Immunoreactivity for substance P in the gasserian ganglion, ophthalmic nerve and anterior segment of the rabbit eye

Summary The distribution of substance P (SP) immunofluorescence was investigated in the Gasserian ganglion, ophthalmic nerve and in the anterior segment of the rabbit eye. About one third of the nerve cell bodies in the Gasserian ganglion exhibited SP immunofluorescence, which was also observed in some nerve fibres of the ophthalmic nerve. In the cornea, some SP-positive iris contained numerous nerve fibres with SP immunofluorescence. In the sphincter area such fibres were circular, while the orientation of the SP fibres was radial in the dilator muscle. Both in the iris and in the ciliary body, the largest vessels were surrounded by nerves exhibiting SP immunofluorescence. A few nerve fibres also appeared in the stroma of the ciliary processes.     

Unusual patterns of benzo[a]pyrene metabolites and DNA-benzo[a]pyrene adducts produced by human placental microsomes in vitro

Human placental microsomes were incubated with [³H]benzo[a]pyrene (BP) and Salmon sperm DNA and the resulting metabolite-nucleoside complexes resolved by Sephadex LH-20 chromatography. The metabolite pattern was analyzed by high-pressure liquid chromatography (HPLC). The incubates were also co-chromatographed with extracts obtained from incubates with rat liver microsomes and [¹⁴C]BP. Phenols, quinones and 7,8-dihydrodiol were detected in the placental incubates. Both 9,10- and 4,5-dihydrodiols were very low as compared with control rat liver samples. Placental microsomes catalyzed the binding of BP metabolites to DNA in vitro, giving rise to two main complexes which co-chromatographed with rat liver-produced peaks attributable to 7,8-diol-9,10-epoxide and 7,8-oxide and/or quinones when metabolized further. The nucleoside metabolite peaks attributable to 4,5-oxide and 9-phenol-4,5-oxide were lacking when compared with the binding pattern catalyzed by rat liver. Both the total binding and specific metabolite-nucleoside adducts in the placenta correlated with fluorometrically measured aryl hydrocarbon hydroxylase (AHH) activity and with the amount of dihydrodiol formed. The results demonstrate that both the metabolite pattern and the nucleoside-metabolite complexes formed by the placental microsomes in vitro differed greatly from those produced by rat liver microsomes. These studies also suggest that it is not possible to predict specific patterns of DNA binding from AHH measurements or even from BP metabolite patterns, especially when comparing different tissues and species.     

Background levels of hydrogen cyanide in human breath measured by infrared cavity ring down spectroscopy

Hydrogen cyanide (HCN) in breath has been suggested as a diagnostic tool for cyanide poisoning and for cyanide-producing bacterial infections. To distinguish elevated levels of breath HCN, baseline data are needed. Background levels of HCN were measured in mixed exhaled air from 40 healthy subjects (26 men, 14 women, age 21–61 years; detection limit: 1.5?ppb; median: 4.4?ppb; range <1.5–14?ppb) by near-infrared cavity ring down spectroscopy (CRDS). No correlation was observed with smoking habits, recent meals or age. However, female subjects had slightly higher breath levels of HCN than male subjects. CRDS has not previously been used for this purpose.     

Gibberellin structure and function: biological activity and competitive inhibition of gibberellin 2- and 3-oxidases

Some gibberellin (GA) analogues, especially with C-16,17 modifications of GA(5), can inhibit growth of plants apparently by acting as competitors with the endogenous substrate of GA biosynthetic enzymes. Here, we directly confirm the competitive action of GA derivatives but also show that some analogues may retain significant bioactivity. A recombinant 3-oxidase from pea, which converts GA(20) to bioactive GA(1), was inhibited by GA(5), and 16,17-dihydro-GA(5) derivatives, especially if the C-17 alkyl chain length was increased by up to three carbons or if the C-13 hydroxyl was acetylated. Genetic confirmation that GA(5) analogues target 3-oxidases in vivo was provided by comparing the growth response of a WT (LE) pea with a 3-oxidase mutant (le-1). Two pea 2-oxidases that inactivate bioactive GAs, were inhibited by GA(1) and GA(3) but were generally insensitive to GA(5) analogues. alpha-Amylase production by barley half-seeds in response to GA analogues provided a method to study their action when effects on GA biosynthesis were excluded. This bioactivity assay showed that 16,17-dihydro GA(5) analogues have some inherent activity but mostly less than for GA(5) (5-50-fold), which in turn was 100-fold less active than GA(1) and GA(3). However, although C-17 alkyl derivatives with one or two added carbons showed little bioactivity and were purely 3-oxidase inhibitors, adding a third carbon (the 17-n-propyl-16,17-dihydro GA(5) analogue) restored bioactivity to that of GA(5). Furthermore, this analogue has lost its capacity to inhibit stem elongation of Lolium temulentum (Mander et al., Phytochemistry 49:1509-1515, 1998a), although it strongly inhibits the 3-oxidase. Thus, the effectiveness of a GA derivative as a growth retardant will reflect the balance between its bioactivity and its capacity to inhibit the terminal enzyme of GA biosynthesis. The weaker growth inhibition in dicots including pea (approximately 10%) than in monocots such as L. temulentum (>35%) is suggestive of taxonomic differences in the bioactivity of GAs and/or their effects on GA biosynthesis.     

Development of biochemical specialization and organelle partitioning in the single-cell C4 system in leaves of Borszczowia aralocaspica (Chenopodiaceae)

The terrestrial plant Borszczowia aralocaspica (Chenopodiaceae) has recently been shown to contain the entire C(4) photosynthesis mechanism within individual, structurally and biochemically polarized chlorenchyma cells rather than in a dual cell system, as has been the paradigm for this type of carbon fixation (Nature 414: 543-546, 2001). Analysis of carbon isotope composition and (14)CO(2) fixation shows that photosynthesis and growth of B. aralocaspica occurs through carbon acquired by C(4) photosynthesis. The development of this unique single-cell C(4) system in chlorenchyma cells was studied by analysis of young (0.2-0.3 cm length), intermediate (ca. 0.5-0.6 cm length), and mature leaves (ca. 3 cm length). The length of chlorenchyma cells approximately doubles from young to intermediate and again from intermediate to the mature leaf stage. In young chlorenchyma cells, there is a single type of chloroplast; the chloroplasts are evenly distributed throughout the cytosol, and all contain starch and rubisco. During leaf development, the activities of phosphoenolpyruvate carboxylase (PEPC; which is cytosolic), rubisco, and pyruvate,Pi dikinase (PPDK) increase on a chlorophyll basis. As leaves mature, chloroplasts differentiate into two distinct structural and biochemical types that are spatially separated into the proximal and distal parts of the cell (the proximal end being closest to the center of the leaf). The early stages of this polarization are observed in intermediate leaves, and the polarization is fully developed in mature leaves. The chloroplasts in the distal ends of the cell have reduced grana and little starch, while those at the proximal ends have well-developed grana and abundant starch. In mature leaves, PPDK is expressed in chloroplasts at the distal end of the cells, while rubisco and adenosine diphosphate glucose (ADPG) pyrophosphorylase are selectively expressed in chloroplasts at the proximal end of the cell. Mitochondrial polarization also occurs during development as nicotinamide-adenine dinucleotide phosphate-malic enzyme (NAD-ME) and the photorespiratory enzyme glycine decarboxylase are expressed in mature but not young leaves and are localized in mitochondria at the proximal end of the cells. The data show that single-cell C(4) develops from a single pool of identical organelles that develop differential biochemical functions and spatial partitioning in the cell during maturation.     

Susceptibility loci for preeclampsia on chromosomes 2p25 and 9p13 in Finnish families

Preeclampsia is a common, pregnancy-specific disorder characterized by reduced placental perfusion, endothelial dysfunction, elevated blood pressure, and proteinuria. The pathogenesis of this heterogeneous disorder is incompletely understood, but it has a familial component, which suggests that one or more common alleles may act as susceptibility genes. We hypothesized that, in a founder population, the genetic background of preeclampsia might also show reduced heterogeneity, and we have performed a genomewide scan in 15 multiplex families recruited predominantly in the Kainuu province in central eastern Finland. We found two loci that exceeded the threshold for significant linkage: chromosome 2p25, near marker D2S168 (nonparametric linkage [NPL] score 3.77; P=.000761) at 21.70 cM, and 9p13, near marker D9S169 (NPL score 3.74; P=.000821) at 38.90 cM. In addition, there was a locus showing suggestive linkage at chromosome 4q32 between D4S413 and D4S3046 (NPL score 3.13; P=.003238) at 163.00 cM. In the present study the susceptibility locus on chromosome 2p25 is clearly different (21.70 cM) from the locus at 2p12 found in an Icelandic study (94.05 cM) and the locus at 2q23 (144.7 cM) found in an Australian/New Zealand study. The locus at 9p13 has been shown to be a candidate region for type 2 diabetes in two recently published genomewide scans from Finland and China. The regions on chromosomes 2p25 and 9p13 may harbor susceptibility genes for preeclampsia.     

Far red end-of-day treatment restores wild type-like plant length in hybrid aspen overexpressing phytochrome A

Shoot elongation in woody plants is modulated by a multitude of light signals, including irradiance, photoperiod and spectral composition, for which the phytochrome system is the probable photoreceptor. In hybrid aspen ( Populus tremula  ×  tremuloides ) overexpression of the oat phytochrome A ( PHYA ) prevents growth cessation in response to short photoperiod, and plants exhibit dwarf growth that is related to reduced cell numbers and reduced gibberellin contents. End-of-day far-red treatment significantly enhances internode elongation in PHYA overexpressors as well as in the wild type, and this was found here to be caused by stimulation of cell division and cell extension. In PHYA overexpressors the effects were substantially larger than in the wild type, and resulted in complete restoration of wild type-like plant length as well as cell numbers, and gibberellin content was greatly increased. No clear effect of far-red end-of-day treatment on gibberellin levels could be detected in the wild type. It thus appears that the far-red end-of-day treatment might modify the responsiveness of the tissue to GA rather than the GA levels. The observed effects were completely reversed by a subsequent irradiation with red light. The present data show that dwarfism due to PHYA overexpression can be completely overcome by far red end-of-day treatment, and the observations indicate that effects of far red end-of-day treatments appear to be mediated by phytochrome(s) other than phytochrome A.     

Bundle sheath diffusive resistance to CO(2) and effectiveness of C(4) photosynthesis and refixation of photorespired CO(2) in a C(4) cycle mutant and wild-type Amaranthus edulis

A mutant of the NAD-malic enzyme-type C(4) plant, Amaranthus edulis, which lacks phosphoenolpyruvate carboxylase (PEPC) in the mesophyll cells was studied. Analysis of CO(2) response curves of photosynthesis of the mutant, which has normal Kranz anatomy but lacks a functional C(4) cycle, provided a direct means of determining the liquid phase-diffusive resistance of atmospheric CO(2) to sites of ribulose 1,5-bisphosphate carboxylation inside bundle sheath (BS) chloroplasts (r(bs)) within intact plants. Comparisons were made with excised shoots of wild-type plants fed 3,3-dichloro-2-(dihydroxyphosphinoyl-methyl)-propenoate, an inhibitor of PEPC. Values of r(bs) in A. edulis were 70 to 180 m(2) s(-1) mol(-1), increasing as the leaf matured. This is about 70-fold higher than the liquid phase resistance for diffusion of CO(2) to Rubisco in mesophyll cells of C(3) plants. The values of r(bs) in A. edulis are sufficient for C(4) photosynthesis to elevate CO(2) in BS cells and to minimize photorespiration. The calculated CO(2) concentration in BS cells, which is dependent on input of r(bs), was about 2,000 microbar under maximum rates of CO(2) fixation, which is about six times the ambient level of CO(2). High re-assimilation of photorespired CO(2) was demonstrated in both mutant and wild-type plants at limiting CO(2) concentrations, which can be explained by high r(bs). Increasing O(2) from near zero up to ambient levels under low CO(2), resulted in an increase in the gross rate of O(2) evolution measured by chlorophyll fluorescence analysis in the PEPC mutant; this increase was simulated from a Rubisco kinetic model, which indicates effective refixation of photorespired CO(2) in BS cells.     

Cytochrome P450 enzyme activity in five herbivorous,non-passerine bird species

We examined hepatic cytochrome P450 activity in wild and hand-reared grey partridges (Perdix perdix), capercaillies (Tetrao urogallus) and ring-necked pheasants (Phasianus colchicus), as well as the enzyme activity in a variety of tissues of hand-reared Japanese quails (Coturnix coturnix japonica) and pigeons (Columba livia). Post-mortem decrease in hepatic enzyme activity in the grey partridge was measured. Hepatic 7-ethoxyresorufin-O-deethylase activity was similar in wild and hand-reared grey partridges and pheasants, but the activity was significantly lower in wild than in hand-reared capercaillies, probably resulting from their phenolic-rich diet. In the tissues of both quails and pigeons 7-ethoxycoumarin-O-deethylase exhibited the highest and 7-pentoxyresorufin-O-deethylase the lowest activity. Hepatic enzyme activity was significantly higher than that in other tissues. In the small intestine some activity could be found, reflecting some intestinal detoxication capacity. Enzyme activity decreased by 34-69% during the 30-min sampling period, which confirmed the importance of equalising sampling time to obtain comparable data. Because the hand-reared birds in this study were fed the same commercial diets, we assumed that the enzyme activity values detected reflect species differences without any induction by dietary secondary compounds.