Anticholinesterase-resistant neuromuscular blockade in sheep given amino-steroid muscle relaxants

Three cases of anticholinesterase-resistant neuromuscular blockade in sheep are described. Neuromuscular blockade produced with pancuronium (n = 2) and vecuronium was only temporarily antagonised with anticholinesterases and signs of residual block-muscular weakness, recumbency, hypoventilation, dyspnoea, dysphagia, 'fade' in evoked responses to repetitive nerve stimulation and rapid exhaustion of cranial nerve reflexes - were observed. Responses to repeated edrophonium and, or neostigmine injections were incomplete and shortlived. These complications probably arose because high loading doses, incremental injections (pancuronium and vecuronium) and infusion (pancuronium) created high plasma drug concentrations at antagonism. This occurred because the facial nerve-levator nasolabialis muscle unit used to monitor drug effects proved resistant to blockade. The effects of relative overdose were probably augmented by the sensitivity of sheep to non-depolarizing neuromuscular blocking agents and the relative insensitivity of monitoring blockade using tactile and visual evaluation of evoked responses.     

Antinociceptive effects of intrathecal 5-HT agonists in sheep

Objectives To examine the role of spinal 5‐hydroxytryptamine (5‐HT) binding sites in nociceptive processing in conscious sheep and to study the role of 5‐HT agonists in mediating analgesia. Study design Prospective controlled study. Animals Nine adult healthy female sheep (Swaledale, Swaledale‐cross or Clun Forest) weighing 45–65 kg. Methods Intrathecal (IT) catheters were implanted at the cervical (n = 5) or lumbar (n = 4) level of the spinal cord under general anaesthesia. At least 1 week later, and at 1 week intervals thereafter, the effects of intrathecal Ringer's solution (control), xylazine (100 µg), 5‐HT creatinine sulphate (200, 400 and 800 µg), RU24969 (200 µg), α‐Methyl‐5‐HT and 1‐(3‐Chlorophenyl)‐biguanide (CPBG) on the mechanical nociceptive threshold (MT) were studied. Results were plotted as mean variable versus time curves. Areas under portions of the curves (0–30 and 0–60 minutes) were measured and expressed as mean ± standard error. Differences between values for control and drug trials were examined using the two‐tailed Student's t‐test. Results Baseline values of MT were lower on the hind limbs than on the forelimbs. Intrathecal Ringer's solution did not alter MT in the cervical or lumbar region. Xylazine (100 µg) produced a characteristic elevation in MT between 5 and 60 + minutes. Lumbar IT injection of 5‐HT (800 µg) raised the MT more than cervical injection, while cervical injection of RU24969 (200 µg) raised the MT more than lumbar administration. Cervical IT injection of α‐Me‐5‐HT (500 µg) produced a marked and significant increase in MT while lumbar application had no effect. CPBG (500 µg) injection caused no significant effect on MT with either cervical or lumbar applications. Conclusions The activation of 5‐HT₁ and 5‐HT₂ receptors particularly at the cervical level appears to be involved in spinal nociceptive processing in the sheep. Clinical relevance These effects, which lasted about 60 minutes, may have an implication in the development of new analgesic strategies for animals.     

Contractile effects of 5-hydroxytryptamine (5-HT) in the equine jejunum circular muscle: functional and immunohistochemical identification of a 5-HT1A-like receptor

REASONS FOR PERFORMING STUDY: Prokinetic drugs used to treat gastrointestinal ileus in man have equivocal results in horses. In man, prokinetic drugs have 5-hydroxytryptamine4(5-HT4) receptors as their target, but little is known about the 5-HT-receptor subtypes in the equine small intestine. OBJECTIVE: Functional and immunohistochemical identification of the serotonin receptor subtype(s) responsible for the 5-HT induced contractile response in the equine circular jejunum. METHODS: Isometric organ-bath recordings were carried out to assess spontaneous and drug-evoked contractile activity of equine circular jejunum. Histological investigations by immunofluorescence analyses were performed to check for presence and localisation of this functionally identified 5-HT receptor subtype. RESULTS: Tonic contractions were induced by 5-HT in horse jejunal circular muscle. Tetrodotoxin, atropine and NG-nitro L-arginine did not modify this response. A set of 5-HT receptor subtype selective antagonists excluded interaction with 5-HT1B, 1D, 2A, 3, 4 and 7 receptors. The selective 5-HT1A receptor antagonists WAY 100635 and NAN 190 caused a clear rightward shift of the concentration-response curve to 5-HT. The contractile effect of 5-CT, that can interact with 5-HT1A, 1B, 1D, 5 and 7 receptors was also antagonised by WAY 100635, identifying the targeted 5-HT receptor as a 5-HT1A-like receptor. Immunohistology performed with rabbit polyclonal anti-5-HT1A receptor antibodies confirmed the presence of muscular 5-HT1A receptors in the muscularis mucosae, and both longitudinal and circular smooth muscle layers of the equine jejunum. CONCLUSIONS: Contractile responses in equine jejunal circular smooth muscle induced by 5-HT involves 5-HT1A-like receptors.     

The vascular responses of the isolated perfused bovine external ear to exogenous histamine and 5-HT (serotonin)

The responses of the vasculature of isolated, non-sensitized, bovine external ears to histamine and serotonin (5-HT) were evaluated while they were being perfused with Krebs-Henseleit solution, Histamine (10^–5 mol/L to 5×10^–3 mol/L) and 5-hydroxytryptamine (5-HT) (10^–9 mol/L to 10^–2 mol/L) caused increased vascular resistance. Mepyramine (10^–7 mol/L), cimetidine (10^–5 mol/L) and atropine (10^–6 mol/L) inhibited the responses to histamine. The responses to 5-HT were inhibited by methysergide (10^–9 mol/L) and potentiated by morphine (10^–5 mol/L). These results suggest the presence of H₁ and H₂ histamine, and 5-HT receptors in bovine auricular vessels, all of which cause net vasoconstriction.     

Early pregnancy alters the metabolic responses to restricted nutrition in sheep

This study investigated whether a 27-day period of nutrition at half-maintenance during early pregnancy (up to Day 14) could alter maternal endocrine responses. Forty-six ewes were fed all or half of their maintenance requirements and slaughtered on Day 14 of the oestrous cycle or pregnancy. We used real time RT-PCR to study gene expression of growth hormone receptor (GHR) and leptin in adipose tissue and GHR, GHR1A and of the insulin-like growth factor I (IGF-I) in the liver. Blood profiles of metabolites and metabolic hormones were also determined. Throughout the experiment, underfed animals presented lower body weight and body condition, greater plasma concentrations of non-esterified fatty acids (NEFA), and lower plasma concentrations of leptin, compared to adequately fed animals. Undernutrition affected the patterns of gene expression in adipose and hepatic tissues, and the responses differed between pregnant and non-pregnant ewes. In adequately fed ewes, pregnancy up-regulated leptin mRNA expression in adipose tissue, a response that was impaired in underfed ewes. The hepatic expression of IGF-I mRNA was increased by pregnancy in underfed animals while no effect was observed in adequately fed ewes. It remains to be determined whether the changes in the endocrine milieu are paralleled by modifications in uterine gene expression that could alter the environment of the embryo during early pregnancy.     

Immune responses of sheep to louping-ill virus vaccine

The immune responses of sheep to single and double doses of commercially available louping-ill virus vaccine were examined. The susceptibility to challenge of sheep which had been vaccinated but showed a poor response was also investigated. Two injections of vaccine were required to provoke an adequate antibody response and maximum titres were obtained when there was an interval of two to eight weeks between injections. After challenge, viraemia could not be detected in animals with an antibody titre of 20 although increase in the concentration of humoral antibodies indicated that infection had occurred. Vaccinated but seronegative sheep and vaccinated animals with an antibody titre of 10 were also clinically resistant to the challenge, although circulation of virus was demonstrated. That vaccination had sensitised those animals to viral antigen was evident from the reduced viraemias, the early rise in humoral antibody titres and subsequent protection afforded compared to unvaccinated control animals. Thus, animals with minimal antibody titres after vaccination are protected, but it is recommended that vaccines eliciting the highest possible antibody responses will be the most useful under field conditions.     

Effect of selenium on sheep lymphocyte responses to mitogens

The effect of selenium (Se) on sheep lymphocyte response to mitogens was studied. In an indoor experiment lambs were fed a basal diet containing 0.13 mg Se kg-1, and supplemented with, respectively, 0.1 or 0.5 mg Se kg-1, either as sodium selenite or as selenomethionine. Enhancement of the proliferative response of lymphocytes to phytohaemagglutinin (PHA), pokeweed mitogen (PWM) and concanavalin A was found in lambs following selenium supplementation at the lower levels. The highest dietary selenium content, however, induced decreased mitogen response. Transient increases in lymphocyte response to PHA and PWM by ewes supplemented with selenium was demonstrated in one field study and a combined effect of selenium and vitamin E was seen in another. There was no stimulatory effect on the mitogen response of lymphocytes from sheep supplemented with dietary vitamin E alone.     

Correlated physiological responses to selection forcarcass lean content in sheep

Correlated responses in physiological traits in lines of Texel-Oxford sheep selected for high or lowcarcass lean content were examined. Serum samples were taken at the end of performance test, 20 weeks of age, from 66 rams when fed ad libitum and every 24 h when fasted for 56 h. The lean line had higher serum concentrations of β-hydroxybutyrate (BHB), non-esterified fatty acids (NEFA) and insulin-like growth factor- 1 (IGF-1) with lower concentrations of triglyceride (TRIG), creatinine (CREA) and UREA, but only the differences in UREA were statistically significant. There were substantial changes in serum concentrations of physiological traits in response to fasting as glucose and IGF-1 decreased while BHB, NEFA, TRIG, CREA and UREA increased. The correlated responses suggested that the lean line preferentially synthesises protein rather than deposits fat during normal feeding. When the animals are fasted, there may be relatively greater use of fat as an energy source in the lean line, rather than using products from protein catabolism as glucose precursors.The accuracy of selection was examined when physiological traits were incorporated into a selectionindex, which included the performance test traits: liveweight, ultrasonic backfat and muscle depths, to predict genetic merit for carcass lean content. UREA may be a useful predictor of genetic merit for lean meat production as it was correlated with estimated carcass lean content and there were substantial differences between the selection lines.     

Physiological and behavioral responses of sheep to gaseous ammonia

Ammonia can accumulate in highly stocked sheep accommodation, for example during live export shipments, and could affect sheep health and welfare. Thus, the objective of this experiment was to test the effects of 4 NH(3) concentrations, 4 (control), 12, 21, and 34 mg/m(3), on the physiology and behavior of wether sheep. Sheep were held for 12 d under a micro-climate and stocking density similar to shipboard conditions recorded on voyages from Australia to the Middle East during the northern hemispheric summer. Ammonia increased macrophage activity in transtracheal aspirations, indicating active pulmonary inflammation; however, it had no effect (P > 0.05) on hematological variables. Feed intake decreased (P = 0.002) in proportion to ammonia concentration, and BW gain decreased (P < 0.001) at the 2 greatest concentrations. Exposure to ammonia increased (P = 0.03) the frequency of sneezing, and at the greatest ammonia concentration, sheep were less active, with less locomotion, pawing, and panting. Twenty-eight days after exposure to NH(3), the pulmonary macrophage activity and BW of the sheep returned to that of sheep exposed to only 4 mg/m(3). It was concluded that NH(3) induced a temporary inflammatory response of the respiratory system and reduced BW gain, which together indicated a transitory adverse effect on the welfare of sheep.     

The epidemiological interpretation of serological responses to leptospiral serovars in sheep

Serum samples were examined for evidence of leptospiral agglutinins from 928 sheep from 45 lines and kidneys from 12 of these lines for evidence of leptospiral infection. All sheep had been submitted for slaughter at meat works in the Manawatu. Serological results were analysed using the results at a minimum serum dilution in the microscopic agglutination test (MAT) of 1:24 and at a minimum dilution of 1:48. It was shown that a minimum dilution of 1:24 resulted in many non-specific or cross-reactions. A minimum dilution of 1:48 was more accurate for detecting the serological prevalence of specific agglutinins to leptospires in ovine sera. Twenty percent of the sheep had titres of 1:48 or greater to hardjo, 3.8% to pomona, 2.6% to tarassovi, 2.3% to copenhageni and 2.7% to ballum. No titres of 1:48 or greater to australis were detected. Serovar hardjo was isolated from the kidneys of three animals in one line. Eighteen months later 291 serum samples and 95 urine samples were collected from live animals on the property from which the three hardjo infected animals originated. No titres to hardjo were detected in the sera of lambs, but a serological prevalence of 44% and 84% to this serovar was demonstrated in the hoggets and ewes respectively. No leptospires were demonstrated in any of the urine samples. These results show that sporadic infection of sheep with hardjo can occur but they also indicate that infection with this serovar is not endemic and that sheep are unlikely to act as maintenance hosts for hardjo in New Zealand.     

Histochemical skeletal muscle fibre types in the sheep

In this study, the differentiation of adult and postnatal muscle fibres in sheep longissimus thoracis muscle has been characterized. By using a variety of histochemical methods, we have investigated the m-ATPase and metabolic activities of skeletal muscle fibres in adult sheep and lambs aged between 1 day and 3 months. Types I, IIA, IIB and IIC fibres were identified. The results showed that the interpretation of the fibre type composition depends on the methods used. The findings also revealed that the fibre types IIA and IIB can be separated histochemically in sheep by using the correct m-ATPase technique, even at early stages of postnatal development, and that the origin of the four different fibres of the adult can be traced back to early postnatal stages.     

Cellular responses in the skin of merino sheep to repeated inoculation with Dermatophilus congolensis

The cellular response in the skin of Merino sheep was examined after three successive inoculations with Dermatophilus congolensis. There was a massive neutrophil influx into the infected epidermis and underlying dermis at 4-10 days after the first inoculation. A lymphocyte-macrophage response occurred at 10-12 days, followed by a plasma cell response at 14-38 days. Resolution of skin lesions after the first inoculation corresponded to the time when the plasma cell response in the skin was most intense. A second inoculation with D. congolensis, 70 days after the first, failed to produce skin lesions typical of dermatophilosis. Typical lesions of dermatophilosis did develop after a third inoculation of the same sheep 140 days after the first inoculation, but the lesions resolved in most sheep within 13 days. Dermatophilosis did not develop in some of these sheep at sites inoculated with 100-1000-fold lower infective doses of D. congolensis, whereas control sheep did develop lesions.     

Systemic responses to challenge infection withHaemonchus contortus in immune Merino sheep

Some systemic responses to single-dose infection with 10 000Haemonchus contortus infective larvae were examined in sheep already shown to have protective immunity against the parasite. The major haematological finding was a neutrophil leukocytosis that occurred after the infections became patent but not during the pre-patent period. There was no definitive eosinophilia and no discernible change in the erythrocyte parameters. Systemic hyperthermia was not conclusively evident during the pre-patent period. Enzyme-linked-immunosorbent assays (ELISA) were used to measure the secondary anti-helminth antibody response in serum during the pre-patent period when the establishment of patent infection is resisted. These ELISAs employed preparations from adult worms to represent the parasitic stages of the worm, preparations from infective larvae to represent the pre-parasitic stages of the worm, and exsheathing fluid, which is the soluble material obtained whenH. contortus larvae undergo ecdysis and transform from the pre-parasitic to the parasitic phase. Antibody responses to the three preparations differed qualitatively, indicating the presence of three different but perhaps overlapping sets of antigens. The three peaks in antibody against exsheathing fluid may reflect the pulses of antigen delivered to sheep as the parasite undergoes its three moults within the host.Abbreviations ELISA enzyme-linked immunosorbent assay - EDTA ethylene diamine tetraacetate     

Endocrine and metabolic responses to halothane and pentobarbitone anaesthesia in sheep

Some metabolic and endocrine responses to anaesthesia in sheep were studied. Adult sheep were anaesthetised with thiopentone and halothane (n=9), acepromazine, thiopentone and halothane (n=8) and pentobarbitone (n=10) on separate occasions. Routine cardiovascular monitoring was carried out and blood samples were taken for assay of cortisol, adrenocorticotrophic hormone (ACTH), arginine vasopressin (AVP), glucose and lactate. Halothane anaesthesia induced hypotension, hypercapnia and respiratory acidosis. Sheep anaesthetised with pentobarbitone were also hypercapnic and acidotic but did not develop hypotension. Plasma cortisol, ACTH and AVP (mean maximum values: cortisol: 83 ng/ml, ACTH 278 ng/ml, AVP 135 pg/ml), increased during halothane anaesthesia but did not change significantly from control values during pentobarbitone anaesthesia (mean maximum values: cortisol: 30 ng/ml, ACTH 71 ng/ml, AVP 7.8 pg/ml). Glucose tended to increase during both halothane and pentobarbitone anaesthesia but lactate decreased. It is not clear what facet of halothane anaesthesia evokes the stress response but it may be associated with cardiovascular depression.     

The epidemiological interpretation of serological responses to leptospiral serovars in sheep.

Serum samples were examined for evidence of leptospiral agglutinins from 928 sheep from 45 lines and kidneys from 12 of these lines for evidence of leptospiral infection. All sheep had been submitted for slaughter at meat works in the Manawatu.

Serological results were analysed using the results at a minimum serum dilution in the microscopic agglutination test (MAT) of 1:24 and at a minimum dilution of 1:48. It was shown that a minimum dilution of 1:24 resulted in many non-specific or cross-reactions. A minimum dilution of 1:48 was more accurate for detecting the serological prevalence of specific agglutinins to leptospires in ovine sera. Twenty percent of the sheep had titres of 1:48 or greater to hardjo, 3.8% to pomona, 2.6% to tarassovi, 2.3% to copenhageni and 2.7% to ballum. No titres of 1:48 or greater to australis were detected. Serovar hardjo was isolated from the kidneys of three animals in one line.

Eighteen months later 291 serum samples and 95 urine samples were collected from live animals on the property from which the three hardjo infected animals originated. No titres to hardjo were detected in the sera of lambs, but a serological prevalence of 44% and 84% to this serovar was demonstrated in the hoggets and ewes respectively. No leptospires were demonstrated in any of the urine samples.

These results show that sporadic infection of sheep with hardjo can occur but they also indicate that infection with this serovar is not endemic and that sheep are unlikely to act as maintenance hosts for hardjo in New Zealand.     

Differences between breeds of sheep in their responses to Bacteroides nodosus vaccines

In a field experiment on irrigated pasture, sheep of several breeds were vaccinated twice, subcutaneously, in the upper neck, with Bacteroides nodosus vaccine containing either depiliated cells (DC vaccine), or whole, piliated cells (WC vaccine) and the responses were measured over the following 14 weeks. DC vaccine was as effective as WC vaccine in protecting against the development of foot-rot under conditions of moderate challenge, although the WC vaccine induced significantly higher pilus agglutinating antibody titres. Foot-rot developed in significantly more vaccinated Merinos (Peppin and Saxon strains) than in Romney Marsh, Dorset Horn or Border Leicester breeds. Agglutinating antibody titres after WC vaccination were significantly lower in the Peppin Merino than in the other sheep for the first 6 weeks, while after DC vaccination the titres remained elevated longer in the Border Leicester and Saxon Merino and were significantly higher from 6 weeks onwards. Reactions at the inoculation sites were generally larger in the British breeds than in the Merinos and among the former the reactions were largest, most numerous and most frequently discharged their contents in the Dorset Horn. Bodyweight gains in all vaccinated sheep were initially reduced, compared with controls, but the differences were no longer significant after the eighth week.     

Niacin supplementation induces type II to type I muscle fiber transition in skeletal muscle of sheep

Background

It was recently shown that niacin supplementation counteracts the obesity-induced muscle fiber transition from oxidative type I to glycolytic type II and increases the number of type I fibers in skeletal muscle of obese Zucker rats. These effects were likely mediated by the induction of key regulators of fiber transition, PPARδ (encoded by PPARD), PGC-1α (encoded by PPARGC1A) and PGC-1β (encoded by PPARGC1B), leading to type II to type I fiber transition and upregulation of genes involved in oxidative metabolism. The aim of the present study was to investigate whether niacin administration also influences fiber distribution and the metabolic phenotype of different muscles [M. longissimus dorsi (LD), M. semimembranosus (SM), M. semitendinosus (ST)] in sheep as a model for ruminants. For this purpose, 16 male, 11 wk old Rhoen sheep were randomly allocated to two groups of 8 sheep each administered either no (control group) or 1 g niacin per day (niacin group) for 4 wk.

Results

After 4 wk, the percentage number of type I fibers in LD, SM and ST muscles was greater in the niacin group, whereas the percentage number of type II fibers was less in niacin group than in the control group (P < 0.05). The mRNA levels of PPARGC1A, PPARGC1B, and PPARD and the relative mRNA levels of genes involved in mitochondrial fatty acid uptake (CPT1B, SLC25A20), tricarboxylic acid cycle (SDHA), mitochondrial respiratory chain (COX5A, COX6A1), and angiogenesis (VEGFA) in LD, SM and ST muscles were greater (P < 0.05) or tended to be greater (P < 0.15) in the niacin group than in the control group.

Conclusions

The study shows that niacin supplementation induces muscle fiber transition from type II to type I, and thereby an oxidative metabolic phenotype of skeletal muscle in sheep as a model for ruminants. The enhanced capacity of skeletal muscle to utilize fatty acids in ruminants might be particularly useful during metabolic states in which fatty acids are excessively mobilized from adipose tissue, such as during the early lactating period in high producing cows.