Remarkable consistency of spinal cord microvasculature in highly adapted diving odontocetes.

Odontocetes are breath-hold divers with a suite of physiological, anatomical, and behavioral adaptations that are highly derived and vastly different from those of their terrestrial counterparts. Because of these adaptations for diving, odontocetes were originally thought to be exempt from the harms of nitrogen gas embolism while diving. However, recent studies have shown that these mammals may alter their dive behavior in response to anthropogenic sound, leading to the potential for nitrogen supersaturation and bubble formation which may cause decompression sickness in the central nervous system (CNS). We examined the degree of interface between blood, gases, and neural tissues in the spinal cord by quantifying its microvascular characteristics in five species of odontocetes (Tursiops truncatus, Delphinus delphis, Grampus griseus, Kogia breviceps, and Mesoplodon europaeus) and a model terrestrial species (the pig-Sus scrofa domesticus) for comparison. This approach allowed us to compare microvascular characteristics (microvascular density, branching, and diameter) at several positions (cervical, thoracic, and lumbar) along the spinal cord from odontocetes that are known to be either deep or shallow divers. We found no significant differences (p < 0.05 for all comparisons) in microvessel density (9.30-11.18%), microvessel branching (1.60-2.12 branches/vessel), or microvessel diameter (11.83-16.079 µm) between odontocetes and the pig, or between deep and shallow diving odontocete species. This similarity of spinal cord microvasculature anatomy in several species of odontocetes as compared to the terrestrial mammal is in contrast to the wide array of remarkable physio-anatomical adaptations marine mammals have evolved within their circulatory system to cope with the physiological demands of diving. These results, and other studies on CNS lipids, indicate that the spinal cords of odontocetes do not have specialized features that might serve to protect them from Type II DCS.

Anatomical and hydraulic responses to desiccation in emergent conifer seedlings.

PREMISE: The young seedling life stage is critical for reforestation after disturbance and for species migration under climate change, yet little is known regarding their basic hydraulic function or vulnerability to drought. Here, we sought to characterize responses to desiccation including hydraulic vulnerability, xylem anatomical traits, and impacts on other stem tissues that contribute to hydraulic functioning. METHODS: Larix occidentalis, Pseudotsuga menziesii, and Pinus ponderosa (all ≤6 weeks old) were imaged using x-ray computed microtomography during desiccation to assess seedling biomechanical responses with concurrently measured hydraulic conductivity (ks ) and water potential (Ψ) to assess vulnerability to xylem embolism formation and other tissue damage. RESULTS: In non-stressed samples for all species, pith and cortical cells appeared circular and well hydrated, but they started to empty and deform with decreasing Ψ which resulted in cell tearing and eventual collapse. Despite the severity of this structural damage, the vascular cambium remained well hydrated even under the most severe drought. There were significant differences among species in vulnerability to xylem embolism formation, with 78% xylem embolism in L. occidentalis by Ψ of -2.1 MPa, but only 47.7% and 62.1% in P. ponderosa and P. menziesii at -4.27 and -6.73 MPa, respectively. CONCLUSIONS: Larix occidentalis seedlings appeared to be more susceptible to secondary xylem embolism compared to the other two species, but all three maintained hydration of the vascular cambium under severe stress, which could facilitate hydraulic recovery by regrowth of xylem when stress is relieved.

Six co-occurring conifer species in northern Idaho exhibit a continuum of hydraulic strategies during an extreme drought year.

As growing seasons in the northwestern USA lengthen, on track with climate predictions, the mixed conifer forests that dominate this region will experience extended seasonal drought conditions. The year of 2015, which had the most extreme drought for the area on record, offered a potential analogue of future conditions. During this period, we measured the daily courses of water potential and gas exchange as well as the hydraulic conductivity and vulnerability to embolism of six dominant native conifer species, Abies grandis, Larix occidentalis, Pinus ponderosa, Pinus monticola, Pseudotsuga menziesii and Thuja occidentalis, to determine their responses to 5 months of record-low precipitation. The deep ash-capped soils of the region allowed gas exchange to continue without significant evidence of water stress for almost 2 months after the last rainfall event. Midday water potentials never fell below -2.2 MPa in the evergreen species and -2.7 MPa in the one deciduous species. Branch xylem was resistant to embolism, with P 50 values ranging from -3.3 to -7.0 MPa. Root xylem, however, was more vulnerable, with P 50 values from -1.3 to -4.6 MPa. With predawn water potentials as low as -1.3 MPa, the two Pinus species likely experienced declines in root hydraulic conductivity. Stomatal conductance of all six species was significantly responsive to vapour pressure only in the dry months (August-October), with no response evident in the wet months (June-July). While there were similarities among species, they exhibited a continuum of isohydry and safety margins. Despite the severity of this drought, all species were able to continue photosynthesis until mid-October, likely due to the mediating effects of the meter-deep, ash-capped silty-loam soils with large water storage capacity. Areas with these soil types, which are characteristic of much of the northwestern USA, could serve as refugia under drier and warmer future conditions.

Embracing 3D Complexity in Leaf Carbon-Water Exchange.

Leaves are a nexus for the exchange of water, carbon, and energy between terrestrial plants and the atmosphere. Research in recent decades has highlighted the critical importance of the underlying biophysical and anatomical determinants of CO2 and H2O transport, but a quantitative understanding of how detailed 3D leaf anatomy mediates within-leaf transport has been hindered by the lack of a consensus framework for analyzing or simulating transport and its spatial and temporal dynamics realistically, and by the difficulty of measuring within-leaf transport at the appropriate scales. We discuss how recent technological advancements now make a spatially explicit 3D leaf analysis possible, through new imaging and modeling tools that will allow us to address long-standing questions related to plant carbon-water exchange.

Vascular development in very young conifer seedlings: Theoretical hydraulic capacities and potential resistance to embolism.

PREMISE OF THE STUDY: Conifers have the highest rates of mortality during their first year, often attributed to water stress; yet, this tree life stage is the least studied in terms of hydraulic properties. Previous work has revealed correlations between xylem anatomy to both hydraulic transport capacity and resistance to hydraulic dysfunction. In this study, we compared xylem anatomical and plant functional traits of Pseudotsuga menziesii, Larix occidentalis, and Pinus ponderosa seedlings over the first 10 wk of growth to evaluate potential maximum hydraulic capabilities and resistance to drought-induced embolism. We hypothesized that, based on key functional traits of the xylem, predicted xylem embolism resistance of the species will reflect their previously determined drought tolerances with L. occidentalis, P. menziesii, and P. ponderosa in order of least to most embolism-resistant xylem. METHODS: Xylem and pit anatomical characteristics and additional hydraulic-related functional traits were compared at five times during the first 10 wk of growth using confocal laser scanning microscopy (CLSM). KEY RESULTS: Based on thickness to span ratio, torus to pit aperture overlap, and torus thickness, primary xylem appeared to be not only more hydraulically conductive but also less embolism-resistant than secondary xylem. By week 10, P. menziesii was predicted to have the most embolism-resistant xylem followed by P. ponderosa and L. occidentalis. CONCLUSIONS: Theoretical measurements suggest that hydraulic transport capacities and vulnerability to embolism varied for each species over the first 10 wk of growth; thus, the timing of germination and onset of limited soil moisture is critical for growth and survival of seedlings.

Quantitative assessment of angiogenic responses by the directed in vivo angiogenesis assay.

One of the major problems in angiogenesis research remains the lack of suitable methods for quantifying the angiogenic response in vivo. We describe the development and application of the directed in vivo angiogenesis assay (DIVAA) and demonstrated that it is reproducible and quantitative. This assay consists of subcutaneous implantation of semiclosed silicone cylinders (angioreactors) into nude mice. Angioreactors are filled with only 18 micro l of extracellular matrix premixed with or without angiogenic factors. Vascularization within angioreactors is quantified by the intravenous injection of fluorescein isothiocyanate (FITC)-dextran before their recovery, followed by spectrofluorimetry. Angioreactors examined by immunofluorescence show cells and invading angiogenic vessels at different developmental stages. The minimally detectable angiogenic response requires 9 days after implantation and >/=50 ng/ml (P < 0.01) of either fibroblast growth factor-2 or vascular endothelial growth factor. Characterization of this assay system demonstrates that the FITC-labeled dextran quantitation is highly reproducible and that levels of FITC-dextran are not significantly influenced by vascular permeability. DIVAA allows accurate dose-response analysis and identification of effective doses of angiogenesis-modulating factors in vivo. TNP-470 potently inhibits angiogenesis (EC(50) = 88 pmol/L) induced by 500 ng/ml of fibroblast growth factor-2. This inhibition correlates with decreased endothelial cell invasion. DIVAA efficiently detects differences in anti-angiogenic potencies of thrombospondin-1 peptides (25 micro mol/L) and demonstrates a partial inhibition of angiogenesis ( approximately 40%) in a matrix metalloprotease (MMP)-2-deficient mouse compared with that in wild-type animals. Zymography of angioreactors from MMP-deficient and control animals reveals quantitative changes in MMP expression. These results support DIVAA as an assay to compare potencies of angiogenic factors or inhibitors, and for profiling molecular markers of angiogenesis in vivo.