Abstract: The effect of His-heme misligation on folding has been investigated for a triple mutant of yeast iso-2 cytochrome c (N26H,H33N,H39K iso-2). The variant contains a single misligating His residue at position 26, a location at which His residues are found in several cytochrome c homologues, including horse, tuna, and yeast iso-1. The amplitude for fast phase folding exhibits a strong initial pH dependence. For GdnHCl unfolded protein at an initial pH<5, the observed refolding at final pH 6 is dominated by a fast phase (tau(2f)=20 ms, alpha(2f)=90 %) that represents folding in the absence of misligation. For unfolded protein at initial pH 6, folding at final pH 6 occurs in a fast phase of reduced amplitude (alpha(2f) approximately 20 %) but the same rate (tau(2f)=20 ms), and in two slower phases (tau(m)=6-8 seconds, alpha(m) approximately 45 %; and tau(1b)=16-20 seconds, alpha(1b) approximately 35 %). Double jump experiments show that the initial pH dependence of the folding amplitudes results from a slow pH-dependent equilibrium between fast and slow folding species present in the unfolded protein. The slow equilibrium arises from coupling of the His protonation equilibrium to His-heme misligation and proline isomerization. Specifically, Pro25 is predominantly in trans in the unligated low-pH unfolded protein, but is constrained in a non-native cis isomerization state by His26-heme misligation near neutral pH. Refolding from the misligated unfolded form proceeds slowly due to the large energetic barrier required for proline isomerization and displacement of the misligated His26-heme ligand.

Abstract: REASONS FOR PERFORMING STUDY: There has been no large study of horses with suspected sacroiliac (SI) joint region pain in which the clinical diagnosis has been supported by either abnormal radiopharmaceutical activity in the SI joint region or by periarticular infiltration of local anaesthetic solution. OBJECTIVES: To describe the clinical features of horses with SI joint region pain, to document the age, breed, sex, discipline, size and conformation of affected horses and to compare these with the author's (SD) normal case population and to document the results of infiltration of local anaesthetic solution around the SI joint region. METHODS: Horses were selected for inclusion in the study based upon the exclusion of other causes of lameness or poor performance, together with clinical signs suggestive of SI joint pain and abnormal radiopharmaceutical activity in the SI joint region and/or a positive response to periarticular infiltration of local anaesthetic solution. RESULTS: Sacroiliac joint region disease was identified in 74 horses between November 1997 and March 2002. Dressage and showjumping horses appeared to be at particular risk (P < 0.001). Affected horses were generally slightly older than the normal clinic population (P < 0.0001), taller at the withers (P < 0.0001) and of greater bodyweight (P < 0.01). There was a significant effect of breed (P < 0.001), with a substantially higher proportion of Warmblood horses (51%) in the SI pain group compared to the normal clinic population (29%). There was no correlation between conformation and the presence of SI joint region pain. The tubera sacrale appeared grossly symmetrical in most (95%) horses. Poor development of the epaxial muscles in the thoracolumbar region and asymmetry of the hindquarter musculature were common. Twenty-six horses (35%) showed restricted flexibility of the thoracolumbar region and 10 (16%) had an exaggerated response to pressure applied over the tubera sacrale. Fourteen horses (19%) were reluctant to stand on one hindlimb for prolonged periods. The majority of horses (75%) had a straight hindlimb flight and only 18% moved closely behind or plaited. In all horses restricted hindlimb impulsion was the predominant feature; invariably this was most obvious when the horse was ridden. Stiffness, unwillingness to work on the bit and poor quality canter were common. Sacroiliac joint region pain was seen alone (47%), or in conjunction with thoracolumbar pain (16%), hindlimb lameness (20%), forelimb lameness (7%) or a combination of problems (10%). Seventy-three horses (99%) had abnormalities of the SI joint region identified using nuclear scintigraphy. Infiltration of local anaesthetic solution around the SI joint region produced profound improvement in gait in all 34 horses in which it was performed. CONCLUSIONS AND POTENTIAL RELEVANCE: Careful clinical examination combined with scintigraphic evaluation of the SI joint region and local analgesia can enable a more definitive diagnosis of SI joint region pain than has previously been possible.

Abstract: The landmark discrimination learning test can be used to assess the ability to utilize allocentric spatial information to locate targets. The present experiments examined the role of various factors on performance of a landmark discrimination learning task in beagle dogs. Experiments 1 and 2 looked at the effects of age and food composition. Experiments 3 and 4 were aimed at characterizing the cognitive strategies used in performance on this task and in long-term retention. Cognitively equivalent groups of old and young dogs were placed into either a test group maintained on food enriched with a broad-spectrum of antioxidants and mitochondrial cofactors, or a control group maintained on a complete and balanced food formulated for adult dogs. Following a wash-in period, the dogs were tested on a series of problems, in which reward was obtained when the animal responded selectively to the object closest to a thin wooden block, which served as a landmark. In Experiment 1, dogs were first trained to respond to a landmark placed directly on top of coaster, landmark 0 (L0). In the next phase of testing, the landmark was moved at successively greater distances (1, 4 or 10 cm) away from the reward object. Learning varied as a function of age group, food group, and task. The young dogs learned all of the tasks more quickly than the old dogs. The aged dogs on the enriched food learned L0 significantly more rapidly than aged dogs on control food. A higher proportion of dogs on the enriched food learned the task, when the distance was increased to 1cm. Experiment 2 showed that accuracy decreased with increased distance between the reward object and landmark, and this effect was greater in old animals. Experiment 3 showed stability of performance, despite using a novel landmark, and new locations, indicating that dogs learned the landmark concept. Experiment 4 found age impaired long-term retention of the landmark task. These results indicate that allocentric spatial learning is impaired in an age-dependent manner in dogs, and that age also affects performance when the distance between the landmark and target is increased. In addition, these results both support a role of oxidative damage in the development of age-associated cognitive dysfunction and indicate that short-term administration of a food enriched with supplemental antioxidants and mitochondrial cofactors can partially reverse the deleterious effects of aging on cognition.

Abstract: Administration of lithium chloride disrupted the intake of flavored solutions but not water in rats. This intake suppression was directly related to the amount of lithium administered (Experiment 1), occurred with both palatable and unpalatable novel saccharin solutions (Experiment 2), but was only observed if subjects were tested starting less than 75 min. after lithium treatment (Experiment 3). Twenty-five daily exposures to saccharin did not attenuate the effect (Experiment 4). However, in saccharin-reared and vinegar-reared rats, lithium did not disrupt consumption of the solutions these subjects had access to throughout life, even though suppressions of intake were observed when these subjects were tested with novel flavors (Experiment 5). The selective disruption of drinking is interpreted as a novelty-dependent sensitization reaction to the discomfort of aversive drug administration.