Comparison of models to identify lame cows based on gait and lesion scores, and limb movement variables.

Bovine lameness results in pain and suffering in cattle and economic loss for producers. A system for automatically detecting lame cows was developed recently that measures vertical force components attributable to individual limbs. These measurements can be used to calculate a number of limb movement variables. The objective of this investigation was to explore whether gait scores, lesion scores, or combined gait and lesion scores were more effectively captured by a set of 5 limb movement variables. A set of 700 hind limb examinations was used to create gait-based, lesion-based, and combined (gait- and lesion-based) models. Logistic regression models were constructed using 1, 2, or 3 d of measurements. Resulting models were tested on cows not used in modeling. The accuracy of lesion-score models was superior to that of gait-score models; lesion-based models generated greater values of areas under the receiving operating characteristic curves (range 0.75 to 0.84) and lower mean-squared errors (0.13 to 0.16) compared with corresponding values for the gait-based models (0.63 to 0.73 and 0.26 to 0.31 for receiving operating characteristic and mean-squared errors, respectively). These results indicate that further model development and investigation could generate automated and objective methods of lameness detection in dairy cattle.

Effect of adding alum or zeolite to dairy slurry on ammonia volatilization and chemical composition.

Development of cost-effective amendments for treating dairy slurry has become a critical problem as the number of cows on farms continues to grow and the acreage available for manure spreading continues to shrink. To determine effectiveness and optimal rates of addition of either alum or zeolite to dairy slurry, we measured ammonia emissions and resulting chemical changes in the slurry in response to the addition of amendments at 0.4, 1.0, 2.5, and 6.25% by weight. Ammonia volatilization over 96 h was measured with six small wind tunnels with gas scrubbing bottles at the inlets and outlets. Manure samples from the start and end of trials were analyzed for total nitrogen and phosphorus, and were extracted with 0.01 M CaCl2, 1.0 M KCl, and water with the extracts analyzed for ammonium nitrogen, phosphorous, aluminum, and pH. The addition of 6.25% zeolite or 2.5% alum to dairy slurry reduced ammonia emissions by nearly 50 and 60%, respectively. Alum treatment retained ammonia by reducing the slurry pH to 5 or less. In contrast, zeolite, being a cation exchange medium, adsorbed ammonium and reduced dissolved ammonia gas. In addition, alum essentially eliminated soluble phosphorous. Zeolite also reduced soluble phosphorous by over half, but the mechanism for this reduction is unclear. Alum must be carefully added to slurry to avoid effervescence and excess additions, which can increase soluble aluminum in the slurry. The use of alum or zeolites as on-farm amendment to dairy slurry offers the potential for reducing ammonia emissions and soluble phosphorus in dairy slurry.

Managing ammonia emissions from dairy cows by amending slurry with alum or zeolite or by diet modification.

Animal agriculture is a significant source of atmospheric ammonia. Ammonia (NH3) volatilization represents a loss of plant available N to the farmer and a potential contributor to eutrophication in low-nitrogen input ecosystems. This research evaluated on-farm slurry treatments of alum or zeolite and compared three diets for lactating dairy cows in their effectiveness to reduce NH3 emissions. NH3 emissions were compared using a group of mobile wind tunnels. The addition of 2.5% alum or 6.25% zeolite to barn-stored dairy slurry reduced NH3 volatilization by 60% and 55%, respectively, compared to untreated slurry. The alum conserved NH3 by acidifying the slurry to below pH 5, while the zeolite conserved ammonia by lowering the solution-phase nitrogen through cation exchange. The use of alum or zeolite also reduced soluble phosphorus in the slurry. NH3 loss from fresh manure collected from lactating dairy cows was not affected by three diets containing the same level of crude protein but differing in forage source (orchardgrass silage vs. alfalfa silage) or neutral detergent fiber (NDF) content (30% vs. 35% NDF). NH3 losses from the freshly excreted manures occurred very rapidly and included the urea component plus some unidentified labile organic nitrogen sources. NH3 conservation strategies for fresh manures will have to be active within the first few hours after excretion in order to be most effective. The use of alum or zeolites as an on-farm amendment to dairy slurry offers the potential for significantly reducing NH3 emissions.

A noninvasive radiotelemetry system to monitor heart rate for assessing stress responses of bovines.

A noninvasive radiotelemetry system was developed to monitor heart rates of cows and to view and analyze data. The system was validated by comparing heart rate data of two restrained heifers collected simultaneously using telemetric and direct electrocardiogram measurements and by acquiring data over 72 h from two dry cows housed in an experimental handling facility consisting of a free-stall pen, a holding pen, a pass-through stall, and a second holding pen. Telemetric and direct measurements in response to pharmacological elevation of heart rates were essentially identical. For cows in the experimental facility, peristimulus-time histograms indexed to standing or lying showed that average heart rates for cows increased 4.0 +/- 1.4 beats/min after cows stood and decreased 4.8 +/- 1.0 beats/min after cows lay. Similarly, the average heart rate for the cow naive to the facility increased from 60 to 86 beats/min and remained elevated for 6.3 min when heart rate was indexed to maximal heart rate within +/- 3 min of entry into the pass-through stall. Heart rate for the naive cow increased consistently from around 60 to over 160 beats/min during repeated agonistic encounters between animals. Heart rate for the other cow was not affected by the encounters. These results show clearly that heart rate can be used to monitor animal anxiety.

Method to monitor the precision of milk yields recorded at individual milking stalls on daily basis.

Automated systems to record individual milk weights of cows at each milking are now common on dairy farms. These data are used by Dairy Herd Improvement Association (DHIA) for testing purposes and by farmers to monitor herd performance. Despite the importance of these data, little effort has been made to monitor the function and reliability of milking stalls and recording equipment on a daily basis. Described is an algorithm that allows for continuous monitoring of the function of individual milking stalls. The basis of the algorithm is the comparison of milk yield of individual cows over a period of days. For each cow, milk production at a selected stall is compared with production of that same cow when milked at any other stall. These ratios are averaged across all cows milked at least once in the selected stall and once at any other stall. Ideally, the mean ratio for each stall should be 1. Stalls where the mean ratio deviates from 1 by more than 5% are assumed to be malfunctioning. This algorithm was implemented as a computer program and was used to identify 3 malfunctioning stalls out of 12 at the USDA dairy facility (Beltsville, MD). The algorithm can easily be incorporated into existing management programs or used as a separate program. In either case, routine use of this algorithm can improve the reliability of DHIA test programs and potentially reduce the incidence of new mastitis infections by enabling detection of malfunctioning milking equipment.

Circadian and ultradian rhythms of body temperature and peripheral concentrations of insulin and nitrogen in lactating dairy cows.

To investigate possible circadian and ultradian periodicities for peripheral insulin and urea in lactating dairy cows, integrated 15-min blood samples were taken sequentially over 48 hr from six cows. In addition, radiotelemetry measurements of body temperature were averaged over the same 15-min periods. Cows were housed in an environmental chamber at 19 degrees C with lights on 0700 to 2300 hr; fed daily at 0900 hr; and milked at 0800 and 2000 hr. For five of the six cows, body temperature showed a circadian rhythm peaking at 2323 hr with an amplitude of 0.34 degree C. For the sixth cow, body temperature was 180 degrees out-of-phase, peaking at 1230 hr with an amplitude of 0.12 degree C. Circadian rhythms for insulin and urea were consistent for all six cows peaking at 1743 hr with an amplitude of 0.74 ng/ml for insulin and at 1034 hr with an amplitude of 3.83 mM for urea. Body temperature and insulin also displayed episodic increases that often exceeded the amplitudes of circadian rhythms. For body temperature, a broad increase in spectral power was seen for periods between 100 and 175 min; time intervals between peaks averaged around 100 min. For insulin, power spectra for individual cows universally indicated rhythms with periods of approximately 45 and 80 min; time intervals between peaks averaged approximately 65 min. For urea, almost all spectral energy was confined to the 24-hr rhythm, although there was evidence of a low-amplitude, 60-min rhythm. In conclusion, when animals are acclimated to a rigidly controlled environment and frequent blood sampling is accomplished with minimal intervention, it is possible to detect rhythms inherent in the regulation of metabolic variables.

Radiotelemetric measurement of body temperature in feedlot steers during winter.

Little is known concerning body temperature regulation in cattle under conditions of low ambient temperature. To investigate the influence of cold on body temperature regulation, core body temperatures of feedlot steers (crossbred Bos taurus) were monitored for two winters in Nebraska, from late December to mid-March in yr 1 and from late December through June in yr 2. In yr 1, radiotransmitters to monitor temperature were implanted in the peritoneum of five steers (360 kg); in yr 2, four steers (320 kg) were used. Body temperatures and ambient temperatures were recorded at 3-min intervals and were mathematically filtered to produce 120 readings/d. For yr 1 and 2, daily maximum (40.09 and 39.66 degrees C), minimum (38.78 and 38.64 degrees C), and average (39.29 and 39.06 degrees C) body temperatures were not affected by ambient temperatures. Body temperatures exhibited circadian rhythms with the minima at approximately 0800 and the maxima at approximately 1900. For both years, sharp peaks in body temperature were often seen in the evening and, for yr 2, to a lesser extent in the morning. The occurrence of peaks was normally congruent, within a 1.5-h window, across steers. Congruent peaks in the evening with peak heights of 1.05 and .77 degrees C occurred on 65 and 56% of the days in yr 1 and 2, respectively. Occurrence of congruent peaks was correlated with dusk; peaks followed dusk by 30 to 60 min. Ambient temperature also influenced the occurrence of peaks; few peaks were observed when average daily ambient temperatures were below -7.5 degrees C. The dynamic changes in body temperature throughout the day, including the peaks in body temperature after dusk, strongly suggest that thermoregulatory systems in steers respond not only to current ambient conditions, but also to more integrative measures such as day length and daily heat load.

Response of catecholamines to manual teat stimulation or machine-milking of Lacaune and Friesen dairy ewes.

Increased sympathetic activity, including peripheral release of catecholamines, has been hypothesized to inhibit the milk ejection reflex by blocking the release of oxytocin, by blocking the effect of oxytocin at the mammary gland, or both. We attempted to relate differences in milking characteristics of Lacaune and East Friesen ewes to the degree of sympathetic arousal, responses of catecholamines and oxytocin to machine-milking and to manual teat stimulation followed by milking. Peripheral concentrations of epinephrine, 67 +/- 6 pg/ml for Lacaunes and 57 +/- 5 pg/ml for Friesens, were essentially constant. Peripheral concentrations of norepinephrine averaged 682 +/- 68 pg/ml for Lacaunes and 250 +/- 54 pg/ml for Friesens. Elevated norepinephrine concentrations during the pretreatment period were associated with the inhibition of oxytocin responses. When norepinephrine concentrations during the pretreatment period exceeded 300 pg/ml for Friescens, or were rising and exceeded 700 pg/ml at initiation of the experimental period for Lacaunes, oxytocin release was inhibited. Results from this study clearly showed that basal concentrations of norepinephrine vary with breed of ewes and provided the first direct evidence that increased sympathetic activity can result in central inhibition of the milk ejection reflex.

Radiotelemetry measurement of body temperatures of feedlot steers during summer.

Heat stress is a common problem for cattle. General consequences of heat stress include increased body temperatures and reduced feed intakes. As a measure of heat stress, core body temperatures of unshaded feedlot steers (crossbred Bos taurus) were monitored from mid-June to early November in Nebraska using transmitters implanted in the peritoneum of 10 steers (initially 10 mo of age). Steers were fed at 0630 and 1430 using a finishing diet of 1.52 NEg Mcal/kg with 13% protein and 4% roughage per day and housed in two open lots with stocking densities of 15.2 or 19.3 m2/steer. Core body temperatures, ambient temperature, relative humidity, and wind speed were measured at 3-min intervals and mathematically filtered to produce 120 readings/ d. For 94 usable daily records, body temperature means (39.04 +/- .12 degrees C), maxima (39.89 +/- .21 degrees C at 1836 +/- .73 h), minima (38.33 +/- .29 degrees C at 0823 +/- .38 h), and patterns were similar among steers. As daily maximum ambient temperatures increased, minimum body temperatures decreased slightly (.04 degree C per 5 degrees C; P < .01). After daily maximum ambient temperatures reached a threshold of 25.6 degrees C, daily maximum body temperatures increased linearly with maximum ambient temperatures (.42 degree C per 5 degrees C; P < .01). Sharp peaks in body temperature were often seen in the late evening (approximately 2200) after ambient temperature had decreased to well below maximum values. These evening peaks occurred on an average of 25% of the days, had amplitudes ranging from .7 to 3.5 degrees C relative to mean daily temperatures and lasted for 1.5 h. From a practical standpoint, we suggest that producers monitor meteorological forecast of peak ambient temperatures and make special efforts, such as spraying animals, when exceptionally hot weather is predicted.

Adrenal responses of Angus x Hereford cattle to the stress of weaning.

To test adrenal responsiveness of beef cattle to stress, plasma norepinephrine, epinephrine, and cortisol responses of Angus x Hereford cattle to weaning were measured in blood samples taken from 1) eight cows and their 4- to 6-mo-old calves before and after separation and, the following day, before and after being reunited and 2) an equal number of control animals. After separation and before they were reunited, dams were returned to their original pen and calves to an adjacent pen. Experimental trials were conducted every 2 wk and included two cow/calf pairs per trial. Norepinephrine concentrations tended to increase in calves after separation, were increased (+82 pg/mL, P < .01) on the 2nd d before animals were reunited, and decreased (-88 pg/mL, P < .01) after animals were reunited. Epinephrine concentrations tended to increase in calves after separation, were increased in calves (+128 pg/mL, P < .001) and dams (+104 pg/mL, P < .001) on the 2nd d before animals were reunited, and decreased after calves (-162 pg/mL, P < .001) and dams (-101 pg/ml, P < .001) were reunited. There were no significant effects of time on catecholamine concentrations for control animals. Cortisol concentrations for treated and control animals tended to increase in sequential samples each day; however, the increases were small (approximately 1 ng/mL) and concentrations remained low (> 5 ng/mL). In conclusion, peripheral catecholamine concentrations in beef calves and epinephrine concentrations in dams increased in response to stress associated with weaning and separation.(ABSTRACT TRUNCATED AT 250 WORDS)

Circadian and ultradian rhythms of peripheral growth hormone concentrations in lactating dairy cows.

To investigate possible circadian and ultradian periodicities for plasma growth hormone in lactating dairy cows, integrated 15-min blood samples taken sequentially over 48 hr from six cows were analyzed by radioimmunoassay. The cows were housed in an environmental chamber at 19 +/- 0.5 degree C, 50% relative humidity, and 16 hr of light and 8 hr of darkness (lights on at 0700 hr); fed daily at 0900 hr; and milked at 0800 and 2000 hr. Peripheral concentrations of growth hormone for all six cows exhibited sinusoidal circadian rhythms with average minima of 4.1 ng/ml at 1820 hr and maxima of 5.3 ng/ml at 0630 hr. Estimated periods of ultradian rhythms for individual cows by spectral analysis, peak identification, and fitting cosine functions using least squares were 71 to 83 min for all cows. No direct relationship between ultradian peaks and milking or feeding was apparent. In conclusion, a circadian rhythm and an ultradian rhythm with a period around 80 min are probably intrinsic to mechanisms regulating peripheral growth hormone concentrations in the lactating dairy cow.

Circadian and ultradian rhythms of peripheral prolactin concentrations in lactating dairy cows.

To investigate possible circadian and ultradian periodicities for peripheral prolactin in lactating dairy cows, integrated 15-min blood samples taken sequentially over 48 h from six cows were analyzed by radioimmunoassay. Cows were housed in an environmental chamber at 19 degrees C with lights on 0700 to 2300, fed daily at 0900, and milked at 0800 and 2000. Peripheral concentrations exhibited sinusoidal circadian rhythms. For two cows, a nadir of 15 ng/ml occurred around 1900 and a zenith of 29 ng/ml around 0730. For four cows, circadian rhythms were 180 degrees out of phase, with a nadir of 28 ng/ml around 0840 and a zenith of 40 mg/ml around 1920. Ultradian peaks, which varied in amplitude within and among animals, showed no consistent relationship with either milking or feeding. Spectral analyses indicated rhythms around 100 min for the first two animals and 150 min for the four animals. In conclusion, there were no simple relations between prolactin rhythms and lights on or lights off. Further, results suggest that changes in the phase of circadian rhythms may be associated with changes in the periods of ultradian rhythms.

Circadian and ultradian rhythms of plasma thyroid hormone concentrations in lactating dairy cows.

Circadian and ultradian rhythms of thyroxine and triiodothyronine were analyzed by radioimmunoassay on integrated 15-min blood samples collected for 48 h from six lactating dairy cows. Body temperatures were recorded every 1.4 min using radiotelemetry. The cows were housed in an environmental chamber at 19.0 +/- 0.5 degrees C (lights on between 0700 and 2300 h), fed daily at 0900 h, and milked at 0800 and 2000 h. Mean concentrations of plasma triiodothyronine and thyroxine exhibited similar circadian rhythms with minima (0500-1300 h) and maxima (1700-0200 h) separated by 12 h. Triiodothyronine peaked 2 h before thyroxine. The thyroxine circadian rhythm preceded a circadian body temperature rhythm by 2 h. Superimposed on the circadian rhythm was an ultradian rhythm with a 90-min period that was 15-20% of the mean thyroid hormone concentrations. Peak thyroxine and triiodothyronine concentrations were 50 +/- 2 and 1.58 +/- 0.17 ng/ml, and minimal concentrations were 42 +/- 2 and 0.94 +/- 0.17 ng/ml, respectively. Our data indicate that peripheral triiodothyronine is regulated independently of peripheral thyroxine concentration.

Circadian and ultradian rhythms of peripheral cortisol concentrations in lactating dairy cows.

To investigate possible circadian and ultradian periodicities for plasma cortisol in lactating dairy cows, integrated 15-min blood samples taken sequentially over 48 h from six cows were analyzed by radioimmunoassay. The cows were housed in an environmental chamber at about 19 degrees C, 50% relative humidity, and 16 h of light and 8 h of darkness (lights on at 0700 h); fed daily at 0900 h; and milked at 0830 and 2000 h. Peripheral concentrations of cortisol for all six cows exhibited weak circadian rhythms--average minima were 3.1 ng/ml at 1800 h, maxima were 4.5 ng/ml at 0530 h--and strong ultradian rhythms with periods around 120 min. Peak to trough amplitudes of ultradian rhythms, 1 to 17 ng/ml, varied among and within cows over time. No direct relation between ultradian peaks and milking or feeding was apparent. Spectral analysis indicated that most power (variation) was centered around 1 cycle every 2 h, i.e., around a period of 120 min. This very strong, easily observable, ultradian rhythm indicates that an oscillation with a period around 120 min is probably intrinsic to mechanisms regulating peripheral glucocorticoid concentrations.

Radiotelemetry temperature responses of mammary gland and body to intramammary injection of Escherichia coli endotoxin or Streptococcus agalactiae in lactating dairy cows.

To investigate the feasibility of using changes in body or mammary temperature to detect mastitis, radiotransmitters were implanted midway between rear udder quarters and in the peritoneal cavity of 5 Holstein cows (1 to 3 months in lactation) housed in an environmental chamber (16 +/- 2 C; lights on 7:00 AM to 11:00 PM). After a 6-week control period, Escherichia coli endotoxin (0.5 mg) was injected after the morning milking into left rear teat cisterns via the teat canal. Wisconsin mastitis test score and somatic cell count in all quarters increased significantly (P < 0.01) by the next milking. Effects were greatest in the endotoxin-exposed quarters. Milk yields for all quarters decreased significantly (P < 0.01) by the first milking after endotoxin injection. Udder and body temperatures at milkings were similar and were not affected by treatment. When temperatures were averaged for the 5 cows for each of 120 time points/d, average temperatures, relative to time of injection of endotoxin, were increased by 0.5 C above baseline at 2.75 hours, peaked at +2.9 C at 6.50 hours, and remained high through 9.25 hours after injection. Power spectra calculated for individual cows on a daily basis universally indicated an increase in power at low frequencies on the day of injection. Subsequently, Streptococcus agalactiae (200 colony-forming units) was injected into right rear teat cisterns. Wisconsin mastitis test score increased at the second milking after injection. Cell count and quarter milk yield decreased by the third milking. As with endotoxin, injection of S agalactiae could not be detected via a change in temperature at milkings.(ABSTRACT TRUNCATED AT 250 WORDS)

Teat stimulation-induced oxytocin and catecholamine release in pregnant and lactating Holstein heifers.

The effect of manual teat stimulation (milking paradigm) on release of oxytocin, epinephrine and norepinephrine was studied in (1) 15 heifers at 100, 150, 200 and 250 d of gestation and at 30 and 90 d of lactation (during machine milking) and (2) simultaneously in six heifers at less than 100 d and in six heifers at greater than 200 d of gestation. Oxytocin responses to teat stimulation, including peak heights and area under the response curves, at 150, 200 and 250 d or at greater than 200 d of gestation were similar and were significantly greater than responses at 100 d or at less than 100 d. Responses to milking were lower at 90 d compared to responses at 30 d. Catecholamines were measured only during gestation and were generally not affected by teat stimulation. Epinephrine levels were significantly higher at 200 and 250 d compared to levels at 100 and 150 d. Baseline oxytocin concentrations and responses to teat stimulation were greatest at 150 d of gestation when epinephrine levels were still low, suggesting that stimulatory mechanisms responsible for the release of oxytocin develop and/or are expressed prior to the development of inhibitory sympathetic mechanisms. For norepinephrine, linear analyses did not show significant responses to teat stimulation overall. However, elevated norepinephrine responses (greater than .2 pmol/ml) following teat stimulation were seen in 28 of 51 trials, and large oxytocin responses (greater than 75 pg/ml/min) were seen predominantly only when norepinephrine responses were low (less than = .2 pmol/ml).

Rhythms in cholesterol, cholesteryl esters, free fatty acids, and triglycerides in blood of lactating dairy cows.

Blood samples from six lactating dairy cows were analyzed to determine whether circulating neutral lipids exhibit rhythmic variations. Plasma neutral lipids were measured by quantitative TLC on every fourth integrated 15-min blood sample taken over 48-h periods. Cows were housed in an environmental chamber at 20 degrees C with 16 h light:8 h dark (lights on at 0700 h), fed daily at 0900 h, and milked at 0830 and 2000 h. Other variables monitored included: body temperature, ammonia nitrogen, urea nitrogen, glucose, triiodothyronine, thyroxine, somatotropin, insulin, cortisol, and prolactin. Mean concentrations of cholesterol, cholesteryl esters, free fatty acids, and triglycerides were 21.4, 175.4, 3.1, and 6.3 mg/dl, respectively. Visual and power spectral analysis of the pulsatile fluctuations in lipids indicated rhythms with periods of 2 to 3 h. Amplitudes of rhythms for free fatty acids and triglycerides were 60% of mean concentrations and for cholesterol and cholesteryl esters were 20% of mean concentrations. The presence of these rhythms was conserved when data were averaged across time by cow. However, because of nonstationary conditions, rhythms identified by spectral analysis were not statistically significant. There was no evidence of circadian patterns in circulating neutral lipid components. All other metabolic and hormonal variables except cortisol exhibited distinct circadian rhythms.

Body temperature of dry cows on pasture: environmental and behavioral effects.

To investigate interrelations among factors potentially affecting body temperature, we observed body temperature, activity, and relative aggregation of 3 to 5 dry cows from a herd of 12 to 15 every 20 min. Ambient conditions were noted every 1 h and fly counts every 3 h. Observations during daylight for 5 d with and without shade indicated effects of shade were minimal and were extended to 24 h without shade at six weekly intervals. Cows lie down during the evening, grazed near sunrise and sunset, and alternated standing and grazing during the day. The aggregation index was greatest during periods of standing and least during grazing; however, it remained constant across periods of similar activity. Cows shifted activity from grazing to standing during periods of increased fly activity. Ambient temperatures ranging from 13 to 35 degrees C had little influence on measured variables. Body temperatures showed one to three daily dips (greater than 1 degree C) and a hysteresis pattern. Almost all dips were associated with transitions from lying to grazing. Body temperature was constant in the morning, rose to a higher level in the afternoon, and remained high until the next morning.

Failure of cortisol injected prior to milking to inhibit milk ejection in dairy cattle.

To test the potential for cortisol to inhibit milk ejection directly, 18 Holstein cows were divided equally into control and treatment groups based on milk yields. For treated animals, a single injection of cortisol was made into the saphenous vein 15 min before milkings. Increasing amounts of cortisol (0, 25, 50, and 100 mg) were injected for one morning and one evening milking, with the exception that the treated cows received only one 100 mg injection. Control animals received injections of 0.9% (w/v) NaCl. Cortisol injections had no effect on milk yields. However, a potential inhibitory mechanism might involve a delay, perhaps due to the necessity of synthesizing a regulatory protein. Therefore, to test the potential for increased cortisol over a period of hours to inhibit milk ejection, six of the nine cows in the treatment group were injected with 100 mg of cortisol at 3.25, 2.25, 1.25 and 0.25 h before sequential morning and evening milkings. In blood samples taken 1 min before and after injections, base-line cortisol concentrations averaged 10.2 mg/ml; after injection they were 984.1 ng/ml, and before subsequent injections they were 37.6 ng/ml. Again cortisol injections had no effect on milk yields.