CAN NUTRIENT ADDITIONS MEDIATE GAPE LIMITED PREDATION IN LARVAL RANA SPHENOCEPHALA?

Some predators are limited in the size prey item that they can catch and consume, and this is referred to as "gape limited". Prey found with these predators may have the ability to respond by growing rapidly to reach a size too large to be preyed upon, and thus reach a size refuge from predation. Due to decreased competition, these prey may have the additional benefit of experiencing an environment with more consumable resources available. One factor that may influence the ability of prey to outgrow the gape limitation of their predators is nutrient availability. Providing an influx of limiting nutrients can result in an increase in abundance of primary producers, and allow herbivore prey living in nutrient enriched environments to grow more quickly than those that live in a nutrient-limited environment. Thus, prey living in nutrient rich environments may be more likely to be able to outgrow the gape limitation of a predator, and reach a predator-free size refuge, than those prey living in areas with less nutrient availability. I was interested in whether nutrient additions can mediate the effects of gape limited predation in an aquatic food web. A mesocosm experiment was performed to evaluate the effects of nutrient influx on Rana sphenocephala tadpoles in the presence of two groups of bluegill fish predators: one that is likely to become gape limited and one that is not. The nutrients added were crystalline NaNO₃ and K₂HPO₄, in a 16:1 N:P ratio, in six different amounts ranging from 0.0 to 4.0 mg/l. The experiment ran for four months to evaluate the larval anuran response (survival, mass, time to metamorphosis) after one pulse of nutrient influx. Algal biomass and zooplankton abundance were evaluated at two times during the experiment as zooplankton feed on algae and bluegill can also feed on zooplankton. The results of this experiment indicate that although nutrient addition was found to significantly impact larval anuran survival, there was no evidence of an interaction between nutrient addition and predation. Thus, there is no strong support for the hypothesis that bottom up forces created by nutrient addition can mediate the top down forces created by predation. Higher abundances of leopard frogs in more nutrient rich environments, as a result of higher survivorship, may change the structure of aquatic communities and have cascading effects on the rest of the food web. Currently we are experiencing an influx of inorganic nutrients reaching our waterways due to anthropomorphic sources. Worldwide human activities, such as agriculture and development, have increased the amount of nitrogen and phosphorus reaching our aquatic systems. This experiment reinforces the idea that increases in these nutrient levels can cause changes in life history characteristics of populations and in community structure of aquatic and terrestrial systems impacted by nutrient additions.M.S

Influence of Chemoreflexes on Respiratory Variability in Healthy Subjects

The background of this study was the hypothesis that respiratory variability is influenced by chemoreflex regulation. In search for periodicities in the variability due to instability of the respiratory control system, spectral analysis was applied to breath-to-breath variables in 19 healthy subjects at rest. During room-air breathing, coherent oscillations in end-tidal CO 2 (P ET CO 2 ) and mean inspiratory flow (V I /T I ) were found in 15 subjects with frequencies mostly below 0.15 cycles per breath. Coherent oscillations in P ET CO 2 and V I /T I were expressed by gain (0.13 to 0.34 L/second · kPa) and phase ( ؊ 170 ؇ to ؉ 8 ؇ ). The oscillations in V I /T I were in phase with inspiratory volume (V I ). A model that describes the effects of chemoreflex feedback to noise in the system could explain these gains and phases, whereas a model without chemoreflex could not. During 100% O 2 breathing, only eight subjects had coherent oscillations in P ET CO 2 and V I /T I . The coherent oscillations in P ET CO 2 and V I /T I were interpreted as a manifestation of chemoreflex activity. We conclude that respiratory variability is not a random process but contains information on chemoreflex properties, such as the chemoreflex gain. The analysis of respiratory variability therefore provides a new tool to study the action of the chemoreflexes without application of external stimuli. Keywords: chemoreceptors; respiratory variability; spectral analysis Is the normal variability of respiratory parameters from breathto-breath a random process? This would imply that, for example, inspiratory volume (V I ) or inspiratory and expiratory time (T I and T E , respectively) are independent of previous breaths. However, due to the circulatory delay from the lungs to the systemic arteries, an "accidental" change in V I causes a change in arterial P O 2 and P CO 2 that only becomes manifest during the following breaths at a normal breathing frequency (1, 2). An adaptation of V I to such a change therefore inevitably leads to a dependency between successive breaths. Conversely, purely random variability of V I implies that V I does not take part in the feedback control of Pa O2 and Pa CO 2 through the chemoreflexes. Several authors have found evidence for a nonrandom breath-to-breath variability of respiratory parameters in the normal steady state (3-5). Significant (auto)correlations have been found between successive values of V I , T I , and T E (4, 6). Specific variability patterns have also been found, mainly in the form of subtle oscillations with a cycle time of approximately 25 seconds to more than 3 minutes (6-10). Clear periodic breathing is seldom observed in healthy subjects during wakefulness The aim of the present study was to derive information on respiratory regulation from the normal breathing pattern in the steady state. The hypothesis was that because of the delays and time constants of the chemoreflexes, continuous regulation tends to induce oscillations in ventilatory drive (represented by mean inspiratory flow, V I /T I ) with a certain coherency with oscillations in end-tidal P CO 2 (P ET CO2 ). To identify such oscillatory components and their mutual relationships, power and cross-spectral analysis was applied to breath-tobreath respiratory variables in 19 healthy subjects at rest. To test the hypothesis that the features of coherent oscillations in P ET CO2 and V I /T I are compatible with the characteristics of chemoreflex-feedback regulation, experimental spectra were compared with theoretical spectra derived from a chemoreflex model. The breathing pattern was also analyzed during 100% O 2 breathing to estimate the contribution of the peripheral chemoreflex (20). METHODS Subjects and Measurements Nineteen healthy nonsmoking medical students were studied with a history free of cardiopulmonary disease and normal physical examination (9 male and 10 female, aged 23 Ϯ 3 years, body mass index 22.2 Ϯ 3.1 kg/m 2 , mean Ϯ SD). The hospital ethical committee approved the protocol. Informed consent was obtained. The subjects knew which measurements were performed. To prevent "conscious" breathing, they were told that the study involved blood pressure regulation. They sat in a comfortable chair in a quiet room and breathed through a cushion-sealed face mask fitted with elastic bands around the head (dead space ‫ف‬ 70 ml). A Lilly type pneumotachograph (Siemens pressure transducer, Munich, Germany) was connected to the mask and hung with an elastic cord to the ceiling. A two-way nonrebreathing valve (S and W, Copenhagen, Denmark) was connected to the pneumotachograph. The inspiratory limb was connected to a stopcock (through a 1-m spirometer tube) which could be switched from room air to 100% O 2 from a 100 L bag. The stopcock was hidden behind a curtain so that the subject did not know which gas was inspired. The experiments began between 9:00 and 10:00 A . M . Recordings started after a 5-minute acclimatizion period. There were two episodes of 30 minutes with more than 5 minutes in between, performed in a random order, during which the subjects breathed either air or 100% O 2 (starting when endtidal P O 2 exceeded 85 kPa). Also measured were P O 2 and P CO 2 in the facemask (partial pressures in dry air, Centronic 200 MGA mass spectrometer, Croydon, United Kingdom), arterial O 2 saturation (Sa O 2 , Ohmeda Biox ear pulse oximeter, Madison, WI), finger arterial pressure (Finapres BMI-TNO, Amsterdam, Netherlands) and a single-channel chest-lead ECG. All signals were recorded on a Bell and Howell T4 recorder (Durham, NC) with airflow, ECG and blood pressure on FM channels and the other signals on a direct record channel using a Kayser Threde K 1180 pulse code modulator (Munich, Germany). The frequency response was 0-625 Hz for FM channels and 0-105 Hz for pulse code modulated channels

Dose fractionation effects in primary and metastatic human uveal melanoma cell lines

PURPOSE: To investigate the effects of split-dose irradiation on primary and metastatic uveal melanoma cell lines, with a clonogenic survival assay. METHODS: Appropriate cell concentrations of four primary and four metastatic human uveal melanoma cell lines were cultured for irradiation with single doses and with two equal fractions separated by 5 hours. After irradiation, colony formation was allowed for 7 to 21 days. Two cutaneous melanomas were also tested for comparison. All survival curves were analyzed using the linear quadratic (LQ) model. Specific parameters for the intrinsic radiosensitivity (alpha-component, SF2), for the capacity of repair of DNA damage (beta-component), as well as the alpha/beta ratio were calculated. RESULTS: After single-dose irradiation a wide range in the values of the alpha- and beta-component was obtained for both primary and metastatic uveal melanomas, which resulted in a wide range of alpha/beta ratios. In contrast, calculations based on split-dose data, with which the beta-component could be estimated independent of the alpha-component, indicated that estimates for the capacity of sublethal DNA damage repair was very similar in all cell lines. This indicated that intrinsic factors dominated the radiosensitivity of these cell lines. Split-dose irradiation had little influence on the intrinsic radiosensitivity (alpha-component), but cell survival increased for all cell lines. For the two cutaneous melanomas comparable split-dose results were obtained. CONCLUSIONS: For both primary and metastatic uveal melanoma cell lines, data from single and fractionated doses indicate large variations in radiosensitivity, which are mainly dominated by the intrinsic radiosensitivities. Doses of approximately 8 Gy in five fractions would be sufficient to eradicate 10(9) cells (approximately 1 cm3) of the most radioresistant tumor cell lines, but this schedule is an overkill for the radiosensitive tumor cell lines. Based on specific morphologic and histologic tumor markers, more individualized dose fractionation schedules could improve the therapeutic ratio for uveal melanomas

Changes in epidermal radiosensitivity with time associated with increased colony numbers

Epidermal clonogenic cell survival and colony formation following irradiation were investigated and related to radiosensitivity. A rapid in vivo/in vitro assay was developed for the quantification of colonies arising from surviving clonogenic cells in pig epidermis after irradiation. Bromodeoxyuridine (BrdU)-labelled cells in full thickness epidermal sheets were visualized using standard immunohistochemistry. In unirradiated skin, approximately 900 BrdU-positive cells mm(-2) were counted. In a time sequence experiment, BrdU-positive cell numbers increased from an average of 900 cells mm(-2) to approximately 1400 cells mm(-2) after BrdU-labelling for 2-24 h. In irradiated skin, colonies containing >/=16 BrdU-positive cells were seen for the first time at days 14/15 after irradiation. The number of these colonies per cm(2) as a function of skin surface dose yielded a cell survival curve with a D(0)-value (+/-SE) of 3.9+/-0.6 Gy. This relatively high D(0)-value is possibly due to a rapid fall off in depth dose distribution for the iridium-192 source and consequently a substantial contribution of hair follicular epithelium to colony formation. At 14/15 days after irradiation, the ED(50) level of 33.6 Gy for the in vivo response of moist desquamation corresponded with 2.7 colonies cm(-2). Surprisingly, the number of colonies increased with time after irradiation with an estimated doubling time of approximately 4 days, while the D(0)-value remained virtually unchanged. This increase in colony numbers could be due to migration of clonogenic cells, to the recruitment of dormant clonogenic cell survivors by elevated levels of cytokines, or to both. Although frequent biopsying caused increased cytokine levels, which had a systemic effect on unirradiated skin, it had no influence on colony formation in irradiated skin. Smaller colonies, containing 4-8 cells or 9-15 cells, were abundant, particularly after higher doses, which resulted in higher D(0)-values. The majority of these small colonies were abortive and did not progress to larger colonies. There was no statistical evidence for significant variations in the interanimal responses

Cellular radiosensitivity of primary and metastatic human uveal melanoma cell lines

PURPOSE: To investigate the radiosensitivity of uveal melanoma cell lines by a clonogenic survival assay, to improve the efficiency of the radiation regimen. METHODS: Four primary and four metastatic human uveal melanoma cell lines were cultured in the presence of conditioned medium. After single-dose irradiation (0-12 Gy), colonies were allowed to form for 6 to 14 days. Two cutaneous melanomas cell lines were also tested for comparison. The survival curves were analyzed by the linear quadratic (LQ) model, and the surviving fraction at a dose of 2 Gy (SF(2)), the SF at 10 Gy (SF(10)), the ratio of initial irreparably damaged DNA (alpha-coefficient) to the capacity to repair sublethally damaged DNA (beta-coefficient), and the plating efficiency were calculated. RESULTS: The melanomas displayed a wide range of initial irreparable DNA damage (alpha-component), as well as a capacity for repair of sublethal DNA damage (beta-component), which ultimately resulted in a wide range of alpha/beta ratios. These findings were similar in both primary and metastatic melanomas and were comparable with data obtained from two cutaneous melanomas. CONCLUSIONS: Cell lines obtained from primary and metastatic human uveal melanomas displayed a wide range of radiosensitivity, similar to that published for cutaneous melanomas. Translating these data to the clinical setting indicates that a fractionated dose of 8 to 10 Gy administered in three to four fractions, as currently delivered in many centers, should be sufficient to eradicate tumors of approximately 1 cm(3)