Effect of supplemental trace minerals on standard and novel measures of bull fertility.

Two studies were conducted to evaluate the effects of trace mineral supplementation on traditional and novel measures of bull fertility. In Experiment 1, 37 mature bulls received one of three dietary supplements daily for 71 d: 1) Supplement without Cu, Zn, and Mn (CON); 2) Supplement with Cu, Zn, and Mn sulfate (SULF); and 3) Supplement with basic Cu chloride, and Zn and Mn hydroxychloride (CHLR). In Experiment 2, 128 Angus or Angus-Hereford bull calves were maintained on a growing diet for 75 d (year 1) or 119 d (year 2) in Calan gate equipped pens without mineral supplementation. Bulls (n = 32 head/treatment) received one of four trace mineral supplements daily for 84 d: 1) Zn with no Cu (ZN), 2) Cu with no Zn (CU), 3) Cu and Zn (ZNCU), or 4) no Cu or Zn (CON). Bull fertility measures included a breeding soundness evaluation (BSE) and novel fertility measures conducted using flow cytometry. In mature bulls, final liver Zn concentration was positively correlated (P = 0.02) with sperm concentration (r = 0.31) and tended (P = 0.06) to be negatively correlated with acrosome damage (r = -0.39). Peripubertal bulls receiving ZNCU had greater ADG than bulls in the CU group (P = 0.05). Each BSE and novel fertility component improved from Day 0-84 in peripubertal bulls and were not affected (P > 0.10) by mineral supplementation. Bulls that received no supplement (CON) had greater (P < 0.01) percentage of sperm with distal midpiece reflex (6.9 vs 4.0% for CON and TM supplement, respectively) and Dag or Dag-like defect (2.6 vs 1.4% for CON and TM supplement, respectively) in their ejaculates. Sperm viability after 30 min of incubation were not affected by trace mineral supplementation, but after 3 h incubation, sperm viability tended to differ (P = 0.06) between treatments and tended to be less for CON bulls (48.5%) compared to ZNCU bulls (55.1%). Among contrast comparisons, trace mineral supplemented bulls had greater (P < 0.05) percentage of viable sperm at 3 h post collection and reactive oxygen resistant sperm than CON bulls. Addition of Zn to trace mineral containing Cu (ZNCU) improved (P < 0.05) percentage of sperm in the ejaculate with high mitochondrial energy potential and viable sperm with intact acrosome membrane. In summary, it appears the homeostasis mechanisms for bull trace mineral maintenance are extremely efficient and mineral supplementation of mature and peripubertal bulls did not have major improvements in any laboratory or chute-side measures of bull fertility, however bulls exposed to breeding or in environments with diet antagonists might respond differently.

Potential of a Cetuximab-based radioimmunotherapy combined with external irradiation manifests in a 3-D cell assay.

Targeting epidermal growth factor receptor (EGFR)-overexpressing tumors with radiolabeled anti-EGFR antibodies is a promising strategy for combination with external radiotherapy. In this study, we evaluated the potential of external plus internal irradiation by [(90) Y]Y-CHX-A″-DTPA-C225 (Y-90-C225) in a 3-D environment using FaDu and SAS head and neck squamous cell carcinoma (HNSCC) spheroid models and clinically relevant endpoints such as spheroid control probability (SCP) and spheroid control dose 50% (SCD50 , external irradiation dose inducing 50% loss of spheroid regrowth). Spheroids were cultured using a standardized platform. Therapy response after treatment with C225, CHX-A"-DTPA-C225 (DTPA-C225), [(90) Y]Y-CHX-A"-DTPA (Y-90-DTPA) and Y-90-C225 alone or in combination with X-ray was evaluated by long-term monitoring (60 days) of spheroid integrity and volume growth. Penetration kinetics into spheroids and EGFR binding capacities on spheroid cells were identical for unconjugated C225 and Y-90-C225. Spheroid-associated radioactivity upon exposure to the antibody-free control conjugate Y-90-DTPA was negligible. Determination of the SCD50 demonstrated higher intrinsic radiosensitivity of FaDu as compared with SAS spheroids. Treatment with unconjugated C225 alone did not affect spheroid growth and cell viability. Also, C225 treatment after external irradiation showed no additive effect. However, the combination of external irradiation with Y-90-C225 (1 µg/ml, 24 hr) resulted in a considerable benefit as reflected by a pronounced reduction of the SCD50 from 16 Gy to 9 Gy for SAS spheroids and a complete loss of regrowth for FaDu spheroids due to the pronounced accumulation of internal dose caused by the continuous exposure to cell-bound radionuclide upon Y-90-C225-EGFR interaction.

Quantum-chemical calculations of the electronic structure of 2-amino-1,3-dicyano-5,6,7,8-tetrahydronaphthalene derivatives.

The UV-Visible absorption spectra of six, newly synthesized donor-substituted 2-amino-1,3-dicyano-5,6,7,8-tetrahydronaphthalene have been measured in methylcyclohexane (MCH) and assigned with the help of quantum-chemical calculations. Our calculations have been performed to assess information regarding the electronic state energy values, corresponding oscillator strengths, x-, y-, z-components of the transition dipole moments and molecular orbitals involved in the main electronic transitions of the studied compounds. Additionally, the experimental absorption transition dipole moments were calculated, on the basis of spectroscopic data, and compared with results of our quantum-chemical calculations. On the basis of the experimental results and quantum-chemical calculations, it was shown that the long-wavelength absorption band involves an overlap of three electronic transitions of different character. For all studied donor-acceptor (D-A) compounds in vapour-phase, the long-wavelength transition (S0→S1) does not possess charge transfer character, whereas the S0→S2 transition possesses electron transfer character e.g., π-electrons of the acceptor moiety are moved to the donor part. Moreover, it is found that the electronic structure of the studied biphenyl derivatives can be approximately described within composite-model of decoupled moieties: donor and acceptor.

Excited-state dynamics of ethyl 5-(4-aminophenyl)-3-amino-2,4-dicyanobenzoate.

The photophysics and excited-state dynamics of ethyl 5-(4-aminophenyl)-3-amino-2,4-dicyanobenzoate (EAADCy) in solvents of varying polarity and viscosity have been studied using femtosecond transient absorption spectroscopy. Global analysis of the time-resolved spectra revealed three processes occurring in an optically excited molecule. The sequence of reactions begins with a transition from an initially excited Franck-Condon state to the nonrelaxed intramolecular charge transfer (ICT(NR)) state which is associated with a partial electron transfer. This process is followed by an additional relaxation to a more relaxed intramolecular charge transfer (ICT(R)) state with stronger charge transfer character and flattened geometry. The lifetime of the flattened charge transfer form (ICT(R)) shortens from 200 to 300 ps in medium polar solvents down to 10 ps in strongly polar solvents. On the other hand, increase of viscosity by 1 order of magnitude leads to deceleration of processes involving twist of the donor and acceptor moieties by a factor of approximately 2.5. Observation of long-lived fluorescence of EAADCy in medium polar solvent suggests that charge transfer is possible only from a hot Franck-Condon state, but not from a relaxed locally excited state which exhibits short-wavelength fluorescence on a nanosecond time scale.

Flow cytometric cell-based assay to preselect antibody constructs for radionuclide conjugation.

Radiolabeled antibodies (Abs) are an attractive tool for targeting and delivering particle emitters for therapy or imaging applications. The labeling of Abs with metal radionuclides requires chelating agents and can cause loss of binding to their ligands. The aim of the present approach was to design an easy-handling flow cytometric cell-based assay to evaluate Ab-binding capacity of conjugates of the therapeutic Ab Cetuximab and to verify the most promising candidate in a competitive radioactive binding experiment. The final setup for flow cytometric assessment of cellular binding capacities of epidermal growth factor receptor (EGFR)/ErbB1-directed Ab conjugates is based on (a) the selection of a robust cell line model (b) the definition of nonsaturated staining concentrations for the unconjugated reference Ab Cetuximab plus implementation of a reasonable isotype control, and (c) the calculation of relative Ab affinities based on the flow cytometric data. Two (FaDu, SAS) out of the three cell lines with different total and cell surface expression levels of EGFR turned out to be adequate models but the application of one cell line was sufficient to estimate reduced binding capacities of conjugates relative to Cetuximab. Only 1/11 conjugate Abs exhibited a fluorescence signal comparable to unconjugated Cetuximab and was applied for radiolabeling with Yttrium-90. Unaltered binding affinity of this conjugate was proven in a competitive radioactive Ab-binding study. We conclude that the flow cytometric assay is reliable and that the relative binding capacity of Cetuximab is neither affected by covalent modification with CHX-A"-DTPA (N-[(R)-2-Amino-3-(p-isothiocyanato-phenyl) propyl]-trans-(S,S)-cyclohexane-1,2-diamine-N,N,N',N",N"-pentaacetic acid) with a final chelator-to-Ab ratio of 5 nor by subsequent radiolabeling. [(90)Y]Y-CHX-A"-DTPA-Cetuximab thus qualifies for preclinical treatment testing as a prerequisite for therapeutic application.

Cellular and molecular properties of (90)Y-labeled cetuximab in combination with radiotherapy on human tumor cells in vitro.

PURPOSE: Anti-EGFR antibody cetuximab (C225) is used in combination with radiotherapy of head and neck squamous cell carcinoma (HNSCC) patients. We investigated whether conjugation of cetuximab with trans-cyclohexyl-diethylene-triamine-pentaacetic acid (CHX-A″-DTPA) and radiolabeling with (90)Yttrium affect the molecular and cellular function of cetuximab and improve its combined effect with external-beam irradiation (EBI). METHODS: The following cell lines were used: HNSCC UT5, SAS, FaDu, as well as A43, Chinese hamster ovary cells (CHO), and human skin fibroblast HSF7. Binding affinity and kinetics, specificity, retention, and the combination of (90)Y-cetuximab with EBI were evaluated. RESULTS: Control cetuximab and CHX-A″-DTPA-cetuximab blocked the proliferation activity of UT5 cells. In combination with EBI, CHX-A″-DTPA-cetuximab increased the radiosensitivity of UT5 to a similar degree as control cetuximab did. In contrast, in SAS and HSF7 cells neither proliferation nor radiosensitivity was affected by either of the antibodies. Binding [(90)Y]Y-CHX-A″-DTPA-cetuximab ((90)Y-cetuximab) to EGFR in HNSCC cells occurred time dependently with a maximum binding at 24 h. Retention of (90)Y-cetuximab was similar in both HNSCC cell lines; 24 h after treatment, approximately 90% of bound activity remained in the cell layer. Competition assays, using cell membranes in the absence of an internalized fraction of cetuximab, showed that the cetuximab affinity is not lost as a result of conjugation with CHX-A″-DTPA. Cetuximab and CHX-A″-DTPA-cetuximab blocked EGF-induced Y1068 phosphorylation of EGFR. The lack of an effect of cetuximab on EGF-induced Akt and ERK1/2 phosphorylation and the inhibition of irradiation (IR)-induced Akt and ERK1/2 phosphorylation by cetuximab were not affected by DTPA conjugation. (90)Y-cetuximab in combination with EBI resulted in a pronounced inhibition of colony formation of HNSCC cells. CONCLUSIONS: Conjugation of CHX-A″-DTPA to cetuximab does not alter the cellular and biological function of cetuximab. (90)Y-labeling of cetuximab in combination with EBI may improve radiotherapy outcome.

Influence of prototropic reactions on the absorption and fluorescence spectra of methyl p-dimethylaminobenzoate and its two ortho derivatives.

The influence of prototropic reactions on the spectral characteristics of methyl p-dimethylaminobenzoate (I) and its o-methoxy (II) and o-hydroxy (III) derivatives has been studied using steady-state spectroscopic technique and quantum-chemical calculations. This study concerns the solvent-induced shift of the absorption, locally excited (LE) and intramolecular charge transfer (ICT) fluorescence bands in the neat tetrahydrofuran (THF) and its hydrochloric acid solutions at different HCl concentrations. On the basis of the experimental results and quantum-chemical calculations, it was shown that in a hydrochloric acid solution the studied molecules exist as a mixture of neutral, mono-, and dicationic forms. Additionally, the results of spectroscopic measurements were used to calculate, according to the Benesi-Hildebrand method, the equilibrium constants of protopropic reactions in the ground, S(0), and excited, S(1), states. Our findings predestine molecules I and II to be used as acid fluorescence probes in a region of 0-2.5 M of [H(+)] concentrations.

Prediction of the energy content of tallgrass prairie hay.

Two experiments were conducted to describe the DE content of tallgrass prairie hay (TPH). In trial 1, steers (n = 13; 277 +/- 15 kg of BW) were used in a 13 x 4 Latin square experiment to measure the DE of 13 samples of TPH fed at 1.5% of BW daily (average feeding level = 0.7 x the maintenance energy requirement). Hays were harvested from a variety of locations in east-central Kansas and represented an array of harvest dates and storage methods. In trial 2, steers (n = 16; 261 + 17 kg of BW) were used in a randomized complete block experiment to assess the effects of TPH intake level on DE. Hay was fed at 1.3, 1.7, 2.1, or 2.5% of BW daily, which corresponded to 0.9, 1.4, 1.6, and 1.9 x the maintenance energy requirement. Steers in both trials were fed soybean meal in amounts calculated to provide ruminally degradable protein (RDP) equal to 11% of digestible OM intake. Hay samples were analyzed for ash, N, NDF, ADF, ADIN, NDIN, acid detergent-insoluble ash, lignin, monosaccharides, and alkali-labile phenolic acids. Chemical components related to DE (P < 0.2) were subjected to iterative regression analysis to predict the DE concentration of the diet. Iterations were ceased when the error mean square of the regression was optimized. At 0.7 x maintenance, the dietary DE concentration (Mcal/kg) was described by: DE = 0.13(CP) - 0.16(ADL) + 2.11 (R(2) = 0.73; S(y*x) = 0.13). Forage OM digestion decreased linearly (P < 0.01) as forage intake increased. Apparent dietary DE concentration decreased by 7.4% when intake was increased from 1 to 2 x maintenance. When RDP was adequate, chemical composition values were useful indicators of forage DE content in our study. Moreover, increased forage intake depressed GE digestion by steers, but ultimately increased total DE intake. Energy digestion varied with forage intake in a predictable manner between 1 and 2 x the maintenance feeding level.

Influence of spectral heterogeneity of prodan and laurdan solutions on the transfer of electronic energy to octadecyl rhodamine B.

Fluorescence quenching and resonance energy transfer have been studied by steady-state fluorescence spectroscopy. The experimental and theoretical values for the rate constants of the electronic energy transfer (kET) and critical radius (R0) were determined for prodan and laurdan as donors and octadecyl rhodamine B as acceptor. The spectroscopic data show, that prodan and laurdan in solution create an inhomogeneous spectroscopic medium in which multi-channel luminescence phenomena take place. This finding indicated that the modified form of the Stern-Volmer relation should be used for analyzing fluorescence quenching data. Results of performed studies point out, that dipole-dipole interaction is responsible for the resonance energy transfer from prodan and laurdan to octadecyl rhodamine B. The relative quenching efficiencies of both dyes depend on polarity of the medium and are higher for more polar solvent (AcN).

Phase transition affects energy transfer efficiency in phospholipid vesicles.

The fluorescence quenching of 6-propionyl-2-dimethylaminonaphtalene (PRODAN) and 6-dodecanoyl-2-dimethylaminonaphtalene (LAURDAN) by octadecyl rhodamine B (ORB) in a model system of small unilamellar vesicles (SUV) of dipalmitoylphosphatidyl-choline (DPPC) was investigated. Non-linear Stern-Volmer behaviour was observed in both systems in the gel phase (25 degrees C) and in the fluid phase (50 degrees C), resulting from association processes and from static quenching. The relative quenching efficiencies of both dyes depend on the phase state of the bilayer and indicate a deeper incorporation of PRODAN and LAURDAN into the membrane in its fluid phase than in its gel phase.

Effect of supplementation frequency and supplemental urea level on dormant tallgrass-prairie hay intake and digestion by beef steers and prepartum performance of beef cows grazing dormant tallgrass-prairie.

Effect of supplementation frequency and supplemental urea level on forage use (Exp. 1) and performance (Exp. 2 and 3) of beef cattle consuming low-quality tallgrass-prairie were evaluated. For Exp. 1 and 2, a 2 x 2 factorial treatment structure was used, such that two supplements (30% CP) containing 0 or 30% of supplemental degradable intake protein (DIP) from urea were fed daily or on alternate days. In Exp. 1 and 2, supplement was fed at 0.41% BW daily or at 0.83% BW (DM basis) on alternate days. For Exp. 3, a 2 x 4 factorial treatment structure was used, such that four supplements (40% CP) containing 0, 15, 30, or 45% of supplemental DIP from urea were fed daily or 3 d/wk. Supplements were group-fed at 0.32% BW daily or at 0.73% BW (DM basis) 3 d/wk. In Exp. 1, 16 Angus x Hereford steers (initial BW = 252 kg) were blocked by BW and assigned to treatment. Urea level x supplementation frequency interactions were not evident for forage intake, digestion, or rate of passage. Forage OM intake (OMI) and total digestible OMI (TDOMI) were not significantly affected by treatment. Total-tract digestion of OM (P = 0.03) and NDF (P = 0.06) were greater for steers supplemented daily. In Exp. 2, 48 Angus x Hereford cows (initial BW = 490 kg) grazing winter tallgrass prairie were used. Significant frequency x urea interactions were not evident for BW and body condition (BC) change; similarly, the main effects were not substantive for these variables. In Exp. 3, 160 Angus x Hereford cows (initial BW = 525 kg) grazing dormant, tallgrass prairie were used. Supplement refusal occurred for cows fed the highest urea levels, particularly for cows fed the supplement with 45% of the DIP from urea 3 d/wk, and supplement refusal increased closer to calving. A frequency x urea interaction (P = 0.02) was observed for prepartum BW changes. As supplemental urea level increased, prepartum BW loss increased quadratically (P = 0.02); however, a greater magnitude of loss occurred when feeding supplements containing > or = 30% of DIP from urea 3 d/ wk. Cumulative BC change followed a similar trend. In conclusion, moderate protein (< or = 30% CP) supplements with < or = 30% of supplemental DIP from urea can be fed on alternate days without a substantive performance penalty. However, infrequent feeding of higher protein (> 30% CP) supplements with significant urea levels (> 15% of DIP from urea) may result in decreased performance compared with lower urea levels.

Influence of conformational effects on spectroscopic properties of 2-dimethylamino-9(4'-dimethylamino) phenyl-9-fluorenol.

Absorption and fluorescence spectra of 2-dimethylamino-9(4'-dimethylamino)phenyl-9-fluorenol have been studied in different solvents. The fluorescence spectra at various temperatures show the excitation wavelength dependence. The spectroscopic data are compared with the results of the quantum chemical calculation obtained using PM3 and INDO/S methods. The fluorescence decay curves of the molecule under study can be only satisfactorily fitted by a biexponential function. The steady state and time resolved spectroscopy studies as well as the theoretical calculation confirm that in methylcyclohexane and ethyl acetate solutions always two absorbing and emitting species exist, namely rotamers alpha and beta in the ground and excited states.

Impact of different wheat milling by-products in supplements on the forage use and performance of beef cattle consuming low-quality, tallgrass-prairie forage.

Two experiments were conducted to evaluate the impacts on forage use and beef cattle performance of incorporating divergent wheat milling by-products in a 30% CP supplement. The by-products were wheat bran (high fiber) and second clears (high starch). The by-products were added as 1) 100% wheat bran; 2) 67% wheat bran, 33% second clears; or 3) 33% wheat bran, 67% second clears to constitute approximately 47 to 49% of the supplement. In Exp. 1, 90 Hereford x Angus cows (BW = 554 kg) grazing winter, tallgrass-prairie range were fed the supplement treatments (2.27 kg/cow daily) from early December 1997 until calving (average calving date = 3/11/98). Cumulative BW and condition changes from trial initiation through calving were not significantly different among treatments. Similarly, significant treatment effects on cow pregnancy rates as well as calf birth weights, ADG, and ending weights were not evident. In Exp. 2, 16 ruminally fistulated Hereford x Angus steers (BW = 484 kg) were blocked by weight and assigned to one of the same three supplement treatments or to a negative control (forage only). Steers had ad libitum access to tallgrass-prairie hay (76.4% NDF, 3.1% CP) and were fed supplement at the same rate (relative to BW) as the cows in Exp. 1. Forage OM, NDF, and digestible OM intakes were lower (P < 0.01) for the negative control than for supplemented steers but were not significantly different among the supplemented steers. Digestion of OM was lower (P = 0.03) for the negative control than for supplemented steers, although significant treatment differences were not evident among the supplemented groups. Digestion of NDF was not affected (P = 0.49) by treatment. Within the context of the amount of supplemental protein offered, changes in the combination of wheat milling by-products in the supplement did not affect cow performance or intake and digestion of low-quality forage.

Differential response of immature and mature neurons to hypoxia in rat mesencephalic cultures.

The effect of hypoxia on immature and mature mesencephalic neurons was studied in in vitro rat cerebral cell cultures on different days. In immature cultures (6-8 days in vitro), exposure to 24 h of hypoxia (10-20 mm Hg pO(2) in the culture medium) did not change the number of neuron-specific enolase (NSE)-immunoreactive (IR) (NSE-IR) neurons but increased the number of tyrosine hydroxylase (TH)-IR (TH-IR) cells, which might be attributed to transient induction of TH. In mature cultures (13-15 days in vitro), 16 h of hypoxia induced a considerable loss of both NSE- and TH-IR cells. A decrease in the number of TH-IR cells 6 and 24 h after hypoxia was more pronounced than that of NSE-IR cells; however, their numbers equalized 48 h after hypoxia, suggesting similar hypoxic vulnerability of dopaminergic and nondopaminergic neurons in mature mesencephalic cultures. In immature cultures, hypoxia slightly stimulated both apoptosis and necrosis, while in mature cultures, it dramatically increased the number of solely necrotic cells.

Differential effects of hypoxia on untreated and NGF treated PC12 cells.

Perinatal hypoxia is known to induce long-lasting changes in the central dopaminergic system. In order to understand the cellular mechanism of these changes, we studied the effects of hypoxia on the levels of dopamine (DA) and tyrosine hydroxylase (TH) mRNA in untreated and NGF treated PC12 cells. On the second day after plating (DAP), cells were exposed to a hypoxic episode (pO2 = 10-20 mm Hg, 24 h), and the levels of DA and TH mRNA were examined on DAP 4 and DAP 8. In untreated cells, hypoxia induced a two fold increase both in DA and TH mRNA levels on DAP 4 which normalized up to DAP 8. This increase correlated with an activation of the hypoxia inducible factor (HIF-1alpha), measured with a reporter gene. In contrast, NGF treated cells responded to hypoxia with an increase of DA level on DAP 8. In these cells neither an increase of the HIF-1alpha activity measured immediately after hypoxia nor a significant increase of the TH mRNA level on DAP 8 were found. The findings indicate that NGF shifts the hypoxia induced changes of DA levels from a short-term to a long-term mode. The long-term increase of dopamine levels is the most likely result of changes connected with cell growth and differentiation and not the result of a long-term TH mRNA level increase.

Density functional study of the S0 (X1Ag) and T1 (a3Au) states of the glyoxal molecule

The density functional theory (DFT) calculations in different approximations have been performed for the geometries and vibrational states of the trans-glyoxal molecule in the ground state S0 (X1Ag) and in the lowest excited triplet state T1 (a3Au, n-pi* type). Eight typical combinations of exchange and correlation functionals have been used. Comparative Hartree-Fock (HF) calculations have also been performed. For the open shell a3Au state the standard spin-unrestricted Hartree-Fock and Kohn-Sham approaches (UHF, UKS) as well as the restricted open-shell versions (ROHF, ROKS) have been applied. The calculated frequencies have been compared, among others, with the data resulting from the most recent phosphorescence exicitation spectra of glyoxal cooled in the supersonic molecular beam, recorded with the cooperation of one of us (JH) for the spin-forbidden S0-T1 transition. The most realistic description of the vibrational frequencies, within the unscaled harmonic approximation, can be obtained using the 3-parameter Becke-93 exchange functional (B3), whereas this description practically does not depend on the correlation functional used. Our calculations support the recently reexamined experimental energy of the symmetric CH-rocking fundamental for the T1 state.

Effects of supplemental degradable intake protein on utilization of medium- to low-quality forages.

Three independent experiments were conducted each using 16 ruminally fistulated beef steers fed bermudagrass (8.2% CP, 71% NDF; Exp. 1), bromegrass (5.9% CP, 65% NDF; Exp. 2), or forage sorghum (4.3% CP, 60% NDF; Exp. 3) hays to evaluate the effects of increasing level of supplemental degradable intake protein (DIP) on forage utilization. In each experiment, steers were blocked by weight and assigned to one of four treatments, and hay was offered to each steer at 130% of average voluntary intake for the preceding 5-d period. Supplemental DIP (sodium caseinate) was placed in the rumen at 0700, immediately before feeding forage. Levels of DIP supplementation were .041, .082, and .124% BW; the control received no supplemental DIP. Following a 10-d adaptation, intake and total fecal output were measured for 7 d. In Exp. 1, neither forage OM intake (FOMI) nor fiber (NDF) digestion were influenced (P > or = .20) by increasing level of DIP supplementation. The DIP supplied by the bermudagrass hay was estimated to be 8.2% of the total digestible OM intake (TDOMI) for control steers. In Exp. 2, increasing level of supplemental DIP did not affect (P > or = .26) FOMI but tended to increase total OM intake linearly (TOMI; P = .10). The tendency for a rise in TOMI coupled with a slight numeric increase in digestion resulted in an increase (linear; P = .06) in TDOMI. In the treatment group in which the maximum TDOMI was observed (supplemental DIP treatment of .082% BW), total DIP intake constituted approximately 9.8% of the TDOMI. In Exp. 3, FOMI, TOMI, organic matter digestion (OMD), and TDOMI were improved (P < .01) by increasing amounts of supplemental DIP. Although there was some evidence of a tendency for a decrease in the magnitude of change in TDOMI in response to increasing DIP supplementation, a clear plateau was not achieved with the levels of supplement provided. When the highest level of supplemental DIP was fed, DIP constituted approximately 12.8% of the TDOMI. In conclusion, significant variation was observed among forage in the amount of DIP needed to maximize intake and digestion when expressed in relationship to the digestible OM.

Early hypoxia modulates the phenotype of dopaminergic cells in rat di- and mesencephalic cell cultures and induces a higher vulnerability of non-dopaminergic neurons to a second hypoxic exposure.

To investigate long-term effects of hypoxia on a cellular level, di- and mesencephalic cell cultures were exposed to hypoxia on in vitro day 2 (incubation in culture medium, pO2 = 10-20 mmHg, 24 h) and on in vitro day 13 (incubation in an electrolyte solution, pO2 = 10-20 mmHg, 8 h). The numbers of neuron-specific enolase immuno-reactive (NSE-IR) and tyrosine hydroxylase immuno-reactive (TH-IR) neurons and the levels of dopamine, its main metabolites and the spontaneous and potassium-stimulated DA release were determined on DIV 15. Hypoxia on DIV 2 did not affect the numbers of NSE-IR and TH-IR neurons, but increased the dopamine content and dopamine release by about 100% in both di-and mesencephalic cultures. In addition, this hypoxia increased the vulnerability of non-TH-IR neurons to the second hypoxic episode applied during more advanced stages of the culture development on DIV 13. On the contrary, hypoxia exposure did not affect the vulnerability of TH-IR cells.

Effects of different supplemental sugars and starch fed in combination with degradable intake protein on low-quality forage use by beef steers.

Twenty ruminally fistulated steers (Exp. 1, 448 kg and Exp. 2, 450 kg) were used in two consecutive randomized complete block experiments with five treatments in each experiment. The purpose was to evaluate the impact of feeding different supplemental sugars or starch in combination with supplemental degradable intake protein (DIP) on the utilization of low-quality tallgrass-prairie hay. In Exp. 1, steers were given ad libitum access to forage and, except for the negative control (NC), received a limited supply (insufficient to maximize forage use) of supplemental DIP (.031% BW/d, DM basis). In addition to the NC, this experiment included four supplementation treatments in which one of four carbohydrate (CHO) sources (starch, glucose, fructose, or sucrose) was fed at .30% BW of DM/d. In Exp. 2, the treatment structure was identical except that the supplemental DIP level (.122% BW, DM basis) was near the level needed to maximize forage use. Forage OM intake (FOMI) was not affected (P> or =.26) by supplementation in Exp. 1 but was increased (P = .05) in Exp. 2. However, no difference (P> or =.46) in FOMI occurred among CHO sources in either experiment. Total OM and digestible OM intakes were increased (P<.01) by supplementation in both experiments. In Exp. 1, no difference (P>.26) in OM digestion (OMD) occurred among treatments. In Exp. 2, supplementation increased (P<.01) OMD. Additionally, sugars yielded a higher (P = .04) OMD than starch, and the monosaccharides yielded a higher (P = .02) OMD than sucrose. In Exp. 1, NDF digestion (NDFD) was decreased (P = .02) by supplementation, but no differences (P> or =.21) occurred among CHO sources. In Exp. 2, NDFD was increased (P = .03) by supplementation. Additionally, sugars led to higher (P = .05) NDFD than starch, and the monosaccharides led to higher (P = .03) NDFD than sucrose. In both experiments, discernible patterns were observable with regard to the effects of supplementation and type of supplemental CHO on ruminal fermentation characteristics. In conclusion, even though some consistency in fermentation profiles for different carbohydrate sources was evident in both experiments, forage intake and digestion responses were not consistent across experiments. This raises the possibility that carbohydrate source may interact with the amount of supplemental DIP fed and, as such, deserves additional investigation.

Effects of level and source of carbohydrate and level of degradable intake protein on intake and digestion of low-quality tallgrass-prairie hay by beef steers.

Ruminally fistulated steers (n = 13; 263 kg) were used in an incomplete Latin square with 13 treatments and four periods to evaluate the effects of level and source of supplemental carbohydrate (CHO) and level of degradable intake protein (DIP) on the utilization of low-quality, tallgrass-prairie hay. Steers were given ad libitum access to forage (5.7% CP, 2.6% DIP, and 74.9% NDF). The supplementation treatments were fashioned as a 2x3x2 factorial arrangement plus a negative control (NC; no supplement). The factors included two DIP levels (.031 and .122% BW) and three CHO sources (starch, glucose, and fiber) fed at two levels (.15 and .30% BW) within each level of DIP supplementation. The effect of supplementation on forage OM intake (FOMI) was dependent (P<.01) on level and source of CHO and level of DIP fed. When DIP was low, forage, total, and digestible OM intakes were generally greater for the starch treatment than for the nonstarch treatments. However, when the DIP level was high, intakes were greater for the nonstarch (i.e., fiber and glucose) treatments. Generally, FOMI decreased (P<.01) when more supplemental CHO was provided. Supplementation typically increased fiber digestion, but the response was dependent (P<.01) on level and source of CHO and level of DIP. Generally, supplements with low levels of CHO improved NDF digestion (NDFD). However, supplements with the high level of CHO decreased NDFD, except for fiber at the high level of DIP. Organic matter digestion was increased by supplementation, but the impact of increasing CHO was dependent (P<.01) on source of CHO and level of DIP. Supplementation treatments had significant impact on ruminal pH, NH3 N, and the total concentration of organic acids as well as their relative proportions. In conclusion, supplemental DIP enhanced the use of low-quality forage; however, the impact of supplemental CHO on low-quality forage use was dependent on source and level of CHO offered, as well as the level of DIP provided.