinnings of these functional roles, we examine the electrostatics of an ion along this extended pore. Homology models are constructed based on the open-state model of KirBac1.1 for four mammalian Kir channels: Kir1.1/ROMK, Kir2.1/IRK, Kir3.1/GIRK, and Kir6.2/KATP. By solving the Poisson-Boltzmann equation, the electrostatic free energy of a K⁺ ion is determined along each pore, revealing that mammalian Kir channels provide a favorable environment for cations and suggesting the existence of high-density regions in the cytoplasmic domain and cavity. The contribution from the reaction field (the self-energy arising from the dielectric polarization induced by the ion's charge in the complex geometry of the pore) is unfavorable inside the long pore. However, this is well compensated by the electrostatic interaction with the static field arising from the protein charges and shielded by the dielectric surrounding. Decomposition of the static field provides a list of residues that display remarkable correspondence with existing mutagenesis data identifying amino acids that affect conduction and rectification. Many of these residues demonstrate interactions with the ion over long distances, up to 40 Å, suggesting that mutations potentially affect ion or blocker energetics over the entire pore. These results provide a foundation for understanding ion interactions in Kir channels and extend to the study of ion permeation, block, and gating in long, cation-specific pores.     

Cloning and characterization of two cellulase genes from Lentinula edodes

The effect of overproduction of the Hsp70 system proteins (DnaK, DnaJ, GrpE) and/or ClpB (Hsp100) from plasmids on the process of formation and removal of heat-aggregated proteins from Escherichia coli cells (the S fraction) was investigated by sucrose density gradient centrifugation. Two plasmids were employed: pKJE7 carrying the dnaK/dnaJ/grpE genes under the control of the araB promoter and pClpB carrying the clpB gene under the control of its own promoter (sigma(32)-dependent). In the wild-type cells the S fraction after 15 min of heat shock amounted to 21% of cellular insoluble proteins (IP), and disappeared 10 min after transfer of the culture to 37 degrees C. In contrast to this, in the clpB mutant the S fraction was larger (35% IP) and its elimination was retarded, nearly 60% of the aggregated proteins remained stable 30 min after heat shock. This result points to the importance of ClpB in removal of the heat-aggregated proteins from cells. Overproduction of the Hsp70 system proteins (exceeding by about 1.5-fold that of wild-type) in wild-type and DeltaclpB cells completely prevented the formation of the S fraction during heat shock. Overproduction of ClpB (exceeding by about eight-fold that of wild-type) in the same background did not prevent protein aggregation after heat shock and only partly compensated for the effect of the mutation in the clpB gene. Monitoring the S fraction during co-production of DnaK/DnaJ/GrpE and ClpB in the DeltaclpB mutant revealed that both the levels of expression and the ratios of ClpB to Hsp70 system proteins had a significant effect on the formation and removal of protein aggregates in heat-shocked E. coli cells. In the presence of excess ClpB, an increase in the levels of DnaK, DnaJ and GrpE was required to prevent aggregate formation upon heat shock or to efficiently remove protein aggregates after heat shock. Therefore, it is supposed that a high level of ClpB under some conditions, especially at insufficient levels of Hsp70 system proteins, may support protein aggregation resulting from heat shock and may lead to stabilization of hydrophobic aggregates.