Intracranial elastance in isoflurane-anesthetized horses

OBJECTIVE: To determine whether high intracranial pressure (ICP) during spontaneous ventilation (SV) in anesthetized horses coincides with an increase in intracranial elastance (ie, change in ICP per unit change of intracranial volume). ANIMALS: 6 adult horses. PROCEDURE: Anesthesia was induced and maintained in each horse for 5 hours with isoflurane at a constant dose equal to 1.2 times the minimum alveolar concentration. Direct ICP measurements were obtained by use of a strain gauge transducer inserted in the subarachnoid space, and arterial blood pressure was measured from a carotid artery. Physiologic responses were recorded after 15 minutes of normocapnic controlled ventilation (CV) and then after 10 minutes of SV. Aliquots (3 mL) of CSF were removed from each horse during SV until ICP returned to CV values. Slopes of pressure-volume curves yielded intracranial elastance. RESULTS: Intracranial elastance ranged from 0.2 to 3.7 mm Hg/mL after removal of the first aliquot of CSF Slopes of pressure-volume curves were largest following removal of the initial CSF aliquot, but shallow portions of curves were detected at relatively high ICPs (25 to 35 mm Hg). A second-order relationship between SV ICP and initial intracranial elastance was found. CONCLUSIONS AND CLINICAL RELEVANCE: In horses anesthetized with isoflurane, small changes in intracranial volume can cause large changes in ICP Increased intracranial elastance could further exacerbate preexisting intracranial hypertension. However, removal of small volumes of CSF may cause rapid compensatory replacement from other intracranial compartments, which suggests steady-state maintenance of an increase in intracranial volume during isoflurane anesthesia in horses.     

Generation and characterization of anti-leptin antisera against synthetic peptides and recombinant protein

The objective of this study was to generate antisera against recombinant bovine leptin and synthetic oligopeptides corresponding to the amino acid sequence 21-40 and 91-110 of bovine leptin. Recombinant bovine leptin was raised in the 293 cells and purified from 10 L of conditioned medium and utilized for immunization. The synthetic peptides were conjugated with keyhole limpet hemocyanin and inoculated into rabbits for antibody generation. Antibody titer was monitored by enzymeimmunoassay, immunoblotting and sandwich binding assay techniques. Each of the antisera, against three different antigens, was found to react with bovine leptin. The titers of anti-peptide antisera were lower than that of anti-recombinant leptin antiserum. Since anti-recombinant leptin antiserum was not neutralized by the leptin peptides 21-40 and 91-110, it is suggested that each antiserum recognizes a distinct epitope. In immunoblot analyses, all antisera exhibited cross-reactivity with human and mouse leptins. However, in the sandwich binding assay, the combination of anti-peptide antisera and anti-recombinant leptin antiserum, originated from bovine leptin, did not cross-react with either human or mouse leptin. The discrepancy of antigenic recognition between the immunoblot analyses and sandwich assay is thought to be dependent on the conformational status of leptin molecules between the species. The antisera generated in this study, which recognized distinct epitopes of bovine leptin, will provide a useful tool for studies of bovine leptin functions.     

Changes of fructose concentrations in seminal plasma and glucose and testosterone concentrations in blood plasma in rams over the course of a year

The present study was performed to examine seasonal changes in the fructose concentrations of seminal plasma and glucose and testosterone concentrations of blood plasma over the course of a year (from November 2004 to November 2005) using 5 Suffolk rams. Osmolality of the seminal plasma was also measured. The fructose concentrations in the seminal plasma increased as the breeding season approached, with the maximum in October (179.8 mg/dl) and the minimum in May (6.9 mg/dl), although there were no significant differences during the year. Osmolality of the seminal plasma in February (304 mOsm) was significantly (P<0.05) lower than in January (325 mOsm), July (327 mOsm), and August (325 mOsm). It was also significantly (P<0.05) lower in November (308 mOsm) than in January and August. The blood plasma glucose concentration in October (79.3 mg/dl) was significantly (P<0.05) higher than in January and February (43.2 and 43.7 mg/dl, respectively). The blood plasma testosterone (T) concentrations were significantly (P<0.05) higher in September (8.5 ng/ml) and October (10.2 ng/ml) than in other months. The fructose concentrations in the seminal plasma appeared to be related to the glucose and T concentrations in the blood plasma. These results show that fructose concentrations in the seminal plasma and blood plasma glucose and T concentrations tended to increase during the breeding season, with the highest concentrations in October.     

Estimation of glomerular filtration rate in calves using the contrast medium iodixanol

To develop a simple procedure for estimating glomerular filtration rate (GFR) in calves, a three-sample method using iodixanol was first compared to that using the standard agent inulin. Iodixanol and inulin were co-administered intravenously to calves at 40 mg I/kg and 40 mg/kg, respectively, and blood was collected 30, 60, 120, and 180 min later. Serum iodixanol and inulin concentrations were separately determined by high performance liquid chromatography and colorimetry. Serum urea nitrogen (UN) and creatinine concentrations were also measured. GFR estimated by iodixanol was consistent with that using inulin in clinically healthy calves. Based on GFR estimations in healthy calves and those renal-loaded with iodixanol, it was found that the serum creatinine concentrations became elevated when GFR decreased to 60% of the reference value. In contrast, serum UN concentrations fluctuated widely, presumably due to extra-renal factors. When GFR was estimated using the three-sample method and compared with the single-blood-sample method, 62/69 (90%) of samples tested were within the agreement plots. The results demonstrated that the single-blood-sample method using iodixanol may be useful in monitoring GFR in calves.     

Seasonal variations in the incidence of pine wilt and infestation by its vector, Monochamus alternatus, near the northern limit of the disease in Japan

During the last few decades, pine wilt disease has spread to cool-climate regions in Japan and, more recently, the potential risk of it spreading into the European midwest has also become a concern. In a coastal pine stand (84.7?ha) in Akita, near the northern limit of pine wilt disease in Japan, we investigated seasonal variations in the incidence of damage caused by the disease to trees and oviposition by the disease’s insect vector, Monochamus alternatus, during a two-year period. Foliage discoloration occurred throughout each year, and its seasonal variation showed a bimodal pattern in Pinus thunbergii (a higher peak in May–June and a smaller peak in October) and a clear peak in June in P. densiflora, which differed from the patterns in seasonal variation seen for warm-climate regions. Oviposition scars by M. alternatus were found in 40–45% of the trees damaged each year. The percentage of trees that had oviposition scars was higher in P. thunbergii than in P. densiflora. This appeared to reflect the difference in seasonal discoloration pattern between the two species. Analysis of the oviposition risk showed that trees that exhibited discoloration starting between July and October had a significantly higher risk or significantly higher oviposition scar densities, particularly for those that became discolored between August and September (2.5–14.6-fold higher risk than during other months). Oviposition scar densities per damaged tree were similar within the period of higher oviposition risk. Considering both oviposition risks and scar densities, we concluded that trees with discoloration that become apparent between July and October are important targets for preventing the spread of pine wilt disease in Akita.     

Synthesis and antifungal activity of imidazole-1-carboxylates

A series of imidazole-1-carboxylates was prepared by reacting various alcohols with trichloromethyl chloroformate and imidazole or N,N'-carbonyl-diimidazole. They were tested for fungitoxic activity in vitro against two phytopathogenic fungi, Botrytis cinerea (grey mould) and Gibberella fujikuroi, and for preventive efficacy against grey mould on cucumber leaves. 1-(4-Substituted phenoxymethyl)-2,2-dimethylpropylimidazole-1-carboxylates showed excellent in-vitro activity against B. cinerea, and moderate activity against G. fujikuroi, and some of them also effectively controlled grey mould in vivo. A 1H-1,2,4-triazole derivative corresponding to an imidazole derivative did not have any activity, while a thiocarboxylate corresponding to an imidazole carboxylate showed excellent activity against both B. cinerea and G. fujikuroi.     

No Promoting Effect of Ethyl Tertiary-butyl Ether (ETBE) on Rat Urinary Bladder Carcinogenesis Initiated with N-Butyl-N-(4-hydroxybutyl)nitrosamine

The effects of ethyl tertiary-butyl ether (ETBE) on two-stage urinary bladder carcinogenesis in male F344 rats initiated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) were investigated at various dose levels with regard to possible promoting activity. Groups of 30 rats were given drinking water containing 500 ppm BBN, as an initiator, for 4 weeks and starting one week thereafter received ETBE by gavage (daily, 7 days/week) at dose levels of 0 (control), 100, 300, 500 or 1000 mg/kg/day until experimental week 36. No statistically significant differences in incidences of preneoplastic lesions, papillomas, and carcinomas of the urinary bladder were evident in rats treated with 100–1000 mg/kg/day ETBE as compared with control values. Furthermore, the average numbers of preneoplastic or neoplastic lesions per unit length of basement membrane in rats given 100–1000 mg/kg/day ETBE were also comparable to control values. However, papillomatosis of the urinary bladder was found in 4 out of 30 rats (13%) in the group given 1000 mg/kg/day ETBE, and soft stones in the urinary bladder were found in 3 out of these 4 rats. The results thus demonstrated that ETBE did not exert promotional activity on urinary bladder carcinogenesis. However, papillomatosis of the urinary bladder developed in small numbers of the rats given ETBE at 1000 mg/kg/day but not in rats given 500 mg/kg/day or lower doses.     

Development of a screening method for genetically modified soybean by plasmid-based quantitative competitive polymerase chain reaction

A novel type of quantitative competitive polymerase chain reaction (QC-PCR) system for the detection and quantification of the Roundup Ready soybean (RRS) was developed. This system was designed based on the advantage of a fully validated real-time PCR method used for the quantification of RRS in Japan. A plasmid was constructed as a competitor plasmid for the detection and quantification of genetically modified soy, RRS. The plasmid contained the construct-specific sequence of RRS and the taxon-specific sequence of lectin1 (Le1), and both had 21 bp oligonucleotide insertion in the sequences. The plasmid DNA was used as a reference molecule instead of ground seeds, which enabled us to precisely and stably adjust the copy number of targets. The present study demonstrated that the novel plasmid-based QC-PCR method could be a simple and feasible alternative to the real-time PCR method used for the quantification of genetically modified organism contents.     

Improvement of DNA extraction method for dried blood spots and comparison of four PCR methods for detection of Babesia gibsoni (Asian genotype) infection in canine blood samples

To eradicate canine babesiosis in epidemic areas, mass-screening of the infection situation of Babesia gibsoni including occult infection is necessary. The development of cost-effective method for storage and transport of blood samples is required. A highly efficient DNA extraction procedure from dried blood spots (DBS) onto Whatman 3MM filter paper was developed for the diagnosis of B. gibsoni infection in dog by PCR. In 3 extraction methods, Chelex-based method in combination with saponin washing and phenol-chloroform-isoamyl alcohol extraction (Saponin-PCI method) provided the best results. Sensitivity of the 4 previously described PCR methods for detection of B. gibsoni infection was also compared using serially diluted blood samples of B. gibsoni-infected dogs. The PCR method using Gib599F/Gib1270R primer pair provided the best performance. To evaluate the stability of DNA in DBS, DBS of B. gibsoni-infected dogs stored at room temperature for 2 months. The stability was superior to whole blood samples stored at -20 degrees C for 2 months. This highly efficient DNA extraction method on DBS using Whatman 3MM filter paper has potential to be cost-effective and high performance tool for storage, and molecular diagnosis of clinical blood sample from dog. This procedure in combination with the PCR method using Gib599F/Gib1270R primer pair may greatly assist in diagnosis of B. gibsoni infection in dog populations that are geographically distant.