Assessment of histamine, bradykinin, prostaglandins E1 and E2 and carrageenin as vascular permeability agents in the horse

The vascular leakage induced by histamine, bradykinin, serotonin and prostaglandin E1 and E2 was assessed. The test agents were injected intradermally into the shaved thoracic skin of horses and the vascular leakage estimated either semi-quantitatively by recording the diameter of the lesions or by measuring the actual volume of extravasated plasma in microliters using iodine-125-labelled human serum albumin (125I-HSA) as a marker in the blood plasma. Using the latter method, the vascular leakage induced by carrageenin and the effect of coadministered prostaglandins E1 and E2 upon the vascular leakage of both histamine and bradykinin were also investigated. No obvious lesions resulted when serotonin (10(-2) mol/l) was injected but histamine and bradykinin produced circular lesions which increased in diameter for approximately 30 min. The size of the lesions and volume of extravasated plasma was dose dependent. On a molar basis, bradykinin (10(-6) mol/l, 10(-5) mol/l) was more potent than histamine but they were equipotent at 10(-4) mol/l. The size of the lesions induced by carrageenin were independent of their anatomical location on the thorax. Except for the second hour, the hourly volume of vascular leakage increased until the fifth hour when the experiment was concluded. The maximum vascular leakage resulting from the injection of prostaglandin E1 or E2 (1, 10, 100 or 1000 ng) was 7 microliters but when co-administered with bradykinin (10(-6) mol/l), the volume of leaked plasma increased from 29 to 78 microliters. No synergy was observed when either prostaglandin was co-administered with histamine (10(-5) mol/l).     

Sedative effects of hydromorphone or oxymorphone with or without acepromazine in dogs

Hydromorphone is an agonist opioid with potency approximately five times that of morphine and half that of oxymorphone. The purpose of this study was to compare hydromorphone with oxymorphone, with or without acepromazine, for sedation in dogs, and to measure plasma histamine before and after drug administration. Ten dogs received IM hydromorphone (H; 0.2 mg kg^?1), oxymorphone (O; 0.1 mg kg^?1), hydromorphone with acepromazine (H; 0.2 mg kg^?1, A; 0.05 mg kg^?1) or oxymorphone with acepromazine (O; 0.1 mg kg^?1, A; 0.05 mg kg^?1) in a randomized Latin‐square design. Sedation score, heart rate, respiratory rate, blood pressure, and SpO₂ were recorded at baseline and every 5 minutes after drug administration up to 25 minutes. Plasma histamine was measured at baseline and at 25 minutes post‐drug administration. Data were analyzed with repeated measures anova . Mean ± SD body weight was 21.62 ± 1.54 kg. Mean ± SD age was 1.07 ± 0.19 years. Sedation score was significantly greater for OA after 5 minutes than O alone (4.1 ± 3.5 versus 1.9 ± 1.5) and for HA after 15 minutes than H alone (8.6 ± 2.9 versus 5.9 ± 2.5). There was no significant difference in sedation between H and O at any time point. There was no significant difference between groups at any time with respect to heart rate, respiratory rate, blood pressure or SpO₂. Mean ± SD plasma histamine (nM ml^?1) for all groups was 1.72 ± 2.69 at baseline and 1.13 ± 1.18 at 25 minutes. There was no significant change in plasma histamine concentration in any group. Hydromorphone is effective for sedation in dogs and does not cause measurable increase in histamine. Sedation with hydromorphone is enhanced by acepromazine.     

Histamine relaxes constricted trachea and bronchi of horse

Horse trachea and bronchi contracted to slow-reacting substance of anaphylaxis, carbachol, 5-hydroxytryptamine, histamine, 2-methyl-histamine, bradykinin and prostaglandin F₂. Prostaglandins E₁ and E₂, isoprenaline and 4-methylhistamine caused relaxation of carbachol-contracted bronchi. Mepyramine (H₁-blocker) specifically inhibited histamine bronchoconstriction. In the presence of mepyramine, histamine caused relaxation of carbachol-contracted bronchi. Metiamide (H₂-blocker) inhibited histamine bronchorelaxation but not relaxation of the trachea. This suggests (1) the presence of both H₁- and H₂-receptors mediating bronchoconstriction and relaxation respectively (2) the existence of an atypical histamine receptor in the trachea. The study suggests that in equine respiratory hypersensitivity under therapy with classical (H₁) antihistaminics, further release of histamine may exert a beneficial broncholytic effect on airways contracted by other chemical mediators of immediate-type inflammation.     

Effects of histamine on chicken airways

This study showed that chicken bronchus is relatively insensitive to histamine. H₁ histamine receptors appear to mediate bronchoconstriction to histamine. In the presence of mepyramine, high doses of histamine (10^–3 to 5×10^–3M) produce weak bronchorelaxation which is susceptible to cimetidine.Cimetidine alone potentiated the bronchoconstriction to histamine. This shows the presence of a small population of histamine H₂-receptors in chicken bronchus, which may modulate bronchospasm during pulmonary allergic reactions.     

Recombinant unpurified rETXH106P/CTB-rETXY196E protects rabbits against Clostridium perfringens epsilon toxin

Epsilon toxin (ETX), produced by Clostridium perfringens types B and D, has been touted as a potential biological weapon and is known to induce fatal enterotoxemia in a variety of livestock animals. For the efficient production of recombinant proteins with the objective of investigating the effects of different recombinant vaccines against ETX, a bicistronic design (BCD) expression system including the ETX coding sequence with mutation of amino acid 106 from Histidine to Proline (ETX^H106P) in the first cistron, followed by Cholera Toxin B (CTB) linked with the ETX coding sequence with mutation of amino acid 196 from Tyrosine to Glutamic acid (ETX^Y196E) in the second cistron, was generated under the control of a single promoter. Rabbits were immunized twice with five inactivated recombinant Escherichia coli (E. coli) vaccines containing 100 µg/ml of the recombinant mutant rETX^H106P/CTB-rETX^Y196E proteins mixed with different adjuvants. Apart from rETX^H106P/CTB-rETX^Y196E-IMS1313-vaccinated rabbits, the neutralizing antibody titers of rETX^H106P/CTB-rETX^Y196E-vaccinated rabbits were higher after the initial immunization than those administered the ETX toxoid or current commercial vaccines. rETX^H106P/CTB-rETX^Y196E mixed with ISA201 induced the highest neutralizing antibody titer of 120 after the first immunization, suggesting that 0.1 ml of pooled sera could neutralize 120× mouse LD₁₀₀ (100% lethal dose) of ETX. Following the second vaccination, rETX^H106P/CTB-rETX^Y196E mixed with ISA201 or GR208 produced the highest neutralizing titer of 800. Rabbits from all vaccinated groups were completely protected from a 2× rabbit LD₁₀₀ of ETX challenge. These results show that these novel recombinant proteins can induce a strong immune response and represent potential targets for the development of a commercial vaccine against the C. perfringens epsilon toxin.     

Kinetics, dose response, tachyphylaxis and cross-tachyphylaxis of vascular leakage induced by endotoxin, zymosan-activated plasma and platelet-activating factor in the horse

Vascular leakage induced by intradermal injection of endotoxin, zymosan-activated plasma (ZAP) and platelet-activating factor (PAF) was measured in nine Thoroughbreds using 125-iodine human serum albumin (¹²⁵I-HSA) as a marker in the blood. ZAP and PAF produced dose-dependent increases in vascular permeability with the maximum occurring within the first 15 min after injection. The vascular leakage induced by endotoxin was also dose-dependent, but the maximum occurred 2 h after intradermal injection. Intradermal sites previously injected with endotoxin were refractory to a second injection of endotoxin for up to 5 days. However, sites injected with endotoxin and re-injected with either ZAP or PAF remained responsive with increased vascular leakage compared to saline injected control sites re-injected with either ZAP or PAF. Diminished response to endotoxin challenge may contribute to the poor prognosis of endotoxaemia in the horse.     

Influence of estrogen on acetylcholinesterase activity in primary cultures of cerebral cells from neonatal rats

To estimate the influence of estrogen on the functional development of the central nervous system during the neonatal period, several doses of estradiol‐17β (E₂) were treated to cultured cells from the cerebral cortex of neonatal rats and the acetylcholinesterase (AChE) activity was examined. E₂ was added to give the following final concentrations: 0, 10^?10, 10^?9, and 10^?8 M. After 72 h of incubation, all cells were obtained from dishes to determine the AChE activity. Although apparent morphological changes were not observed among treatments cultured for 72 h, E₂ suppressed dose‐dependently the spontaneous increase of AChE activity in cerebral cells. Furthermore, a single dose of tamoxifen, an E₂ receptor binding molecule with agonist and antagonist properties, also acted in a similar manner as E₂. These findings suggest that the functional development of the cerebral cortex, at least the cholinergic system, during the neonatal period is regulated by E₂.     

Involvement of ATP‐sensitive K+ channels in the peripheral antinociceptive effect induced by ketamine

ObjectiveTo investigate the contribution of K⁺ channels on peripheral antinociception induced by ketamine.Study designProspective experimental study.Animals110 male Wistar rats weighing 160–200 g.MethodsThe paw pressure required to elicit limb flexion was designated as the nociceptive threshold. Hyperalgesia was induced by intraplantar injection of prostaglandin E₂. All drugs were administered locally into the right hind paw of rats. Ketamine was administered into the right hind paw 2 hours and 55 minutes after local injection of PGE₂. Tetraethylammonium was administered 30 minutes prior to ketamine and the other K⁺ channel blockers, glibenclamide, dequalinium and paxilline, were administered 5 minutes prior to ketamine.ResultsProstaglandin E₂ (2 μg per paw) induced hyperalgesia. Ketamine (10, 20, 40 and 80 μg per paw) elicited a local peripheral antinociceptive effect that was antagonized by a specific blocker of ATP‐sensitive K⁺ channels, glibenclamide (20, 40 and 80 μg per paw). In another experiment, the non‐selective voltage‐dependent K⁺ channel blocker tetraethylammonium (30 μg per paw) and small and large conductance blockers of Ca²⁺‐activated K⁺ channels, dequalinium (50 μg per paw) and paxilline (20 μg per paw), were ineffective at blocking the effect of a local ketamine injection.Conclusions and clinical relevanceAnalysis of these results provides evidence that ketamine, may in part, induce peripheral antinociceptive effects by ATP‐sensitive K⁺ channel pathway activation.     

Effects of continuous intravenous infusion of morphine and morphine‐tramadol on the minimum alveolar concentration of sevoflurane and electroencephalographic entropy indices in dogs

ObjectiveTo compare the effects of continuous rate infusions (CRIs) of intravenous (IV) morphine and morphine-tramadol on the minimum alveolar concentration (MAC) of sevoflurane, and on electroencephalographic entropy indices in dogs.DesignProspective study.AnimalsEight young, healthy German shepherds, weighing 26.3 ± 3.1 kg (mean ± SD).MethodsAnaesthesia was induced and maintained with sevoflurane. A standard tail-clamp technique was used for MAC determination. Within one anaesthetic period, MAC was first determined during sevoflurane anaesthesia alone (MAC_B); then during morphine infusion (MAC_M), (loading dose 0.5 mg kg⁻¹IM; CRI, 0.2 mg kg⁻¹hour⁻¹⁾ then finally during morphine-tramadol infusion (tramadol loading dose 1.5 mg kg⁻¹IV; CRI, 2.6 mg kg⁻¹ hour⁻¹) (MAC_MT). At each change, periods of 45 minutes were allowed for equilibration. Stated entropy (SE), response entropy (RE), and RE-SE differences were measured five minutes prior to and during tail clamping.ResultsThe MAC_B was 2.1 ± 0.3vol%. The morphine and morphine-tramadol infusions reduced MAC to 1.6 ± 0.3vol% and 1.3 ± 0.3vol%, respectively. MAC was decreased below baseline more during morphine-tramadol than during morphine alone (39 ± 9% versus 25 ± 6%, respectively; p = 0.003). All SE and RE and most RE-SE differences were increased significantly (p < 0.05) over pre-stimulation in all groups when the dogs responded purposefully to noxious stimulation. When no response to noxious stimulation occurred, the entropy indices did not change.Conclusion and clinical relevanceIn dogs, combined morphine-tramadol CRI decreased sevoflurane MAC more than morphine CRI alone. Entropy indices changed during nociceptive responses in anaesthetized animals, suggesting that entropy measurements may be useful in determining anaesthetic depth in dogs.     

Effects of methadone, alone or in combination with acepromazine or xylazine, on sedation and physiologic values in dogs

ObjectiveTo evaluate the effects of methadone, administered alone or in combination with acepromazine or xylazine, on sedation and on physiologic values in dogs.Study designRandomized cross-over design.AnimalsSix adult healthy mixed-breed dogs weighing 13.5 ± 4.9 kg.MethodsDogs were injected intramuscularly with physiologic saline (Control), or methadone (0.5mg kg⁻¹) or acepromazine (0.1 mg kg⁻¹) or xylazine (1.0 mg kg⁻¹), or acepromazine (0.05 mg kg⁻¹) plus methadone (0.5 mg kg⁻¹) or xylazine (0.5 mg kg⁻¹) plus methadone (0.5 mg kg⁻¹) in a randomized cross-over design, with at least 1-week intervals. Sedation, pulse rate, indirect systolic arterial pressure, respiratory rate (RR), body temperature and pedal withdrawal reflex were evaluated before and at 15-minute intervals for 90 minutes after treatment.ResultsSedation was greater in dogs receiving xylazine alone, xylazine plus methadone and acepromazine plus methadone. Peak sedative effect occurred within 30 minutes of treatment administration. Pulse rate was lower in dogs that received xylazine either alone or with methadone during most of the study. Systolic arterial pressure decreased only in dogs receiving acepromazine alone. When methadone was administered alone, RR was higher than in other treatments during most of the study and a high prevalence of panting was observed. In all treatments body temperature decreased, this effect being more pronounced in dogs receiving methadone alone or in combination with acepromazine. Pedal withdrawal reflex was absent in four dogs receiving methadone plus xylazine but not in any dog in the remaining treatments.Conclusions and clinical relevanceMethadone alone produces mild sedation and a high prevalence of panting. Greater sedation was achieved when methadone was used in combination with acepromazine or xylazine. The combination xylazine–methadone appears to result in better analgesia than xylazine administered alone. Both combinations of methadone/sedative were considered effective for premedication in dogs.     

Effects of constant rate infusions of dexmedetomidine,remifentanil and their combination on minimum alveolar concentration of sevoflurane in dogs

ObjectiveTo evaluate the effects of constant rate infusions (CRIs) of dexmedetomidine and remifentanil alone and their combination on minimum alveolar concentration (MAC) of sevoflurane in dogs.Study designRandomized crossover experimental study.AnimalsA total of six (three males, three females) healthy, adult neutered Beagle dogs weighing 12.6 ± 1.4 kg.MethodsAnesthesia was induced with sevoflurane in oxygen until endotracheal intubation was possible and anesthesia maintained with sevoflurane using positive-pressure ventilation. Each dog was anesthetized five times and was administered each of the following treatments: saline (1 mL kg^–1 hour^–1) or dexmedetomidine at 0.1, 0.5, 1.0 or 5.0 μg kg^–1 loading dose intravenously over 10 minutes followed by CRI at 0.1, 0.5, 1.0 or 5.0 μg kg^–1 hour^–1, respectively. Following 60 minutes of CRI, sevoflurane MAC was determined in duplicate using an electrical stimulus (50 V, 50 Hz, 10 ms). Then, CRI of successively increasing doses of remifentanil (0.15, 0.60 and 2.40 μg kg^–1 minute^–1) was added to each treatment. MAC was also determined after 30 minutes equilibration at each remifentanil dose. Isobolographic analysis determined interaction from the predicted doses required for a 50% MAC reduction (ED₅₀) with remifentanil, dexmedetomidine and remifentanil combined with dexmedetomidine, with the exception of dexmedetomidine 5.0 μg kg^–1 hour^–1, obtained using log-linear regression analysis.ResultsThe sevoflurane MAC decreased dose-dependently with increasing infusion rates of dexmedetomidine and remifentanil. Remifentanil ED₅₀ values were lower when combined with dexmedetomidine than those obtained during saline–remifentanil. Synergistic interactions between dexmedetomidine and remifentanil for MAC reduction occurred with dexmedetomidine at 0.5 and 1.0 μg kg^–1 hour^–1.Conclusions and clinical relevanceCombined CRIs of dexmedetomidine and remifentanil synergistically resulted in sevoflurane MAC reduction. The combination of dexmedetomidine and remifentanil effectively reduced the requirement of sevoflurane during anesthesia in dogs.     

Plasma histamine concentrations in horses administered sodium penicillin,guaifenesin–xylazine–ketamine and isoflurane with morphine or butorphanol

ObjectiveVarious drugs administered to horses undergoing surgical procedures can release histamine. Histamine concentrations were evaluated in horses prepared for surgery and administered butorphanol or morphine intraoperative infusions.Study designProspective studies with one randomized.AnimalsA total of 44 client-owned horses.MethodsIn one study, anesthesia was induced with xylazine followed by ketamine–diazepam. Anesthesia was maintained with guaifenesin–xylazine–ketamine (GXK) during surgical preparation. For surgery, isoflurane was administered with intravenous (IV) morphine (group M: 0.15 mg kg^–1 and 0.1 mg kg^–1 hour^–1; 15 horses) or butorphanol (group B: 0.05 mg kg^–1 and 0.01 mg kg^–1 hour^–1; 15 horses). Histamine and morphine concentrations were measured using enzyme-linked immunoassay before opioid injection (time 0), and after 1, 2, 5, 30, 60 and 90 minutes. In a subsequent study, plasma histamine concentrations were measured in 14 horses before drug administration (baseline), 15 minutes after IV sodium penicillin and 15 minutes after starting GXK IV infusion. Statistical comparison was performed using anova for repeated measures. Pearson correlation compared morphine and histamine concentrations. Data are presented as mean ± standard deviation. Significance was assumed when p ≤ 0.05.ResultsWith histamine, differences occurred between baseline (3.2 ± 2.4 ng mL^–1) and GXK (5.2 ± 7.1 ng mL^–1) and between baseline and time 0 in group B (11.9 ± 13.4 ng mL^–1) and group M (11.1 ± 12.4 ng mL^–1). No differences occurred between baseline and after penicillin or between groups M and B. Morphine concentrations were higher at 1 minute following injection (8.1 ± 5.1 ng mL^–1) than at 30 minutes (4.9 ± 3.1 ng mL^–1) and 60 minutes (4.0 ± 2.5 ng mL^–1). Histamine correlated with morphine at 2, 30 and 60 minutes.Conclusions and clinical relevanceGXK increased histamine concentration, but concentrations were similar with morphine and butorphanol.     

Actions of histamine and other substances on the airway smooth muscle of swine (in vitro)

Isolated swine tracheal and bronchial smooth muscle strips contracted to carbachol and histamine. 5-HT, PGF₂ and bradykinin were inactive. Isoprenaline (a -adrenoceptor agonist), PGE₁, PGE₂, bradykinin and 4-methylhistamine (4-MeH: a specific H₂-receptor agonist) produced relaxation of carbachol contracted trachea and bronchi. Mepyramine (a specific H₁-antagonist) blocked histamine-induced contractions. After H₁-blockade, histamine induced relaxation of carbachol contracted trachea and bronchi. Metiamide (an H₂-receptor antagonist) antagonizes relaxation to 4-MeH and histamine, but not to isoprenaline. The results of this investigation showed the presence of both H₁- and H₂-histamine receptors mediating constriction and relaxation respectively in the airways of swine.     

Comparative cardiovascular, analgesic, and sedative effects of medetomidine, medetomidine–hydromorphone, and medetomidine–butorphanol in dogs

The purpose of this study was to determine the cardiovascular, analgesic, and sedative effects of IV medetomidine (M, 20 µg kg^?1), medetomidine–hydromorphone (MH, 20 µg kg^?1 ? 0.1 mg kg^?1), and medetomidine–butorphanol (MB, 20 µg kg^?1 ? 0.2 mg kg^?1) in dogs. Using a randomized cross‐over design and allowing 1 week between treatments, six healthy, mixed‐breed dogs (five males and one female) weighing 20 ± 3 kg, were induced to anesthesia by face‐mask administration of 2.9% ET sevoflurane to facilitate instrumentation prior to administration of the treatment combinations. Dogs were intubated and instrumented to enable measurement of heart rate (HR), systolic arterial pressure (SAP), mean arterial pressure (MAP), diastolic arterial pressure (DAP), mean pulmonary arterial pressure (PAP), pulmonary arterial occlusion pressure (PAOP), central venous pressure (CVP), pulmonary arterial temperature (TEMP), and cardiac output via thermodilution using 5 mL of 5% dextrose, and recording the average of the three replicate measurements. Cardiac index (CI) and systemic (SVR) and pulmonary vascular resistances were calculated. After instrumentation was completed, administration of sevoflurane was discontinued, and the dogs were allowed to recover for 30 minutes prior to administration of the treatment drugs. After collection of the baseline samples for blood gas analysis and recording the baseline cardiovascular variables, the test agents were administered IV over 10 seconds and the CV variables recorded at 5, 10, 15, 30, 45, and 60 minutes post‐injection. In addition, arterial blood was sampled for blood gas analysis at 15 and 45 minutes following injection. Intensity and duration of analgesia (assessed by toe‐pinch response using a hemostat) and level of sedation were evaluated at the above time points and at 75 and 90 minutes post‐injection. Data were analyzed using anova for repeated measures with posthoc differences between means identified using Bonferroni's method (p < 0.05). Administration of M, MH, or MB was associated with increases in SAP, MAP, DAP, PAP, PAOP, CVP, SVR, and TEMP and with decreases in HR and CI. No differences in CV variables between treatment groups were identified at any time. PaO₂ increased over time in all groups and was significantly higher when MH was compared with M. At 45 minutes, PaO₂ tended to decrease over time and was significantly lower when MH and MB were compared with M at 15 minutes. Analgesia scores for MH and MB were significantly higher compared with M through 45 minutes, while analgesia scores for MH were significantly higher compared with M through 90 minutes. Sedation scores were higher for MH and MB compared with M throughout 90 minutes. Durations of lateral recumbency were 108 ± 10.8, 172 ± 15.5, and 145 ± 9.9 minutes for M, MH, and MB, respectively. We conclude that MH and MB are associated with improved analgesia and sedation and have similar CV effects when compared with M.     

Absorption and metabolism of estrogens from the stomach and duodenum of pigs

To determine the absorption and metabolism of 17β-estradiol (E₂) by the stomach and liver of the pig, crystalline E₂ was placed in the stomach of prepubertal gilts. Blood samples were subsequently obtained from the hepatic portal and jugular veins and plasma was assayed for E₂, estrone (E₁), 17β-estradiol-glucuronide (E₂G), estrone-glucuronide (E₁G) and estrone-sulfate (E₁S). Concentrations of E₂, E₁, E₂G and E₁S rose in the hepatic portal vein within five min and remained elevated for several hr. Concentration of E₂ represented only 6% of the total estrogen detected in the hepatic portal vein during the sampling period, indicating that most of the E₂ was converted or conjugated prior to entering the hepatic portal vein. The metabolism of E₂ presumably occurred in the stomach mucosa because food had been withheld for 26 hr before infusion of E₂. Concentrations of E₂G, E₁G and E₁S, but not E₂ and E₁, rose in the jugular vein and remained elevated for several hr. The lack of a rise in E₂ and E₁ in the jugular vein indicates that the E₂ and E₁ from the hepatic portal vein were completely converted and/or removed by the liver. Most of E₂ was converted to E₁ and then to E₁G. The infusion of bile containing normal estrogens from pregnant gilts into the duodenum of prepubertal gilts resulted in a peak of E₁G and E₂G in the hepatic portal and jugular veins within a few minutes. This was followed in about 180 min by a second sustained rise. The first peak was essentially abolished by extracting E₁ and E₂ from the bile before infusion. The second peak failed to occur in gilts given antibiotics orally to reduce gut bacteria before infusion of bile.     

Effect of estradiol‐17β during in vitro growth culture on the growth,maturation, cumulus expansion and development of porcine oocytes from early antral follicles

Growing porcine oocytes from early antral follicles can acquire meiotic and developmental competence under suitable culture conditions, but at lower rates compared to full‐grown oocytes. We postulated that estradiol‐17β (E₂) supported the acquisition of meiotic and developmental competence as well as cumulus‐expansion ability during growth culture. Growing oocytes from early antral follicles (1.2 to 1.5 mm in diameter) were grown in vitro for 5 days in a medium containing 0, 10^?7, 10^?6, 10^?5 or 10^?4 mol/L E₂; after in vitro maturation, 35, 58, 47, 74 and 49% of oocytes matured to metaphase II, 25, 79, 77, 90 and 97% acquired cumulus‐expansion ability, and 23, 54, 63, 89 and 64% were fully surrounded by cumulus cells, respectively. Following maturation, electro‐stimulation was applied to the oocytes grown with 10^?5 mol/L E₂. After 6 days of culture, in vitro‐grown oocytes developed to the blastocyst stage at a rate similar to that for full‐grown oocytes (31% and 40%, respectively). Therefore, we suggest that the use of E₂ during growth culture improves the meiotic and developmental competence of oocytes, cumulus‐expansion ability, and cumulus cell attachment to the oocytes.     

The suitability of medetomidine sedation for intradermal skin testing in dogs

The objective of this study was to determine the suitability of medetomidine sedation for facilitating intradermal skin testing in dogs. Quality of sedation and immobilization, and effects of sedation on responses to intradermally injected histamine were evaluated. Ten clinically normal dogs were injected intradermally before and after medetomidine sedation (10 μg kg^?1 intravenously) with diminishing concentrations of histamine (100–10^?5μg mL^?1) and a negative control. Mean wheal responses at injection sites were compared before and during sedation, and no significant suppression of responses occurred during sedation. Medetomidine produced sedation that notably increased the ease of performing multiple intradermal injections in all dogs and sedative effects were rapidly reversed by the antagonist atipamezole. It was concluded that medetomidine may be an excellent sedative for facilitating intradermal skin testing in dogs provided further studies similarly reveal no inhibition of responses to intradermally injected allergens in atopic dogs.     

The Effects of Growth Factor on Testicular Germ Cell Apoptosis in the Stallion

Apoptosis is necessary for both initiation and control of spermatogenesis; however, an increase in apoptosis can lead to subfertility/infertility in stallions, causing substantial financial loss in the equine industry. The ability of stem cell factor (SCF), leukemia-inhibiting factor (LIF), granulocyte–macrophage colony-stimulating factor (GM-CSF), and estradiol (E₂), alone or in combination, to prevent apoptosis of germ cells in short-term equine testicular cultures was examined. Testicular tissue was sectioned into approximately 2-mm cubes and placed in media-filled culture chambers. Concentrations of SCF (100 ng/mL), LIF (10 ng/mL), GM-CSF (5 ng/mL), and E₂ (10⁻⁹ mol/l) were added alone or in combination to each well. After 6 hours in culture, the tissue was fixed and immunohistochemically (terminal deoxynucleotidyl transferase-mediated nick-end labeling; TUNEL) stained for apoptosis detection. Apoptotic cells per 100 Sertoli cell nuclei within seminiferous tubules were counted until the 500^th Sertoli cell nuclei was reached. This counting procedure was used for each slide. An analysis of variance (ANOVA) with a Tukey's test was used to compare apoptotic rates. In comparison with the control, GM-CSF alone lowered apoptosis by 34.77%. GM-CSF–treated tissue combined with SCF and LIF as well as GM-CSF combined with SCF, LIF, and E₂ reduced apoptosis when compared with the control (37.45% and 44.40%, respectively) or other treatment combinations. GM-CSF alone reduced apoptosis; results suggest possible synergy for the combinations of SCF and LIF with GM-CSF and for E₂ with SCF, LIF, and GM-CSF.     

Orbital malignant schwannoma and embryonal rhabdomyosarcoma in rats caused by leakage of locally injected nickel subsulfide to the orbit

Nickel subsulfide (Ni₃S₂) is known to induce intraocular neoplasms when injected intravitreally into the eyes of rats. Here, we found two extraocular orbital neoplasms in two different rat strains, presumably due to the leakage of locally injected Ni₃S₂ to the extraocular orbital tissues. In the F344/DuCrlCrlj rat, an orbital mass arose at 30 weeks after injection, and invaded into the cranium. Histologically, the orbital mass was composed of areas arranged in parallel bundles formed by densely packed elongated or spindle-shaped cells with indistinct cytoplasmic borders, and of areas of hypocellular arrangement consisting of round cells in eosinophilic myxoid-like substances. Metastases were observed in the right submandibular and cervical lymph nodes. The neoplastic cells were immunopositive for S-100 protein and vimentin. Transmission electron microscopy revealed that the neoplastic cells had cellular processes and pericytoplasmic basal laminae. In the RccHan^TM:WIST rat, an orbital mass arose at 36 weeks after injection. Histologically, the mass consisted of rhabdoid-like large round cells with proliferation of small round-to-polygonal cells, and these neoplastic cells infiltrated into the extraocular muscles. Immunohistochemically, the neoplastic cells were positive for desmin and vimentin. Transmission electron microscopy detected immature myofibrils with Z-band structures in the cytoplasm of these neoplastic cells. Consequently, the tumors were diagnosed as an orbital malignant schwannoma in an F344/DuCrlCrlj rat and an orbital embryonal rhabdomyosarcoma in a RccHan^TM:WIST rat. The results of this case report suggest that leakage of Ni₃S₂ to the orbit caused the induction of orbital malignant schwannoma or rhabdomyosarcoma in rats.     

Endotoxaemia in dairy cattle:Mechanism of reticulorumen stasis

The objective of this study was to determine whether blockade of α₋₂ adrenergic receptors would restorereticulorumen motility during toxaemia in cows. Reticulorumen contractions were measured via a water-filled balloon connected to a pressure transducer. Intravenous infusion of endotoxin (100 ng kg⁻¹ over 30 min) significantly decreased the number of reticulorumen contractions. Intravenous infusion of yohimbine (125 μg kg⁻¹ over 30 min) alone did not affect reticulorumen contractions. However, when yohimbine (125 μg kg⁻¹ over 30 min) was infused concurrently with endotoxin (100 ng kg⁻¹ over 30 min), the effects of endotoxin on reticulorumen contraction frequency decreased, suggesting that endotoxaemia causes reticulorumen stasis via a mechanism that involves α₋₂ adrenergic receptors.