This work has focused on understanding in vivo aspects of ASP metabolism particularly postprandially. I showed that ASP was produced directly from human adipose tissue, a process that increased postprandially and was correlated with increased plasma triglyceride clearance and increased fatty acid incorporation into adipose tissue "FIAT". The target of ASP is adipose tissue. Binding studies in freshly obtained adipose tissue demonstrated specific binding to high affinity receptor sites. ASP binding to subcutaneous tissue from obese females demonstrated the greatest binding and highest receptor affinity. In contrast omental tissue particularly from males showed the lowest specific binding and affinity. This suggests an important role for ASP in maintaining regional adipose tissue mass in females and males, hence providing possible explanations for the metabolic complications seen in abdominal obesity. / The strongest evidence for a physiological role of ASP on triglyceride clearance for ASP in vivo was obtained when exogenous intraperitonial hASP was administered to genetically obese mice. Normolipidemic ob /ob mice demonstrated accelerated postprandial TG clearance in the presence of ASP. The effect in the hyperlipidemic: db/db mice, however was markedly greater (2 to 8 times). These findings strongly support the hypothesis that ASP is an important factor in postprandial lipid metabolism and may be a significant factor in determining the pathophysiology of obesity and related dyslipidemias.

Under normal conditions the sinoatrial node serves as the pacemaker of the heart. However, under disease states other pacemaker sites can emerge which compete with the sinoatrial node or with each other. This thesis describes theoretical and experimental studies of pacemaker initiation and interaction. The first aspect of this thesis deals with an arrhythmia called modulated parasystole, which is generated by the interaction between the sinus pacemaker and an ectopic pacemaking focus. A mathematical model was developed to study the dynamics of modulated parasystole using discontinuous circle maps. The mathematical model displayed banded chaos, characterized by zero rotation interval width in the presence of a positive Lyapunov exponent. Banded chaos in the parasystole map produces rhythms characteristic of those found clinically. The second aspect of the thesis deals with spontaneous pacemaker activity and interaction using optical mapping techniques and mathematical models. A macroscopic imaging system was designed and constructed that records fluorescent signals from thin preparations over large areas (1 cm2 ) for long time periods (>30 minutes). Rotating waves (rotors) of cellular activity were observed by mapping calcium in nonconfluent cultures of embryonic chick heart cells. Unlike previous observations of rotors or spiral waves in other systems, the rotors did not persist but exhibited a repetitive pattern of spontaneous onset and offset leading to a bursting rhythm. Similar dynamics were observed in simple excitable media models that incorporated spontaneous initiation of activity and a decrease of excitability as a consequence of rapid activity (fatigue). These results provide a mechanism for bursting dynamics in normal and pathological processes. Activation maps were also obtained from the rabbit atrioventricular (AV) node, a small region of the heart with specialized pacemaking and conducting properties. This work determined sites of delay and spontane

Neuronal nicotinic acetylcholine receptors (nAChRs) are wide spread in the nervous system. Ample evidence indicates that many central nAChRs are located at the nerve terminals, where they act to facilitate neurotransmitter release; however, little is known about how these receptors carry out their function. The focus of my study is to understand the mechanism(s) that underlie the function of neuronal nAChRs. Neuronal nAChRs conduct inward current at negative membrane potentials, but conduct little or no outward current at positive membrane potentials, a process known as inward rectification. Inward rectification prevents the ACh-evoked conductance increase from short-circuiting the action potential at the nerve terminal, thereby ensuring optimal depolarization of the terminal and effective neurotransmitter release. Using the outside-out single channel patch-clamp technique, I demonstrate that intracellular polyamines block neuronal nAChRs with high affinity and in a voltage dependent manner; this is true for native nAChRs expressed by sympathetic neurons as well as recombinant alpha3beta4, alpha4beta2 nAChRs expressed in Xenopus oocytes. Given the physiological concentrations of polyamines inside cells, this block can fully account for the strong macroscopic inward rectification. Furthermore, using a combined approach of site-directed mutagenesis and electrophysiology, I show that the negatively charged residues at the cytoplasmic mouth of the pore (known as the intermediate ring) mediate the interaction of intracellular polyamines with the receptor; partial removal of these residues abolishes the strong inward rectification. Interestingly, I show that the intermediate ring influences the permeation of calcium through the receptor, indicating that a molecular link exists between calcium permeability and inward rectification of neuronal nAChRs. My experiments also show that extracellular polyamines and a polyamine-related toxin, Joro spider toxin, block neuronal n

The excitability properties of multicellular and single cellular cardiac pacemaking tissues was examined using mathematical, electrophysiological and molecular biological techniques. Particular attention was focused on identifying the different ionic channels that control the excitability of a physiological pacemaker, the rabbit atrioventricular node. The first part of the study centers on the excitability and rhythmicity of rabbit atrioventricular cell clusters and embryonic chick heart cell aggregates to compare cardiac pacemaking tissues showing slow- and fast-type action potentials. Subsequent experiments examined the membrane excitability properties of single isolated atrioventricular nodal cells and characterized the principle ionic currents that govern excitability. One of these channels that is directly gated by cyclic nucleotides, the "pacemaker" channel (If), was found to be non-uniformly expressed in different populations of AV nodal cells. The final part of the thesis involved a molecular characterization of cyclic nucleotide-gated ion channel subunits expressed in cardiac tissue during embryonic development. The relationship of these channels to the cardiac "pacemaker" channel (If) and their possible role in controlling cardiac excitability were considered.

No description available.

This study has focussed on the formation and survival of acetylcholine receptor (AChR) clusters at neuromuscular synapses formed in culture between myotomal muscle cells and spinal cord neurons derived from embryos of Xenopus laevis. AChRs were labelled with tetramethylrhodamine-conjugated $ alpha$-bungarotoxin so that the neurite associated receptor patches (NARPs) on the muscle cells could be viewed by fluorescence microscopy. Reduced fluorescence excitation was used in combination with a low light level TV camera and a computer based image calculator to make daily observations on all NARPs. Observations suggest the following conclusions. Neurons retain the capacity to trigger NARP formation as long as they continue to grow. Changes in NARP shape as well as decreases (and increases) in NARP size can occur even in the absence of competitive interactions between neurons and these changes are locally regulated along the contact. The capacity of neurons to maintain NARPs in more proximal portions of their neuritic arbor persists even as growing distal portions continue to induce the formation of new NARPs.

No description available.

The static mechanical properties of the respiratory system have been studied in newborn mammals (body size from rat to pig) and in animals (rats and rabbits) during growth. The elastic recoil pressure at functional residual capacity of the newborn is less than the adult value. The dry lung weight (LW)/body weight (BW) ratio is larger in the newborns (NB) than the adults (A). Functional residual capacity (FRC) is directly related to BW in the newborn. In general the FRC/BW ratio decreases with age while the FRC/LW increases. Respiratory system, lung and chest wall compliance are interspecific constants among newborn mammals. Examination of the NB and A allometric functions suggests that in general changes of respiratory variables are maturation dependent rather than size dependent. From the measurements of respiratory frequency and tracheal dimensions and assuming the calculated tracheal volume to be proportional to dead space the allometric relationship for dead space ventilation may be obtained. By employing the known allometric function for minute ventilation calculation suggests a constant alveolar P(,CO(,2)) and P(,O(,2)) for newborn mammals. / Studies of the onset of breathing and the pattern of breathing at 10 min, 60 min, 90 min, and a few days after birth have been made in infants delivered by cesarean section and vaginally. The pattern of breathing at birth is irregular. The largest contribution to the formation of functional residual capacity in the first 30 seconds of life is made by the first breath. The first breath is deeper and slower with a longer expiration than subsequent breaths. The irregularity of the breathing pattern improves from 10 to 90 min after delivery. Breathing frequency remains high in this period (10-90 min) compared to a few days and tidal volume is increased at a few days compared to the early period of life.

This research serves to answer the following basic question: Are potential toxic effects of hemoglobin solutions due to hemoglobin itself? Homologous and heterologous stroma-free hemoglobin and polyhemoglobin were investigated for their effects on complement activation, blood coagulation, platelet aggregation, blood cell counts, plasma electrolytes, proteins and enzymes in rats. A modified Mayer's method for testing whole complement hemolytic activity (CH$ sb{50}$) was developed. C$ sb3$ and C$ sb{ rm 3a}$ were also measured. Purified homologous or heterologous stroma-free hemoglobin or polyhemoglobin did not cause complement activation when incubated with rat plasma or when infused into rats. In addition, blood coagulation parameters did not change significantly in rats who received purified homologous or heterologous stroma-free hemoglobin or polyhemoglobin. Polyhemoglobin prepared from homologous or heterologous hemoglobin did not induce any changes in total leucocyte, differential or platelet counts. Effects of PolyHb and stroma-free hemoglobin on platelet aggregation were also studied. Plasma electrolytes, proteins, and enzymes tested were all within the normal ranges in rats who received purified homologous or heterologous polyhemoglobin infusions. Infusion of membrane stroma and endotoxins caused complement activation. This suggests that it is not purified hemoglobin or polyhemoglobin, but contaminants in hemoglobin preparations, which are responsible for some of the potential toxic effects observed. This study also has practical implications. It shows that only highly purified hemoglobin preparations should be used as blood substitutes for clinical applications.

The effect of lung deformation upon its mechanical properties was studied by measuring the quasi-static (Cstat) and dynamic (Cdyn) compliances of excised newborn piglet lungs deformed by a vertical pleural surface pressure gradient. Cstat was not affected by deformation. Cdyn was less than Cstat in the deformed and undeformed lungs but the difference was greater in the deformed lungs. This behavior suggests that distortion of the chest wall may increase the external (PV) work of breathing in newborns. / The contribution of lung viscoelasticity to the difference between Cstat and Cdyn of the undeformed lung was examined by measuring stress relaxation, Cstat, and Cdyn of excised lungs from newborn kittens and adult cats. The relative difference between Cdyn and Cstat and the magnitude of stress relaxation decreased with age, indicating that the viscous nature of the newborn's lungs causes larger, rate dependent changes in its compliance compared to the adult's. / Age related changes in the viscoelastic properties of the rat respiratory system were examined in rats between 1 and 40 days old. The rate of stress relaxation increased in the first week and thereafter decreased. Mean rates of stress relaxation were used to predict rate dependent changes in the passive recoil pressure of the respiratory system using an empirical model of viscoelasticity. Predictions were close to actual values suggesting that frequency dependent changes in the dynamic compliance of the newborn's respiratory system can occur independently of mechanisms involving the distribution of ventilation.