bluuebunny/crossref_metadata_2025 huggingface dataset:
Nutrient transfer in arbuscular mycorrhizas: how are fungal and plant processes integrated?
Authors: Smith Sally E. , Dickson Sandy , Smith F. Andrew
Published: September 2002

This review brings together recent work on the coordination of transport
processes between fungus and plant symbionts in arbuscular mycorrhizal (AM)
symbioses, and focuses on new information on the diversity in structure and
function of interfaces and their potential roles in transport processes. We
consider the way that fungal activity is polarised to absorb mineral nutrients
(especially phosphorus, P) in soil, transport them to the root and release
them to the plant. Conversely, the fungal structures within the root appear to
be specialised to absorb sugars, which the external mycelium cannot do. The
external mycelium depends on a supply of lipid, transported from within the
root. High affinity P transporters expressed in the root apices and root hairs
of non-mycorrhizal roots, and most probably mycorrhizal roots, absorb P
actively. This can result in the development of P depletion zones, so that a
low concentration of P at the absorbing surfaces limits further uptake. The
external hyphae of AM fungi extend well beyond the depletion zone, accessing
supplies of P at a distance and in narrow soil pores, that is absorbed
actively by a high affinity P transporter expressed in these small diameter
hyphae. Translocation of P within the hyphae and transfer to the plant results
in much higher rates of uptake (inflows) by mycorrhizal than non-mycorrhizal
roots. The possible role of polyphosphate (polyP) in this process is discussed
in the light of new data. Within the root, P is lost from the fungal
structures to the interfacial apoplast by an unknown mechanism, and is
absorbed by the root cortical cells. The expression of a high affinity P
transporter and H + -ATPase in arbuscule-containing
cells indicates that these are probably the sites of fungus/plant P
transfer. The site of sugar transfer from plant to fungus has not yet been
established. At the whole plant level, plant uptake systems located in the
youngest regions of the root are positioned to absorb P from undepleted soil,
into which the root apex has just grown. In older regions of the roots,
colonised by mycorrhizal fungi, the external mycelium will take over the
absorptive role and overcome the difficulties posed by the slow diffusion of P
in soil.
Sequence analysis and expression patterns of two nifA genes from Frankia EuIK1
Authors: Lee Hyoungseok , Sung Si Bum , Kim Ho Bang , An Chung Sun
Published: September 2002

This paper originates from an address at the 8th International Symposium on Nitrogen Fixation with Non-Legumes, Sydney, NSW, December 2000
Two nifA genes were cloned from
Frankia EuIK1 strain, a symbiont of
Elaeagnus umbellata Thunb. and analysed on the basis of
their deduced amino acid sequences and expression patterns. The complete
nucleotide sequence of 1926 bp of nifA1 and 1524 bp of
nifA2 was determined, respectively. A putative
NifA-binding site was found –95 to about –80 bp upstream of start
codon for nifA1 ORF as
TGT-N10-ACA, but the clpB ORF was
followed by nifA2 ORF with 15 bp of intergenic sequence.
Deduced amino acid sequence showed that two nifA genes
encode typical NifA, having three major domains and two linkers, and their
central domains of NifA1 and NifA2 showed sequence similarity of
70–75% with those from other NifA proteins. However, entire NifA2
ORF is more similar to alternative NifA (60%) than to typical NifA
(53%). Conserved amino acid sequence in helix-turn-helix motif of
typical NifA was also found in NifA1, but it was not conserved in NifA2, which
is also common in alternative NifA proteins. Moreover, the expression of
nifA1 during nodule development was similar to that of
Rhizobium meliloti in that it was expressed at low level
constitutively, while that of nifA2 was similar to the
pattern of nifH, structural gene for nitrogenase
reductase, in that its transcripts level was changed in accordance with
nitrogenase activity. These results indicate that nifA1
and nifA2 might be classified into typical
nifA and alternative nifA,
respectively. This is the first report on the presence of two
nifA genes in Frankia.
Does the routine heat treatment of sugarcane stem pieces for xylem pathogen control affect the nitrogenase activity of an N2-fixing endophyte in the cane?
Authors: Ortega Eduardo , Rodés Rosa , de la Fuente Enrique , Fernández Loiret
Published: September 2002

This paper originates from an address at the 8th International Symposium on Nitrogen Fixation with Non-Legumes, Sydney, NSW, December 2000
In sugarcane propagation the sett pieces are routinely heated at 50˚C
before planting to control the xylem pathogen
Leifsonia xyli ssp. xyli. To
determine whether this treatment also affects the diazotrophic endophyte,
Gluconacetobacter diazotrophicus, found in the
intercellular solution of cane parenchyma, cultures of this bacterium were
similarly heated. The nitrogenase activity of the heated cultures was
monitored by measurement of hydrogen evolution. Activity was destroyed in
cultures heated directly at 45 or 50˚C for 2 h. In contrast, when sett
pieces were first heated for 2 h at 50˚C,
G. diazotrophicus could still be isolated from the
intercellular fluid, and these cultures showed considerable nitrogenase
activity. There was no difference in leaf nitrogen content of plants grown in
nitrogen-poor soil from setts either heated or unheated. It is concluded that
the routine heat treatment of sugarcane setts for pathogen control does not
totally destroy the nitrogenase activity of the diazotrophic endophyte
G. diazotrophicus.
Natural abundance of stable carbon and nitrogen isotopes in Cannabis sativa reflects growth conditions
Authors: Denton Tricia M. , Schmidt Susanne , Critchley Christa , Stewart George R.
Published: September 2002

Stable carbon and nitrogen isotope signatures
(δ13C and δ15N) of
Cannabis sativa were assessed for their usefulness to
trace seized Cannabis leaves to the country of origin
and to source crops by determining how isotope signatures relate to plant
growth conditions. The isotopic composition of Cannabis
examined here covered nearly the entire range of values reported for
terrestrial C3 plants. The
δ13C values of Cannabis from
Australia, Papua New Guinea and Thailand ranged from –36 to
–25‰, and δ 15N values ranged from
–1.0 to 15.8‰. The stable isotope content did not allow
differentiation between Cannabis originating from the
three countries, but δ13C values of
plantation-grown Cannabis differed between well-watered
plants (average δ13C of–30.0‰) and
plants that had received little irrigation (average
δ13C of –26.4‰).
Cannabis grown under controlled conditions had
δ13C values of –32.6 and –30.6‰
with high and low water supply, respectively. These results indicate that
water availability determines leaf 13C in plants grown
under similar conditions of light, temperature and air humidity. The
δ13C values also distinguished between indoor- and
outdoor-grown Cannabis; indoor-grown plants had overall
more negative δ13C values (average
–31.8‰) than outdoor-grown plants (average –27.9‰).
Contributing to the strong 13C-depletion of indoor-grown
plants may be high relative humidity, poor ventilation and recycling of
13C-depleted respired CO2. Mineral
fertilizers had mostly lower δ15N values
(–0.2 to 2.2‰) than manure-based fertilizers (7.6 to
22.7‰). It was possible to link δ15N values
of fertilizers associated with a crop site to soil and plant
δ15N values. The strong relationship between soil,
fertilizer, and plant δ15N suggests that
Cannabis δ15N is determined
by the isotopic composition of the nitrogen source. The distinct d15N values
measured in Cannabis crops make
δ15N an excellent tool for matching seized
Cannabiswith a source crop. A case study is presented
that demonstrates how δ13C and
δ15N values can be used as a forensic tool.
Changes in physiological and morphological traits of roots and shoots of wheat in response to different depths of waterlogging
Authors: Malik Al Imran , Colmer Timothy D. , Lambers Hans , Schortemeyer Marcus
Published: September 2002

The growth reduction of wheat (Triticum aestivum L.)
during and after waterlogging stress depends on the depth of water from the
soil surface. In a pot experiment with 3-week-old plants, soil was waterlogged
for 14 d at the surface, or at 100 or 200 mm below the surface, and pots were
then drained to assess recovery. A fully drained treatment kept at field
capacity served as control. During waterlogging, the relative growth rate of
roots decreased more than that of shoots (by 6–27% for shoots, by
15–74% for roots), and plant growth was reduced proportionally as
the water level was increased. Light-saturated net photosynthesis was reduced
by 70–80% for the two most severe waterlogging treatments, but
was little affected for plants in soil waterlogged at 200 mm below the
surface. The number of adventitious roots formed per stem in plants grown in
waterlogged soil increased up to 1.5 times, but the number of tillers per
plant was reduced by 24–62%. The adventitious roots only
penetrated 85–116 mm below the water level in all waterlogging
treatments. Adventitious root porosity was enhanced up to 10-fold for plants
grown in waterlogged soil, depending on water level and position along the
roots. Porosity also increased in basal zones of roots above the water level
when the younger tissues had penetrated the waterlogged zone. Fourteen days
after draining the pots, growth rates of plants where the soil had been
waterlogged at 200 mm below the surface had recovered, while those of plants
in the more severely waterlogged treatments had only partially recovered.
These findings show that the depth of waterlogging has a large impact on the
response of wheat both during and after a waterlogging event so that
assessment of recovery is essential in evaluating waterlogging tolerance in
crops.
Respiration in the light measured by 12CO 2 emission in 13CO 2 atmosphere in maize leaves
Authors: Loreto Francesco , Velikova Violeta , Di Marco Giorgio
Published: September 2002

The mitochondrial respiration during photosynthesis is difficult to measure
and is indirectly estimated mainly in C 3 plants. Loreto
et al. [(1999)
Australian Journal of Plant Physiology
26, 733–736] have shown that the emission of
12 CO 2 from illuminated leaves
exposed to air containing 13 CO 2
measures photorespiration and mitochondrial respiration in C 3 leaves. This
method was used to measure the mitochondrial respiration in illuminated maize
leaves. The 12 CO 2 emission was
steady after 30 s, a time sufficient to label the CO 2
leakage from bundle sheath cells with 13 CO
2 , but not the mitochondrial respiration in the light.
The emission was low (0.1–0.4 ppm or 0.2–0.4 µmol m –2
s –1 ) in a wide range of leaf temperatures and light intensities, but
increased at light intensities below 200 µmol m –2 s –1 and
at temperatures above 42°C. At 120 s after labelling, the leaf was
darkened and the emission rapidly matched the mitochondrial respiration
measured by gas exchange. The emission of 12 CO
2 in the light was inversely correlated with
photosynthesis. This suggested that most of the respiratory CO
2 was refixed by photosynthesis. The amount of refixed
intercellular 12 CO 2 was
calculated from gas-exchange parameters. It was 60 to 90% of the tota
l12 CO 2 in leaves illuminated and
exposed to temperatures below 42°C. In leaves with reduced
photosynthesis because of exposure to higher temperatures or low light, the
12 CO 2 refixation decreased. The
sum of refixed and emitted 12 CO 2
was close to the mitochondrial respiration in the dark. This suggested that in
these leaves the mitochondrial respiration was not inhibited in the light. In
salt- and water-stressed leaves, however, the sum of refixed and emitted
12 CO 2 was lower than
mitochondrial respiration in the dark, suggesting that the mitochondrial
respiration may be inhibited in the light.
Membrane transport: mechanisms and processes from organelles to whole plants
Authors: Smith F. Andrew
Published: September 2002

Inorganic phosphate (Pi) uptake systems across the plasma membrane of plant
cells have been extensively investigated. Physiological studies have
established that Pi is transported into plant cells via co-transport with
H+ , and in some plants with Na
+ , using the driving force provided by the
electrogenic H + pump in the plasma membrane.
Molecular studies have identified many genes for Pi transporters and are
providing insights into the mechanisms of genetic control of Pi transport.
There still remain, however, questions as to how Pi uptake systems are
regulated at the physiological level. We have found that Pi uptake induces
cytoplasmic acidification, and, conversely, that inducing cytoplasmic
acidification causes the cytoplasmic Pi concentration to decrease. Both of
these responses affect the operation of the H +
-pump. These phenomena are discussed in relation to a possible mechanism for
the physiological control of Pi uptake by plant cells.
Photon flux density dependence of carbon acquisition and demand in relation to shoot growth of kiwifruit (Actinidia deliciosa) vines grown in controlled environments
Authors: Greer Dennis H.
Published: September 2002

Kiwifruit [Actinidia deliciosa (A. Chev.) C.F.
Liang et A.R. Ferguson] vines were grown in four controlled photon
flux densities (PFDs) from 250 to 1100 µmol m
–2 s –1 for 130 d
starting from pre-budbreak to measure relationships between shoot growth and
carbon (C) demand and to assess the effect of PFD on these processes. Leaf
area, stem length, photosynthesis and respiration rates were measured on the
same leaves at regular intervals. From daily C acquisition and accumulation in
biomass, the net C balance per cane was determined throughout the experiment.
High-PFD-grown vines had 13% more leaf area, 250% more leaf
biomass and 30% more stem biomass than low-PFD-grown vines.
High-PFD-grown vines also partitioned relatively more biomass to
photosynthetic tissue than to supporting stem tissue compared with
low-PFD-grown vines. Rates of net photosynthesis were highest on vines grown
at 800 µmol m –2 s
–1 , but respiration rates were highest in
high-PFD-grown vines. Vines grown at 1100 µmol m
–2 s –1 had a net gain
of 119 g sh –1 and 53 g sh
– at 250 µmol m
–2 s –1 , of which 46
and 58%, respectively, was used in shoot biomass growth. Net C balance
was negative for 30 d after budbreak. Over 130 d, high-PFD-grown vines
produced a total surplus of 64 g sh –1 , while
low-PFD-grown vines produced 22 g sh –1 . Results
demonstrate that irradiance has no effects on developmental processes but has
marked effects on vegetative growth rates of kiwifruit vines. Underlining
this, the C economy of these shoots is highly and quantitatively dependent on
the PFD during growth.
Photoinactivation of photosystem II in high light-acclimated grapevines
Authors: Flexas Jaume , Hendrickson Luke , Chow Wah Soon
Published: September 2002

Grapevines are considered well adapted to high irradiance during growth. It is
still controversial, however, whether photoinactivation of photosystem II is
completely avoided in high light-acclimated grapevines growing in the field.
This study examines the functional stability of PSII in leaf discs (floated on
water) of field-grown, high light-acclimated grapevines as a function of
photon exposure. Measuring functional PSII units by flash-induced oxygen
evolution, it was found that the susceptibility of PSII to photoinactivation
was less in sun-exposed leaves than shade leaves of
Vitis riparia Michaux, and enhanced by lincomycin, an
inhibitor of chloroplast-encoded protein synthesis.
Vitis vinifera L. cv. Chardonnay, grown in a glasshouse
with slightly lower irradiance, exhibited an intermediate susceptibility.
Significantly, the dark-relaxed quantum efficiency of PSII, measured as
(Fm –
Fo)/Fm,
where Fm and
Fo are the chlorophyll (Chl)
fluorescence yields for closed and open reaction centres, respectively,
declined much more slowly than did the number of functional PSII units in
V. riparia. Thus, measurements of
(Fm –
Fo)/Fm
may give an impression of little photoinactivation of PSII, even when nearly
half of functional PSII units may be lost. By contrast, the parameter
1/Fo –
1/Fm is a more linear
indicator of functional PSII units. The results indicate that grapevines may
suffer photoinactivation of PSII, at least when leaf discs are floated on
water.
Agrobacterium tumefaciens-mediated transformation of wheat using suspension cells as a model system and green fluorescent protein as a visual marker
Authors: Weir Brian , Gu Xu , Wang Mingbo , Upadhyaya Narayana , Elliott Adrian R. , Brettell Richard I. S.
Published: September 2002

Conditions for Agrobacterium-mediated transformation of
wheat (Triticum aestivum L.) were defined using wheat
suspension cells as a model system and green fluorescent protein (GFP) as a
visual marker. Different strains of
Agrobacterium tumefaciens were compared using
established wheat cell suspension cultures, where the frequency of cell
clusters showing transient activity of GFP ranged from 2 to 52%. High
levels of transient GFP activity and stable transformed callus lines were
obtained with plasmid pTO134 containing a gfp gene with
an enhanced CaMV 35S promoter and a bar gene with a 35S
promoter in combination with Agrobacterium strain AGL0.
These results suggest that the important variables in
Agrobacterium-mediated transformation of wheat cells
include media composition, Agrobacterium strain, plasmid
vector and the addition of virulence-inducing agents such as acetosyringone.
The conditions deemed optimal for transformation of wheat suspension cell
lines were applied to scutella isolated from immature embryos and
scutella-derived calli. Transient GFP expression in these tissues ranged from
10 to 75% and, while quite variable among and within cultivars, stably
transformed scutellum-derived callus was obtained. Further studies with
scutellum-derived calli suggested that variables such as duration of
pre-inoculation culture and co-cultivation, as well as co-cultivation
temperature, were also important. Optimisation of these variables resulted in
the recovery of transformed wheat plants at a transformation frequency of
1.8%, which is comparable with other reports.
Gamete manipulation for development: new methods for conception
Authors: Yanagimachi R.
Published: September 2002

Mammalian oocytes microsurgically injected with spermatozoa can develop into
normal offspring. Apparently the oocyte has the ability to decompose or
eliminate such sperm components as the plasma membrane and acrosomal contents,
which normally do not enter its cytoplasm. Species in which normal offspring
were obtained by direct sperm injection include: human, mouse, rabbit, horse,
sheep, cattle, pig, and monkey. In the mouse, normal offspring can also be
obtained routinely by the injection of round spermatid nuclei into oocytes.
This suggests that all post-meiotic modifications of spermatozoa
(spermiogenesis, sperm maturation, capacitation and the acrosome reaction)
evolved as processes solely dedicated to delivering male genomes into female
gametes. Birth of normal off-spring after injection of spermatocytes into
maturing or mature oocytes suggests that the mechanisms controlling meiosis of
male and female germ cells are similar, if not the same. Spermatozoa do not
need to be morphologically normal or alive in the conventional sense to
participate in embryo development; as long as they have intact genomes, they
are able to produce normal offspring. Chromosomes within the first and second
polar bodies can be used as substitutes for female pronuclei for the
production of normal offspring. The nuclei of adult somatic cells can be used
for production of animals. This procedure, involving introduction of cell
nuclei into enucleated oocytes (genomic cloning), is rather inefficient at
present. Many obstacles must be overcome before it is accepted as a safe,
novel method to reproduce scientifically, medically or economically valuable
animals. For human cloning the prime consideration must be the welfare of the
child.
tiK+ toK+: an embryonic clock?
Authors: Day Margot L. , Winston Nicola , McConnell Josie L. , Cook David , Johnson Martin H.
Published: September 2002

During embryogenesis cells make appropriately timed developmental decisions.
Both ‘hourglass-like’ and ‘clock-like’ mechanisms have
been demonstrated to act as timers in early development. The cell cycle
rhythm, using feedback circuits to drive cells unidirectionally through
checkpoints, is an example of a clock-like timer, but how it operates to time
developmental events is unclear. In other cell types, cyclic oscillations in K+ channel activity, which parallel cell cycle and
circadian rhythms, may be part of the timing mechanism. Changes in K+ oscillations accompany key developmental
transitions and oncogenic transformation. Channel blockade interferes
pharmaco-logically with cell cycle initiation or progression, whereas channel
over-expression can be oncogenic. K+ channel
activity also exists in early mouse oocytes through to at least the blastocyst
stage, and it oscillates in phase with the developmental cell cycles, being
high in M/G1 and low in S/G2. It resembles physiologically the activity of the K+ channels of the eag-
or erg-like families. Reverse
transcriptase–polymerase chain reaction of mouse oocytes has revealed
the presence of transcripts encoding both EAG- and ERG-like proteins
throughout preimplantation devel-opment. Channel activity continues to
oscillate with a cell cycle periodicity in embryos from which the nucleus has
been removed, or after inhibition by puromycin of the cyclin
B–cyclin-dependent kinase 1 driven component of the chromosomal cycle.
Channel oscillatory activity thus appears to be able to function autonomously
of the chromo-somal cycle and may represent a distinct oscillatory timing
activity with possible developmental significance.
Scientific considerations relating to the ethics of the use of human embryonic stem cells in research and medicine
Authors: Pera Martin F.
Published: September 2002

The recent development of embryonic stem (ES) cells from human blastocysts has
the potential to revolutionize many of our approaches to human biology and
medicine. Continued objection to the use of human ES cells on ethical grounds
may inhibit progress or defer this opportunity indefinitely. It is essential
that the ethical discussion proceed on a sound scientific basis. The ethical
controversy surrounding human ES cells concerns their origin from human
blastocysts and the perception of their developmental potential. It is likely
that the worldwide requirement for human ES cells will be met by the
development of a small number of cell lines, as has been the case in the
mouse; current rates of success for human ES cell establishment suggest that
only a modest number of embryos will be required to achieve this goal. It is
in the public interest that human ES cell lines be derived under circumstances
that will enable their widespread distribution with minimum encumbrances to
academic researchers throughout the world. In considering the developmental
potential of ES cells, an important distinction exists between
pluripotentiality, or the ability to develop into a wide range of somatic and
extraembryonic tissues, and totipotentiality, the ability of a cell or
collection of cells to give rise to a new individual given adequate maternal
support. There is no evidence that ES cells from any species can give rise to
a new individual except when combined with cells which are the immediate
progeny of a zygote. These developmental limitations of ES cells appear to
relate to their inability to undergo axis formation and to generate the body
plan. Alternatives to blastocyst-derived ES cells include embryonic germ
cells, adult tissue stem cells, transdetermination of committed somatic cells,
and therapeutic cloning. These research areas are complimentary and
synergistic to ES cell research and it is premature and counterproductive to
suggest that one avenue should be pursued in preference to another. The
combination of cloning and ES cell technology has the potential to address
many important issues in transplantation medicine and research, but a better
understanding of the reprogramming of somatic cells is required before we can
regard ES cells derived from normal nd nuclear transfer blastocysts as
equivalent.
Embryo transfer: how to ensure correct placement in utero
Authors: Hale Lyndon
Published: September 2002

A review of current literature (1990–2000) was undertaken to identify
factors that are important in the correct placement of embryos
in utero. Although there are no absolute prerequisites,
ultrasound monitoring to ensure correct catheter placement, pretreatment
assessment of the cervical canal, minimization of trauma, avoidance of mucus
and the use of a ‘soft’ catheter are all considered important
factors in this process. Early patient mobilization after the transfer
procedure has not been shown to influence the outcome. The establishment of a
benchmark against which to evaluate individual performance and participation
in ‘refresher courses’ if indicated will help to maximize the
implantation rates of the assisted reproductive technology programme as a
whole.
Size distribution and meiotic competence of oocytes obtained from bitch ovaries at various stages of the oestrous cycle
Authors: Otoi Takeshige , Ooka Aya , Murakami Masako , Karja N.W. Kurniani , Suzuki Tatsuyuki
Published: September 2002

The present study was conducted to examine the effects of the stage of the
oestrous cycle on the meiotic competence of canine oocytes and also to
investigate the relationship between the stage of the oestrous cycle and the
relative size distribution of oocytes obtained from bitches at three stages of
the cycle (anoestrus, follicular phase and dioestrus). Only healthy-looking
cumulus–oocyte complexes were used for in vitro
maturation and these were divided into three groups based on diameter
(<110, 110 to <120 and ≥120 µm). The mean
diameter of oocytes from ovaries at anoestrus, the follicular phase and
dioestrus was 103.6, 119.2 and 107.7 µm, respectively. The
percentage of large oocytes (120 µm) collected at the follicular
phase was higher (P<0.01) than that collected at
dioestrus and the percentage of oocytes ≥120 µm collected
from ovaries at dioestrus was higher (P<0.01)
than that collected at anoestrus. After culture for 72 h, significantly more
oocytes reached metaphase II (MII) in the follicular phase than in the other
stages (P<0.01), and more oocytes reached MII in
dioestrus than in anoestrus (P<0.05). In the
≥120 µm group, the frequency of oocytes that resumed meiosis
in the follicular phase was higher (P<0.05) than
in the other stages. However, in the smaller diameter (<120
µm) groups, there were no significant differences between ovaries at
different stages of the oestrous cycle with respect to the proportion of
oocytes reaching each stage of meiosis. Thus, the oestrous cycle stage
influences maturation frequency. Moreover, oocytes demonstrated a size-related
ability to undergo meiotic maturation, irrespective of the stage of the
oestrous cycle. These results suggest that the effects of the stage of the
oestrous cycle may result from differences in the distribution of large
oocytes.
Relationship between the characteristics of frozen-thawed ram spermatozoa and in vitro embryo production
Authors: Morris Lee H. A. , Johnson W. H. , Leibo S. P. , Buckrell B. C.
Published: September 2002

To select rams suitable for ovine in vitro embryo
production (IVP), the predictive values of the screening tests used to
identify unsuitable rams need to be established. The present study examined
some characteristics of frozen–thawed ram spermatozoa that might be
evaluated routinely in a commercial breeding programme. These included sperm
motility, plasma membrane integrity, morphology, and acrosome and capacitation
status of the sperm population. Cryopreserved spermatozoa from four Dorset
rams, which had previously satisfied the selection criteria for inclusion in a
commercial breeding programme, were used for IVP. The overall contribution of
the four rams and the ejaculates within each ram to the variability
(R2) in the production of
blastocysts was very small (2.1% and 2.5% respectively). The
analysis of the sperm characteristics by logistic regression revealed a
significant and positive association between total post-thaw sperm motility,
viability and longevity with in vitro blastocyst
production. However, there was no association between the other surface
characteristics of the spermatozoa measured in this study with embryo
production. Despite the absence of differences between the rams in the low
incidence of polyspermic fertilization, the significant and detrimental
effects of polyspermic fertilization on in vitro
blastocyst production rates were quantified by logistic regression analysis. A
large proportion of the variability within the IVP system was unaccounted for
by the analysis of sperm and oocyte characteristics evaluated in this study.
Thus, the identification of other factors contributing to the variability in
the production of embryos in vitro warrants further
investigation. No single sperm characteristic was sufficient to predict the
ultimate outcome of blastocyst production. Rather, assessments of multiple
characteristics within the IVP system are required to make accurate
predictions.
Differential influence of recombinant non-glycosylated and glycosylated glycodelin on human sperm function: comparative studies with hamster spermatozoa
Authors: Dutta B. , Ain R. , Seshagiri P. B. , Karande A. A.
Published: September 2002

Glycodelin, also known as placental protein 14, has been implicated in
endometriosis-related infertility. To determine the role of glycodelin and its
glycosylated state, the influence of recombinant nonglycosylated-glycodelin
(nongly-glycodelin) and glycosylated-glycodelin
(gly-glycodelin) on human sperm function was evaluated.
Whereas there was a significant (P<0.001) increase in
the capacitation of nongly-glycodelin-treated
spermatozoa compared with untreated controls (28.8 1.0%
v. 21 1.5% respectively), treatment of
spermatozoa with gly-glycodelin markedly
(P<0.001) inhibited capacitation (10.7 0. 3%);
acrosome reaction (AR) remained unaltered in all treatments. In a zona-free
hamster egg penetration assay, the egg penetration index was higher
(P<0.001) with
nongly-glycodelin-treated spermatozoa (3.4 0.3) than
with gly-glycodelin-treated spermatozoa (0.4 0.1) and
untreated spermatozoa (1.6 0.2). A similar influence of glycodelin on
capacitation was observed with hamster spermatozoa. However, the AR rate was
higher (P<0. 01) in
nongly-glycodelin-treated spermatozoa (39.4
1.6%) than in either gly-glycodelin-treated
spermatozoa (19.3 2.0%) or untreated controls (30.0 1.2%).
Moreover, the in vitro fertilization rate was
significantly (P<0.01) higher with
nongly-glycodelin treated-spermatozoa compared with
untreated spermatozoa (77.5 2.3% v. 52.9
4.3%) and gly-glycodelin-treated spermatozoa
(38.3 6.5%; P<0. 05). These results indicate
that whereas nongly-glycodelin improves,
gly-glycodelin inhibits, capacitation and fertilization
potential of human and hamster spermatozoa, and that the glycosylation status
of glycodelin determines its influence on sperm
function.
Immunolocalization of retinol-binding protein, cellular retinoic acid-binding protein I and retinoid X receptor b in the porcine reproductive tract during the oestrous cycle
Authors: Schweigert Florian J. , Siegling Christiane
Published: September 2002

Retinoid-binding proteins and nuclear receptors are expressed in the
reproductive tissues of different species and their expression is hormonally
regulated. In the present study, we demonstrated immunocytochemically the
temporal and spatial localization of retinol-binding protein (RBP), cellular
retinoic acid-binding protein I (CRABPI) and retinoid X receptor β
(RXRβ) in porcine ovary, oviduct and uterus during the oestrous cycle. RBP
and CRABPI were localized in the cytoplasm, whereas RXRβ occurred in the
nucleus. RBP was not detected in either the ovary or the oviduct at any stage
of the oestrous cycle. CRABPI was present in luteal cells of the ovary only
during dioestrus and in glandular and ciliated cells of the oviduct during
oestrus. In the ovary, RXRβ was always present in granulosa cells and
germinal epithelium, with highest levels observed during oestrus. In the
uterus, RXRβ was present throughout the cycle in both the endometrium and
the myometrium. However, changes in RXRβ were observed in the endometrium,
with highest levels observed during dioestrus. RBP and CRABPI could be
observed in the endometrium only during dioestrus. The results show that the
occurrence of retinoid-binding proteins and nuclear receptors in individual
tissues of the reproductive tract are strongly dependent on the stage of the
oestrous cycle. In the oviduct, the expression of CRABPI seems to be dependent
on oestrogen, whereas in the uterus the expression of RBP and CRABPI is
influenced by progesterone. The association of expression in different
sections of the reproductive tissues investigated shows that the presence of
specific proteins involved in retinoid metabolism was dependent on events
associated with ovulation, the migration of the oocyte through the oviduct and
the possible implantation of the blastocyst into the uterus.
Estrogens and human cardiovascular physiology
Authors: Komesaroff Paul A. , Sudhir Krishnankutty
Published: September 2002

The importance of estrogens for cardiovascular physiology in women is
accepted, even if the clinical place for hormonal supplementation after
menopause remains uncertain. However, although men produce significant
quantities of estrogen, relatively little is known about the role of estrogens
in the male cardiovascular system. Recent evidence about the epidemiology of
cardiovascular disease, the mechanisms of action of estrogens, the biological
actions of endogenously produced estrogens in men, and the acute and chronic
effects of estrogens on the vasculature in both men and women, suggests that
these hormones may well play an important role in normal male cardiovascular
physiology. In addition, they may provide useful adjuncts to therapy in
selected groups of men.
Glycine betaine improves survival of fresh bovine spermatozoa
Authors: Zhang Bing Rong , Buhr Mary , Kroetsch Tom , Leibo S. P.
Published: September 2002

Fresh spermatozoa from bulls established as ‘good freezers’ and
‘poor freezers’ (consistently ≥50% or
<20% motile spermatozoa after cryopreservation, respectively)
were incubated for 96 h in Tes/Tris–egg yolk or TALP–egg yolk
media at 37°, 20°, 5° or 0°C. The TALP
extender contained 0, 100 or 200 mM glycine betaine (GB) to test the
hypothesis that GB would efficiently maintain spermatozoa function during
long-term incubation. The percentage of motile spermatozoa declined over time
in a temperature- and medium-dependent fashion. No spermatozoa were motile by
24 h incubation at 37°C or by 72 h incubation at 0°C, and
there were no significant differences in the percentage of motile spermatozoa
from either category of bull when spermatozoa were incubated in any media for
less than 24 h. Spermatozoa from poor freezers were significantly more motile
than spermatozoa from good freezers after 96 h at 20° or 5°C
in TALP alone; however, GB at both 100 and 200 mM increased the percentage of
motile spermatozoa in poor and good freezers and eliminated these differences.
Overall, the presence of GB at either 100 or 200 mM significantly improved the
percentage of motile spermatozoa at 20°, 5° and 0°C,
but not at 37°C.
Patterns of apoptosis in the corpora lutea of the rat during the oestrous cycle, pregnancy and in vitro culture
Authors: Bruce N. W. , Hisheh S. , Dharmarajan A. M.
Published: September 2002

Apoptosis is associated with regression of corpora lutea (CL) in a number of
species. The present work compared apoptotic rates, assessed by low molecular
weight DNA labelling, in rat CL of the oestrous cycle, pregnancy, the
immediate post-partum period, and in vitro culture.
Apoptosis was significantly related to cycle phase with peak activity at
oestrus. This pattern was similar in CL formed from the most recent ovulation
and from the two previous generations. Apoptosis was evident during pregnancy,
albeit at low rates. It declined on the day of parturition and increased on
the following day. Apoptosis was greatly increased in cultured CL to reach
20-fold higher levels than those achieved during the oestrous cycle or
pregnancy. Collectively, these results suggest that apoptosis has lesser
significance in rat CL functional regression than in other species with more
precipitate structural changes. However, rat CL clearly retain the potential
for high rates of apoptosis and so present a useful model for examining
molecular mechanisms of apoptotic cell death.
Plasma thyroid hormones and growth hormone in embryonic and growing emus (Dromaius novaehollandiae)
Authors: Blache D. , Blackberry M. A. , van Cleeff J. , Martin G. B.
Published: September 2002

Growth hormone (GH), thyroxine (T4) and
tri-iodothyronine (T3) are known to be involved in the
regulation of growth and development in a variety of avian species. It has
been suggested that an absence of GH and thyroid hormones in ostriches is the
cause of their neoteny, a phenomenon in which juvenile characteristics are
retained into adulthood. Neoteny is typical of all ratites, the single group
of flightless birds that includes the ostrich, but similar endocrine studies
have not been performed for other members of the group, such as the emu. To
test the neoteny hypothesis further, in the present study we measured the
plasma concentrations of T4, T3
and GH in emus during embryonic development and from hatching to 1 year of
age. Concentrations of T4 and GH increased during the
last weeks of incubation, whereas concentrations of T3
were highly variable. After hatching, the concentrations of both thyroid
hormones were high during the first 3 days of life and then fell to a constant
low level. Plasma concentrations of GH were high at the time of hatching and
decreased gradually over the first 22 weeks of age; thereafter, the
concentrations of GH were highly variable. No correlation was observed between
hormone concentrations and live weight at any time. These results support the
hypothesis that thyroid function is abnormally low in ratites, whereas
patterns of GH secretion are similar to those observed in other birds.
Dysfunction of the thyroid axis could explain, in part, the neotenous physical
aspect of adult emus.
Effects of caffeine and its reactive metabolites theophylline and theobromine on the differentiating testis
Authors: Pollard Irina , Locquet Odette , Solvar Antoinette , Magre Solange
Published: September 2002

A previous study in the rat (Pollard et al. 1990)
established that caffeine, when administered during pregnancy, significantly
inhibited the differentiation of the seminiferous cords and subsequent Leydig
cell development in the interstitium. However, that study could not
distinguish between the direct effects of caffeine and/or the intermediary
secondary toxic effects of metabolites such as theophylline and theobromine.
Because the fetus lacks the appropriate enzyme systems, clearance of toxic
substances takes place via the placenta and maternal liver. Thus, a suitable
in vitro system can effectively differentiate between
primary and secondary drug effects. In the present study, 13-day-old fetal
testis, at the stage of incipient differentiation, were cultured for 4 days
in vitro in the presence of graded doses of caffeine,
theophylline or theobromine. It was found that explants exposed to caffeine or
theobromine differentiated normally, developing seminiferous cords made up of
Sertoli and germ cells, soon followed by the differentiation of functionally
active Leydig cells appearing in the newly formed interstitium. However,
explants exposed to theophylline failed to develop seminiferous cords and, as
a consequence, Leydig cells. In conclusion, insights obtained from different
experimental methods, such as organ culture or whole organism studies, are not
always identical. It may be prudent, therefore, to take into account that
certain experimental techniques, despite providing valuable information, may
require confirmation by other test methods in order to obtain an in-depth
understanding of mechanisms of action involved.
Effect of centrifugation on early embryonic development and parthenogenetic activation of bovine oocytes matured in vitro
Authors: Chung Jin-Tae , Downey Bruce R. , Casper Robert F. , Chian Ri-Cheng
Published: September 2002

This study examined the fertilization, early developmental competence and
capacity for parthenogenetic activation of bovine oocytes matured
in vitro after centrifugation. Immature oocytes were
cultured in tissue culture medium 199 supplemented with 10% fetal
bovine serum and 75 mIU mL–1 FSH + LH at
5% CO2 to facilitate maturation. After culture
for 24 or 30 h, the metaphase-II stage oocytes were centrifuged at 3000, 5000,
7000 or 10000g for 5 min before in vitro fertilization
or parthenogenetic activation. Frozen–thawed bull semen was used for
in vitro fertilization. For parthenogenetic activation,
the oocytes were exposed to 20 M calcium ionophore A23187 for 5 min at room
temperature. Fertilization rates were not different between control and
treatment groups (87.7% v. 74.6%,
73.4%, 75.9% and 76.4% respectively). Also, there were no
differences in early embryonic development between control and treatment
groups (rates of blastocyst formation were 21.1%
v. 20.2%, 28.8%, 31.2% and
24.1% respectively). When the oocytes were centrifuged at various
speeds alone, the activation rate of oocytes was significantly higher
(P<0.05) in the 10 000g treatment group compared with
control (10.8% v. 0.0%). There were no
differences in the activation rates of oocytes between control and treatment
groups at speeds up to 7000g (70.9% v.
71.9%, 78.3% and 77.2% respectively) after centrifugation
and stimulation with Ca2+-ionophore. However, the
activation rate of oocytes was significantly higher
(P<0.05) in the 10 000g treatment group compared with
control (70.9% v. 83.1%). In addition, the
percentage of activated oocytes with diploid formation was significantly
higher in the oocytes after centrifugation at 10 000g and stimulation with
calcium ionophore A23187 than in the control (18.4%
v. 7.1%). These results indicate that
centrifugation of oocytes matured in vitro has no
detrimental effect on fertilization and subsequent early embryonic
development. They also indicate that the oocytes might be parthenogenetically
activated after centrifugation and that high-speed centrifugation may induce
activation of some oocytes. The results suggest that the optimal speed for
centrifugation of bovine oocytes might be ≤7000g to enhance the visibility
of nuclear elements for further micromanipulation.
Technical Report: Effect of commercially available PMSG on maturation, fertilization and embryo development of buffalo oocytes in vitro
Authors: Gupta P. S. P. , Nandi S. , Ravindranatha B. M. , Sarma P. V.
Published: September 2002

In vitro fertilization (IVF) technology provides an
opportunity to produce embryos for genetic manipulation, embryo transfer and
basic research in developmental physiology, and can be exploited for emerging
biotechnologies such as transgenesis and cloning. In the present study, the
effects of different concentrations of commercially available pregnant mare
serum gonadotrophin (PMSG) (Folligon; Intervet, International B.V., Boxmeer,
Holland) in oocyte culture media, on maturation, fertilization and embryonic
development of buffalo oocytes in vitro were
investigated. Oocytes aspirated from abattoir-derived ovaries were cultured in
media containing TCM-199 + PMSG at 0, 2.5, 20, 30, 40 and 50 IU
mL–1 in presence or absence of steer serum
(10%) for 24 h in a CO2 incubator. The maturation
rate was assessed on the basis of degree of expansion of cumulus cells. The
matured oocytes were inseminated with 9–10 x 106
spermatozoa mL–1 in Brackett and Oliphant medium
and the cleavage rate was recorded 40–42 h after insemination. Uncleaved
oocytes were stained with aceto-orcein for evaluation of fertilization rates.
The cleaved embryos were further cultured in TCM-199 + 10% steer
serum on buffalo oviducal cell monolayer for 7 days. Maturation,
fertilization, cleavage and embryonic development were significantly higher
(P<0.05) in oocytes cultured in TCM-199 +
10% steer serum supplemented with 40 and 50 IU PMSG
mL–1. It is concluded that commercially available
PMSG can effectively be used in place of pure follicle-stimulating hormone for
in vitro maturation of buffalo oocytes, making it cost
effective for IVF studies.
The placenta in multiple pregnancy: outstanding issues
Authors: Benirschke Kurt , Masliah Eliezer
Published: September 2002

The present paper reviews the placental development in multiple gestations and
highlights the twin-to-twin transfusion syndrome, its recognition and possible
therapy, and some of the consequences of fetal death of one twin. It presents
data on discordance for structural anomalies in monozygotic twins. These
aspects are discussed with respect to their possible pathogenetic origins.
Antigestogens: the holy grail of contraception
Authors: Baird David T.
Published: September 2002

Progesterone is essential for the establishment and maintenance of pregnancy.
The discovery of antagonists of progesterone offered the possibility of
developing new methods of fertility regulation. It was very quickly
demonstrated that antigestogens would induce bleeding in early pregnancy, and
mifepristone in combination with a prostaglandin is used as a method of
abortion in several countries. Mifepristone will also inhibit ovulation,
prevent implantation and induce menstrual bleeding at the time of expected
menses. However, the development of these contraceptive uses of antigestogens
has been inhibited by political and ethical controversy surrounding abortion.
Surely the time has come to dissociate the ongoing debate about the ethics of
abortion from the method so that the full therapeutic potential of this class
of drugs can be exploited fully.
Mitochondrial DNA and the Y chromosome: parallels and paradoxes
Authors: Cummins Jim
Published: September 2002

Both mitochondrial DNA (mtDNA) and the Y chromosome have been used extensively
by molecular paleoanthropologists in attempts to reconstruct human lineages.
Both are inherited in a haploid manner: mtDNA through the female and the Y
through the male. For mtDNA, maternal inheritance is ensured by a
species-specific mechanism of proteolysis of the sperm midpiece in early
embryogenesis, based on ubiquitination of the mitochondria during
spermiogenesis. Both genomes are thought to lack recombination and are thus
liable to high rates of neutral mutation. For the human Y chromosome, it is
now clear that there has been selection on genes controlling spermatogenesis,
resulting in differential long-term reproductive success. This is corroborated
from studies of genealogies and hunting–gathering societies, although
these lack the rigour provided by the modern molecular markers of inheritance.
Selection is made more complicated by a concentration of genes controlling
secondary sexual characteristics on the X chromosome. Likewise, mtDNA affects
the bioenergetics of gametogenesis and embryo development, as well as
longevity, disease and the aging process. Both Y chromosome and mitochondrial
haplotypes show significant associations with patterns of male infertility
that could distort their use for phylogenetic reconstruction. Moreover, the
molecular analysis of mtDNA is complicated by the presence of numerous nuclear
mitochondrial pseudogenes (Numts) that can be erroneously amplified by
molecular techniques such as PCR. This review examines some of these complex
interactions and suggests that some of the more contentious issues in
understanding human evolution may be resolved by considering the biology of
these genetic markers.