Measurement of Carbonic Anhydrase Isozyme VI (CA-VI) in Bovine Sera,Saliva, Milk and Tissues

Concentrations of bovine carbonic anhydrase isozyme VI (CA-IV) in bovine serum, saliva, normal milk, colostrum, submandibular gland, liver, and mammary gland were determined. CA-VI was purified from bovine saliva and an antibody to CA-VI was generated. The concentrations of CA-VI in the saliva (7.8 ± 7.9 μg/ml), serum (2.1± 5.7 ng/ml), milk (7.9 ± 12.1 ng/ml), submandibular gland (284.7 μg/g protein), liver (921.0 ± 180.7 ng/g protein) and mammary gland (399.6 ± 191.2 ng/g protein) were determined by ELISA. No seasonal change in CA-VI levels was observed in normal milk. The concentration of CA-VI in colostrum (day 1 post partum) was 119 ng/ml and decreased rapidly by 1 month following birth. Mammary gland contained much smaller amounts than the submandibular gland. CA-VI mRNA was detected in the liver and mammary gland of cow by RT-PCR. The ELISA used in this study proved to be a precise and sensitive method for determining CA-VI concentrations in saliva, serum, milk and tissue specimens from cows. The ELISA may enable the study of changes in CA-VI associated with hereditary or metabolic disorders of the salivary gland, mammary gland and liver using small samples of saliva, serum or milk.     

Purification of swine carbonic anhydrase isoenzyme III and measurement of its levels in tissues and plasma

The changes in the levels of carbonic anhydrase isozyme III (CA‐III) in swine plasma and urine have not been previously determined or reported. CA‐III is relatively specific to skeletal muscles, and should therefore be a useful diagnostic marker for muscle diseases. We isolated CA‐III from swine muscle tissues and determined CA‐III levels in the plasma and urine from both healthy and diseased pigs. The levels of CA‐III in the tissues of female swine (age, 3 months) and plasma of young swine (age, 1–5 months) and adult female pigs (age, 2–3 years) were determined using the ELISA system for swine CA‐III. The mean (± SD) levels of CA‐III in the skeletal muscles were 3.8 ± 3.2 mg/g (wet tissue), and in the plasma, 230 ± 193 ng/ml at 1 month, 189 ± 208 ng/ml at 2 months, 141 ± 148 ng/ml at 3 months, 78 ± 142 ng/ml at 4 months and 53 ± 99 ng/ml at 5 months. The mean level of CA‐III in the plasma samples from 2‐ to 3‐year‐old pigs was 18 ± 60 ng/ml. CA‐III in the plasma samples was found to decrease from 1 month until 3 years of age (p < 0.01). We performed far‐western blotting to clarify the cause of the observed decrease in CA‐III in plasma. Our results demonstrated that CA‐III is bound to the transferrin and albumin. In addition, we determined that the levels of CA‐III in plasma and urine samples were higher in diseased swine compared with the healthy pigs.     

The Content of β-endorphin-like Immunoreactivity in Porcine Corpus Luteum and the Potential Roles of Progesterone, Oxytocin and Prolactin in the Regulation of β-endorphin Release from Luteal Cells in vitro

The amount of β‐endorphin‐like immunoreactivity (β‐END‐LI) in porcine corpora lutea from several stages of the oestrous cycle and the effects of progesterone, oxytocin, and prolactin on β‐END‐LI secretion in vitro by luteal cells were studied. Porcine corpora lutea obtained on days 1–5, 6–10, 11–13, 14–18, and 19–21 of the cycle were used to prepare extracts for β‐END‐LI determination. Additionally, corpora lutea from days 11–13 and 14–18 were enzymatically dissociated and isolated luteal cells were used for further study of β‐endorphin secretion in vitro. Cells were cultured in serum‐free defined M 199 medium (10⁶ cells/ml) at 37°C under 5% CO₂ in air, for 12 h. The influences of the following factors on β‐END‐LI secretion by luteal cells were tested: progesterone (10^–9, 10^–7 and 10^–5M ), oxytocin (0.01, 0.1, 1 and 10 ng/ml), and prolactin (0.1, 1, 10 and 100 ng/ml). The β‐END‐LI contents in extracts and media were measured by radioimmunoassay. The tissue concentration of β‐END‐LI was lowest on days 1–5 of the cycle (0.35 ± 0.03 ng/g wet tissue). Subsequently, it constantly increased to the highest value on days 14–18 (16.58 ± 0.52 ng/g wet tissue) and on days 19–21 it declined (11.10 ± 0.52 ng/g wet tissue). Progesterone at a low dose (10^–9 M ) resulted in significant (p < 0.05) increases and decreases in β‐END‐LI secretion by luteal cells from days 11–13 and 14–18, respectively. Higher doses of progesterone (10^–7 and 10^–5 M ) had no effect on β‐END‐LI release, compared with the control group. All dose‐levels of oxytocin used decreased β‐END‐LI secretion by luteal cells on days 11–13 and 14–18 of the cycle. Prolactin at doses of 0.1 and 1 ng/ml on days 11–13, and all doses tested on days 14–18 resulted in decreases in β‐END‐LI release from luteal cells. These results document evident changes in β‐END‐LI content in the pig corpus luteum during its development and indicate the potential roles of progesterone, oxytocin, and prolactin in luteal cell secretion of β‐END‐LI.     

Purification of carbonic anhydrase isozyme VI (CA-VI) from swine saliva.

Salivary or secreted carbonic anhydrase (CA), which constitutes a new class of CA, designated CA-VI, was isolated. Swine CA-VI purified from swine saliva by inhibitor-affinity chromatography and ion exchange chromatography had a specific activity of 5,468 units/mg. The molecular weight was 250,000, as determined by gel filtration under non-denaturing conditions, and the subunit molecular weight was found to be 37,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis, indicating that swine CA-VI consists of 7 subunits. The treatment of the enzyme with endo-N-acetylglucosaminidase F reduced its subunit molecular weight from 37,000 to 35,000 and 32,000. We raised a rabbit antibody against purfied swine CA-VI. Double immunodiffusion showed that anti-swine CA-VI serum reacted with swine CA-VI and swine saliva, but not with hemolysate (containing CA-I and CA-Il) or muscle extracts (containing CA-III). The concentration of CA-VI in swine saliva, measured using single radial immunodiffusion, was 0.027 +/- 0.017 mg/mg total protein.     

Hormonal and metabolic relation to restraint and human handling in growing-fattening steer

Endogenous relationship to restraint and human handling were studied with growing–fattening steers. Thirty‐five crossbred (Japanese Black × Holstein) steers aged 6–10 months were randomly assigned to three pens. They had free access to an Italian ryegrass hay and a restricted amount of high‐concentrate diets (total digestible nutrients (TDN) 70.5%, digestible crude protein (DCP) 10.0%) for the first 6 months of trial. Then they had free access to an oat hay and another vitamin A‐restricted diet (TDN 72.0%, DCP 10.0%) until slaughter. The steers were individually driven into a restraint stall, and bodyweight was recorded. Blood samples were then collected under haltered conditions. These serial handling procedures started at 2 h after the morning feeding were conducted in months 1, 3, 5 (growing stage, GS) and in months 7, 9, 11 (fattening stage, FS) of the trial. Mean peripheral blood concentrations of epinephrine (A; GS, 117.4 ± 76.4 pg/mL; FS, 64.1 ± 34.2 pg/mL), norepinephrine (NA; GS, 257.7 ± 95.0 pg/mL; FS, 125.9 ± 44.1 pg/mL), cortisol (GS, 1.6 ± 0.8 µg/dL; FS, 1.2 ± 0.4 µg/dL), glucose (GS, 83.1 ± 7.5 mg/dL; FS, 71.9 ± 6.9 mg/dL), non‐esterified fatty acid (NEFA; GS, 0.13 ± 0.06 mEQ/L; FS, 0.10 ± 0.06 mEQ/L) and vitamin A (GS, 90.5 ± 24.6 IU/dL; FS, 37.2 ± 21.3 IU/dL) were higher (all P < 0.01) in the GS than in the FS, whereas those of insulin (GS, 1.06 ± 0.82 µU/mL; FS, 1.36 ± 0.61 µU/mL) and leptin (GS, 4.5 ± 1.8 ng/mL human equivalent (HE); FS, 6.8 ± 2.4 ng/mL HE) were lower (both P < 0.01). The metabolite that correlated with A and NA was glucose (A: r = 0.61, P < 0.001; NA: r = 0.53, P < 0.01) in the GS, and the metabolites correlating with A, NA and cortisol were NEFA (A: r = 0.31, P < 0.10; NA: r = 0.32, P < 0.10; cortisol: r = 0.41, P < 0.05) and triglyceride (A: r = ?0.37, P < 0.05; NA: r = ?0.39, P < 0.05) in the FS. Vitamin A was a mediator between A (r = ?0.38, P < 0.05) and NA (r = ?0.42, P < 0.05) and insulin (r = 0.31, P < 0.10) in the GS, and between NA (r = ?0.33, P < 0.10) and leptin (r = 0.38, P < 0.05) in the FS. In conclusion, when changing from the growing to the fattening stages, the stress of handling and restraint had caused the pathways to shift from carbohydrate metabolism to lipid metabolism. In addition, vitamin A seemed to be an important mediator in the endogenous pathways in both stages.     

Pharmacokinetics and tissue disposition of meloxicam in beef calves after repeated oral administration

The objective of this study was to investigate the pharmacokinetics and tissue disposition of meloxicam after repeated oral administration in calves. Thirteen male British × Continental beef calves aged 4 to 6 months and weighing 297–392 kg received 0.5 mg/kg meloxicam per os once daily for 4 days. Plasma meloxicam concentrations were determined in 8 calves over 6 days after first treatment. Calves were randomly assigned to be euthanized at 5, 10, 15 (n = 3/timepoint), and 19 days (n = 4) after final administration. Meloxicam concentrations were determined in plasma (LOQ= 0.025 μg/mL) and muscle, liver, kidney, and fat samples (LOQ = 2 ng/g) after extraction using validated LC–MS–MS methods. The mean (± SD) C_max, C_min, and C_average plasma meloxicam concentrations were 4.52 ± 0.87 μg/mL, 2.95 ± 0.77 μg/mL, and 3.84 ± 0.81 μg/mL, respectively. Mean (± SD) tissue meloxicam concentrations were highest in liver (226.67 ± 118.16 ng/g) and kidney samples (52.73 ± 39.01 ng/g) at 5 days after final treatment. Meloxicam concentrations were below the LOQ in all tissues at 15 days after treatment. These findings suggest that tissue from meloxicam‐treated calves will have low residue concentrations by 21 days after repeated oral administration.     

Assessment of pregnancy‐associated glycoprotein (PAG) concentrations in swamp buffalo samples from fetal and maternal origins by using interspecies antisera

Pregnancy‐associated glycoproteins (PAG) constitute a large family of glycoproteins found in the outer placental epithelial cell layer of the placenta in Eutherian species. In ruminants, they are noted to be structurally closely related among the different species. This study was designed to determine PAG concentrations in maternal and fetal plasma, allantoic and amniotic fluids in buffalo species. Antisera (AS) generated in rabbits against distinct PAG molecules were used in three radioimmunoassay (RIA)‐PAG systems: RIA‐1 (antiserum raised against bovine PAG67kDa; AS#497), RIA‐2 (antiserum raised against caprine PAG55 + 62 kDa; AS#706) or RIA‐3 (antiserum raised against buffalo PAG; AS#859). Samples were collected at a slaughterhouse (n = 67). PAG concentrations determined by RIA‐2 gave significantly higher results in both allantoic and amniotic fluids (12.7 ± 2.1 ng/mL and 24.0 ± 7.3 ng/mL, respectively). Regarding maternal and fetal plasma, PAG concentrations obtained by RIA‐2 (21.8 ± 2.4 ng/mL and 20.2 ± 2.5 ng/mL, respectively) and RIA‐3 (25.0 ± 2.2 ng/mL and 21.9 ± 3.2 ng/mL, respectively) were higher than those obtained by RIA‐1 (15.5 ± 1.4 ng/mL and 16.1 ± 1.8 ng/mL, respectively). The correlation among the three systems was very high. The study clearly reveals the ability of different PAG‐RIA systems to measure PAG concentration in swamp buffalo samples.     

Sulfated cholecystokinin‐8 increases ghrelin secretion but does not affect oxyntomodulin in Holstein steers

The effect of appetite regulatory hormone cholecystokinin (CCK) on the secretions of oxyntomodulin (OXM) and ghrelin, and the effect of ghrelin on the secretions of CCK and OXM were studied in ruminants. Eight Holstein steers, 7 months old, 243 ± 7 kg body weight (BW), were arranged in an incomplete Latin square design (8 animals × 4 treatments × 4 days of sampling). Steers were intravenously injected with 10 µg of sulfated CCK‐8/kg BW, 20 µg of acyl ghrelin/kg BW, 100 µg of des‐acyl ghrelin/kg BW or vehicle. Blood samples were collected from ?60 min to 120 min relative to time of injection. Plasma concentrations of ghrelin, sulfated CCK and OXM were measured by double‐antibody radioimmunoassay. Plasma acyl ghrelin was increased to peak level (428.3 ± 6 pg/mL) at 60 min after injection of CCK compared with pre‐injected levels (203.3 ± 1 pg/mL). These results showed for the first time, that intravenous bolus injection of CCK increased ghrelin secretion in ruminants. In contrast, injection of ghrelin did not change CCK secretion. Administration of ghrelin or CCK has no effect on plasma OXM concentrations. In conclusion, our results show that administration of CCK increased ghrelin secretion but did not affect OXM release in ruminants. Ghrelin did not affect the secretions of CCK and OXM.     

Intravenous anaesthesia using detomidine, ketamine and guaiphenesin for laparotomy in pregnant pony mares

Objective To characterize intravenous anaesthesia with detomidine, ketamine and guaiphenesin in pregnant ponies. Animals Twelve pony mares, at 260–320 days gestation undergoing abdominal surgery to implant fetal and maternal vascular catheters. Materials and methods Pre‐anaesthetic medication with intravenous (IV) acepromazine (30 µg kg^?1), butorphanol (20 µg kg^?1) and detomidine (10 µg kg^?1) preceded induction of anaesthesia with detomidine (10 µg kg^?1) and ketamine (2 mg kg^?1) IV Maternal arterial blood pressure was measured directly throughout anaesthesia and arterial blood samples were taken at 20‐minute intervals for measurement of blood gases and plasma concentrations of cortisol, glucose and lactate. Anaesthesia was maintained with an IV infusion of detomidine (0.04 mg mL^?1), ketamine (4 mg mL^?1) and guaiphenesin (100 mg mL^?1) (DKG) for 140 minutes. Oxygen was supplied by intermittent positive pressure ventilation (IPPV) adjusted to maintain PaCO₂ between 5.0 and 6.0 kPa (38 and 45 mm Hg), while PaO₂ was kept close to 20.0 kPa (150 mm Hg) by adding nitrous oxide. Simultaneous fetal and maternal blood samples were withdrawn at 90 minutes. Recovery quality was assessed. Results DKG was infused at 0.67 ± 0.17 mL kg^?1 hour^?1 for 1 hour then reduced, reaching 0.28 ± 0.14 mL kg^?1 hour^?1 at 140 minutes. Arterial blood gas values and pH remained within intended limits. During anaesthesia there was no change in heart rate, but arterial blood pressure decreased by 10%. Plasma glucose and lactate increased (10‐fold and 2‐fold, respectively) and cortisol decreased by 50% during anaesthesia. Fetal umbilical venous pH, PO₂ and PCO₂ were 7.34 ± 0.06, 5.8 ± 0.9 kPa (44 ± 7 mm Hg) and 6.7 ± 0.8 kPa (50 ± 6 mm Hg); and fetal arterial pH, PO₂ and PCO₂ were 7.29 ± 0.06, 4.0 ± 0.7 kPa (30 ± 5 mm Hg) and 7.8 ± 1.7 kPa (59 ± 13 mm Hg), respectively. Surgical conditions were good but four ponies required a single additional dose of ketamine. Ponies took 60 ± 28 minutes to stand and recovery was good. Conclusions and clinical relevance Anaesthesia produced with DKG was smooth while cardiovascular function in mare and fetus was well preserved. This indicates that DKG infusion is suitable for maintenance of anaesthesia in pregnant equidae.     

Leptin and ghrelin levels in colostrum,milk and blood plasma of sows and pig neonates during the first week of lactation

Radioimmunology was used to determine leptin and ghrelin levels in sow colostrum and milk in relation to those in sow and neonatal pig blood plasma and to the body weight of piglets during the first week of lactation. The highest concentration of leptin was found in colostrum on the second day of lactation (69.3 ± 6.3 ng/mL). Leptin concentrations in sow plasma were significantly lower than in colostrum/milk (2.19 ± 0.9 ng/mL, P = 0.7692) and were stable in the first 7 days of lactation. Total and active ghrelin concentrations in colostrum/milk were stable in the measured time points (6734 ± 261 pg/mL, P = 0.3397; 831 ± 242 pg/mL, P = 0.3988, respectively). Total ghrelin concentrations in sow plasma were lower than in colostrum/milk. These results indicate that pigs follow a unique species‐specific pattern of leptin and ghrelin synthesis, release and existence, and that the mammary gland is an important source of leptin and ghrelin contained in colostrum/milk.     

Communications: Validation of a Solid-Phase Enzyme Immunoassay Technique for the Measure of Plasma Cortisol in Rainbow Trout

Abstract

A kit for a solid-phase enzyme immunoassay (SOPHEIA®) of cortisol in human sera was evaluated and validated for measuring cortisol in plasma of rainbow trout Oncorhynchus mykiss. The accuracy of the SOPHEIA was demonstrated by the recovery of exogenous cortisol concentrations of 25, 50, 100, and 250 ng/mL in charcoal-stripped fish plasma. The amounts (mean ± SE) recovered from triplicate samples were 29.9 ± 2.75, 47.5 ± 3.41, 101.7 ± 12.08, and 232.0 ± 11.06 ng/mL, respectively. The intra- and interassay coefficient of variation (CV = 100 × SD/mean) for cortisol levels in undisturbed fish (26.6 ± 1.18 ng/mL) were 14 and 10%, respectively. The intra- and interassay CV for elevated cortisol levels in stressed fish (330.8 ± 19.90 ng/mL) were 8 and 13%, respectively. Cross-reactivity determined for nine steroids in teleostean fish was negligible. Cortisol concentrations in serial dilutions of pooled fish plasma were parallel to the standard curve. Sensitivity (minimum detection limit) was 3.04 ng/mL. The SOPHEIA compared favorably to radioimmunoassay measurements of cortisol (r = 0.98; P < 0.001).     

Immunomodulatory effects of Alliums and Ipomoea batata extracts on lymphocytes and macrophages functions in White Leghorn chickens: In vitro study

We previously described that supplementary garlic, onion and purple sweet potato (PSP) enhance humoral immune response in White Leghorn chickens. In the present in vitro study, we investigated the effects of garlic (GE), onion (OE) and PSP (PSPE) extracts on proliferation, interleukin (IL)‐2 and interferon (INF)‐γ gene expression of stimulated lymphocytes. The effects on microbicidal activity, reactive oxygen species (ROS) and nitric oxide (NO) productions of stimulated peritoneal macrophages were studied as well. The results showed that GE augmented Concanavalin A (ConA)‐induced splenocytes (4, 8 and 16 µg/mL) and thymocytes (2, 4 and 8 µg/mL) proliferations, and gene expression of IL‐2 (8 and 16 µg/mL) and INF‐γ (16 µg/mL). None of the examined extracts had mitogenic effect nor stimulated bursacytes response to phorbol 12‐myristate 13‐acetate (PMA). Macrophages exhibited superior microbicidal activity and ROS production with GE at 4 and 8 µg/mL and with OE at 25.6 µg/mL. None of the extracts showed stimulatory effects on NO production. The extracts showed concentration‐dependent inhibitory effects on all measured parameters at higher concentrations. Taken together, it is likely that garlic has direct stimulatory effects on immune cell functions, whereas the in vitro inhibitory effects of onion and PSP were likely attributed to high flavonoid contents.     

Blood supply of the ovine adrenal gland and its relevance in adrenal autotransplantation

SUMMARY The arterial supply and venous drainage of 62 left and 5 right ovine adrenal glands is described, and the contribution of individual arteries to successful adrenal gland autotransplantation was evaluated. Arterial flow was measured by direct collection from the draining adrenal vein. Assessment of function of the transplanted adrenal gland was made from survival of the sheep and by the cortisol response to infusion of ACTH and the aldosterone secretory response to infusion of angiotensin II or potassium. For the left adrenal, the principal arterial supply was from the renal artery in 21 (34%), a lumbar artery in 32 (52%), and the anterior mesenteric artery in 3. The total blood flow was 5.0 ± SEM 0.4 mL/min, the flow from the renal branch 2.3 ± 0.3 mL/min, and the principal lumbar branch 2.6 ± 0.3 mL/min. Venous drainage from the left adrenal was via a major adrenal vein to the left renal vein, but additional tributaries to the renal vein were present in 26% . The arterial supply to the adrenal is regional and omission of a branch at transplantation could result in infarction of portion of the gland. By defining arterial supply and measuring blood flow, selection of the appropriate artery or multiple arteries can achieve an adrenal gland autotransplant survival of 90% .     

The effect of medetomidine and its antagonism with atipamezole on stress-related hormones, metabolites, physiologic responses, sedation, and analgesia in goats

Six 3‐year‐old goats (three males and three females) weighing 60.0 ± 18 kg (mean ± SD) were used to investigate the effect of medetomidine (MED; 20 µg kg^?1 IV) and its antagonism with atipamezole (ATI; 100 µg kg^?1 IV) on physiologic responses (heart rate (HR; beats minute^?1), respiratory rate (RR; breaths minute^?1), electrocardiogram (ECG), rectal temperature (T; °C), blood pressure (oscillometric; mm Hg), sedation (SED), posture (REC), analgesia (ALG), and stress‐related hormonal and metabolic responses (epinephrine and norepinephrine (high performance liquid chromatography with electrochemical detection), cortisol (COR; µg dL^?1; radioimmunoassay), glucose (GLU; mg mL^?1; enzymatic colorimetric assay), and free fatty acids (modified enzymatic colorimetric assay)); each goat received ATI or SAL in random order separated by 1 week. Jugular catheters were placed for drug administration and blood sampling (10–12 mL sample^?1) using a lidocaine skin block (20 mg) 2 hours prior to beginning of each trial; during this trial, goats breathed room air. Physiologic parameters were measured, SED, REC, and ALG were scored, and blood samples were collected from jugular catheters at baseline (time = ?30 minutes), 5 minutes post‐MED administration (time = ?25 minutes), 25 minute post‐MED administration and immediately prior to antagonism (time = 0 minute), and at 5, 30, 60, and 120 minutes after administering ATI or SAL. ALG was tested by clamping the withers and metacarpus with hoof testers fitted with a force transducer to measure applied isometric force (lb) (a technique used previously in goats to evaluate analgesia). Continuous variables were analyzed by Repeated Measures analysis of variance (anova ); categorical data were analyzed using a Friedman Repeated Measures anova on ranks. A p‐value of <0.05 was considered significant. If a significant difference was found, a Dunnett's pair‐wise comparison of means was conducted. Differences between ATI and SAL were examined at 5, 30, 60, and 120 minutes using a paired t‐test with a Bonferroni correction. Administration of MED resulted in a decrease in T (38.7 ± 0.3 to 34.5 ± 0.4 °C), HR (78 ± 19 to 55 ± 9), and RR (31 ± 12 to 14 ± 5) over time; an increase in mean arterial blood pressure (90 ± 19 to 132 ± 23), COR (0.254 ± 0.125 to 4.327 ± 1.233), and GLU (82.0 ± 13.2 to 255.9 ± 38.9); and changes in SED (alert to marked sedation), REC (standing to recumbent), and ALG (metacarpus = 5 ± 2 to 14 ± 0; withers = 3 ± 2 to 14 ± 0). GLU was 62–70% higher at 60 and 120 minutes and COR was 336% higher after SAL than after ATI at 120 minutes; at 30, 60, and 120 minutes, T was 4–10% higher after ATI than SAL. There were no other significant differences. REC, SED, and ALG were antagonized after ATI. ATI did not antagonize the effect of MED on HR, RR, or MAP, but stabilized T and antagonized the increase in GLU and COR.     

Sexual maturity,seasonal estrus,and gestation in female Indo-Pacific bottlenose dolphins Tursiops aduncus inferred from serum reproductive hormones

Reproductive hormones in serum concentrations of progesterone, estradiol, and testosterone in female Indo-Pacific bottlenose dolphins (Tursiops aduncus, n = 12) housed in Ocean Park Hong Kong were investigated in the present study. Results showed that, onset of puberty of captive Indo-Pacific bottlenose dolphins was at 5 years while sexual maturity was at 6. Average serum progesterone concentrations in non-pregnant sexually mature individuals was 0.33 (0.25–0.97) ng/mL (interquartile), significantly higher than in immature ones 0.26 (0.25–0.38) ng/mL. This study found significant difference in serum estradiol concentrations between individuals at the onset of puberty (9.5 ± 1.7 pg/mL, ±SD) and not (below detection limit 9 pg/mL). A slightly seasonal breeding pattern, with progesterone values tend to be higher from February to October (0.38 [0.25–1.07] ng/mL) was inferred. During pregnancy, serum progesterone concentrations range from 10.54 ± 8.74 ng/mL (indexed month post-conception [IMPC] 0) to 25.49 ± 12.06 ng/mL (IMPC 2), and display a bimodal pattern with 2 peaks in early- (25.49 ± 12.06 ng/mL, IMPC 2) and late-pregnancy (21.71 ± 10.25 ng/mL, IMPC 12), respectively. Serum estradiol concentrations can seldom be detected in early-pregnancy and increase constantly in mid- (9.45 ± 1.83 pg/mL) and late-pregnancy (11.88 ± 3.81 pg/mL), with a spike (15.45 ± 6.78 pg/mL) 1 month prior to delivery. Serum testosterone concentrations elevate significantly in IMPC 7 (0.36 ± 0.10 ng/mL) compared to other months (0.16 ± 0.10 ng/mL) of the year. The present study provides normal concentration profiles for some reproductive hormones in female Indo-Pacific bottlenose dolphins and can contribute to the breeding monitoring of this species. Also, our study would shed further light on the reproductive physiology of small cetaceans.     

Detection,quantifications, and pharmacokinetics of ponazuril in healthy swine

A study on pharmacokinetics of ponazuril in piglets was conducted after a single oral dose of 5.0 mg/kg b.w. Plasma concentrations were measured by high‐performance liquid chromatography assay with UV detector at 255‐nm wavelength. Pharmacokinetic parameters were derived by use of a standard noncompartmental pharmacokinetic analysis. Samples from six piglets showed good plasma concentrations of ponazuril, which peaked at 5.83 ± 0.94 μg/mL. Mean ± SD area under the plasma concentration–time curve was 1383.42 ± 363.26 h/μg/mL, and the elimination half‐life was 135.28 ± 19.03 h. Plasma concentration of ponazuril peaked at 42 h (range, 36–48 h) after administration and gradually decreased but remained detectable for up to 33 days. No adverse effects were observed during the study period. The results indicate that ponazuril was relatively well absorbed following a single dose, which makes ponazuril likely to be effective in swine.     

Lipid Composition of Hepatic and Adipose Tissues From Normal Cats and From Cats With Idiopathic Hepatic Lipidosis

The purpose of this study was to characterize the lipid classes in hepatic and adipose tissues from cats with idiopathic hepatic lipidosis (IHL). Concentrations of triglyceride, phospholipid phosphorus, and free and total cholesterol were determined in lipid extracts of liver homogenates from 5 cats with IHL and 5 healthy control cats. Total fatty acid composition of liver and adipose tissue was also compared. Triglyceride accounted for 34% of liver by weight in cats with IHL (338 ± 38 mg/g wet liver) versus 1% in control cats {9.9 ±1.0 mg/g wet liver, P < .001). The mass of cholesterol ester was significantly higher in triglyceride-free (TG-free) liver from cats with IHL (741 ± 340 μg/g TG-free wet liver) compared to healthy cats (31 ± 11 μg/g TG-free wet liver, P < .05). Total fatty acid composition of hepatic tissue in the 2 groups differed; pa Imitate was higher (19.5 ± 1.1% of total fatty acids in cats with IHL versus 9.2 ± 2.7% in controls, P < .05), stearate was lower (8.5 ± 0.8% versus 16.8 ± 1.1%, P < .05), oleate was higher (41.2 ± 1.6% versus 31.1 ± 1.8%, P < .05), and arachidonate was lower (1.2 ± 0.2% versus 6.0 ± 0.9%, P < .05). The total fatty acid composition of adipose tissue also differed between the 2 groups; palmitate was higher (25.2 ± 1.2% in cats with IHL versus 21.3 ± 0.6% in controls, P < .05), total monounsaturated fatty acids were higher (48.4 ± 1.0% versus 45.0 ± 0.8%, P < .05), linoleate was lower (13.3 ± 1.6% versus 17.5 ± 0.9%, P < .05), total (n-6) fatty acids were lower (13.8 ± 1.38% versus 18.4 ± 0.83%, P < .05), linolenate was lower (0.2 ± 0.04% versus 0.7 ± 0.06%, P < .05), and total (n-3) fatty acids were lower (0.3 ± 0.02% versus 1.3 ± 0.32%, P < .05). The fatty acid composition of both liver and adipose tissue was similar for stearate, oleate, linoleate, and linolenate in cats with IHL. These results support the hypothesis that the origin of hepatic triglyceride in cats with IHL is the mobilization of fatty acids from adipose tissue.     

Pharmacokinetics of tobramycin following intravenous,intramuscular, and intra‐articular administration in healthy horses

The objectives of this study were to examine the pharmacokinetics of tobramycin in the horse following intravenous (IV), intramuscular (IM), and intra‐articular (IA) administration. Six mares received 4 mg/kg tobramycin IV, IM, and IV with concurrent IA administration (IV+IA) in a randomized 3‐way crossover design. A washout period of at least 7 days was allotted between experiments. After IV administration, the volume of distribution, clearance, and half‐life were 0.18 ± 0.04 L/kg, 1.18 ± 0.32 mL·kg/min, and 4.61 ± 1.10 h, respectively. Concurrent IA administration could not be demonstrated to influence IV pharmacokinetics. The mean maximum plasma concentration (C_max) after IM administration was 18.24 ± 9.23 μg/mL at 1.0 h (range 1.0–2.0 h), with a mean bioavailability of 81.22 ± 44.05%. Intramuscular administration was well tolerated, despite the high volume of drug administered (50 mL per 500 kg horse). Trough concentrations at 24 h were below 2 μg/mL in all horses after all routes of administration. Specifically, trough concentrations at 24 h were 0.04 ± 0.01 μg/mL for the IV route, 0.04 ± 0.02 μg/mL for the IV/IA route, and 0.02 ± 0.02 for the IM route. An additional six mares received IA administration of 240 mg tobramycin. Synovial fluid concentrations were 3056.47 ± 1310.89 μg/mL at 30 min after administration, and they persisted for up to 48 h with concentrations of 14.80 ± 7.47 μg/mL. Tobramycin IA resulted in a mild chemical synovitis as evidenced by an increase in synovial fluid cell count and total protein, but appeared to be safe for administration. Monte Carlo simulations suggest that tobramycin would be effective against bacteria with a minimum inhibitory concentration (MIC) of 2 μg/mL for IV administration and 1 μg/mL for IM administration based on C_max:MIC of 10.     

Plasma and serum concentrations of cytarabine administered via continuous intravenous infusion to dogs with meningoencephalomyelitis of unknown etiology

The objective of this study was to evaluate the plasma and serum concentrations of cytarabine (CA) administered via constant rate infusion (CRI) in dogs with meningoencephalomyelitis of unknown etiology (MUE). Nineteen client‐owned dogs received a CRI of CA at a dose of 25 mg/m²/h for 8 h as treatment for MUE. Dogs were divided into four groups, those receiving CA alone and those receiving CA in conjunction with other drugs. Blood samples were collected at 0, 1, 8, and 12 h after initiating the CRI. Plasma (n = 13) and serum (n = 11) cytarabine concentrations were measured by high‐pressure liquid chromatography. The mean peak concentration (C_MAX) and area under the curve (AUC) after CRI administration were 1.70 ± 0.66 μg/mL and 11.39 ± 3.37 h·μg/mL, respectively, for dogs receiving cytarabine alone, 2.36 ± 0.35 μg/mL and 16.91 + 3.60 h·μg/mL for dogs administered cytarabine and concurrently on other drugs. Mean concentrations for all dogs were above 1.0 μg/mL at both the 1‐ and 8‐h time points. The steady‐state achieved with cytarabine CRI produces a consistent and prolonged exposure in plasma and serum, which is likely to produce equilibrium between blood and the central nervous system in dogs with a clinical diagnosis of MUE. Other medications commonly used to treat MUE do not appear to alter CA concentrations in serum and plasma.     

Comparative pharmacokinetics of sulfamethazine in plasma and parotid saliva of sheep

Salivary output in sheep is large enough to be considered a physiologic body fluid compartment. The hypothesis for this work was that pharmacokinetics of sulfamethazine in saliva was similar to that in plasma. A reliable technique was developed to measure parotid salivary output. Mean output of saliva was 3.18 ± 1.04 L from a single parotid gland per day with a mean flow of 2.21 ± 0.43 mL/min. Using concentrations of sulfamethazine in parotid saliva made it possible to calculate the total passage of sulfamethazine to parotid saliva, which was calculated to be 3.5% of the total dose. Pharmacokinetic variables obtained for sulfamethazine in plasma and in saliva were closely related ( AUC 1408 μg.h/mL and AUC 1484 μg.h/mL; V d_area 0.434 L/kg and V d _area 0.374 L/kg; t _½β 4.30 h and 3.46 h, respectively) and no substantial differences were observed. The convenience of using salivary concentrations of sulfamethazine for drug monitoring is discussed.