Comparative analysis of genetic diversity in the mangrove species Avicennia marina (Forsk.) Vierh. (Avicenniaceae) detected by AFLPs and SSRs

Avicennia marina is an important mangrove species with a wide geographical and climatic distribution which suggests that large amounts of genetic diversity are available for conservation and breeding programs. In this study we compare the informativeness of AFLPs and SSRs for assessing genetic diversity within and among individuals, populations and subspecies of A. marina in Australia. Our comparison utilized three SSR loci and three AFLP primer sets that were known to be polymorphic, and could be run in a single analysis on a capillary electrophoresis system, using different-colored fluorescent dyes. A total of 120 individuals representing six populations and three subspecies were samplcd. At the locus level, SSRs were considerably more variable than AFLPs, with a total of 52 alleles and an average heterozygosity of 0.78. Average heterozygosity for AFLPs was 0.193, but all of the 918 bands scored were polymorphic. Thus, AFLPs were considerably more efficient at revealing polymorphic loci than SSRs despite lower average heterozygosities. SSRs detected more genetic differentiation between populations (19 vs 9%) and subspecies (35 vs 11%) than AFLPs. Principal co-ordinate analysis revealed congruent patterns of genetic relationships at the individual, population and subspecific levels for both data sets. Mantel testing confirmed congruence between AFLP and SSR genetic distances among, but not within, population comparisons, indicating that the markers were segregating inde- pendently but that evolutionary groups (populations and subspecies) were similar. Three genetic criteria of importance for defining priorities for ex situ collections or in situ conservation programs (number of alleles, number of locally common alleles and number of private alleles) were correlated between the AFLP and SSR data sets. The congruence between AFLP and SSR data sets suggest that either method, or a combination, is applicable to expanded genetic studies of mangroves. The codominant nature of SSRs makes them ideal for further population-based investigations, such as mating-system analyses, for which the dominant AFLP markers are less well suited. AFLPs may be particularly useful for monitoring propagation programs and identifying duplicates within collections, since a single PCR assay can reveal many loci at once.

Combining the analyses of introgressive hybridisation and linkage mapping to investigate the genetic architecture of population divergence in the lake whitefish (Coregonus clupeaformis, Mitchill)

Adaptation and reproductive isolation, the engines of biological diversity, are still elusive when discussing the genetic bases of speciation. Namely, the number of genes and magnitude of selection acting positively or negatively on genomic traits implicated in speciation is contentious. Here, we describe the first steps of an ongoing research program aimed at understanding the genetic bases of population divergence and reproductive isolation in the lake whitefish (Coregonus clupeaformis). A preliminary linkage map originating from a hybrid cross between dwarf and normal ecotypes is presented, whereby some of the segregating AFLP markers were found to be conserved among natural populations. Maximum-likelihood was used to estimate hybrid indices from non-diagnostic markers at 998 AFLP loci. This allowed identification of the most likely candidate loci that have been under the influence of selection during the natural hybridisation of whitefish originating from different glacial races. As some of these loci could be identified on the linkage map, the possibility that selection of traits in natural populations may eventually be correlated to specific chromosomal regions was demonstrated. The future prospects and potential of these approaches to elucidate the genetic bases of adaptation and reproductive isolation among sympatric ecotypes of lake whitefish is discussed.

A comparison of peak power in the shoulder press and shoulder throw

The ability to generate peak power is central for performance in many sports. Currently two distinct resistance training methods are used to develop peak power, the heavy weight/slow velocity and light weight/fast velocity regimes. When using the light weight/fast velocity power training method it was proposed that peak power would be greater in a shoulder throw exercise compared with a normal shoulder press. Nine males performed three lifts in the shoulder press and shoulder throw at 30% and 40% of their one repetition maximum (1RM). These lifts were performed identically, except for the release of the bar in the throw condition. A potentiometer attached to the bar measured displacement and duration of the lifts. The time of bar release in the shoulder throw was determined with a pressure switch. ANOVA was used to examine statistically significant differences where the level of acceptance was set at p

Orthographic/phonological facilitation of naming responses in the picture-word task: An event-related fMRI study using overt vocal responding

In the picture-word interference task, naming responses are facilitated when a distractor word is orthographically and phonologically related to the depicted object as compared to an unrelated word. We used event-related functional magnetic resonance imaging (fMRI) to investigate the cerebral hemodynamic responses associated with this priming effect. Serial (or independent-stage) and interactive models of word production that explicitly account for picture-word interference effects assume that the locus of the effect is at the level of retrieving phonological codes, a role attributed recently to the left posterior superior temporal cortex (Wernicke's area). This assumption was tested by randomly presenting participants with trials from orthographically related and unrelated distractor conditions and acquiring image volumes coincident with the estimated peak hemodynamic response for each trial. Overt naming responses occurred in the absence of scanner noise, allowing reaction time data to be recorded. Analysis of this data confirmed the priming effect. Analysis of the fMRI data revealed blood oxygen level-dependent signal decreases in Wernicke's area and the right anterior temporal cortex, whereas signal increases were observed in the anterior cingulate, the right orbitomedial prefrontal, somatosensory, and inferior parietal cortices, and the occipital lobe. The results are interpreted as supporting the locus for the facilitation effect as assumed by both classes of theoretical model of word production. In addition, our results raise the possibilities that, counterintuitively, picture-word interference might be increased by the presentation of orthographically related distractors, due to competition introduced by activation of phonologically related word forms, and that this competition requires inhibitory processes to be resolved. The priming effect is therefore viewed as being sufficient to offset the increased interference. We conclude that information from functional imaging studies might be useful for constraining theoretical models of word production. (C) 2002 Elsevier Science (USA).

Distribution kinetics of solutes in the isolated in-situ perfused rat head using the multiple indicator dilution technique and a physiological two-barrier model

The purpose of this study was to determine the pharmacokinetics of [C-14]diclofenac, [C-14]salicylate and [H-3]clonidine using a single pass rat head perfusion preparation. The head was perfused with 3-[N-morpholino] propane-sulfonic acid-buffered Ringer's solution. Tc-99m-red blood cells and a drug were injected in a bolus into the internal carotid artery and collected from the posterior facial vein over 28 min. A two-barrier stochastic organ model was used to estimate the statistical moments of the solutes. Plasma, interstitial and cellular distribution volumes for the solutes ranged from 1.0 mL (diclofenac) to 1.6 mL (salicylate), 2.0 mL (diclofenac) to 4.2 mL (water) and 3.9 mL (salicylate) to 20.9 mL (diclofenac), respectively. A comparison of these volumes to water indicated some exclusion of the drugs from the interstitial space and salicylate from the cellular space. Permeability-surface area (PS) products calculated from plasma to interstitial fluid permeation clearances (CLPI) (range 0.02-0.40 mL s(-1)) and fractions of solute unbound in the perfusate were in the order: diclofenac>salicylate >clonidine>sucrose (from 41.8 to 0.10 mL s(-1)). The slow efflux of diclofenac, compared with clonidine and salicylate, may be related to its low average unbound fraction in the cells. This work accounts for the tail of disposition curves in describing pharmacokinetics in the head.

Multiple maps and activity-dependent representational plasticity in the anterior Wulst of the adult barn owl (Tyto alba)

In the present study we addressed the issue of somatosensory representation and plasticity in a nonmammalian species, the barn owl. Multiunit mapping techniques were used to examine the representation of the specialized receptor surface of the claw in the anterior Wulst. We found dual somatotopic mirror image representations of the skin surface of the contralateral claw. In addition, we examined both representations 2 weeks after denervation of the distal skin surface of a single digit. In both representations, the denervated digital representation became responsive to stimulation of the adjacent, mutually functional, digit. The mutability and multiple representations indicates that the Wulst provides the owl with sensory processing capabilities analogous to those in mammals.

Gene complementation of airway epithelium in the cystic fibrosis mouse is necessary and sufficient to correct the pathogen clearance and inflammatory abnormalities

Increasingly, cystic fibrosis (CF) is regarded as an inflammatory disorder where the response of the lung to Pseudomonas aeruginosa is exaggerated as a consequence of processes mediated by the product of the CF gene, CFTR. Of importance to any gene-replacement strategy for treatment of CF is the identification of the cell type(s) within the lung milieu that need to be corrected and an indication whether this is sufficient to restore a normal inflammatory response and bacterial clearance. We generated G551D CF mice transgenically expressing the human CFTR gene in two tissue compartments previously demonstrated to mediate a CFTR-dependent inflammatory response: lung epithelium and alveolar macrophages. Following chronic pulmonary infection with P. aeruginosa, CF mice with epithelial-expressed but not macrophage-specific CFTR showed an improvement in pathogen clearance and inflammatory markers compared with control CF animals. Additionally, these data indicate the general role for epithelial cell-mediated events in the response of the lung to bacterial pathogens and the importance of CFTR in mediating these processes.

The distribution and morphological characteristics of catecholaminergic cells in the brain of monotremes as revealed by tyrosine hydroxylase immunohistochemistry

The present study describes the distribution and cellular morphology of catecholaminergic neurons in the CNS of two species of monotreme, the platypus (Ornithorhynchus anatinus) and the short-beaked echidna (Tachyglossus aculeatus). Tyrosine hydroxylase immunohistochemistry was used to visualize these neurons. The standard A1-A17, C1-C3 nomenclature was used for expediency, but the neuroanatomical names of the various nuclei have also been given. Monotremes exhibit catecholaminergic neurons in the diencephalon (All, A12, A13, A14, A15), midbrain (A8, A9, A10), rostral rhombencephalon (A5, A6, A7), and medulla (A1, A2, C1, C2). The subdivisions of these neurons are in general agreement with those of other mammals, and indeed other amniotes. Apart from minor differences, those being a lack of A4, A3, and C3 groups, the catecholaminergic system of monotremes is very similar to that of other mammals. Catecholaminergic neurons outside these nuclei, such as those reported for other mammals, were not numerous with occasional cells observed in the striatum. It seems unlikely that differences in the sleep phenomenology of monotremes, as compared to other mammals, can be explained by these differences. The similarity of this system across mammalian and amniote species underlines the evolutionary conservatism of the catecholaminergic system. Copyright (C) 2002 S. Karger AG, Basel.

The distribution and morphological characteristics of serotonergic cells in the brain of monotremes

The distribution and cellular morphology of serotonergic neurons in the brain of two species of monotremes are described. Three clusters of serotonergic neurons were found: a hypothalamic cluster, a cluster in the rostral brainstem and a cluster in the caudal brainstem. Those in the hypothalamus consisted of two groups, the periventricular hypothalamic organ and the infundibular recess, that were intimately associated with the ependymal wall of the third ventricle. Within the rostral brainstem cluster, three distinct divisions were found: the dorsal raphe nucleus (with four subdivisions), the median raphe nucleus and the cells of the supralemniscal region. The dorsal raphe was within and adjacent to the periaqueductal gray matter, the median raphe was associated with the midline ventral to the dorsal raphe, and the cells of the supralemniscal region were in the tegmentum lateral to the median raphe and ventral to the dorsal raphe. The caudal cluster consisted of three divisions: the raphe obscurus nucleus, the raphe pallidus nucleus and the raphe magnus nucleus. The raphe obscurus nucleus was associated with the dorsal midline at the caudal-most part of the medulla oblongata. The raphe pallidus nucleus was found at the ventral midline of the medulla around the inferior olive. Raphe magnus was associated with the midline of the medulla and was found rostral to both the raphe obscurus and raphe pallidus. The results of our study are compared in an evolutionary context with those reported for other mammals and reptiles. Copyright (C) 2002 S. Karger AG, Basel.

The distribution and morphological characteristics of cholinergic cells in the brain of monotremes as revealed by ChAT immunohistochemistry

The present study employs choline acetyltransferase (ChAT) immunohistochemistry to identify the cholinergic neuronal population in the central nervous system of the monotremes. Two of the three extant species of monotreme were studied: the platypus (Omithorhynchus anatinus) and the short-beaked echidna (Tachyglossus aculeatus). The distribution of cholinergic cells in the brain of these two species was virtually identical. Distinct groups of cholinergic cells were observed in the striatum, basal forebrain, habenula, pontomesencephalon, cranial nerve motor nuclei, and spinal cord. In contrast to other tetrapods studied with this technique, we failed to find evidence for cholinergic cells in the hypothalamus, the parabigeminal nucleus (or nucleus isthmus), or the cerebral cortex. The lack of hypothalamic cholinergic neurons creates a hiatus in the continuous antero-posterior aggregation of cholinergic neurons seen in other tetrapods. This hiatus might be functionally related to the phenomenology of monotreme sleep and to the ontogeny of sleep in mammals, as juvenile placental mammals exhibit a similar combination of sleep elements to that found in adult monotremes. Copyright (C) 2002 S. Karger AG, Basel.

Rates of gene rearrangement and nucleotide substitution are correlated in the mitochondrial genomes of insects

A number of studies indicated that lineages of animals with high rates of mitochondrial (mt) gene rearrangement might have high rates of mt nucleotide substitution. We chose the hemipteroid assemblage and the Insecta to test the idea that rates of mt gene rearrangement and mt nucleotide substitution are correlated. For this purpose, we sequenced the mt genome of a lepidopsocid from the Psocoptera, the only order of hemipteroid insects for which an entire mtDNA sequence is not available. The mt genome of this lepidopsocid is circular, 16,924 bp long, and contains 37 genes and a putative control region; seven tRNA genes and a protein-coding gene in this genome have changed positions relative to the ancestral arrangement of mt genes of insects. We then compared the relative rates of nucleotide substitution among species from each of the four orders of hemipteroid insects and among the 20 insects whose mt genomes have been sequenced entirely. All comparisons among the hernipteroid insects showed that species with higher rates of gene rearrangement also had significantly higher rates of nucleotide substitution statistically than did species with lower rates of gene rearrangement. In comparisons among the 20 insects, where the mt genomes of the two species differed by more than five breakpoints, the more rearranged species always had a significantly higher rate of nucleotide substitution than the less rearranged species. However, in comparisons where the mt genomes of two species differed by five or less breakpoints, the more rearranged species did not always have a significantly higher rate of nucleotide substitution than the less rearranged species. We tested the statistical significance of the correlation between the rates of mt gene rearrangement and mt nucleotide substitution with nine pairs of insects that were phylogenetically independent from one 2 another. We found that the correlation was positive and statistically significant (R-2 = 0.73, P = 0.01; R-s = 0.67, P < 0.05). We propose that increased rates of nucleotide substitution may lead to increased rates of gene rearrangement in the mt genomes of insects.

Synthesis and characterization of delta-atracotoxin-ar1a, the lethal neurotoxin from venom of the Sydney funnel-web spider (Atrax robustus)

delta-Atracotoxin-Ar1a (delta-ACTX-Ar1a) is the major polypeptide neurotoxin isolated from the venom of the male Sydney funnel-web spider, Atrax robustus. This neurotoxin targets both insect and mammalian voltage-gated sodium channels, where it competes with scorpion alpha-toxins for neurotoxin receptor site-3 to slow sodium-channel inactivation. Progress in characterizing the structure and mechanism of action of this toxin has been hampered by the limited supply of pure toxin from natural sources. In this paper, we describe the first successful chemical synthesis and oxidative refolding of the four-disulfide bond containing delta-ACTX-Ar1a. This synthesis involved solid-phase Boc chemistry using double coupling, followed by oxidative folding of purified peptide using a buffer of 2 M GdnHCl and glutathione/glutathiol in a 1:1 mixture of 2-propanol (pH 8.5). Successful oxidation and refolding was confirmed using both chemical and pharmacological characterization. Ion spray mass spectrometry was employed to confirm the molecular weight. H-1 NMR analysis showed identical chemical shifts for native and synthetic toxins, indicating that the synthetic toxin adopts the native fold. Pharmacological studies employing whole-cell patch clamp recordings from rat dorsal root ganglion neurons confirmed that synthetic delta-ACTX-Ar1a produced a slowing of the sodium current inactivation and hyperpolarizing shifts in the voltage-dependence of activation and inactivation similar to native toxin. Under current clamp conditions, we show for the first time that delta-ACTX-Ar1a produces spontaneous repetitive plateau potentials underlying the clinical symptoms seen during envenomation. This successful oxidative refolding of synthetic delta-ACTX-Ar1a paves the way for future structure-activity studies to determine the toxin pharmacophore.