Synthesis, Evaluation, and Characterization of an Ergotamine Imprinted Styrene-Based Polymer for Potential Use as an Ergot Alkaloid Selective Adsorbent

Alkaloid toxicities negatively impact livestock health and production. To assess alkaloid occurrences, adsorbent technologies may offer effective means to their extraction and isolation from a complex feed matrix. In this study, molecularly imprinted polymers (MIPs) were synthesized and evaluated for their specificity of binding to various ergot alkaloids. Co-polymers of styrene and hydroxyethyl methacrylate were synthesized in the absence or presence of an ergotamine (ETA) template, yielding non-imprinted polymer (NIP) and molecularly imprinted polymer (MIP), respectively. The influence of parameters such as pH, temperature, and initial concentration on the adsorption of ergot alkaloids was evaluated along with their application as solid phase extraction materials. Chemical and morphological properties were characterized. Adsorption was generally greater for MIP compared to NIP. Cross-reactivity with related alkaloids existed due to similarities in structure and functional groups and was dependent on the type and concentration of alkaloid and polymer type (alkaloid type × concentration × product; P \u3c 0.05). The pH of the medium had no influence on the binding properties of polymers toward ETA within a pH range of 2–10. Binding was independent of temperature between 36 and 42 °C. When kinetics of adsorption were evaluated, the Langmuir isotherm had a better fit (R2 \u3e 0.95) to adsorption equilibrium data than the Freundlich equation. The maximum amounts adsorbed (Qo) from the Langmuir model were 8.68 and 7.55 μM/g for MIP and NIP, respectively. Fourier transform infrared, scanning and tandem electron microscopy, and Brunauer–Emmett–Teller analysis confirmed a highly porous MIP structure with a greater surface area compared to NIP. Binding characteristics evaluated with computational strategy using molecular docking experiments and in vitro in a complex media (rumen fluid) indicated a stronger ETA adsorption by the tested composition selected among other polymeric materials and affinity of MIP compared with NIP. This study suggested the possible utility of MIP as a solid phase extraction sorbent for applications in analytical chemistry or sensing devices tailored to track ergot alkaloid incidence and the fate of those alkaloids in complex ruminal digestive samples

Development and Utilization of an Isolation Box Test to Characterize Personality Traits of Dairy Calves

The use of an isolation box test (IBT) to characterize personality traits has been used in non-bovine species with success. We aimed to develop an IBT for dairy calves and determine if the behavioral responses to an IBT were associated with personality traits found from traditional tests (novel person, novel object, and a startle tests) and average daily gain (ADG; Kg/d) through weaning. Calf movement while in the IBT was measured via accelerometers attached to 5 locations on the exterior of the box. A total movement index (TMI) was calculated based on accelerometer readings during the IBT. We performed a principal component analysis on the traditional tests and identified 3 influential factors that we labeled as “fearful,” “bold,” and “active.” Calves were weighed biweekly to track liveweight ADG. Factor scores and ADG were regressed against TMI. A significant negative association was found between the TMI and the factor “active,” indicating the validity of IBT as a tool for assessing some personality traits of dairy calves. Furthermore, TMI had a positive association with ADG through the entire experimental period and thus has potential to help predict performance through weaning. IBT has potential to be used as a personality test in research scenarios. Further development is needed to produce an IBT that would be appropriate to measure animals\u27 responses reliably in production settings. An automated test that can accommodate a wide range of ages and developing a computer learning model to interpret output from the IBT would be a possible option to do so

Suitability of a GPS Collar for Grazing Studies

The traditional means of tracking animal location in a field is by visual observation. Not only is this method labor intensive, it is also prone to error as the observer can alter cattle movement, observation periods are often too short to obtain confidence in general daily behavior patterns, and observer fatigue becomes an issue. In the 1990s, the University of Kentucky began using GPS collars on cattle to track their position with the goal of incorporating this information into cattle management practices. One of the key unanswered questions regarding the GPS collars is the accuracy of the position data recorded by the collar. The objective of this work was to assess the capabilities and limitations of using GPS collars to track animal movement in grazed watersheds. Static tests were conducted in an open field, under trees, and near fence lines to ascertain the impacts of various field features on collar performance. Dynamic tests were carried out to examine the errors associated with the collars while operated under real-world conditions. Results from these tests indicate that the collars generally provide data with horizontal accuracies of 4 to 5 m. This information will assist researchers in the development of experiments based on collar capabilities and limitations

Contractile Response of Bovine Lateral Saphenous Vein to Ergotamine Tartrate Exposed to Different Concentrations of Molecularly Imprinted Polymer

Ergot alkaloids, in their active isomeric form, affect animal health and performance, and adsorbents are used to mitigate toxicities by reducing bioavailability. Adsorbents with high specificity (molecularly imprinted polymers: MIP) adsorb ergot alkaloids in vitro, but require evaluation for biological implications. Using ex vivo myography, synthetic polymers were evaluated for effects on the bioactivity of ergotamine tartrate (ETA). Polymers were first evaluated using isotherms. Lateral saphenous veins were collected from 17 steers for four independent studies: dose response of ETA, adsorbent dose response, validation of pre-myograph incubation conditions and MIP/ non-molecularly imprinted polymer (NIP) comparison. Norepinephrine normalized percent contractile response to increasing ETA exhibited a sigmoidal dose response (max: 88.47 and log of the effective molar concentration (EC50) (−log [ETA]) of 6.66 ± 0.17 M). Although sample preparation time affected contractile response (p \u3c 0.001), pre-myograph incubation temperature (39 vs. 21 °C, 1 h) had no effect (p \u3e 0.05). Isothermal adsorption showed a maximum adsorption of 3.27E-008 moles·mg−1 and affinity between 0.51 and 0.57 mg (R2: 0.83–0.92) for both polymers, with no significant difference between polymers (p \u3e 0.05). No significant differences in maximum inhibitory (p = 0.96) and IC50 responses (p = 0.163) between MIP and NIP were noticed. Normalized percent contraction could be predicted from the in vitro adsorption data (R2 = 0.87, p\u3c 0.01), for both polymers. These studies indicate that synthetic polymers are potentially effective adsorbents to mitigate ergot toxicity caused by ergot alkaloids, with little evidence of significant differences between MIP and NIP in aqueous media

Streambank Erosion Associated with Grazing Practices in the Humid Region

The effects of cattle grazing on stream stability have been well documented for the western portion of the U.S., but are lacking for the east. Stream and riparian damage resulting from grazing can include alterations in watershed hydrology, changes to stream morphology, soil compaction and erosion, destruction of vegetation, and water quality impairments. However, few studies have examined the successes of best management practices (BMPs) for mitigating these effects. The objective of this project was to assess the ability of two common BMPs to reduce streambank erosion along a central Kentucky stream. The project site consisted of two replications of three treatments: (1) an alternate water source and a fenced riparian area to exclude cattle from the stream except at a 3.7 m wide stream ford, (2) an alternate water source with free stream access, and (3) free stream access without an alternate water source (i.e., control). Fifty permanent cross-sections were established throughout the project site. Each cross-section was surveyed monthly from April 2002 until November 2003. Results from the project indicated that the incorporation of an alternate water source and/or fenced riparian area did not significantly alter stream cross-sectional area over the treatment reaches. Rather than exhibiting a global effect, cattle activity resulted in streambank erosion in localized areas. As for the riparian exclosures, changes in cross-sectional area varied by location, indicating that localized site differences influenced the processes of aggradation and/or erosion. Hence, riparian recovery within the exclosures from pretreatment grazing practices may require decades, or even intervention (i.e., stream restoration), before a substantial reduction in streambank erosion is noted

Feeding Cannabidiol (CBD)-Containing Treats Did Not Affect Canine Daily Voluntary Activity

Growing public interest in the use of cannabidiol (CBD) for companion animals has amplified the need to elucidate potential impacts. The purpose of this investigation was to determine the influence of CBD on the daily activity of adult dogs. Twenty-four dogs (18.0 ± 3.4 kg, 9 months−4 years old) of various mixed breeds were utilized in a randomized complete block design with treatments targeted at 0 and 2.5 mg (LOW) and at 5.0 mg (HIGH) CBD/kg body weight (BW) per day split between two treats administered after twice-daily exercise (0700–0900 and 1,700–1,900 h). Four hours each day [1,000–1,200 h (a.m.) and 1,330–1,530 h (p.m.)] were designated as times when no people entered the kennels, with 2 h designated as Quiet time and the other 2 h as Music time, when calming music played over speakers. Quiet and Music sessions were randomly allotted to daily a.m. or p.m. times. Activity monitors were fitted to dogs\u27 collars for continuous collection of activity data. Data were collected over a 14-day baseline period to establish the activity patterns and block dogs by activity level (high or low) before randomly assigning dogs within each block to treatments. After 7 days of treatment acclimation, activity data were collected for 14 days. Data were examined for differences using the MIXED procedure in SAS including effects of treatment, day, session (Quiet or Music), time of day (a.m. or p.m.), and accompanying interactions. CBD (LOW and HIGH) did not alter the total daily activity points (P = 0.985) or activity duration (P = 0.882). CBD tended (P = 0.071) to reduce total daily scratching compared with the control. Dogs were more active in p.m. sessions than in a.m. sessions (P \u3c 0.001). During the p.m. session, dogs receiving HIGH tended (P = 0.091) to be less active than the control (CON). During the a.m. and p.m. sessions, CBD reduced scratching compared with CON (P = 0.030). CBD did not affect the activity duration during exercise periods (P = 0.143). These results indicate that, when supplemented with up to 4.5 mg CBD/kg BW/day, CBD does not impact the daily activity of adult dogs, but may exert an antipruritic effect

Consumption of endophyte-infected fescue seed during the dry period does not decrease milk production in the following lactation.

Ergot alkaloids in endophyte-infected grasses inhibit prolactin (PRL) secretion and may reduce milk production of cows consuming these grasses. We investigated the effects of consuming endophyte-infected fescue seed during late lactation and the dry period on mammary growth, differentiation, and milk production. Twenty-four multiparous Holstein cows were randomly assigned to 3 treatment groups. Starting at 90±4 d prepartum, cows were fed endophyte-free fescue seed (control; CON), endophyte-free fescue seed plus 3×/wk subcutaneous injections of bromocriptine (0.1mg/kg of body weight, positive control; BROMO), or endophyte-infected fescue seed (INF) as 10% of the diet on an as fed basis. Although milk yield of groups did not differ before treatment, at dry off (-60 d prepartum) INF and BROMO cows produced less milk than CON. Throughout the treatment period, basal concentrations of PRL and the prepartum increase in plasma PRL were reduced in INF and BROMO cows compared with CON cows. Three weeks after the end of treatment, circulating concentrations of PRL were equivalent across groups. In the subsequent lactation milk yield was not decreased; in fact, BROMO cows exhibited a 9% increase in milk yield relative to CON. Evaluation of mammary tissue during the dry period and the subsequent lactation, by quantitative histology and immunohistochemical analysis of proliferation markers and putative mammary stem or progenitor cell markers, indicated that feeding endophyte-infected fescue seed did not significantly affect mammary growth and development. Feeding endophyte-infected grasses during the dry period may permit effective utilization of feed resources without compromising milk production in the next lactation

Contractile Response of Bovine Lateral Saphenous Vein to Ergotamine Tartrate Exposed to Different Concentrations of Molecularly Imprinted Polymer

Ergot alkaloids, in their active isomeric form, affect animal health and performance, and adsorbents are used to mitigate toxicities by reducing bioavailability. Adsorbents with high specificity (molecularly imprinted polymers: MIP) adsorb ergot alkaloids in vitro, but require evaluation for biological implications. Using ex vivo myography, synthetic polymers were evaluated for effects on the bioactivity of ergotamine tartrate (ETA). Polymers were first evaluated using isotherms. Lateral saphenous veins were collected from 17 steers for four independent studies: dose response of ETA, adsorbent dose response, validation of pre-myograph incubation conditions and MIP/ non-molecularly imprinted polymer (NIP) comparison. Norepinephrine normalized percent contractile response to increasing ETA exhibited a sigmoidal dose response (max: 88.47 and log of the effective molar concentration (EC50) (−log [ETA]) of 6.66 ± 0.17 M). Although sample preparation time affected contractile response (p < 0.001), pre-myograph incubation temperature (39 vs. 21 °C, 1 h) had no effect (p > 0.05). Isothermal adsorption showed a maximum adsorption of 3.27E-008 moles·mg−1 and affinity between 0.51 and 0.57 mg (R2: 0.83–0.92) for both polymers, with no significant difference between polymers (p > 0.05). No significant differences in maximum inhibitory (p = 0.96) and IC50 responses (p = 0.163) between MIP and NIP were noticed. Normalized percent contraction could be predicted from the in vitro adsorption data (R2 = 0.87, p < 0.01), for both polymers. These studies indicate that synthetic polymers are potentially effective adsorbents to mitigate ergot toxicity caused by ergot alkaloids, with little evidence of significant differences between MIP and NIP in aqueous media

Pathways regulating equine skeletal muscle protein synthesis respond in a dose-dependent manner to graded levels of protein intake

Activation of the mechanistic target of rapamycin (mTOR)-controlled anabolic signaling pathways in skeletal muscle of rodents and humans is responsive to the level of dietary protein supply, with maximal activation and rates of protein synthesis achieved with 0.2 to 0.4 g protein/kg body weight (BW). In horses, few data are available on the required level of dietary protein to maximize protein synthesis for maintenance and growth of skeletal muscle. To evaluate the effect of dietary protein level on muscle mTOR pathway activation, five mares received different amounts of a protein supplement that provided 0, 0.06, 0.125, 0.25, or 0.5 g of crude protein (CP)/kg BW per meal in a 5 × 5 Latin square design. On each sample day, horses were fasted overnight and were fed only their protein meal the following morning. A preprandial (0 min) and postprandial (90 min) blood sample was collected and a gluteus medius muscle sample was obtained 90 min after feeding the protein meal. Blood samples were analyzed for glucose, insulin, and amino acid concentrations. Activation of mTOR pathway components (mTOR and ribosomal protein S6 [rpS6]) in the muscle samples was measured by Western immunoblot analysis. Postprandial plasma glucose (P = 0.007) and insulin (P = 0.09) showed a quadratic increase, while total essential amino acid (P 0.05) between the 0.25 and 0.5 g of protein intake for either mTOR or rpS6 activation, indicating that protein synthesis may have reached near maximal capacity around 0.25 g CP/kg BW. This is the first study to show that the activation of muscle protein synthetic pathways in horses is dose-dependent on the level of protein intake. Consumption of a moderate dose of high-quality protein resulted in near maximal muscle mTOR pathway activation in mature, sedentary horses