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Stires, JC, Latz MI.  2018.  Contribution of the cytoskeleton to mechanosensitivity reported by dinoflagellate bioluminescence. Cytoskeleton. 75:12-21.   10.1002/cm.21392   AbstractWebsite

The cytoskeleton is crucial to cell mechanics and sensing the extracellular physical environment. The objective of this study was to examine the role of the cortical cytoskeleton in mechanosensitivity in a unicellular protist, the marine dinoflagellate Lingulodinium polyedra, using its intrinsic bioluminescence as a rapid reporter of mechanotransduction. Pharmacological treatments resolved effects due to immediate cytoskeleton disruption from those due to cytoskeletal remodeling during the light to dark phase transition. The cytoskeleton was visualized by confocal laser scanning microscopy of immunohistochemically labeled microtubules and phalloidin labeled F-actin, and mechanosensitivity assessed based on the bioluminescence response to mechanical stimulation measured during the dark phase. Latrunculin B treatment after the transition from the light to dark phase resulted in some disruption of cortical F-actin, no observed effect on the cortical microtubules, and partial inhibition of the bioluminescence response. Treatment with oryzalin, which depolarizes microtubules, completely disrupted the microtubule network and cortical F-actin, and partially inhibited bioluminescence. These results demonstrate that cells retain some mechanosensitivity despite a disrupted cytoskeleton; link mechanosensitivity to intact F-actin; show a close connection between F-actin and microtubules comprising the cortical cytoskeleton; confirm a strong contribution of the actin cytoskeleton to the translocation of scintillons, vesicles containing the luminescent chemistry; and support the role of the actin cytoskeleton in the association of scintillons with the vacuole membrane.

Lindstrom, JB, Pierce NT, Latz MI.  2017.  Role of TRP Channels in dinoflagellate mechanotransduction. Biological Bulletin. 233:151-167.   10.1086/695421   AbstractWebsite

Transient receptor potential (TRP) ion channels are common components of mechanosensing pathways, mainly described in mammals and other multicellular organisms. To gain insight into the evolutionary origins of eukaryotic mechanosensory proteins, we investigated the involvement of TRP channels in mechanosensing in a unicellular eukaryotic protist, the dinoflagellate Lingulodiniumpolyedra. BLASTPanalysis of the protein sequences predicted from the L. polyedra transcriptome revealed six sequences with high similarity to human TRPM2, TRPM8, TRPML2, TRPP1, and TRPP2; and characteristic TRP domains were identified in all sequences. In a phylogenetic tree including all mammalian TRP subfamilies and TRP channel sequences from unicellular and multicellular organisms, the L. polyedra sequences grouped with the TRPM, TPPML, and TRPP clades. In pharmacological experiments, we used the intrinsic bioluminescence of L. polyedra as a reporter of mechanoresponsivity. Capsaicin and RN1734, agonists of mammalian TRPV, and arachidonic acid, an agonist of mammalian TRPV, TRPA, TRPM, and Drosophila TRP, all stimulated bioluminescence in L. polyedra. Mechanical stimulation of bioluminescence, but not capsaicinstimulated bioluminescence, was inhibited by gadolinium (Gd3+), a general inhibitor of mechanosensitive ion channels, and the phospholipase C (PLC) inhibitor U73122. These pharmacological results are consistent with the involvement of TRP-like channels in mechanosensing by L. polyedra. The TRP channels do not appear to be mechanoreceptors but rather are components of the mechanotransduction signaling pathway and may be activated via a PLC-dependent mechanism. The presence and function of TRP channels in a dinoflagellate emphasize the evolutionary conservation of both the channel structures and their functions.

Deane, GB, Stokes DM, Latz MI.  2016.  Bubble stimulation efficiency of dinoflagellate bioluminescence. Luminescence. 31:270-280.   10.1002/bio.2957   Abstract

Dinoflagellate bioluminescence, a common source of bioluminescence in coastal waters, is stimulated by flow agitation. Although bubbles are anecdotally known to be stimulatory, the process has never been experimentally investigated. This study quantified the flash response of the bioluminescent dinoflagellate Lingulodinium polyedrum to stimulation by bubbles rising through still seawater. Cells were stimulated by isolated bubbles of 0.3–3 mm radii rising at their terminal velocity, and also by bubble clouds containing bubbles of 0.06–10 mm radii for different air flow rates. Stimulation efficiency, the proportion of cells producing a flash within the volume of water swept out by a rising bubble, decreased with decreasing bubble radius for radii less than approximately 1 mm. Bubbles smaller than a critical radius in the range 0.275–0.325 mm did not stimulate a flash response. The fraction of cells stimulated by bubble clouds was proportional to the volume of air in the bubble cloud, with lower stimulation levels observed for clouds with smaller bubbles. An empirical model for bubble cloud stimulation based on the isolated bubble observations successfully reproduced the observed stimulation by bubble clouds for low air flow rates. High air flow rates stimulated more light emission than expected, presumably because of additional fluid shear stress associated with collective buoyancy effects generated by the high air fraction bubble cloud. These results are relevant to bioluminescence stimulation by bubbles in two-phase flows, such as in ship wakes, breaking waves, and sparged bioreactors. Copyright © 2015 John Wiley & Sons, Ltd.

Tesson, B, Latz MI.  2015.  Mechanosensitivity of a rapid bioluminescence reporter system assessed by atomic force microscopy. Biophysical Journal. 108:1341-1351.   10.1016/j.bpj.2015.02.009   AbstractWebsite

Cells are sophisticated integrators of mechanical stimuli that lead to physiological, biochemical, and genetic responses. The bioluminescence of dinoflagellates, alveolate protists that use light emission for predator defense, serves as a rapid noninvasive whole-cell reporter of mechanosensitivity. In this study, we used atomic force microscopy (AFM) to explore the relationship between cell mechanical properties and mechanosensitivity in live cells of the dinoflagellate Pyrocystis lunula. Cell stiffness was 0.56 MPa, consistent with cells possessing a cell wall. Cell response depended on both the magnitude and velocity of the applied force. At the maximum stimulation velocity of 390 mu m s(-1), the threshold response occurred at a force of 7.2 mu N, resulting in a contact time of 6.1 ms and indentation of 2.1 mu m. Cells did not respond to a low stimulation velocity of 20 mu m s(-1), indicating a velocity dependent response that, based on stress relaxation experiments, was explained by the cell viscoelastic properties. This study demonstrates the use of AFM to study mechanosensitivity in a cell system that responds at fast timescales, and provides insights into how viscoelastic properties affect mechanosensitivity. It also provides a comparison with previous studies using hydrodynamic stimulation, showing the discrepancy in cell response between direct compressive forces using AFM and those within flow fields based on average flow properties.

Latz, MI, Rohr J.  2013.  Bathyphotometer bioluminescence potential measurements: A framework for characterizing flow agitators and predicting flow-stimulated bioluminescence intensity. Continental Shelf Research. 61–62:71-84.   10.1016/j.csr.2013.04.033   AbstractWebsite

Bathyphotometer measurements of bioluminescence are used as a proxy for the abundance of luminescent organisms for studying population dynamics; the interaction of luminescent organisms with physical, chemical, and biological oceanographic processes; and spatial complexity especially in coastal areas. However, the usefulness of bioluminescence measurements has been limited by the inability to compare results from different bathyphotometer designs, or even the same bathyphotometer operating at different volume flow rates. The primary objective of this study was to compare measurements of stimulated bioluminescence of four species of cultured dinoflagellates, the most common source of bioluminescence in coastal waters, using two different bathyphotometer flow agitators as a function of bathyphotometer volume flow rate and dinoflagellate concentration. For both the NOSC and BIOLITE flow agitators and each species of dinoflagellate tested, there was a critical volume flow rate, above which average bioluminescence intensity, designated as bathyphotometer bioluminescence potential (BBP), remained relatively constant and scaled directly with dinoflagellate cell concentration. At supra-critical volume flow rates, the ratio of BIOLITE to NOSC BBP was nearly constant for the same species studied, but varied between species. The spatial pattern and residence time of flash trajectories within the NOSC flow agitator indicated the presence of dominant secondary recirculating flows, where most of the bioluminescence was detected. A secondary objective (appearing in the Appendix) was to study the feasibility of using NOSC BBP to scale flow-stimulated bioluminescence intensity across similar flow fields, where the contributing composition of luminescent species remained the same. Fully developed turbulent pipe flow was chosen because it is hydrodynamically well characterized. Average bioluminescence intensity in a 2.54-cm i.d. pipe was highly correlated with wall shear stress and BBP. This correlation, when further scaled by pipe diameter, effectively predicted bioluminescence intensity in fully developed turbulent flow in a 0.83-cm i.d. pipe. Determining similar correlations between other bathyphotometer flow agitators and flow fields will allow bioluminescence potential measurements to become a more powerful tool for the oceanographic community.

Jin, K, Klima JC, Deane G, Dale Stokes M, Latz MI.  2013.  Pharmacological investigation of the bioluminescence signaling pathway of the dinoflagellate Lingulodinium polyedrum: evidence for the role of stretch-activated ion channels. Journal of Phycology. 49:733-745.   10.1111/jpy.12084   AbstractWebsite

Dinoflagellate bioluminescence serves as a whole-cell reporter of mechanical stress, which activates a signaling pathway that appears to involve the opening of voltage-sensitive ion channels and release of calcium from intracellular stores. However, little else is known about the initial signaling events that facilitate the transduction of mechanical stimuli. In the present study using the red tide dinoflagellate Lingulodinium polyedrum (Stein) Dodge, two forms of dinoflagellate bioluminescence, mechanically stimulated and spontaneous flashes, were used as reporter systems to pharmacological treatments that targeted various predicted signaling events at the plasma membrane level of the signaling pathway. Pretreatment with 200 μM Gadolinium III (Gd3+), a nonspecific blocker of stretch-activated and some voltage-gated ion channels, resulted in strong inhibition of both forms of bioluminescence. Pretreatment with 50 μM nifedipine, an inhibitor of L-type voltage-gated Ca2+ channels that inhibits mechanically stimulated bioluminescence, did not inhibit spontaneous bioluminescence. Treatment with 1 mM benzyl alcohol, a membrane fluidizer, was very effective in stimulating bioluminescence. Benzyl alcohol-stimulated bioluminescence was inhibited by Gd3+ but not by nifedipine, suggesting that its role is through stretch activation via a change in plasma membrane fluidity. These results are consistent with the presence of stretch-activated and voltage-gated ion channels in the bioluminescence mechanotransduction signaling pathway, with spontaneous flashing associated with a stretch-activated component at the plasma membrane.

Maldonado, EM, Latz MI.  2011.  Species-specific effects of fluid shear on grazing by sea urchin larvae: comparison of experimental results with encounter-model predictions. Marine Ecology-Progress Series. 436:119-130.   10.3354/meps09249   AbstractWebsite

Small-scale turbulence can alter the rate of plankton predator-prey encounters. Encounter models predict that prey ingestion by slow-swimming zooplankton is enhanced at low levels of turbulence. We investigated whether small-scale turbulence increases ingestion for the slow-swimming, suspension-feeding pluteus larvae of the white urchin Lytechinus pictus and the purple urchin Strongylocentrotus purpuratus. Model predictions of the critical level of turbulence, epsilon(cr), above which encounters due to turbulence are greater than those due to behavior (swimming or suspension feeding) alone, were experimentally tested using short-and long-term grazing treatments. Because urchin larvae are smaller than the smallest eddy scales of turbulence and thus experience turbulence as laminar shear, larvae were exposed to flow conditions using a simple laminar shear flow with dissipation rates, epsilon, of 0, 0.1, 0.4, and 1 cm(2) s(-3). Short-term ingestion of beads by L. pictus larvae was unaffected by epsilon < 1 cm(2) s(-3) but was 30% greater at this level, which was greater than ecr based on flow speeds produced in suspension feeding. Long-term flow treatments with algal prey had no significant effect on grazing or growth. Short-term ingestion of beads by S. purpuratus larvae was unaffected by epsilon < epsilon(cr) based on suspension feeding; the effect of long-term flow exposure on ingestion and growth could not be investigated because of high mortality, suggesting greater sensitivity to flow exposure compared to L. pictus. Experimental results are consistent with model predictions that ecr is high, and thus levels of turbulence in the ocean are not expected to significantly increase ingestion and reduce food limitation in suspension-feeding urchin larvae.

Roth, MS, Latz MI, Goericke R, Deheyn DD.  2010.  Green fluorescent protein regulation in the coral Acropora yongei during photoacclimation. Journal of Experimental Biology. 213:3644-3655.   10.1242/jeb.040881   AbstractWebsite

Reef-building corals inhabit high light environments and are dependent on photosynthetic endosymbiotic dinoflagellates for nutrition. While photoacclimation responses of the dinoflagellates to changes in illumination are well understood, host photoacclimation strategies are poorly known. This study investigated fluorescent protein expression in the shallow-water coral Acropora yongei during a 30 day laboratory photoacclimation experiment in the context of its dinoflagellate symbionts. Green fluorescent protein (GFP) concentration measured by Western blotting changed reversibly with light intensity. The first 15 days of the photoacclimation experiment led to a similar to 1.6 times increase in GFP concentration for high light corals (900 mu mol quanta m(-2) s(-1)) and a similar to 4 times decrease in GFP concentration for low light corals (30 mu mol quanta m(-2) s(-1)) compared with medium light corals (300 mu mol quanta m(-2) s(-1)). Green fluorescence increased similar to 1.9 times in high light corals and decreased similar to 1.9 times in low light corals compared with medium light corals. GFP concentration and green fluorescence intensity were significantly correlated. Typical photoacclimation responses in the dinoflagellates were observed including changes in density, photosynthetic pigment concentration and photosynthetic efficiency. Although fluorescent proteins are ubiquitous and abundant in scleractinian corals, their functions remain ambiguous. These results suggest that scleractinian corals regulate GFP to modulate the internal light environment and support the hypothesis that GFP has a photoprotective function. The success of photoprotection and photoacclimation strategies, in addition to stress responses, will be critical to the fate of scleractinian corals exposed to climate change and other stressors.

Latz, MI, Allen J, Sarkar S, Rohr J.  2009.  Effect of fully characterized unsteady flow on population growth of the dinoflagellate Lingulodinium polyedrum. Limnology and Oceanography. 54:1243-1256.   10.4319/lo.2009.54.4.1243   AbstractWebsite

Dinoflagellate population growth is inhibited by fluid motion, which is typically characterized by some average flow property, regardless if the flow is steady or unsteady. This study compares the effect of fully characterized steady and unsteady flow on net population growth of the red tide dinoflagellate Lingulodinium polyedrum. The unsteady flow fields were generated using oscillatory laminar Couette flow and characterized analytically to provide complete knowledge of the fluid shear exposure over space and time throughout the chamber. Experimental conditions were selected so all cells experienced a similar shear exposure regardless of their position within the chamber. Unsteady flow with maximum shears of 6.4 s(-1) and 6.7 s(-1) and an average absolute shear of 4 s(-1), comparable with levels found at the ocean surface on a windy day, resulted in higher levels of growth inhibition than for steady Couette flow with shears of 4 and 8 s(-1). Over the parameter space studied, growth inhibition increased with increasing treatment duration (5-120 min) but was insensitive to oscillation period (60-600 s) or whether the unsteady flow changed in direction. These results indicate that over the parameter space studied, unsteady flow is more inhibitory to net growth than steady flow, for the same average flow conditions, and demonstrate that flow characterization on the basis only of average flow properties is inadequate for comparing population growth in unsteady and steady flows.

Deheyn, DD, Latz MI.  2009.  Internal and secreted bioluminescence of the marine polychaete Odontosyllis phosphorea (Syllidae). Invertebrate Biology. 128:31-45.   10.1111/j.1744-7410.2008.00149.x   AbstractWebsite

The syllid polychaete Odontosyllis phosphorea produces brilliant displays of green bioluminescence during mating swarms. We studied freshly collected individuals of O. phosphorea in the laboratory to understand the characteristics of its luminescent system. Light emission appeared as an intense glow after stimulation with potassium chloride, and was associated with secreted mucus. The mucus was viscous, blue in color, and exhibited a long-lasting glow that was greatly intensified by addition of peroxidase or ammonium persulfate. The emission spectrum of mucus-associated bioluminescence was unimodal, with a maximum emission in the green spectrum between 494 and 504 nm. The fluorescence emission spectrum was similar, but the fluorescence intensity was low unless it originated from mucus that had already produced light, suggesting that the oxidized product of the light production is the source of fluorescence. Individuals as small as 0.5-1.0 mm produced bioluminescence that was mainly internal and not secreted as mucus. The early occurrence of bioluminescence in the life cycle of members of O. phosphorea suggests that bioluminescence may be used for purposes other than attracting mates. The luminous system was functional at temperatures as low as -20 degrees C and was degraded above 40 degrees C. Mixing hot and cold extracts of the mucus did not result in reconstituting original levels of light emission. Additionally, mucus samples exposed to oxygen depletion by bubbling with argon or nitrogen were still able to produce intense bioluminescence. These results suggest that bioluminescence from the mucus may involve a photoprotein rather than a luciferin-luciferase reaction.

Latz, MI, Bovard M, VanDelinder V, Segre E, Rohr J, Groisman A.  2008.  Bioluminescent response of individual dinoflagellate cells to hydrodynamic stress measured with millisecond resolution in a microfluidic device. Journal of Experimental Biology. 211:2865-2875.   10.1242/jeb.011890   AbstractWebsite

Dinoflagellate bioluminescence serves as a model system for examining mechanosensing by suspended motile unicellular organisms. The response latency, i.e. the delay time between the mechanical stimulus and luminescent response, provides information about the mechanotransduction and signaling process, and must be accurately known for dinoflagellate bioluminescence to be used as a flow visualization tool. This study used a novel microfluidic device to measure the response latency of a large number of individual dinoflagellates with a resolution of a few milliseconds. Suspended cells of several dinoflagellate species approximately 35 mu m in diameter were directed through a 200 mu m deep channel to a barrier with a 15 mu m clearance impassable to the cells. Bioluminescence was stimulated when cells encountered the barrier and experienced an abrupt increase in hydrodynamic drag, and was imaged using high numerical aperture optics and a high-speed low-light video system. The average response latency for Lingulodinium polyedrum strain HJ was 15ms (N>300 cells) at the three highest flow rates tested, with a minimum latency of 12 ms. Cells produced multiple flashes with an interval as short as 5 ms between individual flashes, suggesting that repeat stimulation involved a subset of the entire intracellular signaling pathway. The mean response latency for the dinoflagellates Pyrodinium bahamense, Alexandrium monilatum and older and newer isolates of L. polyedrum ranged from 15 to 22 ms, similar to the latencies previously determined for larger dinoflagellates with different morphologies, possibly reflecting optimization of dinoflagellate bioluminescence as a rapid anti-predation behavior.

Deheyn, DD, Latz MI.  2007.  Bioluminescence characteristics of a tropical terrestrial fungus (Basidiomycetes). Luminescence. 22:462-467.   10.1002/bio.985   AbstractWebsite

Freshly collected samples of luminous mycelium of a terrestrial fungus from Panama were investigated for their bioluminescence characteristics. Taxonomic identification of fungal species could not be determined because of the lack of fruiting bodies. Fluorescence excited by 380 nm illumination had an emission spectrum with a main peak at 480 nm and additional chlorophyll peaks related to the wood substrate. Bioluminescence appeared as a continuous glow that did not show any diel variation. The light production was sensitive to temperature and decreased with temperatures higher or lower than ambient. Bioluminescence intensity was sensitive to hydration, increasing by a factor of 400 immediately after exposure to water and increasing by a factor of 1 million after several hours. This increase may have occurred through dilution of superoxide dismutase, which is a suppressive factor of bioluminescence in fungus tissue. The mycelium typically transports nutritive substances back to the fruiting body. The function of luminescent mycelium may be to increase the intensity of light from the fungus and more effectively attract nocturnal insects and other animals that serve as disseminating vectors for fungal spores. Copyright (C) 2007 John Wiley & Sons, Ltd.

Chen, AK, Latz MI, Sobolewski P, Frangos JA.  2007.  Evidence for the role of G-proteins in flow stimulation of dinoflagellate bioluminescence. American Journal of Physiology-Regulatory Integrative and Comparative Physiology. 292:R2020-R2027.   10.1152/ajpregu.00649.2006   AbstractWebsite

Luminescent dinoflagellates respond to flow by the production of light. The primary mechanotransduction event is unknown, although downstream events include a calcium flux in the cytoplasm, a self-propagating action potential across the vacuole membrane, and a proton flux into the cytoplasm that activates the luminescent chemistry. Given the role of GTP-binding (G) proteins in the mechanotransduction of flow by nonmarine cells and the presence of G-proteins in dinoflagellates, it was hypothesized that flow-stimulated dinoflagellate bioluminescence involves mechanotransduction by G-proteins. In the present study, osmotic swelling of cells of the dinoflagellate Lingulodinium polyedrum was used as a drug delivery system to introduce GDP beta S, an inhibitor of G-protein activation. Osmotically swollen cells produced higher levels of flow-stimulated bioluminescence at a lower threshold of shear stress, indicating they were more flow sensitive. GDP beta S inhibited flow-stimulated bioluminescence in osmotically swollen cells and in cells that were restored to the isosmotic condition following hypoosmotic treatment with GDP beta S. These results provide evidence that G-proteins are involved in the mechanotransduction of flow in dinoflagellates and suggest that G-protein involvement in mechanotransduction may be a fundamental evolutionary adaptation.

Maldonado, EM, Latz MI.  2007.  Shear-stress dependence of dinoflagellate bioluminescence. Biological Bulletin. 212:242-249. AbstractWebsite

Fluid flow stimulates bioluminescence in dinoflagellates. However, many aspects of the cellular mechanotransduction are incompletely known. The objective of our study was to formally test the hypothesis that flow-stimulated dinoflagellate bioluminescence is dependent on shear stress, signifying that organisms are responding to the applied fluid force. The dinoflagellate Lingulodinium polyedrum was exposed to steady shear using simple Couette flow in which fluid viscosity was manipulated to alter shear stress. At a constant shear rate, a higher shear stress due to increased viscosity increased both bioluminescence intensity and decay rate, supporting our hypothesis that bioluminescence is shear-stress dependent. Although the flow response of non-marine attached cells is known to be mediated through shear stress, our results indicate that suspended cells such as dinoflagellates also sense and respond to shear stress. Shear-stress dependence of flowstimulated bioluminescence in dinoflagellates is consistent with mechanical stimulation due to direct predator handling in the context of predator-prey interactions.

Deheyn, DD, Latz MI.  2006.  Bioavailability of metals along a contamination gradient in San Diego Bay (California, USA). Chemosphere. 63:818-834.   10.1016/j.chemosphere.2005.07.066   AbstractWebsite

San Diego Bay is heavily contaminated with metals, but little is known about their biological availability to local marine organisms. This study on 15 elements showed that concentrations of metals associated with sediment increased from the mouth to the back of the Bay while metals in seawater particulates were similar throughout the Bay. Metal bioavailability was assessed over 8 weeks following transplant of the local brittlestar, Ophiothrix spiculata (Ophuroidea, Echinodermata), from outside to inside the Bay. Despite a gradient of contamination, brittlestars accumulated similar levels of metals throughout the Bay, suggesting that metal contamination occurred through dissolved metals as well as through the diet. Sediment transplanted in dialysis tubing in the Bay accumulated metals only when placed on the seafloor bottom, indicating greater metal bioavailability near the bottom; the level of accumulation was similar between the mouth and the back of the Bay. The results are consistent with a circulation pattern in which a bottom layer of seawater, enriched with metals, drains from the back to the mouth of the Bay. There was a positive correlation between metal concentration in brittlestars and tidal range, suggesting increased metal exposure due to bay-ocean water exchange. For brittlestar arms the correlation was higher at the mouth than the back of the Bay, indicating greater metal accumulation in arms from dissolved metals in seawater than from ingestion of metal contaminated diet. In contrast, for brittlestar disks the correlation was higher at the back of the Bay, indicative of metal accumulation mainly through the diet. The results highlight the importance of considering bioavailability and physical processes in environmental quality assessments. (c) 2005 Elsevier Ltd. All rights reserved.

von Dassow, P, Bearon RN, Latz MI.  2005.  Bioluminescent response of the dinoflagellate Lingulodinium polyedrum to developing flow: Tuning of sensitivity and the role of desensitization in controlling a defensive behavior of a planktonic cell. Limnology and Oceanography. 50:607-619. AbstractWebsite

Dinoflagellate bioluminescence is believed to serve a defensive function, decreasing grazing at night. Previous characterization of bioluminescence stimulated by fully developed flows might have underestimated the true sensitivity of bioluminescence by not observing the initial response. Also, it has been suggested that bioluminescence may be more sensitive to time-varying flow than to constant flow conditions. We used developing laminar Couette flow to characterize the sensitivity of the initial bioluminescent response of the dinoflagellate Lingulodinium polyedrum in time-varying flow. Both the absolute sensitivity (threshold) and dynamic sensitivity were consistent with that determined previously in fully developed flows, although there were differences between different cultured isolates of the same species and between those isolates and cells harvested from a unialgal bloom of the same species. When the rate of increase of shear was varied while keeping the maximum shear level similar, the threshold was independent of the rate of increase of shear. Surprisingly, the integrated bioluminescence was strongly dependent on the rate of increase of shear. The mechanism behind the preferential response to rapidly increasing shear was determined to be desensitization. Desensitization may influence which naturally occurring flows strongly stimulate bioluminescence either by allowing cells to avoid producing a primary response in certain slowly changing flows or, more generally, to avoid the cost of repeated stimulation when entrained in environmental flows containing above-threshold shears.

Deheyn, DD, Gendreau P, Baldwin RJ, Latz MI.  2005.  Evidence for enhanced bioavailability of trace elements in the marine ecosystem of Deception Island, a volcano in Antarctica. Marine Environmental Research. 60:1-33.   10.1016/j.marenvres.2004.08.001   AbstractWebsite

This study assessed whether trace elements present at Deception Island, an active submarine volcano in the Antarctic Peninsula, show enhanced biological availability to the local marine community. Using a weak acid extraction method to dissolve organic material and leach associated but not constitutive trace elements of sediments, fifteen elements were measured from seafloor sediment, seawater particulates, and tissues of benthic (bivalves, brittlestars, sea urchins) and pelagic (demersal and pelagic fishes, krill) organisms collected in the flooded caldera. The highest element concentrations were associated with scafloor sediment, the lowest with seawater particulates and organism tissues. In the case of Ag and Se, concentrations were highest in organism tissue, indicating contamination through the food chain and biomagnification of those elements. The elements Al, Fe, Mn, Sr, Ti, and to a lesser extent Zn, were the most concentrated of the trace elements for all sample types. This indicates that the whole ecosystem of Deception Island is contaminated with trace elements from local geothermal activity, which is also reflected in the pattern of element contamination in organisms. Accordingly, element concentrations were higher in organisms collected at Deception Island compared to those from the neighboring non-active volcanic King George Island, suggesting that volcanic activity enhances bioavailability of trace elements to marine organisms. Trace

Latz, MI, Juhl AR, Ahmed AM, Elghobashi SE, Rohr J.  2004.  Hydrodynamic stimulation of dinoflagellate bioluminescence: a computational and experimental study. Journal of Experimental Biology. 207:1941-1951.   10.1242/jeb.00973   AbstractWebsite

Dinoflagellate bioluminescence provides a near-instantaneous reporter of cell response to flow. Although both fluid shear stress and acceleration are thought to be stimulatory, previous studies have used flow fields dominated by shear. In the present study, computational and experimental approaches were used to assess the relative contributions to bioluminescence stimulation of shear stress and acceleration in a laminar converging nozzle. This flow field is characterized by separate regions of pronounced acceleration away from the walls, and shear along the wall. Bioluminescence of the dinoflagellates Lingulodinium polyedrum and Ceratocorys horrida, chosen because of their previously characterized different flow sensitivities, was imaged with a low-light video system. Numerical simulations were used to calculate the position of stimulated cells and the levels of acceleration and shear stress at these positions. Cells were stimulated at the nozzle throat within the wall boundary layer where, for that downstream position, shear stress was relatively high and acceleration relatively low. Cells of C. horrida were always stimulated significantly higher in the flow field than cells of L. polyedrum and at lower flow rates, consistent with their greater flow sensitivity. For both species, shear stress levels at the position of stimulated cells were similar to but slightly greater than previously determined response thresholds using independent flow fields. L polyedrum did not respond in conditions where acceleration was as high as 20 g. These results indicate that shear stress, rather than acceleration, was the stimulatory component of flow. Thus, even in conditions of high acceleration, dinoflagellate bioluminescence is an effective marker of shear stress.

Deheyn, DD, Bencheikh-Latmani R, Latz MI.  2004.  Chemical speciation and toxicity of metals assessed by three bioluminescence-based assays using marine organisms. Environmental Toxicology. 19:161-178.   10.1002/tox.20009   AbstractWebsite

Metal toxicity is a function of the biology of the target organism and the chemical speciation of the metal. The toxicity of 11 metals was assessed with three cell-based bioassays based on marine organisms: the bacterium Photobacterium phosphoreum of the Microtox(R) bioassay, an environmental strain of P. phosphoreum, and photocytes isolated from the brittlestar Ophiopsila californica. Metal speciation was calculated for three commonly used media: NaCl-based Microtox(R) bioassay medium, artificial seawater glycerol, and artificial seawater. Decreased bioluminescence was considered a proxy for cell toxicity. In all three assays the elements Cd and Hg exhibited similar speciation as well as similar toxicity profiles. The element Cu was toxic in all three assays despite different metal speciation for the P. phosphoreum bioassay. The element Ag was toxic to both bacterial strains but not to photocytes despite a similar chemical speciation for all three assays. In general, the Microtox(R) bioassay was sensitive to all metals (except Pb), whereas the photocytes were the least sensitive to the metals. The heightened response of the Microtox(R) bioassay probably resulted from a combination of the limited complexing power of the medium and the greater sensitivity of the bacterial strain. (C) 2004 Wiley Periodicals, Inc.

Stokes, MD, Deane GB, Latz MI, Rohr J.  2004.  Bioluminescence imaging of wave-induced turbulence. Journal of Geophysical Research-Oceans. 109   10.1029/2003jc001871   AbstractWebsite

[1] The ability to measure turbulent processes on small spatial and temporal scales is a long standing problem in physical oceanography. Here we explore a novel means of measuring fluid shear stress using the cell flashing behavior of bioluminescent dinoflagellates. To illustrate this technique, we present estimates of the heterogeneous, time-varying shear stress inside a breaking wave crest. These results have implications for a better understanding of upper ocean wave physics, air-sea gas transfer, and the biology of planktonic near-surface organisms as well as providing a new quantitative fluid visualization tool.

Latz, MI, Nauen JC, Rohr J.  2004.  Bioluminescence response of four species of dinoflagellates to fully developed pipe flow. Journal of Plankton Research. 26:1529-1546.   10.1093/plankt/fbh141   AbstractWebsite

Dinoflagellate bioluminescence provides a nearly instantaneous index of flow sensitivity. This study compared flow sensitivity in four species of morphologically diverse luminescent dinoflogellates (Ceratium fusus, Ceratocorys horrida, Lingulodinium, poyledrum and Pyrocystis fusiformis) using fully, developed lam mar and turbulent pipe flow. Bioliuminescence response thresholds always occurred in 2 laminar flows with wall shear stress levels that, depending on species, ranged from 0.02 to 0.3 N m(-2). With few exceptions, such as breaking waves and wave-forced bottom shears in shallow nearshore areas, these threshold shear stress levels are several orders of magnitude larger than typical oceanic ambient flows. For laminar flows above threshold, species also differed in. the proportion of organisms responding and the minimum, shear stress level where individual flashes reached their highest intensity. Following transition to turbulent flow, there was never a dramatic increase in bioluminescence, even when energetic turbulent length scales were similar to the cell size. On the basis of their bioluminescence response in laminar flow, these species were ranked in order of decreasing sensitivity as C. horrida > P fusiformis > C. fusus > L. polyedrum. This ranking, though not conclusive, is consistent with increased flow sensitivity due to increasing size and the presence of spines. With the exception of a small fraction of the C. horrida population that is sensitive enough to flash within the feeding current of a predator, the present serve as an study suggests that flashes only occur with Predator contact. Nevertheless flow sensitivity index of the response to mechanical agitation during predator contact/handling. Flow sensitivity may be constrained to maximize the response to Predator contact/handling while minimizing stimulation by background oceanic flows to avoid depleting luminescent reserves.

Chen, AK, Latz MI, Frangos JA.  2003.  The use of dinoflagellate bioluminescence to characterize cell stimulation in bioreactors. Biotechnology and Bioengineering. 83:93-103.   10.1002/bit.10647   AbstractWebsite

Bioluminescent dinoflagellates are flow-sensitive marine organisms that produce light emission almost instantaneously upon stimulation by fluid shear in a shear stress dose-dependent manner. In the present study we tested the hypothesis that monitoring bioluminescence by suspended dinoflagellates can be used as a tool to characterize cellular response to hydrodynamic forces in agitated bioreactors. Specific studies were performed to determine: (1) impeller configurations with minimum cell activation, (2) correlations of cellular response and an integrated shear factor, and (3) the effect of rapid acceleration in agitation. Results indicated that (1) at a volumetric mass transfer coefficient of 3 x 10(-4) s(-1), marine impeller configurations were less stimulatory than Rushton configurations, (2) bioluminescence response and a modified volumetric integrated shear factor had an excellent correlation, and (3) rapid acceleration in agitation was highly stimulatory, suggesting a profound effect of temporal gradients in shear in increasing cell stimulation. By using bioluminescence stimulation as an indicator of agitation-induced cell stimulation and/or damage in microcarrier cultures, the present study allows for the verification of hypotheses and development of novel mechanisms of cell damage in bioreactors. (C) 2003 Wiley Periodicals, Inc.

von Dassow, P, Latz MI.  2002.  The role of Ca2+ in stimulated bioluminescence of the dinoflagellate Lingulodinium polyedrum. Journal of Experimental Biology. 205:2971-2986. AbstractWebsite

Many marine dinoflagellates emit bright discrete flashes of light nearly instantaneously in response to either laminar or turbulent flows as well as to direct mechanical stimulation. The Hash involves a unique pH-dependent luciferase and a proton-mediated action potential across the vacuole membrane. The mechanotransduction process initiating this action potential is unknown. The present study investigated the role of Ca2+ in the mechanotransduction process regulating bioluminescence in the dinoflagellate Lingulodinium polyedrum. Calcium ionophores and digitonin stimulated luminescence in a Ca2+-dependent manner in the absence of mechanical stimulation. Mechanically sensitive luminescence was strongly inhibited by the intracellular Ca2+ chelator BAPTA-AM [1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester]; there was only a partial and irreversible dependence on extracellular Ca2+. Ruthenium Red, a blocker of intracellular Ca2+ release channels, inhibited mechanically sensitive luminescence. Luminescence was also stimulated by increasing K+, even in the absence of extracellular Ca2+; K+ stimulation was inhibited both by BAPTA-AM and Ruthenium Red. These results support the hypothesis that Ca2+ mediates stimulated bioluminescence and also indicate the involvement of intracellular Ca2+ stores. Rapid coupling between mechanical stimulation and mobilization of intracellular Ca2+ stores might occur through a mechanism similar to excitation-contraction coupling in skeletal muscle.

Rohr, J, Hyman M, Fallon S, Latz MI.  2002.  Bioluminescence flow visualization in the ocean: an initial strategy based on laboratory experiments. Deep-Sea Research Part I-Oceanographic Research Papers. 49:2009-2033.   10.1016/s0967-0637(02)00116-4   AbstractWebsite

Observations of flow-stimulated bioluminescence have been recorded for centuries throughout the world's oceans. The present study explores, within a laboratory context, the use of naturally occurring bioluminescence as a strategy towards visualizing oceanic flow fields. The response of luminescent plankton to quantifiable levels of flow agitation was investigated in fully developed pipe flow. With two different pipe flow apparatus and freshly collected mixed plankton samples obtained over a year at two separate locations, several repeatable response patterns were identified. Threshold levels for bioluminescence stimulation occurred in laminar flow with wall shear stress levels generally between 1 and 2dyn cm(-2) (0.1-0.2 N m(-2)), equivalent to energy dissipation per unit mass values Of 10(2)-10(3)cm(2)s(-1) (10(-2)-10(-1)m(2)s(-3)). In an attempt to account for different concentrations and assemblages of mixed plankton, mean bioluminescence levels were normalized by an index of the corresponding flow-stimulated bioluminescence potential. This procedure generally accounted for variability between turbulent flow experiments, but was not effective for laminar flow. In turbulent flow, mean bioluminescence levels increased approximately linearly with wall shear stress. The magnitude of the flash response of individual cells, however, remained nearly constant throughout high laminar and turbulent flow, even as the energetic length scales of the turbulence became less than the size of the organisms of interest. Threshold flow stimuli levels determined in the laboratory were compared with oceanic measurements taken from the literature and with numerical simulations of ship wakes, one of the few highly turbulent flows to be well studied. Several oceanic flow fields are proposed as candidates for bioluminescence flow visualization. Published by Elsevier Science Ltd.

Mallipattu, SK, Haidekker MA, von Dassow P, Latz MI, Frangos JA.  2002.  Evidence for shear-induced increase in membrane fluidity in the dinoflagellate Lingulodinium polyedrum. Journal of Comparative Physiology a-Neuroethology Sensory Neural and Behavioral Physiology. 188:409-416.   10.1007/s00359-002-0315-9   AbstractWebsite

Fluid shear stress has been demonstrated to affect the structure and function of various cell types. In mammalian cells, it was hypothesized that shear-induced membrane fluidization leads to activation of heterotrimetric G-proteins. The purpose of this study was to determine if a similar mechanism exists in the dinoflagellate Lingulodinium polyedrum, a single-celled eukaryotic aquatic organism that bioluminesces under shear stress. Membrane fluidity changes in L. polyedrum were monitored using the molecular rotor 9-(dicyanovinyl)-julolidine, whose fluorescence intensity changes inversely with membrane fluidity. Dual-staining with 9-(dicyanovinyl)julolidine and the membrane dye 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene p-toluenesulfonate indicates membrane localization. Subjecting L. polyedrum cells to increasing shear stress reversibly decreased 9-(dicyanovinyl)-julolidine fluorescence, while autofluorescence of the cytoplasmic chlorophyll did not change. The relationship between shear stress (0.63 Pa, 1.25 Pa, 1.88 Pa, and 2.5 Pa) and membrane fluidity changes was linear and dose-dependent with a 12% increase in fluidity at 2.5 Pa. To further explore this mechanism a membrane fluidizing agent, dimethyl sulfoxide was added. Dimethyl sulfoxide decreased 9-(dicyanovinyl)-julolidine emission by 41 +/- 15% and elicited a dose-dependent bioluminescent response at concentrations of 0.2%, 0.5%, 1.0%, and 1.25%. This study demonstrates a link between fluid shear stress and membrane fluidity, and suggests that the membrane is an important flow mechanosensor of dinoflagellates.