Reversal of ketamine/xylazine anesthesia in the rabbit with yohimbine

Ketamine and xylazine used in combination have been shown to be effective, easily administered, cost efficient agents for surgical anesthesia in the rabbit. The effect of xylazine on the central nervous system has been shown to be mediated through alpha-2 adrenergic receptors. Yohimbine, an alpha-2 adrenergic antagonist has been shown to reverse xylazine induced depression and partially antagonize ketamine in other species. We evaluated the antagonistic effect of yohimbine on ketamine/xylazine anesthesia in the rabbit. Six New Zealand White rabbits were anesthetized with intramuscular ketamine (50 mg/kg) and xylazine (10 mg/kg) to establish baseline parameters including respiratory rate, heart rate, and palpebral, pedal and postural reflex activity. Fourteen days later each rabbit was subjected to the same anesthetic regimen followed 30 minutes later by the intravenous administration of yohimbine (0.2 mg/kg). The duration of anesthesia estimated by the time elapsed between the loss and return of the palpebral reflex was reduced in the yohimbine treated trial (means = 29.7 +/- 1.9 minutes) compared to the control trial (means = 67.0 +/- 13.5 minutes). The palpebral reflex returned within 5 minutes following yohimbine treatment. Our results indicated that yohimbine is an effective antagonist of ketamine/xylazine anesthesia in the rabbit. Yohimbine decreases anesthetic duration after intravenous administration and also may aid in the control of undesirable anesthetic effects and overdosage.     

Amino acid sequence homologies between rabbit, rat, and human serum retinol-binding proteins

The main transporting protein for vitamin A in rabbit serum, the retinol-binding protein (RBP), was isolated and its amino acid sequence determined. Rabbit RBP was found to be highly homologous to human RBP, whose amino acid sequence was elucidated earlier, and to rat RBP. The rat RBP sequence was obtained by combining information deduced from the nucleotide sequences of two overlapping cDNA clones with the NH2-terminal sequence of the isolated protein determined by automated Edman degradation. The identity between the three proteins is approximately 90%. The high degree of homology between RBP molecules from different species is probably explained by the fact that RBP participates in at least three types of molecular interactions: in the binding of prealbumin, in the interaction with retinol, and in the recognition of a specific cell surface receptor. All these interactions should lead to a conservation of RBP structure. The amino acid differences between rabbit, rat, and human RBP are discussed in light of the recent elucidation of the three-dimensional structure of human RBP. Hybridization of a probe isolated from a rat RBP cDNA clone to restriction enzyme-digested genomic DNA from rat and mouse suggests that RBP is encoded by a single gene.     

Concerning the structure of 7α,15β-dichloro-5α-cholest-8(14)-en-3β-ol,a novel inhibitor of cholesterol biosynthesis

Treatment of 3β-p-bromobenzoyloxy-14α, 15α-epoxy-5α-cholest-7-ene with gaseous HCI in chloroform at ?25°C gave 3β-p-bromobenzoyloxy-7α, 15β-dichloro-5α-cholest-8(14)-ene in 93% yield. The structure of the latter compound was unequivocally established by the results of X-ray crystallographic analysis.     

Serum haptoglobins in experimental coccidioidomycosis — A preliminary report

Summary The level of haptoglobin was determined in control rats and in rats infected withC. immitis. The haptoglobin levels in the infected group were significantly higher than in those in the control group. The possibility that serial determinations may be of value in following the course of this disease is currently being investigated.This study was supported in part by USPHS Grants A1-06048-01, 5 T1 A1 52–06 and the Dermatologic Research Foundation of California, Inc.     

Effect of pH and salt bridges on structural assembly: molecular structures of the monomer and intertwined dimer of the Eps8 SH3 domain

The SH3 domain of Eps8 was previously found to form an intertwined, domain-swapped dimer. We report here a monomeric structure of the EPS8 SH3 domain obtained from crystals grown at low pH, as well as an improved domain-swapped dimer structure at 1.8 A resolution. In the domain-swapped dimer the asymmetric unit contains two "hybrid-monomers." In the low pH form there are two independently folded SH3 molecules per asymmetric unit. The formation of intermolecular salt bridges is thought to be the reason for the formation of the dimer. On the basis of the monomer SH3 structure, it is argued that Eps8 SH3 should, in principle, bind to peptides containing a PxxP motif. Recently it was reported that Eps8 SH3 binds to a peptide with a PxxDY motif. Because the "SH3 fold" is conserved, alternate binding sites may be possible for the PxxDY motif to bind. The strand exchange or domain swap occurs at the n-src loops because the n-src loops are flexible. The thermal b-factors also indicate the flexible nature of n-src loops and a possible handle for domain swap initiation. Despite the loop swapping, the typical SH3 fold in both forms is conserved structurally. The interface of the acidic form of SH3 is stabilized by a tetragonal network of water molecules above hydrophobic residues. The intertwined dimer interface is stabilized by hydrophobic and aromatic stacking interactions in the core and by hydrophilic interactions on the surface.     

Protein folding and three-dimensional domain swapping: a strained relationship?

Many proteins function as multimeric assemblies into which the folded individual promoters organize as higher order structures. An oligomerization mechanism that appears to impose the coordination of events during folding and oligomer assembly is three-dimensional domain swapping. Recent studies have focused on revealing the structural basis of domain swapping and a possible role for domain swapping in the regulation of protein aggregation and activity.     

Endothelium-dependent and -independent relaxation in the forelimb and hindlimb vasculatures of swine

Limb differences in endothelial function exist between arm and leg vasculatures of humans. The current investigation tested the hypothesis that forelimb and hindlimb vasorelaxation are similar in the absence of limb differences in blood pressure. Conduit arteries (brachials/femorals) and second order arterioles were harvested from 22 miniature Yucatan swine. In vitro assessment of vasorelaxation was determined by administering increasing doses of bradykinin (BK), acetylcholine (ACh), and sodium nitroprusside (SNP). The role of the nitric oxide synthase (NOS) and cyclooxygenase (COX) pathways was assessed in conduit arteries but not resistance arterioles through L-NAME (300 microM) and INDO (5 microM) incubation, respectively. The relaxation responses to BK and ACh were similar in brachial and femoral arteries. SNP relaxation response was greater in the brachial compared to femoral arteries. There were also no significant differences in the relaxation responses of second order arterioles of the forelimb and hindlimb to BK, ACh, and SNP. Incubation of conduit arterial rings in L-NAME produced a greater reduction in BK and ACh relaxation in the brachial (approximately 25%) compared to femoral (approximately 13%) arterial rings. The current results of this investigation suggest that the forelimb and hindlimb vasculatures of swine have relatively similar vasorelaxation responses to both endothelium-dependent and -independent vasodilators.     

Heterogeneous vasodilator responses of human limbs: influence of age and habitual endurance training

Forearm endothelium-dependent vasodilation is impaired with age in sedentary, but not endurance-trained, men. The purpose of this investigation was to determine whether these age- and physical activity-related differences in endothelium-dependent vasodilation also occur in the leg. Brachial and common femoral arterial blood flow were measured with Doppler ultrasound during increasing doses of acetylcholine (1, 4, and 16 microg.100 ml limb tissue(-1).min(-1)), substance P (8, 31, and 125 pg.100 ml limb tissue(-1).min(-1)), and sodium nitroprusside (0.063, 0.25, and 1 microg.100 ml limb tissue(-1).min(-1)) in 23 healthy men (8 younger sedentary, 8 older sedentary, and 7 older endurance trained). Increases in forearm blood flow to the highest dose of acetylcholine and sodium nitroprusside were smaller (P < 0.05) in older sedentary (841 +/- 142%, 428 +/- 74%) compared with younger sedentary (1,519 +/- 256%, 925 +/- 163%) subjects. Similarly, increases in forearm blood flow to sodium nitroprusside (1 microg.100 ml limb tissue(-1).min(-1)) were smaller (P < 0.05) in older endurance-trained (505 +/- 110%) compared with younger sedentary (925 +/- 163%) subjects. In contrast, no differences in leg blood flow responses to intra-arterial infusions of acetylcholine, substance P, or sodium nitroprusside were noted between subject groups. These results demonstrate that 1) acetylcholine- and sodium nitroprusside-induced vasodilation are attenuated in the forearm vasculature and preserved in the leg vasculature of older sedentary subjects and 2) sodium nitroprusside-induced vasodilation remains attenuated in the forearm vasculature of healthy older endurance-trained men but preserved in the leg vasculature of these men.     

The Prp19 U-box crystal structure suggests a common dimeric architecture for a class of oligomeric E3 ubiquitin ligases

Prp19 is an essential splicing factor and a member of the U-box family of E3 ubiquitin ligases. Prp19 forms a tetramer via a central coiled-coil domain. Here, we show the U-box domain of Prp19 exists as a dimer within the context of the Prp19 tetramer. A high-resolution structure of the homodimeric state of the Prp19 U-box was determined by X-ray crystallography. Mutation of the U-box dimer interface abrogates U-box dimer formation and is lethal in vivo. The structure of the U-box dimer enables construction of a complete model of Prp19 providing insights into how the tetrameric protein functions as an E3 ligase. Finally, comparison of the Prp19 U-box homodimer with the heterodimeric complex of BRCA1/BARD1 RING-finger domains uncovers a common architecture for a family of oligomeric U-box and RING-finger E3 ubiquitin ligases, which has mechanistic implications for E3 ligase-mediated polyubiquitination and E4 polyubiquitin ligases.     

Intermittent pneumatic leg compressions enhance muscle performance and blood flow in a model of peripheral arterial insufficiency

Despite the escalating prevalence in the aging population, few therapeutic options exist to treat patients with peripheral arterial disease. Application of intermittent pneumatic leg compressions (IPC) is regarded as a promising noninvasive approach to treat this condition, but the clinical efficacy, as well the mechanistic basis of action of this therapy, remain poorly defined. We tested the hypothesis that 2 wk of daily application of IPC enhances exercise tolerance by improving blood flow and promoting angiogenesis in skeletal muscle in a model of peripheral arterial insufficiency. Male Sprague-Dawley rats were subjected to bilateral ligation of the femoral artery and randomly allocated to treatment or sham groups. Animals were anesthetized daily and exposed to 1-h sessions of bilateral IPC or sham treatment for 14-16 consecutive days. A third group of nonligated rats was also studied. Marked increases in treadmill exercise tolerance (～33%, P < 0.05) and improved muscle performance in situ (～10%, P < 0.05) were observed in IPC-treated animals. Compared with sham-treated controls, blood flow measured with isotope-labeled microspheres during in situ contractions tended to be higher in IPC-treated animals in muscles composed of predominantly fast-twitch white fibers, such as the plantaris (～93%, P = 0.02). Capillary contacts per fiber and citrate synthase activity were not significantly altered by IPC treatment. Collectively, these data indicate that IPC improves exercise tolerance in a model of peripheral arterial insufficiency in part by enhancing blood flow to collateral-dependent tissues.