Technical note: effect of corticotropin-releasing hormone on adrenocorticotropic hormone and cortisol in steers

The objective of this study was to determine an appropriate exogenous dose of bovine corticotropin-releasing hormone (bCRH) to stimulate the physiological effects of the hypothalamic-pituitary-adrenal axis in steers as a method to test the sensitivity of the pituitary and adrenal gland. Twenty 14-mo-old Holstein-Friesian steers were blocked by weight (443.7+/-2.5 kg) and randomly allotted to receive either saline (control) or bCRH (0.1, 0.3, 1.0, or 1.5 microg/kg BW). Animals were housed in a slatted-floor facility (n = 5 per pen). Indwelling jugular catheters, for both the administration of bCRH and blood collection, were fitted on d -1 of the experiment. Saline and bCRH were administered i.v. at time 0. Serial blood samples were collected at -15, 0, 15, 30, 45, 60, 75, 90, 105, 120, 135, 150, 165, and 180 min relative to time 0. Following administration of 0.1 microg of bCRH/kg BW, the peak ACTH response was not significantly different from pretreatment baseline concentrations (mean concentrations as measured at -15 and 0 min before bCRH administration). Mean ACTH concentrations from 0 to 180 min following 0.1 microg of bCRH/kg BW were not significantly different (P = 0.177) from controls. Administration of 0.3, 1.0, and 1.5 microg of bCRH/kg BW increased (P < 0.05) peak ACTH above pretreatment concentrations, and mean ACTH from 0 to 180 min for these treatments were greater (P < 0.05) than for controls. Peak cortisol responses to all bCRH treatments were greater (P < 0.05) than those to pretreatment concentrations. Mean cortisol concentrations from 0 to 180 min were greater (P < 0.05) in all bCRH-treated steers than in controls, but there were no significant differences among the bCRH treatments. The ratio of mean cortisol to mean ACTH for all bCRH doses tested differed (P < 0.05) from control values, indicating reactivity of the adrenals. In conclusion, bCRH challenge may be a useful method for testing the sensitivity of the hypothalamic-pituitary-adrenal axis in steers subjected to stressful husbandry conditions, and a minimum dose of 0.3 microg of bCRH/kg BW is required to stimulate physiological effects of stressor hormones.     

Adrenocorticotropic hormone and cortisol in calves after corticotropin-releasing hormone.

The aim for this study was to analyze responsiveness of the hypothalamo-pituitary-adrenocortical axis to exogenous bovine corticotropin-releasing hormone (bCRH) in calves. Two dose-response studies were carried out, using either bCRH alone (dose rates of 0, .01, .03, and .1 microg bCRH/kg live weight) or in combination with arginine-vasopressin (bCRH:AVP, 0:0, .1:.05, .5:.25, and 1:.5 microg kg live weight). The bCRH was administered i.v. to calves (n = 5 to 7 per dose) housed individually or in groups. Serial blood samples were obtained from before to 300 min after injection and analyzed for plasma ACTH and cortisol concentrations. The lowest bCRH dose that produced a response in all calves was .1 microg/kg. In the experiment using bCRH with AVP, increasing the bCRH dose from .1 to 1 microg/kg resulted in an increase in peak ACTH concentration (321 vs. 2,003 pg/mL) but did not significantly affect the peak cortisol concentration (37 vs. 40 ng/mL). The time to reach the peak cortisol concentration increased with the dose of bCRH with AVP (from 38 to 111 min). The ACTH and cortisol concentrations determined at any time between 20 and 90 min after bCRH injection were correlated to the integrated responses calculated as areas under the ACTH and the cortisol curves (r between .61 and .99, P<.05). In comparison with results from studies in humans, pigs, and sheep, our data showed that the pituitary of calves seems less sensitive to CRH than that of other mammals, despite a greater capacity to produce ACTH. Moreover, the calf's adrenals seem to have a lower capacity to produce cortisol than adrenals of other mammals. As in other species, it seems that AVP enhances the release of ACTH and cortisol. For CRH challenge to be used in calves, we suggest injecting at least .1 microg of bCRH/kg live weight either with or without AVP and taking several blood samples before injection and between 20 and 90 min after injection.     

Pituitary adenylate cyclase-activating polypeptide induces secretion of growth hormone in cattle.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a hypothalamic neuropeptide that stimulates release of growth hormone (GH) from cultured bovine anterior pituitary gland cells, but the role of PACAP on the regulation of in vivo secretion of GH in cattle is not known. To test the hypothesis that PACAP induces secretion of GH in cattle, meal-fed Holstein steers were injected with incremental doses of PACAP (0, 0.1, 0.3, 1, 3, and 10 microg/kg BW) before feeding and concentrations of GH in serum were quantified. Compared with saline, injection of 3 and 10 microg PACAP/kg BW increased peak concentrations of GH in serum from 11.2 ng/ml to 23.7 and 21.8 ng/ml, respectively (P < 0.01). Peak concentrations of GH in serum were similar in steers injected with 3 or 10 microg PACAP/kg BW. Meal-fed Holstein steers were then injected with 3 microg/PACAP/kg BW either 1 hr before feeding or 1 hr after feeding to determine if PACAP-induced secretion of GH was suppressed after feeding. Feeding suppressed basal concentrations of GH in serum. Injection of PACAP before feeding induced greater peak concentrations of GH in serum (19.2 +/- 2.6 vs. 11.7 +/- 2.6 ng/ml) and area under the response curve (391 +/- 47 vs. 255 +/- 52 ng. ml(-1) min) than injection of PACAP after feeding, suggesting somatotropes become refractory to PACAP after feeding similar to that observed by us and others with growth hormone-releasing hormone (GHRH). We concluded that PACAP induces secretion of GH and could play a role in regulating endogenous secretion of GH in cattle, perhaps in concert with GHRH.     

Effect of ACTH and CRH on plasma levels of cortisol and prostaglandin F2alpha metabolite in cycling gilts and castrated boars

The present study was designed to evaluate the effects of synthetic ACTH (1-24, tetracosactid) and porcine CRH on the plasma levels of cortisol and PGF2alpha metabolite in cycling gilts (n = 3) and castrated boars (n = 3). The experiments were designed as crossover studies for each gender separately. Each animal received, during three consecutive days; 1) ACTH (Synacthen Depot) at a dose of 10 microg/kg body weight in 5 ml physiological saline, 2) porcine CRH at a dose 0.6 microg/kg body weight in 5 ml physiological saline or 3) physiological saline (5 ml). The test substances were administered via an indwelling jugular cannula in randomized order according to a Latin square. The administration of ACTH to cycling gilts resulted in concomitant elevations of cortisol and PGF2alpha metabolite with peak levels reached at 70.0 +/- 10.0 and 33.3 +/- 6.7 min, respectively. Similarly, the administration of ACTH to castrated boars resulted in concomitant elevation of cortisol and PGF2alpha metabolite with peak levels reached at 60.0 +/- 0.0 and 20.0 +/- 0.0 min, respectively. Cortisol peaked at 20 min after administration of CRH in both cycling gilts and castrated boars with maximum levels of 149.3 +/- 16.5 nmol/l and 138.3 +/- 10.1 nmol/l, respectively. It can be concluded that administration of synthetic ACTH (tetracosactid) to pigs caused a concomitant elevation of cortisol and PGF2alpha metabolite levels in both cycling gilts as well as castrated boars. The administration of CRH to pigs resulted in an elevation of cortisol levels in both cycling gilts and castrated boars. Conversely, PGF2alpha metabolite levels were not influenced by the administration of CRH either in cycling gilts or in castrated boars.     

Adrenocorticotropin and cortisol levels in the plasma of bovine fetuses and neonates

Plasma ACTH and cortisol levels in the bovine fetuses over the period of 5 to 9 months of gestation and in the neonates immediately after birth and at 5 days old were studied. In the bovine fetuses, the plasma ACTH levels ranged from 60.8 +/- 17.8 to 71.3 +/- 19.7 pg/ml over the period of 5 to 7 months of gestation. It increased rapidly to 239.2 +/- 261.5 pg/ml at 8 months and to 406.9 +/- 409.4 pg/ml at 9 months of gestation. In the neonates immediately after birth it decreased to 182.3 +/- 110.7 pg/ml. The plasma cortisol levels ranged from 3.23 +/- 2.12 to 3.85 +/- 2.52 ng/ml over the period of 5 to 8 months of gestation and increased to 8.10 +/- 4.88 ng/ml at 9 months of gestation. It then increased rapidly to 88.35 +/- 42.78 ng/ml in the neonates immediately after birth. The correlation between plasma ACTH and cortisol levels in the fetuses of 5 to 7 months of gestation was not significant, but in the fetuses of 8 and 9 months of gestation and neonates were significant. However, especially immediately after birth, the increase in plasma cortisol occurred without a concomitant rise in plasma ACTH. According to these findings, it is suggested that the pituitary-adrenocortical axis in bovine fetus matures in the later stage of gestation and an increase of sensitivity in the fetal adrenal gland to ACTH may serve as a trigger for the onset of parturition.     

Plasma concentrations of luteinizing hormone and adrenocorticotropic hormone in blood collected during the luteal and follicular phases of the estrous cycle in cows.

Luteinizing hormone (LH) and ACTH concentrations were measured in plasma from 7 cows to determine whether ACTH secretion changes with the phase of the estrous cycle, and to determine whether any ACTH peaks are associated with LH peaks. Blood was collected every 5 minutes for 190 minutes during the luteal and follicular phases of the estrous cycle. Radioimmunoassays were used to measure ACTH and LH in plasma. Mean concentration of ACTH in all cows did not differ significantly between luteal (35.1 +/- 8.0 pg/ml) and follicular (37.5 +/- 9.4 pg/ml) phases of the estrous cycle. Mean concentration of luteal-phase LH of all cows (2.0 +/- 1.1 ng/ml) was significantly (P less than 0.01) lower than mean concentration of follicular-phase LH (5.4 +/- 1.6 ng/ml). Frequency of peaks in ACTH concentration was low during the sampling period. Mean number of luteal-phase ACTH peaks (0.29 +/- 0.49) was not significantly different from that of follicular-phase samples (0.43 +/- 0.530). Unlike ACTH, mean frequency of LH peaks was significantly (P less than 0.05) higher in plasma from cows in the follicular phase of the estrous cycle (2.9 +/- 0.7), compared with that from cows in the luteal phase (0.29 +/- 0.49).     

Effect of spray-dried plasma and lipopolysaccharide exposure on weaned pigs: II. Effects on the hypothalamic-pituitary-adrenal axis of weaned pigs

A study was conducted with 20 weaned barrows (14 d, 4.98 +/- 0.21 kg) to determine the effect of feeding spray-dried plasma (SDP) after weaning on the pig's stress response to a lipopolysaccharide (LPS) challenge. After weaning, pigs were fed a diet containing 0 or 7% SDP for 7 d. On d 6 after weaning, all pigs were nonsurgically fitted with a jugular catheter. On d 7 after weaning, the pigs were given i.p. injections of either saline or LPS (150 microg/kg BW) followed by serial blood collection every 15 min for a 3-h period. Following the 3-h blood collection, all pigs were killed and tissue was collected for mRNA analysis. Pig weight on d 7 after weaning was not affected by dietary treatment (P > 0.21). Pigs fed the diet with SDP had lower (P < 0.05) levels of hypothalamic corticotropin-releasing hormone (CRH) mRNA, pituitary gland CRH receptor mRNA, and adrenal gland adrenocorticotropin-releasing hormone (ACTH) receptor mRNA. Dietary treatment did not affect pituitary gland proopiomelanocortin (POMC) mRNA. No effect of LPS treatment was observed in any of the mRNA levels examined. For both serum ACTH and cortisol, there was a significant diet x LPS treatment interaction (P < 0.01) such that both the ACTH and cortisol responses to the LPS challenge were greater in the pigs fed the diet with SDP than in the pigs fed the diet without SDP. For pigs given the saline injection, diet did not affect basal serum cortisol concentration; however, basal serum ACTH concentration was lower in those pigs fed the diet with SDP (P < 0.0001). A diet x LPS treatment interaction (P < 0.024) was observed for adrenal gland mRNA expression for steroidogenic acute regulatory (StAR) protein such that the LPS-induced increase in StAR mRNA was greater in the pigs fed SDP than in pigs fed the diet without SDP. These results demonstrate that pigs fed a diet with SDP have an increased activation of the pituitary-adrenal axis following an LPS challenge compared to pigs fed a diet without SDP.     

Cortisol-induced inhibition of ACTH secretion in adult sheep

Previous results from this laboratory have demonstrated that in preterm fetal sheep (117-131 days gestation), stimulated ACTH secretion is highly sensitive and that in term fetal sheep (129-143 days), stimulated ACTH secretion is insensitive to the negative feedback effects of cortisol. The purpose of this study was to quantitate cortisol negative feedback inhibition of stimulated ACTH secretion in adult sheep. Adult, conscious, nonpregnant ewes, chronically prepared with carotid arterial loops, were infused intravenously with vehicle or cortisol at 4 different rates (denoted Groups I, II, III, and IV) for 5 hours. These infusions increased total and unbound plasma cortisol concentrations within the range observed after stimulation of the hypothalamus-pituitary-adrenal axis. One hour after termination of the cortisol or vehicle infusions, ACTH secretion was stimulated by intravenous infusion of sodium nitroprusside for 10 min at a rate of 20 micrograms/kg.min. Cortisol infusions suppressed ACTH responses to nitroprusside in a dose-related manner. After vehicle infusion, nitroprusside increased plasma ACTH to 735 +/- 229 pg/ml. After cortisol infusions, nitroprusside increased plasma ACTH to 292 +/- 63, 101 +/- 30, 73 +/- 12, and 67 +/- 24 in Groups I, II, III, and IV, respectively. Overall, there was a significant negative exponential relationship between plateau plasma cortisol concentration during the cortisol or vehicle infusion and the peak plasma ACTH concentration during the response to nitroprusside infusion (r = -0.81). The highest rate of cortisol infusion increased total and unbound plasma cortisol concentrations to 40.1 +/- 5.7 and 19.5 +/- 5.9 ng/ml and completely suppressed the subsequent ACTH response to nitroprusside.     

Plasma and urine concentrations of marbofloxacin following single subcutaneous administration to cats

The pharmacokinetic properties of marbofoxacin, a third generation fluoroquinolone, were investigated in 12 healthy adult cats after single subcutaneous (SC) administration of 2 mg/kg BW (Part I, n=8 cats) and 4 mg/kg BW (Part II, n=4 cats). In each part of the study blood and urine samples were collected before treatment and thereafter for 5 days. The plasma and urine concentrations of marbofloxacin were determined by HPLC with UV detection. Pharmacokinetic calculations were performed for each treated animal using an open one-compartment-model with first-order elimination after SC dosing. Marbofloxacin in plasma (means): Maximum concentrations (Cmax) of about 1.2 and 3.0 microg/ml were measured 2.3 and 4 hours (tmax) after dosing of 2 and 4 mg/kg BW, respectively. Elimination from the body was low with a total clearance (Cl/F) of approximately 0.1 l/h/kg for both dosages. The half-life (t 1/2) for this process was calculated with 8-10 hours. AUC increased almost proportional when doubling the dose, i.e., 19.77 +/- 6.25 microg * h/ml (2 mg/kg BW) and 51.26 +/- 11.83 microg * h/ml (4 mg/kg BW). Plasma kinetics measured were in accordance with data from literature. Marbofloxacin in urine (means): Maximum drug concentrations were detected 4 and 8 hours after dosing with 70 microg/ml (2 mg/kg BW) and 160 microg/ml (4 mg/kg BW), respectively. Inhibitory effects of the urinary matrix on the antimicrobial activity of the drug were taken into account when performing PK/PD calculations. However, a concentration-dependent bactericidal activity (Cmax/MIC > 8-10) which is claimed for fluoroquinolones was sufficiently met with focus on Escherichia (E.) coli (MIC90 0.5 microg/ml). In the same matrix a threshold value of 1.0 microg/ml was undercut 82 and 116 hours after SC dosing, respectively. Hence, a time-dependent bacteria killing kinetic (T > MIC) which may be of relevance for some Gram-positive germs like Staphylococcus spp. (MIC90 1.0 microg/ml) should be covered, too.     

Adrenocortical response to ACTH in Angora and Spanish goat wethers.

Angora goats do not cope well with stress compared with goats of other breeds. Our hypothesis that this involves subclinical primary hypoadrenocorticism associated with low cortisol release in response to ACTH stimulation was tested by measuring adrenocortical response (plasma cortisol) in six Spanish (37 +/- 2 kg BW) and six Angora wethers (39 +/- 3 kg BW) under simulated acute and chronic ACTH challenges. In Exp. 1 (acute ACTH challenge), wethers were dosed i.v. with high (2.5 IU/kg BW) or low (.4 IU/kg BW) quantities of ACTH. In Exp. 2 (chronic ACTH challenge), ACTH at the rate of .015 IU/(kg BW x min) or saline (.15 M NaCl) was infused i.v. at 15 mL/h for 6 h. The mean baseline plasma cortisol concentration before ACTH stimulation was similar (P > .05) between Angora and Spanish goats in Exp. 1 (averaged over days) and in Exp. 2. The cortisol concentration response area (ng/ (mL x min) x 10(-3)) above the baseline was similar (P > .05) between Angora and Spanish goats during low (7.6 +/- .5 and 9.0 +/- 1.7, respectively) and high (12.8 +/- 1.0 and 16.0 +/- 1.8, respectively) levels of acute ACTH challenge (Exp. 1) and during chronic ACTH challenge (45.1 +/- 5.9 and 41.8 +/- 7.3, respectively; Exp. 2). In conclusion, these data indicate that, under the conditions of this study, adrenocortical responsiveness to ACTH stimulation is not different between Angora and Spanish goat wethers and, thus, may not contribute to stress susceptibility in Angora goats.     

Birth by caesarian section alters postnatal function of the hypothalamic-pituitary-adrenal axis in young pigs

Eight crossbred sows were selected for the present study (n = 4 vaginal delivery and n = 4 Caesarian section [C-section]). Gestation length did not differ between vaginal delivery and C-section pigs (113.6 +/- .1 and 113.2 +/- .3 d, respectively; P > .16). Blood and tissue samples from 38 pigs were collected at birth. All remaining pigs were sustained with vaginal-delivery sows until 2 wk of age (n = 39). At 2 wk of age, remaining pigs were catheterized for blood sample collection to assess pituitary-adrenal responsiveness to an injection of corticotropin-releasing hormone (CRH; 10 microg/kg). Blood samples were collected at -30, -15, 0, 5, 10, 20, 40, 60, and 90 min; pigs received CRH or saline at time 0. Pigs were killed and tissue samples were collected immediately following the last blood sample. Serum concentrations of ACTH and cortisol (CS) were measured. Total RNA was isolated from the pituitary and adrenal glands to evaluate gene expression for mRNA specific for pro-opiomelanocortin (POMC) and for the ACTH receptor. Centrifuged clot:blood ratio was reduced in the C-section pigs at birth (P < .001) and at 2 wk of age (P < .043), compared with the vaginally delivered pigs. Basal serum concentration of ACTH was greater in C-section than in vaginally delivered pigs at birth (P = .01) but did not differ at 2 wk of age (P = .42). Basal serum concentration of CS was not different at birth (P = .86) but was greater in C-section pigs than in vaginally delivered pigs at 2 wk of age (P < .04). Serum concentration of ACTH was not different (P > .99) between the two groups of pigs following the CRH challenge. However, serum concentration of CS was greater (P < .05) in C-section pigs following the CRH challenge. Expression of ACTH receptor mRNA tended to be greater in C-section pigs at birth (P < .063) and at 2 wk of age (P < .016). There was no difference in POMC mRNA between treatments (P > .73); however, there was a developmental increase (P < .001) from birth to 2 wk of age. These data indicate that the birth process plays an important role in postnatal function of the hypothalamic-pituitary-adrenal axis in young pigs.     

Effects of sample handling on adrenocorticotropin concentration measured in canine plasma, using a commercially available radioimmunoassay kit.

A commercially available radioimmunoassay (RIA) kit for measurement of human adrenocorticotropin (hACTH) was validated for use in dogs. Assay sensitivity was 3 pg/ml. Intra-assay coefficient of variation (x 100; CV) for 3 canine plasma pools was 3.0 (mean +/- SD, 33 +/- 0.99 pg/ml), 4.2 (71 +/- 2.4 pg/ml) and 3.7 (145 +/- 3.7 pg/ml) %. Interassay CV for 2 plasma pools measured in 6 assays was 9.8 (37 +/- 3.6 pg/ml) and 4.4 (76 +/- 3.4 pg/ml) %, respectively. Dilutional parallelism was documented by assaying 2 pools of canine plasma at 3 dilutions and correcting the measured result for dilution. Corrected mean concentrations for the first pool were 33 (+/- 0.99), 36 (+/- 4.3), and 33 (+/- 6.8) pg/ml; corrected mean concentrations for the second pool were 145 (+/- 5.4), 141 (+/- 10.8) and 125 (+/- 3.4) pg/ml. Recovery of 1-39hACTH added to canine plasma (6.25, 12.5, 25.0, 50.0, and 100.0 pg/ml) was linear and quantitative (slope = 0.890, R2 = 0.961). To test whether anticoagulant or the protease inhibitor, aprotinin, influences ACTH concentration in canine plasma, ACTH was measured in canine blood collected in 4 tubes containing anticoagulant: heparin (H), heparin + 500 kallikrein inhibitor units (KIU) of aprotinin/ml (HA), EDTA (E), and EDTA + aprotinin (EA). Plasma ACTH concentration was the same when samples containing H and HA, or HA and E were compared, and was significantly (P less than 0.01) lower in samples containing EA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Circadian profile and production rate of melatonin in the cow

Five adult pasture-bred French Friesian cows were used to qualify the circadian profile and characterized pulsatility of plasma melatonin, and to estimate melatonin secretion rate, around the summer solstice. Plasma concentrations of melatonin were low (5 pg/ml) during the photophase, began to rise at sunset (light intensity less than 20 lx) and reached a maximum (about 90 pg/ml) in the middle of the scotophase. The mean amplitude of peaks was 48.67 +/- 23.01 pg/ml, their mean duration was 32.30 +/- 21.50 min and the frequency was 1.5 +/- 0.3 peak/hr during the secretory period (537 +/- 42.3 min). The plasma clearance (ClB) was 0.0247 +/- 0.0013 1/kg per min, the steady state volume of distribution (Vss) was 1.404 +/- 0.225 1/kg, the elimination half life (t1/2 beta) was 66.66 +/- 11.30 min, the mean residence time was 51.37 +/- 9.92 min and the mean production rate was 399.9 +/- 57.37 ng/kg per 24 hr. These results support the concept of linearity for melatonin kinetics in cattle and the plasma clearance value suggest a first-pass hepatic effect.     

Plasma profiles of ivermectin in horses following oral or intramuscular administration

A study was undertaken in order to evaluate and compare ivermectin's (IVM) plasma disposition kinetic parameters after oral or intramuscular (IM) administration in horses. Ten clinically healthy adult horses, weighing 380-496 kg body weight (BW), were allocated to two experimental groups of five horses. Group I, was treated with an oral paste formulation of IVM at the manufacturer's recommended dose of 0.2 mg/kg BW. Group II, was treated IM with an injectable 1% formulation of IVM at a dose of 0.2 mg/kg BW. Blood samples were collected by jugular puncture at different times between 0.5 h and 75 days post-treatment. After plasma extraction and derivatization, samples were analysed by high-performance liquid chromatography with fluorescence detection. A computerized kinetic analysis was performed, and data were compared using the Wilcoxon signed rank test. The parent molecule was detected in plasma between 30 min and either 20 (oral) or 40 (IM) days post-treatment. Significant differences were found for the time corresponding to peak plasma concentrations (tmax) and for absorption half-life. Peak plasma concentrations (Cmax) of 51.3 +/- 16.1 ng/ml (mean +/- SD) were obtained after oral administration and of 31.4 +/- 6.0 ng/ml for the IM route. The values for area under concentration-time curve were 137.1 +/- 35.9 ng day/ml for the group treated orally, and 303.2 +/- 4.3 ng day/ml for the IM treated group. The mean plasma residence times were 4.2 +/- 0.4 and 8.9 +/- 0.7 days for oral and IM-treated groups, respectively. The results of this study show that the route of administration considerably affects the disposition of IVM. A significant difference in bioavailabilty and half-life of elimination of IVM was observed after IM administration compared with oral administration. A close relationship between pharmacokinetic profiles and the clinical efficacy of IVM was established.     

Transport stress modulates adrenocorticotropin secretion from peripheral bovine lymphocytes

The influence of transport stress on the secretion of adrenocorticotropin (ACTH) from peripheral bovine lymphocytes was evaluated by exposing cows to short- (30 min) or long-term (14 h) transport. After transporting animals for 14 h they were given a mandatory rest for 24 h in two different situations, either by off-loading them and allowing them to rest in a stall or by keeping them in the truck. Blood samples were withdrawn before and after transport and after the rest period in long-term transported cows and before and after transport in cows transported for 30 min. Peripheral blood lymphocytes were separated and cultured for 72 h in serum-free medium. Adrenocorticotropin was measured using highly sensitive and specific immunoradiometric assay in culture supernates. We noticed no effect of short-term transport on ACTH secretion from lymphocytes. The ACTH concentration in animals transported for 14 h increased (P < or = 0.01) from 4.72 +/- 0.48 pg x mL(-1)/2 x 10(6) lymphocytes before the transport to 8.24 +/- 1.40 pg x mL(-1) directly after the transport. When animals were off-loaded and rested in a stall for 24 h, ACTH secretion from cultured lymphocytes returned to the basal value of 4.24 +/- 0.31 pg x mL(-1), whereas the animals rested in the truck had ACTH levels of 8.9 +/- 1.43 pg x mL(-1). Phytohemagglutinin, a plant lectin that stimulates lymphocytes, did not affect the lymphocytic ACTH secretion in this study. Heart rate and rectal temperature measured telemetrically increased in cows directly after 14 h of transport but decreased to pretransport values in cows rested for 24 h in cows rested in stalls and those rested inside the truck. This experiment is the first to show lymphocytic ACTH secretion in cows, and the results indicate that ACTH secretion from peripheral lymphocytes could be used as a reliable measurement in stress studies.     

Pharmacokinetics of orally administered ibuprofen in African and Asian elephants (Loxodonta africana and Elephas maximus).

The pharmacokinetic parameters of S(+) and R(-) ibuprofen were determined in 20 elephants after oral administration of preliminary 4-, 5-, and 6-mg/kg doses of racemic ibuprofen. Following administration of 4 mg/kg ibuprofen, serum concentrations of ibuprofen peaked at 5 hr at 3.9 +/- 2.07 microg/ml R(-) and 10.65 +/- 5.64 microg/ml S(+) (mean +/- SD) in African elephants (Loxodonta africana) and at 3 hr at 5.14 +/- 1.39 microg/ml R(-) and 13.77 +/- 3.75 microg/ml S(+) in Asian elephants (Elephas maximus), respectively. Six-milligram/kilogram dosages resulted in peak serum concentrations of 5.91 +/- 2.17 microg/ml R(-) and 14.82 +/- 9.71 microg/ml S(+) in African elephants, and 5.72 +/- 1.60 microg/ml R(-) and 18.32 +/- 10.35 microg/ml S(+) in Asian elephants. Ibuprofen was eliminated with first-order kinetics characteristic of a single-compartment model with a half-life of 2.2-2.4 hr R(-) and 4.5-5.1 hr S(+) in African elephants and 2.4-2.9 hr R(-) and 5.9-7.7 hr S(+) in Asian elephants. Serum concentrations of R(-) ibuprofen were undetectable at 24 hr, whereas S(+) ibuprofen decreased to below 5 microg/ml 24 hr postadministration in all elephants. The volume of distribution was estimated to be between 322 and 356 ml/kg R(-) and 133 and 173 ml/kg S(+) in Asian elephants and 360-431 ml/kg R(-) and 179-207 ml/kg S(+) in African elephants. Steady-state serum concentrations of ibuprofen ranged from 2.2 to 10.5 microg/ml R(-) and 5.5 to 32.0 microg/ml S(+) (mean: 5.17 +/- 0.7 R(-) and 13.95 +/- 0.9 S(+) microg/ml in African elephants and 5.0 +/- 1.09 microg/ml R(-) and 14.1 +/- 2.8 microg/ml S(+) in Asian elephants). Racemic ibuprofen administered at 6 mg/kg/12 hr for Asian elephants and at 7 mg/kg/12 hr for African elephants results in therapeutic serum concentrations of this antiinflammatory agent.     

Effects of parachlorophenylalanine,quipazine and cyproheptadine on growth hormone and adrenocorticotropin secretion in steers

Adrenergic and perhaps dopaminergic neurons provide inhibitory regulation of growth hormone (GH) secretion in ruminants. This suggests that either serotonergic or other neurons regulate the stimulatory release of GH. The nature of neurotransmitter control of adrenocorticotropin (ACTH) secretion in ruminants has not been determined. Parachlorophenylalanine (PCPA; serotonin synthesis inhibitor), quipazine (serotonin receptor agonist) and cyproheptadine (serotonin receptor antagonist) were utilized in Holstein steers to determine whether serotonin receptors mediate stimulatory actions on GH and ACTH secretion. PCPA (100 mg/kg BW) administered each day at 1900 hr for three successive days did not alter mean GH concentrations, amplitude of GH peaks, nor the number of GH peaks. Likewise, PCPA altered none of these parameters for ACTH. Quipazine injected iv at .1 or .5 mg/kg BW increased plasma GH (P<.05) and ACTH (P<.001) concentrations. There was a dose effect of quipazine on both GH (P<.05) and ACTH (P<.001) secretion. Pretreatment of steers with cyproheptadine (.06 and .6 mg/kg BW) reduced the stimulation of GH by quipazine (P<.0001) and decreased basal GH concentrations (P<.0004). Cyproheptadine at .06 mg/kg BW did not alter quipazine effects on ACTH, however, the higher dose decreased the peak ACTH response (P<.02) to quipazine. Studies with quipazine and cyproheptadine indicated that serotonergic mechanisms are likely involved in the regulation of GH and ACTH secretion in steers.     

Treatment with pergolide or cyproheptadine of pituitary pars intermedia dysfunction (equine Cushing's disease)

Medical records of 27 horses (including 13 ponies) treated with pergolide or cyproheptadine for pituitary pars intermedia dysfunction were reviewed to determine the effect of treatment on plasma ACTH, insulin, and glucose concentrations and clinical signs. Prior to treatment, the most common clinical signs were laminitis, hirsutism, and abnormal body fat distribution. The median pergolide dose was 3.0 microg/kg p.o. q24h (range, 1.7-5.5 microg/kg). All horses treated with cyproheptadine were given 0.25 mg/kg p.o. q24h. After pergolide treatment, ACTH concentrations (n = 20; median = 30.4 pg/ml; range, 4.2-173) were significantly lower (P < .01) than those in horses treated with cyproheptadine (n = 7; median = 141.0 pg/ml: range, 10-1,230). Among horses treated with pergolide, there was a correlation between ACTH concentration after treatment and the duration of treatment (P < .001) and pergolide dose (P = .04). Significantly (P = .02) more owners of horses treated with pergolide (85%, 17/20) reported an improvement in clinical signs compared to owners of horses treated with cyproheptadine (28%, 2/7).     

Influence of CRH and ACTH Administration on Endocrine Profile and Ovulation in Sows

Grouping of sows is a stressful event until the ranking is established. The purpose of this study was to simulate stress by repeated administration of porcine corticotropin releasing hormone (CRH) and adrenocorticotropic hormone (ACTH)/tetracosactide and to study its influence on endocrine profile and ovulation. Four multiparous sows were used and blood was collected every 2 h from the onset of pro-oestrus until 12 h after ovulation. The first oestrus after weaning was used to check ovulation and acclimate the sows to their environment. The second oestrus after weaning was used as control. At their third oestrus CRH (0.6 microg/kg) and at their fourth oestrus ACTH (5 microg/kg) were given every 4 h from onset of oestrus until ovulation. The total 'area under the curve' of cortisol was twofold larger in two of four sows during the CRH treatment period, and two- to fourfold larger (p < or = 0.05) during the ACTH treatment period, compared with the corresponding control period. In three sows, there was no clear effect of either CRH or ACTH on the levels of oestradiol 17beta, luteinizing hormone (LH) or on the timing of ovulation. One sow was different in all hormonal patterns and also in the timing of ovulation. In all four sows, ACTH treatment lowered the baseline level of prostaglandin F(2 alpha)-metabolite. Therefore, we conclude that stage of the oestrous cycle seems to be of importance when investigating the influence of exogenous administration of CRH/ACTH on hormonal pattern and ovulation time in the sow.     

Use of a low-dose ACTH stimulation test for diagnosis of hypoadrenocorticism in dogs

BACKGROUND: Although definitive diagnosis of hypoadrenocorticism usually is made by an adrenocorticotrophic hormone (ACTH) stimulation test using 250 microg/dog of synthetic ACTH (cosyntropin/tetracosactrin), increased costs have prompted a search for less-expensive diagnostic methods. HYPOTHESIS: A low-dose ACTH stimulation test (5 microg/kg) will distinguish between dogs with nonadrenal illness and hypoadrenocorticism. Additionally, administration of cosyntropin will not affect the results of another ACTH stimulation test performed 24 hours later. ANIMALS: Eight healthy adult dogs and 29 hospitalized dogs with suspected hypoadrenocorticism. METHODS: In this prospective study, each healthy dog received 4 ACTH stimulation tests. Dogs received either 5 microg/kg or 250 microg/dog of cosyntropin on day 1 and the alternate dose on day 2. The opposite dosing sequence was used after a 2-week washout period (days 15 and 16). Dogs with suspected Addison's disease received 2 ACTH stimulation tests, 24 hours apart, using either a dose of 5 microg/kg cosyntropin or 250 microg/dog on the 1st day and the alternate dose on the 2nd day. RESULTS: In healthy dogs, poststimulation cortisol concentrations on days 2 and 16 and days 1 and 15 were equivalent (90% confidence interval [CI]: 86.7-101.2%). In dogs with suspected Addison's disease, mean (+/-SD) cortisol responses to ACTH in the 5 microg/kg dose (16.2+/-7.7 microg/dL) and 250 microg/dog dose (15.9+/-6.3 microg/dL) were statistically equivalent (90% CI: 91.2-105.4%). CONCLUSIONS AND CLINICAL IMPORTANCE: Low-dose ACTH stimulation testing distinguishes between dogs with nonadrenal illness and hypoadrenocorticism. Additionally, the administration of 2 ACTH stimulation tests on consecutive days does not affect results of the second test.