Recently, various proteoglycans (PGs) have been identified in the lung. The first objective of this thesis was to test the hypothesis that matrix glycosaminoglycans contribute to lung tissue viscoelasticity. Lung parenchymal strips were exposed to specific glycosaminoglycans-degradating enzymes to determine whether the mechanical properties of the tissue were affected. The degradation of heparan sulphate and chondroitin/dermatan sulphate glycosaminoglycans caused significant increases in energy dissipation and dynamic resistance relative to control strips. Hyaluronidase treatment did not alter any of the dynamic or static measures. Since PGs were found to be part of the stress bearing structure, the second part of the thesis aimed at examining whether subjecting the lung to excessive mechanical force can cause alteration in PG composition so as to adapt to the altered stress bearing requirement. To address this hypothesis, the effect of different ventilation regimes on lung tissue mechanics and PGs was examined in an in vivo rat model. After 2 h of mechanical ventilation, lung tissue elastance and resistance were significantly increased in rats ventilated with tidal volume of 30 ml/kg at 0 positive end-expiratory pressure (Vt30PEEP0) as compared to controls (Vt8PEEP1.5). Versican, a basement membrane heparan sulphate PG and biglycan, were all increased in rat lungs ventilated with Vt30PEEP0 as compared to control. These data demonstrated that alterations in lung tissue mechanics with excessive mechanical ventilation are accompanied by changes in all classes of ECM PGs. However, whether the alteration seen in PG composition resulted from excessive mechanical ventilation directly was unclear. In addition the cellular source of these PGs was not determined. Therefore, the aim of the third part of the thesis was to investigate and characterize the effect of mechanical strain on lung fibroblast PG production in vitro. We found cell layer associated versican protein in

A recursive least-squares algorithm was developed to estimate respiratory mechanical parameters with high temporal resolution. This algorithm was used to investigate the time course of bronchoconstriction induced by intravenous histamine injection in the dog. The onset of the response of lung tissue resistance and elastance demonstrated a different time course than airway resistance. This was interpreted in terms of the sequential delivery of the drug first through the pulmonary and then the bronchial circulations. The time course of respiratory mechanical parameters among various alveolar capsules revealed two patterns of inhomogeneity development. The first one was random whereas the second one was progressive with dose. A mathematical derivation elucidated the negative tissue resistance frequently obtained at high levels of constriction. The time courses of respiratory resistance and elastance during bronchoconstriction were transient and scaled with dose. They were reproducible for repeated doses of histamine after indomethacin pre-treatment and were intrinsically modulated by the adrenergic sympathetic system and through the $ rm H sb2$ histamine receptors.

The mechanisms underlying the net fluid/electrolyte gain during pregnancy are not fully understood. Utilizing virgin, pregnant (7, 16 and 21 days gestation), estradiol-17beta- or progesterone-treated female rats, we have examined the role of atrial natriuretic peptide (ANP) and of its receptors in the adaptation of body fluid homeostasis during pregnancy. Pregnancy and progesterone attenuate the ANP-mediated inhibition on aldosterone production in adrenal zona, glomerulosa (ZG) cells. The ribonuclease protection assay and Western analysis revealed that ZG natriuretic peptide guanylyl cyclase-linked (GC) receptors are mainly GC-A and that they are downregulated by pregnancy. In addition, pregnancy downregulates GC-A and GC-B receptors in the uterus; however, it does not modulate GC-linked receptors in the lung. It is concluded that the downregulation of ZG GC-A receptors could lead to a decrease in the aldosterone-suppressant effects of ANP and that the decrease in uterine GC-A and GC-B receptors could lead to a decrease in the tocolytic effects of ANP during pregnancy.

The 5-lipoxygenase product 5-oxo-6,8,11,14-eicosatetraenoic acid (5-oxo-ETE) is a potent eosinophil chemoattractant in vitro. To determine whether it is active in vivo, 5-oxo-ETE was administered intratracheally to BN rats and pulmonary eosinophils were immunostained with an antibody to major basic protein. 5-Oxo-ETE induced a dramatic increase in eosinophils, which reached maximal levels (5 times control) between 15 and 24 h following administration, and thereafter declined. LTB4 had a similar effect to 5-oxo-ETE but appeared to be somewhat less effective. In contrast, LTD4 and LTE4 were inactive. 5-Oxo-ETE-induced eosinophilia was inhibited by 75% following pretreatment of the animals with antibodies to integrins VLA-4 or LFA-1, but was not significantly inhibited by an antibody to Mac-1, nor after pretreatment with receptor antagonists to LTB4 (LY255283) or PAF (WEB 2170). These observations raise the possibility that 5-oxo-ETE may be an important physiological mediator in inflammatory diseases characterized with eosinophil recruitment, such as asthma.

Autoregulation of renal blood flow is mediated by two mechanisms. The myogenic response operates at $ approx$0.1-0.2 Hz and tubuloglomerular feedback (TGF) operates at $ approx$0.03-0.05 Hz. Atrial natriuretic factor (ANF) dilates pre-glomerular resistance vessels, in which autoregulation occurs, and has been reported to inhibit TGF. We tested the potential actions of ANF on TGF-mediated autoregulation using Sprague-Dawley rats anesthetized with isoflurane. Renal perfusion pressure (RPP) was manipulated by a servo-controlled clamp placed on the aorta between the renal arteries. Time constants for constriction and dilatation were consistent with operation of TGF. ANF did not affect either the magnitude or the time constants of the response. / To test the contribution of TGF resetting to the wide dynamic range of steady state autoregulation experiments, the autoregulatory range was first defined by conventional stepwise reduction of RPP. As ANG II is necessary for TGF resetting, the experiment was repeated in the presence of Losartan, a competitive ANG Il-AT$ sb1$ receptor antagonist. The results do not support the hypothesis that TGF resetting contributes to the wide dynamic range of steady state autoregulation. In fact, the lower limit of autoregulation was extended to a lower RPP. In the Losartan experiment, this shift was not apparent, suggesting that it was ANG II-dependent. (Abstract shortened by UMI.)

Familial hypophosphatemic rickets and its murine model, Hyp, are characterized by a specific defect in renal inorganic phosphate reabsorption. In Hyp mice the defect has been localized to the proximal tubule brush border membrane and a constitutively enhanced protein kinase C has been demonstrated. Since the latter is activated through the protein G pathway, we have studied phosphatidyl inositol 4,5-bisphosphate metabolism (PIP$ sb2$) in normal (+/Y) and Hyp primary cultures of proximal tubules. At confluence, ($ sp3$H) -myo-inositol labelled cultures were stimulated with calcitriol or parathyroid hormone (PTH) for one minute. Cells were extracted and the aqueous phase was chromatographed on ion exchange resin. The inositol phosphate fractions were counted for radioactivity. For calcitriol, maximal PIP$ sb2$ metabolism occurred at 10$ sp{-11}$ M and 10$ sp{-12}$ M for +/Y and Hyp, respectively. For both cultures, PTH produced a maximum at 10$ sp{-8}$ M. Pertussis toxin alone stimulated +/Y but not Hyp cultures and produced no additive stimulation with calcitriol or PTH in either culture.

Plasma acylation stimulating protein (ASP) is a small basic protein isolated from human plasma that stimulates triglyceride synthesis (TGS) and glucose transport (GT) in human skin fibroblasts and adipocytes. ASP was measured in fasting samples of 59 age-matched controls and 208 patients with documented coronary artery, disease (CAD). Overall, plasma ASP was higher in the CAD patients than in the controls and within the CAD patients, plasma ASP was highest in the group that had neither an FHTG-type (hypertriglyceridemic normal apoB) nor an FH-type (marked hypercholesterolemia and apoB with normal plasma triglyceride) phenotype. The relations between plasma ASP, triglyceride rich lipoproteins and apoB in the remaining CAD subjects were examined and ASP increased with either increasing triglyceride or apoB. In 208 CAD subjects, 47 subjects had ASP greater than 95th percentile (greater than 65 nM). Within those 47 subjects, 66% had increased plasma apoB (apoB > 105mg/dl). / The relationship between ASP and hyperapoB (apoB > 105mg/dl) was further investigated in cell culture study. Human skin fibroblasts were obtained from normals and hyperapoB patients. The fibroblasts from the three groups of subjects (normal subjects:NASP-NB, hyperapoB subjects with normal plasma ASP:NASP-HB, and hyperapoB subjects with high plasma ASP:HASP-HB) were studied for TGS and GT in response to ASP, PMA (4beta phorbol 12-myristate 13-acetate, a protein-kinase C stimulator) and insulin stimulation. The competition binding of 125I ASP to the fibroblasts from the three groups of subjects was also examined. / The data showed that only HASP-HB cells have a lower response to ASP in TGS and GT. The responses to insulin and PMA in TGS and GT are normal. There is no significant difference among the three groups in the Kd values (NASP-NB Kd: 163 +/- 23nM) for binding to 125I ASP. The Bmax value for the H ASP-HB group is 50% lower than the other two groups (N ASP-NB:NASP-HB:HASP-H B: 334 +/- 81:427 +/- 92:146 +/- 33 pmoles/mg cell protein). / The reduced receptor of ASP on the cell surface of HASP-H B patients can lead to the reduced response to ASP.

The effects of taurine deficiency on aspects of retinal cGMP metabolism were examined. Rats were made taurine deficient by the administration of 1% GES. The activities of the key enzymes of cGMP metabolism, phosphodiesterase and guanylate cyclase, were indirectly determined by the response of retinas to light and to low calcium. Dark-adapted retinas of rats treated with GES for 4 and 8 weeks exhibited significantly smaller decreases in cGMP in response to light exposure as compared to control retinas. Retinas from 4 and 8 week GES-treated rats also exhibited smaller increases in cGMP when they were exposed to 0.1nM calcium. In addition, dark-adapted retinas of 8 week GES-treated rats had 36% less cGMP than dark-adapted control retinas. Since a light-induced decrease of cGMP is of central importance in phototransduction, these results support the hypothesis that abnormalities in the retinal cGMP metabolism of taurine-deficient rats may account for the electro-retinographic deficits observed in these animals.

The purpose of this study was two-fold: (1) to examine the relationship between the amount of airway smooth muscle and the airway responsiveness to inhaled aerosolized methacholine (MCh) in guinea pigs, and (2) to characterize the distribution of airway narrowing following MCh. / In summary: (1) the quantity of airway smooth muscle (ASM) does not appear to determine differences in maximal bronchoconstriction among normal guinea pigs; the lack of a correlation between responsiveness and amount of ASM may be explained by the heterogenous distribution of bronchoconstriction among the airways studied or the modality of challenge; (2) the sensitivity to MCh appears to be related to differences in the amount of ASM in intraparenchymal cartilaginous airways; (3) variability in the EC$ sb{50}$ may also reflect differences in airway cross-sectional area; (4) lung resistance appears to be a good measure of constriction since the morphometric measure of airway narrowing correlated well with resistance; (5) the heterogeneity of airway narrowing does not appear to be determined by differences in ASM.

Generalized spike and wave (SW) activity of feline generalized penicillin epilepsy (FGPE), an experimental model of epilepsy exhibiting a sufficient degree of similarity with human generalized 3 Hz SW discharge, depends upon both cortical and thalamic mechanisms. Cortical neurons under the influence of systemic penicillin become more excitable and respond more vigorously to thalamocortical volleys normally inducing spindles or recruiting responses. This increased response of cortical neurons activates phasic recurrent inhibition which remains intact during FGPE as is shown by the preservation of antidromically evoked recurrent inhibition of pyramidal tract cells. Thus, an oscillation between periods of increased and decreased neuronal firing probability ensues which underlies SW discharge in the EEG. The cortex imposes this oscillatory pattern of neuronal discharge upon the thalamus, which by itself, however, is incapable of responding in this matter to systemic penicillin, as it appears from experiments performed in decorticated cats. Nevertheless, the thalamus is not a mere passive follower of the cortex in SW activity, since thalamocortical inputs remain necessary to maintain it. Furthermore thalamic neurons, like cortical ones, undergo a cyclically alternating pattern of increased and decreased firing probability. They display three different modes of phase-locked timing of their discharges in relation to cortical SWs and the cortical neuronal firing pattern. Two groups of cells fire coincidentally with the "spike" of the SW complex, one group reaching its maximum firing probability slightly before, the other slightly after that of the simultaneously recorded cortical neuron. The third group fires during the "wave" of the SW complex and its firing pattern is therefore about 180(DEGREES) out of phase with that of the cortical and the other two groups of thalamic neurons. Thus the substrate of generalized SW discharge seems to be oscillatory activity within a c