Evaluation of the use of midazolam as a co-induction agent with ketamine for anaesthesia in sedated ponies undergoing field castration.

BACKGROUND: There are limited investigations comparing ketamine to a ketamine-midazolam co-induction. OBJECTIVES: To compare quality and safety of general anaesthesia induced using ketamine alone with anaesthesia co-induced using ketamine and midazolam. STUDY DESIGN: Randomised, double blinded, placebo controlled trial. METHODS: After i.v. detomidine (20 µg/kg) thirty-eight ponies undergoing field castration received either 0.06 mg/kg (0.6 mL/50 kg) midazolam (group M) or 0.6 mL/50 kg placebo (group P) with 2.2 mg/kg ketamine i.v. for anaesthetic induction. Quality of anaesthetic induction, endotracheal intubation, surgical relaxation and recovery were scored using combinations of simple descriptive and visual analogue scales. Time of sedation, induction, start of endotracheal intubation, first movement, sternal recumbency and standing were recorded, as were time, number and total quantity of additional i.v. detomidine and ketamine injections. Cardiorespiratory variables were assessed every 5 min. Adverse effects were documented. Data were tested for normality and analysed with a mixed model ANOVA, Fisher's exact test, unpaired Students' t test and Wilcoxon Rank-sum as appropriate; P<0.05 was considered significant. RESULTS: Group M had better scores for induction (P = 0.005), intubation (P<0.001) and surgical relaxation (P<0.001) and required fewer additional injections of detomidine and ketamine (P = 0.04). Time (minutes) from induction to first movement (P<0.001), sternal recumbency (P =< 0.001) and standing was longer (P = 0.05) in group M. Recoveries were uneventful with no difference in quality between groups (P = 0.78). MAIN LIMITATIONS: Clinical study with noninvasive monitoring undertaken in field conditions. CONCLUSIONS: Ketamine-midazolam co-induction compared to ketamine alone improved quality of induction, ease of intubation and muscle relaxation without impacting recovery quality.

Effect of fixed value positive end expiratory pressure valves on canine thoracic volume and atelectasis.

OBJECTIVES: The objective of this study was to investigate whether a fixed 10 cm H2 O positive end-expiratory pressure valve would increase the aeration of, and reduce atelectasis formation in, the lungs after induction of anaesthesia in dogs undergoing thoracic CT. MATERIALS AND METHODS: 28 dogs were paired based on breed, bodyweight and body condition score and then randomly allocated to either Group Z (0 cm H2 O) or Group P (10 cm H2 O positive end-expiratory pressure valve) immediately after the induction of anaesthesia. All patients received a standardised anaesthetic protocol, and their lungs were manually hyperventilated before image acquisition. Cardiorespiratory parameters were recorded every 5 minutes. Total lung volume, lung density and degree of atelectasis were determined for each dog from the acquired images. RESULTS: The 10 cm H2 O positive end-expiratory pressure valve significantly increased lung volume (mL/kg) (Group Z: 52 ±14; Group P: 83 ±17; P<0·001) whilst significantly reducing lung density (Hounsfield units) (Group Z: -775 ±30; Group P: -856 ±22; P<0·001) and the amount of atelectasis (P=0·004). Dogs in Group P had significantly higher end-tidal carbon dioxide (P<0·05), but there was no difference between the groups for respiratory rate or any cardiovascular variable. CLINICAL SIGNIFICANCE: A fixed-value positive end-expiratory pressure valve provides a simple, cost-effective technique for improving expiratory thoracic CT studies by increasing lung volume and decreasing atelectasis formation.

Thoracoscopic foreign body removal and right middle lung lobectomy to treat pyothorax in a dog.

A three-year-old, 30-kg, spayed female German wirehaired pointer was presented for coughing, pyrexia and lethargy. Thoracic radiographs showed mild right-sided pleural effusion, moderate pneumothorax and a pulmonary lesion in the right middle or caudal lung lobe. A diagnosis of pyothorax was established by fine needle aspiration of the pleural effusion. Thoracoscopic exploration was performed using one-lung ventilation. A vegetal foreign body (grass awn) and an abscess were observed in the distal part of the right middle lung lobe. The foreign body was removed and a right middle lung lobectomy was performed, both thoracoscopically. No complications were noted. The dog was discharged 48 hours after surgery, and no recurrence of the clinical signs was observed during the follow-up time period (three years and three months). Thoracoscopy is a minimally invasive alternative to thoracotomy to explore and successfully treat some non-chronic pyothoraces in dogs, including lesions affecting the right middle lung lobe.

Roles of the C termini of alpha -, beta -, and gamma -subunits of epithelial Na+ channels (ENaC) in regulating ENaC and mediating its inhibition by cytosolic Na+.

The amiloride-sensitive epithelial Na(+) channels (ENaC) in the intralobular duct cells of mouse mandibular glands are inhibited by the ubiquitin-protein ligase, Nedd4, which is activated by increased intracellular Na(+). In this study we have used whole-cell patch clamp methods in mouse mandibular duct cells to investigate the role of the C termini of the alpha-, beta-, and gamma-subunits of ENaC in mediating this inhibition. We found that peptides corresponding to the C termini of the beta- and gamma-subunits, but not the alpha-subunit, inhibited the activity of the Na(+) channels. This mechanism did not involve Nedd4 and probably resulted from the exogenous C termini interfering competitively with the protein-protein interactions that keep the channels active. In the case of the C terminus of mouse beta-ENaC, the interacting motif included betaSer(631), betaAsp(632), and betaSer(633). In the C terminus of mouse gamma-ENaC, it included gammaSer(640). Once these motifs were deleted, we were able to use the C termini of beta- and gamma-ENaC to prevent Nedd4-mediated down-regulation of Na(+) channel activity. The C terminus of the alpha-subunit, on the contrary, did not prevent Nedd4-mediated inhibition of the Na(+) channels. We conclude that mouse Nedd4 interacts with the beta- and gamma-subunits of ENaC.

Identification of multiple proteins expressed in murine embryos as binding partners for the WW domains of the ubiquitin-protein ligase Nedd4.

Nedd4 is a member of a growing family of ubiquitin-protein ligases which consist of a lipid-binding domain, two to four WW domains and a C-terminal ubiquitin-protein ligase domain. The Nedd4 mRNA levels are developmentally regulated and Nedd4 protein is highly expressed in many mouse embryonic tissues. In this study we have used a far-Western screen to identify embryonic proteins that interact with the WW domains in mouse Nedd4. We report here identification of eight Nedd4 WW-domain-interacting proteins from mouse embryonic cDNA expression libraries. Two of the proteins are novel, while two have been identified previously as ligands for a WW domain. All of these proteins contain one or more PY motifs. In seven of the eight proteins, these PY motifs are necessary for their interaction with the WW domains of Nedd4. Using site-directed mutagenesis, and by using individual WW domains of Nedd4 as probes for far-Western analysis, we show that the three WW domains in Nedd4 interact with varying affinities with the PY motifs present in various Nedd4-binding proteins. These results provide evidence that Nedd4 can potentially interact with multiple proteins, possibly simultaneously, through its WW domains.

All three WW domains of murine Nedd4 are involved in the regulation of epithelial sodium channels by intracellular Na+.

The amiloride-sensitive epithelial sodium channel (ENaC) plays a critical role in fluid and electrolyte homeostasis and consists of alpha, beta, and gamma subunits. The carboxyl terminus of each ENaC subunit contains a PPxY motif which is necessary for interaction with the WW domains of the ubiquitin-protein ligase, Nedd4. Disruption of this interaction, as in Liddle's syndrome where mutations delete or alter the PY motif of either the beta or gamma subunits, results in increased ENaC activity. We have recently shown using the whole-cell patch clamp technique that Nedd4 mediates the ubiquitin-dependent down-regulation of Na+ channel activity in response to increased intracellular Na+. In this paper, we demonstrate that WW domains 2 and 3 bind alpha-, beta-, and gamma-ENaC with varying degrees of affinity, whereas WW domain 1 does not bind to any of the subunits. We further show using whole-cell patch clamp techniques that Nedd4-mediated down-regulation of ENaC in mouse mandibular duct cells involves binding of the WW domains of Nedd4 to three distinct sites. We propose that Nedd4-mediated down-regulation of Na+ channels involves the binding of WW domains 2 and 3 to the Na+ channel and of WW domain 1 to an unknown associated protein.

Transmembrane 4 superfamily protein CD151 (PETA-3) associates with beta 1 and alpha IIb beta 3 integrins in haemopoietic cell lines and modulates cell-cell adhesion.

CD151 (PETA-3/SFA-1) is a member of the transmembrane 4 superfamily (TM4SF) of cell-surface proteins and is expressed abundantly both on the cell surface and in intracellular membranes by the haemopoietic cell lines M07e, HEL and K562. In the presence of mild detergent (CHAPS), CD151 co-immunoprecipitated with integrin alpha 4 beta 1, alpha 5 beta 1, alpha 6 beta 1 and alpha IIb beta 3. The association of CD151 with alpha 4 beta 1 and alpha 5 beta 1 seemed to be constitutive, as it was not modified by treatment of M07e cells with cytokines that regulate integrin function by 'inside-out' signalling. CD151 also associated with other tetraspans in an apparently cell-type-specific fashion, as defined by its co-precipitation with CD9, CD63 and CD81 from M07e cells, but not from K562 cells, which express similar levels of these proteins. F(ab')2 fragments of monoclonal antibodies (mAbs) against CD151 caused homotypic adhesion of HEL and K562 cells that was dependent on energy and cytoskeletal integrity and was augmented in the presence of RGDS peptides. The adhesion was not blocked by function-inhibiting mAbs against beta 1 or beta 3 integrins, suggesting that cell-cell adhesion was not mediated by the binding of integrin to a cell-associated ligand. Furthermore, mAb CD151 did not affect adhesion of the cells to fibronectin, laminin, collagen or fibrinogen, which are ligands for alpha 4 beta 1, alpha 5 beta 1, alpha 6 beta 1 and alpha IIb beta 3 integrins. Taken together, these results indicate that the ligation of CD151 does not induce the up-regulation of integrin avidity, but might act as a component of integrin signalling complexes.

Light baffle attenuation measurements in the visible.

Experimental results, obtained using photometry, are obtained to assess the attenuating capability of three baffle geometries. The data obtained include the effect of sensor location in the baffle aperture as well as the type of coating on baffle surfaces. The baffles attenuate best at large viewing angles with surfaces coated with black paint.