The activity of hippocampal pyramidal cells (place cells) is correlated with the position of a rat in a given environment, suggesting that the rat hippocampus may contain a cognitive map of the environment. Internally-generated information about the motion of the rat may be the strongest initial determinant of place-cell firing. With experience in a given environment, cues and landmarks may gain control over place-cell firing, allowing the correction of errors or drift in the self-motion information. This 'binding' of cues to the hippocampal map may involve long-term potentiation (LTP). Because hippocampal LTP is impaired in aged rats, cues and landmarks should be poorly bound to the hippocampal map in aged rats. To test this prediction, aged and adult rats were trained to run back and forth on a linear track. The position of the start box was changed from trial to trial, so that self-motion information (i.e., the distance of the rat from the start box) and visual-cue information (the position of the rat in the room) were mismatched. In this situation, self-motion information controls place-cell firing during the initial portion of each journey. At some point in each journey, however, the hippocampal map realigns, so that place-cell firing is controlled by visual-cue information. If age-related LTP deficits impair the binding of cues to the hippocampal map, this realignment should be delayed in aged rats. As predicted, the realignment occurred nearer to the end of the track in the aged rats. Moreover, the delayed realignment was correlated with the rats' ability to find a hidden goal at a position relative to the visual cues. The results suggest that some of the spatial-learning deficits observed in aged rats may be due to impaired binding of cues to the hippocampal map. In addition, despite the fact that the animals in the present study had never been trained with a stable start box (the anchor point for self-motion information), place-cell firing was controlled by self-motion information for a large portion of each outbound journey. This result supports a strong initial role of self-motion information in determining the hippocampal representation of a rat's position.

<p> Intercalated discs (ICDs) are cardiac-specific structures located at the longitudinal termini of cardiomyocytes. Classically, the functions assigned to ICDs include mechanical and electrical communications among adjacent cardiomyocytes. More recently, it has been increasingly realized that ICDs also function in signal transduction and regulation of the surface expression of ion channels. Accordingly, defects of ICD components are shown to cause a number of human cardiac diseases and changes of ICDs are associated with cardiomyopathy, arrhythmias, and heart failure. The expansion of our knowledge about the development, function and maintenance of ICDs are promoted by identification, cataloging and characterization of the molecular components of the ICDs. In this thesis, I characterize a family of Xin repeat-containing proteins, which are striated muscle-specific and localized to the ICDs in the cardiomyocytes. This thesis provides novel insights into the mechanism of the formation, maintenance and functions of ICDs. </p><p> Our previous studies showed that the Xin repeat-containing proteins play critical role in cardiac morphogenesis and cardiac function. Knocking down the <i>Xin</i> in chicken embryo collapses the wall of developing heart chambers and leads to abnormal cardiac morphogenesis. In mammals, a pair of paralogous genes, <i>Xin&alpha;</i> and <i>Xin&beta; </i>, exists. Ablation of the mouse <i>Xin&alpha;</i> (<i> mXin&alpha;</i>) does not affect heart development. Instead, the mXin&alpha;-deficient mice show adult late-onset cardiac hypertrophy and cardiomyopathy with conduction defects. The ICD structural defects in mXin&alpha;-null mice occur between 1 and 3 months of age and progressively worsen with aging. The mXin&alpha;-deficient hearts up-regulate mXin&beta;, suggesting a partial compensatory role of mXin&beta;. </p><p> In this thesis, I focus on two questions. First, what are the molecular mechanisms of mXin&alpha;'s functions that account for the observed phenotypes in the mXin&alpha;-deficient hearts? And second, what are the functions of mXin&beta;? Through biochemical methods and electron microscopy, I demonstrated that mXin&alpha; binds and bundles actin filaments. In addition, a direct interaction between mXin&alpha; and the adherens junction protein &beta;-catenin facilitates mXin&alpha;'s interaction with the actin filaments. Based on this in vitro characterization of mXin&alpha;, we proposed that mXin&alpha; may act as a direct link between the adherens junctions and actin cytoskeleton, thus providing an important means to strengthening the intercellular adhesion at the ICDs. To characterize mXin&beta;'s roles, I generated and characterized <i> mXin&beta;</i>-knockout mice. I showed that complete loss of mXin&beta; leads to cardiac morphological defects, diastolic dysfunction and heart failure, which lead to severe growth retardation and early postnatal lethality. I also showed that mXin&beta; might be involved in a number of cell signaling pathways and provide multiple lines of evidence to support mXin&beta;'s roles in the formation of ICDs. </p><p> In summary, this thesis provides novel insights into the specialization of the adherens junctions at the ICDs to withstand the contractile forces, and the molecular mechanisms for the establishment, maintenance and function of ICDs. The knowledge gained from the roles of Xin proteins in cardiac development and function will likely provide new insights for improved therapeutic strategies for human cardiomyopathy, arrhythmias and heart failure.</p>

<p> The child care setting represents a crucial environment for infants and children to establish healthy feeding practices in order to prevent overweight and obesity. The objective of this research was to investigate the association between parental care (PC) and child care (CC) on infant feeding practices, food consumption, nutrient intake and growth in infants receiving Special Supplemental Nutrition Program for Women Infants and Children (WIC) assistance. Our hypothesis was that unhealthy feeding practices would be more common in CC compared to PC thus leading to greater weight for length (WFL) and weight for age (WFA) z-score at 1 year of age for infants in CC. This study sampled 105 infants aged 2 to 8 months of age from the Champaign Urbana WIC office from October 2009-August 2011. Mothers completed a 3-day food record and survey at the time of recruitment to assess their infant's feeding practices, nutrient intake, health status, and demographic characteristics. Baseline and follow-up weight and length for these children within the first year of age were collected from the WIC office.</p><p> The major differences in demographic characteristics of the study sample included child care hours per week, maternal employment, household income, and single parent home by CC use. Infants in CC had an average of 29 hours of care per week compared to the 0.64 hours in the PC group (p&lt;0.01). A larger (p&lt;0.01) percentage of mothers were employed in the CC group (73.9%) compared to the PC group (22%). However, the household income was greater (p&lt;0.01) in the PC group ($15,986 &plusmn; $10,284 PC vs $9,967&plusmn; $7,489.5 CC). In addition, there was a higher (p=0.04) percentage of single parents in the CC group (30.5 % PC vs. 50% CC).</p><p> Breastfeeding duration and age of solid food introduction did not differ between care type. Breastfeeding duration was on average 2.3 months while average solid food introduction was 4.4 months. No differences were observed between PC and CC infants in the rates of formula introduction. When comparing food consumption at the time of recruitment, there were no differences in the number of servings per day of food groups, but the CC group showed lower consumption of formula (p=0.03) and breast milk (p=0.18) compared to PC.</p><p> Energy intake did not differ between care type after adjusting for feeding practices and child, maternal and household characteristics. However, there was a pattern of greater energy intake in the PC group. Child age (&beta;=34.8, p&lt;0.01) and number of servings of infant formula (&beta;=86.0, p&lt;0.01) were the strongest predictors of energy intake. There was greater (p=0.05) calcium intake in the CC group (788 mg CC vs. 742 mg PC). Otherwise, there were no differences in macro or micronutrient intakes between CC and PC.</p><p> For growth measures, infants in PC had a significantly greater change in WFL (&beta;=2.06, p=0.05) and WFA (&beta;=1.69, p=0.01) z-score and a greater follow-up z-score, after adjusting for feeding practices and child, maternal and household characteristics. There were no differences by care type in the length for age (LFA) z-score over the first year of life. </p><p> The strongest predictors of the change in WFL z-score were PC use (&beta;=2.06, p=0.05), maternal pre-pregnancy BMI (&beta;=0.14, p&lt;0.01), birth order (&beta;=1.63, p&lt;0.05), maternal age (&beta;=-0.34, p&lt;0.01), birth weight (&beta;=-1.77, p=0.06), non-Black/African American (&beta;=3.09, p=0.02) and male gender (&beta;=-2.12, p=0.06). Change in WFA z-score was significantly affected by CC use (&beta;=1.69, p=0.01), lower birth weight (&beta;=-1.74, p&lt;0.01), greater pre-pregnancy BMI (&beta;=0.09, p&lt;0.01), and less servings of infant formula (&beta;=-0.53, p=0.05). Change in LFA was unaffected by CC use (&beta;=1.69, p=0.11), but significantly affected by lower pre-pregnancy BMI (&beta;=-0.04, p=0.04) and black race (&beta;=-2.54, p=0.05).</p><p> Thus, we concluded that CC use did not affect feeding practices, overall nutrient intake or LFA z-scores for infants receiving WIC assistance. There was significantly greater calcium intake in the CC group. CC use also showed a trend of less formula and breast milk. Infants in PC had a statistically greater change in WFL and WFA compared to those in CC. The main finding in this study is that CC use may have influenced differences in the change in WFL and WFA z-scores, but not overall infant feeding practices, nutrient intake and LFA z-score. Future longitudinal studies are warranted to explore the role of CC use on feeding practices, nutrient intake and growth.</p>

RET is a tyrosine kinase receptor that induces cell survival, proliferation and migration. When bound by its ligand, GDNF (glial derived neurotrophic factor) and its co-receptor, GFRalpha-1 (GDNF family receptor alpha-1), tyrosine residues in the intracellular domain of the receptor are phosphorylated as part of signal transduction. In the kidney, RET is required for branching of the ureteric bud and it also regulates the maturation of the ureter in the urinary tract. Although it is well established in mice that the absence or overexpression of RET leads to severe kidney abnormalities, the cellular basis for these defects remains unexplored. To determine whether the overexpression of RET affected the development of the urinary tract, HoxB7/RET mice were tested for the presence of vesico-ureteric reflux (VUR) in which urine flows retrogradely from the bladder back up the ureter. To further understand how the urinary tract is formed, we looked for the presence of VUR in HoxB7/RET and wildtype mice at E17. These results provide a new understanding of how combined kidney and urinary tract malformations occur in humans.

The chondroitin sulphate epitope 846 of aggrecan is abundant in foetal cartilage, barely detectable in normal adult cartilage, but reappears in the cartilage and body fluids of arthritic patients. This epitope has been proposed to be a marker of aggrecan synthesis during new cartilage formation and repair. The purpose of the present studies was to investigate, in vitro, whether the epitope was truly reflective of aggrecan synthesis, and to understand its role in the cartilage repair process. Foetal bovine chondrocyte cultures were established to study aggrecan synthesis in order to investigate whether the epitope 846 was present on these molecules. These studies showed that the epitope was indeed present on the newly synthesised aggrecan molecules and that these were preferentially retained within the extracellular matrix. The larger size and higher epitope density of the matrix molecules, compared to those molecules which were released into culture medium, suggested a role for the 846-epitope bearing molecules in the formation of new cartilage and during repair. Normal adult articular cartilage cultures were established to investigate whether the epitope could be synthesised by this tissue under conditions where the tissue had been stimulated to repair a damaged matrix, by trypsin-treatment of the cartilage. In these studies, an increase in abundance of the epitope on newly synthesised proteoglycans was observed, further indicating a role for the epitope-bearing molecules in cartilage repair. Explant cultures of osteoarthritic cartilage demonstrated that the release of the 846-bearing molecules from the cartilage was accompanied by the release of some of the newly synthesised 35S-sulphate labelled proteoglycans from the cartilage, and that these molecules contained, in part at least, the epitope. In addition, the release of 846-bearing molecules from the cartilage into culture medium or synovial fluid was reflective of the epitope content of the tissue. The stru

The pharmacokinetics of verapamil were evaluated in the young and elderly after steady state administration of a sustained release formulation. The objective was to determine whether the interindividual metabolic variability of verapamil and its main metabolites could be correlated with the enzymatic activity of CYP3A4 by using dextromethorphan (DM) as an in vivo probe of CYP3A4 activity. / Pharmacokinetic analysis showed a significant age related increase in the clearance of verapamil. Verapamil interindividual variability could not be correlated with formation of 3-methoxy-morphinan. Our results suggest that verapamil metabolism may be significantly mediated by other cytochromes.

Treatment of chick myoblasts with the glucosidase inhibitors bromoconduritol (BCD) or N-methyl-1-deoxynojirimycin (MDJN), but not the mannosidase I inhibitor 1-deoxymannojirimycin (ManDJN), decreased the rate of cell-adhesion to fibronectin and laminin and increased the rate of adhesion to collagen. The adhesion of chick myoblasts is predominantly mediated by integrin(s), as judged by inhibition of adhesion by an anti-integrin antibody (JG22). The effects of BCD, MDJN and ManDJN on myoblast integrin detectable at the myoblast cell surface with JG22 antibody correlated well with their effects on adhesion to fibronectin and laminin, and paralleled the previously reported effects of these agents on myogenesis. Interaction of integrin with the extracellular matrix appears critical for terminal differentiation. Mn$ sp{2+}$ ions increased myoblast adhesion to ECM proteins, and antagonized the effect of BCD and MDJN on myoblast differentiation. / Finally BCD and MDJN were found to act at an early stage in myoblast differentiation, down regulating the expression of both myogenin and muscle $ alpha$-actin mRNA in L$ sb6$ myoblasts.

Idiopathic central precocious puberty may compromise adult height. Gonadotrophin-releasing hormone analogues (GnRHa) suppress puberty and increase final height prediction, but their influence on final height is unclear, especially in girls with pubertal onset just below eight years of age, the traditional limit of normal. / At seven children's hospitals, we identified 53 treated and 24 untreated patients (24 and 7 to final height, respectively) whose pubertal onset was between ages 5--8 years. At baseline, bone age advancement and predicted adult height were similar in the two groups. In both groups, the predicted adult height slightly overestimated the final height. What role, if any, GnRHa therapy played in preventing a shorter adult height is uncertain in these borderline cases. The substantial intra- and inter-observer variability in bone age readings compromised the utility of the height prediction method. / The methodological challenges inherent to this study are identified and discussed.

The intervertebral disc has been identified as a possible contributor to two very common back disorders: degenerative disc disease and scoliotic deformity. Little is known about the composition and turnover of extracellular matrix in the human intervertebral disc and end-plate with these two pathologies. / We studied 121 intervertebral discs representing all age groups and degeneration grades. In addition, fifteen scoliotic discs and seventeen control discs were analyzed. / Synthesis in intervertebral discs, end-plates, and scoliotic tissues were measured by immunoassay measuring the content of a putative aggrecan biosynthesis marker (846) and contents of types I and II procollagen markers (CPI & CPII). The percentage of denatured type II collagen was assessed by the presence of an epitope that is exposed with cleavage of type II collagen. / We identified three matrix turnover phases in the intervertebral disc and end-plate. Phase I (growth) is characterized by active synthesis of matrix molecules and active denaturation of type II collagen. Phase II (maturation and ageing phase) is distinguished by progressive drops in synthetic activity and denaturation of type II collagen. Phase III (degeneration and fibrotic phase) is illustrated by decreased synthesis of aggrecan and type II procollagen and an increase in collagen type II denaturation and type I procollagen synthesis, the later dependent on age and grade of tissue degeneration. / The results of the scoliotic study indicate that synthesis of type II procollagen in the end-plate and annulus was higher in scoliotic discs. Aggrecan synthesis showed a similar trend in the nucleus pulposus of scoliotic disc. By contrast, the contents of hydroxyproline, glycosaminoglycan, and water were significantly lower in scoliotic discs while total protein content was higher in scoliotic discs, indicating that non-collagenous protein content was increased in scoliotic discs. This elevated synthetic activity without an associated collagen denaturation reflects an overall synthetic response without an increased matrix turnover, suggesting that scoliotic changes are due to an altered and ineffective synthetic response to a pathologic mechanical environment. / This thesis represents the first in situ quantitation of human lumbar intervertebral disc matrix composition and turnover with ageing, degeneration, and scoliotic change. These observations are an important advancement in our biochemical understanding of disc ageing, disc degeneration, and adolescent idiopathic scoliosis.

The replacement of the hyaline cartilage model by bone tissue during osseous development involves an extensive resorption of cartilage. The current study examines the role played by matrix metalloproteinases (MMPs) in cartilage resorption in the tibial epiphysis of 8 and 10 day old rats. The entry of cellular, highly vascularized channels requires the degradation of cartilage components. As a first step, aggrecan is cleaved by an unidentified MMP along the edges of the channel walls, as detected by an antiserum directed to a cleavage fragment retained in the tissue; the presence of an active MMP at the channel edge is confirmed by the binding of a recombinant MMP inhibitor, TIMP-2. Second, using histozymography, the collagen component of cartilage is degraded by another MMP, identified as gelatinase B by inhibition studies, at similar channel sites. Thus, two proteinases combine their efforts to produce the resorption associated with channel invasion preceding the appearance of the secondary ossification center in the rat tibial epiphysis.